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CONTENTS
Volume 314, Issue 5804

COVER DEPARTMENTS
Artist’s representation of communication 1347 Science Online
pathways initiated by cell surface receptors 1349 This Week in Science
that influence cell physiology and organelle 1355 Editors’ Choice
function. This joint special issue between 1360 Contact Science
Science and Science’s STKE highlights 1363 Random Samples
new insights into signaling mechanisms 1365 Newsmakers
that control development and reproduction 1471 New Products
(see page 1409). 1472 Science Careers
Image: Christopher Bickel

1410 CONTENTS continued >>

www.sciencemag.org SCIENCE VOL 314 1 DECEMBER 2006 1341

Systems Biology — RNAi

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CONTENTS

SCIENCE EXPRESS
www.sciencexpress.org

MOLECULAR BIOLOGY
P[acman]: A BAC Transgenic Platform for Targeted Insertion of Large DNA
Fragments in Drosophila Melanogaster GEOPHYSICS
K. J. T. Venken, Y. He, R. A. Hoskins, H. J. Bellen Slow Earthquakes Coincident with Episodic Tremors and Slow Slip Events
A method allows efficient site-specific integration of large DNA sequences and thus Y. Ito, K. Obara, K. Shiomi, S. Sekine, H. Hirose
manipulation of proteins in vivo in Drosophila and potentially other organisms. A series of weak low-frequency earthquakes correspond with seismic tremor and slip
>> News story p. 1371 episodes on a subduction zone beneath Japan, perhaps increasing overall stress.
10.1126/science.1134426 10.1126/science.1134454
EVOLUTION ASTROPHYSICS
Homoploid Hybrid Speciation in an Extreme Habitat Spectropolarimetric Diagnostics of Thermonuclear Supernova Explosions
Z. Gompert, J. A. Fordyce, M. L. Forister, A. M. Shapiro, C. C. Nice L. Wang, D. Baade, F. Patat
As postulated by theory, a new species of butterfly evolved when a hybrid of two A survey of supernovae shows that brighter ones have more spherical explosions,
existing species became adapted to an extreme alpine environment. constraining the physics of burning and improving their use as standard candles.
10.1126/science.1135875 10.1126/science.1121656

LETTERS TECHNICAL COMMENT ABSTRACTS

Balancing Communication and Safety S. A. Ehrlich 1387 EVOLUTION
Glossing Over the Complexity of Water G. Kallis, Comment on “Population Size Does Not Influence 1390
M. Kiparsky, A. Milman, I. Ray Mitochondrial Genetic Diversity in Animals”
Mitochondrial DNA and Population Size O. F. Berry; C. J. Mulligan, A. Kitchen, M. M. Miyamoto
J. P. Wares et al. Response E. Bazin et al. full text at www.sciencemag.org/cgi/content/full/314/5804/1390a

BOOKS ET AL. BREVIA

The Other Insect Societies 1391 CLIMATE CHANGE
J. T. Costa, reviewed by R. Gadagkar Old-Growth Forests Can Accumulate Carbon in Soils 1417
The Creation An Appeal to Save Life on Earth 1392 G. Zhou et al.
E. O. Wilson, reviewed by S. Bouma-Prediger Old-growth forests in Southern China accumulated atmospheric
carbon at a rate considerably greater than expected for
Nota Bene: Game On Science Museum, London 1393 broadleaved evergreen forests.

POLICY FORUM RESEARCH ARTICLE

When Patents Threaten Science 1395 ATMOSPHERIC SCIENCE
L. Andrews, J. Paradise, T. Holbrook, D. Bochneak Phytoplankton and Cloudiness in the 1419
Southern Ocean
PERSPECTIVES N. Meskhidze and A. Nenes
The Turing Model Comes of Molecular Age 1397 Oxidation of aerosols released from a phytoplankton bloom
P. K. Maini, R. E. Baker, C.-M. Chuong doubled the number of droplets in overlying clouds and reflected
>> Report p. 1447 solar radiation as much as severe air pollution.

Variable High-Energy γ Rays from the Elliptical 1398

Galaxy M87
REPORTS
A. C. Fabian >> Report p. 1424 ASTRONOMY
Fast Variability of Tera–Electron Volt γ Rays from 1424
When Dry Air Is Too Humid 1399
the Radio Galaxy M87
T. Peter et al.
F. Aharonian et al.
Tools to Tamper with Phosphoinositides 1402 Very-high-energy gamma rays from the radio galaxy M87
S. McLaughlin >> Reports pp. 1454 and 1458 vary daily, implying that they originate close to the central
Delivering New Disease Genes 1403 supermassive black hole. >> Perspective p. 1398
L. R. Cardon >> Report p. 1461 PHYSICS
Edward I. Stiefel (1942–2006) 1406 Solid-State Qubits with Current-Controlled Coupling 1427
F. M. M. Morel and J. T. Groves T. Hime et al.
Manipulation of the mutual inductance between two qubits can be
used to switch their coupling on and off.

CONTENTS continued >>

www.sciencemag.org SCIENCE VOL 314 1 DECEMBER 2006 1343

Good Vibrations... Great Sections!
”Ideally, a tissue slicer should generate large-amplitude and high-frequency movements of the cutting blade
in a horizontal axis, with minimal vibrations in the vertical axis.“ *(According to Prof. P. Jonas, Institute of
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CONTENTS

REPORTS CONTINUED...
APPLIED PHYSICS
Optical Atomic Coherence at the 1-Second Time Scale 1430 1439
M. M. Boyd et al.
A highly stable laser and the ability to trap a large number of atoms
coherently provide a tenfold increase in measuring spectral lines BIOCHEMISTRY
needed for precision applications. Structural Basis for Ribosome Recruitment and 1450
MATERIALS SCIENCE Manipulation by a Viral IRES RNA
Macroscopic Hierarchical Surface Patterning of 1433 J. S. Pfingsten, D. A. Costantino, J. S. Kieft
Porphyrin Trimers via Self-Assembly and Dewetting The structure of a viral RNA containing an internal ribosomal
R. van Hameren et al. entry site suggests how translation can begin in the middle
Upon dewetting, a molecule containing porphyrin and long alkyl of a messenger RNA.
groups can self-assemble in long chains and patterns over areas NEUROSCIENCE
as large as several square millimeters. Rapid Chemically Induced Changes of 1454
CHEMISTRY PtdIns(4,5)P2 Gate KCNQ Ion Channels
Probing the Chiroptical Response of a Single Molecule 1437 B.-C. Suh, T. Inoue, T. Meyer, B. Hille
R. Hassey et al. Neurotransmitters close a potassium channel by changing
Circular dichroism spectra at high resolution reveal that weak the lipid content of the surrounding plasma membrane.
aggregate signals arise because the effects of distinct conformations >> Perspective p. 1402
in a chiral ensemble cancel each other. CELL BIOLOGY
GEOCHEMISTRY PI(3,4,5)P3 and PI(4,5)P2 Lipids Target Proteins with 1458
Organic Globules in the Tagish Lake Meteorite: 1439 Polybasic Clusters to the Plasma Membrane
Remnants of the Protosolar Disk W. D. Heo et al.
K. Nakamura-Messenger et al. Two phospholipid signaling molecules are also essential to anchor
Carbon-rich nanospheres in a primitive meteorite are relatively proteins that have clusters of basic amino acids to the cell membrane.
enriched in the heavy nitrogen isotopes and deuterium, suggesting >> Perspective p. 1402
that these grains have a pre-solar origin. GENETICS
ATMOSPHERIC SCIENCES A Genome-Wide Association Study Identifies IL23R 1461
Increasing Trend of Extreme Rain Events Over India 1442 as an Inflammatory Bowel Disease Gene
in a Warming Environment R. H. Duerr et al.
B. N. Goswami et al. People with a rare sequence variant of the gene encoding the
The frequency and intensity of heavy rainfall events during monsoon receptor for an immunological cytokine have a reduced risk
storms in Central India have increased during the past 50 years as the of inflammatory bowel disease. >> Perspective p. 1403
climate there has warmed. MICROBIOLOGY
EVOLUTION Microfluidic Digital PCR Enables Multigene Analysis 1464
Male Fertility and Sex Ratio at Birth in Red Deer 1445 of Individual Environmental Bacteria
M. Gomendio et al. E. A. Ottesen, J. W. Hong, S. R. Quake, J. R. Leadbetter
Like females, males can affect offspring sex ratio; more-fertile male A DNA analysis method that can link genes to individual organisms
red deer sire more sons and less-fertile males sire more daughters. collected in the wild is used to identify a gut symbiont of the termite.

DEVELOPMENTAL BIOLOGY MEDICINE

WNT and DKK Determine Hair Follicle Spacing 1447 Prevention of Brca1-Mediated Mammary 1467
Through a Reaction-Diffusion Mechanism Tumorigenesis in Mice by a Progesterone Antagonist
S. Sick, S. Reinker, J. Timmer, T. Schlake A. J. Poole et al.
Modeling and experimental tests explain how a growth factor Experiments in mice suggest that a mutation leading to
and its inhibitor determine the density and pattern of hair follicles breast cancer acts in part by altering signaling by the steroid
in the developing mouse. >> Perspective p. 1397 hormone progesterone. >> News story p. 1370

SCIENCE (ISSN 0036-8075) is published weekly on Friday, except the last week in December, by the American Association
for the Advancement of Science, 1200 New York Avenue, NW, Washington, DC 20005. Periodicals Mail postage (publication No.
484460) paid at Washington, DC, and additional mailing offices. Copyright © 2006 by the American Association for the Advancement
of Science. The title SCIENCE is a registered trademark of the AAAS. Domestic individual membership and subscription (51 issues): $139
($74 allocated to subscription). Domestic institutional subscription (51 issues): $650; Foreign postage extra: Mexico, Caribbean (surface
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Change of address: Allow 4 weeks, giving old and new addresses and 8-digit account number. Postmaster: Send change of address to AAAS, P.O. Box 96178, Washington, DC 20090–6178. Single-copy sales:
$10.00 current issue, $15.00 back issue prepaid includes surface postage; bulk rates on request. Authorization to photocopy material for internal or personal use under circumstances not falling within the
fair use provisions of the Copyright Act is granted by AAAS to libraries and other users registered with the Copyright Clearance Center (CCC) Transactional Reporting Service, provided that $18.00 per article is
paid directly to CCC, 222 Rosewood Drive, Danvers, MA 01923. The identification code for Science is 0036-8075. Science is indexed in the Reader’s Guide to Periodical Literature and in several specialized indexes.
CONTENTS continued >>

www.sciencemag.org SCIENCE VOL 314 1 DECEMBER 2006 1345

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www.sciencemag.org

Adapting to stem cell

research policies.

SCIENCE CAREERS
www.sciencecareers.org
CAREER RESOURCES FOR SCIENTISTS

EUROPE: Navigating the Stem Cell Research Maze

S. Webb and E. Pain
Building a stem cell research career in Europe means navigating
the policy maze in each country.
US: It Ain’t Over Till It’s Over
B. Benderly
A bureaucratic snag has stalled California’s postdoc unionization Fit for consumption.
drive.
CANADA: Winning the HHMI International Research Award
A. Fazekas SCIENCENOW
A young biomedical researcher explains how career choices www.sciencenow.org DAILY NEWS COVERAGE
translated into professional success.
Taking the Toxin out of Cotton
GRANTSNET: December 2006 Funding News Engineered plants have edible seeds, providing a possible
J. Fernandez new source of cheap protein.
Read about the latest in research funding, scholarships,
Clocking Cosmic Eyewalls
fellowships, and internships for postdocs and students.
Scientists measure the speed of a spinning black hole.
Chimps Go Ape Over Older Females
Findings give clues to evolution of human mate preference.

SPECIAL SECTION

Cell Signaling
SCIENCE’S STKE
www.stke.org
SIGNAL TRANSDUCTION KNOWLEDGE ENVIRONMENT

EDITORIAL GUIDE: Signal Reception and Transmission

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New pathways and updates to the Database of Cell Signaling
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Separate individual or institutional subscriptions to these products may be required for full-text access.

www.sciencemag.org SCIENCE VOL 314 1 DECEMBER 2006 1347

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EDITED BY STELLA HURTLEY AND PHIL SZUROMI

<< Fast Flickering

Among the very few known extragalactic emitters of very high
energy tera–electron volt (TeV) γ rays are blazars, which are
galaxies with relativistic particle jets that point toward
Earth. It has been suggested that the TeV γ rays originate in
those jets. By monitoring the nearby radio galaxy M87,
whose twin jets are oriented in the plane of the sky rather
than pointed at us, Aharonian et al. (p. 1424, published
online 26 October; see the Perspective by Fabian) show that
γ rays in active galaxies are actually produced near the cen-
tral black hole. M87 is bright in γ rays up to 10 TeV, and its
brightness varies daily. Such fast variations imply the source of
the γ rays lies near the Schwarzschild radius of the supermassive
black hole that lies at the heart of the M87 galaxy. Although this
behavior may fit some leptonic models for γ-ray production, an
alternative mechanism of proton curvature radiation near to the black
hole is proposed.

Phytoplankton Clouds values by an order of magnitude. Such capabil- achieved. Hime et al. (p. 1427) demonstrate
Phytoplankton produce compounds that can ities facilitate high-precision unit standardiza- on-and-off control on a pair of superconduct-
become aerosols, which suggests that biologi- tion and enhanced measures of fundamental ing-flux qubits coupled through their mutual
cal productivity might exert an important con- physical constants. inductance. With both qubits also coupled to a
trol on cloudiness over the ocean if these nearby superconducting quantum interference
aerosols act as cloud condensation nuclei. device (SQUID), their mutual inductance and
Meskhidze and Nenes (p. 1419, published Monsoon Violence the extent of the coupling strength could be
online 2 November) combine satellite observa- Most climate models have predicted that controlled by varying the working parameters
tions of surface ocean chlorophyll a content extreme rainfall events will become more of the SQUID.
and cloud cover to show that biological produc- common as air temperature rises, but obser-
tivity can have a significant effect on shallow vational evidence of this trend has been
marine clouds. Cloud droplet number concen- hard to find. Goswami et al. (p. 1442) Lining Up at
trations over a phytoplankton bloom in the used a daily rainfall data set for central
Southern Ocean doubled, and cloud effective India to show that there was an increase in the Front
radius was reduced by 30%, which led to a the frequency and intensity of heavy rain Self-assembly of molecules can
large change in the short-wave radiative flux at events, and a decrease in the frequency of create nanoscale features on flat
the top of the atmosphere. light to moderate rain events, for the mon- surfaces, but the maximum extent
soon seasons from 1951 to 2000. The of a single domain is usually on
mean rainfall did not show a significant the order of tens of micrometers.
In Tune for a Second trend because the increasing contribution Van Hameren et al. (p. 1433)
High-resolution spectroscopy generally from heavy events was offset by a decreas- show that disk-like molecules, in
CREDITS (TOP TO BOTTOM): AHARONIAN ET AL.; GOSWAMI ET AL.

requires a trade-off between the size of the ing one from light ones. These findings which three porphyrin groups bear-
ensemble being probed and the coherence of suggest that severe rain events over India ing long alkyl groups assemble
that sample during the course of the measure- will become more common if global warm- around a central core, form very
ment, so that increasing the sample size to ing continues as expected. long aligned chains over areas
raise signal strength often broadens the signal of several square millimeters
of interest. Boyd et al. (p. 1430) have used an through a dewetting process. On
optical trap to inhibit the random motion of
strontium atoms in order to maintain coher-
Controlled Coupling mica, single-column stacks form lines parallel to
the evaporation front of smaller droplets, whereas
ence of the photoexcited sample for ~1 second. of Qubits for larger droplets, longer evaporation times
By careful frequency stabilization of the probe Performing logical operations on quantum com- cause larger lines of aggregates to grow normal
laser, an absorption line at ~1014 hertz could puters will require the coupling and decoupling to the evaporation front. Patterns formed on
be measured with a corresponding width of of qubits so that individual qubits can be pre- rougher glass surfaces were less regular but could
~1 hertz. The attained ratio of frequency to pared in a given quantum state, allowed to still be used to align liquid crystal molecules.
linewidth, or quality factor, exceeds previous interact, and be read out once the final state is Continued on page 1351

www.sciencemag.org SCIENCE VOL 314 1 DECEMBER 2006 1349

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This Week in Science
Continued from page 1349

More Boys Preferred

Biases in sex ratio at birth have led to the suggestion that females may manipulate the sex of their
offspring. Gomendio et al. (p. 1445) now show that males may also influence offspring sex ratio.
In red deer, more fertile males tend to produce proportionally more sons who are likely to inherit
high fertility rates. Sperm collected during the rut from males living in natural populations was
used for artificial insemination to minimize known female effects on sex ratio. Such male contribu-
tions to biases in offspring sex ratio suggest an evolutionary scenario in which conflicts of interest
between males and females in relation to the sex of their offspring may play an important role.

Turing Patterning in the Mouse Hairs

More than 50 years ago, Alan Turing provided a theoretical explanation of biological pattern for-
mation through a hypothesis of reaction-diffusion, whereby patterns, such as that for hair follicles
or feather distribution, can form as a result of positive and negative feedback regulation of an
inhibitor and activator. Turing models have since been used to account for patterns in many chemi-
cal systems, but have not been successful in explaining biological pattering in developmental
model systems such as the fly. Sick et al. (p. 1447, published online 2 November; see the Perspec-
tive by Maini et al.) have now examined hair follicle arrangements in mice that arise through the
WNT activator protein and its inhibitor DKK and show through computation modeling that reac-
tion-diffusion can account for the patterning observed.

Not Lost in Translation

The canonical mechanism for initiation of protein synthesis in
eukaryotes involves a nucleotide cap on messenger RNA
(mRNA) that is recognized by an initiation protein factor.
However, a variety of pathogenic viruses and cellular mRNAs
bypass the canonical mechanism by using structured RNA
sequences, called internal ribosomal entry sites (IRESs), to
initiate translation. Pfingsten et al. (p. 1450) have deter-
mined the structure of the ribosome-binding domain of an
IRES at 3.1 angstrom resolution. The RNA prefolds to create a
specific ribosome-binding structure. By docking the structure
onto cryoelectron microscopic reconstructions of an IRES-
ribosome complex, contacts were identified that drive bind-
ing and induce conformational change in the ribosome.

Of Genes and Gut Reactions

Inflammatory bowel diseases (IBDs) such as Crohn’s disease and ulcerative colitis are thought to be
caused by an inappropriate immune response to commensal intestinal bacteria. There is strong evi-
dence that these disorders have a genetic component; for example, individuals carrying specific
sequence variants of the NOD2/CARD15 gene are at increased risk. Now, in a genome-wide associ-
ation study, Duerr et al. (p. 1461, published online 26 October; see the Perspective by Cardon)
find that a rare sequence variant of the gene encoding the receptor for interleukin-23 (IL23R)
significantly lowers an individual’s risk of developing IBDs. Interleukin-23 is a cytokine that has
attracted increasing attention because of its role in a wide range of chronic inflammatory diseases
in mouse models, including IBDs, multiple sclerosis, and arthritis.

Progesterone and Breast Cancer

Mutations in the breast cancer susceptibility gene BRCA1 greatly increase a woman’s risk of develop-
ing breast and ovarian cancers. Why do these mutations predominantly affect hormone-responsive
tissues when the mutant gene is widely expressed throughout the body? Poole et al. (p. 1467; see
CREDIT: PFINGSTEN ET AL.

the news story by Marx) suggest that this tissue specificity is caused in part by BRCA1-mediated
effects on signaling by the hormone progesterone. Mammary epithelial cells (MECs) of Brca1/p53-
deficient mice accumulated high levels of progesterone receptors, probably through defective
degradation by the proteasome, and developed aberrant proliferation of the MECs. Treatment with
the progesterone antagonist mifepristone (RU 486) prevented or delayed mammary tumor develop-
ment in the mice.

www.sciencemag.org SCIENCE VOL 314 1 DECEMBER 2006

Science is a way of thinking
much more than it is a body of
knowledge.
Carl Sagan
Scientist (1934-1996)

Our core strengths include not only technologies that support superior products and services, but also the spark of ideas that
lights the way to a brighter future. Shimadzu believes in the value of science to transform society for the better. For more than
a century, we have led the way in the development of cutting-edge technology to help measure, analyze, diagnose and solve
problems. The solutions we develop find applications in areas ranging from life sciences and medicine to flat-panel displays.
We have learned much in the past hundred years. Expect a lot more.
EDITORIAL

Responding to Fraud
Our journal—as well as science with a small “s”—went through a disappointing and troubling
experience with the two stem cell papers from the South Korean research group led by Dr. Woo
Donald Kennedy is the
Suk Hwang. As a result of an investigation by a committee from Seoul National University,
Editor-in-Chief of Science.
the first paper from this group, Science 303, 1669 (2004), was found to be fraudulent and was
subsequently retracted by Science. A second paper, Science 308, 1777 (2005), published a year
later, was retracted for the same reasons.
What Science did then entailed two steps. First, we compiled a chronological anthology of
the editorial review process for both papers; it included all submissions; correspondence among
editors, our Board of Reviewing Editors, peer reviewers, authors, and agencies responsible for
regulatory oversight in South Korea; and notes on telephone conversations. This material was
reviewed by an internal review committee of six in-house
editors. This archive and their comments were then sent to
an outside committee consisting of three members of our
“The report sends us some tough
external Senior Editorial Board (John Brauman, George messages about what Science
Whitesides, and Linda Partridge), a former Science senior should do to confront a present
editor who is now the U.S. Executive Editor at Nature reality and prepare for a more
(Linda Miller), and two distinguished biologists who work
in the stem cell community (Doug Melton and John challenging future.”
Gearhart). The committee was asked to make a thorough
and unsparing analysis of Science’s handling of both papers and to make recommendations
for changes in procedure that might protect both the journal and the scientific community from
further unfortunate outcomes of this kind.
The report, and a short response from Science, are available at www.sciencemag.org/cgi/
content/full/314/5804/1353/DC1. The report is notable for its thoroughness, insight, and candor.
It reaches several conclusions; some of these apply to our journal and to those of us who edit
and publish it, and others are relevant for the larger community of scientists. The good news for
Science is that its editors and peer reviewers not only followed the procedures in place here
and at other top-tier journals, but made a substantially greater effort than for most papers to
ensure that the science was sound. The not-so-good news is that the report sends us some tough
messages about what Science should do to confront a present reality and prepare for a more
challenging future. It points out forcefully that the environment for science now presents
increased incentives for the production of work that is intentionally misleading or distorted by
self-interest. It urges us to give special attention to a relatively small number of papers that are
likely to be especially visible or influential.
We are now formulating ways to respond to this advice. The report recommends developing
a risk assessment template. We have been conducting discussions among ourselves and
with committee members to develop criteria for deciding which papers deserve particularly
careful editorial scrutiny. Papers that are of substantial public interest, present results that are
unexpected and/or counterintuitive, or touch on areas of high political controversy may fall
into this category. We are also considering the kinds of special attention that might be given to
these high-risk papers. These might include higher standards for including primary data,
demands for clearer specification of the roles of all authors, and more intensive evaluation of
the treatment of digital images. The report makes no bones about the fact that for some papers
that meet the higher risk standard, the experience will be time-consuming and expensive for the
journal and “may lead to conflict with authors.”
This is not the first time that scientific journals have had to adapt their procedures to new real-
ities in the world they live in. After 9/11 and the subsequent anthrax releases in the United States,
journals developed guidelines for recognizing and dealing with papers that might present inter-
national security problems. As we did then, we will be looking for ways to meet a new challenge,
while maintaining the integrity of the review process and minimizing damage to the expectations
of our authors and the speed of our publication process. We invite your comments and plan to
keep you informed as we develop particular policies in response to these recommendations.
– Donald Kennedy

10.1126/science.1137840

www.sciencemag.org SCIENCE VOL 314 1 DECEMBER 2006 1353

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EDITORS’CHOICE
EDITED BY STELLA HURTLEY AND JAKE YESTON

ECOLOGY/EVOLUTION

Going to the Dingoes

In the past 200 years, since the arrival of Europeans in Australia, 18 of the
continent’s mammal species have become extinct. These extinctions have
been chiefly attributed to introduced, non-native predators, especially
foxes and cats. Johnson et al. present evidence that the success of these
medium-sized introduced predators has been the direct result of
persecution by humans of Australia’s native large predator,
the dingo. In areas where dingoes have been left alone,
foxes and cat populations are kept at bay, and the
diversity and abundance of native marsupials are
greater. Thus, top predators can maintain biodiversity
at middle trophic levels and may help ecosystems to
resist invasion by alien species. By allowing dingo
populations to recover in regions where they have been
Dingo (top), rat-kangaroo (right). persecuted, it might be possible to insure remaining small
marsupials against further decline and extinction. — AMS
Proc. R. Soc. London Ser. B 10.1098/rspb.2006.3711 (2006).

CELL BIOLOGY disease, and other protein misfolding diseases CHEMISTRY

might be similarly amenable to equivalent inter-
Toward the Chaperome Catalyst Compatibility
ventions. — SMH
The expression of misfolded or aberrant proteins Cell 127, 803 (2006). The isolation and purification procedures that
on the cell surface could wreak havoc with the follow synthetic chemical reactions often produce
immune system. Cells have therefore developed GEOLOGY substantial quantities of waste material. Research
an efficient quality-control system, which diverts has thus increasingly focused on methods for
misfolded membrane and secretory proteins
Sediment Sources carrying out multiple reaction steps in a single
from the secretory pathway by retaining and The Nile River drains much of northeast Africa, and vessel. However, the mutual incompatibility of
degrading them at the entry portal to the secre- its sediments reflect erosion across the continent. many catalysts, in particular Lewis acids and bases,
tory pathway, the endoplasmic reticulum (ER). Dams such as the Aswan have caused efficient col- presents a major challenge to this approach.
One well-studied example of quality control lection of these sediments in the human-made Poe et al. present an encapsulation technique
involves the cystic fibrosis transmembrane con- lakes that form behind them. Garzanti et al. exam- that allows the mixing of a polymeric amine cata-
ductance regulator (CFTR), misfolding of which is ined the mineralogy and amount of sand dumped lyst with a nickel-centered Lewis acid while avoid-
CREDITS (TOP TO BOTTOM): MARTIN HARVEY/PETER ARNOLD, INC.; KARL VERNES; CORBIS

responsible for disease in a large proportion of by the Nile into these ing the complexation reaction that
sufferers. However, sometimes quality control is lakes and found that would deactivate both. The
too stringent, and functional, though mutant, ~200 million metric poly(ethyleneimine) base is
proteins are retained. Wang et al. used a system- tons of sediment are treated with a cross-linking agent
atic approach to examine the folding pathway transported per year, in a methanol/cyclohexane emul-
and protein interaction partners of CFTR and the several times the quanti- sion, yielding a microcapsule
common disease variant CFTR ΔF508, which, ties estimated previ- morphology that conserves cat-
even though functional, is retained in the ER. ously. The sand is mainly alytic activity in the condensation
A variety of chaperone proteins, which help to composed of basaltic reaction of benzaldehyde and
promote protein folding, are present in the ER, rock or feldspar and nitromethane. Addition of a
and a chaperome of over 30 proteins involved in metamorphic minerals, bis(diamino)nickel catalyst to the
The Aswan Dam.
CFTR folding and transport was identified from indicative of the reaction mixture promotes a
among more than 200 interacting proteins. Ethiopian highlands, an Michael addition of dimethyl mal-
In particular, Aha1, a Hsp90 co-chaperone area of abundant deforestation and farming that onate to the dehydrated product in ~80% yield.
ATPase regulator, was found to be important in receives monsoon rainfall during summer. Thus, a Moreover, the compatibility of the two catalysts is a
retaining mutant CFTR. When levels of Aha1 relatively small area of the Nile drainage, greatly boon to selectivity as well as efficiency; the nickel
were reduced, mutant CFTR managed to escape affected by humans, supplies most of the sedi- complex staves off a side pathway that would lead
from the ER and reached the plasma membrane. ments carried by the river to artificial lakes. — BH to a double nitromethane adduct. — JSY
Interfering with CFTR-specific chaperone mecha- Earth Planet. Sci. Lett. 10.1016/j.epsl.2006.10.001 J. Am. Chem. Soc. 128, 10.1021/ja066476l (2006).
nisms may thus be a useful strategy to correct (2006). Continued on page 1357

www.sciencemag.org SCIENCE VOL 314 1 DECEMBER 2006 1355

EDITORS’CHOICE
Continued from page 1355

IMMUNOLOGY paracortex, transferring to a similar network of

dendritic cells once they had entered the lymph
Trekking Lymph Node Tracks
node follicle. It will now be interesting to elucidate
Lymph nodes are crucial staging posts from which the molecular cues that govern migration along
immune responses are launched throughout the these cellular highways and byways. — SJS
body. To achieve this, naïve lymphocytes must Immunity 25 10.1016/j.immuni.2006.10.011
locate and respond to their specific antigens, (2006).
which are relatively scarce. The active migratory
tendency of lymphocytes helps to achieve this, PHYSICS
and the structural organization of the lymph node
itself also improves the chances of antigen
Fast Track for Fusion
encounter. Bajenoff et al. now find that organized The search for controlled nuclear fusion for
networks of stromal cells provide trackways for energy production has been hindered by sub-
lymphocytes to travel around lymph nodes. With a stantial engineering and fundamental physical
combination of microscopy and real-time intravi- challenges. One approach has been to confine
tal imaging, T cells were seen to enter the lymph a hot plasma with magnetic fields in a device
node paracortex by interacting with fibroblastic called a Tokamak and then to heat the plasma
until nuclear reactions become self-sustaining.
As the plasma is heated, however, the highest-
velocity ions can drive wave motions and insta-
bilities that disrupt its integrity. Worse yet, the
fast ions can escape with their energy rather
than contributing to the heating process.
Bindslev et al. report a diagnostic technique in
which beams of electromagnetic waves at fre-
quencies of ~110 GHz are scattered off the ions
in the TEXTOR (Tokamak Experiment for Technol-
ogy-Oriented Research) reactor in Germany.
The energy spectrum of the scattered photons
T cell (blue) on the FRC network (red and green). from this collective Thomson scattering process
reveals the velocity distribution of the fast ions.
reticular cells (FRCs). Inside the lymph node, the By acquiring spectra at different times during
FRC formed a three-dimensional network along the heating of the plasma, the authors can
which a large proportion of T cells could crawl. uncover the evolution of the fast ion dynamics.
Antigen-presenting dendritic cells also associated Diagnostic tools such as this are expected to be
with the FRC network, which is consistent with especially important when ITER (the Interna-
the idea that this would optimize the rate of en- tional Thermonuclear Experimental Reactor)
counter between the two types of cell. B cells were commences operation in 2016. — DV
also seen to move along the FRC tracks within the Phys. Rev. Lett. 97, 205005 (2006).

<< Shedding Light on Immunosuppression

CREDITS: BAJENOFF ET AL., IMMUNITY 25 10.1016/J.IMMUNI.2006.10.011 (2006)

Ultraviolet (UV) radiation from sunlight has been implicated in skin

cancer and—perhaps not coincidentally—suppresses the immune
response. UV-dependent immune suppression depends on its absorp-
www.stke.org tion by an epidermal photoreceptor. Trans-urocanic acid (UCA) iso-
merizes to cis-UCA in response to UV exposure, and epidermal UCA
acts as a UV photoreceptor that can mediate immune suppression. The mechanism whereby cis-
UCA affects the immune response, however, has been unclear. Cis-UCA forms a ring-like struc-
ture in solution, and Walterscheid et al., who serendipitously observed that the serotonin recep-
tor antagonist ketanserin blocked UV- and cis-UCA–mediated immune suppression, now find
that cis-UCA can bind to human serotonin receptors. Cis-UCA stimulated calcium mobilization
in cells that express the serotonin receptor, and this calcium mobilization was blocked by
ketanserin. UV- or cis-UCA–induced immune suppression in mice was blocked by antibodies
directed against serotonin (as well as by antibodies directed against cis-UCA) and by serotonin
receptor antagonists. Thus, the ability of cis-UCA to suppress the immune response—and that
of UV radiation—are mediated through the serotonin receptor. — EMA
Proc. Natl. Acad. Sci. U.S.A. 103, 17420 (2006).

www.sciencemag.org SCIENCE VOL 314 1 DECEMBER 2006 1357

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appreciation of science and technology; and strengthen support for TOR Richard Stone +66 2 662 5818 ([email protected]) JAPAN NEWS BUREAU
the science and technology enterprise. Dennis Normile (contributing correspondent, +81 (0) 3 3391 0630, FAX
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See pages 102 and 103 of the 6 January 2006 issue or access dent +91 (0) 11 2271 2896; [email protected])
www.sciencemag.org/feature/contribinfo/home.shtml AFRICA Robert Koenig (contributing correspondent, [email protected])

S ENIOR E DITORIAL B OARD J. M. Claverie, CNRS, Marseille Bernhard Keimer, Max Planck Inst., Stuttgart David S. Schimel, National Center for Atmospheric Research
Jonathan D. Cohen, Princeton Univ. Elizabeth A. Kellog, Univ. of Missouri, St. Louis Georg Schulz, Albert-Ludwigs-Universität
John I. Brauman, Chair, Stanford Univ. Stephen M. Cohen, EMBL Alan B. Krueger, Princeton Univ. Paul Schulze-Lefert, Max Planck Inst., Cologne
Richard Losick, Harvard Univ. Robert H. Crabtree, Yale Univ. Lee Kump, Penn State Terrence J. Sejnowski, The Salk Institute
Robert May, Univ. of Oxford F. Fleming Crim, Univ. of Wisconsin Mitchell A. Lazar, Univ. of Pennsylvania David Sibley, Washington Univ.
Marcia McNutt, Monterey Bay Aquarium Research Inst. William Cumberland, UCLA Virginia Lee, Univ. of Pennsylvania George Somero, Stanford Univ.
Linda Partridge, Univ. College London George Q. Daley, Children’s Hospital, Boston Anthony J. Leggett, Univ. of Illinois, Urbana-Champaign Joan Steitz, Yale Univ.
Vera C. Rubin, Carnegie Institution of Washington Judy DeLoache, Univ. of Virginia Michael J. Lenardo, NIAID, NIH Thomas Stocker, Univ. of Bern
Christopher R. Somerville, Carnegie Institution Edward DeLong, MIT Norman L. Letvin, Beth Israel Deaconess Medical Center Jerome Strauss, Virginia Commonwealth Univ.
George M. Whitesides, Harvard University Robert Desimone, MIT Olle Lindvall, Univ. Hospital, Lund Tomoyuki Takahashi, Univ. of Tokyo
Dennis Discher, Univ. of Pennsylvania Richard Losick, Harvard Univ. Marc Tatar, Brown Univ.
B OARD OF R EVIEWING E DITORS W. Ford Doolittle, Dalhousie Univ. Ke Lu, Chinese Acad. of Sciences Glenn Telling, Univ. of Kentucky
Jennifer A. Doudna, Univ. of California, Berkeley Andrew P. MacKenzie, Univ. of St. Andrews Marc Tessier-Lavigne, Genentech
Julian Downward, Cancer Research UK Raul Madariaga, École Normale Supérieure, Paris Michiel van der Klis, Astronomical Inst. of Amsterdam
Joanna Aizenberg, Bell Labs/Lucent Rick Maizels, Univ. of Edinburgh
R. McNeill Alexander, Leeds Univ. Denis Duboule, Univ. of Geneva Derek van der Kooy, Univ. of Toronto
Michael Malim, King’s College, London Bert Vogelstein, Johns Hopkins
David Altshuler, Broad Institute Christopher Dye, WHO Eve Marder, Brandeis Univ.
Arturo Alvarez-Buylla, Univ. of California, San Francisco Richard Ellis, Cal Tech Christopher A. Walsh, Harvard Medical School
William McGinnis, Univ. of California, San Diego Christopher T. Walsh, Harvard Medical School
Richard Amasino, Univ. of Wisconsin, Madison Gerhard Ertl, Fritz-Haber-Institut, Berlin Virginia Miller, Washington Univ.
Meinrat O. Andreae, Max Planck Inst., Mainz Douglas H. Erwin, Smithsonian Institution Yasushi Miyashita, Univ. of Tokyo Graham Warren, Yale Univ. School of Med.
Kristi S. Anseth, Univ. of Colorado Barry Everitt, Univ. of Cambridge Edvard Moser, Norwegian Univ. of Science and Technology Colin Watts, Univ. of Dundee
Cornelia I. Bargmann, Rockefeller Univ. Paul G. Falkowski, Rutgers Univ. Andrew Murray, Harvard Univ. Julia R. Weertman, Northwestern Univ.
Brenda Bass, Univ. of Utah Ernst Fehr, Univ. of Zurich Naoto Nagaosa, Univ. of Tokyo Daniel M. Wegner, Harvard University
Ray H. Baughman, Univ. of Texas, Dallas Tom Fenchel, Univ. of Copenhagen James Nelson, Stanford Univ. School of Med. Ellen D. Williams, Univ. of Maryland
Stephen J. Benkovic, Pennsylvania St. Univ. Alain Fischer, INSERM Roeland Nolte, Univ. of Nijmegen R. Sanders Williams, Duke University
Michael J. Bevan, Univ. of Washington Jeffrey S. Flier, Harvard Medical School Helga Nowotny, European Research Advisory Board Ian A. Wilson, The Scripps Res. Inst.
Ton Bisseling, Wageningen Univ. Chris D. Frith, Univ. College London Eric N. Olson, Univ. of Texas, SW Jerry Workman, Stowers Inst. for Medical Research
Mina Bissell, Lawrence Berkeley National Lab R. Gadagkar, Indian Inst. of Science Erin O’Shea, Harvard Univ. John R. Yates III, The Scripps Res. Inst.
Peer Bork, EMBL John Gearhart, Johns Hopkins Univ. Elinor Ostrom, Indiana Univ. Martin Zatz, NIMH, NIH
Dianna Bowles, Univ. of York Jennifer M. Graves, Australian National Univ. Jonathan T. Overpeck, Univ. of Arizona
Robert W. Boyd, Univ. of Rochester Christian Haass, Ludwig Maximilians Univ. Walter Zieglgänsberger, Max Planck Inst., Munich
John Pendry, Imperial College Huda Zoghbi, Baylor College of Medicine
Dennis Bray, Univ. of Cambridge Dennis L. Hartmann, Univ. of Washington Philippe Poulin, CNRS
Stephen Buratowski, Harvard Medical School Chris Hawkesworth, Univ. of Bristol Maria Zuber, MIT
Mary Power, Univ. of California, Berkeley
Jillian M. Buriak, Univ. of Alberta Martin Heimann, Max Planck Inst., Jena David J. Read, Univ. of Sheffield
Joseph A. Burns, Cornell Univ. James A. Hendler, Univ. of Maryland Les Real, Emory Univ. B OOK R EVIEW B OARD
William P. Butz, Population Reference Bureau Ove Hoegh-Guldberg, Univ. of Queensland Colin Renfrew, Univ. of Cambridge
Doreen Cantrell, Univ. of Dundee Ary A. Hoffmann, La Trobe Univ. John Aldrich, Duke Univ.
Trevor Robbins, Univ. of Cambridge
Peter Carmeliet, Univ. of Leuven, VIB Evelyn L. Hu, Univ. of California, SB Barbara A. Romanowicz, Univ. of California, Berkeley David Bloom, Harvard Univ.
Gerbrand Ceder, MIT Olli Ikkala, Helsinki Univ. of Technology Nancy Ross, Virginia Tech Londa Schiebinger, Stanford Univ.
Mildred Cho, Stanford Univ. Meyer B. Jackson, Univ. of Wisconsin Med. School Edward M. Rubin, Lawrence Berkeley National Lab Richard Shweder, Univ. of Chicago
David Clapham, Children’s Hospital, Boston Stephen Jackson, Univ. of Cambridge Gary Ruvkun, Mass. General Hospital Ed Wasserman, DuPont
David Clary, Oxford University Daniel Kahne, Harvard Univ. J. Roy Sambles, Univ. of Exeter Lewis Wolpert, Univ. College, London

1360 1 DECEMBER 2006 VOL 314 SCIENCE www.sciencemag.org

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Keeping Book on Baby

Baby books, those little pamphlets that record baby’s first steps
and first words, now interest more than doting relatives. Scholars
ANOTHER PHYSICS DEPARTMENT DOWN are finding the books a new source of close-up information on
Reading University last week became the 21st British university to announce the the early lives of infants from different times.
closure of its physics department since 1997. Despite protests from staff and stu- Last month, the University of California, Los Angeles (UCLA),
dents and a petition from more than 2000 researchers around the world, the uni- Louise M. Darling Biomedical Library held a reception to intro-
versity council voted on 20 November to accept no more physics students. The duce its collection of
department will close in 2010. more than 500 baby
U.K. universities are largely government-funded, with the amounts deter- books dating back to
mined by numbers of students and quality of research. Reading Vice-Chancellor 1884. It’s a historical
Gordon Marshall said in an open letter that the physics department is losing treasure trove that
about $1 million a year because it is not getting enough new students—28 this charts shifting atti-
year against a target of 50—or enough research income. tudes about public
The closure of science and math departments (Science, 4 February 2005, health and parent-
p. 668) prompted the U.K. government last month to announce $140 million to hood, says archivist 1930
help key departments over the next 3 years. But it won’t be enough to help Russell Johnson. “A baby
Reading, Marshall says. Philip Diamond of the Institute of Physics in London says space for the father book
economics favors big departments these days; a half-dozen now account for half to make entries
of all U.K. physics students, and “small ones are just vulnerable.” doesn’t show up
until around World
War II,” he notes.
The books may
WELL-WIRED WHALES >> have emerged as part
of a late–19th cen-
CREDITS (TOP TO BOTTOM): EVENING POST PHOTOGRAPHERS; LOUISE M. DARLING BIOMEDICAL LIBRARY/UCLA; ROYALTY-FREE/CORBIS

Some whales have a tury public health

specialized brain cell campaign to “Save the Babies,” according to Jacqueline H. Wolf,
that hitherto has been a medical historian at Ohio University in Athens. “Baby books
seen only in humans represented a change in cultural thinking,” she says. “Infants
and great apes—lead- were not weak and susceptible, as people had long argued.
ing some scientists to Rather, infant death was preventable.”
suggest that cetaceans Russell says UCLA welcomes donations from all eras, espe-
evolved their relatively Humpback cially because the library often battles collectors of famous
advanced brains before whale. children’s book illustrators when copies come up on eBay.
primates did.
Humans, chimps, and gorillas share a type of cortical nerve cell—called a
spindle neuron—that is lacking in all other primates. The cells appear to con-
nect regions implicated in higher cognitive functions to other parts of the brain.

NETWATCH >>
Neuroscientist Patrick Hof and neuroendocrinologist Estel Van der Gucht of
Mount Sinai School of Medicine in New York City have now discovered spindle
neurons, in areas homologous to their location in human brains, in several large-
brained cetaceans including humpback and fin whales. The researchers estimate
Diseases on the Move
that bigger-brained whales evolved spindle neurons 22 million to 30 million Visitors to the new site HEALTHmap can pinpoint the latest out-
years ago. Because the common ancestor of great apes only dates to about 15 mil- breaks of more than 50 human and animal illnesses, from
lion years ago, the pair concludes that these cells must have evolved independ- avian influenza to chikungunya fever, a mosquito-spread dis-
ently in apes and whales. Reporting online this week in The Anatomical Record, ease of Asia and Africa. Created by epidemiologist John
they speculate that whale talents such as the formation of social groups as well as Brownstein of Harvard Medical School in Boston and software
singing and other communicative skills are linked to the enhanced connectivity developer Clark Freifeld of Children’s Hospital Boston, the site
provided by spindle neurons. automatically picks up and charts fresh case reports and other
Clever but smaller-brained dolphins don’t have spindle neurons. John Allman, data from sources such as the World Health Organization,
a neurobiologist at the California Institute of Technology in Pasadena, says the Google News, and the disease alert Web site ProMed-Mail. You
neurons are probably “adaptations that support fast communication … in very can sort the information by disease and country and click on
large brains.” Allman says his group is looking to see whether elephants also the world map to summon the original report or article. >>
have the cells. www.healthmap.org

www.sciencemag.org SCIENCE VOL 314 1 DECEMBER 2006 1363

NEWSMAKERS
EDITED BY YUDHIJIT BHATTACHARJEE

Up Next THE FIRST PODSTER. There you are, standing by

your poster at the big annual meeting, when the
Big Kahuna in your field walks up. If only you had some multime-
dia to make a quick impression. Next time, try attaching some video
iPods to your poster, says graduate student Pascal Wallisch of the
University of Chicago in Illinois, who unveiled what may be the
world’s first “Podster” at October’s meeting of the Society for
Neuroscience in Atlanta, Georgia.
Wallisch got the idea after a poster session at the 2005 neuro-
science meeting in which he used his laptop to show videos of his
research on the primate visual system. Reaching around to point at
the screen was awkward, however, and the batteries ran out. So this
year, he replaced the laptop with two iPods, each loaded with
videos to explain different features of his experiment. It gave visi-
tors a more interactive experience, he says, and left his hands free.
Passers-by loved the concept, says Wallisch, as did two Apple
Computer reps from a nearby booth—until they found out he’d used
a Microsoft product to create the poster’s text and graphics. “Then
they just left,” Wallisch says.

ON CAMPUS sity of having political motives and claiming Slowing the buildup of
CLEVERNESS CONTROVERSY. A Danish that the errors in his research were trivial. greenhouse gases, he
IQ researcher who was suspended after his said, is the “world’s
research suggested that men have slightly POLITICS most pressing chal-
higher IQs than women has been found U.K. SCIENCE MINISTER. An educator turned lenge.” Wicks has
guilty of “official misconduct” but reinstated politician is the new U.K. minister for science called the United
in his job. and innovation. Malcolm Wicks, 59, succeeds Kingdom’s and the
Psychologist Helmuth Nyborg, 69, of the David Sainsbury, who stepped down last month world’s failure to
University of Aarhus was suspended last after 8 years marked by a doubling of U.K. address nuclear waste
spring following criticism of a report from a spending on research. “an absolute disgrace,”
CREDITS (TOP TO BOTTOM): BEN SCOTT; COURTESY OF MALCOLM WICKS; KLAUS GOTTFREDSON; PETER SHENDEROV

longitudinal study called the Skanderborg The son of a Labour Member of Parliament although he has also
project. Nyborg reported in a June 2005 paper and a Labour MP himself since 1992, Wicks said that nuclear power could be a clean
in the Journal of Personality and Individual most recently guided a strategic plan for a dra- alternative to fossil fuels. Wicks attended the
Differences that when IQ test scores of matic shift to low-CO2-emitting power sources London School of Economics but has no formal
62 Danes he has been as energy minister in the Blair government. training in science.
following since the
1970s are properly
analyzed, they reveal
a roughly four-point
Rising Stars >>
advantage for males. IN TANDEM. Sometimes a little sibling rivalry
Although Nyborg is can be a good thing. Last week, Kevin
not alone in reporting Shenderov, a 19-year-old senior at New York
such a sex difference, University, followed the footsteps of his brother
some scholars criti- Eugene by winning a Rhodes Scholarship.
cized the research on Kevin (left) intends to pursue a doctorate in
methodological immunology at Oxford University—just as
grounds. University officials set up a committee Eugene (center) is now doing.
to investigate, and in July, it reported that there The brothers credit their parents, Peter, a
was no evidence of fraud. In September, the medical physicist, and Faina, who is complet-
university declared that Nyborg had demon- ing a doctorate in pharmacy, for inculcating a love of science. Both worked at Memorial
strated “grossly negligent behavior” and issued Sloan-Kettering Cancer Center in New York City when the boys were growing up, and “the
him a “severe reprimand” before revoking his dinner conversation pretty much always centered around what was going on at the hospital,”
suspension. Colleagues from around the world says Eugene, 23, who won a Rhodes 2 years ago. “Science was the family’s bread and butter.”
have rallied to his defense, accusing the univer- Peter says the brothers have pushed each other but remain close. “This kind of competition is
pulling them together,” he says. “The accomplishment is the icing on the cake.”
Got a tip for this page? E-mail [email protected]

www.sciencemag.org SCIENCE VOL 314 1 DECEMBER 2006 1365

NEWS>>
THIS WEEK Progesterone and
breast cancer
A new way to
engineer flies

1370 1371
SCIENTIFIC CONDUCT ence: The Web postings are individual
actions not directed by the state. The Chi-
China’s Fraud Buster Hit by Libel nese government takes an ambiguous
stance: It blocks access in China to New
Threads’ U.S.–based site, www.xys.org,
Judgments; Defenders Rally Round but allows access to mirror sites.
Fang’s recent setbacks came on consecu-
BEIJING— China’s self-appointed science dubious claims in the Chinese media about tive days. On 21 November, a Beijing inter-
cop, Fang Shi-min, was dealt a pair of set- “nucleotide supplements.” Fang then mediate court ruled that an article Fang
backs last week in his high-profile crusade started using his Web site, Xin Yu Si (“New wrote in 2005 defamed the late Liu Zihua, a
against academic misconduct. Two Chinese Threads”), to debunk pseudoscience Sichuan provincial government employee.
courts handed down libel judgments a n d expose alleged misconduct, from In a dissertation written in France in the
against Fang, known by his nom de guerre résumé padding to data fabrication (Science, 1930s, Liu presented calculations based on
Fang Zhouzi, and the newspapers and Inter- 10 August 2001, p. 1039). the eight trigrams of an ancient divination
net sites that have featured his writings on By Fang’s tally, New Threads has aired text, I Ching (Book of Changes), predicting
the existence of a 10th major planet in the
solar system. Liu’s prognostication was res-
urrected after last year’s announced discov-
Back to the wall. Libel ery of 2003UB313 (now officially a dwarf
judgments have cast a pall planet named Eris). A Sichuan newspaper
over Fang Zhouzi’s fraud ran a story extolling Liu’s prophecy.
fighting. In an essay, Fang labeled Liu’s prediction
“pseudoscience” and noted that a Chinese
astronomer discredited it in the 1940s. Liu’s
widow and son sued Fang and several news-
papers and Internet content providers for
libel. The court judged Fang’s words “insult-
ing” to Liu and ordered him to apologize
publicly and pay Liu’s family $2500 plus
legal fees. The family did not respond to an
interview request.
Then on 22 November, a court in Xi’an
slapped another libel judgment on Fang,
ordering him and Beijing Keji Bao (Beijing
Sci-Tech Report) to pay Xi’an Fanyi Univer-
sity $18,750 and its president Ding Zuyi
$1250 in damages plus legal fees. In 2004,
Chinese newspapers ran stories citing a
pseudoscience and fraud. Fang’s revela- allegations against more than 500 individu- “report” in the Los Angeles Times lauding
tions have cost several scientists their jobs als. Fang uncovered some cases himself, but Ding as one of China’s most respected univer-
and reputations. most were e-mailed to him by others. Few sity presidents and his private college for
With Fang now on the defensive, his exposures have led to official investiga- training translators as the 10th-ranked univer-
backers are setting up two funds to help foot tions, and fewer still have resulted in pun- sity in China. In a 2005 article in Beijing
the costs of litigation. “If you strike false ishment—the most notable being the dis- Sci-Tech Report, Fang quoted an education
science, false science [makers] will strike missals earlier this year of an assistant dean ministry spokesperson, who stated that inves-
you,” says Guo Zhengyi, a science writer of Qinghua University’s medical school in tigations showed the report to be “a self-paid
and a co-organizer of one foundation. Guo Beijing and a dean at Tongji University in advertisement.” Ding sued Fang for libel.
and others say they hope that, by drawing Shanghai, both for having falsified their Ding could not be reached for comment.
attention to what they call “absurd” court résumés and exaggerated achievements. Fang is appealing another libel verdict
CREDIT: R. STONE/SCIENCE

rulings, they may force the government to The anonymous allegations published by a Wuhan court last July. In this case,
crack down on corruption. on New Threads trouble some people, Xiao Chuan-guo, a urology professor at
Fang received a Ph.D. in biochemistry who liken them to dazibao, or posters, Huazhong University of Science and Tech-
and did a postdoc in the United States used during the Cultural Revolution to nology in Wuhan and a clinical associate
before becoming a science essayist. He got denounce “class enemies.” Fang and his professor at New York University School of
fired up about fraud in 2001, after reading supporters contend there’s a big differ- Medicine, sued after Fang accused him in an

1366 1 DECEMBER 2006 VOL 314 SCIENCE www.sciencemag.org

FOCUS Pruning science in
the classroom
South Africa’s
turnaround on HIV
Saving the
saola

1374 1378 1380

essay last year of counting conference concluded that Fang had begun to “misguide some individual cases or organize seminars
abstracts as publications in international the public” with less-than-solid accusations. to discuss legal punishments against proved
journals to inflate his achievements. Fang In response to the Wuhan ruling, Zhang misconduct makers.”
also challenged Xiao’s claim that a surgical Feng, a Florida-based financial analyst and Fang vows to continue “using sharp-
procedure he invented is recognized inter- college classmate of Fang’s, along with eight tongued criticism” to expose misconduct
nationally and has won neurourology’s other expatriates, last month established the and folly. But he doubts that his freelance
“highest award.” The presiding judge ruled Organization for Scientific and Academic fraud busting can play a “decisive role” in
that Fang’s criticisms “seriously lacked Integrity in China to raise money for Fang cleaning up Chinese academia. To be more
facts” and ordered him to apologize publicly and other anticorruption campaigners. So effective, he says, he intends to report future
and pay Xiao $3750 in compensation. A far, the nonprofit has received more than allegations, when appropriate, to a new dis-
final ruling is expected in early December. $10,000 in donations. And in China, Guo ciplinary office at China’s Ministry of Sci-
Xiao told Science that the accusations are and others are creating a separate science ence and Technology and wait for a response
groundless and that Fang “intentionally con- fraud-fighting fund. Fang’s lawyer, Peng before posting them online.
fused” Xiao’s urology awards. Xiao says he Jian, hopes the foundations will raise money –JIA HEPENG AND HAO XIN
supported Fang until 2002, after which he to “implement systematic investigations into Jia Hepeng is a writer in Beijing.

DEVELOPMENTAL BIOLOGY

Fraud Investigation Clouds Paper on Early Cell Fate

A surprising report in a contentious area of embryos are teased apart and cells trans- ally went on to give rise to the placental tis-
developmental biology has sparked a scien- planted from one embryo to another have sues. The other, which had less Cdx2 expres-
tific misconduct investigation at the Univer- suggested that until about a week after fer- sion, went on to form the eventual fetus. The
sity of Missouri, Columbia. Until that tilization, mammalian embryo cells are scientists argued that their observations might
inquiry is complete, the results of the impli- quite interchangeable. There is an ongoing help explain why cloning in mammals is so
cated paper, published in Science earlier this debate, however, over whether very early inefficient. If Cdx2 expression is disrupted by
year, remain in limbo. embryo cells—when the embryo is at the cloning, they speculated, then embryos might
Contrary to prevailing dogma, the report four- or eight-cell stage—might have a ten- have a hard time developing further.
claimed that mouse embryo cells have dis- dency toward one fate or another, although Proponents of so-called alternative
tinct fates from the time of the very first they are not yet committed. The results nuclear transfer were also excited by the
cell division. If true, those findings would published in February indicated a much results. This technique has gained some sup-
dramatically change the current under- earlier differentiation than anyone else in port among people otherwise opposed to
standing of mammalian embryo develop-
ment—and could also play a role in on-
going political and ethical debates over
cloning and stem cells.
But the senior author of the paper now
says the results are not trustworthy and pre-
dicts that the paper will be retracted as soon
as the university completes its investiga-
tion. The paper, which was accepted as Surprisingly clear. The paper reported finding the Cdx2 protein (green) concentrated in just one of the cells
Science editors were embroiled in the scan- in two-cell mouse embryos.
dal surrounding Woo Suk Hwang’s human
cloning papers in early 2006 (see Editorial the field had suggested. stem cell research and human nuclear trans-
on p. 1353), again raises questions about The corresponding author, R. Michael fer because it hypothetically offers a way to
the limitations of the peer-review process in Roberts, is an expert in bovine embryology derive stem cells without destroying an
detecting fraud. and until this year had not been involved in embryo. If scientists use egg cells or somatic
CREDIT: R. M. ROBERTS ET AL. (2006)

Published in the 17 February issue of the debate. In the paper, Roberts, with post- cells lacking Cdx2 for their nuclear transfer,
Science (p. 992), the paper caused an doctoral fellows Kaushik Deb and Hwan Yul the reasoning goes, the resulting cells would
immediate stir. It is well known that embry- Yong and microscope technician Mayandi be unable to form a true embryo.
onic cells of insects and amphibians have Sivaguru, claimed that in most mouse The Roberts results seemed to help the
distinct fates from the first cell divisions, embryos there was a distinct difference supporters’ case: If the gene is so crucial
but the picture for mammalian embryos has between cells from the first cell division on. from the very beginning, then that would
been far murkier (Science, 6 May 2005, One cell had strong expression of a gene strengthen the argument that cells lacking it
p. 782). Experiments in which mouse called Cdx2, the paper claimed, and eventu- could not be called an embryo.
▲

www.sciencemag.org SCIENCE VOL 314 1 DECEMBER 2006 1367

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SCIENCESCOPE
The results, however, “were so drastically that if they had, they likely would have
different from any of the results obtained by caught the problems. But Richard Behringer Party Animals
any other group” that most people viewed them of M. D. Anderson Cancer Center in Hous- AMSTERDAM—The Dutch Party for Animals
skeptically from the start, says Magdalena ton, Texas, is not sure the problems could gained two seats in the 150-member Second
Zernicka-Goetz of the University of Cam- have been spotted ahead of time. Although Chamber of Parliament last week after draw-
bridge, U.K. However, it wasn’t immediately Behringer now says he can see the evidence ing 1.9% of the votes nationwide. The group,
clear why the results were so different, she of duplicated images, he says at first read- one of whose goals is the eventual elimination
says. A different strain of mice or a different ing the paper seemed solid, if surprising. of animal experimentation, appears to be
labeling technique might have been the “I can understand why referees would the first political party devoted to animal
cause, she says. say OK,” he says. welfare. Its platform includes a ban on trans-
Others in the field were less willing to Kelner and Science Editor-in-Chief genic animals, better oversight of animal
suspend their disbelief. Within weeks of the Donald Kennedy add that even if the data are experiments, including better housing and
paper’s publication, Roberts says, several found to have been manipulated, the new daily checks by independent vets, and more
scientists wrote to Science, to Roberts, and image-analysis techniques would not have research on alternatives.
to the University of Missouri, pointing out picked it up. Those techniques can flag –MARTIN ENSERINK
problems with the data. Some of the images unmatched pixels that are signs of deletions
seemed suspiciously similar to each other, or cut-and-paste manipulations. But dupli- Dawkins Versus the Gods
they said. In others, the staining didn’t seem cated images—like those in the Hwang After scanning the titles in a local bookshop,
to line up exactly with the cells. By late paper—are harder to spot, Kelner says. Oxford University geneticist Richard Dawkins
April, Roberts says, the university had In retrospect, Roberts says he wishes he discovered that “real science” was “out-
started an investigation. had been more cautious with the results his numbered three to one by pseudoscience.”
It was soon clear that there Concerned that “the enlightenment is under
was reason to worry about the threat,” the author of The God Delusion has
data’s veracity, Roberts says. “In created and will help fund the Richard Dawkins
my view, there are a number of Foundation for Science and Reason. The new
questionable images,” he says. charity, with U.S. and U.K. branches, will sup-
But until the university investi- port research on “the psychological basis of
gation is complete, he says, the unreason,” produce videos and books, and run
team will not be able to explain a Web site (richarddawkins.net/foundation).
the details of what is wrong or Another goal, “to oppose … well-financed
retract the paper. Roberts says efforts to teach creationism in science classes,”
the university is being very cau- will put it up against the U.K.–based Truth in
tious about assigning any blame Science, which recently sent “intelligent
before the investigation is com- design” promotional packs to 5700 British sec-
plete. All the co-authors have ondary schools. Truth in Science claims it
since left the university. Two received 59 positive responses.
have found other jobs, and a third –ELIOT MARSHALL
has apparently dropped out of
contact. In the meantime, Science Hope for German GM crops
issued an “Editorial Expression BERLIN—In a move to support plant
of Concern” to alert the commu- researchers, the German agriculture minister
nity that it should not trust the has apparently agreed to ease rules controlling
p u bl i s h e d r e s u l t s ( S c i ence, Senior author. R. Michael Roberts says the paper will likely be the planting of genetically modified (GM)
27 October, p. 592). retracted as soon as the University of Missouri finishes its investigation. crops. German media reported last week that
Some critics question why the the minister, Horst Seehofer, will propose
paper was published in the first place or why lab members presented to him. “I didn’t go allowing the government to pay for damages
CREDIT: NORMA MCCORMACK/UNIVERSITY OF MISSOURI, COLUMBIA

image-analysis techniques—which Science into this with preconceived ideas. I got into it resulting from any gene-altered pollen that
editors said they put in place at the beginning by happenstance,” he says. The research was escapes from government-funded research
of the year—didn’t spot the apparent aimed at determining whether Cdx2 was plots. Under current rules, the farmers or
problems. Davor Solter of the Max Planck involved in turning on another gene in researchers who plant GM seeds are liable for
Institute for Immunobiology in Freiburg, bovine embryos, he explains, and the mouse any pollen that might contaminate a neigh-
Germany, one of the scientists who wrote to embryos were used as controls to analyze bor’s field, preventing it from being sold as
Science, contends that the review process Cdx2 expression. “But the results looked so GM-free. The proposal, contained in a measure
was flawed. Science editors declined to dis- beautiful, you couldn’t come to any other that could be presented to legislators early
cuss the specifics of the review process, conclusion.” Since questions about the paper next year, would also restrict public access to
which is confidential, but Katrina Kelner, were raised, he says, “I’ve obviously ques- information about where GM crops are
deputy editor for biology, says, “Science tioned myself and my judgment. I haven’t planted. Despite overwhelming public opposi-
published the paper based on feedback we had a good night’s sleep since February.” tion to GM foods, research minister Annette
got from the field.” The University of Missouri is expected to Schavan has been pushing for such rules.
Solter speculates that leading scientists finish its investigation later this month. –GRETCHEN VOGEL
in the field did not review the paper, noting –GRETCHEN VOGEL

www.sciencemag.org SCIENCE VOL 314 1 DECEMBER 2006 1369

NEWS OF THE WEEK

MEDICINE

Squelching Progesterone’s Signal May Prevent Breast Cancer

A woman who carries a mutated BRCA1 increased cell proliferation—much as the mifepristone, however, were still tumor-
gene faces a daunting decision: She can opt breasts of pregnant woman do when high free at 12 months of age.
for constant monitoring hoping to catch progesterone levels prepare the mammary Lee and her colleagues then took a closer
any cancer early, while it’s still curable, or glands for lactation. What’s more, all the look at the epithelial cells that give rise to
she can elect to have her breasts or ovaries rodents developed mammary cancers by breast cancer. “A lot more cells” from the
removed to prevent cancer from develop- the age of 8 months. Mice treated with double-mutant mice had progesterone
ing in the first place. Results described on receptors, she says, than did cells from nor-
page 1467 now suggest that one day there mal animals or from animals in which only
may be a third option: using drugs rather the p53 gene had been knocked out.
than surgery to prevent BRCA1-mediated Further work on cultured mouse and
breast cancers. human cells revealed that the progesterone
BRCA1 is a so-called tumor suppressor, receptor is broken down less readily when
a gene that in its normal form protects BRCA1 activity is missing. As a result,
against cancer. One way it does this is by “the [hormone’s] signal goes on much
helping cells repair DNA damage that longer,” Lee says. The excessive cell
might otherwise result in cancer-causing g rowth this produces provides extra
mutations. The new work, which comes chances for cancer-promoting DNA muta-
from Eva Lee and her colleagues at the Uni- tions to occur, especially because BRCA1
versity of California, Irvine, points to loss also handicaps the cell’s DNA repair
another cancer-preventing role for BRCA1. machinery. The participation of the proges-
By aiding in the degradation of the receptor terone receptor in BRCA1-mediated breast
through which progesterone exerts its cancer could help explain why tumors
effects, the gene’s protein product appar- occur specifically in the breast and ovaries
ently checks the hormone’s growth-pro- even though the gene is mutated in cells
moting action on breast tissue. throughout the body. Those other cells
Lee’s team also showed that mifepris- don’t carry progesterone receptors.
tone, a drug that induces abortions by Lee points out that mifepristone itself
inhibiting the progesterone receptor, blocks may not be suitable for long-term use in
the development of mammary tumors in cancer prevention because it acts on steroid
mice that have had the rodent version of receptors besides the one for progesterone.
BRCA1 inactivated in their mammary It might therefore cause unacceptable side
glands. “The paper has a mechanism [of effects such as immune suppression. Other
BRCA1 activity] and has clinical implica- more specific progesterone blockers are
tions. It’s potentially important,” says Eliot under development, Lee notes.
Rosen of Georgetown University School of There is uncertainty about how accu-
Medicine in Washington, D.C., who is also rately the new mouse model reflects
studying the interaction between BRCA1 human breast cancer. Lee cites findings by
and progesterone. Jeff Boyd’s team at Memorial Sloan-
Previous work had raised suspicions Kettering Cancer Center in New York City
that progesterone fosters breast cancer that tissue adjacent to human breast tumors
development. For example, women tak- with BRCA1 mutations shows elevated
ing both estrogen and progesterone to progesterone expression compared to
treat menopausal symptoms have a higher t i s sue from nor mal breast. However,
risk of developing breast cancer than Christine Clarke and her colleagues at the
women who take estrogen only. And University of Sydney at Westmead Millen-
working with human breast cancer cells nium Institute in Westmead, Australia,
in lab cultures, Rosen’s team found that actually saw a decrease in progesterone
normal BRCA1 inhibits the action of the receptors in tissue removed by mastectomy
progesterone receptor, although how has from BRCA1 carriers.
CREDIT: A. J. POOLE ET AL./ SCIENCE

been unclear. The two situations aren’t quite compa-

In the current work, Lee and her col- rable. “The status of tissue around tumors
leagues created mice that lacked function- is different from that of tissue taken from
Releasing the brakes. The ducts from mouse
ing copies of the rodent versions of both mammary tissue in which both the p53 and BRCA1
normal breast,” Clarke says. But that
BRCA1 and p53, another tumor suppressor genes have been inactivated (bottom) show issue, and likely many others, needs to be
that is frequently mutated in breast cancers. increased growth and branching compared to ducts resolved before cancer prevention trials of
Although the female mice had never been from either normal mice (top) or animals in which progesterone inhibitors can begin.
mated, their mammary tissue showed only p53 is inactive (middle). –JEAN MARX

1370 1 DECEMBER 2006 VOL 314 SCIENCE www.sciencemag.org

SCIENCESCOPE
MOLECULAR BIOLOGY
EPA, Berkeley Think Small
Three Methods Add Up to One New The U.S. Environmental Protection Agency
(EPA) has decided to regulate silver ions, the
Way to Genetically Engineer Fruit Flies bacteria-killing nanoparticles used in prod-
ucts as diverse as shoe liners and food storage
When Koen Venken began a Ph.D. project on colleagues spliced a stretch of DNA into a containers. The agency will require Samsung,
fruit fly genetics at Baylor College of Medi- portion of a P element and found that the new which sells an ion-emitting device in a wash-
cine in Houston, Texas, 4 years ago, he quickly DNA was easily incorporated into the fly’s ing machine, to spell out possible environ-
became frustrated by limitations of the stan- genome. The P element, however, can’t inte- mental impacts under rules that apply to pes-
dard techniques for genetically engineering grate long stretches of DNA like the ones ticides, even though the agency does not yet
the insects. So he turned his attention to devel- Venken wanted to work with when he joined know whether the device involves nanomateri-
oping a novel procedure. The result, described the Baylor lab of geneticist and HHMI inves- als. “The fact that EPA seems to be addressing
in a paper published online by Science this tigator Hugo Bellen. this is a good thing,” says physicist Andrew
So Venken took components of the Maynard of the Woodrow Wilson International
P element, including ones that allow it to Center for Scholars in Washington, D.C. The
integrate DNA into the fly genome, and upcoming federal notice could clarify whether
added them to loops of bacterial DNA the decision heralds broader federal limits on
called plasmids. These bacterial artificial nanotechnology.
chromosomes (BACs) can more stably Meanwhile, scientists in Berkeley, Cali-
retain larger amounts of foreign DNA than fornia, say their work will not be affected by a
a P element alone. To add into those BACs proposed city rule, the first of its kind in the
the DNA he wanted to insert into the flies, United States, that would require the registra-
Venken next turned to a technique called tion of nanoparticles. The council is expected
recombineering, which was developed to discuss the measure next week.
about 8 years ago. Recombineering –ELI KINTISCH
involves allowing a BAC to recombine with
an intended transgene within bacteria, iso- South Korean Flu Mystery
lating that BAC, and then using other bacte- An outbreak of the H5N1 strain of bird flu in
ria to make multiple copies of it. South Korea may reignite debate over how the
Finally, Venken used a third existing disease is spread. Researchers had long
technique to control where in the fly argued about whether the H5N1 strain, which
genome the BAC-ferried gene would land. has killed 258 humans since it started sweep-
When he injected the BACs into fruit fly ing through Asia in 2003, is spread by wild
BAC-ed up. A new method allows researchers to embryos, Venken also injected messenger birds or the movement of infected poultry and
insert lots of DNA into fruit flies. RNA that encodes an enyzme made by a contaminated crates and vehicles.
bacterial virus called a phage. This enzyme Last week, the South Korean government
week (www.sciencemag.org/cgi/content/ normally inserts a phage’s DNA into spe- confirmed H5N1 as the culprit behind the
abstract/1134426), appears to be a powerful cif ic sites on a bacterial genome, but in deaths of 6000 chickens on a farm in Iksan,
new way of making transgenic flies, one that these circumstances, it integrates the BAC- south of Seoul, the first known H5N1 outbreak
will likely make it easier to study fruit fly carried gene at similar DNA sequences in Korea since 2003. The route of infection in
genes that were previously too large to work engineered into the fly genome. that incident has never been proven, although
with and to compare the behavior of similar Using this combination of methods, South Korea’s Ministry of Agriculture & Forestry
genes belonging to different fly species. Venken, Bellen, and their colleagues have has since warned on its Web site about the
The work eases two roadblocks that have inserted DNA stretches as long as 133,000 potential for H5N1 transmission from migratory
long troubled the fly community: inserting bases into the fly genome. “It’s now not clear birds to domestic chickens to people.
genes longer than about 20,000 DNA what the upper limit is,” says Daniel Barbash, Casting the blame on migratory fowl
bases—which make up more than 5% of the a geneticist at Cornell University. Bellen’s “could lead to the vilification of wild birds”
insect’s genes—and controlling where in the lab is now assembling a library of fly DNA in and attempts to slaughter them or disturb
genome those genes land, which impacts how the novel BACs for interested researchers. their habitat, warns ornithologist Nial Moores
they get expressed. “The real advantage here The new approach “is a significant tech- of Birds Korea, a conservation group. “The real
is that it’s a way of putting in really big bits of nical advance,” says Rubin. It “allows us to danger comes from poultry infecting wild birds
DNA” into the fly, says Michael Ashburner, a do certain things we couldn’t do before,” and not the other way around,” Moores says.
fly geneticist at the University of Cambridge, such as study the effects of whole gene com- He’s particularly worried about major winter-
United Kingdom. plexes, like the homeotic genes that affect ing grounds at the mouth of the Geum River,
Traditionally, geneticists create trans- early development. And several scientists 5 kilometers from Iksan, currently home to the
CREDIT: GETTY IMAGES

genic flies with help from a piece of fly DNA note that this blended approach might work world’s largest concentration of Baikal teal. “If
called the P element. Roughly 2 decades ago to genetically modify other organisms. “The avian influenza gets transmitted into this flock,
Gerald Rubin, now director of the Howard potential to export this system to … other it could be devastating,” Moores says.
Hughes Medical Institute’s (HHMI’s) Janelia animals is quite high,” says Barbash. –DENNIS NORMILE
Farm in Loudoun County, Virginia, and his –JENNIFER COUZIN

www.sciencemag.org SCIENCE VOL 314 1 DECEMBER 2006 1371

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NEWS OF THE WEEK

DRUG RESEARCH offers a different idea: The world should

create markets where they currently don’t
exist. For instance, companies could be
WHO Panel Weighs Radical Ideas enticed with research g rants from a
“Global Tropical Disease Fund” or the
Lifesaving antiretroviral drugs have been may consider, is a hotly debated proposal promise of guaranteed sales should they
available for a decade in wealthy countries, for an international treaty to open up drug develop an effective new drug.
yet millions of HIV-infected people south of discovery, championed since 2002 by James The industry also contributes through a
the equator still can’t get them. The medicine Love, director of the Consumer Project on model called the public-private partnership
cupboard is equally bare for people afflicted Technology in Washington, D.C. Under (PPP). Over the past 10 years, more than
by tropical illnesses such as visceral leishma- Love’s “R&D Treaty,” countries would two dozen PPPs have sprung up to tackle
niasis, sleeping sickness, and Chagas agree to spend a minimum percentage of diseases of the poor. Enlisting industry,
disease, for which there are no truly good gross domestic product on medical academia, governments, and foundations,
therapies. Western medical sci- these partnerships, such as the TB Alliance
ence has not done well by the and the Medicines for Malaria Venture
world’s poor, and some critics
World Pharmaceutical Market by Region (MMV), have produced many new candi-
blame this on its reverence for $ billions (2005) date drugs (Science, 13 January, p. 167).
intellectual property (IP). Is it N. America (268.8) And the IP protection regime has not been
time to overhaul the IP protec- Europe (180.4) an obstacle, says MMV president Chris
tion system? A new working Japan (69.3) Hentschel: “If people spent less time think-
group hosted by the World Health Oceania (7.7) ing about IP and more about other things,
Former USSR (5.0)
Organization (WHO) will con- we would make more progress.”
Southeast Asia (28.8)
sider that question in a series of Latin America (26.6) But others point out that health PPPs have
meetings beginning next week in India (6.7) a narrow base: 60% of their funding comes
Geneva, Switzerland. Africa (6.7) from a single source, the Bill and Melinda
Critics of the current IP pro- Middle East (4.9) Gates Foundation; governments con-
tection system hope that WHO’s tribute very little. Moreover, industry
Intergovernmental Working tends to help PPPs that work on dis-
Group (IGWG) on Public No profit? A minuscule pharmaceuti- eases that affect both the poor and
Health, Innovation and Intellec- cal market in developing countries people from rich countries, such as
tual Property will reform—or limits R&D on drugs against try- malaria and TB, says Els Torreele,
even hack down—the still- panosomes, which cause African project manager at the Drugs for
expanding worldwide patent sleeping sickness and Chagas disease. Neglected Diseases Initiative. Given
system. They say it puts life- the scope of the problem, something
saving new drugs beyond the reach of poor research, including more radical is needed, she says.
patients and hampers development of new a portion for neg- Whether the IGWG can deliver a
medicines for tropical diseases. But others, lected diseases. In solution remains to be seen. The
including the pharmaceutical industry, addition, the treaty group’s predecessor at WHO, the
argue that the IP protection system isn’t the would promote open Commission on Public Health, Inno-
real problem and that the talks in Geneva access to research vation, and Intellectual Property
risk distracting people from practical solu- findings and possibly Rights, issued a raft of recommenda-
tions. The IGWG—whose members will add R&D incentives. For instance, govern- tions in April—such as increasing contribu-
include representatives of governments as ments could award big monetary prizes for tions to PPPs and building clinical trial capac-
well as nongovernmental organizations— those who invent important new medicines. ity—but could not agree on some key patent
appears “motivated by anticapitalism rather Manufacturers would then be free to pro- issues. Some predict that when the IGWG
than logical thinking about how to get drugs duce and market them cheaply. issues its final report to the World Health
to patients,” says Trevor Jones, a former The treaty, recommended in a letter to Assembly in May 2008, it may propose ways
director of research and development at the the World Health Assembly by 162 scien- to implement the less controversial parts from
Wellcome Foundation. tists, health experts, and others last year, “is the April review rather than a radical reform.
Patents are designed to spur the inven- widely seen as the end of the pharmaceuti- But Love thinks the world may be ready
SOURCE: CIPIH, IMS HEALTH; PHOTO: GETTY IMAGES

tion of new products. But they also allow cal industry as we know it,” says Anne- for a change. He notes that, although the
companies to charge high prices, putting Laure Ropars, a researcher at the George U.S. government has generally aligned
people without purchasing power at a dis- Institute for International Health in London. itself with the pharmaceutical industry, it
advantage. Many critics say it is not enough No wonder the industry is vehemently strongly supported increased access to
to help the poor get access to drugs; the sys- opposed. The treaty would create an HIV drugs in Africa. It also unexpectedly
tem’s incentives must be changed. To pro- “extremely complicated international voted for the resolution introduced by
duce new drugs for neglected diseases, they bureaucracy,” says Eric Noehrenberg of the Kenya and Brazil that called the IGWG into
say, the world needs a new R&D system that International Federation of Pharmaceutical existence. (The drug companies and the
rewards not market sales but the potential to Manufacturers and Associations in European Commission opposed the plan.)
save lives and improve health. Geneva, adding that the award system Love is hoping for another surprise.
One such framework, which the IGWG would never work. Instead, Noehrenberg –MARTIN ENSERINK

www.sciencemag.org SCIENCE VOL 314 1 DECEMBER 2006 1373

NEWSFOCUS

Many U.S. educators think that a streamlined

science curriculum with fewer topics per grade
is a necessary first step toward boosting
student achievement

Doing More With Less

WHAT DO SCHOOLCHILDREN NEED TO Science to School (nap.edu), is that students accountable for student achievement in that
know to be scientifically literate? Scientists receive a “fragile foundation” in science. That subject. The second is the National Assess-
and educators keep coming up with new fragile foundation is exposed in both national ment of Educational Progress (NAEP), a non-
answers to that deceptively simple question. assessments of what students know and in binding, quadrennial federal assessment
As states gear up for two nationwide assess- international comparisons with their peers. of student achievement in grades 4, 8, and
ments of student achievement in science, Those poor performances are fueling a 12 across several subjects. Although called
many educators think that the time is ripe to campaign by the National Science Teachers the nation’s report card, its results are not
take another hard look at what children Association (NSTA) to develop a national broken out by schools and districts, and there
should be taught. But others worry that consensus around what NSTA Executive are no penalties for poor performance.
reviving debate on that contentious topic Director Gerald Wheeler calls “science One big sticking point is that, thanks to the
may divert attention and resources from the anchors”: a small number of concepts that country’s 200-year history of local control
bigger challenge of actually improving stu- educators agree are essential for students to over education, there isn’t a national curricu-
dent performance in science. understand at any particular grade level. lum. Casserly and many educators would like
Everybody agrees that current practices “There are way too many things in the stan- to see voluntary national standards that would
aren’t good enough. “It is the height of dards,” Wheeler says, “and too much diver- reduce variations among the 50 states and
national folly to think that America can main- gence in what’s being taught across the coun- 15,000 local school districts. Two Senate bills
tain any competitive edge in science the way try.” He sees the anchors as a de facto core cur- introduced earlier this year would move the
we are now teaching and testing it,” asserts riculum drawn from topics that most schools country in that direction by asking expert pan-
Michael Casserly, executive director of the are already teaching, “like Newton’s law of els to identify common ground among state
Council of the Great City Schools in Wash- gravity, or evolution and natural selection.” curricula and standards. One bill (S. 3790),
ington, D.C., after urban schools last month Wheeler hopes to influence two testing from Senator Hillary Clinton (D–NY), would
reported low performance in science. There’s regimens that dominate U.S. elementary and even develop a model math and science cur-
also consensus that the curriculum is a big secondary school education. The first, the riculum and sample assessment questions.
part of the problem. A September report by a 2001 federal No Child Left Behind Act The other (S. 2357), by Senator Edward
panel of experts assembled by the National (NCLB), requires states to test students in “Ted” Kennedy (D–MA), would help states
CREDIT: AAAS PROJECT 2061

Academies’ National Research Council grades 3 through 8 each year in reading and align their curricula and standards to national
(NRC) deplores curricula that “contain too mathematics. Its importance derives from the benchmarks. Neither bill attracted much
many disconnected topics that are given equal sanctions facing schools whose students do attention this year, but that’s likely to change
priority, with too little attention to how … not make sufficient progress each year. Next next year, when Kennedy takes over as chair
[knowledge] is enhanced from grade to year, science will be added to that lineup, of the Senate panel with jurisdiction over fed-
grade.” The result, says the panel in Taking although the law doesn’t hold schools eral education efforts.

1374 1 DECEMBER 2006 VOL 314 SCIENCE www.sciencemag.org

NEWSFOCUS

Clear on the concept. AAAS’s Ted Willard leads a Lewis is very supportive of the state’s science,” says Janice Earle, a senior pro-
teachers’ workshop on using the Atlas of Science attempt to upgrade science instruction. But gram director for elementary and secondary
Literacy. she wonders why her state has just adopted its education at the National Science Founda-
third set of science standards since she began tion (NSF), which funded the recent NRC
Do it again teaching 14 years ago. “Why are we doing this report and supported the creation of both
Standards-based instruction is not a new idea. again?” she asks. “Science is science.” 1990s standards documents. One conse-
And this is not the first time the concept is quence is what Shuler calls “the science
being invoked to help raise student achieve- Hydra-headed science wars,” in which experts lobby to make sure
ment in science. In the early 1990s, scientists If only it were that simple. For one thing, most their specialty is adequately represented in
and educators rallied around the idea of experts agree that the nationwide standards any standards document.
describing the important concepts in biology, that came out in the 1990s weren’t really a That effect is magnified as each state (Iowa
chemistry, physics, and the earth sciences that bare-bones version of what students needed to is the lone exception) develops its own stan-
all U.S. elementary and secondary school stu- master. “We pared down by 40% the amount dards, says Nelson, who runs a science, math,
dents need to master, as well as the nature of of material that was being taught, we esti- and technology education program at Western
scientific thought. The movement crested mated,” says George “Pinkie” Nelson, former Washington University in Bellingham. “It’s
with the appearance of two acclaimed docu- director of AAAS’s Project 2061, which easier to put things in than to take them out,”
ments: the 1993 Science Benchmarks for All developed Benchmarks and another AAAS he notes. Expanding the standards, in turn, has
Americans from AAAS (which publishes product, called the Atlas of Science Literacy, led to ever-larger textbooks, as publishers
Science), and the National Academies’ 1996 that presents the concepts in Benchmarks as a scramble to make sure their materials cover all
National Science Education Standards. Edu- cluster of interlocking maps to help teachers the topics state and local school districts had
cators hoped the standards would ensure not prepare lessons on any particular topic. “But crammed into their standards.
only that teachers covered the most important there was still way too much material.” Sally But despite their heft, those textbooks
topics but also that there would be a seamless Goetz Shuler, executive director of the often fail to capture the idea that science is, in
transition from one grade to the next—and, in National Science Resources Center, a joint the words of the recent NRC report, “not only
a highly mobile society, that children wouldn’t effort of the National Academies and the a body of knowledge, but also a way of know-
be shortchanged if they moved from one dis- Smithsonian Institution in Washington, D.C., ing” about the world. That approach includes
trict to another. calls them “a good first effort. … They were a formulating and testing hypotheses, adjusting
So far, so good. “Before the standards, lot better than the mile-wide, inch-deep” cur- one’s understanding to fit the data, and then
teachers pretty much taught whatever they riculum most states were offering at the time. blending that new information with what the
wanted to,” says Megan Lewis, who teaches “But they were still way too complicated,” she student already knows to come up with a bet-
physical sciences, chemistry, and physics to adds, “especially beyond the fifth grade.” ter understanding of any particular phenome-
high school students in the rural Glen Lake, One problem in developing science stan- non. It’s a process that doesn’t fit neatly into a
Michigan, school district. But because many dards is the multiple f ields that must be lecture, or even an experiment, the report
officials see standards as a threat to local con- included. “Remember, it’s the sciences, not points out. And it’s something that few stu-
trol over education, they are no dents have a chance to experience
more than voluntary yardsticks at any level.
that states are free to adopt, “My students have a hard time
modify, or ignore. Over the past f iguring out how things really
decade, state and local education work,” says Thomas Lord, a plant
authorities have used those docu- biologist and science educator at
ments as a starting point for com- Indiana University of Pennsylva-
piling their own standards. Unfor- nia in Indiana, Pennsylvania.
tunately, the results have been less “When I teach the water cycle, for
than ideal. example, I ask them about the role
Take Lewis’s home state. that plants play in the process. Not
Michigan was one of the pioneers one kid mentions photosynthesis.
in the standards movement, And these are science majors.”
adopting science guidelines in Lord worries that any standards,
1991. In 2000, the document was especially something concise
revamped and renamed the such as science anchors, could
Michigan Curriculum Framework. simplify the curriculum to the
Since then, it’s undergone another point of squeezing out the real sci-
metamorphosis, emerging this ence students need to learn.
CREDIT: PROVIDED BY MEGAN LEWIS

summer as the Michigan Merit

Curriculum. The current version, From zero to 10
which describes what students On track. Megan Lewis To be sure, the standards are just
should know at each grade level, is (left) helps her high one element in reforming sci-
linked to tougher statewide gradu- school students with ence education, an effort that
ation requirements that, for the an experiment demon- includes improved teacher train-
first time, mandate 3 years of high strating Newton’s laws ing and professional develop-
school science. of motion. ment and stronger ties between

www.sciencemag.org SCIENCE VOL 314 1 DECEMBER 2006 1375

NEWSFOCUS

school districts and univer- Anchors aweigh. NSTA’s Gerry Wheeler (center) talks
sity science faculty. “Gerry’s with Wendy Benz (left) and Chad Sechrist at a regional
idea of putting together some teachers’ conference in Baltimore, Maryland.
big ideas is an important
one,” says Nelson. “And new Michigan’s Lewis doesn’t have the luxury
standards are fine. But I think of time. As a classroom teacher, she’s respon-
that it’s zero on a 10-point sible for making sure her students understand
scale of improving U.S. sci- the subject matter and can pass the high-
ence education.” stakes tests. As a result, she suggests, only
Wheeler agrees that sci- partly in jest, that some of the money being
ence anchors won’t suddenly used to rework science standards might be
make students smarter or better spent on her students. “With $10,000, I
give teachers a better under- could buy enough [PASCO] probes for every
standing of fundamental sci- kid in my class,” she says, referring to equip-
entif ic concepts that they ment that allows experimental data on temper-
never learned adequately ature, acceleration, and other features of the
before entering the class- physical world to be collected and analyzed.
room. But he thinks that the Wheeler agrees that the interaction of stu-
anchors might be attractive to dent and teacher is paramount to improving
states preparing for two upcoming major A matter of time how schoolchildren learn science. “A clear set
assessments. “It’s a way of identifying the NSF’s Earle thinks there are valid reasons to of standards aligned to the state assessment is
low-hanging fruit. It’s a marketing tech- be optimistic about the latest efforts to clar- a key first step,” he reiterates. “But unfortu-
nique,” he admits. “Once people buy into ify what students need to know in science. nately, we have to take about four first steps,”
the concept, then maybe we can get them to “I’m sensing that maybe it is time to think he adds, ticking off the need for better materi-
develop better assessment items, and pro- about taking the next step,” Earle opines. als, improved professional development, and
fessional development, around them. If that “The standards have been out there for a higher teacher retention rates. “Otherwise,
happens, then I think we will be moving in decade or so, and it takes people a while to there’s going to be a lot of finger-pointing at
the right direction.” digest them.” But don’t expect anything to the fourth grade teacher whose students didn’t
The science component of NCLB begins happen quickly, she counsels: “U.S. educa- do well enough on the science assessment.
in 2007–’08, for students in one grade at each tion is not efficient, by definition. We have a And that doesn’t help anybody.”
of three levels: elementary, middle, and high very decentralized system.” –JEFFREY MERVIS
school. But the results won’t be counted as
part of the law’s requirement that students
show adequate yearly progress (AYP). “Not ASTROPHYSICS
being part of AYP means that science may
remain on the back burner,” Wheeler fears. A
more promising target may be NAEP, which Burst-Hunter’s Rich Data Harvest
will be given next in 2009. Earlier this year,
an expert panel (Wheeler chaired its steering
committee) sketched out a new “framework”
Yields a Cosmic Enigma
of what the test should cover, as well as new The 2-year-old Swift gamma ray satellite has delighted astrophysicists with its
ways to measure that knowledge. versatility—and surprised them with observations that don’t fit the models
The draft NAEP framework (nagb.org)
drew explicitly from the two 1990s docu- SAN FRANCISCO, CALIFORNIA—In the quest Now, Swift has given astrophysicists a
ments, says Senta Raizen of the National for the secrets of cosmic explosions known as major surprise. Its observation of a GRB ear-
Center for Improving Science Education run gamma ray bursts (GRBs), no satellite has lier this year has challenged the standard clas-
by WestEd, a California-based nonprofit with been more successful than Swift. sification system for such bursts.
a contract from NAEP’s oversight body to Launched 2 years ago in November, Swift GRBs are intense pulses of extremely
revise the assessment. “Our hypothesis was, has outperformed expectations, providing high-frequency radiation emanating from
if it’s in both documents that it’s in,” says NASA with a steady stream of news to distant space. Such bursts were first detected
Raizen, who co-chaired the project’s plan- report. Swift not only has been recording its almost 4 decades ago by satellites designed to
ning committee. “If only one, then we’ll think predicted budget of bursts (about 100 a year) seek signs of nuclear weapons tests. When
about it. So there’s nothing fundamentally but also has gathered abundant data on other gamma radiation arrived from space instead
new about the content.” Raizen emphasizes astrophysical phenomena, from nearby black of the ground, baffled astrophysicists groped
that NAEP isn’t trying to tell states what to holes in active galaxies to the most energetic for explanations. Among the more specula-
teach—“we don’t have a national curriculum magnetic flare ever detected on a star. “Swift tive suggestions was that the bursts signaled
in this country”—and that NAEP provides has been a scientific bonanza,” says high- the demise of faraway civilizations that had
only “a snapshot” of what students have energy astrophysicist Ilana Harrus of annihilated themselves in nuclear wars.
CREDIT: NSTA

learned. But she agrees that it can “serve as a NASA’s Goddard Space Flight Center in In the 1990s, however, observations from
model” for the upcoming NCLB assessments Greenbelt, Maryland. “It’s the satellite that the orbiting Compton Gamma Ray Observa-
that states must devise. keeps on giving.” tory provided enough information to pin down

1376 1 DECEMBER 2006 VOL 314 SCIENCE www.sciencemag.org

NEWSFOCUS

key details, eventually establishing that some In another online paper, Johan Fynbo of ther away, too far for the supernova associ-
bursts were associated with supernova explo- the Niels Bohr Institute in Copenhagen, Den- ated with it to be visible.
sions. Others seemed to result from cosmic col- mark, and an international team of collabora- Data from Swift and other instruments can
lisions, perhaps between neutron stars, the tors including Berkeley’s Bloom suggest that be used to estimate the intrinsic brightness of the
small, dense spheres left behind by supernovae. the 14 June burst implies a new type of explo- burst, Schaefer and Xiao point out. Comparing
Bursts believed to be associated with neu- sive star death, producing a GRB but no the intrinsic brightness with the observed
tron stars were typically short—lasting less supernova. If so, the burst represents the first brightness gives a good measure of distance.
than 2 seconds. “Long” bursts lasted from sec- of a whole new category of GRBs. Gehrels’s Various indicators all suggest a high brightness
onds to minutes and were generally “softer”— and Fynbo’s papers have both been accepted for the burst, leading Schaefer and Xiao to
meaning lower in energy—than the short, for publication in Nature. assign it a redshift of about 2, far enough away
“hard” higher-energy bursts. Long bursts have Other astrophysicists, however, say it’s too to explain the lack of a supernova sighting. They
been clearly linked to supernovae, but the soon to junk the two-category system or invent calculate the odds of such a lineup of a galaxy
short bursts’link to neutron stars is more spec- new stellar death processes. “These guys are with a more distant burst to be 1 in 125; because
ulative. “Short bursts lack a smoking gun,” going off and making claims that you have a Swift has recorded more than 190 bursts, find-
says Joshua Bloom, a GRB investigator at the whole new class of [GRB] popu- ing one such alignment is not
University of California, Berkeley. lation never before seen,” says surprising. “It’s fully consistent
In June, Swift further blurred the line Bradley Schaefer of Louisiana with chance coincidence,”
between short and long by finding a long, soft Schaefer says.
burst with no apparent connection to a super- A similar conclusion
nova, several astrophysicists reported at a So rare. “Long” gamma ray burst appeared in a paper published
Swift spotted in June had puzzling
recent meeting here.* That new burst and other 10 November in Astrophysical
“short” properties.
Swift observations challenge the standard Journal Letters. B. E. Cobb
two-category scheme and raise questions and colleagues at Yale Univer-
about the nature of GRB progenitors. sity determined the likelihood
“I see a growing crisis of classification,” of lineup coincidence to be
said Bloom. “We don’t just have long bursts and from about 1 in 50 to 1 in 200.
short bursts anymore that map directly to these Consequently, from one to
progenitors. We actually have counterexamples four such coincidences would
in both cases that are really throwing a monkey be expected in the bursts observed
[wrench] in the works.” by Swift so far. “The conclusion
The prime culprit behind the category cri- that [the 14 June burst] requires
sis is a burst recorded on 14 June that lasted a ‘new paradigm’ for gamma
103 seconds, far into the range generally ray burst formation should be
regarded as long. Observers eagerly awaited approached with caution,” Cobb
the appearance of stellar brightening signaling and colleagues wrote.
the supernova explosion responsible for the Actually, a second such possi-
burst. But the supernova never showed. ble coincidence had already been
Further observations suggested that the recorded before the 14 June event.
14 June event was not a typical long burst in A burst detected in May also was
other ways. The initial pulse was high in energy technically “long”—at 4 seconds
but was then followed by a softer afterglow. in duration—with no sign of a
“That was kind of reminiscent of other supernova. But that event was
short bursts we’ve seen,” Swift principal State University (LSU), Baton Rouge. For fainter and poorly observed, Schaefer and
investigator Neil Gehrels of NASA Goddard such an extraordinary claim, he says, “you Xiao noted, and the burst might also have orig-
said at the astrophysics meeting, suggesting ought to have at least good evidence.” inated far behind the presumed host galaxy.
that it belonged in the “short” category. In fact, Schaefer says, the mystery of the In any case, Swift’s findings have surely
CREDITS: SPECTRUM ASTRO; (INSET) ADAPTED FROM SWIFT/SONOMA

“Short isn’t the right word,” Gehrels said, but missing supernova could have a simple complicated the older views of GRBs, pro-
in many respects “it appears to group with solution: The burst may have occurred far viding much precise data that astrophysicists
the short bursts, and that could explain the enough away to make the stellar explosion will have to digest to get a clearer under-
lack of a supernova.” too dim to notice. Lack of a supernova standing of the sources and properties of
In a paper posted online, Gehrels and col- seems mysterious only because the burst those cosmic flashes.
laborators argue that the 14 June burst was estimated to be at a redshift of 0.125, “Our real goal here is to attempt to uncover
requires a new categorization system. “This relatively nearby in cosmic terms. But in a the progenitors of gamma ray bursts, whether
combination of a long-duration event without paper posted on the online astrophysics they be long-duration gamma ray bursts or
an accompanying supernova … opens the preprint archive (astro-ph/0608441), short-duration gamma ray bursts,” says
door on a new gamma ray burst classification Schaefer and LSU collaborator Limin Xiao Bloom. “We’re trying to understand the diver-
scheme that straddles both long and short point out that that distance estimate is based sity of the phenomenon. And because of Swift
bursts,” the Swift scientists wrote. solely on the measured redshift of the near- and other satellites, we’re now in the position
est galaxy along the line of sight to the to really ask these questions in detail.”
*High Energy Astrophysics Division of the American burst. Perhaps that galaxy was not actually –TOM SIEGFRIED
Astronomical Society, 4–7 October. the burst’s host and the burst was much far- Tom Siegfried is a writer in Los Angeles, California.

www.sciencemag.org SCIENCE VOL 314 1 DECEMBER 2006 1377

GLOBAL HEALTH
a lack of leadership on HIV/AIDS,” says
South Africa Bolsters HIV/AIDS Francois Venter, who heads the Southern
African HIV Clinicians Society. But he and

Plan, but Obstacles Remain others cautioned that the devil is in the
details, some of which were not available as
Science went to press. And no one was
Ridicule at the Toronto AIDS Conference spurred South Africa’s Cabinet to order a new expecting that the announcement of a new
plan to battle the epidemic action plan would end the debate on South
Africa’s HIV/AIDS policies.
PRETORIA AND SOWETO, The need for more effective government
SOUTH AFRICA—When small programs is clear. Although a draft of the plan
baskets of garlic, lemons, and cited evidence that “HIV incidence has
beets highlighted the South started to decrease,” a November report by
African exhibit at the XVI the Joint United Nations Programme on
International AIDS Confer- HIV/AIDS and the World Health Organiza-
ence in Toronto last summer, tion says that HIV prevalence—at nearly
many delegates were out- 19% of South Africa’s adult population in
raged. They viewed the dis- 2005—“has not yet reached a plateau.” The
play—intended to show the nation’s 5.5 million infected people include a
importance of nutrition in quarter of a million children under age 15, the
bolstering immune systems— report said. It also warned of “a continuing,
as trivializing the response to rising trend in HIV infection levels” among
the epidemic that now infects pregnant women using prenatal clinics.
5.5 million South Africans and
kills an estimated 800 of them A history of controversy
a day. As the meeting ended, International dissatisfaction with the coun-
Stephen Lewis, the United try’s HIV/AIDS policy is rooted in a series of
Nations special envoy for government controversies and miscues over
HIV/AIDS in Africa, lashed the past decade. In 1997, an attempt to fast-
out at aspects of South Africa’s track clinical trials of a drug called Virodene
AIDS policies as “wrong, ended in disgrace when a review panel found
immoral, [and] indefensible.” that the substance was toxic and had been pre-
The ridicule in Toronto maturely tested on humans. Three years later,
was followed by a sharply in early 2000, Mbeki sent a letter to the White
critical letter to President Thabo House and to the U.N. Secretary-General
Mbeki from 82 prominent suggesting that factors other than HIV could
international scientists, includ- cause AIDS and asserting that it would be a
ing Nobelist David Baltimore, Under fire. South Africa’s embattled health minister, Manto Tshabalala- “criminal betrayal” to “mimic foreign
virologist Robert Gallo, and Msimang, has been the target of protesters who called for her dismissal. approaches to treating HIV/AIDS.”
11 South African researchers. Later that year, delegates to the Inter-

CREDITS (TOP TO BOTTOM): RAJESH JANTILAL/AFP/GETTY IMAGES; HOWARD BURDITT/REUTERS

Arguing that garlic and lemons “are not alter- ter—as the nation’s point person for devel- national AIDS Conference in Durban were
natives to effective medications,” the oping a more effective HIV/AIDS strategic stunned that Mbeki and his health minister
researchers warned that “many people are … plan for the next 5 years. continued to question the connection between
dying unnecessarily” in South Africa because The deputy president planned to outline HIV and AIDS and failed to support ARV
they do not have access to antiretroviral the framework of that new plan on 1 Decem- therapy. “The government took a strange
(ARV) drugs to slow the progression of the ber, World AIDS Day. A draft of the wide- position opposed by well-established sci-
disease. Although the country has the world’s ranging plan, obtained by Science, features ence,” recalls the chair of the Durban meeting,
largest ARV program, it now reaches only commitments to bolster prevention pro- pediatric AIDS researcher Hoosen Coovadia
about a quarter of the South Africans who are grams to sharply reduce the number of of the University of KwaZulu-Natal.
estimated to need the drugs. people being infected with HIV; better Meanwhile, as the epidemic worsened, the
Reflecting the outcry, some of South c o o r d i n a t e t h e g ove r n m e n t ’s o f t e n - government came under increasing pressure
Africa’s leading newspapers called in Septem- f r a g mented response to the epidemic; to take decisive action. In 2002, Mbeki began
ber for the resignation of the garlic-promoting support AIDS vaccine and antimicrobial to distance himself from the denialists and
health minister, Manto Tshabalala-Msimang. research; and significantly expand ARV endorsed the concept of making ARVs avail-
At about the same time, South Africa’s ruling treatment—although the exact ARV target able to pregnant women and rape survivors.
Cabinet, unhappy to again be the focus of numbers were still being developed. Late in 2003, a panel developed an ARV roll-
international scientific scorn, decided to Many South African scientists, clini- out plan, which went into effect the following
revive the near-moribund South African cians, and activists welcomed the long- spring and now covers about 214,000 per-
National AIDS Council. It named Deputy overdue initiative to revamp HIV/AIDS sons. Noting that the South African ARV pro-
President Phumzile Mlambo-Ngcuka— programs. “For years, we had been con- gram reaches more people than that of any
rather than the controversial health minis- fronted with obfuscation and confusion and other country, Medical Research Council

1378 1 DECEMBER 2006 VOL 314 SCIENCE www.sciencemag.org

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(MRC) President Anthony MBewu contends Antiretroviral Therapy in Low- and Middle-Income Countries
that South Africa’s recent initiatives on 1400

People receiving therapy (thousands)

HIV/AIDS treatment and prevention have
begun to blunt the epidemic. 1200 North Africa and the Middle East
Europe and Central Asia
Trying to rehabilitate the government’s 1000 East, South, and Southeast Asia
international image on HIV/AIDS policy, Latin America and the Caribbean
800 Sub-Saharan Africa
Cabinet officials are avoiding any public
expression of AIDS denialism. Government 600

spokesperson Themba J. Maseko told Science 400

that “the position of the government is based
200
on the understanding that HIV causes AIDS.”
Even South Africa’s most outspoken AIDS 0
End 2002 Mid-2003 End 2003 Mid-2004 End 2004 Mid-2005 End 2005
activist, Zackie Achmat, credits the ruling
African National Congress party with exert- African challenge. A researcher tests blood at the Perinatal HIV
ing pressure to suppress AIDS denialism Research Unit in Soweto, South Africa. The number of people receiving
within its ranks. Although Mbeki has not antiretroviral therapy in South Africa—and sub-Saharan Africa as a
made a definitive statement of his own posi- whole—has climbed in the past few years, but the need is still great.
tion, Maseko says the president fully supports
the Cabinet’s recent HIV/AIDS decisions on
developing a new action plan. Campaign, the AIDS Law Project, and the Uphill battle
University of the Witwatersrand’s Repro- Although generally heartened by plans to
A targeted approach ductive Health and HIV Research Unit— boost HIV/AIDS prevention and treatment
After the Toronto AIDS meeting, with the delivered a 28-page critique in November of programs, South African scientists and
health minister hospitalized with a respira- an early draft of the action plan. That activists caution that obstacles remain. For
tory ailment, the Cabinet asked Deputy response, obtained by Science, argued that some, the chief problem is the recuperat-
President Mlambo-Ngcuka to try to mend the proposed ARV targets “represent approx- ing health minister, Tshabalala-Msimang,
fences with interest groups and develop a imately 20 percent of those requiring treat- who remains responsible for implementing
stronger HIV/AIDS plan. Declaring that the ment, and should be revised upwards. All the new HIV/AIDS plan. In November, she
nation’s AIDS policy debate was at “a criti- epidemiologic data suggests that there are issued a statement lashing out at her crit-
cal point,” she held meetings this fall with approximately 800,000 people who need ics and reaff irming her commitment to
leading clinicians, scientists, and activists to ARVs at the moment … and that an addi- nutrition and traditional medicine in
try to resolve “difficulties and misunder- tional 500,000 people will require treatment HIV/AIDS treatment.
CREDITS (TOP, LEFT TO RIGHT): WHO/UNAIDS (2005); PERINATAL HIV RESEARCH UNIT, SOWETO; (BOTTOM) LERATO MADUNA/REUTERS

standings” and forge a consensus. annually going forward.” University of Cape Town economist
The most diff icult single issue in The critique also argues that the nation’s Nicoli Nattrass, an expert on the impact
reaching such an agreement has been set- research facilities are “largely uncoordinated of the epidemic on South Africa, believes
ting targets for ARV treatments. (The gov- when it comes to research on HIV” and rec- that HIV/AIDS activists “have won an
ernment bears most of the costs of ARV ommends that a national health supervisory important ideological battle,” but—with
drugs at public clinics, but international council find ways to improve coordination. the health minister still involved in imple-
organizations and donors pay for ARV costs The draft HIV/AIDS plan did not mentation—“the counterinsurgency
at many private or religious facilities, and address the coordination issue, but it con- remains strong.”
medical insurance covers other individu- firmed that basic and clinical research into Clinicians and researchers are eager to
als.) The government originally planned to the epidemic were national priorities. see evidence that the government will back
announce specific ARV targets on World Although shy on details, the draft specifically up its new HIV/AIDS commitments with
AIDS Day as part of the new called for boosting more funding and improvements in health
plan. Indeed, one draft listed a research into micro- facilities. “Setting ambitious targets is
goal of tripling the current ARV bicides and AIDS good, but you have to have the resources
numbers, to 650,000 adults vaccines, a research and plans to meet those targets,” says
and 100,000 children, by 2011. strength of the coun- researcher Coovadia.
But as soon as the plan’s early try. MRC President Complicating the challenge, tuberculo-
drafts began circulating, AIDS MBewu told Science sis is rife among South Africans infected
activists and clinician groups that “HIV/AIDS with HIV, and new drug-resistant strains
began lobbying for much higher i s the nation’s top are threatening to spread rapidly. “We’ve
numbers, and they persuaded research priority.” got two epidemics clashing in a dangerous
the deputy president to delay way. We can’t carry on with business as
announcing specific targets until Action plan. Deputy
usual,” says immunologist Linda-Gail
a compromise could be worked President Phumzile Bekker, co-director of the Desmond Tutu
out, probably early next year. Mlambo-Ngcuka is HIV Center in Cape Town.
Four influential AIDS groups South Africa’s new point Clinician Venter agrees that “a lot more
that sought the delay—the South- person in bolstering needs to be done to get control of this epi-
ern African HIV Clinicians Soci- the nation’s HIV/AIDS demic.” Still, he says, “it helps to have sup-
ety, the activist Treatment Action action plan. port at the top.” –ROBERT KOENIG

www.sciencemag.org SCIENCE VOL 314 1 DECEMBER 2006 1379

Vanishing breed. A young captive saola shortly
before its death in Hanoi in 1993.

central Vietnam.
The odds are against Long and company.
“Foundations can easily raise funds for pri-
mates, tigers, elephants, rhinos,” says Dang.
“For the saola, we can’t even get money to
educate the public, to tell people to stop hunt-
ing it.” As Vietnam’s action plan notes starkly,
“resources and attention afforded to the saola
are currently insufficient to protect it from
extinction in the immediate future.”

Trophy hunting
Whereas biologists are captivated by the
saola’s unicornlike mystique, villagers in
Truong Huong, on the edge of the Pu Mat
Nature Reserve, are blasé about the beast.
WILDLIFE CONSERVATION Few in this ethnic Thai community have seen
a saola, and when they do, the outcome for the

The Saola’s Last Stand demure herbivore is almost invariably bad.

In a wooden house built on stilts, Lo Van
Tinh, a farmer, sits cross-legged with four
Wildlife experts say the rare Southeast Asian ungulate may soon disappear; a generations of family huddled around him
Vietnamese lab is undertaking a controversial attempt to clone it and describes how, one day 10 years ago, he
was hunting turtles in a mountain river. His
PU MAT NATURE RESERVE, VIETNAM— Do More is at stake than one obscure relict dog spotted a saola mother and calf upstream
Tuoc climbs a steep riverbank, entering the species. The ecosystem that shelters the saola and gave chase. The mother escaped, but her
realm of the elusive saola. The creature, a is home to an array of creatures, including at calf was cornered and assumed a defensive
cousin of cows, goats, and antelopes, is so least two kinds of muntjac deer found posture. Although a saola in captivity betrays
rare that even Tuoc, who discovered the nowhere else in the world. Saving this unique no fear of humans, at the sight of a dog it
species in 1992, has never spied one in the menagerie “would be a success story for other snorts and hunkers head down, brandishing
wild. The forest ecologist finds safe footing countries to follow,” says Barney Long, a con- its long, straight horns, says saola expert
on the slick slope and grabs a handful of servation biologist with the World Wide Fund William Robichaud, a zoologist with the
broad, dark-green Araceae leaves. “Saola for Nature (WWF) who is working with local Nakai-Nam Theun National Protected Area
like to eat these,” Tuoc says. “At least, we scientists and officials to protect the saola in in Laos. That renders the saola easy to shoot,
have seen bite marks.” and for a juvenile, easy to grab. “I caught it
A decade ago, the saola made headlines with my hands,” says Tinh. The saola did not
as the first large mammal new to science in survive the 2-day hike back to Truong Huong,
more than half a century. Recent sleuthing so Tinh and his family ate it. It was like beef,
suggests that the exotic ungulate is sliding although not as tasty, he says.
toward extinction. At most, 250 saola are In a home in nearby Truong Chinh village,
thought to roam the Annamites (called the a pair of saola horns hangs in a place of honor
Truong Son Mountains in Vietnam) of cen- next to a poster of a smiling Vietnamese

CREDITS (TOP TO BOTTOM): WWF-CANON/DAVID HULSE; MUTSUMI STONE

tral Vietnam and Laos. model. The dark-brown horns, about 40 cen-
Now scientists are embarking on a last- timeters in length, are more than twice as long
ditch effort to save the critically endangered as the head, which has short, coarse, chestnut-
species. Vietnam’s National Saola Conserva- brown hair. To local people, the slightly
tion Action Plan, expected to be approved by diverging horns resemble the parallel wooden
the government later this month, prescribes posts that support a spinning wheel (hence the
measures, including a hunting ban, that are name: Sao means “post,” and la means “spin-
deemed essential for the saola’s survival. ning wheel”). Streaks of white hair above the
Meanwhile, a Vietnamese team is pursuing a eyes look like garish mascara.
conservation option of last resort: an attempt It was in a home just like this that Tuoc dis-
to clone the saola. But somatic cloning is covered the saola. In May 1992, he was part of
supremely difficult even in the best-studied a team dispatched by the Ministry of Forestry,
mammals—and “we know almost nothing with WWF support, to Vu Quang forest,
about the saola,” says zoologist Nguyen Xuan roughly 100 kilometers southeast of Pu Mat,
Dang of the Institute of Ecology and Biologi- On the track of unknown animals. Ecologist Do to survey biodiversity in advance of Vu
cal Resources in Hanoi. Tuoc, Araceae in hand, at the Pu Mat Nature Reserve. Quang’s designation as a nature reserve.

1380 1 DECEMBER 2006 VOL 314 SCIENCE www.sciencemag.org

NEWSFOCUS

Tuoc, schmoozing with the local villagers, Museum piece. A saola head in a
wangled an invitation to a young hunter’s hunter’s home near Pu Mat. Saola
home, where the team was shown a peculiar sightings in Laos and Vietnam are
skull and horns. “I immediately thought it was dwindling.
a new species of antelope,” Tuoc says. But it
was puzzling, as antelope prefer dry areas, and of a saola for his Lak Xao Zoo.
much of the Truong Son range is soaked by In January 1996, Cheng pro-
seasonal monsoons. Excited by the find, he cured an adult female. “I had
asked local hunters to look for other speci- the good fortune to observe her
mens. Two more pairs of horns soon material- daily,” says Robichaud. The
ized, convincing the scientists that they had saola, he says, marked territory
indeed found a new species, which they by flaring open a fleshy flap
anointed the “Vu Quang ox.” covering her maxillary glands
WWF funded a follow-up survey that on either side of the snout and
November that turned up about two dozen stroking the underside across
pairs of horns and an intact saola skin. DNA rocks, depositing a pungent,
analysis of the mitochondrial cytochrome b musky paste. The massive
gene revealed a new bovid genus, and a scent glands are thought to be
paper in Nature in 1993 unveiled Pseudoryx the largest of any living mam-
nghetinhensis. (Subsequent DNA analyses mal.
suggest that cattle are its closest cousins.) “Her most striking and
The animal was confirmed in Laos through endearing aspect,” Robichaud
villager sightings and trophy horns in 1993. says, “was her utter calmness
The species name is an amalgamation of the in the presence of humans.”
two Vietnamese provinces where specimens Soon after arriving at Lak Xao,
were first uncovered. The common name Vu the saola allowed people to
Quang ox soon gave way to saola, a less stroke her and fed from their
parochial designation and one with histori- hands. “She was tamer and more
cal roots. The first known written reference approachable than any domestic live-
to the species is in an early 20th century Lao- Nghe An
Pu Mat stock I’d ever been around,” he says.
French dictionary, which defines saola as an But after a mere 18 days in captivity,
“antelope of the rocks,” says Robichaud. the saola died suddenly, and no
The saola was the first large mammal dis- autopsy was performed—although she
covery since the kouprey, a wild ox in South- Thua was found to be bearing a male fetus.
east Asia, in 1937. As an encore, Tuoc and col- Thien Saola are so rarely seen in the wild that it
leagues first described the large-antlered (for- LAOS Hue wasn’t until 1998 that one was first caught on
merly giant) muntjac in 1994 and the diminu- Quang film in its habitat, by a camera trap near a
tive Truong Son muntjac in 1997. (Both were THAILAND Nam mineral-rich spring in Pu Mat. Robichaud
discovered simultaneously in Laos.) With and Robert Timmons, an independent con-
three mammal species under his belt, Tuoc has servation biologist in Southeast Asia, have
become a legend in cryptozoology, the study VIETNAM suggested that the survivors are descendents
of previously unknown, presumed, or mythi- of a Pleistocene bottleneck, when their wet
CAMBODIA
cal creatures. “I’ve been very lucky,” he says. evergreen forests receded during cool, dry
Presumed
ice ages. “The current distribution of saola
Zoological riddle saola may reflect where these ice age refugia
Ever since the saola’s appearance, its biology, distribution were,” says Robichaud.
like the animal itself, has remained an enigma. Humans now have the saola on the ropes.
In June 1993, Tuoc and colleagues at the For- In 1992, scientists pegged the population at
est Inventory and Planning Institute in Hanoi Survival at the Smithsonian’s Conservation 500 to 1000 in Vietnam, says Long. The esti-
took custody of two young saola that had been and Research Center near Front Royal, Vir- mate in Vietnam’s action plan—“probably”
captured in Vu Quang. The animals ate several ginia. Or the problem could be as simple as an fewer than 200—could be a large under- or
dozen kinds of plants and put on weight fast, “inappropriate” diet, says Wildt, whose team overestimate, he says. But Long says a decline
Tuoc says. But after 2 months, they suc- has pioneered techniques for breeding deli- is evident “from the amount of hunted tro-
cumbed to infections. In all, 20-odd saola cate creatures such as the Elds deer and the phies that we see” and the lack of sightings in
CREDIT (TOP): MUTSUMI STONE

have been captured in Vietnam and Laos. All black-footed ferret. “A careful examination of areas where the saola once roamed. Saola are
but two that were released into the wild died why these animals die after capture is really also killed in snares set for more lucrative
quickly in captivity. needed,” he says. game such as bears, which fetch a high price
The saola’s fragility is no big surprise. What little is known about the saola has for their gall bladders. Vietnam’s action plan
“Certain animals in captivity, especially ungu- been gleaned primarily from the short-lived would ban snares in saola territory.
lates, are highly sensitive to stress,” says captives. In the mid-1990s, Cheng Syavong, a Habitat fragmentation further endangers
David Wildt, head of the Center for Species Lao general, offered a reward for the capture the species. The action plan notes that the

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nearly completed Ho Chi Minh Highway, the reproductive cycle, no idea how long
which will link northern and southern Viet- pregnancy lasts,” he says.
nam, “must be viewed as the single largest Nguyen and his collaborators have filled
threat to the connectivity of Saola populations in some gaps. For instance, they’ve estab-
and their habitat.” With support from the lished that the saola has 50 chromosomes.
World Bank, the Dutch Development Organi- (Cows have 60, buffalo 84.) Nguyen now
zation, and the U.S. Agency for International hopes to unravel how saola nuclei are repro-
Development, WWF is working with Viet- grammed. During reprogramming, an egg
namese authorities to protect forests in two turns back the clock on an adult nucleus by
provinces, Thua Thien Hue and Quang Nam, removing chemical signatures of develop-
where the largest saola subpopulation, ment, which returns it to an embryonic
approximately 50 individuals, is found. This state—an essential step in somatic cloning.
“Saola Conservation Landscape” abuts “We’re interested in early molecular events
forests in Laos, providing contiguous habitat in saola and closely related species,” says
for some of the few dozen saola thought to live Renard.
across the border. When all the problems of interspecies
As an additional safeguard, Vietnam’s cloning—such as different chromosome
national action plan would forbid keeping numbers and different mitochondrial
saola in captivity until 2010, unless one is con- DNA—are solved, then “cloning the saola
fiscated from a hunter or liberated from a will be possible,” predicts Takashi Nagai, a
snare and is too injured to be released into the reproductive biologist at the National Insti-
wild. To Wildt, this is a risky strategy. “I don’t tute of Livestock and Grassland Science in
go along with the philosophy of leave them Tsukuba, Japan, who is working with
only in the wild and hope for the best,” he says. Nguyen to conserve the genetic line of Viet-
He suggests that saola experts convene a namese miniature pigs. Nguyen says he will
workshop that would take a hard look at cap- persevere: “I’m a patient man.”
tive breeding. “It’s not like this has never been Some biologists, however, deem the
done before,” he says. effort hasty—or misguided. “Cloning is a
tool for last-ditch heroics,” says Wildt. “It’s
A genetic “Hail Mary” too premature to consider it” for the saola,
Long and others argue that without a robust he says. “I don’t see any conservation bene-
effort to shield the saola from hunters and fit from cloning the saola,” adds Long. “The
preserve its habitat, the animal is doomed. money … would be much better spent trying
For all they know, the species may already Eleventh-hour heroics? Bui Xuan Nguyen hopes to to protect the species in the wild.” (Nguyen
have passed the point of no return. clone a saola. So far, his team’s early saola embryos says his funding is “modest.”) To Long, the
That possibility is the main justification have failed to develop. battle must be fought in the Truong Son
for a controversial, high-tech bid to keep the Mountains. “If we lose the saola,” he says,
species on life support. On the tree-lined endangered species conservation. By then, “it will be a symbol of our failure to protect
grounds of the Vietnamese Academy of Sci- the saola had become an icon in Vietnam. this unique ecosystem.”
ence and Technology in central Hanoi, a Nguyen struck up a collaboration with That could jeopardize unknown species.
team at the Institute of Biotechnology led by Tuoc’s forest institute. “When someone “In Vietnam, there is still a lot of terrain not
Bui Xuan Nguyen is trying to clone the finds a saola, the institute calls us and we yet surveyed,” says Dang. Only in 2005, the
saola. immediately go take tissue samples,” kha-nyou, a bizarre, smallish rodent, was
Nguyen knows the project is a long shot. Nguyen says. They have samples from one described from a specimen found in a Lao
But his lab has a chance at succeeding: He male and two females, including 30 imma- market; an expedition brought back the first
and his staff have been collaborating with ture eggs from one of the females that died. live specimen last May. “There are small and
top reproductive biologists in France, Japan, They’ve held on to most of the eggs in the medium-sized animals waiting to be discov-
CREDITS (TOP TO BOTTOM): MUTSUMI STONE; BUI XUAN NGUYEN

and elsewhere for 30 years and have racked event that, someday, they might be able to ered,” Dang says.
up achievements in embryo transfer and in attempt in vitro fertilization. But Nguyen Optimists about the saola’s fate are about
vitro fertilization in animals such as cows has decided that “we cannot wait for a live as rare as the animal itself—but Tuoc is one
and rabbits. Nguyen is also credited with female.” Working with Patrick Chesné from of them. Natural enemies like the dhole are
having developed a technique for rapidly the lab of Jean-Paul Renard of the National becoming scarcer, he says. Provided that
freezing eggs and sperm that is particularly Institute for Agricultural Research in Paris, snares are removed and vital habitat is pre-
handy for preserving samples in the field. Nguyen has used nuclear transfer to inject served, the saola should be able to rebound,
Building on this work, Nguyen is spearhead- saola DNA into cow, goat, and swamp buf- Tuoc says. “Maybe I’ll never see one in the
ing an effort to set up a lab network in South- falo eggs. They have obtained early wild,” admits the cryptozoologist extraordi-
east Asia next year to “cryobank” frozen embryos—blastocysts—but these fail to naire. “But I think—no, I hope—it will sur-
germ cells of rare species. develop. “We don’t have any idea how to get vive.” For the saola, survival will mean van-
Soon after the cloning of the sheep Dolly past this stage,” Nguyen says. A fundamen- ishing back into the misty sanctuary that hid
in 1997, Nguyen says, he thought the revolu- tal hurdle is the dearth of knowledge about it so well until humans came along.
tionary technique might be applicable to saola biology. “We have no information on –RICHARD STONE

www.sciencemag.org SCIENCE VOL 314 1 DECEMBER 2006 1383

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COMMENTARY
Looking below Parenting natural Turning pattern of
eusociality phenomena hair growth

1391 1393 1397

LETTERS I BOOKS I POLICY FORUM I EDUCATION FORUM I PERSPECTIVES

LETTERS Glossing Over the Complexity

edited by Etta Kavanagh of Water
Balancing Communication and Safety ALTHOUGH WE APPLAUD THE RECOGNITION GIVEN BY
Science to Freshwater Resources, the recent Special Section
(25 Aug., pp. 1067–1090) missed an opportunity to high-
DEMOCRATIC SOCIETIES ARE NOW ENGAGED AGAINST TERRORIST ACTIVITIES. light the multifaceted nature of water resources
In such an environment, there is tension between the desire to withhold scientific research. Framing “the” water problem as a search to
information from those who would use it for ill and the need to not stifle fundamen- quench a universal thirst (“A thirsty world”) glosses over
tal research in the life sciences or the open communication of results. To inhibit the critical differences in the causes of, and thus the solu-
pursuit of science may suggest safety from those prepared to tions to, water problems across regions. It forces the dis-
“Freedom for use science for harmful purposes, but any sense of security is cussion into the domains of supply augmentation and
false. Freedom for research and communication is more nec- engineering and marginalizes underlying drivers of
research and essary than ever, and the best defense against those who “thirst” such as rapid urbanization, economic transitions,
communication is would employ science as a weapon is scientific excellence. geopolitical factors, or poverty.
more necessary There will inevitably be worldwide communication of the Lack of access to water in many African countries, for
than ever…” results of scientific studies, but open communication is vital example, is less the outcome of a first-order water scarcity
—Ehrlich to peer review and an independent evaluation of research, than of a second-order scarcity of social resources (1).
including oversight by the executive and legislative branches As the News story “Running out of water—and time”
of government as well as the public. Open communication is also essential for (J. Bohannon, p. 1085) suggests, Gaza suffers at least as
public-health and public-safety planning, for the robust growth of business and tech- much from geopolitical factors that inhibit access to money
nology, and for research that will be beneficial for society. Such openness is addition- and nearby water as from the “environmental problem” of
ally necessary for the development of countermeasures against sinister applications “running out of water.” Water transfers or desalination help
of science. Preventing publication, even if that could be accomplished, will not pre- overcome local/regional scarcity, but with important enviro-
vent the misuse of science because sanctions will not deter those who have a malev- nmental, social, and economic costs (“Going against the
olent intent. Secrecy instead poses the danger of enforced ignorance. flow,” R. Stone, H. Jia, News Focus, p. 1034; “Desalination
The life-sciences community has generally garnered public trust. To ensure freshens up,” R. F. Service, News, p. 1088). For example,
the continued success of the scientific enterprise, it is critical to maintain and fur- Israel’s water management is becoming “sustainable”
ther that trust against the possibility of public misunderstanding, particularly in (“Seeking sustainability: Israel’s evolving water management
an ever-changing scientific and political environment. To preserve their credibility, strategy,” A. Tal, Perspective, p. 1081) only from a narrow
members of the scientific community must remain sensitive to the potential that technical perspective that treats as exogenous the growth in
information could be misused by individuals and communities to endanger public its arid south and neglects the environmental and third-
safety and health or otherwise jeopardize national security; continuing education party impacts of overexploiting the Jordan River. First-
and responsible engagement in the wider body politic are required. order scarcity metrics (“Global hydrological cycles and
Life scientists enjoy a virtually unrestricted exchange of information; shared world water resources,” T. Oki, S. Kanae, Review,
information has been a safeguard and a cornerstone. But legitimate threats to our p. 1068), especially global ones, overlook such specifici-
national security necessitate that there be appropriate oversight of scientific research ties and are of limited policy use.
and publication. Restraints of the kind set forth by President Reagan in National The interdisciplinary water research community has
Security Decision Directive (NSDD) 189 (1) are fit. However, perfect regulation is shifted its attention to context-specific and proactive
impossible because it assumes perfect compliance. While the scientific community approaches such as watershed management, ecological
continues to accept responsibility for principled research and communication, and engineering, demand management, reallocation, and
regulation as a management tool, the public and the government must recognize that collaborative/adaptive planning (2). We understand that the
true national security requires scientific accomplishment and that scientific excel- Special Section was not meant to be an exhaustive review
lence requires the open communication of research and results. SUSAN A. EHRLICH* of freshwater issues. But institutional, political, and
Judge, Arizona Court of Appeals, 1501 West Washington Street, Phoenix, AZ 85007, USA, and a member of economic options deserve more than cursory mention in
the National Science Advisory Board on Biosecurity. Science, since it is primarily these, rather than technical
*The views expressed are the author’s alone.
fixes alone, that “offer a measure of hope for the future”
(“A thirsty world,” J. Yeston et al., p. 1067).
Reference GIORGOS KALLIS, MICHAEL KIPARSKY,
1. Available at www.fas.org/irp/offdocs/nsdd/nsdd-189.htm. ANITA MILMAN, ISHA RAY

www.sciencemag.org SCIENCE VOL 314 1 DECEMBER 2006 1387

LETTERS

Energy and Resources Group (ERG), University of California at ary time (hundreds of millions of years). It is tinely exploited for conservation purposes.
Berkeley, Berkeley, CA 94720–3050, USA. E-mail: gkallis@
berkeley.edu
rare that conservation biologists are inter- This would not be the case if selective sweeps
ested in how mtDNA diversity is distributed were as dominant a force as implied by Bazin
References at such a level. Rather, it is standard practice et al. Despite their claims, Bazin et al.’s results
1. J. Lundqvist, M. Falkenmark, C. Folke, L. Gordon, L. Ohlsson, that genetic diversity is interpreted in the con- have limited relevance to most standard appli-
New Dimensions in Water Security (FAO AGL/MISC/25/2000, text of a relevant, almost always closely cations of mtDNA in conservation.
UN Food and Agriculture Organization, Rome, 2000).
2. P. H. Gleick, Water Int. 25, 127 (2000).
related, control group (1). This practice is OLIVER F. BERRY
designed to account as best as possible for the School of Animal Biology, University of Western Australia,
3. The Letter was written with input from the ERG Water Group.
Stirling Highway, Crawley, Western Australia 6009, Australia.
potentially confounding historical, demo-
graphic, mutational, and selective variables
Mitochondrial DNA and that influence genetic diversity.
References
1. J. L. Bouzat, Genetica 110, 109 (2000).

Population Size Second, it is well established that the geo- 2. J. C. Avise, Phylogeography: The History and Formation of
Species (Harvard Univ. Press, Cambridge, MA, 2000).
graphical distribution of mtDNA diversity as
3. C. Moritz, Trends Ecol. Evol. 9, 373 (1994).
IN THEIR REPORT “POPULATION SIZE DOES NOT determined by lineage-sorting, and not just 4. P. D. N. Hebert, A. Cywinska, S. L. Ball, J. R. deWaard, Proc.
influence mitochondrial genetic diversity diversity per se, is informative with respect to R. Soc. London B 270, 313 (2002).
in animals” (28 Apr., p. 570), E. Bazin et al. biodiversity conservation (2–4). Use of this cri- 5. M. Turelli, A. A. Hoffmann, Nature 353, 440 (1991).
present compelling evidence that selective terion is recognized to address the very differ-
sweeps occur in animal mitochondrial DNA ences in accumulation or maintenance of genetic IN A META-ANALYSIS OF GENETIC POLY-
(mtDNA) and reduce genetic diversity below diversity within different taxa described by morphism, E. Bazin et al. suggest that mito-
the level expected at mutation-drift equilib- Bazin et al.—otherwise known as the “how chondrial DNA (mtDNA) is more profound-
rium in some taxa. They also assert that this much divergence is enough” question (3). ly affected by nonneutral evolution than
evidence implies that mtDNA has limited Clearly, conservation biologists should nuclear loci (“Population size does not influ-
relevance to biodiversity and conservation not ignore selective sweeps; they do occur, ence mitochondrial genetic diversity in ani-
studies. I contest this claim on two fronts. and sometimes rapidly (5). However, mtDNA mals,” Reports, 28 Apr., p. 570). This inter-
First, the selective sweeps that they detect diversity is abundant at the population, species, pretation has already led some to conclude
occur at very deep phylogenetic levels (phyla and genus level of animals (2), and it is here that mtDNA is of little utility in studies of
to class), which translate into deep evolution- that it can be, and is, most relevant and rou- evolution and conservation. It is well known

1388 1 DECEMBER 2006 VOL 314 SCIENCE www.sciencemag.org

LETTERS
2.0
Neutrality indices simulated for a single nonrecom-

▲
that multiple evolutionary processes must be 1.5 bining 1 kb coding region. For each interspecific
considered in interpreting patterns of genetic distance class, 100 coalescent simulations were
diversity at any gene region (1, 2). However, performed comparing an ingroup taxon of n = 10

NI
1.0 and expected within-species pairwise divergence of
dismissing mtDNA as a more biased analyti-
2% to a single outgroup taxon. Simulations assume
cal tool is neither necessary nor justified (3). a transition:transversion ratio of 2 and a relative sub-
First, it is inappropriate to approximate 0.5
stitution rate of 2:1:20 for the first, second, and third
effective population size (Ne) from census size, codon positions, respectively. Thick horizontal bars
as is implied by Bazin et al.’s “intuitive” pre- 0.0 indicate medians, and boxes include 50% of the
dictions. Bottlenecks, fluctuating population 0 5 10 15 20 25 distributions. The vertical line indicates the cutoff
size, reproductive strategies, and geographic Total % divergence between species point used by Bazin et al. in their meta-analysis.
structure, none of which can be inferred reli-
ably from present census size, profoundly more distant outgroup comparisons. animals—can make to studies of conservation,
impact Ne and genetic diversity (4). Indeed, Bazin et al. rightfully emphasize the neces- taxonomy, and historical demography.
invertebrate taxa and fish generally have sity of adequately testing for deviation from JOHN P. WARES,1 PAUL H. BARBER,2
greater census size than tetrapods, but there is the neutral model for mtDNA, as with all loci. JEFFREY ROSS-IBARRA,1 ERIK E. SOTKA,3
also greater diversity in life history and repro- Further, the meta-analytical tools developed ROBERT J. TOONEN4
1Department of Genetics, University of Georgia, Athens, GA
ductive strategies, traits that alter patterns of by Bazin et al. and others can help assess the 30606, USA. 2Department of Biology, Boston University,
sequence divergence within and among taxa. time scale of selective sweeps relative to Boston, MA 02215, USA. 3Grice Marine Laboratory, College of
Second, the neutrality index (NI) may demographic events commonly considered by Charleston, Charleston, SC 29412, USA. 4Hawai’i Institute of
be inappropriate for distantly related taxa evolutionary biologists (e.g., effects of glacia- Marine Biology, University of Hawai’i at Manoa, Kane’ohe, HI
96744, USA.
because the high substitution rate and site tion, high variance in reproductive success,
heterogeneity of mtDNA often lead to muta- and recent/incipient speciation). All genetic References
tional saturation in protein-coding genes (see data come with complications, but we argue 1. S. Y. W. Ho, M. J. Phillips, A. Cooper, A. J. Drummond, Mol.
figure). This saturation biases the NI toward that it is inappropriate and unnecessary to dis- Biol. Evol. 22, 1561 (2005).
2. A. S. Gerber, R. Loggins, S. Kumar, T. E. Dowling, Annu. Rev.
values <1 as species divergence increases. miss the contribution that mtDNA sequence
Genet. 35, 539 (2001).
The smaller number of invertebrate mtDNA data—still one of the most powerful universal 3. D. Rubinoff, B. S. Holland, Syst. Biol. 54, 952 (2005).
genomes currently available tends to force sources of genetic variation for nonmodel 4. R. Frankham, Genet. Res. 66, 95 (1995).

www.sciencemag.org SCIENCE VOL 314 1 DECEMBER 2006 1389

LETTERS

influenced the level of polymorphism observ- selection, especially in invertebrate species,

able within species. where adaptation might be the rule. The age
Letters to the Editor 2) We agree that effective population size of the most recent mtDNA ancestor, in partic-
Letters (~300 words) discuss material published
can be very different from census population ular, should not be connected to any climatic,
in Science in the previous 6 months or issues of
general interest. They can be submitted through size, to an extent largely variable between geologic, or biotic event unless confirmation
the Web (www.submit2science.org) or by regular species. Our analysis, however, recovers a is obtained from nuclear markers.
mail (1200 New York Ave., NW, Washington, DC positive relationship between nuclear genetic E. BAZIN, S. GLÉMIN, N. GALTIER
20005, USA). Letters are not acknowledged upon diversity and indicators of species abun- CNRS UMR 5171–Génome, Populations, Interactions,
receipt, nor are authors generally consulted before Adaptation–Université Montpellier 2, 34095 Montpellier
dance, indicating that effective and census Cedex 5, France.
publication. Whether published in full or in part, populations sizes are correlated. The lack of
letters are subject to editing for clarity and space.
relationship with mtDNA markers can there-
TECHNICAL COMMENT ABSTRACT
fore hardly be due to the census versus effec-
tive size problem, especially given the much
Comment on “Population Size Does
Response larger data set analyzed.
Berry and Wares et al. independently com- 3) It is true that very distant outgroups can
Not Influence Mitochondrial Genetic
ment on our conclusion that mtDNA might bias the NI analysis because of saturation of the Diversity in Animals”
not be a reliable marker of species popula- synonymous divergence (ds), as neatly demon- Connie J. Mulligan, Andrew Kitchen,
tion size and diversity. They introduce four strated by Wares’simulations. Our data set does Michael M. Miyamoto
arguments: (i) age of selective sweeps, (ii) not show strong variation of mitochondrial dS Bazin et al. (Reports, 28 April 2006, p. 570) found no
census size versus effective size, (iii) dis- across taxa: the average dS is 0.262 in inverte- relationship between mitochondrial DNA (mtDNA) diver-
tance to outgroup, and (iv) the usefulness of brates versus 0.266 in vertebrates. sity and population size when comparing across large
mtDNA despite selective sweeps. 4) We do not mean to argue that mtDNA groups of animals. We show empirically that species
with smaller populations, as represented by eutherian
1) Despite the fact that our study is based markers should be abandoned; there are mammals, exhibit a positive correlation between mtDNA
on comparisons between distantly related taxa, many practical reasons why they can be and allozyme variation, suggesting that mtDNA diversity
the selective sweeps we think have contributed useful. We strongly caution mtDNA users, may correlate with population size in these animals.
to decreased mtDNA diversity in large popula- however, that within-species mtDNA varia- Full text at
tions must be recent ones, because they have tions are likely to be influenced by natural www.sciencemag.org/cgi/content/full/314/5804/1390a

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BOOKS ET AL.
who can “take you to wonderful, unheard-of
EVOLUTION and even amorous worlds” (10). The Other
Insect Societies is a new avatar of the magic
A Subaltern View of Eusociality lamp, complete with a high-power genie. It pro-
vides over 1000 entries in its subject index and
Raghavendra Gadagkar over 2000 in both the taxonomic and author
indices. And in contrast to the contributors to
the Choe and Crespi volume, Costa tells read-

I
n the early 1980s, a group of scholars con- What should we do next? It often helps to
sisting largely of Indian historians set up start from a new perspective. To this end, ers a great deal about the source of his facts—
the Subaltern Studies Group and per- some are offering bold new theoretical who did what, when, why, where, and how.
suaded Oxford University Press, New Delhi, approaches (5–8). But perhaps we also need I doubt that many people would read the
to launch a new publication series, Subaltern fresh data from previously neglected kinds of book from cover to cover or benefit from
Studies: Writings of South Asian History and
Society. Inspired and led by their chief men-
tor, Ranajit Guha, many, now well-known,
historians (among them Gyan Prakash, Gaya-
tri Chakravorty Spivak, Partha Chaterjee,
Shahid Amin, and Gyanendra Pandey) pur-
sued a relatively new brand of historiography
(1). The principal novelty of their approach
was to focus on ordinary people—the
masses, the peasants, and other marginalized
groups. They created a history from “below”
rather than the usual narrative of the kings,
leaders, and other elites. Two decades and ten
volumes later, it is clear that the subaltern
studies have yielded a valuable new perspec-
tive on history, one perhaps especially useful
for understanding and managing present-day
social and cultural problems. Gathering together to gather. In mud puddling, males gather from wet soil supplementary nutrients
In his 1971 book The Insect Societies, needed by the females to produce more eggs. This aggregation of common albatross butterfly (Appias albina)
Edward O. Wilson (2) picked “eusocial- was photographed at the Chinnar Wildlife Sanctuary, Kerala, India.
ity”—a term coined by Suzanne Batra (3)
and given a second lease on life by Charles insect societies. This is the approach James T. doing so. It is more likely that readers who are
Michener (4)—to describe the most orga- Costa offers in The Other Insect Societies. already wedded to specific taxa will devour
nized of animal societies, those in which Costa (the director of the Highlands Biological the chapters on their favorites with pleasure
group members share a composite nest Station, North Carolina, and a professor at and profit. I am rather optimistic that, parallel-
and exhibit cooperative brood Western Carolina University) ing the effects of the subaltern studies of
care, overlap of generations, launches the entomological Indian historians, a focus on other insect soci-
and reproductive castes. Wilson The Other Insect equivalent of subaltern stud- eties will provide valuable fresh perspectives
vested eusociality with such Societies ies, focusing deliberately on useful even for understanding present-day
an elite status that, overnight, by James T. Costa species that have failed to eusocial species.
students of ants, bees, wasps, make it to Wilson’s elite grade Although I found much to praise in the
Harvard University Press,
and termites felt they be- Cambridge, MA, 2006.
of eusociality. book, if I were to write a 100-page review—
longed to a privileged new 811 pp. $59.95, £38.95, Readers will f ind in the and one could envision such a review; after
community of entomologists €55.30. ISBN 0-674-02163-0. book a fascinating wealth of all, it’s a 700-page book—I would probably
ideally poised to solve the information about the obscure devote 90 pages to extol its virtues and some
Darwinian paradox of altru- social lives of earwigs, grass- 10 pages to criticize and disagree with the
ism. They (I should say, we) hoppers, crickets, mantids, cock- author. I would dispute some of his interpre-
have done well: Hundreds of species of roaches, aphids, treehoppers, bugs, thrips, tations, regret his failure to cite certain
eusocial insects have been studied in depth, beetles, caterpillars, sawflies, and even some papers, question some of his assignments of
and we now have a reasonably sophis- non-insect arthropods (spiders, centipedes, priority, and reject his calls to abandon less
ticated understanding of the forces that millipedes, and crustaceans). Costa’s book will entrenched terms (e.g., subsocial, commu-
mold the evolution of insect societies. Nev- inevitably be compared with The Evolution nal) while retaining eusociality. Costa pro-
ertheless, no one would claim that the prob- of Social Behavior in Insects and Arachnids, ceeds at an unduly leisurely pace, which is
CREDIT: E. KUNHIKRISHNAN

lems concerning the evolution of sociality edited by Jae C. Choe and Bernard Crespi made more problematic by the absence of
and altruism are entirely solved. (9)—Wilson and Burt Hölldobler both summaries at the end of individual chapters.
mention that work in their introductory A thematic, rather than taxonomic, treatment
comments on the book. In my review of of the subject matter probably would have
The reviewer is at the Centre for Ecological Sciences,
Indian Institute of Science, Bangalore, 560012, India. the Choe and Crespi volume, I likened it to been more enticing and easier to follow; it
E-mail: [email protected] Aladdin’s magic lamp and the index to a genie might also have allowed Costa to weave the

www.sciencemag.org SCIENCE VOL 314 1 DECEMBER 2006 1391

BOOKS ETAL.

chapters together into a more unif ied I would not claim that what is already research, there is still some hope that we can
account. I would not endorse the claim that known about the non-eusocial insect societies, bring the reading and writing of books back
the “hope for a universal ecological expla- as painstakingly and thoroughly detailed in into fashion among younger biologists.
nation of cooperative breeding may be Costa’s book, makes us substantially wiser
doomed.” Although I have now used my about the evolution of insect social behavior. References
quota of 100 words of criticism and disagree- Instead, I suspect that the book will draw 1. D. Ludden, Ed., Reading Subaltern Studies: Critical History,
ment proportionate with the length of this attention to these other insect societies and Contested Meaning, and the Globalisation of South Asia
(Permanent Black, Delhi, 2005).
review, I cannot ignore the author’s most make their study fashionable and feasible. A 2. E. O. Wilson, The Insect Societies (Harvard Univ. Press,
remarkable statement. After criticizing S. few hours with Costa’s book will bring any Cambridge, MA, 1971).
Mukerji, for not knowing in 1927 that the beginner up to date with a century’s worth of 3. S. W. T. Batra, Indian J. Entomol. 28, 375 (1966).
position of the spinning apparatus and the 4. C. D. Michener, Annu. Rev. Entomol. 14, 299 (1969).
scattered literature on almost everything that
5. E. O. Wilson, B.Hölldobler, Proc. Natl. Acad. Sci. U.S.A.
mechanism of spinning in embiids (webspin- is known about any of the many obscure 102, 13367 (2005).
ners) had already been discovered and pub- groups of insects discussed. One could rea- 6. D. L. Cassill, J. Bioecon. 200, 1 (2006).
lished by M. Rimsky-Korsakov in 1910, sonably expect a new graduate student to read 7. P. Nonacs, K. M. Kapheim, Proc. XV Congr. Int. Union Study
Soc. Insects, Washington, DC, p. 28 (2006); http://iussi.
Costa incredibly goes on to say that “Perhaps the appropriate chapter in the book and confex.com/iussi/2006/techprogram/P1602.HTM.
we should not be surprised at such errors; embark on a study of the corresponding group 8. J. H. Hunt, The Evolution of Social Wasps (Oxford Univ.
after all, these inconspicuous insects long for her dissertation. Press, Oxford, forthcoming).
9. J. C. Choe, B. J. Crespi, Eds., The Evolution of Social Behavior
remained out of reach for most temperate- There is also an altogether different reason in Insects and Arachnids (Cambridge Univ. Press, Cambridge,
zone entomologists.” It seems mind- why I am delighted to see The Other Insect 1997).
boggling that such an invidious statement Societies in print. If an early-career academic 10. R. Gadagkar, Trends Ecol. Evol. 13, 122 (1998).
was written in the first place, let alone that it like James Costa can write a 700-page
passed the scrutiny of referees and editors. account that covers relatively little of his own 10.1126/science.1135094

ENVIRONMENT AND RELIGION

Hoping to Establish Common Ground

for Saving Biodiversity
Steven Bouma-Prediger

E
dward O. Wilson is no stranger to read- proclaimed “secular humanist.” Nonetheless,
ers of Science or the general public. given the scope and pace of ecological degra-
One of the most famous scientists liv- dation, Wilson suggests that Christians and
ing today, Wilson is the author of more than secularists “set aside our differences in order
20 books, two of which have won the Pulitzer to save the Creation.” His new book is, as it
Prize for nonfiction. In his 40 years as a were, an olive branch extended to Christians,
faculty member at Harvard University, he especially conservative Christians in North Worth saving.
has put forward important scientific theories America, to make common cause in the effort
(including island biogeography to preserve biodiversity.
and sociobiology) and coined The Creation Wilson’s argument, in es- problem [of biological catastrophe] would
novel terms (such as biophilia). An Appeal to Save Life sence, is this. His first premise soon be solved.” So writing to his imagined
Through it all, Wilson has been on Earth is that “the Creation—living Baptist pastor, Wilson acknowledges that
an articulate and passionate Nature—is in deep trouble”; “you have the power to help solve a great
advocate for the conservation by Edward O. Wilson indeed, we are facing a “bio- problem about which I care deeply.”
of the natural world. Norton, New York, 2006. logical catastrophe.” Evidence Wilson devotes most of the book to an
Though now retired, Wilson’s 185 pp. $21.95, C$27.50. for this claim runs throughout attempt to persuade his reader to care for the
influence is still considerable. ISBN 0-393-06217-1. the book, from a discussion of planet and its biota. For example, he argues
So any book from him is note- alien species in 16th-century that because we humans are inextricably
worthy. But his The Creation: An Appeal to Hispaniola to the current “pauperization of dependent on a plethora of other species for
Save Life on Earth is certain to draw added Earth” evident in tropical rainforests. We face a our very survival, “even the most recalcitrant
attention, not least because the book is written stark choice: either “conserve Earth’s natural people must come to view conservation as
as a personal letter to a hypothetical Baptist heritage, or let future generations adjust to a simple prudence in the management of
CREDITS: NASA, JUPITER PHOTOS

pastor. Although raised in Alabama as a South- biologically impoverished world.” Earth’s natural economy.” In addition to self-
ern Baptist, Wilson long ago gave up that The second premise, and the reason for interest, however, Wilson insists that each
faith; he is now and has long been a self- this particular book, is Wilson’s belief that species is “a masterpiece of biology, and well
“religion and science are the two most power- worth saving.” He further argues that many
ful forces in the world today,” and thus “if reli- organisms, such as the pitchfork ant, evoke
The reviewer is at the Department of Religion, Hope
College, 126 East 10th Street, Holland, MI 49423, USA. gion and science could be united on the com- wonder and that such wonder motivates care.
E-mail: [email protected] mon ground of biological conservation, the Moving still farther beyond prudence, Wilson

1392 1 DECEMBER 2006 VOL 314 SCIENCE www.sciencemag.org

BOOKS ETAL.

NOTA BENE: EXHIBITS Invaders technique intact after a quarter century of abstinence and to
achieve rare tactical superiority over the next generation.
Delightful Digital Diversions As the hardware continued on the path to increasing portability
and invaded the home, games themselves diversified in several direc-
rom Space War! to Nintendo DS Lite, computer and video gaming tions—action, puzzle, simulation—and evolved into the vast array on

F is now venerable enough to have an entire exhibition devoted to its today’s market. Because of the limits of space, Game On samples this
history (Game On)—which is a bit of a shock in itself. The first diversification sparingly, but nonetheless instructively. Even though
game in the exhibition at London’s Science Museum, Space War!, was some old favorites are inevitably missing, the narrative provided by
designed by Steve Russell and friends at the Massachusetts Institute of the generous graphics fills in the gaps. The artwork is by the British
Technology in the distant days of 1962. The equip- illustrator Jon Burgerman; its centerpiece is a giant
ment required to play that game (see photograph) is Game On timeline charting the history of gaming in the form of a
arresting: The computer alone (a PDP-1, donated to fantastical segmented creature spread along the entire
curated by Conrad Bodman
MIT by the Digital Equipment Corporation in the hope and Lucian King for the wall of the first hall.
that the students would “use it productively”) occupies Barbican Art Gallery The exhibition is perhaps not as much about the sci-
the space of several wardrobes, there is a cumbersome ence of computer gaming as about how technology and
Science Museum, London,
monitor with a 2-foot tube, and it’s all hooked up to an design, graphic arts and characterization have inter-
through 25 February 2007.
electric typewriter. This bulky construction is the only www.sciencemuseum.org. acted to give life to one of the major social phenomena
true anachronism in the show. uk/exhibitions/gameon/ and wealth generators of the post-industrial world. In
Three of us—one from the pre-gaming generation the United Kingdom alone, more than half the popula-
and two 12-year-old connoisseurs of the hand-held tion (with an almost even gender split) regularly uses
console—went to review almost half a century of com- electronic games. The enhancement of 3D skills and the
puter game evolution. The exhibition, sponsored by hand-eye-brain coordination that these games allegedly
Nintendo, is displayed in a series of rooms in roughly offer have put gaming on the compulsory training
chronological order, and it beeps, buzzes, and flashes schedules of pilots in some air forces. The pervasive
like an arcade. Most of the exhibits are fully opera- influence of computer games and debates on their pos-
tional, allowing the visitor hands-on experience. (The sible wider social effects (e.g., violence, loss of social
exception, frustratingly, is the hand-held equipment, skills, sexism, and ethnicism) are tentatively explored in
much of which is displayed beneath a glass dome.) two sections of the exhibition devoted to games culture
From the PDP-1, one rapidly moves into more in the United States, Europe, and Japan.
familiar territory. The next phase of miniaturization was accomplished Where next? The exhibition ends with questions on the future of
within little more than a decade. The game machines of the 1970s, computer gaming. In the coming months a number of ancillary
housed in the fruit-machine cabinets and knee-high tables of arcades events and talks, some by games pioneers, are scheduled at the Sci-
and bar rooms, have been the stock in trade of the industry ever since. ence Museum, to explore and develop these and other themes of
Despite minimal instructions, our two 21st-century gamers had no the exhibition.
trouble familiarizing themselves with the games of that decade. And, it –Andrew M. Sugden, Hugh A. Russell, Rowan M. A. Sugden
was gratifying to the oldest member of the team to discover his Space 10.1126/science.1137012

also asserts that “the Creation, whether you to save biodiversity. In this regard Wilson per- other Christian) readers to care for Earth.
believe it was placed on this planet by a single ceptively criticizes a naïve faith in technology Absent any more explicitly religious argu-
act of God or accept the scientific evidence as the supposed savior of all that ails us. ments, I’m not at all sure his hypothetical
that it evolved autonomously during billions The book, however, has some serious reader will be convinced.
of years, is the greatest heritage, other than the shortcomings. First, the term “the Creation” is Notwithstanding the above criticisms, The
reasoning mind itself, ever provided to a religious term that logically implies a Cre- Creation is an important book. At a time when
humanity.” This heritage, he insists, we ator. In his use of this term, does Wilson intend there is much evidence that Christians are tak-
humans are duty-bound to preserve. to imply the existence of God? Also, Wilson ing their biblical call to be earthkeepers more
There is much to admire in this book. Wil- seems to use “the Creation” interchangeably seriously, Wilson offers an irenic invitation to
son is at his best when describing the natural with “living Nature” and “Earth,” but it is far form “an alliance for life.” Christians have
world in all its wondrous detail. The expertise from clear that these all mean the same thing. every good reason to join with Wilson and
of this world-class biologist is evident. Wilson Furthermore, Wilson speaks often of “reli- meet on common ground—both the beliefs
is also persuasive in communicating a sense of gion and science” when what he means is we share and the Earth we are called to serve
urgency about the looming ecological crisis. Christianity and biology. Few would hold that and protect (Genesis 2:15). At a time when
His feel for the biological vital signs of our religion can be reduced to Christianity or nat- “life on this planet can stand no more plunder-
home planet rings tellingly true. Lastly, Wil- ural science reduced to biology. In addition, ing,” may we so meet.
CREDIT: ANDREW SUGDEN

son honestly confronts what he calls “the Wilson’s conceptions of science, religion, and
ignorance and self-absorption” of humanity. their interrelationship are problematic. Is it References
He dismantles the claims of those, both secu- really the case that “without science there had 1. E. O. Wilson, On Human Nature (Harvard Univ. Press, Cam-
bridge, MA, 1978).
lar and religious, who either see no harm in to be religion”? Lastly, at the end of the book, 2. B. Hölldobler, E. O. Wilson, The Ants (Harvard Univ. Press,
ecological degradation or believe that zoos, one wonders whether the arguments Wilson Cambridge, MA, 1990).
aquariums, and botanical gardens can suffice puts forward will persuade his Baptist (and 10.1126/science.1135704

www.sciencemag.org SCIENCE VOL 314 1 DECEMBER 2006 1393

POLICYFORUM
SCIENCE AND LAW
Patents should not be used to protect laws of

When Patents Threaten Science nature, products of nature, or mathematical

formulas.

Lori Andrews,1* Jordan Paradise,2 Timothy Holbrook,1 Danielle Bochneak1

W hat if each generation of scien-

tists was forbidden to use—or
even think about—the theorems,
principles, and natural phenomena that had
been discovered or proven by the previous
Patents on Scientific Building Blocks and
Processes
The USPTO has issued various patents that
could interfere with the work of basic scien-
tists, social scientists, and engineers. These
eling salesman problem with a computer pro-
gram that used a standard statistical algo-
rithm outputting a set of optimal data points
given certain inputs and constants (11).
Although the inventor includes superficial
generation of scientists? Researchers may range from correlation patents, such as the language referencing a machine, what is actu-
soon find themselves in that position as the one in the Metabolite case, to patents on cer- ally claimed is the first step to solving any
U.S. Patent and Trademark Office (USPTO) tain ways of analyzing data. Patents can chill optimization or linear programming problem.
comes dangerously close to issuing patents research if the patent holder forbids other The patent holder can, until the patent expires
on the basic building blocks of science itself. researchers from using the scientific fact or in 2021, demand a royalty from any industrial
A U.S. Supreme Court decision in June natural phenomenon, or charges an exces- engineer, facilities planner, telecommunica-
2006, Laboratory Corporation v. Metabolite sive fee for access to that knowledge. tions analyst, or other researcher who uses
Laboratories (1), and a solicitation by the One patent claims the use of a computer to this algorithm with computer assistance.
USPTO in July 2006 for comments on pro- derive a solution to any optimization algo- Another patent claims “a method of psy-
posed guidelines for patent examiners (2–4) rithm. Optimization problems have tradition- chological testing of a person, comprising:
have raised questions about the delicate bal- ally been expressed in terms of the hypotheti- (a) instructing the person to produce a draw-
ance between a common body of knowledge cal traveling salesman who has to travel his ing which includes at least one pictorial rep-
and the exclusive rights over scientific infor- route with the minimum expenditure of time resentation of each of at least a majority of
mation embodied in a patent. and money. Commentators expressed the the following items: a hand, an eye, a tree,
The patent at issue in the Metabolite case opinion that no one would ever attempt to a fish, a star, a spiral; a half-circle, and a
covered the following process: Use any test patent such an obvious and important method zigzag; and (b) subjecting to psychological
(whether patented or unpatented) to measure of problem-solving (10). But in 2005, a patent interpretation the drawing produced in
the level of the amino acid homocysteine in a was issued for the process of solving the trav- response to step (a)” (12). Although the
body fluid and then, if the level is elevated patent mentions specific pictures, a cognitive
above the norm, conclude that vitamin B defi- science researcher who substitutes his own
ciency is likely. The Court of Appeals for the drawings may be found liable of infringe-
Federal Circuit held that LabCorp induced Current ment under the doctrine of equivalents (13).
infringement of that patent (and thus was liable The patent thus covers a basic psychological
for over $2 million in damages) based on the research evaluation technique.
publication to physicians of a law of nature— The patent entitled “[d]atabase and sys-
the relation between levels of homocysteine tem for storing, comparing and displaying
and vitamin deficiency (5). Astonishingly, the genomic information” encompasses the
Federal Circuit also held that physicians (or very manner in which a computer user may
researchers) would infringe the patent merely access genomic libraries for viewing (14).
by thinking about the relation between homo- Included in the claims is the “method of
Magnetic field
cysteine and vitamin deficiency when they comparing genetic complements of differ-
analyzed an alternative homocysteine test (5). ent types of organisms” by means of
As the Supreme Court contemplated the NONPATENTABLE? electronic sequence libraries (14). Such
merits of the Metabolite case, legal scholars a broad patent may restrict meaningful
wrote commentaries (6, 7), and major news- access to and analysis of genetic sequence
papers ran editorials (8, 9) addressing the information that would otherwise be freely
PHOTO CREDIT: NATIONAL MUSEUM OF AMERICAN HISTORY

problems in the current interpretation of available to researchers who wish to com-

patentable subject matter by the USPTO and pare, for example, the genes of mice to the
federal courts. In June 2006, the Supreme genes of humans.
Court dismissed the appeal for procedural Patents that claim the correlation be-
reasons (1), which allowed this patent on a tween the existence of a genetic mutation
biological fact to stay in effect. and the predisposition to a disorder are also
problematic (15). One patent claims the
1Institute
process of assessing a patient’s risk of
for Science, Law, and Technology, Chicago-Kent
College of Law, Chicago, IL 60661, USA. 2Consortium on PATENTABLE? developing certain neurological or neu-
Law and Values in Health, Environment, and the Life ropsychiatric disorders based on the pres-
Sciences, University of Minnesota Law School, Minne- What can be patented? (Top) Natural relationship ence of specific polymorphisms (16).
apolis, MN 55455, USA. However, mutations are natural occur-
between magnetic field and electrical current.
*Author for correspondence. E-mail: [email protected] (Bottom) Samuel Morse’s telegraph machine. rences and, if patients have the mutation,

www.sciencemag.org SCIENCE VOL 314 1 DECEMBER 2006 1395

POLICYFORUM

they necessarily have the predisposition. netic material, the Court actually distin- result” (23). However, this is clearly over-
Similar to the patent at issue in Metabolite, guished between a product of nature and a inclusive and in direct conflict with exist-
a patent covering a natural correlation patentable genetically modified bacterium ing Supreme Court precedent (1). To be
could apply to researchers who study the cell that did not exist in nature. The Court patentable, there must be something more—
mutation and its effects, or who design tests reiterated that “a new mineral discovered a human invention that produces a result
aimed at targeting the mutation, or who in the ear th or a new plant found in beyond the law of nature or product of
even think about this relation. the wild is not patentable…. Likewise, nature itself.
Einstein could not patent his celebrated
Ignoring Supreme Court Precedent law that E = mc2; nor could Newton have Conclusion
A close look at patent policy and U.S. patented the law of gravity. Such discover- Scientists may not have paid sufficient
Supreme Court jurisprudence—as well as ies are ‘manifestations of . . . nature, free attention to the privatization of common
an understanding of the nature of the scien- to all men’” (21). knowledge because, in the past, they felt
tific enterprise—provides a foundation for Even if a patent applicant exercised con- that research activities did not require
assuring that the laws of nature and products siderable innovation discovering a law of approval from patent holders. The 2002
of nature remain freely available to all. Not nature or product of nature, neither is Madey v. Duke decision put an end to such
every discovery or innovation is entitled to patentable under existing Supreme Court protection (24). Scientists can be influen-
patent protection. U.S. patent law dictates precedent. A person might expend money tial by helping policy-makers understand
that patent applicants must satisfy a number and creativity building a telescope, but he that open access to basic laws of nature,
of requirements in order to be issued a patent should not be able to patent the new planet products of nature, and mathematical for-
by the USPTO. An invention must be of eli- he discovers through the telescope. mulae is necessary for scientists to explore
gible subject matter (17). Justices Breyer, Stevens, and Souter, dis- and innovate. The U.S. Supreme Court has
In the 1854 O’Reilly v. Morse case, the senting in the Metabolite case, said: “The recognized that fact, but, increasingly, the
U.S. Supreme Court expressed its concern justification for the principle does not lie in USPTO in granting such patents and the
that granting Samuel Morse broad rights to a any claim that ‘laws of nature’ are obvious, Federal Circuit in upholding them seem to
law of nature, beyond its particular applica- or that their discovery is easy, or that they are have forgotten it.
tion (the telegraph), would afford Morse the not useful. To the contrary, research into
References and Notes
right to exclude others from making new such matters may be costly and time-con- 1. 126 S. Ct. 2921 (2006).
innovations that Morse himself did not suming; monetary incentives may matter; 2. “Request for comments on interim guidelines for exami-
invent or even contemplate. Accordingly, and the fruits of those incentives and that nation of patent applications for patent subject matter
eligibility,” Fed. Reg. 70, 75451 (2006).
the Court stated that Morse’s claim to “a research may prove of great benefit to the 3. Fed. Reg. 71, 34307 (2006).
monopoly in [electro-magnetism’s] use, human race. Rather, the reason for the exclu- 4. See also “Interim guidelines for examination of
however developed, for the purpose of print- sion is that sometimes too much patent pro- patent applications for patent subject matter eligibility,”
ing at a distance” was “too broad, and not tection can impede rather than ‘promote the Off. Gaz. Pat. Office 1300, 142 (15 November 2005).
5. Metabolite Labs., Inc. v. Lab. Corp. of Am. Holdings,
warranted by law” (18). The Court explained Progress of Science and useful Arts,’ the 370 F. 3d 1354 (Fed. Cir. 2004).
that patent law did not support overly broad constitutional objective of patent and copy- 6. R. S. Eisenberg, Nat. Biotechnol. 24, 317 (2006).
patent rights to scientific principles because right protection” (1). 7. L. Andrews, Chron. Higher Educ. 52, (24), B20
(17 February 2006).
such monopolies “would be unjust to the The idea that a patent could block future 8. Wall Street Journal, 1 March 2006, p. A14.
public…and defeat the manifest object of innovation, to the detriment of the public, is 9. New York Times, 22 March 2006, p. A24.
the law” (18). pertinent because the USPTO is granting 10. B. Klemens, Math You Can’t Use (Brookings Institution
The Supreme Court continued to police patents that could block scientific inquiry. Press, Washington, DC, 2006), pp. 46–61.
11. U.S. Patent 6,904,421 (2005).
the line between invention and scientific Although the discoveries of natural phe- 12. U.S. Patent 5,190,458 (1993).
principle in Parker v. Flook, rejecting a nomenon may be necessary precursors to 13. Graver Tank & Mfg. Co. v. Linde Air Products Co.,
patent that claimed a method for calculating invention, improperly tying up these dis- 339 U.S. 605, 609 (1950).
14. U.S. Patent 5,966,712 (1999).
updates in the catalytic conversion process coveries with patent rights will only drive 15. J. Paradise et al., Science 307, 1566 (2005).
as merely a mathematical formula (19). up the costs of such subsequent innova- 16. U.S. Patent 6,660,476 (1999).
The Court reasoned that such a scientific tions, if not thwart them altogether. 17. 35 U.S.Code § 101.
18. O’Reilly v. Morse, 56 U.S. 62 (1854).
principle, though useful, simply “reveals The USPTO and lower courts are respon- 19. Parker v. Flook, 437 U.S. 584 (1978).
a relationship that has always existed” (19). sible for granting and enforcing patent rights 20. Gottschalk v. Benson, 409 U.S. 63, 68 (1972).
Likewise, the Supreme Court in Gottschalk v. that run contrary to U.S. Supreme Court 21. Diamond v. Chakrabarty, 447 U.S. 303 (1980).
Benson held that a claim to the conversion of precedent (22). Merging the U.S. Court 22. S. Merrill et al., Eds., A Patent System for the 21st
Century (National Research Council, National Academies
numerical data into binary code in any type of Claims and the U.S. Court of Customs Press, Washington, DC, 2004), pp. 25–27.
of general-purpose digital computer was and Patent Appeals to create the Federal 23. State Street Bank & Trust Co. v. Signature Fin. Group 149
unpatentable because it was “so abstract and Circuit in 1982 seems to have accelerated F. 3d 1368, 1373 (Fed. Cir. 1998), cert. denied 525 U.S.
1093 (1999).
sweeping” that it was an attempt at patenting this expansion by creating a specialized, 24. Madey v. Duke University, 307 F. 3d 1351, 1362
an idea rather than an inventive process (20). arguably pro-patent court. (Fed. Cir. 2002).
In 1980 the Supreme Court handed Patent applicants who seek to patent laws 25. Funded by the Robert Wood Johnson Foundation
down a seminal decision in Diamond v. of nature often point to a Federal Circuit Investigator Awards in Health Policy Research Program;
the Office of Science, U.S. Department of Energy under
Chakrabarty (21). Often mischaracterized opinion, State Street Bank & Trust Co. v. award number DE-FG02-06ER64276; and an NSF grant,
as opening the door for patents claiming Signature Financial Group, which suggests SES 0508321.
isolated and purified versions of naturally that a law of nature is patentable if it
occurring products, including human ge- produces a “useful, concrete, and tangible 10.1126/science.1135872

1396 1 DECEMBER 2006 VOL 314 SCIENCE www.sciencemag.org

PERSPECTIVES
DEVELOPMENTAL BIOLOGY

The Turing Model Comes of Molecular analyses of hair follicle formation

provide evidence to support the most well-
known mathematical model for biological
Molecular Age pattern formation.

Philip K. Maini, Ruth E. Baker, Cheng-Ming Chuong

W hat are the underlying mecha-

nisms that give rise to complex
patterns in biology? Despite
recent advances in biotechnology and mathe-
matical modeling, this still remains a largely
amenable to experimental manipulation.
Nagorcka was the first to propose the Turing
model to explain hair pattern formation (4),
but at that stage the molecular biology was
lagging behind the theory. It was only in
The model predicts that moderate overex-
pression of activator (WNT) increases fol-
licular density, whereas moderate overex-
pression of inhibitor (DKK) during the ini-
tial inductive wave increases the interfollic-
open question. As reported on page 1447 of 1998 that Jung et al. made the first efforts to ular spacing. Sick et al. have verified these
this issue, Sick et al. have made a major link known molecular morphogens with a predictions experimentally, providing strong
advance toward answering this question by reaction-diffusion mechanism for feather evidence for a genetic underpinning of a
identifying key molecular players in hair fol- germ formation (5). They showed how the Turing reaction-diffusion model.
licle growth and by confirming the validity size, number, and distribution of appendages Together the papers of Jung et al. and
of perhaps the best-known mathematical could be modulated by altering morphogen Sick et al. show that the skin progenitors are
model for biological pattern formation (1). concentrations (6). stem cells, in that they are multipotent and
In a seminal paper, Alan Turing proposed Sick et al. investigated the regulation of may assume appendage or interappendage
that spatial patterns result from a phenome- hair follicle patterning in developing murine fates depending on the local chemical envi-
non he termed “diffusion-driven instability” skin. They propose that the protein WNT ronment at the time of specification. In this
(2). He showed mathematically that small and its inhibitor DKK are morphogens in the sense, the molecular components identified
spatial fluctuations in an otherwise well- Turing sense. Expression of the protein by these experiments appear to be acting as
mixed system of reacting and diffusing chem- Dkk1, which inhibits WNT, is actually con- morphogens in the true Turing sense.
icals could become unstable, and that ampli-
fication of these fluctuations could lead to a Activator concentration Inhibitor concentration Cell density Chemical concentration
10 10
spatial pattern of chemicals that he termed
morphogens (i.e., substances that stimulate
the development of form or structure in an 0 0
y

y
organism). He proposed that this spatial
arrangement could serve as a prepattern for
development. Turing’s work was ground- −10 −10
−10 0 10 −10 0 10 −10 0 10 −10 0 10
breaking because the mathematical nature of x x x x
the resulting patterns is wholly counterintu-
Biological pattern formation. Two mechanisms can show similar results. (Left) Outcome of a reaction-
itive; since their discovery, they have moti- diffusion model (7) in which activator and inhibitor react and diffuse. Small random fluctuations in the ini-
vated much mathematical research. However, tial field lead to coinciding spatial patterns of activator and inhibitor concentration. (Right) Results of a cell
the model has been the subject of controversy chemotaxis model (9) in which cells and chemical both diffuse, with cells also moving up gradients in chem-
because it has been deemed too simplistic and ical concentration. Again, small random fluctuations in the initial field lead to coinciding spatial patterns in
the search for real biological examples has cell density and chemical concentration. Blue indicates low concentration levels; red indicates high levels.
been neglected. Moreover, although diffu-
sion-driven instability has been shown to be trolled by secreted WNTs, and both WNTs In principle, a reaction-diffusion model
present in chemistry, there is substantial evi- and DKKs are secreted into the extracellular can set up a chemical prepattern before we
dence in the fruit fly Drosophila to refute the space where they diffuse, thereby acting can visualize changes in cell distribution.
model for biology (3). The report by Sick et over longer distances. Given that the WNT That is, it determines sites at which cells will
al., by providing the first compelling biologi- proteins are substantially larger than the cluster: Regions of high cell density coin-
cal evidence for the Turing model, is thus a DKKs, one would expect a large difference cide with those of increased morphogen
landmark publication. in their rates of diffusion. This makes possi- concentration—although the model does not
The formation of skin appendages (hairs, ble the classical “short-range activation, specify how this rearrangement occurs. On
feathers, etc.) is an excellent paradigm long-range inhibition” phenomenon that the other hand, it is possible for cellular
for patterning because these systems are underlies diffusion-driven instability (7). aggregations to form without such a prepat-
Because hair follicle patterning occurs in tern via simple chemotactic movement in
waves, the authors used a reaction-diffusion response to gradients in chemical concentra-
CREDITS (TOP TO BOTTOM):

P. K. Maini is at the Center for Mathematical Biology,

University of Oxford, Oxford OX1 3LB, UK, and the Oxford
model to set up an initial pattern of follicles. tion. By way of illustration, the patterns
Center for Integrative Systems Biology, University of Then, along the same lines as Mooney and formed by these two different mechanisms
Oxford, Oxford OX1 3QU, UK. R. E. Baker is at the Center Nagorcka (8), they assumed these follicles are shown in the figure. It is immediately
for Mathematical Biology, University of Oxford, Oxford to be chemical sources giving rise to a sec- obvious how similar such patterns are.
OX1 3LB, UK. C.-M. Chuong is in the Department of
Pathology, University of Southern California, Los Angeles, ond wave of hair follicle formation on a This highlights one of the difficulties in
CA 90033, USA. E-mail: [email protected] larger domain (due to the growth of skin). mathematical modeling: determining which

www.sciencemag.org SCIENCE VOL 314 1 DECEMBER 2006 1397

PERSPECTIVES

is the “correct” model. Now that WNT and example of the Turing model in biology. (10.1126/science.1130088).
2. A. M. Turing, Philos. Trans. R. Soc. London Ser. B 237, 37
DKK have been identified as possible mor- Turing models have been proposed to (1952).
phogens, this issue can be addressed experi- describe other types of patterns observed 3. M. Akam, Nature 341, 282 (1989).
mentally. The key requirement, then, is that in developmental biology. Two applications 4. B. N. Nagorcka, Biosystems 16, 323 (1983–1984).
5. H.-S. Jung et al., Dev. Biol. 196, 11 (1998).
the results of such experiments are used to currently receiving much attention from 6. T.-X. Jiang, H.-S. Jung, R. B. Widelitz, C.-M. Chuong,
test and refine models, ruling some out if the experimentalists are pigmentation patterning Development 126, 4997 (1999).
data allow us to do so. The WNT-DKK inter- in fish and skeletal development in the 7. A. Gierer, H. Meinhardt, Kybernetik 12, 30 (1972).
8. J. R. Mooney, B. N. Nagorcka, J. Theor. Biol. 115, 299
action does appear to be qualitatively of the mouse limb. Although the evidence for a (1985).
form necessary for a Turing-type system, but Turing diffusion-driven instability in these 9. M. R. Myerscough, P. K. Maini, J. D. Murray, K. H. Winters,
it is now imperative that we try to overcome systems is not as strong as that presented by in Dynamics of Complex Interconnected Biological
the experimental challenges in measuring Sick et al., their report should stimulate fur- Systems, T. L. Vincent, A. I. Mees, L. S. Jennings, Eds.
(Birkhäuser, Boston, MA, 1990), pp. 65–83.
key parameters (rates of production, decay, ther work in biological pattern formation. 10. Supported by Research Councils UK, Lloyds Tercentenary,
diffusion coefficients, etc.) so that quantita- Microsoft Corporation, and St. Hugh’s College, Oxford
tive tests can be performed to determine (R.E.B.) and by NIH (C.-M.C.).
References and Notes
whether the system actually is of Turing 1. S. Sick, S. Reinker, J. Timmer, T. Schlake, Science 314,
type. This would then be the first definitive 1447 (2006); published online 2 November 2006 10.116/science.1136396

CREDIT: (LEFT) NASA/CHANDRA X-RAY OBSERVATORY CENTER/MIT; (RIGHT) NATIONAL RAIDO ASTRONOMY OBSERVATORY; (RIGHT) NASA/SPACE TELESCOPE SCIENCE INSTITUTE/UNIVERSITY OF MARYLAND
ASTRONOMY

Variable High-Energy γ Rays High-energy γ rays emanating from intense jets

of matter that are associated with certain
from the Elliptical Galaxy M87 galaxies provide clues to jet formation.

A. C. Fabian

A lmost 90 years ago, astronomer

Heber Curtis recorded the presence
of a “curious straight ray” con-
nected to the nucleus of the giant elliptical
galaxy M87. Since then, researchers have
may reveal new details of how the emissions
from this galaxy are powered and how the jet
is created (1).
The first hints of highly energetic teraelec-
tron-volt (TeV) emission from M87 were
close to the black hole. In the optical images,
there is a peculiar knot about 100 pc (1 pc =
3.26 light years) along the jet (see right panel
of the figure) where some slow variations
have been seen, but this is unlikely to be the
acquired high-resolution images of this reported by the High Energy Gamma Ray source of the TeV photons because it would
famous jet at wavelengths from the radio to Astronomy (HEGRA) collaboration in 1998 (2). require the jet to be unreasonably tightly colli-
x-ray bands (see the figure). In these images, Since 2003, regular observations of M87 have mated there.
the jet appears on only one side of the been made by the High Energy Stereoscopic Variable TeV γ emission has been seen
galaxy nucleus because it is moving in our System (H.E.S.S.), sited in Namibia (3). from other galactic nuclei with jets known as
direction at very close to the speed of light; Aharonian et al. now find evidence of fast varia- blazars. In these objects, we are looking

Chandra x-ray VLA radio HST optical

Energetic jet. The M87 jet imaged at x-ray wavelengths by the Chandra spacecraft (left), at radio wavelengths by the Very Large Array (middle), and at optical wave-
lengths by the Hubble Space Telescope (right). The view of each panel is 32 arc sec by 21 arc sec. Total length of the arc is 2000 pc.

the side pointed away from us is so dim as tions in the TeV γ emission from the source in more or less straight down the jet, which rel-
to be invisible. Rapid motion of nearby H.E.S.S. observations made during a bright ativistically boosts both the energy of the
gas and stars reveals that the central engine in phase of the jet in 2005. emission and its observed intensity. For typi-
this energetic nucleus is a massive black The high-energy γ emission varies on a cal conditions, if the jet from M87 were a
hole. On page 1424 of this issue, Aharonian time scale of about 1 day, which is comparable blazar we would have to be observing the jet
et al. now report observations of M87 at the to the time it takes light to cross the black hole within an angle of about 6°. However, the
highest energies of the γ-ray band, which and is therefore the shortest natural time scale M87 jet is generally considered to be point-
of the system. This is about 10 times as fast as ing at 30° to 40° or so away from our line of
The author is at the Institute of Astronomy, Madingley Road, variations seen from M87 at any other wave- sight, which puts us out of the extreme blazar
Cambridge CB3 0HA, UK. E-mail: [email protected] length, which points to an origin for the γ rays situation. Measuring the angle with confi-

1398 1 DECEMBER 2006 VOL 314 SCIENCE www.sciencemag.org

PERSPECTIVES

dence is difficult, but evidence for apparent In terms of energy output, the TeV emis- prime targets for these instruments, but objects
faster-than-light motion in the jet seen in sion in M87 is only a few times 1040 erg s–1, such as M87 and other radio galaxies where the
Hubble Space Telescope images (4) does whereas the jets probably have a mechanical jet is not pointing directly our way will make for
support a smaller value of <20°. However, power 100 to 1000 times as large, as indi- interesting observing. If all goes well, we shall
the more the jet points directly at us the cated by the bubbles created by them in the understand how the jet in M87 operates before
longer it must really be: We see a projected surrounding hot gas (7). Therefore, TeV the centenary of its discovery takes place.
length of about 2 kpc, and if it is at 20° it emission could be a minor energy loss from
must then be 6 kpc long. It would be surpris- the jet, unless much more TeV power is References
1. F. Aharonian et al., Science 314, 1424 (2006). Published
ing if one of the nearest powerful jets were beamed out of our line of sight. Even if it is online 26 October 2006; 10.1126/science.1134408.
pointed almost directly at us. All of this minor in power, it may still be of great impor- 2. M. Beilicke et al., New Astron. Rev. 48, 407 (2004).
argues against M87 being a blazar. tance if it is our only direct probe of the 3. F. Aharonian et al., Astron. Astrophys. 403, L1 (2003).
4. J. A. Biretta, W. B. Sparks, F. Macchetto, Astrophys. J.
What mechanism accelerates matter and acceleration region. 520, 621 (1999).
the resulting radiation to TeV energies is not NASA’s Gamma Ray Large Area Space 5. R. D. Blandford, R. L. Znajek, Mon. Not. Roy. Astron. Soc.
clear. Indeed, the details of how a relativistic Telescope (GLAST) should be launched in less 179, 433 (1977).
jet is produced and accelerated remain a than a year, detecting γ rays up to 0.3 TeV, 6. R. L. Znajek, Mon. Not. Roy. Astron. Soc. 185, 833
(1978).
mystery, despite many years of study. Jet out- which nicely complements the ground-based 7. A. J. Young, A. S. Wilson, C. G. Mundell, Astrophys. J.
flows are commonly found where matter Cerenkov telescopes like H.E.S.S. that operate 579, 560 (2002).
accretes via a disc onto a central object, at higher energy. Blazars will be among the 10.1126/science.1136199
whether it be a star or a black hole. The speed
of the outflow appears roughly proportional
to the escape velocity from the object. The ATMOSPHERE
radio and optical emission in the M87 jet is
polarized and is therefore synchrotron radia-
tion that is produced when energetic elec-
trons (or positrons) spiral around magnetic
When Dry Air Is Too Humid
fields. Sufficiently energetic electrons then Thomas Peter, Claudia Marcolli, Peter Spichtinger, Thierry Corti, Marcia B. Baker, Thomas Koop
scatter the synchrotron photons up to TeV
energies, particularly when highly beamed, Analyses of upper tropospheric humidity are forcing reassessment of how ice clouds form.
as in a blazar. Whether such jets also contain
substantial numbers of protons is uncertain,
as is whether protons can produce the
observed hard spectrum of high-energy γ
rays observed from M87.
An exciting possibility is that the TeV
A s moist air rises to colder regions in
the atmosphere, the humidity rises
above its equilibrium value over ice.
To relax this metastability, the air releases its
water vapor via ice cloud formation. Such
tion above a critical threshold value.
Nucleation can occur homogeneously from
aqueous solution droplets, or heteroge-
neously on particles known as ice nuclei. At
upper-troposphere temperatures, homoge-
emission comes from between the black hole atmospheric ice clouds form in two steps: neous freezing sets in at a supersaturation of
and the inner end of the radio jet, that is, from First, ice nucleates in or on existing aerosol ~60% (4); lower supersaturations are suffi-
the acceleration zone. The jet can be traced particles; second, these ice particles grow cient for heterogeneous nucleation. After
down to within a few hundreds of times the through condensation of supersaturated water nucleation, vapor molecules condense onto
radius of the event horizon of the black hole vapor onto the ice surfaces. Recent field the ice particles, causing them to grow and
in very-high-resolution radio images, mean- observations (1–3) call into question the the gas phase to become depleted in water
ing that the acceleration happens close to the basic principles underpinning the current until equilibrium is reached (see the green
black hole. It is often speculated that jets are understanding of ice cloud formation and curves in the figure).
accelerated and collimated by magnetic alter the assessment of water distribution in Large-scale regions of persistent super-
fields brought in with matter accreting onto the upper troposphere. saturation up to 60% outside ice clouds are
the black hole. Close to the black hole, the jet The governing quantity for nucleation not unexpected in the absence of ice nuclei.
may be completely dominated by magnetic and growth is the excess activity relative to Yet values even above 100% have been
fields. The particles necessary for us to see the equilibrium humidity over ice, also observed in cloud-free regions (1) (red curves
the jet are either created (as electron-positron called ice supersaturation (expressed as in the figure). These values are far above the
pairs) or picked up further along the jet from a percentage). The equilibrium humidity critical value for homogeneous ice nucle-
the accretion flow and from the interstellar decreases strongly with falling temperature. ation (5) or cloud chamber data (6).
medium of the host galaxy. A black hole does Hence, when an ascending air mass cools, it At least as puzzling are supersaturations
not have an intrinsic magnetic field, but a rap- can become supersaturated with respect to of 30% reported to persist inside ice clouds
idly spinning black hole surrounded by mag- ice. Ice nucleation requires a supersatura- and contrails (2) for at least 1 hour of aircraft
netic fields can drag the fields around with measurement time (3) (orange curves in the
it (5) and, as in a pulsar, can in principle gen- figure). Such large supersaturations are ex-
CREDITS (TOP TO BOTTOM):

Th. Peter, C. Marcolli, P. Spichtinger, and T. Corti are at the

erate enormous electric fields (1017 V or Institute for Atmospheric and Climate Science,
pected to relax rapidly as a result of fast vapor
more), which would have more than suffi- Eidgenössische Technische Hochschule (ETH) Zürich, 8092 condensation (7) unless continuous cooling
cient potential to create TeV photons (6). The Zürich, Switzerland. E-mail: [email protected] M. B. remains sufficiently strong. To achieve such
accreting gas probably conducts too well and Baker is in the Department of Atmospheric Sciences, cooling, the clouds would have to rise by sev-
University of Washington, Seattle, WA 98195, USA. T.
would short-circuit the generation of extreme Koop is in the Department of Chemistry, Bielefeld eral kilometers in the measurement time,
electric fields, however. University, D-33615 Bielefeld, Germany. which contradicts the observations.

www.sciencemag.org SCIENCE VOL 314 1 DECEMBER 2006 1399

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PERSPECTIVES

Measuring water in the upper tropo- librium value might lead to perceived super- tuate faster than aircraft-borne instruments
sphere is difficult. A major international saturation. Laboratory data show (16) that can resolve, causing apparent supersatura-
effort to assess water vapor measurements in below 200 K, cubic ice (a metastable form of tion by averaging over glaciated and ice-
the upper troposphere and stratosphere con- ice) nucleates first and might persist in free supersaturated patches. Verification
cluded that, on the basis of laboratory cali- clouds. The equilibrium vapor pressure for of such effects will have to await higher-
brations, typical mean accuracies of aircraft cubic ice is ~10% higher than that for stable resolution instrumentation.
and balloon instruments were on the order of hexagonal ice (17). None of these hypotheses is likely to be
10% (8). However, direct comparisons in the But even after an ice crystal has nucle- the sole explanation for the observed high
upper troposphere suggest that differences ated and transformed into hexagonal ice, supersaturations outside and inside ice
between various instruments on aircraft and surface effects might hinder its growth. It clouds. The uncertainties in the expres-
balloons often exceed 25%, especially when
temperatures are very low (9). Also, bal-

Altitude (km)
14
loon-borne instruments appear to yield
mostly lower supersaturations than do air- 13 Conventional nucleation
and growth
craft instruments (10). Nonetheless, large
12
supersaturations were observed during all
recent aircraft and balloon campaigns; these
studies used a range of instruments based
Altitude (km)

14
on different measurement principles (11).
Hence, only a fraction of the observed 13
Suppressed nucleation
supersaturations can be ascribed to instru- 12
mental inaccuracies.
The theoretical assumptions underlying
Altitude (km)

modeling of ice cloud formation should also 14

be reassessed. How can we explain ice
nucleation in light of the extreme supersatu- 13
rations observed in cloud-free regions, and Suppressed growth
12
ice growth in light of the persistent supersat-
urations observed within dense ice clouds?
Ice supersaturation (%)

Outside ice clouds, the supersaturation at 120 Suppressed nucleation

which ice nucleation occurs depends on the Nucleation
equilibrium vapor pressure of supercooled 60
Suppressed growth
threshold
water. Measured data only exist down to ~235 K 0
and must be extrapolated to lower tempera- Conventional nucleation and growth
tures (12), possibly leading to errors of up to 0 Time (minutes to hours)
20%. Moreover, although air masses appear to Rising air and ice cloud formation. The top three panels sketch three scenarios for the formation of ice
always contain sufficient numbers of aerosol clouds along an ascending air parcel trajectory; the bottom panel sketches the effect of these scenarios on
particles for cloud formation, the composition supersaturation. According to conventional understanding, ice particles nucleate (star), grow, and reduce the
of these aerosols might inhibit ice nucleation. supersaturation (green curves). Recent observations suggest suppressed nucleation (red curves) or suppressd
The homogeneous ice nucleation threshold of growth (orange curves) in large parts of the atmosphere.
60% was established for atmospherically rele-
vant salt solutions and sulfuric acid, but only is usually assumed that most water mole- sions used to retrieve the supersaturation
for a few organic species. Cloud chamber data cules hitting the crystal are built into the from the data must be resolved and their
indicate that aerosols containing only organic lattice, but recent laboratory data indicate usage clarified (12). Uncertainties in the
and elemental carbon may be almost com- that this is the case for fewer than 10% (18) aircraft and balloon data must be deter-
pletely unable to nucleate ice (13). Alter- or even as few as 0.4% (19) of water mole- mined accurately. In addition, mesoscale
natively, if water-rich aerosols were fully cules. Furthermore, gas-phase species meteorological fluctuations must be char-
covered with organic surfactants, nucleation such as nitric acid may selectively block acterized, theories of ice nucleation and
might be suppressed if it started preferentially the growth sites on ice crystals (2), al- growth must be reassessed, and state-of-
at the surface (14). In field experiments, the though this hypothesis needs support from the-art numerical models must be com-
presence of organic pollutants has indeed laboratory studies. pared. Six years after a major effort to char-
been associated with impeded ice particle Further reasons for the observed supersat- acterize the distribution of upper atmos-
formation (15). However, laboratory data of urations may be related not to the properties pheric water (8), the issue is again open,
surface nucleation and field data on particle of individual ice particles, but to effects on and, because of its climatic importance,
composition and surface morphology of larger scales. The presence of ice nuclei in more pressing than ever.
upper tropospheric aerosols are too limited to cloud-free air may initiate ice nucleation
allow any conclusions to be drawn. below the homogeneous-nucleation thresh- References and Notes
Inside sufficiently dense ice clouds, con- old, leading to clouds with low ice particle 1. E. J. Jensen et al., Atmos. Chem. Phys. 5, 851 (2005).
densation should rapidly reduce vapor pres- number densities, in which supersaturations 2. R. S. Gao et al., Science 303, 516 (2004).
3. S. H. Lee et al., J. Geophys. Res. 109, D20209,
sures in excess of the equilibrium vapor might be sustained for relatively long periods. 10.1029/2004JD005033 (2004).
pressure of ice. However, errors in this equi- Also, conditions within clouds might fluc- 4. T. Koop et al., Nature 406, 611 (2000).

www.sciencemag.org SCIENCE VOL 314 1 DECEMBER 2006 1401

PERSPECTIVES

5. In a cooling event, when aerosol particles are exposed to Water Vapour (www.aero.jussieu.fr/~sparc/ Sci. U.S.A. 99, 15873 (2002).
a supersaturation of 60%, the characteristic time for ice WAVASFINAL_000206/WWW_wavas/Cover.html). 15. P. J. DeMott et al., Proc. Natl. Acad. Sci. U.S.A. 100,
nucleation is ~1 min. This drops to <1 s for the atmo- 9. H. Vömel, in Report from the NDACC Meeting on Atmo- 14655 (2003).
spheric measurements described in (1). spheric Water Vapour Measurement, G. Braathen, Ed. 16. B. J. Murray, D. A. Knopf, A. K. Bertram, Nature 434, 202
6. J. P. D. Abbatt et al., Science 313, 1770 (2006); (Univ. of Bern, Bern, Switzerland, 2006); (2005).
published online 30 August 2006 (10.1126/science. www.iapmw.unibe.ch/research/collaboration/ 17. J. E. Shilling et al., Geophys. Res. Lett. 33, L17801,
1129726). ndsc-microwave/workshop/2006). 10.1029/2006GL026671 (2006).
7. The characteristic time to consume the supersaturation 10. H. Vömel et al., J. Geophys. Res. 107, 18. P. Pratte, H. van den Bergh, M. J. Rossi, J. Phys. Chem. A
due to vapor condensation is determined by how rapidly 10.1029/2001JD000707 (2002). 110, 3042 (2006).
gas-phase water molecules can diffuse to and accommo- 11. A. Korolev, G. A. Isaac, J. Atmos. Sci. 63, 2865 (2006). 19. N. Magee, A. M. Moyle, D. Lamb, Geophys. Res. Lett. 33,
date on ice surfaces. For the ice clouds described in (2), 12. D. M. Murphy, T. Koop, Q. J. R. Meteorol. Soc. 131, 1539 L17813, 10.1029/2006GL026665 (2006).
this time should be ~1 min. (2005).
8. D. Kley, J. M. Russell III, C. Phillips, Eds., SPARC 13. O. Möhler et al., Meteorol. Z. 14, 477 (2005).
Assessment of Upper Tropospheric and Stratospheric 14. A. Tabazadeh, Y. S. Djikaev, H. Reiss, Proc. Natl. Acad. 10.1126/science.1135199

CELL BIOLOGY

Tools to Tamper with Rapidly depleting or elevating specific

CREDITS (TOP TO BOTTOM):

phosphoinositide lipids in the plasma
membrane reveals their role in controlling
Phosphoinositides cellular processes.

Stuart McLaughlin

T he lipid phosphatidylinositol 4,5-

bisphosphate (PIP2) both regulates
many different processes at the cell’s
plasma membrane (1) and is the source of
three “second messenger” molecules that
which binds to and activates
the channel (see the figure).
Alternatively, PLC-catalyzed
hydrolysis of PIP2 produces
the second messengers inosi-
K+ channel
PLASMA MEMBRANE

transmit signals throughout the cell. This tol 1,4,5-trisphosphate (IP3)

multiplicity of functions can make it diffi- and diacylglycerol (DAG) (see K-Ras
cult to sort out whether a particular regula- the figure). This leads to the
tory activity of PIP2—for example, activat- activation of protein kinase C
Phosphatase
ing an ion channel or anchoring a peripheral and Ca2+-calmodulin, which
CYTOPLASM
protein to the plasma membrane—is due to also affect KCNQ channel Rapamycin
direct interaction or indirect effects related activity. Suh et al. engineered a
IP3 Ca2+
to the second messengers. Moreover, the lipid phosphatase that, when PIP3
toolbox for investigating the role of lipids in bound to a rapamycin analog, PLC PIP2
Farnesyl
PIP
cell signaling is limited compared to that can translocate rapidly from PI3K
DAG
available for studying specific proteins. Two the cytoplasm to the plasma Basic cluster Other lipid
reports in this issue highlight the utility of a membrane, where it removes a
new tool that rapidly depletes the plasma 5' phosphate from PIP2 (see PIP2, potassium channels, and peripheral proteins. Lipid phos-
membrane of PIP2 without producing the the figure). Depleting the PIP2 phatase-based tools that manipulate the amount of phosphatidylino-
three signaling molecules. On page 1454, causes the KCNQ current sitol 4,5-bisphosphate (PIP2) in the plasma membrane of living cells
Suh et al. (2) show that PIP2 acts directly on measured in cultured mam- (by translocating to the membrane upon addition of rapamycin) reveal
that this phosphoinositide regulates a potassium channel and helps
a K+ ion channel to modulate its activity in malian cells to fall promptly to
2+ anchor K-Ras and other GTPases to the plasma membrane. PIP is also
the plasma membrane, and on page 1458, zero, without changing Ca , the source of three signaling molecules: Phosphorylation by2 phos-
Heo et al. (3) demonstrate that phosphoinosi- DAG, or IP3 concentrations. phatidylinositol 3-kinase (PI3K) yields phosphatidylinositol 3,4,5-
tides probably target guanosine triphos- The second report focuses trisphosphate (PIP ); hydrolysis by phospholipase C (PLC) yields diacyl-
3
phatases (GTPases) to the plasma membrane. on the role of PIP2 and phos- glycerol (DAG) and inositol 1,4,5-trisphosphate (IP3). PIP, phos-
This and related tools described in the phatidylinositol 3,4,5-trisphos- phatidylinositol 4-phosphate.
papers open the door for new investigations phate (PIP3) in binding mem-
of phosphoinositide function in living cells. bers of the Ras, Rab, Arf, and Rho families of get a wide variety of peripheral proteins to the
The ion channel report starts from the ear- GTPases to the inner leaflet of the plasma plasma membrane—but how? Work from sev-
lier observation that muscarinic receptor–trig- membrane. Heo et al. imaged the subcellular eral laboratories, including those of Meyer (3)
gered phospholipase C (PLC) activation location of 125 fluorescently tagged GTPases and Grinstein (4), established that a cluster of
closes KCNQ (Kv7) K+ channels (2). PLC in cultured mammalian cells to survey about eight positively charged lysine residues
could directly induce closing of this channel plasma membrane–targeting mechanisms. in the carboxyl terminus of the GTPase K-Ras,
by depleting the plasma membrane PIP2, One of their important observations is that of which acts in concert with a farnesyl lipid moi-
the 50 GTPases that localize to the plasma ety, targets and anchors the protein to the inner
membrane, 40 contain a cluster of basic (pos- leaflet of the plasma membrane (see the fig-
The author is in the Department of Physiology, Stony
Brook University, Stony Brook, NY 11794–8661, USA. itively charged) amino acids. ure). Is this a general electrostatic effect medi-
E-mail [email protected] Unstructured clusters of basic residues tar- ated by the predominant acidic (negatively

1402 1 DECEMBER 2006 VOL 314 SCIENCE www.sciencemag.org

PERSPECTIVES

charged) phosphatidylserine lipids in the ing peripheral proteins with clusters of basic membrane, and how the new tools can be used
plasma membrane, or do the much less abun- residues to the plasma membrane. The results to investigate these functions, read these
dant phosphoinositides PIP2 and PIP3 play a on GTPases suggest that the phosphoinositides important reports (2, 3, 7).
key role in targeting? PIP2 and PIP3 may be more important (3).
Heo et al. used the same conjugated form The new lipid phosphatase (and kinase) References
of a constitutively active yeast lipid phos- tools should also prove useful for investigat- 1. G. Di Paolo, P. De Camilli, Nature 443, 651 (2006).
phatase to study the potential role of PIP2 ing the binding of other proteins with mem- 2. B.-C. Suh, T. Inoue, T. Meyer, B. Hille, Science 314, 1454
in GTPase–plasma membrane interactions. brane-sticky clusters of basic residues [for (2006); published online 21 September 2006
(10.1126/science.1131163).
Using the phosphatase, rather than PLC, to example, the scaffolding protein gravin and 3. W. D. Heo et al. Science 314, 1458 (2006); published
deplete the plasma membrane of PIP2 had lit- the protein kinase Src] and the many other online 9 November 2006 (10.1126/science.1134389).
tle effect on the membrane association of the plasma membrane processes involving PIP2 4. T. Yeung et al., Science 313, 347 (2006).
GTPases (PLC-mediated hydrolysis of PIP2 and PIP3 (1). Indeed, a different group re- 5. W. Cho, R. V. Stahelin, Annu Rev. Biophys. Biomol.
Struct. 34, 119 (2005).
leads to activation of protein kinase C and cently used essentially the same phosphatase
6. S. McLaughlin, D. Murray, Nature 438, 605 (2005).
Ca2+-calmodulin, both of which cause K-Ras tool to investigate how PIP2 affects both endo- 7. P. Varnai, B. Thyagarajan, T. Rohacs, T. Balla, J. Cell Biol.
to move off the membrane). This was not sur- cytosis and the transient receptor potential 175, 377 (2006).
prising, because previous work showed that melastatin 8 (TRPM8) Ca2+ conducting chan-
Published online 9 November 2006;
K-Ras binds well to phospholipid vesicles nel (7). If you are interested in the many func- 10.1126/science.1136314
with putative physiological levels (20%) of tions of the phosphoinositides in the plasma Include this information when citing this paper.
monovalent phosphatidylserine (4). The sur-
prise came when cells were also treated with
inhibitors of phosphatidylinositol 3-kinase to GENETICS
reduce both PIP2 and PIP3 in the membrane:
K-Ras4B and the other GTPases with a clus-
ter of basic residues translocated from the
Delivering New Disease Genes
plasma membrane. The authors conclude Lon R. Cardon
that both phosphoinositide molecules target
and anchor clusters of basic residues to the Complex diseases represent an extraordinary challenge to geneticists, but recent results are
plasma membrane. revealing some successful strategies.
This conclusion may generate controversy
for two reasons. First, it challenges the com-
mon wisdom that PIP3 “is present in negligi-
ble amounts in resting cells” (1). Second, it
challenges the hypothesis, first put forth by
Silvius, that the cluster of basic residues on a
S ince the human genome was sequenced,
advances in human genetics research
have steadily built momentum toward
identifying genes that influence common
human diseases. Validation of millions of
large number of genetic variants, or single-
nucleotide polymorphisms (SNPs); then,
at each SNP site, compare the frequencies
among cases and controls (Duerr et al. exam-
ined more than 300,000 SNPs). Sites that dif-
GTPase associates with the plasma membrane genetic variants, rapid advances in genotyp- fer significantly between cases and controls
because its inner leaflet may contain a ing technologies, and are then validated in independent samples. In
higher mole fraction of phosphatidylser- the ongoing estab- practice, of course, genome-wide association
Enhanced online at
ine than the cytoplasmic leaflets of intra- www.sciencemag.org/cgi/ lishment of reposi- is more complicated. Individual differences
cellular organelle membranes, and thus content/full/314/5804/1403 tories of large, pop- in most common diseases are thought to
has a more negative surface potential (5). ulation-based patient arise from a relatively small number of
However, data in Yeung et al. (4) support samples have created genes (numbering in the tens to hundreds),
the conclusion that it is PIP2 and PIP3 (3), expectations of imminent discoveries of prized most of which contribute only modestly to
rather than phosphatidylserine, that likely tar- disease genes. Have these activities truly laid the overall disease risk (2). Therefore, in a
get K-Ras to the plasma membrane. the foundation for gene identification? On large-scale association screen, most disease
Experiments on model phospholipid mem- page 1461 of this issue, Duerr et al. (1) demon- gene variants are expected to produce only
branes and theoretical calculations explain why strate an association between variants in the a small “signal” that is difficult to detect
clusters of basic residues in proteins require IL23R gene and Crohn’s disease, a common among a large number of SNPs. One big
neither structure nor specific sequences to lat- inflammatory condition of the gastrointestinal question is how many (if any) of the signals
erally sequester polyvalent PIP2 and PIP3. tract. Their results show that complex disease will be large enough to separate from the
These clusters produce a local positive electro- genes are finally yielding their secrets and pro- “noise” (3, 4). The answer from the IL23R
static potential that extends about 1 nm from vide crucial validation of the long journey to study is not many, but even a few may be
the region and acts as a deep “basin of attrac- gene discovery. enough to help uncover significant novel dis-
tion” for multivalent negatively charged phos- Duerr et al. used a genome-wide associa- ease-associated variants.
phoinositides (6). Uncertainty about several tion approach premised on a simple idea: The genome-wide association screen of
factors—for example, the free concentration Assay genomic DNA from a sample of cases Duerr et al. revealed three SNPs with evi-
CREDITS (TOP TO BOTTOM):

and lateral distribution of lipids in biological (diseased patients) and controls for a very dence for disease association more than 100
membranes or the proximity of acidic residues times as large as that of the next most statisti-
to the basic cluster—means that experiments cally significant SNP. In genome-wide asso-
on model membranes cannot be used to tease The author is at the Wellcome Trust Centre for Human ciation scans, such SNPs are usually either
Genetics, University of Oxford, Oxford OX3 7BN, UK, and
out the relative importance of phosphatidylser- Fred Hutchinson Cancer Research Center, Seattle, WA artifacts due to genotyping error, or rarely
ine or phosphoinositides in targeting or anchor- 98109, USA. E-mail: [email protected] observed examples of variants with large

www.sciencemag.org SCIENCE VOL 314 1 DECEMBER 2006 1403

!
" # $ $ %
&

!"
#$#%#&&'%&
( ))***

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PERSPECTIVES

phenotypic effects. Distinguishing these two low frequency, such as the IL23R variant The comulative effect of all of these
outcomes is challenging because there is little studied by Duerr et al., which has a fre- aspects of the IL23R study is to lend confi-
statistical and experimental information to quency of 2 to 3%. Larger sample sizes will dence in the results, which is not always read-
guide the process (5). Duerr et al. found that be needed to identify more disease loci. ily apparent in other designs. A recent study
two of the three SNPs with the strongest evi- There are at least two important lessons of association between the KIBRA gene and
dence for association are in a well-known sus- in the strategy used to uncover IL23R human memory (13) adopted an entirely dif-
ceptibility gene (CARD15) for Crohn’s dis- and the macular degeneration genes. First, ferent design, using pooled DNA samples in
ease (6, 7), and the third is a nonsynonymous though the initial genome-wide association multistage genotyping, analyzing different
coding change in IL23R, a gene encoding a discoveries may uncover relatively few “low- memory measures in the primary and repli-
receptor for the proinflammatory cytokine hanging fruit”—especially with small sam- cation phases, and studying patient samples
interleukin-23. Thus, two of the top three hits ple sizes—they may lead to identifying from different geographic locations, some
validated the genome-wide association more stubborn variants with smaller effect with high levels of population substructure.
proof-of-principle, and the third was either an size (10–12). Second, every SNP counts. These phenotypic and sampling differences
artifact or a true positive result. may eventually support the
Replication of genome-wide 16000 Frequency distribution of SNP generality of the reported find-
association tests in a genome scan
association findings is essential to 14000 ing, but they will complicate
distinguish these possibilities. Noise interpretations of external con-
Unfortunately, replication has not 12000 Irrevelant SNPs firmation because the hypo-
been an area of strength in genetic 10000 Gene variants Known theses generated may be diffi-
related to disease CD gene
association studies (4). In this case, + + cult to falsify—negative results
Frequency

CARD15
however, Duerr et al. replicated 8000 could reflect either measure-
Novel
the IL23R finding unambiguously, 6000 CD gene
ment differences or lack of repli-
revealing strong statistical support + cation, whereas strictly positive
IL23R
for the same SNP, with the same 4000 associations would require firm
risk allele, and in the same specific 2000 concordance in phenotypes and
Three exceptional SNPs (+)
phenotypes in larger independent Re-examine scan data sampling to demonstrate con-
0
0
for other loci in IL23R
samples of disease patient cases, 1 2 3 4 5Test6 statistic
7 8 9 10 11 12 13 sistency.
controls, and families. Together Not all genome-wide associa-
with the recent identification of tion studies will be as successful
genes associated with age-related Additional disease- as the IL23R finding. Sample
macular degeneration by similar
associated variants + . size will be a key determinant of
discovered ( )
techniques (8, 9), the IL23R find- outcome, as will genetic, popula-
ing should help future studies, as Cascade of discovery. Common human diseases are thought to have few disease tion, and phenotypic heterogene-
loci (red) relative to the number of single-nucleotide polymorphisms (SNPs) tested
researchers can better understand (green), and most of the true loci will have a small effect. The greatest statistical ity. In addition, it is increasingly
the statistical profiles of genuinely power is thus attributed to loci that occur least frequently in the genome. Duerr et important to present data and
associated disease alleles. al. found three SNPs associated with Crohn’s disease (CD), two in a known disease results for all analyses con-
The finding of an association gene and one in the IL23R gene. This allowed them to extract other real SNP- ducted, which is one of the few
between IL23R and Crohn’s dis- disease associations embedded in the noise of the initial genome-wide scan. shortcomings of the Duerr et al.
ease has already led to other dis- report. In any case, results from
coveries. Duerr et al. examined the chromoso- Had the one highly significant SNP not been the current generation of genetic studies
mal locations of their remaining genome- included in the genome-wide association should help provide a foundation for the next
wide association SNPs to see if others were in panel by Duerr et al., the entire discovery set of problems, involving detection of rare
the same region as the IL23R gene. Many might have been delayed or even missed genetic variants, leveraging the genetics to
were, some of which had supportive but not Dozens of genome-wide association stud- better understand environmental risk factors,
striking statistical evidence for association. ies are under way, and many more are and ultimately, using this hard-won informa-
Replication studies and further statistical planned. So why have Crohn’s disease and tion to improve public health.
analysis suggest that several of these SNPs age-related macular degeneration been the
contribute independently to Crohn’s disease. particular traits to yield such striking results? References
1. R. H. Duerr et al., Science 314, 1461 (2006); published
These loci of smaller effect were detectable One reason relates to phenotypic specificity. online 26 October 2006 (10.1126/science.1135245).
because of the initial discovery of the highly To reduce pathogenic and genetic hetero- 2. K. T. Zondervan, L. R. Cardon, Nat. Rev. Genet. 5, 89 (2004).
significant variant (see the figure). geneity, Duerr et al. focused initially on 3. W. Y. Wang, B. J. Barratt, D. G. Clayton, J. A. Todd, Nat.
Rev. Genet. 6, 109 (2005).
Should we be surprised that the genome- patients with a specific type of Crohn’s dis- 4. L. J. Palmer, L. R. Cardon, Lancet 366, 1223 (2005).
wide association revealed only one unam- ease and further conditioned on genetic back- 5. D. G. Clayton et al., Nat. Genet. 37, 1243 (2005).
biguous novel SNP for the disease of inter- grounds having known differences in Crohn’s 6. Y. Ogura et al., Nature 411, 603 (2001).
7. J. P. Hugot et al., Nature 411, 599 (2001).
est? Probably not. The Crohn’s disease sam- disease prevalence. Similarly, the findings on 8. R. J. Klein et al., Science 308, 385 (2005).
ple size of 547 cases and 548 controls is age-related macular degeneration resulted 9. A. DeWan et al., Science 314, 989 (2006).
small for such a study, having the statistical from a series of refined phenotypic classifi- 10. M. Li et al., Nat. Genet. 38, 1049 (2006).
11. B. Gold et al., Nat. Genet. 38, 458 (2006).
power to detect only large genetic effects, of cations. This careful attention to phenotypes 12. J. Maller et al., Nat. Genet. 38, 1055 (2006).
which there are likely few in the genome. The reduces a primary source of heterogeneity 13. A. Papassotiropoulos et al., Science 314, 475 ( 2006).
problem of statistical power is especially and is thus directly beneficial to genome-
salient when the associated variant occurs at wide association studies. 10.1126/science.1136668

www.sciencemag.org SCIENCE VOL 314 1 DECEMBER 2006 1405

PERSPECTIVES

RETROSPECTIVE
E. I. Stiefel was a bioinorganic chemist whose
insights into the roles of metals in living
Edward I. Stiefel (1942–2006) systems have led to practical applications
and new research ideas.
François M. M. Morel and John T. Groves

E dward I. Stiefel, an unusual chemist

who bridged industry and academia,
pure chemistry and applications, and
bioinorganic and environmental chemistry,
died on 4 September 2006 in Robert Wood
Catalytic Materials Group. He initiated dis-
cussions concerning the active centers of
nitrogenase and other molybdenum enzymes
that bridged “industrial” and “biological”
catalysis. This new insight led him to wonder
graduate-level course on “Metals in Bio-
logy” (jointly with J. T. Groves); and, in
collaboration with colleagues, an innovative
multidepartmental course on “Life in the Uni-
verse,” which was instrumental in doubling the
Johnson Hospital, New Brunswick, New whether tungsten-associated enzymes might number of chemistry majors. Students in Ed’s
Jersey. His insights into the functions of metals exist that would be analogous to known courses revered him. The evening freshman
in living systems led to a new understand- molybdenum enzymes. Stiefel predicted the seminars kept students on the edge of their seats
ing of key biological processes, new techno- existence of such enzymes before their dis- and often went on well into the night.
logical applications, and covery in Pyrococcus furiosus, a Ed held 30 U.S. patents and published
a better appreciation microbe that thrives in the boiling more than 150 scientif ic articles. His
of the interactions waters of hot springs (2). review article on “The Coordination and
among the cycles of Bioinorganic Chemistry of Moly-
trace and major ele- bdenum” has been cited in more
ments on Earth, both “In every problem he tackled, than 800 publications. And his
today and in the past. he had a phenomenal contributions to science and his
Ed grew up in inspiration to students and col-
Brooklyn, New York. ability to see the big leagues are continuing. He is the
He received his Ph.D. picture.” coauthor of several articles that
with Harry Gray at are in various stages of publica-
Columbia University —Harry Gray tion, some of which bring toge-
in 1967 and then served ther the various strands of his
on the faculty at the While at Exxon, Ed multifaceted career. One upcoming article
State University of New developed many new deals with the function of iron storage
York at Stony Brook, molybdenum compounds proteins in ocean regions where life is
before becoming a senior investiga- that have proven useful as limited by the availability of iron (3). He is
tor at the Charles F. Kettering catalysts and lubricating a coeditor of the book Biological Inorganic
Research Laboratory. In 1980, oil additives. For example, Chemistry: Structure and Reactivity, which
he joined the Catalytic Materials he was the principal in- has just been published (4) and is destined
Group at Exxon’s Corporate Stra- ventor of the commercially to become a classic.
tegic Research Center in Annan- important “thiomolybdate” Ed Stiefel was a joyful man, a compas-
dale, New Jersey. Following his additive for lubricating oils. sionate friend, an inspiring teacher, and a
retirement from ExxonMobil in This compound had previ- generous colleague. At a recent colloquium
2001, Ed accepted a position ously been used as a catalyst at Princeton, his close friend and early men-
at Princeton University as the first for the desulfurization of crude tor Harry Gray most concisely described
holder of the Ralph W. Dornte Chair, a dis- oil and was found to make the what it meant to have Ed Stiefel as a col-
tinguished visiting lecturer position with the oil more slippery and less prone to oxidize. league: “A discussion of a research problem
rank of professor. His knowledge of the bioinorganic chem- with Ed was really special. He understood
Ed’s research interests involved the bio- istry and biology of metalloenzymes proved the roles metals play in living systems better
inorganic, coordination, and environmental to be invaluable when he became one of the than anyone. In every problem he tackled,
chemistry of transition-metal ions, particu- principal architects of the cleanup after the he had a phenomenal ability to see the big
larly iron and molybdenum. While at Ketter- Exxon Valdez oil spill in Alaska in 1989. This picture. He was a scholar’s scholar.” He is
ing, he codiscovered bacterioferritin, the iron massive effort successfully applied the prin- survived by his wife Jeannette, a frequent
storage protein of prokaryotes (1), and made ciples of bioinorganic chemistry and micro- visitor and friend of the Princeton Chemistry
pioneering advances in our understanding of biology to a very-large-scale environmental Department, and their daughter Karen.
nitrogenase enzymes, illuminating the role remediation project. Ed’s interdisciplinary
of molybdenum in catalyzing the conversion interests also shaped the science agenda and References
of atmospheric nitrogen to ammonia. recruitment strategies of Exxon’s Corporate 1. E. I. Stiefel, G. D. Watt, Nature 279, 81
(1979).
His arrival at Exxon made an immediate Strategic Research Center. Ed’s genius lay in 2. S. Mukund, M. W. W. Adams, J. Biol. Chem. 265,
impact on the company, vastly increasing the his ability to understand complex chemical 11508 (1990).
CREDITS: EXXONMOBIL

interdisciplinary research by the company’s issues over a wide range of scales, from 3. M. Castruita et al., in preparation.
molecular to cellular to global. 4. I. Bertini, H. B. Gray, E. I. Stiefel, J. S. Valentine, Eds.,
Biological Inorganic Chemistry: Structure and Reactivity
At Princeton, Ed was instrumental in (University Science Books, Sausalito, CA 2006).
The authors are in the Department of Geosciences and
Chemistry, Princeton University, Princeton, NJ 08544, USA. developing three very popular courses: a fresh-
E-mail: [email protected]; [email protected] man seminar entitled “Elements of Life”; a 10.1126/science.1136957

1406 1 DECEMBER 2006 VOL 314 SCIENCE www.sciencemag.org

Science & Engineering
Vi s u a l i z a t i o n Challenge
Cal l for E ntri es

Sc i en ce a n d en gi n eer in g’s m o st p owe r f u l stat e m e n ts

a re n ot ma d e f ro m wo rd s a lo n e
Entry Deadline: May , 

When the left brain collaborates with the right brain, Winners in each category will be published in the Sept.
science emerges with art to enhance communication and 28, 2007 issue of Science and Science Online, and will be
understanding of research results—illustrating concepts, displayed on the NSF Web site.
depicting phenomena and drawing conclusions.
The National Science Foundation (NSF) and the journal Awa rd Cate g o ri e s :
Science, published by the American Association for the Il lus tr atio ns ,
Advancement of Science, invite you to participate in Inf o r matio na l G r a ph ics ,
the ﬁfth annual Science and Engineering Visualization
Inte r active M e d ia ,
Challenge. The competition recognizes scientists,
engineers, visualization specialists and artists for
N o n - Inte r active M e d ia ,
producing or commissioning innovative work in visual Ph oto gr a ph s
communication.

Complete Entry Information: www.nsf.gov/news/special_reports/scivis/

SPECIALSECTION

INTRODUCTION

Size, Mates, and Fates

THE SCIENCE PERSPECTIVES AND ASSOCIATED CONNECTIONS MAPS IN THE
Database of Cell Signaling at Science’s Signal Transduction Knowledge Environ-
ment (www.sciencemag.org/sciext/cellsignaling06/) highlight pathways initiated
by three different types of receptors: brassinosteroid receptors that control plant
size, G protein–coupled receptors that control mating responses in yeast, and Notch
receptors that control cell fate in animals. Cell Signaling
Brassinosteroids are plant hormones that contribute to cell growth and division,
differentiation, and reproductive development. As Belkhadir and Chory (p. 1410) CONTENTS
describe, the brassinosteroid receptor BR1 is a plasma membrane–localized leucine-
rich–repeat receptor kinase that initiates a kinase cascade, ultimately controlling gene Perspectives
expression. Experiments in Arabidopsis thaliana have revealed that the active recep- 1410 Brassinosteroid Signaling:
tor complex is a serine-threonine kinase that initiates a process that inactivates BIN2, A Paradigm for Steroid Hormone
the plant homolog of glycogen synthase kinase 3 (GSK-3). Arabidopsis BIN2 is Signaling from the Cell Surface
localized to the nucleus, whereas in animals, GSK-3 is cytosolic. BIM, which resem- Y. Belkhadir and J. Chory
bles the animal transcription factor Myc, interacts with the brassinosteroid response 1412 G Protein Signaling in Yeast:
factor BES to enhance its activity. Thus, at each step of the brassinosteroid pathway— New Components, New Connections,
from the steroid hormone–like ligands to the transcription factors in the nucleus— New Compartments
there are similarities with animal pathways; however, no known pathway in animals J. E. Slessareva and H. G. Dohlman
assembles this cast of characters in quite the same way. 1414 Notch, a Universal Arbiter of
Even well-characterized pathways continue to reveal new secrets. The yeast Cell Fate Decisions
mating response is an extensively studied G protein–coupled receptor pathway, in M. Ehebauer et al.
which the Gβγ subunits have been in the spotlight as the subunits that activate the
mitogen-activated protein kinase cascade. Now, Slessareva and Dohlman (p. 1412)
Connections Maps
Brassinosteroid Signaling Pathway
describe how the Gα subunit participates in transmitting the mating signal by inter-
Y. Belkhadir, X. Wang, J. Chory, Sci. STKE,
acting with the phosphoinositide 3-kinase (PI3K) at the endosome to stimulate the
http://stke.sciencemag. org/cgi/cm/
production of phosphoinositide 3-phosphate. Not only was this a previously stkecm;CMP_19131.
unknown function and location for Gα in the yeast mating response, but the reg-
ulatory subunit of PI3K appears to serve as a noncanonical Gβ subunit for the Arabidopsis Brassinosteroid Signaling Pathway
endosomally located Gα, a discovery that will undoubtedly stimulate a search for Y. Belkhadir, X. Wang, J. Chory, Sci. STKE,
http://stke.sciencemag.org/cgi/cm/stkecm;CMP_19349.
similar types of interactions in other systems.
The Notch signaling pathway is crucial to animal development, and aberrant Pheromone Signaling Pathways in Yeast
activity of this pathway is associated with certain types of leukemia. Ehebauer et al. H. G. Dohlman and J. E. Slessareva, Sci. STKE,
http://stke.sciencemag.org/cgi/cm/stkecm;CMP_13999.
(p. 1414) explain how the transmembrane Notch receptor interacts with the trans-
membrane ligand on adjacent cells, which leads to cleavage and release of the Notch Signaling Pathway
Notch intracellular domain (NICD) that translocates to the nucleus and regulates M. Ehebauer, P. Hayward, A. Martinez-Arias,
gene expression. The cleavage of Notch remains an open area of research, with var- Sci. STKE,
ious candidates for the protease that performs the first cleavage in the extracellular http://stke.sciencemag.org/cgi/cm/stkecm;CMP_19043.
domain and questions surrounding whether the second cleavage event occurs at the
See also related STKE material on page 1347 or at
plasma membrane or after internalization. Notch acts in concert with other mor- www.sciencemag.org/sciext/cellsignaling06/
phogenic signals to control cell fate; thus, understanding Notch signaling within
CREDIT: C. BICKEL/SCIENCE

the more complex signaling network remains a critical avenue of investigation.

This knowledge may not only yield insight into animal development but also open
the doors to new therapeutic opportunities in cases where the loss of differentiation
contributes to disease.
– NANCY R. GOUGH, ELIZABETH M. ADLER, L. BRYAN RAY

www.sciencemag.org SCIENCE VOL 314 1 DECEMBER 2006 1409

Cell Signaling
dependent of BAK1, a BRI1-BAK1 hetero-
PERSPECTIVE oligomer may be the active signaling complex.
SERK1, a BAK1 homolog, also interacts with

Brassinosteroid Signaling: A Paradigm BRI1, and genetic experiments have implicated

this protein in the signaling pathway (12). In
addition to the plasma membrane, a signaling-
for Steroid Hormone Signaling competent BRI1-BAK1 hetero-oligomer was
detected in bona fide endocytic compartments
from the Cell Surface of plant protoplasts (12). Furthermore, overex-
pression of BAK1 appears to augment the inter-
Youssef Belkhadir and Joanne Chory* nalization of BRI1 in heterologous cell cultures.
Thus, the function of the SERK family in BR
Plants use the coordinated action of several small-molecule hormones to grow and develop signaling may be to facilitate the entry of BRI1
optimally in response to a changing environment. Among these hormones are the brassinosteroids into these intracellular compartments. The phys-
(BRs), the polyhydroxylated steroid hormones of plants. BRs bind a small family of leucine-rich iological role of an endosome-localized BRI1-
repeat receptor kinases at the cell surface, thereby initiating an intracellular signal transduction BAK1 hetero-oligomer is not currently known.
cascade that results in the altered expression of hundreds of genes. Only the BRI1 locus was defined by loss-of-
function mutations that caused BR-resistant dwarf-
rassinosteroids (BRs) are small growth- Forward genetic screens for dwarf mutants ism; however, several additional components of the

B promoting molecules found at low con-

centrations throughout the plant kingdom.
In a technical tour de force, researchers in the late
that were not rescued by exogenous addition of
BL led to the identification of multiple mutant
alleles of a single locus, BRI1 (brassinosteroid
pathway were identified by analysis of gain-of-
function phenotypes or plants with increased sensi-
tivity to BRs. A bin2 (brassinosteroid insensitive 2)
1970s purified a bioactive steroid from Brassica insensitive 1) (8, 9) (Fig. 1). BRI1 is a leucine- dwarf mutant phenotype results from a semi-
napus bee-collected pollen. The most active BR rich repeat receptor kinase (LRR-RK) that has dominant mutation. BIN2 is one member of an
was identified by single-crystal x-ray analysis as a an extracellular domain with an N-terminal sig- Arabidopsis subfamily of glycogen synthase kinase
steroidal lactone and was named brassinolide nal peptide followed by 24 imperfect leucine- 3 (GSK3, also known as Shaggy-like kinases).
(BL) (1). The role of BRs as plant hormones was rich repeats (LRRs), a single transmembrane Although single loss-of-function alleles revealed no
clarified in the mid-1990s with the discovery of domain, and an intracellular serine-threonine effect on BR signaling (6, 15), reduced expression
Arabidopsis thaliana mutants that were deficient kinase domain followed by a short C-terminal of the entire subfamily results in plants with en-
in BR biosynthesis. BR-deficient mutants are tail. BRI1 is the major BR-binding activity of hanced BR responses, supporting a role for these
extremely dwarfed with very small curled leaves; Arabidopsis. The minimal BR-binding domain three kinases as negative regulators of BR sig-
the mutants can be rescued to wild-type stature by is a 94–amino acid subdomain that includes the naling (15). BIN2 localizes to multiple subcellular
exogenous application of BL (2, 3) (Fig. 1). 70–amino acid “island” just proximal to LRR20 compartments but appears to exert its largest effects
Analysis of these mutants showed that BRs play a and the atypical LRR, LRR21 (Fig. 1) (10). As on BR signaling when it is retained in the nucleus.
role in cell expansion and division, differentiation, such, the signaling pathway defined by BRI1 BSU1 (BRI1 suppressor 1) was identified as a
and reproductive development. The regulation of represents a paradigm for steroid perception at dominant suppressor of a weak bri1 mutant (4, 6).
these processes by BRs allows plants to develop the cell surface and may have implications for BSU1 overexpression substantially suppresses the
a body plan that is optimal for their ambient the understanding of the growing field of plasma dwarf phenotypes associated with either the bri1
environment and may confer increased adaptation membrane steroid signaling in metazoans. or bin2 mutations, which suggests that BSU1 is a
to various stresses. The ease of analysis of the BR- In the absence of steroid, the kinase activity of positive regulator of BR signaling that acts on the
deficient phenotype allowed the identification of BRI1 is inhibited by both cis and trans mech- same process or downstream of BIN2. BSU1
the plasma membrane–localized BR receptor and anisms. BKI1 (BRI1 kinase inhibitor 1) is a encodes a nuclear-localized serine-threonine phos-
much of the intracellular signaling pathway that plasma membrane–associated phosphoprotein that phatase with an N-terminal domain comprising
regulates BR-responsive gene expression (4–6). interacts directly with the kinase domain of BRI1 Kelch repeats. Loss of BSU1 function has no
However, key open questions remain with respect (6, 11). Binding of BRs to preformed BRI1 homo- effect on Arabidopsis, but when the expression
to how the BR signaling pathway contributes to oligomers triggers the rapid dissociation of BKI1 of BSU1 and three related genes is reduced by
the adaptive plasticity of plant growth. from the plasma membrane (11). In vitro, BKI1 RNA interference, the resulting plant is dwarfed
BRs have structures similar to those of animal interferes with the interaction of BRI1 with its (16, 17). The substrates of BIN2 and BSU1 are
steroid hormones. BR biosynthesis enzymes signaling partner, a second plasma membrane– likely to be a family of plant-specific transcrip-
share sequence identity with mammalian steroid localized LRR-RK called BAK1 [BRI1-associated tion factors. The founding members of this fam-
biosynthetic enzymes (e.g., DET2 is a plant receptor kinase 1, also known as SERK3 (somatic ily, BES1 (bri1 ems 1) and BZR1 (brassinazole
ortholog of mammalian steroid 5a-reductases, embryogenesis receptor kinase 3)] (12). resistant 1), are 89% identical and contain
and most of the other biosynthetic genes are Although the precise sequence of events is not multiple predicted GSK3 phosphorylation sites
cytochrome P450s) (7). The major branch of the clear, BR binding also allows autophosphoryl- (4, 15). Recent models propose that the balanced
biosynthetic pathway, from campesterol to BL, ation of critical serine and threonine residues activities of BIN2 and BSU1 directly control the
was determined using a combination of Arabi- located in the activation loop of the kinase do- phosphorylation state of BES1 and BZR1. BIN2-
dopsis genetics and feeding experiments with BR main of BRI1 (13, 14). This, in turn, alleviates the induced phosphorylation of BES1 inhibits its
biosynthetic intermediates. autoinhibitory effect of the C-terminal tail on transcriptional activity through impaired multi-
kinase activity and allows further autophospho- merization and DNA binding activity at BR-
Plant Biology Laboratory and Howard Hughes Medical Institute, rylation of the receptor, which increases the af- responsive target promoters. In contrast, in the
Salk Institute for Biological Studies, La Jolla, CA 92037, USA. finity of BRI1 for BAK1 (13). BAK1 has a presence of BRs, BES1 and BZR1 are dephos-
*To whom correspondence should be addressed. E-mail: short extracellular domain comprising five phorylated by the combined inactivation of
[email protected] LRRs. Although BR binding to BRI1 is in- BIN2 and the phosphatase activity of BSU1,

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SPECIALSECTION
ic actions of BRs on plant tolerance to tempera-
ture, salt, and pathogens. Although the signaling
pathway defined by BRI1 is known mostly for
controlling the rapid increase in tissue mass, it
remains possible that the other effects of BRs are
mediated by different signaling pathways. Finally,
every signaling component, from BRI1 to the
regulated transcription factors, is redundantly
encoded in the genome. Perhaps redundancy in
BR signaling could increase the robustness of the
pathway to mutations; alternatively, partial redun-
dancy due to overlapping expression patterns of
signaling components may help to fine-tune the
pleiotropic BR response. A systematic analysis of
the function and expression patterns of family
members will help to unravel these questions.
Ultimately, the goal of this work is to un-
derstand how plant size is controlled and to be able
to manipulate plant growth for human benefit. BRs
play an important role in the processes that control
plant size, but other small-molecule hormones also
contribute to cell expansion and division. Although
there is redundancy within a given hormonal path-
way, loss of response to any one plant hormone
cannot be compensated for by the action of another
hormone. To add another level of complexity, the
entire program can be altered by changes in the
environment’s ambient light or temperature. Plants
provide an accessible model for answering
questions of organ or organismal size because the
number of cell types and different organs is small,
organ size is easily manipulated by environment,
and many of the individual signaling pathways are
known. Questions that can be addressed include
how the levels of plant hormones change through-
out development and in response to an ever-
changing environment, and how these different
pathways interact within individual cells. Ultimate-
Fig. 1. A model of BR control of Arabidopsis size. Arabidopsis biosynthetic mutants that do not
produce brassinolide (BL) are dwarf (upper left photo) but can be rescued to full stature by ly, we may answer the age-old question of how
exogenous application of BL (upper right photo). BR signaling mutants (lower left photo) cannot be the size of an organism is determined.
rescued by exogenous BL. In the absence of BRs, the kinase domains of the BRI1 homodimer are References and Notes
inhibited by both their own C-terminal domain and by an interaction with BKI1. This allows the 1. M. D. Grove et al., Nature 281, 216 (1979).
GSK3 homolog, BIN2, to phosphorylate and inactivate the brassinosteroid response transcription 2. J. Li et al., Science 272, 398 (1996).
3. M. Szekeres et al., Cell 85, 171 (1996).
factors (BRFs), including BES1 and BZR1. Direct binding of BL to BRI1 homodimers results in 4. G. Vert et al., Annu. Rev. Cell Dev. Biol. 21, 177 (2005).
conformational changes of the kinase domain, leading to the phosphorylation of the C-terminal 5. Y. Belkhadir et al., Brassinosteroid Signaling Pathway,
domain of BRI1 and phosphorylation of BKI1, which causes displacement of BKI1 from the plasma Sci. STKE (Connections Map, as seen October 2006)
membrane and the release of autoinhibition of BRI1. These events lead to BRI1’s association with (http://stke.sciencemag.org/cgi/cm/stkecm;CMP_19131).
BAK1 and consequent activation of the receptor. The active signaling receptor complex inhibits the 6. Y. Belkhadir et al., Arabidopsis Brassinosteroid Signaling
Pathway, Sci. STKE (Connections Map, as seen October 2006)
activity of BIN2 by an unknown mechanism, allowing dephosphorylation of the BRFs by BSU1 and (http://stke.sciencemag.org/cgi/cm/stkecm;CMP_19349).
activation or repression of their target genes and optimal plant growth (lower right photo). The 7. S. Fujioka, T. Yokota, Annu. Rev. Plant Biol. 54, 137 (2003).
horseshoe-shaped representations of BRI1 and BAK1 LRR domains, as well as the putative LRR (red 8. J. Li, J. Chory, Cell 90, 929 (1997).
domains) interactions, are inferred from structural models of LRR-containing proteins. The atypical 9. S. D. Clouse et al., Plant Physiol. 111, 671 (1996).
LRR21 is represented by a yellow domain. The BL docking into the binding site is speculative. 10. T. Kinoshita et al., Nature 433, 167 (2005).
11. X. Wang, J. Chory, Science 313, 1118 (2006).
Phosphorylation events are indicated by a circled P. 12. R. Karlova, S. C. De Vries, Sci. STKE 2006, pe36 (2006).
13. X. Wang et al., Dev. Cell 8, 855 (2005).
14. X. Wang et al., Plant Cell 17, 1685 (2005).
which allows them to homo- or heterodimerize and the physiological role of the endosomal BRI1- 15. G. Vert, J. Chory, Nature 441, 96 (2006).
and bind more efficiently to the BR-responsive BAK1 hetero-oligomer needs to be determined. 16. Z. Y. Wang et al., Dev. Cell 2, 505 (2002).
elements to either positively or negatively regu- Second, a major gap exists in the pathway between 17. Y. Yin et al., Cell 109, 181 (2002).
18. L. Li, X. W. Deng, Trends Plant Sci. 10, 266 (2005).
late BR-responsive target genes (15, 18). events at the plasma membrane and in the nucleus. 19. Supported by USDA and NSF grants (J.C.) and by the Howard
Despite tremendous progress in understanding Understanding how BIN2 is inactivated and how Hughes Medical Institute. Y.B. is a Howard Hughes Medical
the molecular and cellular effects of BRs, key its subcellular localization may be altered in Institute Fellow of the Life Sciences Research Foundation.
issues remain unanswered. First, details of the response to BRs may help fill in this gap. A third We thank S. Savaldi-Goldstein for Arabidopsis pictures.
receptor activation mechanism need to be clarified area of investigation revolves around the pleiotrop- 10.1126/science.1134040

www.sciencemag.org SCIENCE VOL 314 1 DECEMBER 2006 1411

Cell Signaling
protein purification and in situ localization
PERSPECTIVE
studies. The existence of these resources has
profoundly transformed our approach to the
G Protein Signaling in Yeast: identification and functional characterization of
cell signaling regulators. Indeed, yeast is the only

New Components, New Connections, experimental system where it is possible to track

the expression, localization, and activity of nearly
every component of the G protein signaling
New Compartments pathway, from the cell surface to the nucleus
(Fig. 1).
In the pheromone response pathway, the Gbg
Janna E. Slessareva and Henrik G. Dohlman*
subunits (Ste4 and Ste18) were long regarded as
the sole signal-transmitting component of the G
Signaling by cell surface receptors and heterotrimeric guanine nucleotide–binding proteins protein heterotrimer. The Ga subunit (Gpa1)
(G proteins) is one of the most exhaustively studied processes in the cell but remains a major focus was thought to regulate the amounts of free
of molecular pharmacology research. The pheromone-response system in yeast (see the Gbg: releasing Gbg when in the activated
Connections Map at Science’s Signal Transduction Knowledge Environment) has provided numerous GTP-bound form and sequestering Gbg when
major advances in our understanding of G protein signaling and regulation. However, the basic in the inactivated GDP-bound form. Evidence
features of this prototypical pathway have remained largely unchanged since the mid-1990s. New of a positive signaling role for Gpa1 came from
tools available in yeast are beginning to uncover new pathway components and interactions and studies showing that GTPase-deficient (and thus
have revealed signaling in unexpected locations within the cell. constitutively active) Gpa1 mutants, including
Gpa1Q323L, activate mating-specific gene tran-
scription and morphological changes in the
any extracellular signals are detected by also emerging as a versatile model for systems- absence of added pheromone (6).

M cell surface receptors and further trans-

mitted inside the cell by G proteins,
which serve as molecular switches (1). Activated
level analysis of complex signaling networks.
The yeast genome was the first among eukary-
otes to be sequenced. There exist comprehen-
However, the question still remained: How
does activated Ga transmit its signal and con-
tribute to the mating response pathway? A study
receptors promote exchange of guanosine tri- sive microarrays for transcription analysis, gene published this year brings us closer to answering
phosphate (GTP) for guanosine diphosphate deletion arrays for genome-scale phenotypic this question (7). A systematic analysis of nearly
(GDP) bound to G protein a subunits, triggering analysis, and gene fusion arrays for systematic 5000 gene deletion strains revealed seven genes
the dissociation of the G protein a and bg
subunits and thus allowing them to signal. In
yeast, mating pheromones activate a G protein
and protein kinase cascade that includes Fus3
and Kss1, two members of the mitogen-
activated protein (MAP) kinase family (2). G
protein activation is eventually terminated by the
intrinsic guanine triphosphatase (GTPase) activ-
ity of Ga subunits, a process that is accelerated
by proteins known as regulators of G protein
signaling (or RGS proteins) (1). Thus, the
intensity of the signal depends on the opposing
actions of receptors and RGS proteins. Proper
functioning of G protein–mediated signal trans-
duction mechanisms is extremely important,
because defects in these pathways are implicated
in a wide variety of diseases (3). Corresponding-
ly, a substantial fraction of all pharmaceuticals act
directly or indirectly on G protein–coupled re-
ceptors (4). Thus, it is possible that a new gen-
eration of drugs will act on RGS proteins and
other newly discovered regulators of G protein
activity (5).
Fig. 1. New features of RGS and G protein signaling in yeast. (Top) Activation of the receptor
The yeast Saccharomyces cerevisiae repre-
(Ste2) by mating pheromone leads to GTP binding and dissociation of the G protein a subunit
sents an attractive and appropriate model for (Gpa1) from the Gbg subunits (Ste4 and Ste18). Gbg activates multiple effectors at the plasma
investigating basic mechanisms of G protein membrane, including components of a MAP kinase signaling cascade. The RGS protein Sst2 binds
and MAP kinase signaling (2) and in particular to the receptor and accelerates Gpa1 GTP hydrolysis. Casein kinase I (Yck1 and Yck2) phos-
for finding new pathway regulators. Yeast is phorylates the receptor, displacing Sst2 and promoting receptor endocytosis. (Bottom) Gpa1 is also
present at the endosome. Activation by endocytosed receptor or another unknown factor may lead
Department of Biochemistry and Biophysics, University of to GTP binding and dissociation of Gpa1 from the Gb-like protein Vps15. Activated Gpa1-GTP then
North Carolina, Chapel Hill, NC 27599–7260, USA. binds directly to the phosphoinositide 3-kinase (PI3K) Vps34. Elevated PI3P recruits proteins
*To whom correspondence should be addressed. E-mail: containing FYVE domains and PX domains to the endosome. Note that Vps15 and Vps34 are part of
[email protected] a multiprotein complex that also includes Vps30 and Vps38 or Atg14.

1412 1 DECEMBER 2006 VOL 314 SCIENCE www.sciencemag.org

SPECIALSECTION
required for Gpa1Q323L-mediated responses. division arrest that normally precedes mating. signaling complexes that are distinct from those
Two of the genes, VPS34 and VPS15, encode Thus, whereas Ga signaling at the plasma known to exist at the plasma membrane. In sup-
the catalytic and regulatory subunits, respec- membrane is clearly important for the early steps port of this model, Gpa1Q323L promotes trans-
tively, of the sole phosphatidylinositol 3-kinase of the pheromone response, Ga signaling at the location of the PI3P-binding protein Bem1 to the
in yeast. Surprisingly, Vps34 and Vps15 are endosome appears to convey a fundamentally endosome (7). Bem1 functions as an adaptor
found at intracellular compartments, such as different signal. protein that promotes activation of other sig-
endosomes and Golgi, rather than at the plas- How does segregation of Gpa1 to two dis- naling components, including the small GTPase
ma membrane where receptors and G proteins tinct membrane compartments relate to ob- Cdc42 and the MAP kinase Fus3 (18). The
are normally thought to reside. Activated Ga served functional differences? As mentioned adaptor protein Ste5 serves a similar role in
localizes along with Vps34 at the endosome, above, there could be differences in the ac- transmitting the Gbg signal at the plasma mem-
binds to Vps34 directly, and triggers increased tivation step. Alternatively, there could be dif- brane (19).
production of the Vps34 product, phosphati- ferences in how the two pools of G protein are If history is any guide, lessons learned in
dylinositol 3-phosphate (PI3P). These obser- inactivated. For example, it is possible that the yeast will prove applicable to signaling events
vations suggest that Vps34 is a bona fide Ga internal pool of G protein is no longer at- in more complex organisms. The identification
effector capable of generating a second mes- tenuated by RGS proteins. Alternatively, both of Sst2 and other RGS family members in the
senger. The regulatory subunit Vps15 has a mechanisms might contribute. Recent studies mid-1990s led to a dramatic rethinking of how
seven WD40 domain repeat structure and in a variety of organisms suggest coordination G protein signaling pathways are organized and
binds directly to the inactive GDP-bound form between the receptors that activate the signal regulated. This view has now been further re-
of Gpa1, both hallmark features of known Gb and the RGS proteins that inactivate the fined with the realization that receptors and
subunits. These findings imply that Vps15 signal. RGS proteins show selectivity for RGS proteins cooperate to modulate G protein
functions as an alternate Gb, but one acting particular Ga subunits, but some RGS pro- activity. Moreover, the old dogma of G protein–
at the endosome instead of at the plasma teins can physically interact with the receptors mediated signaling only at the plasma membrane
membrane. In further support of this model, it (14). One particularly intriguing example of must now be modified to include signaling from
was shown that Vps15 is needed to target Ga receptor-RGS cooperation is provided by the internal compartments. These discoveries ben-
to the endosome, whereas the canonical Gb plant protein AtRGS1, which has both RGS- efited from the development of powerful genome-
subunit Ste4 targets Ga to the plasma mem- and receptor-like domains, suggesting that it scale proteomic and genomic tools available in
brane (7). functions both as a G protein activator and yeast. Our ability to fully exploit these tools is still
Although it is widely accepted that G pro- inactivator (15). More recent findings reveal evolving, but we can be confident that they will
teins signal at the plasma membrane, it has long that the yeast RGS protein Sst2 can bind eventually provide us with a truly global or
been recognized that they are also present at directly to the pheromone receptor Ste2 at systems-level understanding of how cells respond
intracellular compartments (8). Previous studies the plasma membrane (16). Presumably Sst2 to changes in their environment.
have implicated these intracellular G proteins in interaction with its cognate receptor ensures
the regulation of membrane traffic (9). It has also that pathway regulation is both rapid and
become increasingly evident that membrane receptor-specific. Regulation is receptor-specific References and Notes
trafficking and signal transduction events are because of selective binding to Sst2. Regu- 1. C. R. McCudden, M. D. Hains, R. J. Kimple, D. P.
Siderovski, F. S. Willard, Cell. Mol. Life Sci. 62, 551
closely linked and that G protein signals can lation is rapid, because receptor binding po-
(2005).
originate from intracellular compartments as sitions the RGS domain of Sst2 in close 2. Y. Wang, H. G. Dohlman, Science 306, 1508 (2004).
well as from the plasma membrane (10, 11). proximity to its substrate, Gpa1. In this sce- 3. Z. Farfel, H. R. Bourne, T. Iiri, N. Engl. J. Med. 340, 1012
Consequently, endosomes should be considered nario, RGS positioning close to Ga sets the (1999).
not only as a site of membrane sorting but also as threshold for pheromone response, prevent- 4. A. Wise, K. Gearing, S. Rees, Drug Discov. Today 7, 235
(2002).
a site of cell signaling (12). ing signaling at low pheromone concentra- 5. D. L. Roman et al., Mol. Pharmacol. 71, 169 (2006).
How is the internal pool of G proteins reg- tions. As signaling at the plasma membrane 6. M. Guo et al., Mol. Cell 12, 517 (2003).
ulated? One possibility is that Ga is initially wanes, endocytosed receptors might continue 7. J. E. Slessareva, S. M. Routt, B. Temple, V. A. Bankaitis,
activated by receptors at the plasma membrane to signal from the endosome or other internal H. G. Dohlman, Cell 126, 191 (2006).
8. M. Sato, J. B. Blumer, V. Simon, S. M. Lanier, Annu. Rev.
and is then delivered to intracellular compart- compartments, as reported for other receptors Pharmacol. Toxicol. 46, 151 (2006).
ments. Another possibility is that the agonist- (10). Phosphorylation of the receptor pro- 9. J. B. Helms, FEBS Lett. 369, 84 (1995).
bound receptor transits to the endosome and motes endocytosis and also results in disso- 10. A. Sorkin, M. Von Zastrow, Nat. Rev. Mol. Cell Biol. 3,
activates Ga directly. Yet a third possibility is ciation of the RGS protein, suggesting that 600 (2002).
11. A. Mor, M. R. Philips, Annu. Rev. Immunol. 24, 771
that Ga is activated by a distinct guanine nu- the RGS protein does not follow the receptor
(2006).
cleotide exchange factor located at the endo- as it undergoes endocytosis. Thus, one way 12. M. Miaczynska, L. Pelkmans, M. Zerial, Curr. Opin. Cell
some. In fact, in other organisms a number of that endosomal signaling may differ from Biol. 16, 400 (2004).
nonreceptor accessory proteins have been shown plasma membrane signaling is if the RGS 13. G. G. Tall, A. M. Krumins, A. G. Gilman, J. Biol. Chem.
278, 8356 (2003).
to activate Ga in vitro; of these a physiolog- protein is no longer in close proximity to the
14. M. Abramow-Newerly, A. A. Roy, C. Nunn, P. Chidiac, Cell.
ical role has been demonstrated for mammalian G protein. Signal. 18, 579 (2006).
Ric-8A, which activates Ga proteins during cell Lastly, there may be as yet undiscovered 15. J.-G. Chen et al., Science 301, 1728 (2003).
division (1, 8, 13). pathway regulators acting at the endosome, per- 16. D. R. Ballon et al., Cell 126, 1079 (2006).
Why would having segregated pools of Ga haps in response to PI3P production. Unlike 17. S. Misra, G. J. Miller, J. H. Hurley, Cell 107, 559
(2001).
be physiologically important? Signaling by many other second messengers, however, PI3P 18. D. M. Lyons, S. K. Mahanty, K. Y. Choi, M. Manandhar,
Gpa1Q323L may provide some clues. Although cannot diffuse freely and remains bound to E. A. Elion, Mol. Cell. Biol. 16, 4095 (1996).
it mimics most aspects of pheromone binding at membranes at the site of synthesis. Therefore, 19. E. A. Elion, J. Cell Sci. 114, 3967 (2001).
the cell surface, Gpa1Q323L at the endosome pref- PI3P might serve to recruit proteins with PI3P- 20. Supported by NIH grant P01-GM065533.
erentially activates just one of the two mating- binding domains to the endosome (17). Such
specific MAP kinases and fails to trigger the cell protein recruitment may allow assembly of 10.1126/science.1134041

www.sciencemag.org SCIENCE VOL 314 1 DECEMBER 2006 1413

Cell Signaling
dominant, ligand-independent activating muta-
PERSPECTIVE tions of Notch1 that require the S3 cleavage and
are associated with T cell acute lymphoblastic

Notch, a Universal Arbiter of leukemias (10). These mutations provide a clear

link between a signaling event and a specific dis-
ease but also serve as a basis to test therapies that
Cell Fate Decisions target the g-secretase activity of the Presenilin
complex. In other instances, the normal activity
Matthias Ehebauer,1 Penelope Hayward,2 Alfonso Martinez Arias2* of Notch provides a means for tampering with
cancers. For example, Wnt signaling–mediated
Members of the Notch family of receptors act as membrane-tethered transcription factors that are colorectal tumors can be curbed by inhibiting
tightly associated with binary cell fate decisions. Notch signaling acts as a molecular gate that Notch signaling to induce the differentiation of
allows cells to adopt or forfeit a particular fate. Interaction of Notch with ligands triggers a these tumors (11).
sequence of proteolytic cleavages that release the intracellular domain to the nucleus; this Genetic analysis has demonstrated the uni-
mechanism is a target of therapies for leukemias associated with Notch activation. Although the versality of Notch signaling as an arbiter of cell
molecular mechanism of Notch activation is well characterized, further analysis in an appropriate fate. In this process, Notch acts in a permissive
cellular context will provide new insight into Notch signaling. manner: It does not determine the fate of a cell
but rather whether a given fate is adopted (2, 4).
he formation of an organism relies on like repeats (2–4). Notch signaling is deceptively In addition to this signature function, there is

T the generation of diverse cell types that

provide the bricks for creating tissues
and organs. Observations initially made during
simple (Fig. 1C). The interaction of Notch with
DSL proteins induces a cleavage (S2) within the
Notch extracellular domain near the transmem-
evidence that Notch plays other roles in de-
velopment. For example, it is a central element of
a molecular oscillator that establishes segmenta-
studies of insect nervous system development brane region, generating a substrate for a ligand- tion in vertebrates (12) and, probably, in other
highlight two simple and linked strategies that independent cleavage (S3) catalyzed within the segmented organisms (13). In this task, it appears
underlie cell fate determination: (i) Cells attain transmembrane domain by the Presenilin com- that Notch also acts permissively through a
their fates through binary choices, and (ii) ini- plex. The S3 cleavage frees the intracellular do- mechanism similar to that involved in cell fate
tially, all cells in a given population can adopt an main of Notch (NICD) to enter the nucleus (5), decisions, coordinating more than determining
alternative fate, but only some maintain this new where it interacts with members of the CBF1, the activity of different cells. The notion that
fate stably, whereas others revert to the default Su(H), and Lag1 (CSL) family of transcription Notch has mostly a modulatory role has recently
fate (Fig. 1A). The population of cells involved in factors (3). CSL proteins are transcriptional re- been elegantly shown in the differentiation of
making this binary choice is called an equivalence pressors that are turned into activators through mouse embryonic stem cells (14). In a few in-
group, and the process of cell fate pruning usually interaction with NICD (3, 6). The CSL-NICD stances, Notch has also been shown to act in an
involves lateral inhibition, in which the cell that complex recruits other proteins, which regulate instructive fashion (15).
adopts the alternative fate blocks this choice in its the stability of the complex, ensuring a tight con- We understand in detail how NICD is re-
neighbors (1). These strategies for cell fate deter- trol over the signaling event. The CSL-NICD leased from the membrane and how it interacts
mination occur in numerous developmental pro- complex modulates the transcription of different with DNA binding effectors. What else is there
cesses in different cell types. Signaling by Notch targets, but its activity is frequently linked to the to be learned? One avenue of research will be
receptors is intimately associated with these expression of members of the Enhancer of Split, to determine how this functional cassette is
events and may provide a universal mechanism Hairy/Enhancer of Split–related, Hairy and En- plugged in to normal and pathological cell spec-
for cell fate determination (2). hancer of Split [E(spl)/HER/HES] family of ification networks. Also, increasing evidence
The central elements of Notch signaling are transcriptional regulators, which act as a core suggests that endocytosis and trafficking of
featured in an STKE Connections Map (3). The extension of the pathway (3). There is increasing ligands and receptors play central roles in
Notch receptors encompass a cohort of trans- evidence that Notch can signal in a CSL- Notch activation (Fig. 1C) (16, 17). Notably, en-
membrane proteins with an extracellular domain independent manner and that this also plays a docytosis of DSL ligands is a prerequisite for
composed of tandemly arranged epidermal role in development and disease (7). their functional interaction with the receptor, and
growth factor (EGF)–like repeats (36 in the ca- Detailed observations of receptors and lig- the S3 cleavage of Notch requires both endocy-
nonical form of Notch); a membrane-proximal set ands raise some questions that challenge the tosis and trafficking to an internal compartment.
of specific cysteine-rich repeats, the Lin12-Notch apparent simplicity of the pathway. For example, Furthermore, studies of Drosophila mutants
(LN) repeats; and an intracellular moiety that although a Notch receptor found in insects and suggest that endocytosis is used in two different
contains seven conserved ankyrin repeats, which higher eukaryotes has 36 EGF-like repeats and a manners to regulate Notch signaling: It ensures
mediate much of the function of the receptor (Fig. conserved intracellular domain, the nematode re- that there is no ligand-independent signaling,
1B) (2). Notch receptors are membrane-tethered ceptor and two of the mammalian forms (Notch 3 and it regulates ligand-dependent activity. In
transcription factors that are released into the and Notch 4) have fewer repeats and smaller in- the next few years, trafficking will probably
nucleus upon activation by the ligand (2, 4). The tracellular domains. The functional consequences emerge as a central element in the biology of
Notch ligands, collectively known as Delta, of these differences are not yet clear. Moreover, Notch and its therapeutic potential in Notch-
Serrate, and Lag2 (DSL) (3), are also transmem- the DSL ligands fall into two structurally different dependent diseases. Finally, an important area
brane proteins with extracellular arrays of EGF- classes, Delta-like and Serrate, which appear to of future research will involve interactions of
mediate different interactions with Notch. The Notch with other signaling pathways, in par-
1 STKE Connections Map (3) might serve as a ticular with Wnt (18, 19).
Department of Biochemistry, University of Cambridge, Cam-
bridge CB2 1GA, UK. 2Department of Genetics, University of framework to guide research into such questions. It is likely that in the near future, placing
Cambridge, Cambridge CB2 3EH, UK. Mutations that lead to Notch malfunction Notch signaling in an appropriate cellular context
*To whom correspondence should be addressed. E-mail: have been associated with various diseases (8), elucidating its interactions with other signal-
[email protected] including cancer (9). Of particular interest are ing pathways will not simply add detail to the

1414 1 DECEMBER 2006 VOL 314 SCIENCE www.sciencemag.org

SPECIALSECTION
Fig. 1. Notch receptor signaling and architecture. (A) All cells
within an equivalence group are in a particular state (yellow) and
possess similar amounts of Notch and Delta. A combination of
intrinsic and extrinsic signals induces a new cell state (light red)
and breaks this balance by increasing the abundance of Delta in
one or a few cells (dark red). Consequently, Notch signaling is
activated in neighboring cells, suppressing the alternative fate and
reverting these neighboring cells to their original fate through
lateral inhibition. (B) Domain architecture of the Notch receptor
family. The organization of human Notch1 is shown as an example.
All Notch receptors consist of two polypeptides at the cell surface,
except Drosophila, in which a linear form of the receptor might
exist at the cell surface. RAM, recombination signal-binding protein
for J-k (RBPJk)–associated molecule region; ANK, ankyrin; NLS,
nuclear localization sequence; Pro-Glu-Ser-Thr (PEST), degradation
motif. (C) Ligand binding to the Notch receptor triggers signaling
through successive proteolytic cleavages. The ligand (DSL) binds to
the EGF repeats (orange) in the extracellular domain of the Notch
receptor. A protease cleaves Notch at the S2 site, removing the bulk
of the extracellular domain. The identity of this protease is not
clear; it may be a disintegrin and metalloprotease protein (ADAM),
tumor necrosis factor–a converting enzyme (TACE), or Kuzbanian
(Kuz). The membrane-tethered intracellular domain is then cleaved
by the Presenilin complex at site S3, either in the plasma
membrane or after endocytosis, freeing the NICD. NICD then
moves to the nucleus, where it forms a binary complex with CSL,
which recruits coactivators (CoA) to form the transcription-
activating complex. Notch signaling can be regulated by
endocytosis. NICD becomes monoubiquitylated (Ub), targeting
the receptor to the lysosome for degradation. NICD in the nucleus
is degraded in a proteasome-dependent manner, although the
location of degradation is not clear. Solid arrows indicate
demonstrated routes; dashed arrows indicate routes tentatively
inferred from data.

molecular outline we have at the moment (3) but 3. M. Ehebauer, P. Hayward, A. Martinez-Arias, Notch 12. P. C. Rida, N. Le Minh, Y. J. Jiang, Dev. Biol. 265, 2 (2004).
rather reveal new vistas of this central signaling signaling pathway. Science’s STKE (Connections Map, as 13. A. Stollewerk, M. Schoppmeier, W. G. Damen, Nature
seen August 2006) (http://stke.sciencemag.org/cgi/cm/ 423, 863 (2003).
pathway. The focus so far has been on the stkecm;CMP_19043). 14. S. Lowell, A. Benchoua, B. Heavey, A. G. Smith, PLoS Biol.
elements of a linear cascade and their interac- 4. R. Kopan, J. Cell Sci. 115, 1095 (2002). 4, e121 (2006).
tions, but further consideration would suggest 5. E. H. Schroeter, J. A. Kisslinger, R. Kopan, Nature 393, 15. S. Bray, Semin. Cell Dev. Biol. 9, 591 (1998).
that it takes more than a collection of biochem- 382 (1998). 16. R. Le Borgne, A. Bardin, F. Schweisguth, Development
6. E. C. Lai, EMBO Rep. 3, 840 (2002). 132, 1751 (2005).
ical reactions to get cells to make functioning 7. A. Martinez Arias, V. Zecchini, K. Brennan, Curr. Opin. 17. R. Le Borgne, Curr. Opin. Cell Biol. 18, 213 (2006).
organisms. Gen. Dev. 12, 524 (2002). 18. A. W. Duncan et al., Nat. Immunol. 6, 314 (2005).
8. T. Gridley, Hum. Mol. Genet. 12, R9 (2003). 19. P. Hayward et al., Development 132, 1819 (2005).
References and Notes 9. F. Radtke, K. Raj, Nat. Rev. Cancer 3, 756 (2003). 20. Our research is supported by the Wellcome Trust.
1. P. Simpson, Development 109, 509 (1990). 10. A. P. Weng et al., Science 306, 269 (2004).
2. F. Schweisguth, Curr. Biol. 14, R129 (2004). 11. J. H. van Es et al., Nature 435, 959 (2005). 10.1126/science.1134042

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BREVIA
were 0.54 and 0.68 Mg C ha−1 year−1, after
considering the uncertainty introduced by the
lack of thickness-change monitoring. We took
Old-Growth Forests Can more than enough samples to detect the ob-
served SOC change. In fact, statistical analysis
Accumulate Carbon in Soils shows that 20 samples taken every 8 to 10
years of sampling interval (or 100 samples
every 5 years) would be sufficient to detect the
Guoyi Zhou,1*† Shuguang Liu,2* Zhian Li,1 Deqiang Zhang,1 Xuli Tang,1
observed SOC change rate in these forests at a
Chuanyan Zhou,1 Junhua Yan,1 Jiangming Mo1
95% confidence level. More samples would
be required at shorter sampling intervals to de-
ld-growth forests have traditionally We conducted a study to measure the long- tect the observed change, given the observed

O been considered negligible as carbon

sinks because carbon uptake has been
thought to be balanced by respiration (1). We
term dynamics (1979 to 2003) of SOC stock in
old-growth forests [age > 400 years (3)] at the
Dinghushan Biosphere Reserve (23°09′21″N to
spatial variability of SOC concentration and
bulk density.
The driving forces for this observed high
show that soils in the top 20-cm soil layer in 23°11′30″N and 112°30′39″E to 112°33′41″E) rate of SOC increase in the old-growth forests
preserved old-growth forests in southern in Guangdong Province, China. The estimation are not clear at present and deserve further
China accumulated atmospheric carbon at an of SOC stock change requires a series of study. This study suggests that the carbon cycle
unexpectedly high rate from 1979 to 2003. measurements of SOC concentration, bulk den- processes in the belowground system of these
This phenomenon indicates the need for future sity, and soil thickness taken at different points forests are changing in response to the changing
research on the complex responses and ad- in time (4, 5). In this study, we observed long- environment. This result directly challenges the
aptation of belowground processes to global term changes in SOC concentration and bulk prevailing belief in ecosystem ecology regard-
environmental change. density but did not measure changes in soil ing carbon budget in old-growth forests (1) and
Understanding the locations and driving thickness in the old-growth forests. Although supports the establishment of a new, non-
forces of carbon sources and sinks at plot-to- soil thickness dynamics were not monitored, equilibrium conceptual framework to study
global scales is critical for the prediction and their possible contribution to the uncertainty in soil carbon dynamics. Our study further high-
management of the global carbon cycle and the results was analyzed and quantified by using lights the need to focus on the complexity of
ultimately the behavior of the Earth’s climate upper and lower bounds of possible SOC change the belowground processes, as advocated in
system (2). Major uncertainties remain in the (Materials and Methods). previous research (6, 7), and the importance of
geospatial distribution of terrestrial carbon Results show that SOC concentration in establishing long-term observation studies on
sources and sinks and the mechanisms that the top 20-cm soil layer increased between the responses of belowground processes to
drive the distribution and its change. Research 1979 and 2003 from about 1.4% to 2.35% at global change.
efforts have largely been focused on the in- an average rate of 0.035% each year, which
vestigation and quantification of the impacts was significantly different from 0 at a = 0.05. References and Notes
of climate variability and land use activities At the same time, the mean bulk density of the 1. E. P. Odum, Science 164, 262 (1969).
on the carbon cycle at various spatial and top 20-cm soil layer decreased significantly 2. Intergovernmental Panel on Climate Change, Climate
temporal scales. The soil carbon balance of (a = 0.05), with an average rate of 0.0032 g Change 2001: The Scientific Basis (Cambridge Univ.
old-growth forests has received little atten- cm−3 year–1. Measurements on a total of 230 Press, Cambridge, 2001).
3. C. D. Shen et al., Chin. Sci. Bull. 44, 251 (1999).
tion. It is generally accepted that soil organic composite soil samples collected between 4. W. M. Post, R. C. Izaurralde, L. K. Mann, N. Bliss, Clim.
carbon (SOC) levels in old-growth forests 1979 and 2003 suggested that SOC stock in Change 51, 73 (2001).
are in a steady state (1). To our knowledge, the top 20-cm soil layer increased significantly 5. F. Conen, M. V. Yakutin, A. D. Sambuu, Glob. Change
Biol. 9, 1515 (2003).
the long-term dynamics of SOC in old-growth during that time (P < 0.0001), with an average
6. R. Lal, Science 304, 1623 (2004).
forests and the validity of the above percep- rate of 0.61 Mg C ha−1 year−1 (Fig. 1). The 7. C. A. Johnston et al., Front. Ecol. Environ. 2, 522
tion have not been tested. lower and upper bounds of this average rate (2004).
8. G.Z. acknowledges support from the Chinese Ecosystem
Research Network (CERN), the Chinese Academy of
Science (project KSCX2-SW-120), and the Natural Science
Foundation of China (project 30470306). S.L.’s work was
supported by the USGS Geographic Analysis and
Monitoring Program and the Earth Surface Dynamics
Program. Work was performed under USGS contract
03CRCN0001.

Supporting Online Material

www.sciencemag.org/cgi/content/full/314/5804/1417/DC1
Materials and Methods
References

18 May 2006; accepted 13 September 2006

10.1126/science.1130168

1
South China Botanical Garden, Chinese Academy of Sci-
ences, Guangzhou, 510650, China. 2SAIC, U.S. Geological
Survey (USGS) Center for Earth Resources Observation and
Fig. 1. Temporal changes of (left) soil organic carbon concentration, bulk density, and (right) soil Science, Sioux Falls, SD 57198, USA.
organic carbon stock in the top 20-cm soil layer in broadleaved old-growth forests in Dinghushan *These authors contribute equally to this work.
Nature Reserve. Upper and lower bounds contain the uncertainty introduced by the lack of †To whom correspondence should be addressed. E-mail:
monitoring of soil thickness during the study period. Error bars indicate standard deviation. [email protected]

www.sciencemag.org SCIENCE VOL 314 1 DECEMBER 2006 1417

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RESEARCH ARTICLE
bloom is ideal for exploring the temporal
relationship between ocean productivity and
regional clouds.
Phytoplankton and Cloudiness Temporal correlation between [Chl a] and
Reff. We restrict our analysis to liquid-water
in the Southern Ocean clouds. Figure 1 shows surface [Chl a] and
effective radius (Reff) in the SO for the 6
years of available data. Figure 1A demon-
Nicholas Meskhidze1*† and Athanasios Nenes1,2 strates that the observed enhancement of
primary productivity near South Georgia
The effect of ocean biological productivity on marine clouds is explored over a large Island is a localized phenomenon that typi-
phytoplankton bloom in the Southern Ocean with the use of remotely sensed data. Cloud cally occurs between September and Feb-
droplet number concentration over the bloom was twice what it was away from the bloom, and ruary. Despite its regularity, Fig. 1 shows
cloud effective radius was reduced by 30%. The resulting change in the short-wave radiative flux substantial annual variations in the bloom’s
at the top of the atmosphere was –15 watts per square meter, comparable to the aerosol temporal appearance, spatial extent, and
indirect effect over highly polluted regions. This observed impact of phytoplankton on clouds is strength, with strong anticorrelation between
attributed to changes in the size distribution and chemical composition of cloud condensation [Chl a] and Reff. In 2001 and 2002, the
nuclei. We propose that secondary organic aerosol, formed from the oxidation of phytoplankton- smallest R eff coincided with the largest
produced isoprene, can affect chemical composition of marine cloud condensation nuclei and enhancement in [Chl a], whereas the largest
influence cloud droplet number. Model simulations support this hypothesis, indicating that summertime Reff was observed during the
100% of the observed changes in cloud properties can be attributed to the isoprene secondary austral summer of 2000 and 2001 with neg-
organic aerosol. ligible phytoplankton levels. Such systemat-
ically significant temporal anticorrelation of
arine aerosols strongly affect prop- the island are a natural laboratory for in- [Chl a] and Reff suggests a link between

M erties and lifetime of stratiform

clouds, influencing Earth’s radiation
budget and climate. The role of oceanic biota
vestigating the effects of marine productivity
on clouds. Waters in this area can support
massive phytoplankton blooms (5, 6), with
ocean biological productivity and marine
cloud properties; however, the results shown
in Fig. 1 alone cannot be used to ascertain a
in modifying chemical composition and size chlorophyll a concentrations ([Chl a]) more causal relationship, given that other factors
distribution of marine cloud condensation than an order of magnitude higher than the such as variation in background aerosol size
nuclei (CCN) has been one of the most background (7, 8). Because the surface [Chl a] distribution and cloud dynamics may also
intriguing questions in climate studies. Pro- in this area can be used as a reliable proxy for affect Reff. Phytoplankton productivity and
duction of sulfate from the oxidation of di- the primary production (5, 9), satellite re- clouds could also be influenced by large-
methylsulfide (DMS), proposed by Shaw (1) trievals can provide the link between ocean scale atmospheric circulation; such a mech-
and explored by Charlson et al. (2) [the CLAW productivity and clouds. Strong and persistent anism, if it exists, would exhibit a high
hypothesis, named after the authors of the paper, westerlies (10) make it possible to examine correlation between [Chl a] and regional
Charlson, Lovelock, Andreae, and Warren (2)] cloud properties upwind and downwind of cloud properties even if ocean productivity
and primary emissions of biogenic organic the bloom, and the periodic nature of the had no effect on clouds.
matter from wave breaking (3, 4) have been
suggested as possible mechanisms by which
phytoplankton can modulate properties of
marine clouds. In this work, remotely sensed
data from the Moderate Resolution Imaging
Spectroradiometer (MODIS) and Sea-viewing
Wide Field-of-view Sensor (SeaWiFS) were
combined with the National Center for Envi-
ronmental Prediction (NCEP)–generated mete-
orological fields to examine the effects of
ocean productivity on cloud microphysical
and radiative properties and explore an al-
ternative pathway by which phytoplankton
may affect marine CCN. The data analysis is
carried out in the Southern Ocean (SO) near
South Georgia Island. Because of its unique
spatial location and topography, waters near

1
School of Earth and Atmospheric Sciences, Georgia Institute
of Technology, Atlanta, GA 30332, USA. 2School of Chemical
and Biomolecular Engineering, Georgia Institute of Technol-
ogy, Atlanta, GA 30332, USA.
*Present address: Department of Marine, Earth, and At-
Fig. 1. The 8-day averaged (A) SeaWiFS-observed chlorophyll a and (B) MODIS-retrieved cloud
mospheric Sciences, North Carolina State University, Raleigh,
NC 27695, USA. effective radius. Data for [Chl a] is gridded at a resolution of 9 by 9 km and zonally averaged
†To whom correspondence should be addressed. E-mail: between 49°S and 54°S; data for Reff is gridded at a resolution of 1° by 1° and averaged in the
[email protected] area of 49° to 54°S and 35° to 41°W. White areas in (A) indicate missing data.

www.sciencemag.org SCIENCE VOL 314 1 DECEMBER 2006 1419

RESEARCH ARTICLE
Effect of phytoplankton on clouds. To where LWC is cloud liquid water content; rw of variation of the SO marine cloud micro-
constrain the effect of phytoplankton on is the density of liquid water; H is the cloud physics over the bloom may be comparable
clouds, we adopt an approach similar to thickness; t and Reff are MODIS-observed to anthropogenic indirect aerosol effects.
Chylek et al. (11). A rectangular geograph- cloud optical depth and effective droplet ra- Other factors that may potentially affect
ical area was selected in the SO from 55°W dius, respectively; and k is the constant ~0.8 CDNCs near the bloom are long-range trans-
to 21°W and 42°S to 60°S, then divided into (14). H was estimated as the distance between ported Patagonian dust and sea salt. Because
153 cells of 2° by 2°. The background colors the cloud lifting condensation level, zLCL winds in the SO typically flow eastward
in Fig. 2A correspond to the monthly (used as a cloud base proxy), and the cloud- (10), the presence of mineral dust will be
averaged surface [Chl a] for the year with top height. The zLCL is a strong function of manifested by noticeable decrease in aerosol
the largest bloom on record (Fig. 1). In- relative humidity and temperature and is optical thickness (AOT) from west to the
creasing grid box number on Fig. 2B tra- calculated using NCEP reanalysis surface east. Figure 2B shows that this is not the
verses through a row of cells in an eastward data (15). Cloud-top height was computed case. The enhanced productivity near South
direction. Sharply increased [Chl a] between by matching adiabatic liquid water path Georgia Island is primarily controlled by
48°S and 56°S indicates enhanced marine (LWP) with the MODIS-observed LWP. Be- ocean upwelling, not by dust-Fe fertilization
productivity near South Georgia Island. cause the average Reff in the study area was (5, 6, 9). Therefore, dust is not responsible
Figure 2B suggests that average Reff for typically less than the threshold effective for changes in either ocean productivity or
background clouds in this area is ~14 mm, radius for precipitation (~14 to 15 mm) (16), Reff. An increase in submicrometer-sized
with a sharp decrease (~10 mm) in the vicin- we assumed that precipitation did not cause sea-salt particles can certainly affect CDNC,
ity of the bloom. Although MODIS-retrieved substantial deviation of observed LWP from but this must be accompanied by an increase
Reff may be biased to larger sizes compared the adiabatic value. The calculated liquid in surface wind speed over the bloom, which
with in situ measurements, it is reasonable water cloud thickness was between 250 and we did not observe (Fig. 2B). Although the
to expect that errors in observed relative 400 m, consistent with observations (17, 18). general trend of increase in CDNC between
changes of Reff are small (12). This figure The uncertainty in H does not contribute 42°S and 54°S and decrease further south-
demonstrates that, on average, water clouds considerably to the Reff variability. ward can be associated with the variation in
near the bloom region have effective droplet Figure 2B shows that the calculated the surface wind speed (Fig. 2B), there is no
radii 30% smaller than those of background monthly averaged CDNC outside the bloom clear relationship between the two near the
clouds over the SO. area is ~55 cm−3, whereas in the bloom re- bloom region.
Cloud droplet number concentration gion, CDNC increases sharply approaching Analysis of Fig. 2 indicates strong cou-
(CNDC) can be used as a direct micro- twice the background level. Comparison of pling between observed changes in marine
physical link between the biology and cloud these values with the average summertime biological productivity and microphysical
properties. From the remote sensing data, CDNCs for the Southern Ocean Cloud properties of warm clouds over the bloom.
CDNC (cm−3) can be estimated as (13, 14): Experiment (SOCEX) shows that the esti- We examined the possibility that both
mated CDNC outside the bloom area com- processes are driven by the same large-scale
pares well with 57 cm−3 reported for the influence.
SOCEX baseline conditions, whereas cal- Role of meteorology. Strong winds (asso-
3LWC
CDNC ¼ culated CDNCs over the bloom are close to ciated with cyclonic circulation) can cause
4prw R3ef f k ones reported for the clouds affected by vertical mixing and upwelling of nutrient-
t 1
¼ ð1Þ anthropogenic emissions (109 cm−3) (19). rich waters from below the mixed-layer
2pkR2ef f H This comparison suggests that the magnitude depth, fueling photosynthesis and causing

Fig. 2. (A) Monthly averaged (11 December 2001 to 8 January 2002) 4-km (green), MODIS cloud-top effective radius of liquid droplets (blue), estimated
resolution SeaWiFS-observed surface [Chl a]. The black color over the ocean cloud droplet number concentration (red), aerosol optical thickness (black), and
denotes the missing data due to clouds. The South Georgia Island boundary NCEP reanalysis–generated surface wind speed (purple) as a function of the grid
(54.3° to 55.0°S, 35.3° to 38.3°W) is located between cells 111 and 112. (B) The cell number. Tick markers at every 17 cells correspond to the starting point of the
2° by 2° square monthly averaged SeaWiFS surface chlorophyll concentration next west-to-east row in (A). Broken line indicates the missing data.

1420 1 DECEMBER 2006 VOL 314 SCIENCE www.sciencemag.org

RESEARCH ARTICLE
large-scale phytoplankton blooms. Such Reff in the inside region is markedly different wave radiative transfer code (24); Rc and Ac are
deep water entrainments may also be asso- from that of the outside region; inside, the monthly averaged MODIS-observed cloud
ciated with a depression in sea-surface tem- effect of [Chl a] on Reff has by far the stron- albedo and cloud fraction, respectively; and
peratures (SST) (20, 21) that may last up to 2 gest impact of all parameters examined. (ii) DlnN db is relative change in calculated
weeks and considerably influence properties Can changes in meteorological parameters CDNC [DlnNdb = (Nd – Nb)/ Nd], where Nd
of marine stratocumulus clouds (22). Cyclones affect Reff while changing [Chl a]? Linear and Nb are average droplet numbers over the
would therefore generate a correlation (but no multiple regression analysis suggests that in bloom and in the background air, respec-
causality) between [Chl a] and cloud properties. the outside region, the change in Reff is tively. Because of variability of background
The results of linear multiple regression analysis primarily associated with variability in mete- CDNC in zonal direction (Fig. 2B), Nb was
of satellite-retrieved and model-generated pa- orological parameters, whereas in the inside estimated separately for each longitude using
rameters shown in Table 1 suggest that cloud region, [Chl a] is the single most important a linear fit between the locations farthest
properties over the bloom are not influenced by parameter controlling the Reff (Table 1). from the bloom (square numbers 1 and 17,
cyclonic winds. To quantify the possible influ- As a result of this analysis, we concluded 18 and 34, and so on, in Fig. 2A).
ence of phytoplankton on clouds, the analysis that over the bloom, the relationship between Figure 3 shows considerable variation in
was carried out separately for the region with the ocean productivity and Reff of warm TOA mean short-wave forcing in the study
enhanced productivity (48°S to 56°S) near clouds is unique to these two variables and area resulting from changes in properties of
South Georgia Island and the areas with rel- does not extend to large-scale meteorological liquid clouds. However, the remarkable fea-
atively low [Chl a] (42°S to 48°S and 56°S to parameters—i.e., biological productivity is ture of Fig. 3 is the very strong cooling near
60°S), hereafter referred to as the inside and the prime cause for changes in cloud micro- the bloom, reaching ~ –15 W m−2. Such a
outside regions, respectively. physical and radiative parameters. large change in TOA short-wave radiation is
The analysis addresses two main ques- Radiative forcing. The perturbation in comparable in magnitude with the aerosol
tions: (i) Do meteorological parameters and short-wave radiation (DF) at the top-of-the indirect effect in highly polluted regions
[Chl a] affect Reff differently in the inside and atmosphere (TOA) within our study area is (24–26), highlighting the need for improved
the outside regions? Table 1 shows a strong estimated as (23) quantification of interactions between marine
difference in the relationship of meteorologi- biota, aerosols, clouds, and climate. Clearly,
cal parameters and [Chl a] with Reff in the 1 the link between ocean productivity and
inside and outside regions. Outside, Reff is DF ¼ − Fin Ac Rc ð1 − Rc ÞDln Ndb ð2Þ change in cloud properties is in the modifi-
3
mainly controlled by large-scale atmospheric cation of CCN. We next examined whether
parameters (i.e., column precipitable water where Fin is the monthly averaged solar production of secondary organic aerosol
vapor, SST, surface wind speed, and AOT), flux at the top of marine liquid clouds cal- (SOA) from the oxidation of phytoplankton-
and correlation between Reff and [Chl a] is culated with the NASA Global Modeling produced isoprene can lead to considerable
minor. The relationship between [Chl a] and Initiative (GMI) with implemented short- changes in marine CCN.
Phytoplankton isoprene SOA and its effect
on CCN. Although organosulfur emissions and
Table 1. The influence of meteorological parameters and [Chl a] on the Reff in the Study area of the the transfer of surface active organic matter
Southern Ocean. Meteorological parameters used as independent variables were selected according to from the oceanic surface layer to the atmo-
Kaufman et al. (56). We analyzed dependence of Reff on (i) MODIS-retrieved AOT, SST, cloud-top sphere have been studied in detail, little is
temperature (indicator of cloud height), and total precipitable water vapor (indicator of convergence); known about the effect of phytoplankton-
(ii) SeaWiFS-observed [Chl a]; and (iii) NCEP and National Center for Atmospheric Research (NCAR) produced nonmethane hydrocarbons (NMHC)
reanalysis–generated surface wind speed, equivalent potential temperature difference between 500 and on marine aerosol. Oceans are known to be a
925 hPa, and the broad-scale vertical motion at 850 hPa. The logarithm of the AOT is used to reduce
potential source of NMHC and particularly
nonlinearity in the regression (56). The parameters are ranked by order of importance based on the
correlation with Reff in the outside region. Columns to the right of correlation coefficients show the
change in Reff associated by the multiple regression with the changes in meteorological parameters and
[Chl a]. To compare multiple regression coefficients of variables of different magnitudes and dispersions,
we standardized all variables by subtracting the mean and dividing the result by the standard deviation.
Therefore, coefficients given in the “Change in Reff” columns show the average amount of change in Reff
when each meteorological parameter and [Chl a] change by one standard deviation, while keeping
others constant. The range around the sample regression coefficients was determined for 95%
confidence interval.
Outside the bloom area
Inside the bloom area
(42°S to 48°S and
(48°S to 56°S)
56°S to 60°S)
Correlation to Change in Correlation to Change in
Reff Reff Reff Reff
Total column precipitable water vapor 0.67 ± 0.08 0.48 ± 0.10 –0.02 ± 0.12 –0.02 ± 0.10
Sea-surface temperature 0.65 ± 0.08 0.63 ± 0.09 0.01 ± 0.12 0.01 ± 0.11
Surface wind speed –0.52 ± 0.09 –0.50 ± 0.11 –0.14 ± 0.12 –0.15 ± 0.10
ln(AOT) –0.43 ± 0.10 –0.41 ± 0.09 –0.01 ± 0.12 –0.01 ± 0.13 Fig. 3. Change in TOA short-wave radiation. The
Potential temperature difference 0.34 ± 0.10 –0.40 ± 0.11 –0.09 ± 0.12 –0.08 ± 0.11 radiative effect was evaluated for the change in
Vertical velocity –0.27 ± 0.10 0.23 ± 0.09 –0.10 ± 0.12 –0.12 ± 0.10 albedo of warm marine clouds. Calculations are
Eastern wind at 850 hPa 0.22 ± 0.10 0.27 ± 0.12 0.10 ± 0.12 0.01 ± 0.11
carried out using monthly averaged MODIS-
observed data at 1° by 1° resolution and the
Cloud-top temperature –0.02 ± 0.11 –0.01 ± 0.09 –0.02 ± 0.12 –0.02 ± 0.11
GMI-supplied monthly averaged solar flux at 4°
[Chl a] 0.18 ± 0.14 0.19 ± 0.10 –0.48 ± 0.11 –0.49 ± 0.09
by 5° resolution.

www.sciencemag.org SCIENCE VOL 314 1 DECEMBER 2006 1421

RESEARCH ARTICLE
isoprene (27–29); observed atmospheric con- that both blooms had similar average [Chl a], isoprene and SOA concentrations over the
centrations of isoprene in remote SO are high were located in the SO at comparable lati- bloom.
(~0.25 parts per billion by volume) (30)— tudes, had similar SSTs and mixed-layer To evaluate the role of this potentially
about one-fifth the amount of typical boundary- depths, and occurred in the same season, important organic aerosol source on cloud
layer isoprene concentration over the Amazon such large discrepancies in dissolved iso- microphysics, we calculated CDNC using a
(31). Atmospheric oxidation of isoprene may prene concentrations could arise from the cloud parcel model (44). The simulations
lead to formation of SOA (32–34). Because difference in phytoplankton species. Labora- were performed for different concentrations
SOA concentrations in remote marine regions tory studies show that isoprene production of organic aerosol, non–sea-salt (nss) sulfate,
are very small (35), ocean-emitted isoprene rates between diatoms, dinoflagellates, and and updraft velocities. The range of SOA
could contribute considerably to the organic cocolithophores can vary over orders of mag- concentrations examined (0 to 250 ng m−3)
fraction of marine CCN. We propose that nitude (28, 39, 40). Production rates sug- corresponds to the estimated SOA over the
isoprene SOA can affect CCN composition gested in (36) and used in Table 2 for CwA are background air and at the center of the bloom
and contribute to the observed changes in applicable to species typical for the oligo- (Table 2). In all simulations, aerosol number
cloud properties. To evaluate the plausibility trophic oceans (cyanobacteria, picoeucariotes, remains constant; therefore, addition of SOA
of this hypothesis, we estimated atmospheric and coccolithophores) and are likely not corresponds to the condensation growth and
concentrations of isoprene, the resulting SOA, representative of the blooms in the SO where aging of marine aerosols (44). Ambient mea-
and its subsequent effect on CCN. Table 2 microphytoplankton typically contributes surements and chamber experiments showed
summarizes [Chl a] inside the bloom, esti- >70% of total cell counts, followed by nano- that 2-methyltetrols and C5 alkene triols are
mated sea-air fluxes, and concentrations of and picophytoplankton (41–43). Inside the some of the main particulate-phase oxidation
isoprene in the marine boundary layer (MBL). bloom, diatoms were five times more abun- products of isoprene under low-NOx con-
Seawater-dissolved isoprene in the bloom dant than the next contributor, dinoflagellates; ditions (32, 45–48). Because the reaction
region (CwA ) was estimated with the approach species measured (in order of their abun- pathways leading to production of isoprene
of Palmer and Shaw (36). dance) were Thallasiosira sp. 1, Nitzschia SOA and chemical properties of the oligo-
For comparison, in Table 2 we include spp., and Chaetoceros spp. (41). In the ab- mers with high molecular weight remain
observed [Chl a] and dissolved isoprene sence of any local data and good similarity little understood (45), we assumed that
concentrations for the iron-enriched North with SOFeX-N (42), dissolved isoprene in ~20% of organic particulate mass is water
Patch of the Southern Ocean Iron Enrich- the bloom was estimated with SOFeX-N mea- soluble with chemical properties corre-
ment Experiment (SOFeX-N) (37). The sured isoprene concentration and scaled with sponding to 2-methyltetrols (44). Cloud-base
average surface ocean [Chl a] measured for [Chl a]. Estimated sea-air fluxes of isoprene updraft velocity is constrained by estimated
SOFeX-N and for the bloom in our study are orders of magnitude higher than the CDNCs outside the bloom region. The back-
area are comparable (Table 2), yet SOFeX-N monthly averaged fluxes (0.4 × 108 to 8 × ground CDNC (the low left corner on Fig. 4,
isoprene concentration was more than three 108 molecules cm−2 s−1) suggested for the A and B) is well represented by trimodal
orders of magnitude higher than estimated SO (36), indicating that the global marine marine aerosol size distribution and chemical
for the bloom. The chlorophyll content of isoprene flux of ~0.1 Tg C year−1 (36) composition (44) with updraft velocities
seawater, as sensed by the satellite, is related should be viewed as the low-end estimate. between 0.35 to 0.5 ms−1, typical of marine
to the rate of isoprene production (38). Given In Table 2, we also include estimated MBL stratiform clouds (15, 49). Figure 4 shows
that the excess amount of organic mass
increases CDNC; modification of the ambi-
Table 2. Ocean chlorophyll a, fluxes, and atmospheric concentrations of isoprene. [Chl a] inside the ent size and CCN activity of marine aerosols
bloom is based on SeaWiFS observed chlorophyll a data retrieved at 9-km resolution. Maximum and due to addition of organic mass can explain
minimum concentrations correspond to the middle and the edge of the bloom. CwA was estimated up to 60% of the droplet number concentra-
according to Palmer and Shaw (36). The isoprene production rate was calculated by multiplying tion over the bloom.
suggested rates of 1.8 ± 0.7 mmol isoprene produced (grams phytoplankton chlorophyll a)−1 day−1 In addition to enhanced concentration of
by SeaWiFS [Chl a]. The range of dissolved isoprene is from Wingenter (57). CwB was estimated using dissolved isoprene, ocean waters in blooms
SOFeX-N measured surface values and scaled with SeaWiFS [Chl a]. The sea-air isoprene flux F was have commonly been characterized by ele-
parameterized as F = kI(Csw – CBL/H), where kI is the piston velocity, Csw is the estimated seawater vated levels of DMS, atmospheric oxidation
concentration of isoprene in the bloom, CBL is marine boundary layer isoprene concentration, and H of which is a major source of nss sulfate in
is the Henry’s law constant for isoprene (58). For the typical range of atmospheric and oceanic
remote marine air. Model simulations show
isoprene concentrations, the second term in the equation is at least an order of magnitude smaller
(38) and therefore is ignored here. The kI ¼ 0:39U10 2
(Sc=660)−0:5 (59), where U10 is NCEP (left side of Fig. 4) that for the range of nss
reanalysis–generated monthly averaged wind speed at 10-m height, and Sc is the Schmidt number sulfate measured over the SO (50) sea-salt
calculated using isoprene molar volume and MODIS-observed ocean temperatures. Amazon fluxes CCN and nss CCN may account for the en-
are the median fluxes over the tropical forest site of the Peruvian Amazon (31). Assuming that hanced CDNC over the bloom. However,
isoprene oxidation had no significant impact on OH levels in the MBL (60), the average MBL Fig. 4 also shows that when mixed with
isoprene concentration above the bloom was calculated with the use of FB, the average isoprene SOA, even the minimal concentration of nss
lifetime of 2 hours and boundary layer height of 600 to 1000 m. The SOA concentration was sulfate can fully account for the observed
estimated with the use of global isoprene SOA yield of 3% (35, 46). enhancement of CDNC. This is important,
considering that regions with high isoprene
Estimated MBL productivity may not coincide with elevated
Dissolved isoprene Isoprene flux
[Chl a] (mg m−3) concentration levels of DMS (30) or under conditions
concentration (nM) (10 molecules cm−2 s−1)
8
(ng m−3) with biological net DMS consumption in a
Bloom SOFeX C wA SOFeX C wB FA FB Amazon Isoprene SOA bloom (51).
Our model results suggest that phyto-
Average 3.0 2.4 0.03 31.4 36.3 1.8 2370 18200 1920 50
plankton isoprene emissions could contribute
Max 12.7 2.6 0.13 >40 145 8.6 9470 20000 7700 230
to the organic fraction of marine CCN and be
Min 0.1 0.1 0.003 <10 6.1 0.2 395 7000 320 5
a viable mechanism by which ocean biota

1422 1 DECEMBER 2006 VOL 314 SCIENCE www.sciencemag.org

RESEARCH ARTICLE
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15. M. G. Lawrence, Bull. Am. Meteorol. Soc. 86, 225 61. This work was supported by a NASA New Investigator
required to provide accurate estimates of sea-
(2005). Award, an NSF CAREER award and a Blanchard-Milliken
air fluxes of biogenic volatile organic com- 16. H. Shao, G. Liu, J. Geophys. Res. 109, D07205 Young Faculty Fellowship. N.M. acknowledges support
pounds (VOCs) in different parts of the (2004). from a NASA Goddard Visiting Fellowship. We also thank
ocean. Work is also needed to constrain the 17. C. S. Bretherton et al., Bull. Am. Meteorol. Soc. 85, 967 C. O’Dowd, M. C. Facchini, S. N. Pandis, and three
chemical composition of SOA in marine en- (2004). anonymous reviewers for their thoughtful comments.
18. N. Meskhidze, A. Nenes, W. C. Conant, J. H. Seinfeld,
vironments and its effect on aerosol acti- J. Geophys. Res. 110, D16202 (2005). Supporting Online Material
vation. Future campaigns may provide the 19. S. S. Yum, J. G. Hudson, J. Geophys. Res. 109, D06204 www.sciencemag.org/cgi/content/full/1131779/DC1
Materials and Methods
evidence for the importance of this new (2004).
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anthropogenic air pollutants (53, 54) and D. S. Covert, Geophys. Res. Lett. 31, L06113 (2004). Include this information when citing this paper.

www.sciencemag.org SCIENCE VOL 314 1 DECEMBER 2006 1423

REPORTS
(J2000.0). This is, within the quoted statistical
error and the systematic pointing uncertainty
of the H.E.S.S. telescopes (~20′′ in both the
Fast Variability of Tera–Electron Volt right ascension and declination directions)
compatible with the nominal (radio) position
g Rays from the Radio Galaxy M87 (13) of the nucleus of M87 (a = 12h30m 49.4s,
d = +12°23′28′′). Considering the angular res-
olution of H.E.S.S., the source is consistent with
F. Aharonian,1 A. G. Akhperjanian,2 A. R. Bazer-Bachi,3 M. Beilicke,4* W. Benbow,1 D. Berge,1 a pointlike object with an upper limit for a
K. Bernlöhr,1,5 C. Boisson,6 O. Bolz,1 V. Borrel,3 I. Braun,1 A. M. Brown,7 R. Bühler,1 Gaussian surface-brightness profile of 3 arc min
I. Büsching,8 S. Carrigan,1 P. M. Chadwick,7 L.-M. Chounet,9 G. Coignet,10 R. Cornils,4 (99.9% confidence level). At the distance of M87
L. Costamante,1,23 B. Degrange,9 H. J. Dickinson,7 A. Djannati-Ataï,11 L. O'C. Drury,12 (16 Mpc), this corresponds to a radial extension
G. Dubus,9 K. Egberts,1 D. Emmanoulopoulos,13 P. Espigat,11 F. Feinstein,14 E. Ferrero,13 of 13.7 kpc, which can be compared with the
A. Fiasson,14 G. Fontaine,9 Seb. Funk,5 S. Funk,1 M. Füß ßling,5 Y. A. Gallant,14 B. Giebels,9 large-scale structure of M87 as seen at radio
J. F. Glicenstein, P. Goret, C. Hadjichristidis, D. Hauser,1 M. Hauser,13 G. Heinzelmann,4
15 15 7
wavelengths (Fig. 1). A constraint on the size of
G. Henri,16 G. Hermann,1 J. A. Hinton,1,13 A. Hoffmann,17 W. Hofmann,1 M. Holleran,8 the TeV emission region that is ~106 times as
S. Hoppe,1 D. Horns,17 A. Jacholkowska,14 O. C. de Jager,8 E. Kendziorra,17 M. Kerschhaggl,5 strong is deduced from the observed short-term
B. Khélifi,9,1 Nu. Komin,14 A. Konopelko,5† K. Kosack,1 G. Lamanna,10 I. J. Latham,7 flux variability, as shown below.
R. Le Gallou,7 A. Lemière,11 M. Lemoine-Goumard,9 J.-P. Lenain,6 T. Lohse,5 J. M. Martin,6 The differential energy spectra obtained for
O. Martineau-Huynh,18 A. Marcowith,3 C. Masterson,1,23 G. Maurin,11 T. J. L. McComb,7 the 2004 and 2005 data sets (Fig. 2) are both
E. Moulin,14 M. de Naurois,18 D. Nedbal,19 S. J. Nolan,7 A. Noutsos,7 K. J. Orford,7 well fit by a power-law function dN /dE º E −G.
J. L. Osborne,7 M. Ouchrif,18,23 M. Panter,1 G. Pelletier,16 S. Pita,11 G. Pühlhofer,13 M. Punch,11 The spectrum measured in 2005 is found to be
S. Ranchon,10 B. C. Raubenheimer,8 M. Raue,4 S. M. Rayner,7 A. Reimer,20 J. Ripken,4 L. Rob,19 hard (G ~ 2.2) and reaches beyond 10 TeV, with
L. Rolland,15 S. Rosier-Lees,10 G. Rowell,1 V. Sahakian,2 A. Santangelo,17 L. Saugé,16
S. Schlenker,5 R. Schlickeiser,20 R. Schröder,20 U. Schwanke,5 S. Schwarzburg,17
S. Schwemmer,13 A. Shalchi,20 H. Sol,6 D. Spangler,7 F. Spanier,20 R. Steenkamp,21 1
Max-Planck-Institut für Kernphysik, Post Office Box 103980,
C. Stegmann,22 G. Superina,9 P. H. Tam,13 J.-P. Tavernet,18 R. Terrier,11 M. Tluczykont,9,23 D 69029 Heidelberg, Germany. 2Yerevan Physics Institute, 2
Alikhanian Brothers Street, 375036 Yerevan, Armenia. 3Centre
C. van Eldik,1 G. Vasileiadis,14 C. Venter,8 J. P. Vialle,10 P. Vincent,18 H. J. Völk,1 d’Etude Spatiale des Rayonnements, CNRS/UPS, 9 avenue du
S. J. Wagner,13 M. Ward7 Colonel Roche, BP 4346, F-31029 Toulouse Cedex 4, France.
4
Universität Hamburg, Institut für Experimentalphysik, Luruper
Chaussee 149, D 22761 Hamburg, Germany. 5Institut für
The detection of fast variations of the tera–electron volt (TeV) (1012 eV) g-ray flux, on time Physik, Humboldt-Universität zu Berlin, Newtonstrasse 15, D
scales of days, from the nearby radio galaxy M87 is reported. These variations are about 10 times 12489 Berlin, Germany. 6Laboratoire Univers et Théories,
as fast as those observed in any other wave band and imply a very compact emission region with UMR 8102 du CNRS, Observatoire de Paris, Section de
a dimension similar to the Schwarzschild radius of the central black hole. We thus can exclude Meudon, F-92195 Meudon Cedex, France. 7University of
Durham, Department of Physics, South Road, Durham
several other sites and processes of the g-ray production. The observations confirm that TeV g rays DH1 3LE, UK. 8Unit for Space Physics, North-West Uni-
are emitted by extragalactic sources other than blazars, where jets are not relativistically beamed versity, Potchefstroom 2520, South Africa. 9Laboratoire
toward the observer. Leprince-Ringuet, IN2P3/CNRS, Ecole Polytechnique, F-91128
Palaiseau, France. 10Laboratoire d’Annecy-le-Vieux de Phy-
sique des Particules, IN2P3/CNRS, 9 Chemin de Bellevue, BP
o far, the only extragalactic objects known by the High Energy Gamma Ray Astronomy

S to emit g radiation up to energies of Tera

electron volts (1 TeV = 1012 eV) are
blazars. These are active galactic nuclei (AGN)
(HEGRA) collaboration (6). No emission above
400 GeV was observed by the Whipple col-
laboration (7) from 2000-2003.
110 F-74941 Annecy-le-Vieux Cedex, France. 11Astro Particule
et Cosmologie, UMR 7164 (Université Paris 7, CNRS, CEA,
Observatoire de Paris), 10 rue Alice Domon et Léonie Duquet,
75025 Paris Cedex 13, France. 12Dublin Institute for
Advanced Studies, 5 Merrion Square, Dublin 2, Ireland.
with a plasma jet emanating from the vicinity of The observations reported here were per- 13
Landessternwarte, Universität Heidelberg, Königstuhl, D 69117
the black hole and pointing close to the observer’s formed with the High Energy Stereoscopic Sys- Heidelberg, Germany. 14Laboratoire de Physique Théorique
line of sight. Because of the bulk relativistic tem (H.E.S.S.) located in Namibia. H.E.S.S. is et Astroparticules, IN2P3/CNRS, Université Montpellier II,
motion of the plasma in the jet, the energy and an array of four imaging atmospheric-Cherenkov CC 70, Place Eugène Bataillon, F-34095 Montpellier Cedex 5,
France. 15DAPNIA/DSM/CEA, CE Saclay, F-91191 Gif-sur-Yvette
luminosity of emitted photons are boosted by telescopes used for the measurement of cosmic Cedex, France. 16Laboratoire d’Astrophysique de Grenoble,
relativistic effects, making blazars detectable up g rays of energies between 100 GeV and several Institut National des Sciences de l’Univers/CNRS, Université
to TeV energies. 10 TeV [see (8) for more details]. The obser- Joseph Fourier, BP 53, F-38041 Grenoble Cedex 9, France.
17
The nearby radio galaxy M87 is located in vations of M87 were performed between 2003 Institut für Astronomie und Astrophysik, Universität Tübingen,
the Virgo cluster of galaxies at a distance of ~16 and 2006, yielding a total of 89 hours of data Sand 1, D 72076 Tübingen, Germany. 18Laboratoire de
Physique Nucléaire et de Hautes Energies, IN2P3/CNRS,
Mpc (z = 0.0043) and hosts a central black hole after quality selection cuts. After calibration (9), Universités Paris VI and VII, 4 Place Jussieu, F-75252 Paris
of (3.2 ± 0.9) × 109 solar masses (1). The 2-kpc the H.E.S.S. standard analysis was applied to Cedex 5, France. 19Institute of Particle and Nuclear Physics,
scale plasma jet (2) originating from the center the data using hard event selection cuts (10). Charles University, V Holesovickach 2, 180 00 Prague 8,
of M87 is resolved at different wavelengths More information about the standard analysis, Czech Republic. 20Institut für Theoretische Physik, Lehrstuhl
IV, Weltraum und Astrophysik, Ruhr-Universität Bochum, D
(radio, optical, and x-ray). The observed incli- as well as a more recent, alternative analysis 44780 Bochum, Germany. 21University of Namibia, Private
nation of the jet, at an angle of ~30° relative to the technique (11) which gives consistent results, Bag 13301, Windhoek, Namibia. 22Universität Erlangen-
observer’s line of sight (3), demonstrates that can be found in (12). Nürnberg, Physikalisches Institut, Erwin-Rommel-Str. 1, D
M87 is not a blazar and hence would represent a An excess of 243 g-ray events is measured 91058 Erlangen, Germany. 23European Associated Labora-
tory for Gamma-Ray Astronomy, jointly supported by CNRS
new class of TeV g-ray emitters. M87 has also from the direction of M87 in the whole data set, and Max-Planck-Gesellschaft.
been suggested as an accelerator of the enigmatic corresponding to a statistical significance of 13
*To whom correspondence should be addressed. E-mail:
ultra-high-energy (1020 eV) cosmic rays (4, 5). SDs, establishing M87 as a TeV g-ray source [email protected]
Previously, weak evidence for E > 730 GeV g-ray (Fig. 1). The position of the excess (right as- †Present address: Purdue University, Department of Physics,
emission from M87 in 1998 and 1999 with a cension, a; declination, d) was found to be a = 525 Northwestern Avenue, West Lafayette, IN 47907–2036,
statistical significance of 4.1 SDs was reported 12h30m 47.2s ± 1.4s, d = +12°23′51′′ ± 19′′ USA.

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an average g-ray flux of a factor of ~5 as high as angular size of ~ 3 arc min (13.7 kpc ≈ 4.3 × the luminosity of the core emission (20) and
in 2004. 1022 cm) centered on the M87 nucleus position superluminal blobs being detected downstream.
The total g-ray flux above 730 GeV (Fig. 3) already excludes the core of the Virgo cluster Modeling the high-energy radiation properties
for the individual years from 2003 to 2006 in- (15) and outer radio regions of M87 as TeV g-ray of this region (by synchrotron and inverse-
dicates variability on a yearly basis (14) corre- emitting zones. Further, the observed variability Compton scenarios), several authors favor sizes
sponding to a statistical significance of 3.2 SDs, on time scales of Dt ~ 2 days requires a very in the range of 0.1 to 1 pc (for moderate values
being derived from a c2 fit of a constant function. compact emission region because of the light- of the Doppler factor ranging between 2 and 5)
The variability is confirmed by a Kolmogorov crossing time. The characteristic size is limited to (20–22). Formally, though, there is no robust
test comparing the distribution of photon arrival R ≤ c × Dt × d ≈ 5 × 1015 d cm ≈ 5 × d Rs, where lower limit on the size of HST-1; therefore, we
times to the distribution of background arrival d is the relativistic Doppler factor (16) of the cannot exclude HST-1 as a source of TeV g rays.
times, yielding a statistical significance for burst- source of TeV radiation and Rs ≈ 1015 cm is the However, it would be hard to realize the short-
like (nonconstant) behavior of the source of 4.5 Schwarzschild radius of the M87 supermassive term variability of the TeV g-ray emission in
SDs. Unexpectedly, variability on time scales of black hole. For any reasonable value of the relation to HST-1, at least within the framework
days (flux doubling) was found in the high-state Doppler factor (i.e., 1 < d < 50, as used in the of current models. Because the size of the g-ray
data of 2005 (Fig. 3A), with a statistical sig- modeling of TeV g-ray blazars), this implies a production region does not exceed R ≤ 5 × 1015d
nificance of more than 4 SDs. This is the fastest drastic constraint on the size of the TeV g-ray cm, the location of HST-1 along the jet at 0.85
variability observed in any wave band from M87 source, which immediately excludes several arc sec from the nucleus, which corresponds to
and strongly constrains the size of the emission potential sites and hypotheses of g-ray produc- d ≈ 65 pc ≈ 2 × 1020cm, implies that the energy
region of the TeV g radiation, which is further tion. First of all this concerns the elliptical galaxy would be channeled from the central object into
discussed below. No indications for short-term M87 (15) and the g-ray production due to dark the g-ray production region within an unrealis-
variability were found in the data of 2003, 2004, matter annihilation (17). The most obvious tically small opening angle ~ R/d ≈ 1.5 × 10−3 d
and 2006, which is not unexpected given the candidate for efficient particle acceleration (18), degree.
generally lower statistical significances of the namely the entire extended kiloparsec jet, is also The only remaining and promising possi-
g-ray excesses in those years. excluded. Although compatible with the TeV bility is to conclude that the site of TeV g-ray
These observational results (location, spec- source position, even the brightest knot in the jet production is the nucleus of M87 itself (23). In
trum, and variability) challenge most scenarios (knot A) appears excluded, with its typical size contrast to the established TeV g-ray blazars, the
of very-high-energy g-ray production in extra- on the order of one arc sec (about 80 pc ≈ 2.5 × large-scale jet of M87 is seen at a relatively
galactic sources. Although the luminosity (≈ 3 × 1020 cm) resolved in the x-ray range (19). large jet angle (q ~ 30°), which suggests a quite
1040 erg/s) of TeV g rays is quite modest and An interesting possibility would be the modest Doppler boosting of its radiation.
does not cause any problems with the global peculiar knot (HST-1) in the jet of M87 (see Nevertheless, because of the proximity of M87,
energy budget of the active galaxy M87, several supporting online text and fig. S2), a region of both leptonic (24) and hadronic (5, 25) models
models can be dismissed. The upper limit on the many violent events, with x-ray flares exceeding predicted detectable TeV g-ray emission. How-

Fig. 1. Shown are the sky map as well as the position and extension limit radio data (32) measured with the Very Large Array, together with the TeV
of the TeV g-ray emission from M87. (A) Smoothed TeV g-ray excess map position with statistical and 20′′ pointing uncertainty errors (white cross)
(color coded, 0.1° integration radius) as measured by H.E.S.S. The size and again the 99.9% confidence level extension upper limit (red circle).
(68% containment radius) of the H.E.S.S. point-spread function (PSF) is The size of the emission region deduced from the short-term variability is
also indicated. The red circle indicates the intrinsic extension upper limit smaller by a factor of ~106. The black cross marks the position and
(99.9% confidence level) of 3 arc min of the TeV g-ray excess corre- statistical error of the g-ray source reported by HEGRA. The green ellipse
sponding to 13.7 kpc in M87. The contour lines show the 90-cm radio indicates the host galaxy seen in the optical wavelengths with an
emission (32). The white box marks the cutout shown in (B). (B) The 90-cm extension of 8.3 × 6.6 arc min in diameter.

www.sciencemag.org SCIENCE VOL 314 1 DECEMBER 2006 1425

REPORTS
ever, these scenarios typically produce a soft IR radiation luminosity supports the hypothesis component of an irregular magnetic field.) No
energy spectrum of TeV g rays, clearly in con- of an advection-dominated accretion disk (i.e., correlation with fluxes at other wavelengths is
trast to the hard spectrum measured by H.E.S.S. an accretion disk with low radiative efficiency) expected in this model. Although the size of
Leptonic models can be adapted in various in M87 (28) and generally excludes a strong the g-ray production region, R ~ 3 Rs ~ 3 × 1015
ways to match the new results. Within syn- contribution of external inverse-Compton emis- cm, perfectly matches the observed variability
chrotron self-Compton (SSC) scenarios (26), sion on IR light to the TeV g-ray flux. scale, and the model allows extension of the
one method is to consider the possibility of If one accepts the hypothesis that protons g-ray spectrum to 10 TeV without any signif-
differential Doppler-boosting in the jet near the can be accelerated as high as 1020 eV in jets of icant correlation at other wavelengths, the
core region, a phenomenon clearly expected radio galaxies, then (hadronic) proton synchro- main problem of the model is the suggested
in the jet formation zone, which extends over tron models (5, 25) cannot be excluded, con- magnetic field. It is orders of magnitude larger
<0.1 pc from the nucleus (27). Emitting plasma sidering the presented data. An alternative g-ray than the B field expected from the accretion
blobs of small sizes with Doppler factors production mechanism is curvature radiation of process, given the very low accretion rate as it
between 5 and 30 and magnetic fields well ultra-high-energy protons in the immediate follows from the bolometric luminosity of the
below equipartition can account for the ob- vicinity of the supermassive black hole. This core as well as the estimates of the power of
served TeV g-ray emission. An additional flux novel mechanism can simultaneously explain the jet in M87.
contribution from inverse-Compton scattering both the hard spectrum and fast variability of the The time scale of the short-term variability
of background photons, coming from scattered observed TeV g-ray emission. Rapidly rotating of the TeV g rays is on the order of the light-
disk emission or from dust, can further reduce black holes embedded in externally supported crossing time of the black hole (located at the
the range of Doppler factors toward moderate magnetic fields can generate electric fields and center of M87), which is a natural time scale of
values. accelerate protons to energies up to 1020 eV the object. Therefore, the results reported here
The TeV g-ray photons (independent of their (29–31). Assuming that acceleration of protons give clear evidence for the production of TeV
production mechanism) might be absorbed by takes place effectively within 3 Schwarzschild g rays in the immediate vicinity of the black
the pair-absorption process gTeV + gIR → e+ e− radii Rs, and if the horizon threading magnetic hole of M87.
on the local infrared (IR) radiation field in the field is not much below 104 G, one should
TeV g-ray emission region. Because no signature expect g-ray radiation due to proton curvature
References and Notes
for an absorption can be identified in the energy radiation extending to at least 10 TeV. (The 1. F. Macchetto et al., Astrophys. J. 489, 579 (1997).
spectrum up to 10 TeV, one can derive an upper electron curvature radiation is less likely be- 2. H. L. Marshall et al., Astrophys. J. 564, 683
limit on the luminosity of the infrared radia- cause of severe energy losses even in a tiny (2002).
tion field at 0.1 eV (corresponding to a wave-
length of ~10 mm, most relevant for absorption
of 10 TeV g rays) to be L(0.1 eV) ≤ 3.6 × 1038
(R/1015 cm) erg/s, where R is the size of the
TeV g-ray emission region. Such a low central

Fig. 2. The differential energy spectrum of M87

obtained from the 2004 and the 2005 data [using
standard event selection cuts (10)], covering a
range of ~400 GeV to ~10 TeV. Spectra for the
2003 and 2006 data sets could not be derived
because of limited event statistics. Flux points with
a statistical significance less than 1.5 SDs are given
as upper limits (99.9% confidence level). The
corresponding fits of a power-law function dN/dE =
I0 × (E/1 TeV)−G are indicated as lines. The photon Fig. 3. Gamma-ray flux above an energy of 730 GeV as a function of time. The given error bars
indices are G = 2.62 ± 0.35 (2004 data) and G = correspond to 1 SD statistical errors. (B) The average flux values for the years 2003 to 2006 as
2.22 ± 0.15 (2005 data). Aside from the measured with H.E.S.S., together with a fit of a constant function (red line). The flux reported by HEGRA
difference in the flux normalization by a factor is also drawn (a systematic error must be taken into account when comparing results from the two
of ~5 [I0 = (2.43 ± 0.75) × 10−13 cm−2 s−1 TeV −1 instruments). (A) The night-by-night fluxes for the four individual months (February to May) of the
in 2004 and I0 = (11.7 ± 1.6) × 10−13 cm−2 s−1 high-state measurements in 2005, with significant variability on (flux doubling) time scales of ~2 days.
TeV −1 in 2005], no variation in spectral shape is The green points in (B) correspond to the 0.2 − 6 keV x-ray flux of the knot HST-1 [solid, (20)] and the
found within errors. The systematic error on the nucleus [dashed, (33)] as measured by Chandra; the lines are linear interpolations of the flux points. No
photon index and flux normalization are estimated unambiguous correlation between the flux of x rays and TeV g rays can be identified (the x-ray/TeV data
to be DG = 0.1 and DI0 /I0 = 0.2, respectively. were not gathered simultaneously).

1426 1 DECEMBER 2006 VOL 314 SCIENCE www.sciencemag.org

REPORTS
3. G. V. Bicknell, M. C. Begelman, Astrophys. J. 467, 597 the direction of movement (relativistic beaming). The 32. F. N. Owen et al., Proceedings of The Universe at Low
(1996). boost is a function of the observation angle q relative to Radio Frequencies, ASP Conf. Ser., 199 (2000); http://xxx.
4. P. L. Biermann et al., Nucl. Phys. B Proc. Suppl. 87, 417 this direction and is described by the Doppler factor d = lanl.gov/abs/astro-ph/0006152.
(2000). [G(1 − bcosq)]−1, where G = (1 − b2)−1/2 is the Lorentz 33. Provided by D. Harris, private communication.
5. R. J. Protheroe et al., Astropart. Phys. 19, 559 factor of the emission region. 34. The support of the Namibian authorities and of the
(2003). 17. E. A. Baltz et al., Phys. Rev. D 61, 3514 (2000). University of Namibia in facilitating the construction
6. F. Aharonian et al., HEGRA collaboration, Astron. 18. L. Stawarz et al., Astrophys. J. 626, 120 (2005). and operation of H.E.S.S. is gratefully acknowledged, as
Astrophys. 403, L1 (2003). 19. E. S. Perlman, A. S. Wilson, Astrophys. J. 627, 140 is the support by the German Ministry for Education and
7. S. Le Bohec et al., Astrophys. J. 610, 156 (2004). (2005). Research (BMBF), the Max Planck Society, the French
8. W. Hofmann, Proc. 29th Int. Cosmic Ray Conf. (Pune), 20. D. E. Harris et al., Astrophys. J. 640, 211 (2006). Ministry for Research, the CNRS-IN2P3, and the
10, 97 (2005). 21. D. E. Harris et al., Astrophys. J. 586, L41 (2003). Astroparticle Interdisciplinary Programme of the CNRS,
9. F. Aharonian et al., H.E.S.S. collaboration, Astropart. 22. L. Stawarz et al., Mon. Not. R. Astron. Soc. 370, 981 the UK Particle Physics and Astronomy Research Council
Phys. 22, 109 (2004). (2006). (PPARC), the Institute of Particle and Nuclear Physics of
10. W. Benbow, Proceedings: Towards a Network of 23. W. Forman et al., Astrophys. J. 635, 894 (2005). the Charles University, the South African Department of
Atmospheric Cherenkov Detectors VII (Palaiseau), 163 24. M. Georganopoulos et al., Astrophys. J. 634, L33 Science and Technology and National Research Foundation,
(2005). (2005). and the University of Namibia. We thank D. Harris for
11. M. de Naurois, Proceedings: Towards a Network of 25. A. Reimer et al., Astron. Astrophys. 419, 89 (2004). providing the Chandra x-ray light curve of the M87
Atmospheric Cherenkov Detectors VII (Palaiseau), 149 26. D. L. Band, J. E. Grindlay, Astrophys. J. 308, 576 nucleus.
(2005); http://arxiv.org/abs/astro-ph/0607247. (1986).
Supporting Online Material
12. Materials and methods are available as supporting online 27. W. Junor, J. A. Biretta, M. Livio, Nature 401, 891 www.sciencemag.org/cgi/content/full/1134408/DC1
material on Science Online. (1999).
Materials and Methods
13. C. Ma et al., Astronom. J. 116, 516 (1998). 28. C. S. Reynolds, T. di Matteo, A. C. Fabian, U. Hwang,
SOM Text
14. M. Beilicke et al., Proc. of TEXAS Symposium on C. R. Canizares, Mon. Not. R. Astron. Soc. 283, L111 Figs. S1 and S2
Relativistic Astrophysics (Stanford University), Paper (1996).
Table S1
#2403 (2004), see http://arxiv.org/abs/astro-ph/ 29. A. Levinson, Phys. Rev. Lett. 85, 912 (2000).
References
0504395. 30. E. Boldt, M. Loewenstein, Mon. Not. R. Astron. Soc. 316,
15. C. Pfrommer, T. A. Enßlin, Astron. Astrophys. 407, L73 29 (2000). 28 August 2006; accepted 11 October 2006
(2003). 31. F. A. Aharonian, A. A. Belyanin, E. V. Derishev, V. V. Published online 26 October 2006;
16. Emission from a region that is moving with a relativistic Kocharovsky, Vl. V. Kocharovsky, Phys. Rev. D 66, 10.1126/science.1134408
speed b = v/c (c is the speed of light) is boosted along 023005 (2002). Include this information when citing this paper.

to read out the flux states of the coupled qubits,

Solid-State Qubits with we reduced the coupling energy and hence the
splitting of the two energy levels of the excited

Current-Controlled Coupling states to zero. Indeed, as predicted, we can even

change the interaction from antiferromagnetic to
ferromagnetic. Furthermore, we showed that the
T. Hime,1 P. A. Reichardt,1 B. L. T. Plourde,1,2 T. L. Robertson,1* C.-E. Wu,1† transition probability from the symmetric ground
A. V. Ustinov,1‡ John Clarke1§ state to an antisymmetric excited state vanishes at
the anticrossing, in qualitative agreement with
The ability to switch the coupling between quantum bits (qubits) on and off is essential for calculations.
implementing many quantum-computing algorithms. We demonstrated such control with two flux Each flux qubit consists of a superconduct-
qubits coupled together through their mutual inductances and through the dc superconducting ing loop interrupted by three Josephson tunnel
quantum interference device (SQUID) that reads out their magnetic flux states. A bias current junctions (2). When the applied magnetic flux
applied to the SQUID in the zero-voltage state induced a change in the dynamic inductance, Fq is at the degeneracy point (n + ½)F0 (where n
reducing the coupling energy controllably to zero and reversing its sign. is an integer such that |Fq – nF0| ≤ F0/2, F0 ≡
h/2e is the flux quantum, h is the Planck constant,
he past few years have seen major ad- prepared quantum states remain coherent for and e is the electron charge), a screening current

T vances in the field of superconducting

quantum bits (qubits). This family in-
cludes those based on electrical charge (1),
times up to several microseconds (8). Coupling
two or more qubits together results in entangled
states (9–15) with energy spectra that exhibit the
Iq can flow around the loop in either direction,
represented by the states |↑> and |↓>. The ground
and first excited states of the qubit correspond to
magnetic flux (2–4), charge and phase (5), and avoided crossings (anticrossings) predicted by symmetric and antisymmetric superpositions of
the phase difference across a Josephson junction quantum mechanics (16). In addition to studying the two current states and are separated by an
(6). Arbitrary superpositions of the single-qubit quantum coherence in many-body systems, energy D. When Fq ≠ (n + ½)F0, the energy
states can be prepared and manipulated by mi- there is considerable interest in arrays of qubits difference increases to n = (D2 + e2)½, where e =
crowaves to produce Rabi oscillations, Ramsey for quantum computing. Because quantum com- 2Iq[Fq – (n + ½)F0]. The state of the qubit is
fringes, and echoes long-familiar in atomic putation requires both the manipulation of single measured by coupling the flux generated by Iq
physics and nuclear magnetic resonance (7). The qubits and the entanglement of many qubits, the to a dc SQUID. Two flux qubits are coupled
ability to switch the coupling (17–21) between through their mutual inductances to each other
1
Department of Physics, University of California, Berkeley, qubits on and off in a scalable architecture would and to the SQUID. The interaction of two pairs of
CA 94720–7300, USA. 2Department of Physics, Syracuse
University, Syracuse, NY 13244–1130, USA.
enable many quantum-computing algorithms. states produces four new states: a ground state
We conducted experiments on two flux |0> and three excited states |1>, |2>, and |3>.
*Present address: Proteus Biomedical, 750 Chesapeake
Drive, Redwood City, CA 94063, USA. qubits biased at the same frequency. In this re- Each of these states consists of a linear superpo-
†Present address: Department of Physics, National Tsing- gime, the antiferromagnetic interaction between sition of four basis states (22): the symmetric
hua University, Hsinchu 300, Taiwan. the qubits produces an anticrossing and thus a triplet |↑↑>, |S> = (|↑↓> + |↓↑>)/2½, and |↓↓> and
‡Permanent address: Physikalisches Institut III, Universität splitting in the energy spectrum of the first and the antisymmetric singlet |A> = (|↑↓> – |↓↑>)/2½.
Erlangen-Nürnberg, Erwin-Rommel-Strasse 1, D-91058
Erlangen, Germany.
second excited states. By varying the bias cur- The two qubits A and B and their readout dc
§
To whom correspondence should be addressed. E-mail: rent in the zero-voltage state of the superconduct- SQUID are shown schematically in Fig. 1A. The
[email protected] ing quantum interference device (SQUID) used qubits have loop inductances LqA and LqB and

www.sciencemag.org SCIENCE VOL 314 1 DECEMBER 2006 1427

REPORTS
are coupled through a mutual inductance Mqq. qubit A can be chosen to be positive, negative, or SQUID so that we could distinguish their flux
The surrounding dc SQUID consists of a loop zero. The coupling energy K takes the form (19) signals. We measured the SQUID critical
with inductance LS and two Josephson junc- current by applying current pulses (Fig. 1C).
tions, each with critical current I0. The SQUID is K = K0 + KS = 2IqAIqB For each measurement, using 105 current pulses,
coupled to qubits A and B through mutual in- (1) we adjusted the height of the first plateau to
× (– Mqq – MqAS MqBS/L)
ductances MqAS and MqBS. We can pass a bias obtain a 50% probability of switching out of
current Ib through the SQUID and bias the qubits the zero-voltage state. We applied a pulse of
with independent applied fluxes FA and FB; where IqA and IqB are the qubit screening microwave flux to the qubits before each current
these determine the applied SQUID flux FS. By currents. pulse to drive transitions between quantum
varying the bias current through the SQUID in Figure 1B shows our experimental realiza- states of the individual or coupled qubits, pro-
the zero-voltage state, we showed theoretically tion of the two qubits and their common ducing peaks and dips in the SQUID switching
that one can control the coupling energy K be- SQUID. Our qubits have much larger areas than probability; we plotted the microwave frequen-
tween the two qubits (19). The energy K = K0 + the three-junction qubits that have been de- cy versus the applied flux to obtain energy
KS has two contributions: a fixed energy K0 scribed by other groups (3, 12), and consequent- spectra.
through the mutual inductance of the two qubits, ly we must take into account their geometrical In Fig. 2A, we show the joint frequency
and a controllable energy KS through their mutual inductances in simulating their characteristics spectrum of the qubits. The two flux lines
inductances to the SQUID. In the zero-voltage (23). These large areas, together with the on- enable us to keep the total flux applied to the
state of a SQUID with appropriate parameters, the chip flux lines, enable us to apply independent SQUID nearly constant by applying fluxes of
inverse dynamic inductance L–1 = Re(∂J/∂FS)Ib is flux biases using modest currents (~0.3 mA/F0). opposite sign to the qubits (24). Each spectrum
nonlinear and can be positive, negative, or zero, We deliberately gave the two qubits slightly arises from transitions from the ground state
depending on the values of ΦS and Ib; Re indi- different areas and mutual inductances to the to the first excited state. Except near their
cates the real part, and J is the current circulating apparent intersection, the spectra are excellent
in the SQUID loop. As a result, the sign of the fits (dashed lines) to the prediction n = (D2 +
flux change coupled to (for example) qubit B e2)½, yielding DΑ/h = 8.872 ± 0.005 GHz and
through the SQUID by a given flux change in DΒ/h = 8.990 ± 0.004 GHz (where errors are
SD). An expanded view of the spectra near
their intersection at 11.25 GHz (Fig. 2B)
reveals an avoided crossing. The lower and
upper spectra correspond to transitions from the
ground state |0> to the first excited state |1> and
the second excited state |2>, respectively. We
fitted a hyperbolic curve to each data set to find a
splitting of 122.6 ± 0.8 MHz.
The peaks in the lower spectrum of Fig. 2B
vanish near the anticrossing, implying that the
matrix elements vanish for transitions from |0>
to |1>. The origin of this effect lies in the sym-
metry of the eigenstates (fig. S1). For K < 0, the
contribution of the antisymmetric singlet state
at the anticrossing vanishes for |0>, |2>, and
|3>, leaving only contributions from the sym-
metric triplet states, whereas the converse is
true for the state |1>. Consequently, transitions
from the symmetric ground state |0> to the
antisymmetric excited state |1> are forbidden.

Fig. 2. Frequency versus flux for qubits. (A)

Fig. 1. Coupled flux qubits. (A) Two qubits, A and Spectra of qubits A and B with their fluxes ad-
B, surrounded by the dc SQUID used to measure justed independently to separate their degeneracy
their magnetic flux states and control their in- points while keeping the flux applied to the SQUID
ductive coupling. (B) The SQUID and the two qubits nearly constant. Data were acquired in a 400-MHz
are fabricated on a Si chip from Al thin films in the bandwidth around the calculated peak centers.
same process, using two-angle evaporation; an On this scale, the spectra appear to intersect at
intervening oxidation process forms the Josephson 11.25 GHz. Color bar indicates peak heights. (B)
junctions. The SQUID junctions are 215 × 250 nm2 Spectrum shown in (A) expanded to reveal the
and the qubit junctions are 180 × 205 nm2 (two anticrossing of the spectra of |1> and |2> of the Fig. 3. Measured peak heights and calculated
larger junctions) and 150 × 170 nm2 (smaller coupled qubits; dots indicate the positions of transition probabilities for transitions from the
junction). Film widths are 1 mm. Flux lines 1 and 2, maximum peak heights. Lower and upper spectra initial state |0> to the final states |1> and |2>.
connected (separately) in series, apply independent correspond to transitions from the ground state Flux dependence of measured peak heights taken
magnetic fluxes to the qubits and SQUID. The chip |0> to the excited states |1> and |2>, respec- from the spectra in Fig. 2B and of calculated
is enclosed in a superconducting box, and cooled to tively. Frequency splitting at the anticrossing is square of matrix elements |T10|2 and |T20|2. |T20|2
50 mK in a dilution refrigerator. (C) Current pulse Ib 122.6 ± 0.8 MHz. Note the absence of data for |1> is fitted to the peaks at the maximum peak height
used to determine the critical current. near the anticrossing. [measured in arbitrary (arb.) units].

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This behavior is illustrated in Fig. 3, where we [supporting online material (SOM) text]. We and would provide all the necessary ingredients
plot the measured peak heights taken from Fig. show our central result in Fig. 4D, where we to implement scalable universal quantum logic.
2B. For the transitions from |0> to |1>, the plot the splitting versus Ipb for two different Independent flux lines for the qubits are key to
amplitude of the peaks becomes vanishingly intersection frequencies. For both data sets, the this scalable architecture; it is worth emphasizing,
small at the anticrossing, whereas the peaks are splitting decreases smoothly as Ipb is increased. however, that these fluxes remain constant, and
enhanced for the transitions from |0> to |2>. In the case of the data obtained at 10.75 GHz, one needs only to switch a small current (~ 1 mA)
Because the peak heights represent the proba- the splitting goes almost to zero as Ipb is in the SQUID to turn the interaction on and off.
bility of a transition for each measurement, we increased, and then increases. We believe that Note added in proof: S. H. W. van der
expected them to scale as the square of the this result implies that the coupling was re- Ploeg et al. (preprint available at http://arxiv.
matrix element Tf 0 = <f |sz(A) + sz(B)|0>, where duced to zero and subsequently changed sign org/abs/cond-mat/0605588) reported two flux
f = 1,2 is the final state and sz(A) and sz(B) are as Ipb was increased. Higher values of Ipb qubits in which the coupling was controlled by
the Pauli spin operators, characterizing the cou- caused the SQUID to switch prematurely. The means of a coupler loop and demonstrated that
pling of the microwave excitation to the qubits. two solid curves are the results of our simulations the sign of the ground state could be changed
Figure 3 also shows the dependence of | T10|2 that used only the measured and calculated from antiferromagnetic to ferromagnetic. Spec-
and |T20|2 on flux. There is a clear qualitative parameters listed in the caption to Fig. 4D. The troscopy of excited states was not described.
agreement between the peak heights and the calculated curves overestimate the splitting at zero
transition probabilities. bias current by about 28% and at the prebias References and Notes
In Fig. 4, A to C, at the slightly lower current by about 15%. Given the many parameters 1. Y. Nakamura, Y. A. Pashkin, J. S. Tsai, Nature 398, 786
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2. J. E. Mooij et al., Science 285, 1036 (1999).
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current to the SQUID. The bias current was is remarkably good. The dashed curves show fits 4. J. R. Friedman, V. Patel, W. Chen, S. K. Tolpygo,
switched on before the microwave pulse was to the data using common values of SQUID J. E. Lukens, Nature 406, 43 (2000).
applied (Fig. 4D, inset); this prebias current Ipb critical current and prebias current. The fits are 5. D. Vion et al., Science 296, 886 (2002).
6. J. M. Martinis, S. Nam, J. Aumentado, C. Urbina, Phys.
was low enough to ensure that the probability excellent. Rev. Lett. 89, 117901 (2002).
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state would be negligible. Within 10 ns of the two qubits and to switch their coupling on and off (Clarendon Press, Oxford, 1961).
microwaves being switched off, we increased with a single SQUID solely by means of its bias 8. P. Bertet et al., Phys. Rev. Lett. 95, 257002 (2005).
9. A. J. Berkley et al., Science 300, 1548 (2003).
the bias current to provide the readout pulse. current represents an efficient architecture for a 10. T. Yamamoto, Yu. A. Pashkin, O. Astafiev, Y. Nakamura,
We fitted hyperbolas to the data and corrected quantum computer. In particular, we have shown J. S. Tsai, Nature 425, 941 (2003).
for the flux shift generated by the bias current previously (19) that a quantum controlled-NOT 11. Yu. A. Pashkin et al., Nature 421, 823 (2003).
in the SQUID during the measurement process logic gate can be implemented with this principle 12. A. Izmalkov et al., Phys. Rev. Lett. 93, 037003
(2004).
13. R. McDermott et al., Science 307, 1299 (2005).
14. J. B. Majer, F. G. Paauw, A. C. J. ter Haar, C. J. P. M.
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16. J. Von Neumann, E. Wigner, Z. Phys. 30, 467
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18. D. V. Averin, C. Bruder, Phys. Rev. Lett. 91, 057003
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19. B. L. T. Plourde et al., Phys. Rev. B 70, 140501(R)
(2004).
20. P. Bertet, C. J. P. M. Harmans, J. E. Mooij, Phys. Rev. B
Fig. 4. Control of the frequency 73, 064512 (2006).
splitting of |1> and |2>. Spectra 21. A. O. Niskanen, Y. Nakamura, J. S. Tsai, Phys. Rev. B 73,
obtained for an intersection fre- 094506 (2006).
quency of 10.75 GHz for three 22. M. J. Storcz, F. K. Wilhelm, Phys. Rev. A. 67, 042319
values of SQUID prebias current (2003).
23. T. L. Robertson et al., Phys. Rev. B 73, 174526
Ipb applied during the microwave (2006).
excitation. Splitting is (A) 135 ± 24. B. L. T. Plourde et al., Phys. Rev. B 72, 060506(R)
2 MHz, (B) 103 ± 2 MHz, and (C) (2005).
56 ± 5 MHz. (D) Splitting versus 25. M. Kamon, M. J. Tsuk, J. K. White, IEEE Trans. Microw.
Ipb for two intersection frequen- Theory Tech. 42, 1750 (1994).
26. We thank F. Wilhelm for helpful discussions and I. Siddiqi
cies. Solid curves are predictions for thoughtful comments on the manuscript. This work
using the calculated (25) param- was supported by the NSF under grant EIA-020-5641, Air
eters Mqq = 0.75 picohenry (pH), Force Office of Scientific Research under grant F49-620-
MqAS = 87.2 pH, MqBS = 63.8 pH, 02-1-0295, Army Research Office under grant DAAD-19-
LS = 423 pH; measured parame- 02-1-0187, Advanced Research and Development
ters IqA = ½deA/dFqA = 146.0 ± Activity, and Bavaria California Technology Center.
0.2 nA, IqB = ½ deB /dFqB =
147.8 ± 0.2 nA, FS(11.25 GHz) = 0.27 F0, FS(10.75 GHz) = 0.28 F0; and the estimated maximum Supporting Online Material
SQUID critical current 2I0 = pDs/2eRNN = 1.21 ± 0.054 mA, where Ds = 175 ± 5 meV is the energy gap of www.sciencemag.org/cgi/content/full/314/5804/1427/DC1
SOM Text
Al, and RNN = 228 ± 10 ohms is the resistance of the SQUID at voltages much greater than D s /e. Fig. S1
Uncertainties in the low-temperature impedances prevent precise determination of the currents, and we
fitted the data using 2I0 = 0.844 mA and scaling the bias current by a factor of 0.767. Inset shows pulse 28 August 2006; accepted 18 October 2006
sequence. 10.1126/science.1134388

www.sciencemag.org SCIENCE VOL 314 1 DECEMBER 2006 1429

REPORTS
optical trap (13), such that external motions do not
Optical Atomic Coherence at the decohere the superposition of the two states. Using
optically cooled 87Sr atoms in a zero-differential–
1-Second Time Scale Stark shift one-dimensional (1D) optical lattice and
a cavity-stabilized probe laser with a sub-hertz
spectral width, we have achieved probe-time–
Martin M. Boyd, Tanya Zelevinsky, Andrew D. Ludlow, Seth M. Foreman,
limited resonance linewidths of 1.8 Hz at the
Sebastian Blatt, Tetsuya Ido,* Jun Ye†
optical carrier frequency of 4.3 × 1014 Hz. The
Highest-resolution laser spectroscopy has generally been limited to single trapped ion systems ratio of these frequencies, corresponding to a Q ≈
because of the rapid decoherence that plagues neutral atom ensembles. Precision spectroscopy of 2.4 × 1014, is the highest obtained for any co-
ultracold neutral atoms confined in a trapping potential now shows superior optical coherence herent spectral feature.
without any deleterious effects from motional degrees of freedom, revealing optical resonance This ultrahigh spectral resolution allows us
linewidths at the hertz level with a good signal-to-noise ratio. The resonance quality factor of to perform experiments in the optical domain
2.4 × 1014 is the highest ever recovered in any form of coherent spectroscopy. The spectral resolution analogous to radio-frequency nuclear magnetic
permits direct observation of the breaking of nuclear spin degeneracy for the 1S0 and 3P0 optical clock resonance (NMR) studies. Under a small mag-
states of 87Sr under a small magnetic bias field. This optical approach for excitation of nuclear spin netic bias field, we make direct observations of
states allows an accurate measurement of the differential Landé g factor between 1S0 and 3P0. The the magnetic sublevels associated with the
optical atomic coherence demonstrated for collective excitation of a large number of atoms will have a nuclear spin. Furthermore, we have precisely
strong impact on quantum measurement and precision frequency metrology. determined the differential Landé g factor be-
tween 1S0 and 3P0 that arises from hyperfine
mixing of 3P0 with 3P1 and 1P1. This optical
he relative rates of coherent interaction atomic clocks (14) benefit directly from the measurement approach uses only a small mag-

T and decoherence in a quantum system are

of fundamental importance for both quan-
tum information science (1) and precision
enhanced signal size and the high resonance
quality factor Q. Tests of atomic theory can be
performed with increased precision. The availa-
netic bias field, whereas traditional NMR ex-
periments performed on a single state (either
1
S0 or 3P0) would need large magnetic fields to
metrology (2). Enhancing their ratio, which is ble spectral resolution also enables a direct induce splitting in the radio frequency range.
equivalent to improving spectral resolving optical manipulation of nuclear spins that are Because the state mixing between 3P0, 3P1,
power, characterizes much of the recent progress decoupled from the electronic angular momen- and 1P1 arises from both hyperfine interactions
in these fields. Trapped ions have so far provided tum. Nuclear spins can have an exceedingly long and external fields, the use of a small field
the best platform for research in this direction, relaxation time, making them a valuable alter- permits an accurate, unperturbed measurement
resulting in a number of seminal achievements native for quantum information processing and of mixing effects. Optical manipulation of
(3–8). The principal advantage of the ion system storage. Two ground-state nuclear spins can, for nuclear spins shielded by two spin-paired
lies in the clean separation between the internal example, be entangled through dipolar interac- valence electrons, performed with a superior
atomic state and the external center-of-mass tions when photoassociation channels to high- spatial and atomic state selectivity, may pro-
motion, leading to long coherence times asso- lying electronic states (such as 3P1) are excited vide an attractive choice for quantum infor-
ciated with both internal and external degrees of (15). Combined with a quantum degenerate gas, mation science.
freedom. A large ensemble of neutral atoms the enhanced precision in measurement will Optical atomic clocks based on neutral atoms
offers obvious benefits in the signal size and further strengthen the prospects of using optical benefit directly from a large signal-to-noise ratio
scalability of a quantum system (9, 10). Multi- lattices to engineer condensed matter systems (S/N) and a superior line Q. Resolving nuclear
atom collective effects can also dramatically (for example, allowing massively parallel quan- sublevels with optical spectroscopy permits
enhance the coherent matter/field interaction tum measurements). improved measurements of systematic errors
strength (11). However, systems based on Much of the recent interest in alkaline earth associated with the nuclear spin, such as linear
neutral atoms normally suffer from decoher- atoms (and similar atoms and ions, such as Yb, Zeeman shifts, and tensor polarizability that
ence resulting from coupling between their Hg, In+, and Al+) arises from the study of the manifests itself as nuclear spin–dependent trap
internal and external degrees of freedom (12). forbidden optical transitions, both for metro- polarization sensitivity. Tensor polarizability of
In this article, we report a record-level spectral logical applications and as a means for quantum the 3P0 state is one of the important potential
resolution in the optical domain based on a control, with an important achievement being systematic uncertainties for fermion-based clocks
doubly forbidden transition in neutral atomic highly effective narrow line laser cooling (16–18). and is one of the primary motivations for recent
strontium. The atoms are confined in an optical The spin-forbidden 1S0-3P1 transition has been proposals involving electromagnetically induced
trapping potential engineered for accurate extensively studied as a potential optical frequen- transparency resonances or dc magnetic field–
separation between these degrees of freedom cy standard in Mg (19), Ca (20), and Sr (21, 22) induced state mixing in bosonic isotopes (27–29).
(13). The large number of quantum absorbers and has recently been explored as a tool for high- The work reported here has permitted control of
provides a dramatic enhancement in signal size resolution molecular spectroscopy through pho- these systematic effects to ~5 × 10−16 (30). Given
for the recovered hertz-linewidth optical reso- toassociation in ultracold Sr (15). The doubly the superior S/N from the large number of
nance profile. forbidden 1S0-3P0 transition is weakly allowed quantum absorbers, we expect this system to be
The demonstrated neutral-atom coherence as a result of hyperfine-induced state mixing, competitive among the best performing clocks in
properties will affect a number of research fields, yielding a linewidth of ~1 mHz for 87Sr with a terms of stability. Accuracy is already approach-
with some initial results reported here. Optical nuclear spin of 9=2 . This transition is a particularly ing the level of the best atomic fountain clocks
attractive candidate for optical domain exper- (31, 32), and absolute frequency measurement is
JILA, National Institute of Standards and Technology and iments, where long coherence times are desirable, limited by the Cs clock–calibrated maser signal
University of Colorado, and Department of Physics, and is currently being aggressively pursued for the available to us by means of a fiber link (33). An
University of Colorado, Boulder, CO 80309–0440, USA.
realization of an optical atomic clock (23–26). all-optical clock comparison is necessary to re-
*Present address: National Institute of Information and
Communications Technology, Koganei, Tokyo, Japan.
Furthermore, because of the lack of electronic veal its greater potential.
†To whom correspondence should be addressed. E-mail: angular momentum, the level shifts of the two To fully exploit the ultranarrow hyperfine-
[email protected] states can be matched with high accuracy in an induced transition for high-precision spectrosco-

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REPORTS
py, it is critical to minimize decoherence from spread among ~100 lattice sites. The vacuum- nearly equal amplitudes, corresponds to the trap
both fundamental and technical origins. The limited lattice lifetime is >1 s. The atoms are oscillation frequency in the transverse plane.
~100-s coherence time available from the 87Sr confined in the Lamb-Dicke regime along the With atoms confined in the lattice, the lin-
atoms is not yet experimentally practical as a axis of the optical lattice. The Lamb-Dicke early polarized (parallel to the lattice polarization)
result of environmental perturbations to the probe parameter, or the square root of the ratio of 698-nm laser drives the p transitions (Fig. 1B) for
laser phase at long time scales, but atomic co- recoil frequency to trap oscillation frequency, is probe times between 0.08 and 1 s, depending on
herence in the optical domain at 1 s can already ~0.3. Both the axial and radial trap frequencies the desired spectral resolution limited by the
greatly improve the current optical clock and are much larger than the 1S0-3P0 transition Fourier transform of the probe time. The effect of
quantum measurements. To achieve long atomic linewidth, leading to the spectral feature com- the probe laser is detected in two ways. First, after
coherence times, we trap atoms in an optical posed of a sharp optical carrier and two sets of some atoms are excited to the long-lived 3P0 state
lattice with a zero net ac Stark shift between the resolved motional sidebands. One pair of side- by the probe laser, the remaining 1S0 population
two clock states, enabling a large number of bands is observed ±40 kHz away from the is measured by exciting the strong 1S0-1P1
neutral atoms to be interrogated free of pertur- carrier, corresponding to the axial oscillation transition with a resonant pulse at 461 nm. The
1
bations. The tight atomic confinement enables frequency in the lattice. The red-detuned side- S0-1P1 pulse scatters a large number of signal
long probing times and permits spectroscopy band is strongly suppressed, indicating that photons and heats the 1S0 atoms out of the lattice,
free of broadening by atomic motion and photon nearly all atoms are in the motional ground state leaving only the 3P0 atoms. Once the 1S0 atoms
recoil. along the lattice axis. The second pair of have been removed, the 3P0 population is
For the highest spectral resolution, it is nec- sidebands at ±125 Hz from the carrier, with determined by driving the 3P0-3S1 and 3P2-3S1
essary for the probe laser to have a narrow
intrinsic linewidth and a stable center frequency.
A cavity-stabilized 698-nm diode laser is used as
the optical local oscillator for 1S0-3P0 spectros-
copy (34). The linewidth of this oscillator has
been characterized by comparison with a second
laser operating at 1064 nm via an optical
frequency comb linking the two distant colors.
A heterodyne optical beat signal between the two
lasers, measured by the frequency comb, reveals
a laser linewidth of <0.3 Hz (resolution-
bandwidth–limited) at 1064 nm for a 3-s
integration time. This result demonstrates the
ability of the frequency comb to transfer optical Fig. 1. (A) Partial 87Sr energy-level diagram. Solid arrows show relevant electric dipole transitions
phase coherence (~1 rad/s) across hundreds of with wavelengths in nanometers. Dashed arrows show the hyperfine interaction–induced state mixing
terahertz. Our frequency comb is also refer- between 3P0 and 3P1 and between 3P0 and 1P1, which provides the nonzero electric dipole moment for
enced to a hydrogen maser calibrated by the the doubly forbidden 698-nm transition. (B) The mixing alters the Landé g factor of the 3P0 state such
National Institute of Standards and Technology that it is ~50% larger than that of 1S0, resulting in a linear Zeeman shift for the 1S0-3P0 transition in
(NIST) F1 Cs fountain clock (31), which allows the presence of a small magnetic field. The large nuclear spin of 87Sr (I = 9=2 ) results in 10 sublevels for
the 1S0 and 3P0 states, providing 28 possible transitions from the ground state.
us to accurately measure the probe laser fre-
quency to 3 × 10−13 at 1 s. Additionally, the
698-nm laser has been compared with an inde- Fig. 2. Spectroscopy of the
pendent laser system operating at the same 1
S0-3P0 transition in 87Sr. A
wavelength, revealing a resolution-bandwidth– pair of Helmholtz coils pro-
limited laser linewidth of 0.2 Hz, which in- vides a variable field along
creases to ~2 Hz for a 30-s integration time. the lattice (and probe) po-
After removing the linear drift, the stability of larization axis, allowing a
this local oscillator is ~1 × 10−15 from 1 to 1000 s, measurement of the field-
which is limited by the thermal noise of the dependent transition line-
cavity mirrors. Thus, the probe laser provides the width as shown in (A) where
optical coherence needed to perform experiments an 80-ms interrogation pulse
at the 1-s time scale. is used, limiting the width
87
Sr atoms are captured from an atomic beam to ~10 Hz. Error bars in (A)
and cooled to 1 mK by means of a magneto- indicate measurement un-
optical trap (MOT) acting on the strong 1S0-1P1 certainty in linewidth. (B)
transition (Fig. 1A). This step is followed by a A representative spectrum
when the ambient field is well
second-stage MOT with the use of the narrow
1 controlled. Here, a longer
S0-3P1 intercombination line that cools the
probe time is used (~480 ms
atoms to ~1.5 mK. During narrow line cooling, or a 1.8-Hz Fourier limit)
a nearly vertical 1D lattice is overlapped with but the linewidth is limited
the atom cloud for simultaneous cooling and to 4.5 Hz by residual mag-
trapping. The lattice is generated by a ~300-mW netic fields and possibly
standing wave with a 60-mm beam waist at the residual Stark shifts. (C) A field of 0.77 G is applied along the polarization axis, and the individual
wavelength of 813.428(1) nm, where the 1S0 Fourier-limited (10 Hz) p transitions are easily resolved. Data are shown in red, and a fit of 10 evenly
and 3P0 ac Stark shifts from the trapping field are spaced transitions is shown in blue. The calculated transition probabilities based on Clebsch-Gordan (C-G)
equal (35). The cooling and loading stages take coefficients are included in the inset. In (B) and (C), the population is scaled by the total number of atoms
~0.7 s and result in a sample of 104 atoms, available for spectroscopy (~104).

www.sciencemag.org SCIENCE VOL 314 1 DECEMBER 2006 1431

REPORTS
transitions (Fig. 1A), resulting in atomic decay to ultrahigh spectral resolution has allowed a recent probed the spectra of a single resolved sublevel
the ground state via 3P1 for a second measure- measurement of systematic effects for the optical (mF = 5=2 in this case) using p polarization with
ment with the use of the 1S0-1P1 pulse. The sec- clock transition at the 9 × 10−16 level (30). the time window extended to 480 ms. Figure 3,
ond measurement provides superior S/N because The high-resolution spectroscopy enables A and B, shows some sample spectra of the
only atoms initially excited by the 698-nm probe direct measurement of the differential Landé g isolated 1S0 (mF = 5=2 )–3P0 (mF = 5=2 ) transition
laser contribute to the fluorescence signal, and factor (Dg) between 3P0 and 1S0. To observe this with a Fourier-limited linewidth of 1.8 Hz,
the zero background is not affected by shot-to- state mixing effect, we applied a small magnetic representing a line Q of ~ 2.4 × 1014. This Q is
shot atom number fluctuations. Combining both field (<1 G) along the direction of the lattice reproduced reliably, as evidenced by the histo-
approaches permits signal normalization against polarization, and the probe laser polarization was gram of linewidths measured in the course of
atom number fluctuations. again fixed along this quantization axis to drive p 1 hour (Fig. 3C). Typical linewidths are ~1 to 3 Hz,
Although the 3P0 and 1S0 states are magnet- transitions. Figure 2C shows a direct observation with the statistical scatter owing to residual probe
ically insensitive to first order, the hyperfine- of the hyperfine-induced state mixing in the form laser noise at the 10-s time scale.
induced state mixing, which allows the otherwise of 10 resolved transition components, with their To further explore the limit of coherent atom-
forbidden transition, modifies the 3P0 nuclear g relative amplitudes influenced by the Clebsch- light interactions, we have also performed two-
factor by ~50%. This effect results in a linear Gordan coefficients. The narrow linewidth of pulse optical Ramsey experiments on an isolated
Zeeman shift in the 1S0-3P0 transition of about the forbidden transition allows this nuclear- p transition. When a system is lifetime-limited, the
–100 Hz/G per magnetic sublevel mF (36, 37), magnetic-resonance–like g-factor experiment Ramsey technique can achieve higher spectral
where we use the convention that the g factor and to be performed optically at small magnetic resolution at the expense of S/N, leading to use-
nuclear magnetic moment carry the same sign fields. The magnitude of Dg can be measured ful information on the decoherence process.
and 1 G = 10−4 T. This effect is shown sche- by mapping out the line splitting versus magnetic By performing the experiment in the lattice, the
matically in Fig. 1B, where the 10 nuclear spin field. Alternatively, 18 s+ and s– transitions (Fig. Ramsey interrogation pulse can be prolonged,
sublevels are resolved for the 1S0 and 3P0 states in 1B) can be used to extract both the magnitude resulting in a markedly reduced Rabi pedestal
the presence of a magnetic field. The linear and sign [relative to the known 1S0 g factor (38)] width as compared to free-space spectroscopy.
Zeeman shift is an important issue for high- of Dg, without accurate calibration of the field. The reduced number of fringes greatly simplifies
resolution spectroscopy, because the magnetic Using the latter approach, we find Dg = –108.8(4) identification of the central fringe for applications
sensitivity can cause undesirable broadening of Hz/G per mF. The measured Dg permits de- such as frequency metrology. The fringe period
the transition, as well as line center shifts due to termination of the 3P0 lifetime of 140(40) s, in (in Hz) is determined by the sum of the pulse
unbalanced population distribution among the agreement with recent ab initio calculations interrogation time tR and the free-evolution time
sublevels. To achieve the narrowest resonance, (39, 40). The uncertainty is largely dominated by between pulses TR and is given by 1=ðtR þ TR Þ.
the ambient magnetic field must be compensated inconsistencies among hyperfine mixing models Figure 3D shows a sample Ramsey spectrum,
with three orthogonal sets of Helmholtz coils. An (36, 37). where tR = 20 ms and TR = 25 ms, yielding a
example of this zeroing process is shown in Fig. The linewidth of each spectral feature in fringe pattern with a period of 20.8(3) Hz and
2A, where the transition linewidths are measured Fig. 2C is Fourier-limited by the 80-ms probe fringe FWHM 10.4(2) Hz. For the same tran-
under various field strengths. After zeroing the time to ~10 Hz. With the nuclear spin degen- sition with tR raised to 80 ms and TR to 200 ms
field, narrow resonances as in Fig. 2B are eracy removed by a small magnetic field, in- (Fig. 3D, inset), the width of the Rabi pedestal is
routinely obtained. The displayed transition line- dividual transition components allow exploration reduced to ~10 Hz, and the recorded fringe
width of 4.5 Hz [full width at half maximum of the ultimate limit of our spectral resolution linewidth is 1.7(1) Hz.
(FWHM)] represents a resonance Q of ~1014. by eliminating any broadening mechanisms This linewidth is recovered without substan-
The good S/N for the narrow line resonance due to residual magnetic fields or light shifts, tial degradation of the fringe signal size, sug-
achieved without any averaging or normalization the likely limitation for data such as in Fig. 2B. gesting that the spectral resolution is limited by
arises from the contribution of 104 atoms. The To reduce the Fourier limit for the linewidth, we phase decoherence between light and atoms and
not by effects such as trap lifetime. A limit of
1 to 2 Hz is consistent with our measurements of
Fig. 3. Spectroscopy of the probe laser noise integrated over the time
the isolated 1S0 (mF = scales used for spectroscopy. Other potential
5=
2)– P0 (mF = =2) tran-
3 5
limitations to the spectral width include Doppler
sition. Resolving indi- broadening resulting from the relative motion
vidual sublevels allows between the lattice and probe beams and
spectroscopy without mag- broadening as a result of tunneling in the lattice.
netic or Stark broaden- Future measurements will be improved by
ing. Spectra in (A) and locking the probe laser to one of the resolved
(B) are taken under iden- nuclear spin transitions to further suppress
tical experimental condi- residual laser fluctuations. Although the S/N
tions by means of a pulse associated with a sublevel resonance is reduced
time of 480 ms, and
when compared to a measurement involving all
linewidths of 1.5(2) and
2.1(2) Hz are achieved. degenerate sublevels, >103 atoms still contribute
(C) A histogram of the to the signal, which allows measurement to
linewidths of 28 traces proceed without averaging. Clearly, reaching the
obtained within ~1 hour. atom shot-noise limit and performing quantum
The average linewidth is state preparations will further enhance the S/N.
near the 1.8-Hz Fourier The line Q of ~2.4 × 1014 achieved here
limit (dashed red line). provides practical improvements in the fields of
(D) Ramsey fringes with precision spectroscopy and quantum measure-
a 20.8(3)-Hz period and 10.4(2)-Hz fringe width, with data shown as open circles. Inset shows a Ramsey ment. The neutral atom–based spectroscopic
pattern with a 1.7(1)-Hz fringe FWHM. system now parallels the best ion systems in

1432 1 DECEMBER 2006 VOL 314 SCIENCE www.sciencemag.org

REPORTS
terms of fractional resolution but greatly sur- 11. A. T. Black, H. W. Chan, V. Vuletic, Phys. Rev. Lett. 91, associated with the lattice and the probe beams,
passes the latter in signal size. For optical fre- 203001 (2003). magnetic shift, density shift, and blackbody shift. For
12. J. Ye, D. W. Vernooy, H. J. Kimble, Phys. Rev. Lett. 83, further details, see M. M. Boyd et al.; preprint available
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(2005). 30. The magnetic field–induced frequency uncertainty is Program. T.Z. is a National Research Council postdoctoral
9. D. L. Haycock, P. M. Alsing, I. H. Deutsch, J. Grondalski, determined from the product of the measured residual fellow. T.I. acknowledges support from the Japan Science
P. S. Jessen, Phys. Rev. Lett. 85, 3365 (2000). magnetic field by means of the clock transition linewidth and Technology Agency.
10. I. Bloch, M. Greiner, in Advances in Atomic Molecular and experimentally determined frequency shifts versus
and Optical Physics (Academic Press, San Diego, CA, given magnetic fields along three orthogonal directions. 10 August 2006; accepted 18 October 2006
2005), vol. 52, pp. 1–47. The total systematic uncertainty includes Stark shifts 10.1126/science.1133732

(1), especially if combined with physical pro-

Macroscopic Hierarchical Surface cesses such as dewetting and contact-line
pinning. Here we report the spontaneous forma-

Patterning of Porphyrin Trimers via tion of periodic patterns of exceptionally long

(up to 1 mm) columnar stacks of porphyrin dye
molecules at a solid/liquid interface, with highly
Self-Assembly and Dewetting defined spatial and parallel ordering. These self-
assembled patterns were then used to align
Richard van Hameren,1 Peter Schön,1 Arend M. van Buul,1 Johan Hoogboom,2 liquid crystals (LCs) in domains measuring sev-
Sergiy V. Lazarenko,1 Jan W. Gerritsen,1 Hans Engelkamp,1 Peter C. M. Christianen,1 eral square millimeters.
Hans A. Heus,1 Jan C. Maan,1 Theo Rasing,1 Sylvia Speller,1 Alan E. Rowan,1 Porphyrin dye molecules can self-organize
Johannes A. A. W. Elemans,1* Roeland J. M. Nolte1 on a surface into small columnar stacks of sub-
micrometer length (13, 14). These architectures
The use of bottom-up approaches to construct patterned surfaces for technological applications is are generated as a result of combined self-
appealing, but to date is applicable to only relatively small areas (~10 square micrometers). We assembly and dewetting, which take place simul-
constructed highly periodic patterns at macroscopic length scales, in the range of square taneously when a drop-casted solution of the
millimeters, by combining self-assembly of disk-like porphyrin dyes with physical dewetting porphyrin molecules is evaporated on a surface.
phenomena. The patterns consisted of equidistant 5-nanometer-wide lines spaced 0.5 to In order to enhance the columnar stacking and
1 micrometers apart, forming single porphyrin stacks containing millions of molecules, and were hence the length of the assemblies, we have
formed spontaneously upon drop-casting a solution of the molecules onto a mica surface. On glass, synthesized compound 1 (Fig. 1) (15), which
thicker lines are formed, which can be used to align liquid crystals in large domains of square consists of three porphyrin moieties that are
millimeter size. linked via amide bonds to a central benzene
core, a motif that is known to form extended
he formation of complex submicrometer applications, such as detection arrays and optical hydrogen-bonded networks (16–19). Each por-

T patterns on surfaces that extend over

macroscopic distances underlies the fab-
rication of integrated circuits and microelectro-
elements, well-defined symmetrical patterns can
be exploited, especially if the methods decrease
the number of processing steps needed or avoid
phyrin was equipped with three aliphatic hydro-
carbon chains to increase the solubility of the
stack in organic solvents.
mechanical devices (1–3). However, for many surface-invasive steps that scratch, rub, or etch The high tendency of 1 to form aggregates
the surface. Examples of complex pattern for- can be directly observed, in that at a concentra-
1
Institute for Molecules and Materials, Radboud University mation by noninvasive techniques are still few, tion of 8 mg/ml, the chloroform solution formed
Nijmegen, Toernooiveld 1, 6525 ED, Nijmegen, Nether- usually require large polymeric molecules, and a gel. This strong aggregation is highly de-
lands. 2Department of Chemistry, Massachusetts Institute
of Technology, 77 Massachusetts Avenue, Cambridge, MA are often of small spatial extent (4–12). Self- pendent on the presence of the alkyl chains,
02139, USA. assembly of molecules on a surface can be a because porphyrin trimers without these chains
*To whom correspondence should be addressed. E-mail: simple, versatile, and less time-consuming ap- appeared not to gelate the solvent. In the proton
[email protected] proach and may lead to defect-free structures nuclear magnetic resonance (NMR) spectrum of

www.sciencemag.org SCIENCE VOL 314 1 DECEMBER 2006 1433

REPORTS
confirmed that the lines consisted of molecules
of 1, and an emission spectrum characteristic
of a porphyrin aggregate (13) was obtained
(inset, Fig. 3C).
Because mica is birefringent, we could not
study the assembly processes optically in real
time. We could, however, visualize the line-
formation process in real time on a glass surface
([1] = 4.8 × 10−5 M, 10-ml droplets), using an
optical polarization microscope equipped with a
charge-coupled device camera (movie S1). Dur-
ing the evaporation process, the front of the
droplet was pinned several times, and upon its
withdrawal, deposited material was observed
(Fig. 3D). Simultaneously with the pinning, the
formation of linear aggregates as a result of the
Fig. 1. Porphyrin trimer 1. (A) self-assembly processes was already visible
Chemical structure of 1. (B) within the droplet, perpendicular to the front,
Schematic representation of 1. before its withdrawal (fig. S2 and movie S1).
(C) Schematic representation of However, the greater roughness of the glass
a columnar stack of 1. (D) substrate as compared to mica caused the pattern
Computer-generated model of to be less well-defined.
the hydrogen-bonding network The formation of the highly ordered line pat-
in a columnar stack of 1 (two of terns is governed by a combination of molecular
the three porphyrins of each self-assembly and other physical processes (13).
molecule of 1 are omitted for
The strong self-assembly of 1, which is gov-
clarity).
erned by a balanced combination of hydrogen
bonding and p−p stacking interactions, is
essential for the growth of columnar stacks of
almost millimeter length. No members of a wide
family of porphyrin macrocycles (hexamers, do-
decamers, and porphyrin trimers with ester
instead of amide linkers) were able to form sim-
ilar periodic patterns (13). The two primary
1 in CDCl3 ([1] = 10−4 M), very broad peaks one single domain [for example, 650 ± 40 nm in physical processes that play a major role in pat-
were observed, suggesting large aggregate for- a domain with a size of 3 mm2 (Fig. 2D)], but tern formation are contact-line pinning between
mation. The addition of d6–dimethyl sulfoxide between several domains the value of the pe- the edge of a droplet and the surface (the so-
to this solution caused a sharpening of the NMR riodicity varied from 0.5 to 1 mm. The lines were called coffee-stain mechanism) and spinodal
signals, which is the result of the solvent break- oriented parallel with respect to the local solvent dewetting (9, 20–24). The latter effect is observed
ing up the hydrogen-bonding network (Fig. 1D) front, which can be deduced from the broader when the surface of a thin film on a flat substrate
and dissolving the aggregates that are present in contact pinning lines on the sample (Fig. 2E). In (such as mica) is unstable and deforms spontane-
chloroform. addition, at the boundaries of these ordered ously. Surfaces subject to this kind of dewetting
At the solid/liquid interface, the expected domains, patterns more reminiscent of normal are known to dewet via the formation of an un-
columnar stacking of 1 was confirmed by means spinodal dewetting were observed (Fig. 2F). A dulating bicontinuous pattern (24). We postulate
of scanning tunneling microscopy (STM) (fig. clear correlation is seen between these two re- that this undulating pattern governs the spatial
S1). The aggregation behavior at a surface was gions, because most columns in the periodic distribution of the linear aggregates (Fig. 3E). The
further studied with atomic force microscopy domain appear to grow out from the spinodal small defects observed in Fig. 2A support this
(AFM). We drop-casted a diluted solution of 1 in dewetting domain. postulation, because their presence does not
chloroform ([1] = 4.8 × 10−6 M, 3-ml droplets) When larger droplets ([1] = 4.8 × 10−6 M, interrupt the periodicity of the patterns.
on mica. After evaporation, very large domains 10 ml in size) were deposited under similar The initial physical phenomena, contact-line
(up to ~3 mm2) containing a highly ordered conditions, the longer evaporation time formed pinning and solvent evaporation, caused the
pattern of equidistant, nearly parallel, wire-like porphyrin lines with different dimensions and molecules dissolved in the droplet to flow
architectures were observed (Fig. 2, A and B). orientations from those described above (Fig. toward the contact line (9). In the case of the
The height of the lines was 4.5 ± 0.4 nm (Fig. 3A). A periodicity of 13.4 ± 0.7 mm and a line small droplets (3 ml), the contact line was pinned
2C), which corresponds remarkably well to the height of 55.4 ± 0.6 nm were observed. The several times, leaving behind thin layers of
calculated diameter of 1, indicating a pronounced latter value indicates that each line in this pat- deposited molecules of 1 at these positions (Fig.
shape persistence of the molecule. These obser- tern consisted of a bundle of columnar stacks 2, E and G) (9). After repeated retractions of the
vations indicate that the lines consisted of a of 1. Because of the larger dimensions, this solvent front, thin films remained between the
columnar stack one molecule thick, with each of pattern could be visualized via optical mi- pinned contact lines, which were then subject to
the lines containing millions of molecules. We croscopy (Fig. 3B), which clearly demonstrated spinodal dewetting. Combined with the propen-
will argue below that this self-organization of that the orientation of the lines, which were sity of 1 to form one-dimensional (1D) aggre-
molecules on a macroscopic scale results from a up to 0.8 mm long (fig. S3), was now orthog- gates, this dewetting gave rise to the formation of
hierarchical dewetting process. onal with respect to the solvent front. Scanning the highly defined periodic patterns, with the
Analysis of several samples revealed a nar- confocal fluorescence microscopy studies contact pinning lines directing their orientation
row spatial distribution of the periodicity within [Fig. 3C, excitation wavelength l = 411 nm] (Fig. 2G). Within each domain, the local spinodal

1434 1 DECEMBER 2006 VOL 314 SCIENCE www.sciencemag.org

REPORTS

Fig. 2. Patterns formed

on mica after the evap-
oration of 3-ml droplets
of compound 1 in chlo-
roform. (A) AFM image
(scan size = 25 × 25
mm2) of a pattern of
highly ordered equi-
distant parallel lines.
(B) AFM image (scan
size = 10 × 10 mm2),
with the inset showing
the cross section indi-
cated in the AFM image.
(C) Bar diagram show-
ing the height distribu-
tion of the line pattern
within a single domain,
with the Gaussian fit
demonstrating a line
height of 4.5 nm, with
standard deviation (s) =
0.4 nm. (D) Bar diagram
showing the spatial dis-
tribution of lines in a
single domain (size = 3
mm2). Each color repre-
sents a different posi-
tion in a single domain.
The Gaussian fit demonstrates that within this complete domain, the lines are the patterned lines. During the evaporation of the droplet (A→B), the contact
650 nm apart and s = 40 nm. (E) AFM image (scan size = 14 × 14 mm2) line is pinned several times, resulting in the formation of contact pinning lines
showing that the periodic pattern is parallel to the (bold) contact pinning line. (designated with 1, 2, and 3). After retraction of the solvent front, a thin film
(F) AFM image (scan size = 40 × 40 mm2) of a domain transition between a remains in which a pattern of thin lines is formed as a result of self-assembly
highly ordered and a less ordered domain. (G) Mechanism of the formation of and dewetting.

Fig. 3. Patterns
formed after the evapo-
ration of 10-ml droplets
of compound 1 in chlo-
roform. (A) AFM image
(scan size = 95 × 95
mm2) of a line pattern
on mica. (B) Optical
micrograph of the pat-
tern formed on mica
(scale bar, 100 mm).
(C) Scanning confocal
fluorescence microsco-
py image of the lines;
the inset shows the
characteristic fluores-
cence spectrum of a
porphyrin aggregate,
lmax 1 = 665 nm and
lmax 2 = 726 nm. (D)
Optical micrograph of
the coffee-stain–like
pattern formed during
the evaporation of a so-
lution of 1 in chloro-
form on glass ([1] =
4.8 × 10−5 M); the
bottom part is still covered with solution (dark blue). The whitish stripes are formation of the patterned lines. The presence of aggregates preformed in
the aggregates that remain after retraction of the droplet. (E) Proposed solution hinders the retraction of the solvent front from 1 to 2, causing partial
formation of periodic patterns on flat mica; spinodal dewetting causes an pinning of the contact line. In combination with the molecular self-assembly,
undulating pattern in the solvent, which governs the positioning of the this partial pinning results in an orientation and growth of linear aggregates
aggregates and thus the spatial distribution of the lines. (F) Mechanism of the orthogonal to the local solvent front; contact pinning lines are not observed.

www.sciencemag.org SCIENCE VOL 314 1 DECEMBER 2006 1435

REPORTS

Fig. 4. Application of
the patterns formed by
1 on a glass substrate
as alignment layers for
5CB; polarizing micros-
copy images of a LC cell
between crossed polar-
izers (denoted by P and
A). (A) LC ordering
parallel to the analyzer.
(B) Texture after rota-
tion of the sample over
45°. The local orienta-
tion of the 5CB mole-
cules, deduced from
the SHG rotational an-
isotropy patterns (insets), is depicted schematically in both images.

dewetting determined the periodicity, which in showed that the alignment was interrupted by 2. T. Verbiest et al., Science 282, 913 (1998).
all cases was between 500 nm and 1 mm. concentric circles, which were the contact pinning 3. V. Percec et al., Nature 419, 384 (2002).
4. R. D. Deegan et al., Nature 389, 827 (1997).
In the case of the larger droplets (10 ml), the lines (Fig. 3D). 5. M. A. Ray, H. Kim, L. Jia, Langmuir 21, 4786 (2005).
evaporation of the solvent took longer and al- The contact pinning lines themselves do not 6. K. Mougin, H. Haidara, Langmuir 18, 9566 (2002).
lowed the formation of larger aggregates already align the LC molecules but remain visible, which 7. J. V. Barth et al., Angew. Chem. Int. Ed. 39, 1230 (2000).
in solution, which were subsequently deposited indicates that the formed aggregates are the ones 8. M. A. Ray, H. Kim, L. Jia, Langmuir 21, 4786 (2005).
9. R. D. Deegan, Phys. Rev. E Stat. Phys. Plasmas Fluids
(Fig. 3, A and B). Apparently, there was not that act as a command layer. Second harmonic Relat. Interdiscip. Topics 61, 475 (2000).
enough material present at the contact line to generation (SHG) measurements confirmed that 10. J. Huang, F. Kim, A. R. Tao, S. Conner, P. Yang, Nat.
completely pin it (9). The resulting partial pin- the mesogenic molecules were uniformly aligned Mater. 4, 896 (2005).
ning of the solvent front hindered the retraction parallel to the radially oriented stacks of 1 [that is, 11. S. Vyawahare, K. M. Craig, A. Scherer, Nano Lett. 6, 271
(2006).
of the contact line, and in contrast to the ex- perpendicular to the contact lines, in exceptionally 12. I. I. Smalyukh et al., Phys. Rev. Lett. 96, 177801 (2006).
periments with the smaller droplets, contact large domains (1 cm2) (Fig. 4)]. As for most 13. M. C. Lensen et al., Chem. Eur. J. 10, 831 (2004).
pinning lines were then not formed (Fig. 3F) anisotropic surfaces that show LC alignment (31), 14. C. R. L. P. N. Jeukens et al., Nano Lett. 4, 1401 (2004).
(9). The partial pinning caused a flow of mole- the alignment was probably due to (i) a minimi- 15. Materials and methods are available as supporting
material on Science Online.
cules toward and orthogonal to the contact line. zation of elastic energy and (ii) the presence of
16. S. Ranganathan et al., Chem. Commun. 2001, 2544
The concomitant local increase in the concen- molecular interactions between the LC molecules (2001).
tration of molecules of 1 led to growth of the and the oriented columnar stacks (32). In the case 17. M. L. Bushey, T. Q. Nguyen, W. Zhang, D. Horoszewski,
lines from a direction opposite to the molecular of the ordered porphyrin patterns, however, C. Nuckolls, Angew. Chem. Int. Ed. 43, 5446 (2004).
flow, resulting in an orthogonal orientation with dipole-dipole interactions in particular are ex- 18. A. J. Wilson, M. Musada, R. P. Sijbesma, E. W. Meijer,
Angew. Chem. Int. Ed. 44, 2275 (2005).
respect to the solvent front. The combination of pected to have a large effect, because the head-to- 19. M. P. Lightfoot, F. S. Mair, R. G. Pritchard, J. E. Warren,
(i) the tendency of the molecules to form 1D tail orientation of the amide functions within the Chem. Commun. 1999, 1945 (1999).
aggregates, (ii) the occurrence or absence of linear aggregates, as shown in Fig. 1D, creates a 20. A. Sharma, R. Khanna, Phys. Rev. Lett. 81, 3463
contact line pinning, and (iii) spinodal dewetting macroscopic dipole moment parallel to the (1998).
21. A. Sharma, R. Khanna, J. Chem. Phys. 110, 4929 (1999).
effects resulted in the observed surface pat- stacking axis (17). The use of periodic patterns 22. G. Reiter, Science 282, 888 (1998).
terning in the two cases. created by controlled self-organization may lead 23. A. M. Higgins, R. A. L. Jones, Nature 404, 476 (2000).
Previous reports have described organized as- to a viable and cheap alternative to current meth- 24. A. Sharma, J. Mittal, R. Verma, Langmuir 18, 10213
semblies of polymers (25), dendrimers (3, 26, 27), ods of forming alignment layers. (2002).
25. M. H. Stenzel, C. Barner-Kowollik, T. P. Davis, J. Polym.
and block copolymers (28), leading to crystal-like The remaining challenge in exploiting this Sci. Part Polym. Chem. 44, 2363 (2006).
domains on surfaces. In none of these cases, phenomenon will now be to further control the 26. S. D. Hudson et al., Science 278, 449 (1997).
however, have 1D single molecular stacks spon- self-assembly in such a way that surface patterns 27. V. Percec et al., Nature 430, 764 (2004).
taneously organized into periodic dissipative pat- can be oriented at will. Control over the periodic 28. S. I. Stupp et al., Science 276, 384 (1997).
29. H. Yabu, M. Shimomura, Adv. Funct. Mater. 15, 575 (2005).
terns been observed. Unlike our experiments, arrays might be accomplished by patterned heat-
30. Z. Lin, S. Granick, J. Am. Chem. Soc. 127, 2816 (2005).
constructing such patterns requires invasive tech- ing of the surface with the use of laser gratings or 31. M. Behdani et al., Appl. Phys. Lett. 80, 4635 (2002).
niques such as lithography or sliding glass plates by application of an electric field to align the 32. T. Rasing, I. Musevic, Surfaces and Interfaces of Liquid
(29, 30). high intrinsic dipole moments of the stacks. Crystals (Springer, Heidelberg, 2004).
The line patterns obtained with the large drop- Extra stabilization of the patterns can be achieved 33. Supported by grants from the Netherlands Organization
of Scientific Research (NWO) to R.J.M.N. (TOP grant), to
lets were investigated as possible LC alignment by introducing cross-linkable groups (such as J.A.A.W.E. (VENI grant), to P.S. (FOM/ALW), and to A.E.R.
layers. LC cells, consisting of one glass plate cinnamate, thiophene, or methacrylate units) in (VIDI grant); the National Research School Combination
covered with the aggregates and a non-rubbed the alkyl chains, which would allow modification Catalysis (NRSC-C) to R. van H. and to R.J.M.N.; and the
counter-plate spin-coated with a commercially of the patterns after their deposition on the surface. Royal Netherlands Academy of Science to R.J.M.N.
available polyimide, were prepared and filled with The self-assembly/dewetting technique could also Supporting Online Material
4-cyano-4′-pentyl biphenyl (5CB) molecules in the be applied in conjunction with conventional www.sciencemag.org/cgi/content/full/314/5804/1433/DC1
Materials and Methods
isotropic phase to avoid flow alignment. Polarizing (photo-)lithographic or stamping methods. Figs. S1 to S4
microscopy showed that the cells contained Reference
aligned LC domains of several square millimeters References and Notes Movie S1
(Fig. 4) in the regions of the linear aggregates and 1. H. O. Jacobs, A. R. Tao, A. Schwartz, D. H. Gracias, 25 July 2006; accepted 28 September 2006
no alignment in other areas. Closer inspection G. M. Whitesides, Science 296, 323 (2002). 10.1126/science.1133004

1436 1 DECEMBER 2006 VOL 314 SCIENCE www.sciencemag.org

REPORTS
0.001). Recently, significantly higher average
Probing the Chiroptical Response dissymmetry values have been observed in ag-
gregates of conjugated polymers with chiral side
of a Single Molecule chains (7, 8), and experiments probing specific
fine-structure components of 5D0 → 7F1 tran-
Ruthanne Hassey, Ellen J. Swain, Nathan I. Hammer, Dhandapani Venkataraman, sitions in Europium chelates have shown average
Michael D. Barnes* g values between −0.7 and −1.8 (9). The latter
result is particularly interesting in that such a
Chirally sensitive measurement techniques have generally been restricted to bulk samples. Here, large ensemble-averaged g value can be ob-
we report the observation of fluorescence-detected circular dichroism (FDCD) from single (bridged- served in such systems primarily because of the
triarylamine) helicene molecules by using an excitation wavelength (457 nanometers) in the diminished sensitivity of inner-shell electronic
vicinity of an electronic transition that shows circular dichroism in bulk samples. The distributions transitions to environmental factors in rare earth
of dissymmetry (g) parameters by analysis of signals from pure M- and P-type diastereomers are ions. Thus, an important unresolved question is
almost perfect mirror images of one another, each spanning a range of both positive and negative whether the generally weak chiroptical signature
values. In addition, we observe a well-defined structure in the histogram of dissymmetry from molecular systems is a result of intrinsic
parameters suggestive of specific molecular orientations at the polymer interface. These single- molecular properties or the result of averaging
molecule results highlight strong intrinsic circular dichroism responses that can be obscured by over extrinsic heterogeneities such as molecular
cancellation effects in ensemble measurements of a randomly oriented bulk sample. orientation or different local environments.
Answers to these questions are vital to enhanc-
ptical probes of chirality in organic sys- ecules deposited on the surface of a polymer ing the purity of chiroptical response from a

O tems are nearly as old as organic chem-

istry itself, tracing back to the early
1800s in initial studies of chiral natural products.
film, and we show how the dichroic response is
distributed for pure P- and M-type diaster-
eomers. The measured distributions—each con-
molecule of interest, thus potentially improv-
ing the practicability of such materials in device
applications.
Optical rotary dispersion and circular dichroism structed from several hundred single-molecule The power of single-molecule spectroscopy
are now ubiquitous tools for characterizing and measurements—are almost perfect mirror images to disentangle heterogeneities in a complex phys-
quantifying natural and synthetic chiral systems of each other and are characterized by significantly ical system is well known (10–12). Many elegant
(1, 2). More sophisticated gas- and condensed- larger average chiroptical response than observed published works on orientational dynamics (13),
phase probes have recently brought fresh in- in bulk solution. Surprisingly, the measured spectroscopy of conjugated polymers (14, 15),
sights to the role of solvation and local dielectric distributions span a range of dissymmetry param- and quantum dot systems (16), for example, have
environment in the problem of light-matter in- eters encompassing both positive and negative shown how a molecule-by-molecule approach can
teractions with chiral materials (3–5). However, values. In addition, we find a well-defined provide detailed information on the photophysics
because conventional techniques for probing structure in the histograms suggestive of specific of complex systems. Recently, Venkataraman,
chiroptical properties of molecular systems are molecular orientations at the polymer interface. Riehl, and co-workers demonstrated the syn-
based on scattering or absorption, they neces- The chiroptical response of a molecular sys- thesis and bulk chiroptical characterization of
sarily require extensive ensemble averaging, tem is quantified by the dissymmetry param- a new kind of helicene molecule based on a
thus obscuring information on the specific chir- eter, g. In terms of fundamental molecular bridged triarylamine structure (Fig. 1B) in
optical signature for an individual molecule as properties, g is defined as 4R/D, where R = which the right (P)– or left (M)–handed heli-
well as the heterogeneity of the response asso- Im[〈g |μ| e〉 〈e |m| g〉], |e〉 and |g〉 are electronic cal structure is enforced by the presence of a
ciated with a particular system. Here, we report states involved in the optical transition, μ and m camphanate group at the indicated position (16).
observation of fluorescence-detected circular are the electric and magnetic dipole operators, This camphanate group serves only to main-
dichroism (FDCD) on individual helicene mol- and D = |〈g|μ|e〉|2 is the dipole oscillator strength, tain chirality and does not absorb or emit light
thus giving the range of possible g values of ±2 at the wavelengths used and therefore is not
Department of Chemistry, University of Massachusetts- (6). Experimentally, g is determined by measured expected to contribute to the chiroptical proper-
Amherst, Amherst, MA 01003, USA. differential absorbance or luminescence intensity ties of the helicene molecules. A very small
*To whom correspondence should be addressed. E-mail: as g = 2[eL − eR]/[eL + eR]; typically, ensemble- ensemble-averaged circular polarized luminescence
[email protected] averaged values of g are quite small (0.01 to (De/e ≈ 0.001) was observed from solution-

Fig. 1. (A) Experimental schematic. Linear-polarized laser radiation is peri- (B) was collected in an epi-configuration with a 1.4-NA oil objective and high-
odically modulated between right- and left-handed circularly polarized light by sensitivity CCD camera. (C) Typical (in-focus) fluorescence image and defocused
rotation of a quarter waveplate. Fluorescence from single M2 or P2 molecules image (inset) from a M2 sample.

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REPORTS
phase samples of these species (17), similar in stability of the helicenes is significantly en- Plotted as histograms (Fig. 2B), the distribu-
magnitude to the circular dichroism at the same hanced by the use of a Zeonex supporting film tions of the dissymmetry parameter, g, obtained
excitation wavelengths. Our approach here was over clean glass. In a typical fluorescence image from single M2 and P2 molecules are notable
to use ultradilute solutions to immobilize single obtained from such a sample, ~50 diffraction- mirror images of one another and appear to be the
helicene molecules at the surface of a polymer limited fluorescent spots in a ≈300-mm2 area are sum of a broad symmetric (about g = 0) com-
film and image the fluorescence-detected circu- seen, which show temporal instabilities (blinking ponent spanning a range of g ≈ ±1.3 and a much
lar dichroism by using single-molecule spec- and discrete photobleaching) characteristic of narrower component centered at g ≈ ±0.15. Even
troscopy and imaging techniques. single molecules (Fig. 1C). The defocused image though the compounds were isolated as pure
Figure 1A shows a schematic of our ex- shown in the inset of Fig. 1C illustrates the diastereomers, we considered the possibility that
perimental apparatus, which is similar in many bidirectional nature of the transition moment in the unexpected shape of the distributions could
regards to that described by Kahr and co-workers these chiral species. Similar in nature to de- be associated with the opposite diastereomer
for imaging circular dichroism in inorganic focused images observed in quantum dot sys- formed by partial isomerization in methanol
crystals (18). Using an epi-illumination configu- tems (21), the spatial fluorescence intensity solution, or by reaction with condensed water
ration on a Nikon TE300 microscope with 1.4– patterns for the single helicene molecules are vapor on the film during sample preparation.
numerical aperture (NA) objective, we delivered distinctly different from well-known defocused However, further experiments starting from pure
right- or left-handed circularly polarized light images of single linear dipoles or multichromo- M2 powder under strict anhydrous conditions
from a CW Ar+ ion laser (457 nm; ≈100 mW phoric systems (22). with cyclohexane showed that the broad dis-
nominal power) to the sample by orienting a Figure 2A shows representative fluorescence tribution persisted. Thus, we conclude that the
multi-order quarter waveplate (QWP) on a intensity traces from the M2 and P2 helicenes symmetric component present in both M2 and P2
rotation stage at ±45° with respect to the under excitation with right- and left-handed g distributions at 457-nm excitation is a photo-
(horizontal) input polarization axis, with 5 to 10 circularly polarized laser radiation. In these physical property of the molecular system and
sequential charge-coupled device (CCD) camera examples, the sample is illuminated with al- not the result of sample degradation. Our ex-
exposures (Roper Scientific PhotonMax) for ternating right- and left-circular polarized laser perimental bias toward molecules with higher
each QWP orientation (19). The molecules under radiation every 10 frames, with a 1-s integration photostability might, however, bias the dis-
study here were pure M- and P-type diastereo- time per frame. The dissymmetry factor, g, in the tribution of dissymmetry parameters to higher
mers of helicenes [as verified by 1H–nuclear single-molecule FDCD signal was defined as (absolute) values.
magnetic resonance (NMR)] derivatized with a 2[(IL – IR)/(IL + IR)], where IR and IL are the The structure of the histograms is intriguing
camphanate moiety to enforce a specific chirality measured fluorescence intensities associated with and suggests a nonrandom distribution of chiral
(Fig. 1B). Synthetic details are given elsewhere right or left circularly polarized excitation, re- axes at the polymer-air interface. Differences
(20). For consistency in notation with respect to spectively (23, 24). Single-molecule FDCD dis- between normalized g distributions for P2 cast
previously published work, we refer to the dif- symmetry parameters were determined for each from methanol versus cyclohexane solutions
ferent diastereomers here simply as M2 and P2 right and left circular polarization alternation are subtle and limited to the wings of the two
(17). The 457-nm excitation wavelength excites cycle, and only fluorescent molecules with
transitions within the lowest electronic absorp- sufficient photochemical stability to follow in-
tion band, where bulk solution and solid-film tensity trajectories for at least 1.5 modulation
circular dichroism for these molecules are ob- cycles were used in the analysis. Although some
served. Solutions of the two diastereomers were variations in g are observed within a given single-
dissolved in semiconductor-grade methanol or molecule intensity trajectory, the gross value of g
cyclohexane and diluted to nominal concentra- appears to be well defined for a given molecule
tions of 10−11 M; film-based samples were pre- during the measurement duration, suggesting that
pared by drop-casting ≈200 ml of the ultradilute a particular single-molecule g factor is deter-
solution onto a thin polycylcoolefin (Zeonex) mined primarily by a predominantly static local
polymer film. We find that the photochemical environment.

Fig. 2. (A) Representa-

tive fluorescence inten-
sity traces as a function
of excitation polarization
state for dye-doped poly-
mer nanosphere (black),
single P2 molecule (blue),
and single M2 molecule
(red). The dissymmetry
parameters extracted
from the P2 and M2
traces shown here were
0.54 and −0.63, respec- Fig. 3. Comparison of normalized dissymmetry
tively. (B) Normalized parameter histograms for P2 dispersed from cy-
histograms of FDCD dis- clohexane (dashed blue line and open circles) and
symmetry parameters methanol (solid blue line and triangles). The solid
determined from single- black curve is a fit to the P2/cyclohexane data with
molecule fluorescence measurements. The red curve with open squares represents data from M2, and the a three-component Gaussian fit. The lower graphic
blue curve with open circles represents data from P2. For comparison, results from our control experiment illustrates three different molecular orientations
with dye-doped 20-nm polymer nanospheres are shown (black curve with crosses). Data from ~500 single at the surface: camphanate-down, tripod, and
molecules were used in the construction of the histograms. camphanate-up.

1438 1 DECEMBER 2006 VOL 314 SCIENCE www.sciencemag.org

REPORTS
distributions (Fig. 3) despite the solubility of In summary, we observe that for a given 10. W. E. Moerner, J. Phys. Chem. B 106, 910 (2002).
Zeonex in the hydrocarbon but not the alcohol. bridged-triarylamine helicene diastereomer, there 11. C. Bai, C. Wang, X. S. Xie, P. G. Wolynes, Proc. Natl.
Acad. Sci. U.S.A. 96, 11075 (1999).
Analysis of the M2 and P2 g distributions in- exists a significant probability of finding rela- 12. S. Nie, R. N. Zare, Annu. Rev. Biophys. Biomol. Struct.
dicated three distinct components, with similar tively large positive and negative g values whose 26, 567 (1997).
amplitudes but opposite signs for the two dia- distribution appears as a sum of three distinct 13. A. P. Bartko, K. W. Xu, R. M. Dickson, Phys. Rev. Lett. 89,
stereomers. We propose that these three com- components. We hope this result will spark 026101 (2002).
14. Z. H. Yu, P. F. Barbara, J. Phys. Chem. B 108, 11321
ponents are associated with three distinct stable further experimental and theoretical efforts in (2004).
molecular frame orientations at the surface shown the study of individual chiral fluorophores. Our 15. P. Kumar et al., J. Phys. Chem. B 107, 6252 (2003).
in Fig. 3: camphanate down or up, and tripod work demonstrates the feasibility of interrogating 16. M. Nirmal et al., Nature 383, 802 (1996).
(chiral axis perpendicular to the optical axis). the fundamental nature of the interaction of light 17. J. E. Field, G. Muller, J. P. Riehl, D. Venkataraman, J. Am.
Chem. Soc. 125, 11808 (2003).
These configurations correspond to the helicene with chiral molecules at the single–quantum 18. K. Claborn, E. Puklin-Faucher, M. Kurimoto, W. Kaminsky,
frames parallel or perpendicular to the surface. We system level and provides valuable insights into B. Kahr, J. Am. Chem. Soc. 125, 14825 (2003).
speculate that the opposite-handed component is the photophysics of chiral fluorophores. This 19. The degree of circular polarization at the sample was
associated with the “camphanate-up” orientation result may also pave the way to the develop- determined to be >96%. Tests for artifactual FDCD
performed on single DiI18 molecules randomly oriented
that places the helicene frame in direct contact ment of advanced new materials for efficient
in the x-y sample plane showed a distribution of
with (or solvated by) the polymer film. Thus, the polarized light–emitting diodes (POLEDs) in dissymmetry parameters similar to that of the polymer
two in-plane orientations should be distinguish- next-generation display technologies (26). nanosphere results.
able from each other. Similar effects have been 20. J. E. Field, T. J. Hill, D. Venkataraman, J. Org. Chem. 68,
observed by Vaccaro and co-workers in cavity 6071 (2003).
References and Notes 21. X. Brokmann, L. Coolen, M. Dahan, J. P. Hermier, Phys.
ringdown polarimetry measurements on (S)-(−)- 1. M. Srinivasarao, Chem. Rev. 99, 1935 (1999). Rev. Lett. 93, 107403 (2004).
propylene oxide, which show (for this particular 2. N. Berova, K. Nakanishi, R. W. Woody, Eds., Circular 22. A. P. Bartko, R. M. Dickson, J. Phys. Chem. B 103, 11237
system) that the specific rotation changes sign in Dichroism: Principles and Applications (Wiley-VCH, New (1999).
the transition from the gas-phase to solvated York, ed. 2, 2000). 23. J. P. Riehl, F. S. Richardson, Chem. Rev. 86, 1 (1986).
3. T. Muller, K. B. Wiberg, P. H. Vaccaro, J. Phys. Chem. A 24. F. S. Richardson, J. P. Riehl, Chem. Rev. 77, 773 (1977).
molecule (3), illustrating the sensitivity of the 25. D. Mucha, K. Stadnicka, W. Kaminsky, A. M. Glazer,
104, 5959 (2000).
chiroptical response to molecular environment. 4. M. Y. Sfeir et al., Science 312, 554 (2006). J. Phys. Condens. Matter 9, 10829 (1997).
This effect is also analogous to the observation 5. R. Fasel, M. Parschau, K. H. Ernst, Nature 439, 449 26. S. H. Chen et al., Nature 397, 506 (1999).
in crystals of tartaric acid of a variation in optical (2006). 27. Support from the U.S. Department of Energy Office of
6 Contributions from electric dipole–electric quadrupole Basic Energy Sciences (grant 05ER15695), NSF-
activity that is bisignate in nature (25). Here, we sponsored MRSEC, NSF CHE 0134287, and the
interactions that enter into the chiroptical response at the
are restricted to molecular orientations induced same order of perturbation theory as electric dipole– Intelligence Community Postdoctoral Research Fellowship
by interactions with the substrate and as a result magnetic dipole interactions in the absence of ensemble Program is gratefully acknowledged. E.J.S. acknowledges
observe distinct contributions to the average g averaging are expected to be small. support from the Bates Summer Research Fellowship.
value. This result suggests that in solution phase, 7. A. Satrijo, S. C. J. Meskers, T. M. Swager, J. Am. Chem.
Soc. 128, 9030 (2006). 23 August 2006; accepted 16 October 2006
the measured g value represents a weighted 8. J. N. Wilson et al., J. Am. Chem. Soc. 124, 6830 (2002). Published online 2 November 2006;
average of all possible orientations and inter- 9. R. W. Schwartz, H. G. Brittain, J. P. Riehl, W. Yeakel, F. S. 10.1126/science.1134231
actions with the solvent. Richardson, Mol. Phys. 34, 361 (1977). Include this information when citing this paper.

particles (IDPs), suggesting that some inter-

Organic Globules in the Tagish stellar organic materials have survived intact
(2–4). However, analytical limitations have left

Lake Meteorite: Remnants of the the nature of these materials poorly known.
Tagish Lake is a meteorite whose chemis-
try and mineralogy are intermediate between
Protosolar Disk CI and CM2 carbonaceous chondrites (5). It
was collected immediately after its fall was
Keiko Nakamura-Messenger,1,2* Scott Messenger,1 Lindsay P. Keller,1 witnessed, minimizing terrestrial contamina-
Simon J. Clemett,1,3 Michael E. Zolensky1 tion (5). It has been linked to outer belt as-
teroids from its orbit, reflectance spectrum,
hydrated mineralogy, and abundant carbona-
Coordinated transmission electron microscopy and isotopic measurements of organic globules in
ceous matter, having 2.6 weight percent or-
the Tagish Lake meteorite shows that they have elevated ratios of nitrogen-15 to nitrogen-14 (1.2
ganic carbon (5–7). Tagish Lake organic matter
to 2 times terrestrial) and of deuterium to hydrogen (2.5 to 9 times terrestrial). These isotopic
often occurs as submicrometer, hollow glob-
anomalies are indicative of mass fractionation during chemical reactions at extremely low
ules (8). Similar objects were first observed in
temperatures (10 to 20 kelvin), characteristic of cold molecular clouds and the outer protosolar
meteorite extracts in 1961 (9) and have re-
disk. The globules probably originated as organic ice coatings on preexisting grains that were
cently been reported in several carbonaceous
photochemically processed into refractory organic matter. The globules resemble cometary carbon,
chondrites (10). However, owing to analytical
hydrogen, oxygen, and nitrogen (CHON) particles, suggesting that such grains were important
constituents of the solar system starting materials.
1
Robert M. Walker Laboratory for Space Science, Astro-
arbonaceous chondrite meteorites con- gone complex histories of processing, dilution, materials Research and Exploration Science Directorate,

C tain rare micrometer-sized mineral grains

from evolved stars (stardust) (1). These
meteorites also contain remnants of interstellar
and isotopic exchange with solar system ma-
terials, obscuring its original chemical and phys-
ical state. Rare microscopic inclusions with
NASA Johnson Space Center, Houston, TX 77058, USA.
2
ESC Group/Jacobs Sverdrup, NASA Johnson Space Center,
Houston, TX 77058, USA. 3ESC Group/ERC Inc., NASA
Johnson Space Center, Houston, TX 77058, USA.
organic matter, marked by anomalous H and N highly anomalous H and N isotopic composi- *To whom correspondence should be addressed. E-mail:
isotopic compositions. This material has under- tions occur in meteorites and interplanetary dust [email protected]

www.sciencemag.org SCIENCE VOL 314 1 DECEMBER 2006 1439

REPORTS
and sample limitations, the origins of these processing. However, globules that are attached interstellar clouds have D/H ratios enriched by
objects have not been well understood. to each other have similar H and N isotopic factors of 102 to 105 relative to HD/H2 (13).
We performed coordinated in situ micro- compositions (Fig. 3, A and B), suggesting that Grain surfaces are also predicted to become
structural, chemical, and isotopic studies of Tagish the globules aggregated before incorporation enriched in D/H via single-atom addition of D
Lake globules to establish whether their origins into the Tagish Lake parent body. (14). D-enrichment may also have occurred in
were products of chemical processes in the The isotopic compositions of the globules the protoplanetary disk in the region of the
meteorite parent body, the solar nebula, a cold are indicative of mass fractionation during Kuiper Belt [>30 astronomical units (AU)],
molecular cloud, or circumstellar environment. By chemical reactions at low temperatures (10 to possibly reaching values of protostellar cores
determining their sources, the globules can provide 50 K). In cold molecular clouds, ion-molecule at distances of >100 AU (15). Nitrogen
direct probes of primordial chemical processes. chemical reactions are promoted by cosmic isotopic fractionation is expected at extremely
Fresh samples of Tagish Lake matrix were ray ionization. Gas-phase molecules in cold low T (10 K), with 15N/14N ratios of poly-
sectioned by ultramicrotomy in high-purity S
into 50- to ~70-nm-thick sections (11). We ob-
served numerous, mostly submicrometer, hol- Fig. 1. EFTEM images of
low organic globules in carbonate-free sections Tagish Lake organic glob-
ules. (A) Bright-field TEM
of the meteorite (Fig. 1). Although the average
image of three organic
concentration was about one per 100 mm2,
globules (G15-1 1.3 mm,
aggregates of two to five globules are common. G15-2 0.7 mm, and G15-3
The globule diameters (140 to 1700 nm) vary 0.55 mm) embedded in
significantly more than their wall thickness saponite matrix. (B) Car-
(100 to 200 nm). All but one of the globules ap- bon K-edge EFTEM image
peared hollow in thin sections. High-resolution of the area shown in
transmission electron microscopy (TEM) im- Fig. 3A, showing carbon-
aging and electron energy-loss spectroscopy containing material in
(EELS) show that the globules consist of struc- high contrast. Globule
turally amorphous C that lacks long-range order G15-1 (left) shows micro-
or development of graphite-like domains. The structure that may be
distribution of C, N, and O in the globules was subgrains within its wall.
obtained using energy-filtered TEM (EFTEM) N and H isotopic images
imaging. The walls and cores of the organic of these globules are
globules are almost always free of other matrix shown in boxed areas of
materials. Fig. 3, A and B. (C) Car-
Twenty-six of the globules identified by bon K-edge EFTEM image
TEM and 1100 mm2 of surrounding matrix ma- of globule G8-3 showing
terial were subjected to C and N isotopic im- a typical hollow structure
of an individual organic
aging with a NanoSIMS 50L ion microprobe
globule. N and H isotopic
[(11), table S1]. Remarkably, all of the globules
images of this globule
had elevated 15N/14N ratios, with d15N values are shown in boxed areas
ranging from 200 to 1000‰, significantly of Fig. 3, C and D. (D)
exceeding bulk 15N/14N ratios of CI and CM2 This carbon postedge EFTEM image of globule aggregate G21-1 reveals its internal structure, indicating that it
chondrite meteorites (7, 12) (Fig. 2) and Tagish has incorporated several ~50-nm globules, shown by arrows.
Lake organic matter (77‰) (7). Although ac-
counting for only ~1.5% of the area analyzed,
the globules accounted for 80% of the highly Fig. 2. Carbon and nitro-
15
N-rich material (d15N > 400‰). Eight of these gen isotopic compositions
globules were also measured for H isotopic of Tagish Lake organic
compositions, and all were D-rich, with dD globules compared with
values ranging from 1800 to 8100‰. All ranges observed among
globules are clearly spatially resolved in H and whole-rock samples of CI1
N isotopic images from the surrounding matrix (d15N = 31 – 52‰), CM2
material, with the exception of two globules that (d15N = 13 – 47‰), whole
are adjacent to D-rich matrix material (Fig. 3). rock Tagish Lake (diamond),
The C isotopic compositions of the globules had and Tagish Lake organic
a narrower range (d13C ~ –77 to +16‰), gen- matter (square) (7, 12). The
erally below the value of bulk Tagish Lake anomalous CM2 meteorite
organic matter (–9‰)(7). Bells has a bulk d15N value
of 335‰ (12).
The fact that the globules all exhibited N
and H isotopic anomalies that greatly ex-
ceeded the surrounding meteorite matrix rules
out their possible formation in the Tagish
Lake parent body. The globules have highly
variable H and N isotopic ratios, even those
within a few mm of each other (Fig. 3, C and
D), making it unlikely that the isotopic var-
iations resulted primarily from parent body

1440 1 DECEMBER 2006 VOL 314 SCIENCE www.sciencemag.org

REPORTS
cyclic aromatic hydrocarbon (PAH) molecules original interstellar organics with solar system susceptible to chemical oxidation. Interesting-
and grain surfaces predicted to be enhanced materials. ly, two of the globules are adjacent to D-rich
by up to a factor of two (16). These conditions Isotopic measurements of meteorites and matrix material (Fig. 3, B and D) that may have
occur in cold molecular clouds and at the IDPs at micrometer scales have revealed a originated from the globules. This is not
outermost regions of protoplanetary disks (>100 much greater isotopic variability compared observed in the N isotopic images, implying
AU). The N isotopic anomalies in the globules with values of bulk organic extracts, in some that the 15N enrichments are carried by lower-
likely resulted from chemical fractionation, cases reaching values of cold molecular cloud solubility phases than the D-rich material, such
not from nucleosynthesis, because they are molecules (dD = 50,000‰) (3, 4). In several as insoluble macromolecular material, PAHs,
enriched in both D/H and 15N/14N and also cases, D- and 15N-rich “hotspots” were asso- or amines. This would support the prediction
lack the large C isotopic anomalies charac- ciated with carbonaceous materials (4, 20, 21). that PAHs obtain the strongest 15N enrichments
teristic of evolved stars. Their narrow range of Here we show that in the Tagish Lake meteorite, in protostellar cores (16).
13 12
C/ C ratios limits the degree of accompany- the primary carriers of the most highly anoma- Ice grains in cold molecular clouds are either
ing C isotopic fractionation to <50‰ (Fig. 2), lous H and N are distinctive submicrometer primarily H2O-rich polar ices containing CO2,
consistent with expectations that significant C organic globules. CH3OH, H2CO, and NH3, or nonpolar ices
isotopic fractionation would not be preserved Since forming, these globules experienced containing CO, CO2, N2, and O2 (24, 25). The
(14, 17). wide ranges of thermal and chemical con- globules are more likely to have formed by
Carbonaceous chondrites contain a diversity ditions, ultimately residing in an aqueous solu- condensation of polar molecular ices, where
of organic compounds, including hundreds of tion that formed the phyllosilicates matrix of abundant free H (H/H2 > 1) was available to
solvent extractable species and a dominant the Tagish Lake parent body (5). Hydrothermal form hydrocarbons and deuterate molecules on
insoluble macromolecular material (18). These alteration occurred among many carbonaceous grain surfaces. The isotopic variations of the
materials are moderately enriched in D and/or chondrites, leaving variable imprints on their globules may reflect the sensitivity of H and N
15
N (typically dD <1000‰; d15N <200‰), with organic matter (18, 22). Tagish Lake organic isotopic fractionation to temperature, radiation
significant isotopic variations among different matter is unusually poor in soluble species that environment, and chemical composition.
classes of organics and meteorites (19). In gen- may have been lost by low-temperature chem- Interstellar ice analogs are readily converted
eral, however these anomalies are well below ical oxidation, resulting in the production of into complex refractory organic compounds by
those expected for cold molecular cloud ma- carbonate (23). Because the organic globules exposure to ultraviolet (UV) radiation (26, 27).
terials, probably reflecting histories of al- are only found in carbonate-free regions of The penetration depth of UV radiation is remark-
teration, mixing, and isotopic exchange of Tagish Lake, this suggests that the globules are ably similar to the wall thickness of the organic
globules (100 to 200 nm). The interiors of these
objects may have been preexisting ice grains that
were protected from radiation processing by the
organic mantle. These cores remained more
volatile than the coatings and would have
volatilized at a later stage, leaving the organic
globules with hollow cores.
Alternatively, the hollow structures of the
globules may have resulted from asteroidal
aqueous alteration. This is suggested by exper-
imental production of hydrophobic, vesicle-rich
materials from UV-irradiated interstellar ice
analogs exposed to alkaline solutions, similar
to conditions proposed for aqueous alteration of
carbonaceous chondrites (28, 29). However,
these materials are generally much larger and
weaker than the Tagish Lake organic globules.
Whatever their formation process, the
organic globules very likely originated at the
outer regions of the protosolar disk in the re-
gion of the Kuiper Belt or in the preceding
cold molecular cloud, well beyond the influ-
ence of the Sun and the nascent planetary
system. Consequently, similar organic globules
should also have been incorporated into com-
etary parent bodies. Interestingly, many par-
ticles detected during the Giotto and Vega
encounters with comet Halley were primarily
composed of the elements C, H, O, and N
(CHON particles) (30). The size range (40 to
Fig. 3. (A) Nitrogen isotopic image of section G8-3 containing a uniformly 15N-enriched globule 2000 nm) and bulk compositions of CHON
aggregate. The box is the field of view of Fig. 1A. (B) Hydrogen isotopic image of the globule particles match the properties of the organic
aggregate in Fig. 3A. The aggregate is uniformly enriched in D/H and is adjacent to D-rich matrix. globules studied here, suggesting that such
(C) Nitrogen isotopic image of section G15 containing three globules with differing 15N- grains were prevalent throughout the proto-
enrichments. The boxes are fields of view of Fig. 1A and Fig. 1B. (D) Hydrogen isotopic image of planetary disk. Microscopic organic globules
the globules in Fig. 1C. All three globules are D-rich, but the magnitudes of the H and N isotopic may thus have been a common form of
anomalies are not correlated. prebiotic organic matter delivered to the early

www.sciencemag.org SCIENCE VOL 314 1 DECEMBER 2006 1441

REPORTS
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the data and the methodologies used so far.

Increasing Trend of Extreme Short-duration extreme rain events are a conse-
quence of small-scale convective instabilities

Rain Events Over India in a in a moist atmosphere. Although a fraction of

extreme rain events is triggered in the back-
ground of synoptic disturbances (17) and is
Warming Environment preferentially located around the tracks of mon-
soon lows and depressions, a large fraction
B. N. Goswami,1* V. Venugopal,2 D. Sengupta,2 M. S. Madhusoodanan,2 Prince K. Xavier2 arises from processes like severe thunderstorms
and is more uniformly distributed in space and
Against a backdrop of rising global surface temperature, the stability of the Indian monsoon time. Even if the total number of extreme events
rainfall over the past century has been a puzzle. By using a daily rainfall data set, we show (i) over a homogeneous large-scale environment
significant rising trends in the frequency and the magnitude of extreme rain events and (ii) a were to have an increasing trend, no significant
significant decreasing trend in the frequency of moderate events over central India during the trend may appear in data from a single station
monsoon seasons from 1951 to 2000. The seasonal mean rainfall does not show a significant because of the inherently large variability and/or
trend, because the contribution from increasing heavy events is offset by decreasing moderate sampling issues (18–23). Therefore, we exam-
events. A substantial increase in hazards related to heavy rain is expected over central India in the ined the trend of daily heavy and very heavy rain
future. events over a relatively large region.
We used daily gridded rainfall data at 1°-by-1°
nalysis of rain gauge data shows that Extreme rainfall results in landslides, flash resolution from the India Meteorological De-

A Indian monsoon rainfall has remained

stable over the past century even though
the global mean surface temperature has risen
floods, and crop damage that have major
impacts on society, the economy, and the
environment. Although prediction of such
partment (IMD), based on 1803 stations (24, 25)
that have at least 90% data availability, for the
period 1951–2000. The interannual variability
steadily (1–3). Although the amount of sum- extreme weather events is still fraught with of JJAS all-India rainfall (AIR2) from this data
mer monsoon rain [June to September (JJAS) uncertainties, a proper assessment of likely set (Fig. 1B) is similar to AIR, which is a long-
seasonal mean all-India rainfall, AIR] has some future trends would help in setting up infra- term data set based on 306 stations (26). Daily
interdecadal variability (4), it has no significant structure for disaster preparedness. anomalies of rainfall at each grid box were
long-term trend (Fig. 1). Physical considerations The number of severe cyclonic storms over constructed as deviations of observed daily
and model studies indicate that tropospheric the north Indian Ocean (IO) has shown an in- values from a smoothed climatological annual
warming leads to an enhancement of moisture creasing trend in the past 3 decades (12, 13), cycle (the sum of the mean and first three
content of the atmosphere (5) and is associated consistent with similar findings over other harmonics of the daily climatology). The
with an increase in heavy rainfall events (6–11). basins (12). However, no coherent signal has climatological mean and variance of daily
1
emerged from investigations of the trend of summer monsoon rainfall have large spatial
Indian Institute of Tropical Meteorology, Doctor Homi daily station rainfall data over India (13–16), variability across the country (Fig. 1A and fig.
Bhabha Road, Pashan, Pune 411 008, India. 2Centre for
Atmospheric and Oceanic Sciences, Indian Institute of with some stations showing an increasing trend S1). However, over central India (CI, 74.5°E
Science, Bangalore, Karnataka 560 012, India. whereas others show a decreasing trend. The to 86.5°E and 16.5°N to 26.5°N, containing
*To whom correspondence should be addressed. E-mail: ambiguity in the existence of a trend in mon- 143 grid boxes) the mean and the standard de-
[email protected] soon rainfall extremes may be partly related to viation are reasonably homogeneous (spatially

1442 1 DECEMBER 2006 VOL 314 SCIENCE www.sciencemag.org

REPORTS

Fig. 1. (A) Climatological mean summer monsoon rainfall (mm/day). The box no trend. The AIR2 (blue) is the normalized seasonal mean AIR on the basis of the
indicates the CI region used in our analysis. (B) Normalized (by the interannual new gridded rainfall data (24). The seasonal mean and standard deviation are
standard deviation) JJAS AIR based on 306 stations (26) from 1871 to 2003 94.0 cm and 9.1 cm, respectively. The CIR (red) is the normalized seasonal mean
(bars). The mean is 84.9 cm, and the standard deviation is 8.4 cm. The solid black over CI on the basis of the gridded rainfall data set, the mean and the standard
line represents an 11-year running mean indicating interdecadal variability but deviation of which are 69.5 cm and 11.2 cm, respectively.

uniform). Therefore, we select CI as the region Fig. 2. (A) Temporal variation

to examine the trend of extreme rainfall over (1951 to 2000) in the variance of
India. daily anomalies during summer
The gridded daily data are smoother than the monsoon seasons (June 1 to Sep-
individual station data because of averaging tember 30), together with its linear
over a 1°-by-1° box. The maximum 1-day rain- trend (dashed line). (B) Coefficient
fall during the summer monsoons of 1951 to 2003 of variability of daily precipitation
in any box over CI is 58.2 cm. The seasonal during summer monsoon season
mean over CI is 5.7 mm of rain in a day (mm/day), and its trend (thin line) together
whereas the standard deviation of the daily with JJAS SST anomalies averaged
over tropical IO and their trend
anomalies is 11.5 mm/day. Although a fixed
(bold line). Statistically significant
threshold for defining extreme events is not
trends (0.01 significance level) are
appropriate over regions where the mean climate calculated on the basis of a t test,
has large spatial variability (27, 28), a fixed with a sample size of 50, under a
threshold can be used to define extreme rain null hypothesis of no trend.
events over CI, where the seasonal mean climate
as well as the daily variability is reasonably
homogeneous (Fig. 1A and fig. S1). We used
100 mm/day in a 1°-by-1° box as a threshold to
define a heavy rain event, whereas a threshold of
150 mm/day was used to define a very heavy
event.
The temporal variance of daily rainfall anom-
alies averaged over CI shows a significant in-
creasing trend (at 0.01 significance level) during
1951 to 2000 (Fig. 2A). The increasing trend of
the coefficient of variability, defined as the ratio
of the standard deviation to the mean, of daily be correlated on a year-to-year basis. The long- (R ≥ 100 mm/day) and very heavy (R ≥ 150
monsoon rainfall (Fig. 2B) is a consequence of term increase of daily rainfall variance is likely mm/day) events over CI shows clear and sig-
the absence of a trend in the seasonal mean due to the warming trend of tropical IO JJAS nificant (at 0.01 significance level) increasing
(Fig. 1) and an increasing trend in the standard SST (Fig. 2) and the associated increase in trends (Fig. 3) (29), whereas that of moderate
deviation. A trend in daily rainfall variance is water vapor (5). events shows a significant (at 0.1 significance
related to a trend in large-scale moisture availa- The frequency histogram of daily rainfall at level) decreasing trend. There is a 10% increase
bility (5), which in turn is due to gradual warming each 1° by 1° box (R) over CI during the sum- per decade in the level of heavy rainfall activity
of sea surface temperature (SST) (7). However, mer monsoons of 1951 to 1970 and 1981 to since the early 1950s (Fig. 3A), whereas the
interannual changes in moisture content over CI 2000 was separately constructed (plotted as number of very heavy events has more than dou-
can be influenced by regional-scale land surface line curves in fig. S2) to assess the increase in bled (Fig. 3B), indicating a large increase in
processes as well as by atmospheric teleconnec- variance in recent decades compared with those disaster potential. These findings are in tune with
tions associated with remote SST such as the El of the 1950s and 1960s. The tails of the histo- model projections (6–11) and some observations
Niño and Southern Oscillation (ENSO). Al- gram indicate a larger number of extreme events (30) that indicate an increase in heavy rain events
though El Niño events are generally associated (≥100 mm/day of rain) during 1981–2000. On and a decrease in weak events under global
with positive SST anomaly over the tropical IO, the other hand, the number of light to moderate warming scenarios.
they lead to drying of the atmosphere over CI events (≥5 mm/day but <100 mm/day) have In order to see whether the unambiguous in-
through large-scale subsidence. As a result, decreased during 1981 to 2000 compared with crease in the frequency of heavy and very heavy
daily CI rainfall variance and IO SST need not 1951 to 1970. In fact, the frequency of heavy events is also accompanied by an increase in the

www.sciencemag.org SCIENCE VOL 314 1 DECEMBER 2006 1443

REPORTS
intensity of heavy events, we examined the rain with events of higher intensity contributing to assess the role of sampling and variability, we
intensity between 99 and 99.99 percentiles (31) the higher percentiles. For instance, the average examined the number of heavy rain events over
of summer monsoon rainfall (Fig. 4A). The intensity of the heaviest four events in each regions of increasing size (fig. S3). We find that
rainfall intensity that contributed to the 99.75 monsoon season (Fig. 4B) shows an ~10% per for regions smaller than about 800 km by 800
percentile in the early 1950s seems to contribute decade increase over the 50-year period (18 to km, it is difficult to find significant trends in
to only the 99.5 percentile in the early 1990s, 26 cm), significant at 0.01 significance level. heavy rain events. On the other hand, the whole
Although the above results present strong of India cannot be taken as one unit to in-
evidence of an increase in the number of ex- vestigate such trends. The northeast and the west
treme monsoon weather events over India over coast are regions of high mean (Fig. 1A) and
the past half century, the Indian monsoon cli- high variability (fig. S1), and local orography
mate (seasonal mean monsoon rainfall) remains has a strong influence on the rainfall over both
stable for the same period (Fig. 1). The findings regions. Therefore, trends in extreme rainfall
in Fig. 3 help us piece this puzzle together. Note due to a warming environment are difficult to
that although the frequency histograms for the discern in these regions.
two periods (1951 to 1970 and 1981 to 2000) In spite of considerable year-to-year variabil-
have significant differences (fig. S2), the mean ity, there are significant increases in the frequen-
rainfall during these periods is nearly identical cy and the intensity of extreme monsoon rain
at 5.75 mm and 5.69 mm, respectively. The heavy events in central India over the past 50 years.
events (≥100 mm/day of rain) contribute about Although desirable for applications, it is difficult
6.4% to the seasonal mean, whereas moderate to detect signals of climate change in extreme
events (from 5 mm/day to <100 mm/day) con- rain events at individual stations; instead, as we
tribute about 85.8%. Although the relative con- show, one needs a sufficiently large area to dis-
tributions to the mean from these two classes do cern a trend reliably. The observed trends suggest
not balance in a given year, the contribution enhanced risks associated with extreme rainfall
from the decreasing trend of moderate events is over India in the coming decades.
partially offset by that from increasing heavy
Fig. 3. Temporal variation (1951 to 2000) in the rain events (7). Consequently, the seasonal total References and Notes
number (N) of (A) heavy (R ≥ 100 mm/day, bold does not show any statistically significant change 1. J. T. Houghton et al., Eds., Climate Change 2001: The
line) and moderate (5 ≤ R < 100 mm/day, thin over longer time scales. Scientific Basis (Cambridge Univ. Press, Cambridge,
2001).
line) daily rain events and (B) very heavy events Previous attempts to detect trends in extreme
2. A. N. Rayner et al., J. Geophys. Res. 108,
(R ≥ 150 mm/day) during the summer monsoon rain events by using station data were incon- 10.1029/2002JD002670 (2003).
season over CI. The statistical significance of the clusive, probably because of the large year-to- 3. M. E. Mann, R. S. Bradley, M. K. Hughes, Geophys. Res.
trends (dashed lines) was calculated as in Fig. 2. year variability in Indian monsoon rainfall. To Lett. 26, 759 (1999).
4. B. N. Goswami, in The Asian Monsoon, B. Wang, Ed.
(Praxis, Springer, Berlin, 2005), ch. 7, pp. 295–327.
5. K. E. Trenberth, J. Fasullo, L. Smith, Clim. Dyn. 24, 741
(2005).
6. M. R. Allen, W. J. Ingram, Nature 419, 224 (2002).
7. K. E. Trenberth, A. Dai, R. M. Rasmussen, D. B. Parsons,
Bull. Am. Meteor. Soc. 84, 1205 (2003).
8. K. J. Hennessy, J. M. Gregory, J. F. B. Mitchell, Clim. Dyn.
13, 667 (1997).
9. H. B. Gordon, P. H. Whetton, A. B. Pittock, A. M. Fowler,
M. R. Haylock, Clim. Dyn. 8, 83 (1992).
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(2000).
11. V. A. Semenov, L. Bengtsson, Clim. Dyn. 19, 123
(2002).
12. P. J. Webster, G. J. Holland, J. A. Curry, H.-R. Chang,
Science 309, 1844 (2005).
13. K. Rupa Kumar, K. Krishnakumar, R. G. Ashrit, S. K.
Patwardhan, G. B. Pant, in Climate Change and India,
P. R. Shukla et al., Eds. (Tata McGraw Hill, New Delhi,
2002), pp. 24–75.
14. K. Krishnakumar et al., paper presented at the
Workshop on Indices and Indicators for Climate Extremes,
National Climate Data Center, Ashville, NC, 3 to 6 June
1997.
15. S. Sen Roy, R. C. Balling Jr., Int. J. Climatol. 24, 457
(2004).
16. L. V. Alexander et al., J. Geophys. Res. 111,
10.1029/2005JD006290 (2006).
17. P. A. Francis, S. Gadgil, Meteorol. Atmos. Phys. 94, 27
10.1007/s00703-005-0167-2 (2006).
18. X. Zhang, W. D. Hogg, E. Mekis, J. Clim. 14, 1923
(2001).
19. X. Zhang, F. W. Zwiers, G. Li, J. Clim. 17, 1945
(2004).
20. D. A. Stone, A. J. Weaver, F. W. Zwiers, Atmos. Ocean 38,
321 (2000).
Fig. 4. Temporal variation (1951 to 2000) in (A) 99.4 to 99.99 percentiles of seasonal rainfall and (B) 21. C. Frei, C. Schär, J. Clim. 14, 1568 (2001).
the mean rainfall of the four highest rain events every season (R1..4). Color bar in (A) indicates rain 22. P. Frich et al., Clim. Res. 19, 193 (2002).
intensity in mm/day. The statistical significance of the trend (dashed line) was calculated as in Fig. 2. 23. P. Ya. Groisman et al., J. Clim. 18, 1326 (2005).

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REPORTS
24. M. Rajeevan, J. Bhate, J. D. Kale, B. Lal, Curr. Sci. 91, 29. The frequency histogram of 1980 to 2000 rain is Government of India, for partial support for this work,
296 (2006). consistently above that of 1951 to 1970 rain for and J. Srinivasan for useful discussions.
25. Materials and methods are available on Science Online. intensities greater than 70 mm/day (fig. S2B). The
26. B. Parthasarathy, A. A. Munot, D. R. Kothawale, “Monthly number of events larger than 70 mm/day shows a trend
and seasonal rainfall time series for all India, homogeneous significant at the 0.1 significance level. Supporting Online Material
divisions and meteorological subdivisions, 1871-1994” 30. M. Brunetti, M. Colacino, M. Maugeri, T. Nanni, Int. J. www.sciencemag.org/cgi/content/full/314/5804/1442/DC1
(Technical Report RR065, ISSN 025201075, Indian Institute Climatol. 21, 299 (2001). Materials and Methods
of Tropical Meteorology, Pune, India, 1996). 31. For each year, the calculation of percentiles is based on Figs. S1 to S3
27. M. Haylock, N. Nicholls, Int. J. Climatol. 20, 1533 122 days (monsoon season) and 143 grid points.
2000). 32. We thank the IMD for making the daily gridded rainfall 3 July 2006; accepted 23 October 2006
28. M. J. Manton, Int. J. Climatol. 21, 269 (2001). data available, the Department of Ocean Development, 10.1126/science.1132027

Male Fertility and Sex Ratio depend exclusively on body size, but also on the
ability of males to fertilize females after copula-
tion. Male fertility is advertised by antler size and
at Birth in Red Deer complexity, so more-fertile males also have
larger and more elaborate sexual characters,
which may be inherited by their sons (7).
Montserrat Gomendio,1* Aurelio F. Malo,1 Ana J. Soler,2 Maria R. Fernández-Santos,2 We tested the hypothesis that more-fertile
Milagros C. Esteso,2 Andrés J. García,2 Eduardo R. S. Roldan,1*† Julian Garde2† red deer males produce more sons. The key
challenge was to disentangle male and female
Efforts to test sex ratio theory have focused mostly on females. However, when males possess
effects by designing an experiment to retain the
traits that could enhance the reproductive success of sons, males would also benefit from the
inter-male variation in fertility rates found in
manipulation of the offspring sex ratio. We tested the prediction that more-fertile red deer males
natural populations while minimizing differences
produce more sons. Our findings reveal that male fertility is positively related to the proportion of
between females (8). Thus, our experimental de-
male offspring. We also show that there is a positive correlation between the percentage of
sign was aimed at eliminating several female
morphologically normal spermatozoa (a main determinant of male fertility) and the proportion of
factors known to influence sex ratios: (i) We
male offspring. Thus, males may contribute significantly to biases in sex ratio at birth among
avoided the possibility that females may bias sex
mammals, creating the potential for conflicts of interest between males and females.
ratio in response to male quality by artificially
inseminating females so that they had no direct
he Trivers and Willard hypothesis (1) for izes the ovum. Thus, mammalian males may experience with the males. (ii) We minimized

T sex allocation predicts that parents

should increase the production of the
sex with the higher fitness benefit. This hypothesis
have more control over the mechanisms of
sex determination than they do in other taxa.
In mammals, male fertility may have a great
differences in body condition by using a sample of
females that were all in good physical condition,
were kept under similar environmental conditions,
has been applied most often to mothers, who have influence on the reproductive success of sons. and had access to an unlimited food supply. (iii)
a strong influence on offspring quality through Ungulates are good models to test sex ratio All females were inseminated at the same time in
maternal care. It can also apply to any trait that theory because they are sexually dimorphic in relation to ovulation, avoiding the confounding
parents transmit to offspring that has a differential body size, variance in reproductive success is effects of insemination time. In contrast, by using
effect on the reproductive success of sons and greater among males, and the reproductive sperm collected during the rut from males living
daughters. Thus, among birds, offspring sex ratios success of sons is more strongly influenced by in natural populations, we ensured a representative
may be adjusted in relation to the attractiveness of maternal investment. Early studies on red deer sample of the large degree of variation in male
the father, because sons will inherit large sexual (Cervus elaphus) found support for the predic- fertility previously described (6).
ornaments and will achieve high reproductive tion that high-quality mothers should produce When the entire study sample is considered,
success (2). However, it is assumed that such sons (4), but subsequent studies have generated a similar number of male and female offspring
manipulation is under female control, because in inconsistent results (5). Our previous studies have were produced (Table 1). However, among
birds females are the heterogametic sex. shown that in natural populations of red deer, males, differences in fertility rates and in the
The possibility that males may also facul- males differ markedly in their fertility rates, and proportion of male offspring were substantial.
tatively adjust sex ratio has seldom been more-fertile males have faster swimming sperm Male fertility rates ranged from 24 to 70%, and
considered. In haplodiploid insects, the off- and a greater proportion of normal spermatozoa the proportion of male offspring ranged from 25
spring sex depends on whether the ovum is (6). Thus, male reproductive success may not to 72% (Table 1).
fertilized or not, and males may constrain sex
ratios because males with poor-quality ejacu-
lates fail to fertilize the ova (3). In mammals, Table 1. Descriptive statistics [mean, standard deviation (SD), and range] for male fertility rates,
males are the heterogametic sex, and off- proportion of male offspring sired, percentage of normal sperm, sperm swimming-velocity
parameters, and number of hinds inseminated per male (n = 14 red deer stags). VCL, curvilinear
spring sex is determined by whether an X- or
velocity; VSL, straight-line velocity; VAP, average path velocity.
Y-chromosome–bearing spermatozoon fertil-
Range
Parameters Mean SD
1
Reproductive Ecology and Biology Group, Department of Evo- min–max
lutionary Ecology, Museo Nacional de Ciencias Naturales [Consejo Fertility rate (%) 50.39 13.06 24–70
Superior de Investigaciones Científicas (CSIC)], 28006-Madrid,
Spain. 2Instituto de Investigación en Recursos Cinegéticos [CSIC- Proportion of male offspring 0.50 0.14 0.25–0.72
Universidad de Castilla-La Mancha–Junta de Communidades de Morphologically normal spermatozoa (%) 80.07 8.78 65–95
Castilla-La Mancha (JCCM)], 02071-Albacete, Spain. VCL (mm/s) 126.87 28.48 85–163
*To whom correspondence should be addressed. E-mail: VSL (mm/s) 67.86 27.31 28–111
[email protected] (M.G.); [email protected] VAP (mm/s) 88.74 26.52 53–122
(E.R.S.R.)
Hinds inseminated per male 24.57 16.00 11–69
†These authors contributed equally to this work.

www.sciencemag.org SCIENCE VOL 314 1 DECEMBER 2006 1445

REPORTS
Fig. 1. (A) Relation
between male fertility
[(number of hinds
pregnant/number of
hinds inseminated) ×
100] and proportion
of male offspring sired.
(B) Relation between
percentage of normal
spermatozoa and pro-
portion of male off-
spring sired.

There was a significant relation between male could influence sperm shape, size, and func- smaller antlers (7), their ability to defend females
fertility and the proportion of male offspring sired tion. Furthermore, it has recently been shown for a long period of time may be constrained.
(squared correlation coefficient r 2 = 0.41, P = that males with deletions in the Y chromosome Furthermore, in Mediterranean populations food is
0.013). More-fertile males sired a greater number produce Y-bearing spermatozoa with morpholog- scarce during the mating season, and males either
of sons, and less-fertile males sired more daughters ical abnormalities that are less efficient at fertiliza- defend harems or establish territories where food
(Fig. 1A). There was also a significant relation tion, resulting in sex ratio biases toward females resources are concentrated (16). Females move
between the percentage of morphologically (14). Thus, red deer males with low fertility rates between territories and harems while searching for
normal spermatozoa and the proportion of male may have a lower proportion of morphologically food; thus, repeated copulations with the same
offspring sired per male (r 2 = 0.37, P = 0.021) normal spermatozoa as a consequence of genetic female may be rare. Finally, frequent copulations
(Fig. 1B). In contrast, no significant relation was information on the Y chromosome, which would may lead to sperm depletion among low-fertility
found between sperm velocity parameters and the also impair the chances of fertilization of Y- males given their limited sperm numbers; there is
proportion of male offspring sired (P > 0.05). bearing spermatozoa. On the contrary, males with evidence that in natural populations, frequent
Thus, of the two main determinants of male high fertility rates may produce more-competitive copulation leads to sperm depletion and decreases
fertility—sperm swimming velocity and the Y-bearing spermatozoa. Alternatively, females male siring success (17). Thus, in natural popula-
proportion of normal spermatozoa (6)—the latter could influence the fertilization success of X- tions, differences in fertility rates are likely to
was found to be associated with sex ratio. This and Y-bearing spermatozoa depending on the contribute substantially to differences between
may be the case because the proportion of normal fertility of the male. This would require that males in lifetime reproductive success.
spermatozoa is more likely to be inherited by sons females be able to assess ejaculate quality (and Our findings suggest that mammalian males
(9) than sperm swimming velocity, which may be more specifically the proportion of normal can manipulate the sex ratio of their offspring,
influenced to a greater extent by environmental spermatozoa) and bias sex ratio accordingly, thus creating an unforeseen evolutionary sce-
factors (10). Thus, males with a higher propor- given that in our experimental design females nario that includes conflicts of interest between
tion of normal spermatozoa may benefit from were prevented from evaluating male quality or males and females. For instance, a fertile male
producing sons who will inherit the trait that copulatory behavior. This hypothesis assumes may benefit from producing sons, but the costs
will increase their fertility, and they will thus that differences in fertilization success between of raising a male may be high for a female in
achieve high reproductive success. In contrast, X- and Y-bearing spermatozoa are caused, not by poor physical condition (18). This level of con-
low-fertility males will benefit from producing differences in competitiveness between them (as flict may improve our ability to explain biases in
daughters who will not inherit their father’s proposed by the previous hypothesis), but by sex ratio at birth.
poor ejaculate quality. female selection in the reproductive tract.
There are two possible mechanisms by which Our experimental approach reveals unexpected- References and Notes
males may adjust sex ratio. First, although it is ly large differences in fertility rates between males 1. R. L. Trivers, D. E. Willard, Science 179, 90 (1973).
assumed that mammalian males produce equal from natural populations when females are artificial- 2. H. Ellegren, L. Gustafsson, B. C. Sheldon, Proc. Natl.
Acad. Sci. U.S.A. 93, 11723 (1996).
numbers of X- and Y-bearing spermatozoa as a ly inseminated. Are such differences in male fertility
3. H. J. Henter, J. Evol. Biol. 17, 886 (2004).
consequence of meiotic cell division, ejaculates may likely to occur in natural contexts? In the wild, low- 4. T. H. Clutton-Brock, S. D. Albon, F. E. Guinness, Nature
differ in the proportion of Y-bearing spermato- fertility males could compensate by transferring 308, 358 (1984).
zoa (11), resulting in biases in sex ratio at birth. more spermatozoa per ejaculation. This is un- 5. A. J. M. Hewison, J. M. Gaillard, Trends Ecol. Evol. 14,
Thus, high- and low-fertility males could differ in likely to occur because in natural populations, 229 (1999).
6. A. F. Malo et al., Biol. Reprod. 72, 822 (2005).
the proportion of Y-bearing spermatozoa in the low-fertility males have smaller testes that produce 7. A. F. Malo, E. R. S. Roldan, J. Garde, A. J. Soler, M. Gomendio,
ejaculate. Second, Y-bearing spermatozoa could be fewer spermatozoa (6), a trait that is known to have Proc. R. Soc. London B Biol. Sci. 272, 149 (2005).
at an advantage in relation to X-bearing spermato- a major influence on fertility (15). Thus, the dif- 8. Materials and methods are available as supporting
zoa when produced by more-fertile males, whereas ferences in fertility rates when all females are material on Science Online.
9. J. Smital, J. Wolf, L. L. De Sousa, Anim. Reprod. Sci. 86,
the opposite may occur among less-fertile males. inseminated with equal sperm numbers are likely 119 (2005).
Differences between males in the competitive- to be exacerbated when differences in sperm num- 10. S. J. Kilgallon, L. W. Simmons, Biol. Lett. 1, 253 (2005).
ness of X- and Y-bearing spermatozoa could arise bers come into play in natural contexts. Alterna- 11. J. E. Chandler, A. M. Canal, J. B. Paul, E. B. Moser,
through differential expression of genes carried in tively, low-fertility males could enhance their Theriogenology 57, 1327 (2002).
12. C. W. LaMunyon, S. Ward, Proc. Natl. Acad. Sci. U.S.A.
the sex chromosomes (12). Such postmeiotic fertilization success by copulating more often with 94, 185 (1997).
expression of germ line–specific X- or Y-linked the same female, but the opportunities to do so may 13. P. J. Wang, D. C. Page, J. R. McCarrey, Hum. Mol. Genet.
genes has recently been demonstrated (13) and be limited. Because low-fertility males have 14, 2911 (2005).

1446 1 DECEMBER 2006 VOL 314 SCIENCE www.sciencemag.org

REPORTS
14. M. A. Ward, P. S. Burgoyne, Biol. Reprod. 74, 652 18. M. Gomendio, T. H. Clutton-Brock, S. D. Albon, JCCM. We thank A. Cockburn, J. M. Cummins, C. LaMunyon,
(2006). F. E. Guinness, M. J. A. Simpson, Nature 343, 261 E. Martínez, and J. M. Vázquez for helpful discussions.
15. M. Gomendio, A. H. Harcourt, E. R. S. Roldan, in Sperm (1990).
Competition and Sexual Selection, T. R. Birkhead, 19. Empresa Medianilla allowed work at Finca Las Lomas. Supporting Online Material
A. P. Møller, Eds. (Academic Press, London, 1998). Funding was provided by Ministerio de Ciencia y Tecnologia, www.sciencemag.org/cgi/content/full/314/5804/1445/DC1
16. J. Carranza, P. Fernandez-Llario, M. Gomendio, Ethology Ministerio de Educacion y Ciencia, European Regional Materials and Methods
References
102, 793 (1996). Development Fund, and Instituto Nacional de Investigacion
17. B. T. Preston, I. R. Stevenson, J. M. Pemberton, K. Wilson, y Tecnologia Agraria. A.F.M. was supported by a studentship 26 July 2006; accepted 26 October 2006
Nature 409, 681 (2001). from MCYT, and A.J.S. and M.R.F.-S. received support from 10.1126/science.1133064

WNT and DKK Determine the early hair follicle bud (5) (Fig. 1B), where-
as Dkk4, a further functional inhibitor of WNT
signaling (11, 12), shows strong epithelial ex-
Hair Follicle Spacing Through a pression at discrete loci before hair placode
formation (Fig. 1C). Weak expression in the
Reaction-Diffusion Mechanism early hair follicle bud indicates that Dkk4
expression marks the forming follicle (Fig.
Stefanie Sick,1 Stefan Reinker,2* Jens Timmer,2 Thomas Schlake1† 1C). Five LEF/TCF consensus binding motifs
are found within 700 base pairs (bp) upstream
Mathematical reaction-diffusion models have been suggested to describe formation of animal of the transcriptional start site of Dkk4, and
pigmentation patterns and distribution of epidermal appendages. However, the crucial signals and regulation of the promoter by the canonical WNT
in vivo mechanisms are still elusive. Here we identify WNT and its inhibitor DKK as primary signaling pathway was suggested by transfec-
determinants of murine hair follicle spacing, using a combined experimental and computational tion studies (Fig. 1D). Hence, the available
modeling approach. Transgenic DKK overexpression reduces overall appendage density. Moderate data support the role of WNT and DKK as
suppression of endogenous WNT signaling forces follicles to form clusters during an otherwise primary determinants of hair follicle spacing
normal morphogenetic program. These results confirm predictions of a WNT/DKK-specific patterns.
mathematical model and provide in vivo corroboration of the reaction-diffusion mechanism for If WNTs and WNT inhibitor(s) represent the
epidermal appendage formation. two components required by the RD hypothesis,
it should be possible to derive, from a WNT/
he development of regularly arranged RD mechanism are discussed in SOM text 2 DKK-specific RD model (SOM text 3), pre-

T body parts has long fascinated experimen-

tal biologists and theoreticians alike. One
area of long-standing debate has been the for-
and fig. S1. In developing murine skin, mes-
enchymal Dkk1 expression is found adjacent to
dictions about the outcome of experimental
alterations of activating and inhibitory functions.

mation of epidermal appendages such as feathers

and hairs. Theoretical models have provided
seemingly simple solutions to complex develop-
mental processes (1); in order to achieve regular
patterns, the reaction-diffusion (RD) hypothesis
of Alan Turing postulates a pair of activator
and inhibitor with special characteristics (2)
[supporting online material (SOM) text 1].
However, it remains largely unclear whether
such predictions can be substantiated in molec-
ular and mechanistic terms (3). Because canon-
ical WNT signaling is essential for the induction
of hair and feather follicles (4, 5) and forced
stimulation of this pathway is sufficient to induce
supernumerous appendages (6, 7), the pathway
represents an appealing candidate for the primary
signal that dictates follicle distribution. Here we
set out to analyze its role in hair follicle ar-
rangement by verifying predictions of a biolog-
ically adapted RD model.
The WNT pathway is active from the ear-
liest stages of follicular development (5, 8)
(Fig. 1A). Expression of the WNT inhibitor
Dkk1 is directly controlled by secreted WNTs
(9, 10). Further aspects of this pathway and the

1
Fig. 1. WNT signaling and expression of Dkk genes are associated with hair follicle formation.
Max-Planck Institute of Immunobiology, Stuebeweg 51, (A) WNT signaling in mesenchymal cells is associated with developing hair follicles (arrowhead).
79108 Freiburg, Germany. 2Institut für Physik, Universität
BATgal mice harboring a WNT-responsive lacZ gene were used as a reporter. (B) Mesenchymal Dkk1
Freiburg, Hermann-Herder-Strasse 3, 79104 Freiburg,
Germany. expression adjacent to epithelial placodes and buds (arrowheads). (C) Strong epithelial Dkk4
expression at discrete loci prior to hair placode formation. Expression rapidly declines after follicle
*Present address: Novartis Institutes for Biomedical Research,
Basel, Switzerland. budding (arrow). (A to C) Scale bars, 100 mm. (D) Reporter gene expression [relative light units
†To whom correspondence should be addressed. E-mail: (RLUs) ± SEM] after endogenous (white) and stimulated (black) canonical WNT signaling. *P < 0.0001
[email protected] (t test) for stimulated WNT signaling (black columns).

www.sciencemag.org SCIENCE VOL 314 1 DECEMBER 2006 1447

REPORTS
Our computational modeling showed that mod- ductive wave increases the interfollicular spacing is still contentious. During a subsequent induc-
erate overexpression of activator during either (Fig. 2A). In line with the prediction of defective tive wave, increased inhibitor expression blocks
the initial or a subsequent inductive wave in- pattern formation after further enhancement of the development of new follicles in the inter-
creases follicular density (Fig. 2, A and B). By inhibitor expression, hair and feather follicle follicular space (Fig. 2B). In addition, excess in-
contrast, strong overexpression of activator com- induction is indeed blocked in the presence of hibitor gives rise to ringlike zones of high
pletely disrupts the patterning process. Moderate strong Dkk1 expression (4, 5); however, the role activator levels around preexisting appendages
overexpression of inhibitor during the initial in- of WNTs in appendage formation and patterning (Fig. 2B). If levels of activator above a threshold

Fig. 2. A WNT/DKK-specific
model of a reaction-diffusion
system predicts changes in ep-
idermal appendage distribution
after transgenic interference
with endogenous signaling. Di-
agrams for excess activator or
inhibitor production correspond
to moderate overexpression
that was restricted to the form-
ing appendages. (A) Modeling
of the first inductive wave. The
calculated distribution of acti-
vator is shown for a 100 × 100
area (arbitrary units). (B) Model-
ing of a subsequent inductive
wave. The first wave is shown in
blue, the second in red. The
calculated distribution of activa-
tor is depicted for a 200 × 200
area. The inset reflects the
difference in activator distribution between second and first inductive wave for a first wave follicle. An activator and inhibitor insensitivity of first wave follicles might
reflect the in vivo situation most closely.

Fig. 3. Suppression of WNT signaling increases

interfollicular spacing in Foxn1::Dkk2 mice. (A)
Schematic of the timing of Foxn1 promoter
activity and the induction of the three major hair
follicle types. (B) E18.5 back skin reveals the
existence of successive inductive waves that can
be distinguished by the developmental stage of
follicles (1, guard; 2 and 3, awl; 4, zigzag). (C)
Levels of Dkk2 expression. TG, Foxn1::Dkk2 trans-
genic mice. (D) Appearance of wild-type and dif-
ferently affected Foxn1::Dkk2 mice. (E) Strongly
affected Foxn1::Dkk2 mice only develop large
guard hair follicles. (F) Densities of guard hair
follicles (black) and of the full complement of
follicles (white) are reduced after WNT signaling is
suppressed. (G) The density of guard hair follicles
is reduced at E14.5, that is, immediately after the
first inductive wave, after WNT signaling is sup-
pressed. Emerging follicles are characterized by
localized WNT signaling, visualized by whole-
mount lacZ staining of skin from BATgal mice of
the indicated genotypes (top views). (B, C, E, and G)
Scale bars, 100 mm.

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REPORTS
determine where appendages form, this suggests test these theoretical predictions (Fig. 3 and Dkk2 mRNA, mice appeared almost hairless; at
the possibility of follicle cluster formation. The SOM text 5 and fig. S2). In mouse, temporally lower levels, the hair coat is present but struc-
ringlike zones of high activator levels are con- well defined successive waves of induction dur- turally abnormal (Fig. 3D). Suppression of hair
verted to discrete spots (Fig. 2B) if preexisting ing embryogenesis and early postnatal life give follicle formation at high levels of Dkk2 expres-
follicles become insensitive to activator and rise to three major hair follicle types (designated sion is consistent with previous results for Dkk1
inhibitor (SOM text 4). Thus, moderate overex- guard, awl, and zigzag) (13) (Fig. 3A). In (5) and conforms to the prediction of our com-
pression of inhibitor by the appendages predicts a compliance with the model, new follicles are putational modeling. However, guard hair follicle
quantitative as well as a qualitative read-out: The initiated in between previously induced append- induction still took place even in severely affected
number of follicles is reduced, and their distri- ages (Fig. 3B). We analyzed four Foxn1::Dkk2 mice (Fig. 3, D and E) because of the delayed
bution is changed. mouse lines in detail, and their phenotype was onset of Foxn1 promoter activation relative to
We used transgenic expression of inhibitor correlated to the level of transgene expression guard hair follicle induction (Fig. 3A and fig. S2).
during hair follicle formation to experimentally (Fig. 3, C and D). At high levels of ectopic The number of follicles per square milli-
meter is reduced by about 30% in adult Foxn1::
Dkk2 (medium) mice (Fig. 3F). Regarding the
first inductive wave that gives rise to guard hair
follicles, a similar reduction is evident (Fig. 3F).
In Foxn1::Dkk2 (strong) mice, which only
develop guard hair follicles, the density is
further reduced (Fig. 3F). The emergence of
hair follicle buds is accompanied by localized
WNT signaling in mesenchymal cells of the
prospective dermal papilla (Fig. 1A). As ex-
pected, the density of these cell clusters is re-
duced in Foxn1::Dkk2 transgenic mice (Fig. 3G).
However, we could not detect any difference
between mice with high and medium mRNA
levels. Thus, the significant decrease of guard
hair follicle density in Foxn1::Dkk2 (strong) as
compared with Foxn1::Dkk2 (medium) mice
suggests the existence of more than one inductive
wave for guard hair follicles; apparently, all but
the first wave are blocked by strong transgene
expression. For the first inductive wave, the
results confirm the predictions of our model with
respect to the quantitative effects of inhibitor
overexpression.
As suggested by our simulations, WNT and
DKK proteins may control the natural increase
of follicle density during subsequent inductive
waves (Fig. 2B and SOM text 6 and figs. S3
and S4). In order to investigate inhibitor effects
on these waves, we next examined transgenic
mice with an abnormal hair coat in more detail.
These mice are characterized by a misdistribu-
tion of hair shafts. Whereas emerging hair shafts
are almost evenly distributed over the skin
surface in wild-type mice, bundles of hair shafts
are separated by large areas of interfollicular
epidermis in Foxn1::Dkk2 mice (Fig. 4A). His-
tological sections confirmed that usually more
than three follicles are tightly clustered in trans-
genic mice (Fig. 4B); all follicles within a
cluster gave rise to bona fide hair shafts. By
contrast, follicles are clearly separated from each
other in wild-type skin (Fig. 4B). Strong follicle
clustering was also observed in Foxn1::Dkk1
mice (fig. S5), which corroborated its inde-
pendence of the inhibitor’s identity. A ringlike
Fig. 4. A normal sequence of inductive waves gives rise to hair follicle clusters after moderate pattern of WNT signal-receiving cells around
suppression of WNT signaling. (A) Hair distribution on the back of wild-type and Foxn1::Dkk2 mice. preexisting follicles confirms the model’s pre-
(B) Hair follicle distribution in the back skin of 10-day-old wild-type and Foxn1::Dkk2 animals. (C) diction for activator distribution and indicates an
Aberrant distribution of WNT signal–receiving cells in Foxn1::Dkk2 mice at E17.5. Canonical WNT initiation of cluster formation at about embry-
signaling is visualized by whole-mount lacZ staining of back skin from BATgal mice of the indicated onic day 17.5 (E17.5) (Fig. 4C). Unequivocal
genotypes (top views). (D) New follicles (arrowheads) emerge at E18.5 and P1 in Foxn1::Dkk2 mice. morphological indications of hair follicle cluster
All scale bars, 100 mm. formation were observed at E18.5 and postnatal

www.sciencemag.org SCIENCE VOL 314 1 DECEMBER 2006 1449

REPORTS
day 1 (P1) (Fig. 4D and fig. S6). Thus, hair fol- However, a more sophisticated systems biology 16. K. Kratochwil, J. Galceran, S. Tontsch, W. Roth,
licle clusters in Foxn1::Dkk2 mice form during a approach will be needed in the future to include R. Grosschedl, Genes Dev. 16, 3173 (2002).
17. K. Kratochwil, M. Dull, I. Farinas, J. Galceran,
normal inductive program by misdistribution of the full complexity and dynamics of the WNT R. Grosschedl, Genes Dev. 10, 1382 (1996).
the normal complement of epidermal append- signaling pathway (25) in a model of interfollic- 18. C. Jamora, R. DasGupta, P. Kocieniewski, E. Fuchs, Nature
ages (Fig. 4D and SOM text 7 and fig. S6). ular patterning. In conclusion, our combined 422, 317 (2003).
We note that further signaling pathways are experimental and computer modeling approach 19. L. Grotewold, U. Ruther, EMBO J. 21, 966 (2002).
20. K. Kobielak, H. A. Pasolli, L. Alonso, L. Polak, E. Fuchs,
involved in interfollicular patterning. Studies on presents compelling evidence for WNT signaling J. Cell Biol. 163, 609 (2003).
feather development demonstrated that fibroblast and a reaction-diffusion mechanism as key 21. M. Plikus et al., Am. J. Pathol. 164, 1099 (2004).
growth factors (FGFs) promote follicle forma- determinants of hair follicle spacing patterns. 22. J. Behrens et al., Nature 382, 638 (1996).
tion, whereas BMP and EGF signaling confers 23. T. Mustonen et al., Dev. Biol. 259, 123 (2003).
24. P. Zhou, C. Byrne, J. Jacobs, E. Fuchs, Genes Dev. 9, 700
interfollicular fate (14, 15). Because these path- (1995).
ways appear to be downstream of WNT signal- References and Notes
1. K. Amonlirdviman et al., Science 307, 423 (2005). 25. R. DasGupta, A. Kaykas, R. T. Moon, N. Perrimon, Science
ing and exert feedback control (5, 14, 16–19), 2. A. Turing, Philos. Trans. R. Soc. London B Biol. Sci. 237, 308, 826 (2005).
we propose that they mainly mediate and mod- 37 (1952). 56. We thank B. Hammerschmidt for her excellent technical help;
3. S. Kondo, R. Asai, Nature 376, 765 (1995). B. Kanzler, E. Huber, and J. Wersing for producing transgenic
ulate WNT signals, thereby contributing to the
4. C. H. Chang et al., Mech. Dev. 121, 157 (2004). mice; C. Bleul and T. Boehm for the Foxn1 promoter
stabilization and refinement of the patterning construct; C. Niehrs for murine Dkk1 cDNA; R. Kemler for Lef1
5. T. Andl, S. T. Reddy, T. Gaddapara, S. E. Millar, Dev. Cell
process. Indeed, ablation of the BMP receptor 2, 643 (2002). and b-catenin expression plasmids; and T. Boehm for helpful
IA appears to have no major impact on hair 6. S. Nonchev et al., Development 122, 543 (1996). discussions and comments on the manuscript.
follicle induction and distribution (20). How- 7. U. Gat, R. DasGupta, L. Degenstein, E. Fuchs, Cell 95,
Supporting Online Material
ever, suppression of BMP signaling that may 605 (1998).
www.sciencemag.org/cgi/content/full/1130088/DC1
8. S. Reddy et al., Mech. Dev. 107, 69 (2001).
reduce Dkk1 expression (19) causes an increase 9. A. Niida et al., Oncogene 23, 8520 (2004).
Materials and Methods
in hair follicle density (21), consistent with our SOM Text
10. M. N. Chamorro et al., EMBO J. 24, 73 (2005).
Figs. S1 to S6
simulations (fig. S3C). Given that LEF1 plays 11. V. E. Krupnik et al., Gene 238, 301 (1999).
References
an important role in WNT signaling, which 12. B. Mao, C. Niehrs, Gene 302, 179 (2003).
13. F. W. Dry, J. Genet. 16, 287 (1926). 5 May 2006; accepted 18 October 2006
controls ectodysplasin signals (5, 22), our results 14. H. S. Jung et al., Dev. Biol. 196, 11 (1998). Published online 2 November 2006;
may also explain the misdistribution of follicles 15. R. Atit, R. A. Conlon, L. Niswander, Dev. Cell 4, 231 10.1126/science.1130088
in EdaA1 and Lef1 transgenic mice (23, 24). (2003). Include this information when citing this paper.

Ideal model systems for understanding

Structural Basis for Ribosome IRES RNA-driven translation are the mecha-
nistically streamlined intergenic region (IGR)

Recruitment and Manipulation IRESs of the virus family Dicistroviridae (4).

The IGR IRESs drive the association of the
ribosomal subunits without any of the protein
by a Viral IRES RNA factors that comprise the canonical translation
initiation apparatus (Fig. 1A) (5). Hence, this
Jennifer S. Pfingsten, David A. Costantino, Jeffrey S. Kieft* one structured RNA (molecular size ~ 66 kD)
supplants over 1000 kD of structured ini-
Canonical cap-dependent translation initiation requires a large number of protein factors that act tiation factor proteins, operating as an all-
in a stepwise assembly process. In contrast, internal ribosomal entry sites (IRESs) are cis-acting RNA translation initiation apparatus (6–9).
RNAs that in some cases completely supplant these factors by recruiting and activating the The full-length IGR IRES folds in solution
ribosome using a single structured RNA. Here we present the crystal structures of the ribosome- into two structurally independent domains
binding domain from a Dicistroviridae intergenic region IRES at 3.1 angstrom resolution, providing (10–13). The larger domain (regions 1 and
a view of the prefolded architecture of an all-RNA translation initiation apparatus. Docking of the 2, Fig. 1A and fig. S1) is the ribosome-
structure into cryo–electron microscopy reconstructions of an IRES-ribosome complex suggests a binding domain. It folds into a compact
model for ribosome manipulation by a dynamic IRES RNA. structure (10) that binds directly to the 40S
subunit (10, 12, 13). Cryo–electron micros-
n eukaryotes, there are two known mecha- mRNA 5′ end. Rather, structured RNA se- copy (cryo-EM) reconstructions of an IGR

I nisms for the initiation of protein synthesis

(Fig. 1A). The canonical mechanism re-
quires a modified nucleotide cap on the 5′ end
quences called internal ribosomal entry sites
(IRESs) recruit and activate the translation
machinery, functionally replacing many pro-
IRES bound to the ribosome reveal that the
IGR IRES binds over the mRNA-binding
groove, making contact to and changing the
of the mRNA, which is recognized by an tein factors (2). IRESs are essential for in- structure of both ribosomal subunits (40S
initiation factor protein (eIF4E). This protein fection by many medically and economically and 60S) (14). However, these cryo-EM struc-
recruits other factors that assemble the ribo- important viruses such as hepatitis C (HCV), tures do not reveal the structure of the IRES,
some on the mRNA in a stepwise process (1). hepatitis A, polio, foot-and-mouth disease, rhi- how the IRES structure creates a ribosome-
In contrast, internal initiation of translation novirus, coxsackievirus-B3, and HIV-1 (3). binding site, or which IRES structural fea-
does not require a cap or recognition of the IRESs also drive the translation of eukaryotic tures specifically contact and manipulate the
mRNAs, encoding factors involved in develop- ribosome.
ment, growth regulation, apoptosis, transcrip- To address these questions and develop a
Department of Biochemistry and Molecular Genetics, University
of Colorado at Denver and Health Sciences Center, Mail Stop tion, translation, and other important cellular model for the structural basis of IGR IRES-
8101, Post Office Box 6511, Aurora, CO 80045, USA. processes (3). The molecular rules underlying driven translation, we have solved the struc-
*To whom correspondence should be addressed. E-mail: this RNA structure–driven mechanism remain ture of the ribosome-binding domain of the
[email protected] elusive. Plautia stali intestine virus (PSIV) IGR IRES

1450 1 DECEMBER 2006 VOL 314 SCIENCE www.sciencemag.org

REPORTS
RNA using x-ray crystallography to a resolu- these bases are kept invariant to maintain the nearly a right angle to P2.2 and adjacent to
tion of 3.1 Å with Rwork = 25.3% and Rfree = specific intramolecular contacts that knit the SL IV (Fig. 2C). Classic pseudoknot folding
29.4% (Fig. 1B). In the structure, regions structure together. is characterized by two stacked helices and
1 and 2 pack side-by-side with stem-loop IV The two stem-loops are positioned to- two single-stranded loops; the 5′-most loop
(SL IV) and SL V emerging from the same gether to contact a relatively small area on crosses the major groove, whereas the other
side of the structure (Fig. 1B). Both stem- the ribosome by a set of specific intra- crosses the minor groove (15). Hence, in the
loops have been shown through mutagenesis molecular contacts involving pseudoknot III IGR IRES, this classic pseudoknot archi-
and footprinting experiments to make func- (PK III) and helix P2.2, which form a tecture is maintained, despite the fact that
tionally critical direct contact to the 40S continuous helical stack that extends into PK both loops contain a large amount of em-
subunit (10, 12, 13), and isolated region 2 II (Fig. 1B). The placement of SL IV is fa- bedded structured RNA (fig. S3).
has been shown to bind to the 40S subunit cilitated by underwinding of helix P2.2, which Region 1 does not participate directly in
(12). Thus, the position of SL IV and SL V opens the normally deep and narrow major the interactions that create the 40S subunit–
identifies the side of the structure that is groove (Fig. 2A). The degree of underwind- binding site; rather, it packs against region 2
the 40S subunit–binding interface (“top” of ing is such that a single turn of helix covers a in position to contact the 60S subunit (vide
the structure in Fig. 1B) and indicates that total rise of ~ 42 Å, as compared to ~34 Å for infra). Regions 1 and 2 pack in an interaction
the 40S subunit recognition surface prefolds canonical A-form RNA. This feature is in- in which the large L1.2 loop (green, Fig. 2D)
before subunit binding. The foundation of duced by four key bases that stack into the cradles one strand of helix P2.2 (red, Fig.
this prefolded IRES architecture is helix P2.2 helix, forming two noncanonical base pairs 2D). One strand of this loop (L1.2A) lies
(red, Fig. 1B), which is contacted by bases and anchoring the 3′ end of SL IV (J2.3, along the minor groove of P2.2, forming
from multiple IRES elements (J2.3, J2.2, Fig. 2A). The 5′ end of SL IV is anchored by stabilizing A-minor interactions [for discus-
L1.2A, and L1.2B, cyan and green, Fig. 1B). a G-U wobble pair (G6110-U6082, Fig. 2B) sions of A-minor interactions, see (16)]. The
This folded core corresponds to areas of (12). From these anchors in the P2.2 major other strand of the loop (L1.2B) forms the
strong protection in hydroxyl-radical probing groove, the stem of SL IV is closed with a other half of the cradle; apparently poised to
experiments (fig. S2), providing additional U•U pair, the bases of which are flipped out enter the major groove, it loops back to the
evidence that we have captured the authentic from the P2.2 major groove (U6083-U6096, minor groove to form more stabilizing A-
prefolded, unbound form of the IRES RNA Fig. 2B). Whereas SL IV nestles in the major minor interactions. The tight, complex pack-
(10). Many of the most-conserved bases in groove, SL V is positioned by several con- ing of RNA that is essential for function is
the IGR IRESs cluster in this highly struc- served bases that force the stem of SL V to especially evident in this region, where five
tured core region. Our structure suggests that emerge from the minor groove, placing it at strands of RNA trace in close proximity. The

A B
SL V
Cap- IGR IRES-
dependent: dependent:
3 40S 40S
tRNA
4G 2
SL IV
4E 4A 4B VpG AUG ORF
m7G AUG ORF
Small subunit ORF 5'
recruitment P1.1
P2.3
P2.1
4G
3 40S
tRNA
Region 2 J2.2
4E 2 J2.3 L1.2A
m7G 4A 4B
AUG ORF
PK III
Scanning J2.1
highly flexible
region
3 40S PK III PK II
4G tRNA
4E 2
4A 4B
m7G AUG ORF
P2.2
PK II
Region 1
GTP GDP 60S 3'
60S Factor release
L1.2B (to region 3)
2 3

40S 40S L1.1

4G tRNA VpG AUG ORF P1.2
4E 4A 4B
m7G AUG ORF ORF
60S 60S

Fig. 1. IRES-driven initiation and the structure of the PSIV IGR IRES. contain two functionally critical pseudoknots (10, 12, 13). Because PK III
(A) Ribosome recruitment strategies used in cap-dependent and IGR- is nested inside PK II, this forms an RNA tertiary structure called a
IRES–dependent translation initiation. ORF, open reading frame; GTP, double-nested pseudoknot (15). Figure S1 contains a detailed secondary
guanosine triphosphate; GDP, guanosine diphosphate. At right is the structure with sequence information. (B) Structure of the ribosome-
protein-independent pathway used by this IRES RNA. The inset is a binding domain, colored to match the inset in (A). J, junction; P, paired/
cartoon of the secondary structure of the ribosome-binding domain, helix; L, loop. The gray hexagon shows structure that was weakly visible
colored to match the structure of (B). Parts colored gray did not appear in and hence conformationally flexible. The RNA crystallized in a domain-
the crystal structure and were not built into the final model. The swapped dimer (fig. S4) in which the functionally essential structural
secondary structure consists of two regions (regions 1 and 2), which features are preserved.

www.sciencemag.org SCIENCE VOL 314 1 DECEMBER 2006 1451

REPORTS

A 3' C other parts of regions 1 and 2 (helix P1.1 and

5' loop L1.1) are only weakly visible in the
5'
electron density. We built RNA structure into
G6115
C6114 this weak density, but the structure is poorly
defined in these regions, indicating confor-
P2.2 A6128 mational flexibility. Hence, although we can
SL V
~42 PK III clearly see where L1.1 lies in relation to the
U6097
angstroms rest of the structure, we cannot report its high-
A6105 resolution structure. This flexibility corre-
A6135
G6110 U6082
A6129 sponds to the fact that the RNA crystallized
A6098 U6130 as a domain-swapped dimer in which the
A6131 native interactions between regions 1 and 2
are preserved (for a detailed discussion, see
fig. S4). Hence, the IRES contains regions
3' of stable, highly structured RNA and other
regions of flexible, less stable structure. We
B G6110 D A6046
L1.2B examined the functional significance of this
U6082 A6049 U6045 U6044 observation by combining our structure with
A6050 existing cryo-EM reconstruction data (14).
P2.2 U6083 To identify the specific IGR IRES RNA
U6096 structures that contact the ribosome, we
A6050 docked the crystal structure into published
A6049 5' cryo-EM reconstructions (Fig. 3). To generate
SL IV 3'
A6030 a model for docking, we returned the SL IV
U6097 G6029 PK II and SL V stems to their wild-type lengths
A6135 G6030 [these were changed to induce crystallization
A6033 A6034 (17)]. With the use of the location of the 3′ end
L1.2A P2.2
of the IRES RNA, assignments of region 3
and regions 1 and 2 into the cryo-EM maps
Fig. 2. Structural details of the IRES. (A) Underwound helix P2.2 (red) is shown from the minor (14), footprinting and directed hydroxyl-
groove side. A6105 (red) stacks into the helix, forming a noncanonical A•A N7-amino base pair with radical probing data (12, 18), and the overall
A6098 of J2.3 (cyan). A6135 (red) stacks on A6098 and forms a reverse (parallel) Watson-Crick A-U agreement of the crystal structure with the
pair with conserved base U6097 of J2.3 (cyan). In this and subsequent panels, base pairing is shape of the cryo-EM density, the docking ori-
indicated with double-ended arrows and stacking with a thick dashed line. (B) Within the P2.2 major entation was unambiguous (fig. S5) (12, 14, 18).
groove, both U6097 (cyan) and U6082 (purple) pair with bases in P2.2 (red), whereas U6083 and
Based on the good, but not perfect, match of
U6096 pair with each other at the end of SL IV (cyan). Region 1 bases (green) buttress this structure
the structure to the cryo-EM density, we con-
through A-minor interactions. (C) The U6082-G6110 pairing extrudes U6130 from the helix and
induces a sharp turn in the backbone. Bases A6129 and A6128 stack on the minor groove of the PK clude that the IRES does not need to undergo
III helix, starting the base stacking that extends into SL V. (D) The two strands of L1.2 splay apart, a global structural rearrangement to match our
with L1.2A lying in the minor groove of P2.2, stabilizing the inter-region packing through A-minor structure. Rather, local structural shifts (such
interactions (e.g., A6033 and A6034). Bases U6044 and U6045 of L1.2B continue to stack on the PK as a change in the relative angle of helices or a
II stack, whereas A6046-A6048 form a u-turn–like structure (27), allowing A6049 and A6050 to shift of region 1 relative to region 2) occur
reach the minor groove of P2.2. upon binding. The docking and fit are robust
enough to identify the IRES RNA structural
domains that contact the 40S and 60S subunits
and to suggest a dynamic mechanism of IRES
Fig. 3. Interaction of A B action.
40S
the IGR IRES RNA with Cryo-EM showed that small ribosomal
the ribosome. (A) The protein S5 (rpS5) makes contact with the
PSIV IGR IRES ribo- 60S IGR IRES (14), and our docked structure re-
some affinity domain veals that SL IV and SL V are the features that
structure docked into the 60S 40S
interact with rpS5 (Fig. 3A). These are the
cryo-EM representation only IRES contacts to rpS5, indicating that
of the IRES bound to this is the keystone interaction that drives
the 80S ribosome, with SL V L1.1 40S subunit binding, induces conformational
the 60S ribosome den-
change, and begins the translation initiation
sity computationally re-
moved. The 40S subunit rpS5 rpL1/
process. This observation is supported by data
is in yellow, the cryo-EM H77 showing that mutation of these loops abro-
density of the IRES is in (L1 stalk) gates 40S subunit binding and translation ini-
gray, and the IRES crys- SL IV tiation activity (10, 12, 13). The apical loop
tal structure is colored sequences of SL IV and SL V are conserved
as in Fig. 1. The posi- almost universally among the IGR IRESs,
tions of rpS5 (40S subunit) and SL IV and SL V (IGR IRES) are shown. (B) Detailed view of the interaction of underscoring that their specific interaction
the IGR IRES to the 60S subunit within the 80S ribosome–IRES complex, with the 40S subunit density with rpS5 is critical for function (fig. S6).
computationally removed. The L1 stalk of the 60S subunit contacts IRES loop L1.1 and perhaps P1.1. For The HCV IRES also contacts rpS5, and the
both (A) and (B), the orientation is indicated in the insets. HCV IRES domain that contacts rpS5 also

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REPORTS
changes the conformation of the 40S subunit important contact to the 60S subunit predicts for by the folded core and contacts with rpS5
(19–21), suggesting that this interaction is that mutating L1.1 will block translation ini- and the L1 stalk suggests that other contacts
central to this mode of IRES RNA-driven tiation after 40S subunit binding. To test this may be nonspecific in nature (fig. S6).
translation. prediction, we constructed two mutants and Our structure of the PSIV IGR IRES
In the docked structure, IRES loop L1.1 used them in preinitiation complex assays ana- ribosome-binding domain, combined with a
(and perhaps adjacent helices) is positioned lyzed on sucrose gradients (Fig. 4, A to C). wealth of published biochemical, functional,
to make direct contact with the large subunit’s Both mutants produce IRES–40S subunit com- and low-resolution structural data, suggests a
L1 stalk, which contains rpL1 and ribosomal plexes but fail to progress to 80S ribosomes, mechanistic model for the structural basis of
RNA helix H77 (Fig. 3B) (14). That L1.1 and demonstrating that L1.1 is critical for recruit- IGR IRES-driven initiation that involves pro-
the adjacent P1.1 helix are poorly ordered ment of the 60S subunit. Neither of these grammed regions of stable and flexible struc-
suggests this part of the IRES is not stably mutations globally misfold the RNA (Fig. 4B), ture (Fig. 4D). We propose that P1.1 and L1.1
folded until it interacts with the 60S subunit. which suggests that the effect is due to the loss remain flexible when the IRES binds to the
During translation, the L1 stalk interacts with of the direct L1.1 interaction with the 60S 40S subunit through structured SL IV and
the T loop of E site–bound tRNA (22, 23), subunit. SL V (Fig. 4D, step 1). Support for this idea
which suggests that IGR IRES loop L1.1, The IRES interactions with rpS5 and the comes from a close examination of the IRES
when bound in the 80S ribosome, adopts a L1 stalk are the only intimate contacts to structure docked into the cryo-EM density of
structure that mimics this portion of tRNA regions 1 and 2 in our docked structure. This the 40S subunit–bound IRES. The part of the
structure. Like the 40S subunit–binding stem- does not preclude that other interactions may density that corresponds to L1.1 and P1.1 is
loops, IRES loop L1.1 is highly conserved, occur, especially upon proposed subtle con- weak or missing (14) (red oval, Fig. 4D). In
demonstrating the importance and specificity formational changes to the IRES. However, the 40S-bound form, IRES region 3 overlaps
of its interactions with the L1 stalk (fig. S6) the fact that the conservation of nucleotides in with the P and A sites (14). 60S subunit bind-
(4, 11). Our observation that L1.1 makes an regions 1 and 2 of the IRES is fully accounted ing (Fig. 4D, step 2) results in structural
changes in the IRES and a shift that with-
draws region 3 from the A and P sites, as well
as a change in the structure of the ribosome’s
L1 stalk that is similar to changes associated
with elongation factor binding (14, 24, 25).
These IRES structural changes are explained
by a 60S subunit binding–induced organiza-
tion of L1.1 and perhaps P1.1, evident in the
appearance of additional cryo-EM density
around L1.1 and P1.1 upon 60S subunit
binding (Fig. 4D) (14). Furthermore, the fact
that regions 1 and 2 of the IRES make rel-
atively few inter-region contacts suggests that
the two regions can shift relative to one
another. Change in the L1.1 and P1.1 structure
thus could be linked to PK II and domain 3
through the P2.1 helix. Other ribosome fea-
tures positioned near the IRES may also be
part of this overall mechanism (14). Pestova
et al. have reported that the IGR IRES RNAs
also can recruit 80S ribosomes directly (18)
in what must be a coupled series of events
within the context of the assembled 80S
ribosome.
Although there is great diversity in IRES
structure, combining stable and flexible regions
may be a strategy used by other IRESs. This
structural characteristic is observed in the HCV
Fig. 4. IRES structural changes and a proposed mechanism of ribosome recruitment. (A) Sequences IRES despite a very different overall RNA ar-
of two L1.1/P1.1 mutants. COMP, Watson-Crick complement. (B) Native gel analysis of these chitecture (26). Thus, the structure presented
mutants shows that they do not globally misfold, because they run very close to wild-type (WT) here provides the basis for experiments aimed
RNA. Previously published (10) native gel analysis of mutants, in which SL IV and V were changed at understanding the basic tenets of RNA-based
to GAAA tetraloops (which does not change the fold) and in which PK II was altered (which causes
translation initiation.
global misfolding), is shown for comparison at right. (C) Assembly assays of L1.1/P1.1 mutants
analyzed on a sucrose gradient. The locations of 40S- and 80S-bound IRESs are indicated. CPM,
counts per minute. (D) Interactions with the 40S subunit are shown in yellow, and those with the
60S subunit are shown in blue. The IRES is shown as a space-filling representation inside the References and Notes
corresponding gray cryo-EM density, colored as in previous figures. The folded, unbound IRES binds 1. J. W. B. Hershey, W. C. Merrick, in Translational Control
to the 40S subunit through SL IV and SL V interacting with rpS5, inducing a conformational change of Gene Expression, N. Sonenberg, J. W. B. Hershey,
in the subunit and docking region 3 into the P site (step 1). Subsequent 60S subunit binding M. B. Mathews, Eds. (Cold Spring Harbor Laboratory
Press, Cold Spring Harbor, NY, 2000), pp. 33–88.
induces a series of conformational changes in both the IRES and the ribosome, including a putative
2. C. U. Hellen, P. Sarnow, Genes Dev. 15, 1593
organization of IRES structures L1.1 and P1.1 (step 2). The appearance of additional IRES density in (2001).
the cryo-EM map of the 80S ribosome–bound IRES (indicated with the red oval) supports the 3. S. Bonnal, C. Boutonnet, L. Prado-Lourenco, S. Vagner,
existence of this structural switch. Nucleic Acids Res. 31, 427 (2003).

www.sciencemag.org SCIENCE VOL 314 1 DECEMBER 2006 1453

REPORTS
4. E. Jan, Virus Res. 119, 16 (2005). 17. Materials and methods are available as supporting discussions and advice. We especially thank C. Spahn for
5. T. V. Pestova, C. U. Hellen, Genes Dev. 17, 181 material on Science Online. supplying various cryo-EM density files and R. Batey for
(2003). 18. T. V. Pestova, I. B. Lomakin, C. U. Hellen, EMBO Rep. 5, the iridium (III) hexammine. R. Batey, M. Churchill,
6. J. Sasaki, N. Nakashima, J. Virol. 73, 1219 (1999). 906 (2004). D. Bentley, L. Krushel, and R. Zhao provided a critical
7. J. Sasaki, N. Nakashima, Proc. Natl. Acad. Sci. U.S.A. 97, 19. S. Fukushi et al., J. Biol. Chem. 276, 20824 (2001). reading of this manuscript. This work was supported by a
1512 (2000). 20. C. M. Spahn et al., Science 291, 1959 (2001). grant from NIH and funding from the UC Cancer Center
8. E. Jan et al., Cold Spring Harbor Symp. Quant. Biol. 66, 21. J. S. Kieft, K. Zhou, R. Jubin, J. A. Doudna, RNA 7, 194 in support of the x-ray facility. Structure factors and
285 (2001). (2001). coordinates have been deposited in the Protein Data
9. J. E. Wilson, T. V. Pestova, C. U. Hellen, P. Sarnow, Cell 22. R. K. Agrawal et al., J. Cell Biol. 150, 447 (2000). Bank under accession code 2IL9.
102, 511 (2000). 23. M. M. Yusupov et al., Science 292, 883 (2001). Supporting Online Material
10. D. Costantino, J. S. Kieft, RNA 11, 332 (2005). 24. M. G. Gomez-Lorenzo et al., EMBO J. 19, 2710 (2000).
www.sciencemag.org/cgi/content/full/1133281/DC1
11. Y. Kanamori, N. Nakashima, RNA 7, 266 (2001). 25. R. K. Agrawal, A. B. Heagle, P. Penczek, R. A. Grassucci, Materials and Methods
12. T. Nishiyama et al., Nucleic Acids Res. 31, 2434 J. Frank, Nat. Struct. Biol. 6, 643 (1999).
Figs. S1 to S7
(2003). 26. J. S. Kieft et al., J. Mol. Biol. 292, 513 (1999).
Table S1
13. E. Jan, P. Sarnow, J. Mol. Biol. 324, 889 (2002). 27. F. M. Jucker, A. Pardi, RNA 1, 219 (1995). References
14. C. M. Spahn et al., Cell 118, 465 (2004). 28. We acknowledge the staff at beamline ALS 8.2.1 for
15. C. W. Hilbers, P. J. Michiels, H. A. Heus, Biopolymers 48, assistance, R. Zhao for managing the University of 1 August 2006; accepted 30 October 2006
137 (1998). Colorado (UC) at Denver and Health Sciences Center Published online 23 November 2006;
16. P. Nissen, J. A. Ippolito, N. Ban, P. B. Moore, T. A. Steitz, x-ray facility, D. Farrell for computer administration, and 10.1126/science.1133281
Proc. Natl. Acad. Sci. U.S.A. 98, 4899 (2001). M. Churchill, R. Batey, and A. Ferré-D’Amaré for useful Include this information when citing this paper.

been used to assess the roles of PI(4,5)P2 in

Rapid Chemically Induced Changes of cytoskeleton–plasma membrane adhesion
(8). We fused a truncated Inp54p to FKBP
PtdIns(4,5)P2 Gate KCNQ Ion Channels already tagged with cyan fluorescent protein
(CFP)–FKBP (CF) (9, 10). The CFP label of
the resulting fusion protein CF-Inp54p (CF-
Byung-Chang Suh,1* Takanari Inoue,2* Tobias Meyer,2 Bertil Hille1† Inp) exhibited good cytosolic localization in
NIH3T3 cells. CF-Inp was then cotransfected
To resolve the controversy about messengers regulating KCNQ ion channels during phospholipase along with Lyn11-FRB (LDR), a membrane-
C–mediated suppression of current, we designed translocatable enzymes that quickly alter the anchored FRB (7), and YFP-PH(PLC-d), a yel-
phosphoinositide composition of the plasma membrane after application of a chemical cue. The low fluorescent protein (YFP)–tagged pleckstrin
KCNQ current falls rapidly to zero when phosphatidylinositol 4,5-bisphosphate [PtdIns(4,5)P2 or homology (PH) domain from phospholipase
PI(4,5)P2] is depleted without changing Ca2+, diacylglycerol, or inositol 1,4,5-trisphosphate. C–d1 (PLC-d1) serving as a PI(4,5)P2/IP3
Current rises by 30% when PI(4,5)P2 is overproduced and does not change when biosensor (11, 12). The addition of iRap led to
phosphatidylinositol 3,4,5-trisphosphate is raised. Hence, the depletion of PI(4,5)P2 suffices to the translocation of the fluorescent CF-Inp
suppress current fully, and other second messengers are not needed. Our approach is ideally suited from the cytosol to the plasma membrane and
to study biological signaling networks involving membrane phosphoinositides. a reciprocal translocation of YFP-PH(PLC-d)
from the plasma membrane to the cytosol
(Fig. 1, B and H), demonstrating inducible
hosphoinositide phospholipids are major Can we rule out that increases of the many accumulation of the Inp54p enzyme at the

P signaling molecules of cell membranes.

Many cellular proteins are inhibited when
phosphatidylinositol 4,5-bisphosphate [PI(4,5)P2]
signaling molecules downstream of PLC or
other changes in phosphoinositides are essential
for closing channels instead of or together with
plasma membrane and in situ PI(4,5)P 2
depletion. To analyze the kinetics, we mea-
sured the fluorescence intensities of CF-Inp
is hydrolyzed by phospholipase C (PLC) and the loss of PI(4,5)P2? We used a new method to and YFP-PH(PLC-d) in a cytosolic region
reactivated when phosphatidylinositol 4- deplete plasma membrane–associated PI(4,5)P2 (Fig. 1C), which revealed quick translocation
phosphate [PI(4)P] 5-kinase restores PI(4,5)P2 in living cells within seconds without activat- of both probes [half-time (t1/2) = 12.3 ± 1.2 s
(1, 2). For example, KCNQ K+ channels are ing PLC. Upon addition of a dimerizing drug, and 14.7 ± 2.5 s; n = 12 cells from four
closed by muscarinic-receptor–triggered PLC PI(4,5)P2 was selectively depleted in living cells experiments]. Control experiments with either
activation (3, 4). Closure of KCNQ2/KCNQ3 without the production of diacylglycerol (DAG), a phosphatase-dead mutant of Inp54p [CF-Inp
channels increases the excitability of central inositol 1,4,5-trisphosphate (IP3), or calcium Asp281→Ala281 (D281A)] (13) or a fusion
and peripheral neurons, and hypomorphic signals. protein lacking the phosphatase (CF) showed
mutations in KCNQ subunits underlie famil- The chemical dimerizer strategy uses het- no translocation of YFP-PH(PLC-d), although
ial epilepsies, deafness, and arrhythmias (5). erodimerization of protein domains from iRap did induce translocation of the two CFP
Whether the depletion of PI(4,5)P2 suffices to FK506 binding protein (FKBP) and from constructs to the plasma membrane (Fig. 1, D
close KCNQ channels upon receptor activation mTOR (FRB) by the immunosuppressant to H).
remains controversial because much of the rapamycin (6) or an analog called iRap (7). We then studied whether PI(4,5)P2 deple-
supporting experimental evidence is indirect. This approach has previously permitted us to tion suppresses currents in KCNQ K+ channels
develop a high-speed membrane translocation in human embryonic kidney tsA-201 (tsA)
1
Department of Physiology and Biophysics, University of of Rho guanosine triphosphatases (GTPases) cells. As controls, iRap alone had only minor,
Washington School of Medicine, Seattle, WA 98195, USA. (7). Now we depleted PI(4,5)P2 by inducible reversible effects on current amplitude and no
2
Department of Molecular Pharmacology, Stanford Uni- membrane translocation of Inp54p, a yeast effect on the voltage dependence of activation
versity, Clark Center, 318 Campus Drive, Stanford, CA
94305, USA. inositol polyphosphate 5-phosphatase that of KCNQ current (iRap, –24.2 ± 1.2 mV;
*These authors contributed equally to this work.
specifically cleaves the phosphate at the 5 controls, –23.9 ± 0.7 mV; n = 6 cells). Fur-
†To whom correspondence should be addressed. E-mail: position of PI(4,5)P2 (8) (Fig. 1A). Con- thermore, in cells expressing the full Inp54p
[email protected] stitutively membrane-targeted Inp54p has translocation system together with the M1

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REPORTS
subtype of muscarinic receptors (M1R) and application of iRap to cells expressing The goal of our study was to use an
the red fluorescent protein (RFP)–PH(PLC-d) channels and the full Inp54p translocation enzyme distinct from PLC to determine
probe, only iRap and not the muscarinic agonist system suppressed KCNQ current rapidly, whether depletion of PI(4,5)P2 suffices to
oxotremorine-M (Oxo-M) translocated the fully, and irreversibly (Fig. 2, B and C). turn off KCNQ channels. Therefore, we had
CF-Inp enzyme to the plasma membrane (Fig. Suppression was not seen when the Inp54p to rule out the alternative possibility that
2A). The fast, irreversible translocation of enzyme or the membrane anchor was some essential downstream products of PLC,
CF-Inp reached half-maximal rate at 1.2 ± omitted, although Oxo-M acting via trans- such as IP3, DAG, or calcium transients,
0.1 mM iRap (n = 5 to 7 cells) (fig. S1). Both fected M1Rs was still effective at suppress- might also be generated by membrane trans-
Oxo-M and iRap caused translocation of ing current reversibly. The saturated rate location of Inp54p. In tsA cells where PLC
RFP-PH(PLC-d), reflecting rapid depletion constant for suppression of KCNQ current was blocked with the inhibitor U73122 or
of PI(4,5)P2, by PLC in one case and by the by iRap was the same as for Oxo-M, where IP3 accumulation was prevented by
phosphatase in the other (Fig. 2A). Only equivalent to a 5-s t 1/2, and reached half- overexpressing IP3 5-phosphatase (14), the
the effect of Oxo-M was reversible. We maximal at 0.6 ± 0.1 mM iRap (n = 5 to 9 iRap-induced translocation of CF-Inp, the
were now ready to test our hypothesis. The cells) (Fig. 2D). depletion of PI(4,5)P2 (fig. S2, A and B),
and the suppression of KCNQ current were
unaltered (Fig. 2, E and F). Further, in tsA
Fig. 1. In situ PI(4,5)P2 cells expressing the YFP-labeled C1 domain
hydrolysis induced by iRap of protein kinase C–g, a DAG indicator (15),
heterodimerization in NIH3T3 iRap induced no translocation of the in-
cells. (A) FRB is anchored to dicator, but Oxo-M did (fig. S2C). Hence,
plasma membrane via myris- CF-Inp/iRap does not induce DAG produc-
toylation and palmitoylation tion. Finally, we measured intracellular calci-
modification sequence Lyn11. um using fura-2 as a calcium indicator in tsA
Inp54p is recruited from the cells expressing M1Rs. Application of Oxo-M
cytosol upon addition of iRap, led to a transient calcium rise that was abol-
forming the tripartite complex ished by overexpressing IP3 5-phosphatase
FRB-iRap-FKBP. Membrane (fig. S2D). On the other hand, the application
recruitment of Inp54p rapidly of iRap led to no calcium elevation. Analogous
induces specific dephospho- experiments in NIH3T3 cells also showed
rylation at the 5 position of neither DAG production nor calcium elevation
PI(4,5)P2 [PtdIns(4,5)P 2].
with CF-Inp/iRap.
PtdIns(4)P, PI(4)P. (B, D, and
As a further test of the phosphoinositide
F) Time-series confocal fluores-
cent images of cells expressing hypothesis, we used a similar strategy to in-
LDR, YFP-PH(PLC-d) and either crease PI(4,5)P2 in the plasma membrane with
CF-Inp (B), CF-Inp(D281A) (D), a lipid kinase. We made a translocatable
or CF (F). Images before and construct CF-PIPK by combining most of the
after the 30-s addition of iRap enzyme PI(4)P 5-kinase type I–g (PIPKI-g)
(5 mM). Scale bar, 10 mm. (C, (16) with CFP-FKBP. In NIH3T3 cells and
E, and G) Cytosolic fluores- tsA cells, CF-PIPK fluorescence was largely
cence intensities of CFP (blue) in the cytosol at rest and moved to the plasma
and YFP (yellow) for the cells membrane irreversibly upon addition of iRap
shown in B (C), D (E), or F (G). (fig. S3). In tsA cells also expressing KCNQ
Normalized fluorescence inten- subunits, the KCNQ current was slowly aug-
sities are shown as a function mented after the addition of iRap (exponen-
of time. DF/F0, fluorescence tial time constant t = 138 ± 23 s; n = 4 cells),
change divided by initial fluo- as might be predicted if there is a slow
rescence. (H) Normalized fluo- increase of membrane PI(4,5)P2 (Fig. 3A).
rescence intensity of CFP and The midpoint of channel activation did not
YFP in the cytosol from more change (iRap, –28.5 ± 2.0 mV; controls, –27.3 ±
than four independent experi- 1.8 mV; n = 5 cells). However, current sup-
ments with the same condi-
pression by subsequent addition of Oxo-M
tions as [(B) to (G)] before
was greatly slowed and incomplete after only
(white bars) and after (colored
bars) 30-s iRap treatment. The 20 s of application (Fig. 3B), as if accumu-
black box refers to both the lation and speeded synthesis of PI(4,5)P2
solid orange and solid blue decreased the depletion by PLC. The effects
bars, and the white box refers of the translocated CF-PIPK enzyme were
to the white bars, whether paralleled in cells overexpressing the non-
outlined with orange or blue. translocating full-length original enzyme
Mean values are shown and PIPKI-g (fig. S4). Again, in those cells, the
error bars indicate SEM. current suppression by Oxo-M was slowed and
only partial [compare to similar experiments
with PIPKI-b (17)]; current suppression by the
coexpressed CF-Inp/iRap system was also
slowed but remained nearly complete. A
kinase-dead version of the kinase construct

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REPORTS
Fig. 2. Modulation of KCNQ current induced by CF-Inp and
iRap in tsA cells. (A) Translocation of CF-Inp (blue) and RFP-
PH(PLC-d) (red) upon addition of Oxo-M (10 mM) or iRap (5
mM) to cells cotransfected with M1R. Scale bar, 10 mm.
Asterisks indicate times of the four images shown. (B)
Channel modulation by iRap (5 mM) or Oxo-M (10 mM) in
cells expressing CF constructs with LDR or CF-Inp alone.
Insets show the current waveforms at indicated times a to d;
the dotted line indicates zero current. Vm, membrane voltage
protocol; I−20 mV, KCNQ current recorded at −20 mV. (C)
Current inhibition by iRap and Oxo-M in cells expressing
combinations of LDR and CF constructs. (D) Rate constants of
current inhibition with various iRap concentrations. Inset
shows the rate constants (s−1) for inhibition by iRap (5 mM) or
Oxo-M (10 mM) in CF-Inp–expressing cells (n = 3 to 10 cells).
(E) Effect of Oxo-M and iRap on KCNQ channel in a cell
treated with the PLC inhibitor U73122 (3 mM). (F) Current
inhibition by Oxo-M and iRap in cells treated with U73122 or
overexpressing IP3 5-phosphatase (n = 3 to 7 cells). Error
bars in (C), (D), and (F) indicate SEM.

(Asp253→Ala253, CF-PIPKi) (18) increased Fig. 3. Increase of KCNQ

KCNQ current little and had almost no effect current by activation of
on subsequent suppression by muscarinic ago- PI(4)P 5-kinase. (A) Mod-
nist (Fig. 3, A and B). ulation of KCNQ2/KCNQ3
The ability of translocated CF-PIPK to channels in tsA cells ex-
augment KCNQ current suggests that KCNQ pressing CF-PIPK (top) or
channels are not fully saturated by resting CF-PIPKi (bottom) con-
levels of plasma membrane–associated structs by application of
PI(4,5)P2. We tested this further by taking ad- iRap (5 mM) for 1 min.
vantage of a reported difference in PI(4,5)P2 The dashed line indicates
current amplitude before
affinities for different KCNQ subunit iso-
iRap. Insets show the
forms: Increasing concentrations of short-chain
current waveforms at indi-
PI(4,5)P2 are reportedly needed to activate chan- cated times a to c. (B) Per-
nels expressed from KCNQ3 alone, KCNQ2 cent inhibition by Oxo-M
and KCNQ3 together, and KCNQ2 alone, and time constant of inhi-
respectively (19). When we compared these bition after previous iRap
channel types for responses to CF-PIPK/iRap, translocation of CF-PIPK
KCNQ2 current was augmented by 54 ± 12% or CF-PIPKi. (C) Effects of
(n = 5 cells), KCNQ2/KCNQ3 current by 27 ± CF-PIPK or CF-PIPKi on
8% (n = 10 cells), and KCNQ3 channels by iRap-induced augmentation of current in homomeric KCNQ2, heteromeric KCNQ2/KCNQ3, or homomeric
only 5 ± 2% (n = 5 cells) (Fig. 3C). Aug- KCNQ3 channels (n = 4–10 cells). Error bars in (B) and (C) indicate SEM.
mentation of current after iRap addition must
be due to accumulation of extra PI(4,5)P2. The
experiments of Fig. 3C with KCNQ2/KCNQ3 Triply phosphorylated PI(3,4,5)P3 is a as effectively as PI(4,5)P2 in excised mem-
heteromers suggest, therefore, that the degree of potent second messenger for growth factor brane patches (22). Therefore, we asked
saturation by resting PI(4,5)P2 is about 70%, signaling (21). Roles in ion channel regulation whether PI(3,4,5)P3 affects KCNQ currents
which is in good agreement with preliminary are less studied. Applied PI(3,4,5)P3 restores in intact cells. TsA cells grown in normal
published estimates of 72% (20) and 81% (17). the function of G protein–activated, inward serum-containing medium have some plasma
KCNQ2 channels are even less saturated, and rectifier Kir3.1/3.4 K+ channels (but not membrane–associated PI(3,4,5)P3 as reported
KCNQ3 channels are more saturated. constitutively active Kir2.1 channels) about by the PI(3,4,5)P3 indicator YFP-PH(Akt) (23),

1456 1 DECEMBER 2006 VOL 314 SCIENCE www.sciencemag.org

REPORTS
The phosphoinositide dependence of many
cellular functions is now directly testable.

References and Notes

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J. Gen. Physiol. 123, 663 (2004).
if phosphatidylinositol 3-kinase (PI 3-kinase) (fig. S6). 21. L. C. Cantley, Science 296, 1655 (2002).
activity is lowered by growth in serum-free With intact cells, we have shown that de- 22. T. Rohacs, J. Chen, G. D. Prestwich, D. E. Logothetis,
conditions, by treatment with 0.5 to 1 mM of creases of PI(4,5)P2 quickly turn off current J. Biol. Chem. 274, 36065 (1999).
the inhibitor wortmannin, or by overexpres- in KCNQ2/KCNQ3 channels by more than 23. C. D. Kontos et al., Mol. Cell. Biol. 18, 4131
(1998).
sion of the dominant-negative PI 3-kinase reg- 95% in the complete absence of the cascade 24. K. Hara et al., Proc. Natl. Acad. Sci. U.S.A. 91, 7415
ulatory subunit Dp85 (fig. S5, A to C) (24). of IP3, calcium, and DAG signals normally (1994).
Still, the current and its suppression by Oxo-M generated by the activation of PLC. Con- 25. Q. Hu, A. Klippel, A. J. Muslin, W. J. Fantl, L. T. Williams,
remained normal (fig. S5, B and D) (3). To versely, an increase of PI(4,5)P2 augments Science 268, 100 (1995).
26. K. R. Auger, L. A. Serunian, S. P. Soltoff, P. Libby,
raise PI(3,4,5)P3, we made CF-iSH, a trans- the current, whereas synthesis of PI(3,4,5)P3 L. C. Cantley, Cell 57, 167 (1989).
locatable construct combining CFP-FKBP does not change the amplitude. During the 27. W. D. Heo et al., Science 314, 1458 (2006).
with the inter–Src homology 2 (iSH2) domain activation of CF-Inp, rapid dephospho- 28. Supported by NIH grants NS08174 and AR17803
from p85, which complexes in cells with rylation of the PI(4,5)P2 pool would generate (B.H.) and MH64801 and GM63702 (T.M.). T.I. is a
endogenous PI 3-kinase p110 (Fig. 4A) (25). a bolus of additional PI(4)P. Modeling shows recipient of a fellowship from the Quantitative
Chemical Biology Program. We thank J. York for the
In NIH3T3 cells and tsA cells, the CF-iSH that this dephosphorylation will produce a Inp54p plasmid and advice on construction of
construct translocated rapidly to the plasma large transient elevation of PI(4)P, followed CF-Inp, Alliance for Cellular Signaling for the p85
membrane (t1/2 = 14.4 ± 3.6 s, n = 8 cells) by a maintained plateau of extra PI(4)P. plasmid, T. Martin and Y. Aikawa (University of
upon addition of iRap and induced mem- Nevertheless, our experiments show that this Wisconsin) for the PIPKI-g plasmid and advice on
construction of CF-PIPK, T. Wandless and J. Grimley
brane translocation of YFP-PH(Akt) (t1/2 = elevated PI(4)P is not able to sustain KCNQ for the iRap compound, J. Duman for help with calcium
36.5 ± 7.6 s) (Fig. 4, B and C). Never- current when PI(4,5)P2 is depleted. These measurements, and K. Mackie for advice and
theless, the PI(4,5)P2 indicator YFP-PH(PLC-d) results give clarity to our hypothesis that the discussion. Some images used the University of
was not translocated (Fig. 4D), indicating function of KCNQ channels is dependent on Washington Keck Imaging Center.
that CF-iSH initiates synthesis of PI(3,4,5)P3 plasma membrane–associated PI(4,5)P2. Our
at the plasma membrane with little depletion study also shows the utility of the iRap Supporting Online Material
of PI(4,5)P2. This agrees with reports that translocation strategy for perturbing the lipid www.sciencemag.org/cgi/content/full/1131163/DC1
the amount of PI(3,4,5)P3 made by receptor composition of the plasma membrane. This Materials and Methods
activation of endogenous 3-kinases is only a method is noninvasive, inducible, rapid, and Figs. S1 to S6
References
few percent of the available PI(4,5)P2 (26). specific, a combination not found when using
Adding iRap to tsA cells expressing CF-iSH RNA interference, antibodies, or pharmacol-
12 June 2006; accepted 25 August 2006
and grown in serum-free conditions had little ogy. We have used this strategy in an ac- Published online 21 September 2006;
effect on the amplitude of KCNQ2/KCNQ3 companying paper studying the plasma 10.1126/science.1131163
current (Fig. 4E). It did reduce the sub- membrane targeting of small GTPases (27). Include this information when citing this paper.

www.sciencemag.org SCIENCE VOL 314 1 DECEMBER 2006 1457

REPORTS
motifs. We also confirmed, for the examples of
PI(3,4,5)P3 and PI(4,5)P2 Lipids Rit and KRas, that polybasic targeting motifs
alone can be sufficient for PM targeting (Fig. 1D).
Target Proteins with Polybasic To test whether polybasic clusters are an-
chored to the PM by binding to PI(4,5)P2 (14),

Clusters to the Plasma Membrane we hydrolyzed PM PI(4,5)P2 by rapid targeting

of Inp54p, a 5′ specific PI(4,5)P2 phosphatase
(15), to the PM. This method is based on a
Won Do Heo,1 Takanari Inoue,1 Wei Sun Park,1 Man Lyang Kim,1 Byung Ouk Park,2 PM-localized FK506-binding protein (FKBP12)–
Thomas J. Wandless,1 Tobias Meyer1* rapamycin–binding (FRB) construct and a cyto-
solic Inp54p enzyme conjugated with FKBP12
Many signaling, cytoskeletal, and transport proteins have to be localized to the plasma membrane (CF-Inp) that can be translocated to the PM by
(PM) in order to carry out their function. We surveyed PM-targeting mechanisms by imaging the chemical heterodimerization by using a rapamycin
subcellular localization of 125 fluorescent protein–conjugated Ras, Rab, Arf, and Rho proteins. analog, iRap (16).
Out of 48 proteins that were PM-localized, 37 contained clusters of positively charged amino In experiments where we monitored PI(4,5)P2
acids. To test whether these polybasic clusters bind negatively charged phosphatidylinositol using a yellow fluorescent protein (YFP)–
4,5-bisphosphate [PI(4,5)P2] lipids, we developed a chemical phosphatase activation method to conjugated pleckstrin homology (PH) domain
deplete PM PI(4,5)P2. Unexpectedly, proteins with polybasic clusters dissociated from the PM only from phospholipase Cd (PLCd) (17), PM trans-
when both PI(4,5)P2 and phosphatidylinositol 3,4,5-trisphosphate [PI(3,4,5)P3] were depleted, location of CF-Inp triggered a rapid and near
arguing that both lipid second messengers jointly regulate PM targeting. complete dissociation of the YFP-PLCd-PH do-
main from the PM (Fig. 2A). Despite this marked
mall guanosine triphosphatases (GTPases) without requiring polybasic amino acids (as does reduction in PI(4,5)P2 concentration, only a small

S from the Ras, Rho, Arf, and Rab sub-

families often exert their role at the PM
where they control diverse signaling, cytoskel-
that of HRas) (Fig. 1, A and C) (5, 12). Arf6
lacked a specific targeting motif and was only
localized to the PM in its guanosine triphosphate
fraction of the polybasic-nonlipid tail fragment of
Rit dissociated from the PM (Fig. 2B), which
suggests that PI(4,5)P2 is not alone responsible
etal, and transport processes (1–3). KRas, (GTP)–bound form (fig. S2) (13). The sequence for PM targeting. Parallel experiments suggested
CDC42, and other family members require a homology comparison of PM-localized small that phosphatidylinositol 3,4,5-trisphosphate
cluster of positively charged amino acids for PM GTPases in Fig. 1C shows that closely homolo- [PI(3,4,5)P3] might be involved, because stimu-
localization and activity (2, 4). In vitro studies gous small GTPases can have different targeting lation of cells with platelet-derived growth factor
indicate that the physiological PM binding part-
ner of such polybasic clusters could be phosphati-
dylserine, which has one negative charge, or the
less abundant lipid second messenger PI(4,5)P2,
which has four negative charges (5–7). We took
a genomic survey approach and investigated
PM-targeting mechanisms by confocal imaging
of 125 cyan fluorescent protein (CFP)–tagged
constitutively active small GTPases (8). Expres-
sion in NIH3T3 and HeLa cells showed that 48
small GTPases were fully or partially localized to
the PM (Fig. 1A and fig. S1).
Thirty-seven of these PM-localized small
GTPases had C-terminal polybasic clusters
consisting of four or more Lys or Arg residues
at positions 5 to 20 from the C terminus (Fig. 1B
and fig. S1). Polybasic clusters were found in
three forms: They were present together with
N-terminal myristoylation consensus sequences
(as in Arl4) (9) or with C-terminal prenylation
consensus sequences (as in KRas) (5, 6, 10), or
they lacked lipid modifications (as in Rit) (11).
We called these three combinations polybasic-
myristoyl, polybasic-prenyl, and polybasic-
nonlipid PM-targeting motifs, respectively. A
number of remaining PM-targeted small GTPases
Fig. 1. A survey of the subcellular localization of 125 small GTPases shows that most PM-localized
had a combined prenylation and palmitoylation
small GTPases have targeting motifs with clusters of polybasic amino acids. (A) Confocal images of
consensus sequence that mediated PM targeting the subcellular localization of CFP-conjugated small GTPases in NIH3T3 cells (full set of images in
1
NIH3T3 and HeLa cells in fig. S1). The four main PM-targeting motifs are represented in the image
Department of Molecular Pharmacology, 318 Campus panels. (B) Correlation between PM localization and the presence of lysine residues in a region 5 to
Drive, Clark Building, Stanford University Medical School,
Stanford, CA 94305, USA. 2Division of Applied Life Science
20 amino acids from the C terminus. (C) Phylogenetic tree of 48 small GTPases that were identified
(BK21 Program) and Environmental Biotechnology Na- to be partially or fully localized to the PM. Individual membrane targeting elements are color
tional Core Research Center, Gyeongsang National Uni- coded: red for polybasic clusters and blue, green, and orange for palmitoyl, prenyl, and myristoyl
versity, Jinju 660-701, Korea. consensus sequences, respectively. (D) Twenty-amino-acid-long C-terminal tail fragments of Rit and
*To whom correspondence should be addressed. E-mail: KRas are PM-localized. Lack of PM targeting of a KRas tail fragment without the polybasic region
[email protected] (right). Scale bars, 10 mm.

1458 1 DECEMBER 2006 VOL 314 SCIENCE www.sciencemag.org

REPORTS
(PDGF) led to a small increase in PM localization (LY29) (Fig. 2C; control experiments in fig. S3). the PM (Fig. 2, D and E). The effect of reducing
of the Rit tail and this small effect could be Strikingly, the combined reduction of both either PI(4,5)P2 or PI(3,4,5)P3 concentration
reversed by addition of an inhibitor of phos- PI(4,5)P2 and PI(3,4,5)P3 concentration trig- alone on Rit tail localization was relatively small
phoinositide 3-kinase (PI 3-kinase), LY294002 gered a dissociation of most Rit tail protein from (Fig. 2E).
We also found that both PI(4,5)P2 and
PI(3,4,5)P3 have to be lowered to significantly
dissociate a tail fragment of Rin and a polybasic
effector domain peptide from myristoylated alanine-
rich C kinase substrate (MARCKS) protein
(MARCKS ED) from the PM (Fig. 2, D and E).
This MARCKS ED peptide was included be-
cause it has been extensively used for in vitro
biochemical studies of the interaction between
polybasic amino acids and phosphatidylserine
and PI(4,5)P2 (7, 14, 18, 19). HRas, which has a
prenyl-palmitoyl PM-targeting motif without a
polybasic cluster, did not dissociate from the
PM after depletion of PI(4,5)P2 and PI(3,4,5)P3
(Fig. 2E). Control experiments with the same con-
structs in HeLa cells showed similar results (fig.
S4), and an analysis of the dissociation kinetics
showed that PM dissociation occurs within
minutes after CF-Inp activation and LY29 ad-
dition (fig. S5A). We also verified that PM dis-
sociation of polybasic proteins occurred when we
combined PI(4,5)P2 depletion with addition of
wortmannin, an alternative PI 3-kinase inhibitor,
or with expression of a dominant negative PI 3-
kinase inhibitory construct (20) (figs. S6 and S7).
Fig. 2. Depletion of PI(4,5)P2 and PI(3,4,5)P3 dissociates Rit, Rin, and MARCKS ED with polybasic- Additional control experiments are shown in figs.
nonlipid targeting motifs from the PM. (A) Development of a chemically inducible translocation
S8 to S12, including in vitro lipid-blot assays that
method to deplete PI(4,5)P2 from the inner leaflet of the PM (22, 23). The PI(4,5)P2 biosensor YFP-
showed enhanced affinity of proteins with poly-
PLCd-PH was cotransfected to monitor depletion of PI(4,5)P2. (B) Depletion of PI(4,5)P2 caused
only a small reduction in the PM localization of YFP-conjugated Rit tail. (C) A small, but significant, basic clusters for PI(3,4,5)P3 over PI(4,5)P2.
PDGF receptor–mediated increase in Rit tail PM localization can be reversed by addition of the PI3- These control experiments strengthen the argu-
kinase inhibitor LY29 (before and 9 min after addition of 5 mM iRap). The bar graph shows a ment that PI(4,5)P2 and the less abundant
quantification of the same experiment. The PM dissociation index is the relative ratio of internal PI(3,4,5)P3 both serve as PM anchors for pro-
over PM fluorescence; that is, F1cyt/F1PM * F0PM/F0cyt, with F0 and F1 as the fluorescent intensities teins with polybasic clusters.
before and after PI(4,5)P2 and PI(3,4,5)P3 depletion. (D) Joint reduction in PI(4,5)P2 and PI(3,4,5)P3 Insights into the electrostatic binding mech-
triggered a near-complete dissociation of Rit tail and MARCKS ED from the PM. (E) Quantitative anism between positively charged polybasic
analysis of the CF-Inp and/or LY29-triggered PM dissociation of MARCKS ED, and Rit and Rin tails. clusters and negatively charged polyphospho-
PLCd-PH and HRas tails are shown as controls. The inactive LY29 analog LY30 was used as a control inositides came from a sequence comparison of
(24). Scale bars, 10 mm. nonprenylated PM-targeted small GTPases. Most
of these proteins contain two or three subclusters
of polybasic residues in the C-terminal tail. Each
subcluster spans about four or five amino acids,
and the mean distance between subclusters is
nine amino acids (Fig. 3A). Consistent with a
need for multiple subclusters, the removal of a
flanking subcluster in the Rit tail was sufficient to
abolish PM targeting (Rit tail 199 to 219) (Fig.
3B). This suggests that PM targeting results from
additive binding energy of individual subclusters
that each electrostatically interact with a PI(4,5)P2
or PI(3,4,5)P3 lipid. Selectivity for polyphos-
phoinositides over phosphatidylserine may occur
because of opposing high relative-charge den-
sities of polyphosphoinositides and polybasic
subclusters (7).
Fig. 3. Polybasic subclusters and hydrophobic amino acids are required for PM targeting by polybasic- We then investigated differences between the
nonlipid targeting motifs. (A) Statistical analysis of the relative sequence location of positively charged targeting mechanisms of the three polybasic-
amino acids in nonprenylated small GTPases. (B) Loss of PM targeting of Rit tail fragment after removal nonlipid, polybasic-myristoyl, and polybasic-
of a single subcluster with positively charged amino acids. (C) Identification of a tryptophan residue in prenyl PM-targeting motifs. A distinct feature
the polybasic regions of Rit and Rin that mediates PM over nuclear localization. Full-length small of the polybasic-nonlipid targeting motifs is
GTPases, as well as tail fragment mutants, are shown. (D) Identification of two additional hydrophobic their similarity to nuclear localization sequences.
amino acid residues that contribute to the PM targeting of Rit. Scale bars, 10 mm. We found hydrophobic amino acids to be im-

www.sciencemag.org SCIENCE VOL 314 1 DECEMBER 2006 1459

REPORTS
Fig. 4. Depletion of PI(4,5)P2 and
PI(3,4,5)P3 dissociates proteins with
polybasic-myristoyl and polybasic-
prenyl targeting motifs from the
PM. (A) Aligned sequences of the
N terminus (6 amino acids) and C
terminus (20 amino acids) of six
ARF family members. Confocal im-
ages of an Arl7 mutant construct
lacking the N-terminal myristoyla-
tion motif (left), a C-terminal tagged
Arl7 control construct (middle), and
a mutant construct with an internal
CFP tag and a flexible C-terminal
polybasic Arl7 tail (right). (B) PM lo-
calization motifs in geranylgeranylated
Rab35. Confocal images from left to
right: localization of wild-type Rab35,
wild-type tail fragment, wild-type tail
lacking geranylgeranylation motif
(ΔCC), Rab35 tail ΔCC mutant with
KRas caax (Rab35 tail caax), and
Rab35 tail caax mutant lacking poly-
basic amino acids (Rab35 tail caax
ΔpB). (C) Depletion of PI(4,5)P2 by
CF-Inp activation without L29 addi-
tion causes only a minimal PM dissociation of Arl7, KRas tail fragment, and Rab35. (D) Quantitative analysis of the much larger PM dissociation of
polybasic-lipid modified proteins after depletion of PI(4,5)P2 and PI(3,4,5)P3 by CF-Inp activation and addition of LY29. Scale bars, 10 mm.

portant for selective PM localization, because Rab35 (fourth panel, Fig. 4B). We also confirmed these lipid second messengers function as
site-directed mutagenesis of a single hydrophobic that the PM targeting of RAB35 requires a signaling hubs in cellular control systems.
amino acid (Trp204Ala in Rit or Trp202Ala in Rin) polybasic cluster (last panel, Fig. 4B). This
led to a complete loss of PM targeting and a shows that the polybasic-geranylgeranyl motifs References and Notes
1. M. Fivaz, T. Meyer, Neuron 40, 319 (2003).
strong nuclear localization (Fig. 3C). A loss of of Rab35 can be equally effective in PM targeting
2. M. N. Teruel, T. Meyer, Cell 103, 181 (2000).
PM targeting but with less nuclear targeting as the polybasic-farnesyl motif of KRas, which 3. J. L. Guan, Science 303, 773 (2004).
could also be observed for Leu207Ala and supports the notion that both types of prenylation 4. D. Michaelson et al., J. Cell Biol. 152, 111 (2001).
Phe211Ala Rit mutants (Fig. 3D) and for hydro- motifs can be grouped into a single polybasic- 5. K. A. Cadwallader, H. Paterson, S. G. Macdonald,
phobic amino acid mutants of the small GTPases prenyl PM-targeting motif. J. F. Hancock, Mol. Cell. Biol. 14, 4722 (1994).
6. F. Ghomashchi, X. Zhang, L. Liu, M. H. Gelb, Biochemistry
GEM and RAD (fig. S13). Thus, hydrophobic We then tested whether PI(4,5)P 2 and 34, 11910 (1995).
amino acids strengthen PM binding of polybasic- PI(3,4,5)P3 also regulate polybasic-myristoyl and 7. S. McLaughlin, D. Murray, Nature 438, 605 (2005).
nonlipid motifs and prevent the polybasic clus- polybasic-prenyl targeting motifs. As for the 8. W. D. Heo, T. Meyer, Cell 113, 315 (2003).
ter from functioning as a nuclear localization polybasic-nonlipid targeting motif, depletion of 9. A. Schurmann et al., J. Biol. Chem. 269, 15683 (1994).
10. E. Choy et al., Cell 98, 69 (1999).
sequence. PI(4,5)P2 alone triggered only a minor reduction 11. C. H. Lee, N. G. Della, C. E. Chew, D. J. Zack, J. Neurosci.
The polybasic-myristoyl PM-targeting motifs in PM localization of the Arl7 polybasic- 16, 6784 (1996).
have the distinct feature of a separated N-terminal myristoyl targeting motif and the KRas and 12. O. Rocks et al., Science 307, 1746 (2005).
myristoylation consensus sequence and a C- Rab35 polybasic-prenyl motifs (Fig. 4C). Deple- 13. H. Radhakrishna, J. G. Donaldson, J. Cell Biol. 139, 49
(1997).
terminal polybasic cluster. Mutant constructs tion of both PI(4,5)P2 and PI(3,4,5)P3 triggered 14. S. McLaughlin, J. Wang, A. Gambhir, D. Murray, Annu.
showed that effective PM targeting of Arl7 re- significant PM dissociation of all polybasic- Rev. Biophys. Biomol. Struct. 31, 151 (2002).
quired an N-terminal myristoyl motif (left panel, myristoyl and polybasic-prenyl constructs tested 15. T. Inoue, W. D. Heo, J. S. Grimley, T. J. Wandless, T. Meyer,
Fig. 4A), as well as a flexible C-terminal poly- (Fig. 4D) with a kinetics similar to that of Rit Nat. Methods 2, 415 (2005).
16. D. Raucher et al., Cell 100, 221 (2000).
basic tail (middle versus right panel, Fig. 4A), (fig. S5), which suggests that PI(4,5)P2 and
17. T. P. Stauffer, S. Ahn, T. Meyer, Curr. Biol. 8, 343 (1998).
which suggests that the two ends of the protein PI(3,4,5)P3 have the same role for PM localiza- 18. J. Wang, A. Arbuzova, G. Hangyas-Mihalyne, S. McLaughlin,
synergistically support PM targeting. tion for all three types of polybasic PM-targeting J. Biol. Chem. 276, 5012 (2001).
The polybasic-prenyl PM-targeting motif motifs. HRas was again included as a control 19. C. Chapline, K. Ramsay, T. Klauck, S. Jaken, J. Biol. Chem.
includes proteins such as KRas for which a 20– protein without a polybasic cluster. 268, 6858 (1993).
20. S. Poser, S. Impey, K. Trinh, Z. Xia, D. R. Storm, EMBO J.
amino acid tail sequence is sufficient for farnesyla- Our study shows that polybasic PM-targeting 19, 4955 (2000).
tion and PM targeting (Fig. 1D), as well as Rab35 motifs are built from two parts, an unspecific 21. M. C. Seabra, C. Wasmeier, Curr. Opin. Cell Biol. 16, 451
for which an intact GTPase domain is required membrane-targeting part that can be hydropho- (2004).
for geranylgeranylation (21) and for PM target- bic amino acids, myristoyl groups, or prenyl 22. B.-C. Suh, T. Inoue, T. Meyer, B. Hille, Science 314,
1454 (2006); published online 21 September 2006;
ing (second and third panels, Fig. 4B). We groups and a polybasic targeting part that 10.1126/science.1131163.
further compared the roles of farnesylation and provides PM specificity by binding of posi- 23. Correspondence about the iRap inducible enzyme systems
geranylgeranylation by creating a Rab35 mutant tively charged amino acid clusters to negative- should be directed to T. Inoue (e-mail: jctinoue@
with a consensus CAAX farnesylation sequence ly charged PI(4,5)P2 and PI(3,4,5)P3 lipids in stanford.edu).
24. T. W. Poh, S. Pervaiz, Cancer Res. 65, 6264 (2005).
in place of the geranylgeranylation sequence. the PM. This gives PI(4,5)P2 and PI(3,4,5)P3 a 25. We thank A. Salmeen, M. F. Teruel, M. Fivaz, and
This mutant showed PM localization indistin- ubiquitous role in regulating signaling, cyto- other members of the Meyer laboratory for support;
guishable from that of the geranylgeranylated skeletal, and transport proteins and argues that A. R. Koh and S. H. Ryu (POSTECH) for critical reading the

1460 1 DECEMBER 2006 VOL 314 SCIENCE www.sciencemag.org

REPORTS
manuscript; S. McLaughlin (SUNY Stonybrook) and B. Hille This work was supported by grants from National Institute Figs. S1 to S13
(U. Washington) for discussions; and James Whalen for of Mental Health and National Institute of General References
assisting with the in vitro lipid-binding assays. T.I. is a Medical Sciences, NIH, to T.M.
recipient of a fellowship from the Quantitative Chemical 28 August 2006; accepted 11 October 2006
Biology Program. B.O.P. was supported by a grant from Supporting Online Material Published online 9 November 2006;
KOSEF/MOST EB-NCRC (grant no. R15-2003-012-01001-0) www.sciencemag.org/cgi/content/full/1134389/DC1 10.1126/science.1134389
and by scholarships from the Brain Korea 21 program. Materials and Methods Include this information when citing this paper.

A Genome-Wide Association Study didate. In addition to Arg381Gln, nine other

markers in IL23R and in the intergenic region

Identifies IL23R as an Inflammatory

between IL23R and the adjacent IL-12 receptor,
beta-2 gene (IL12RB2), had association P-
values < 0.0001 in the non-Jewish, ileal CD
Bowel Disease Gene case-control cohort (Table 1 and table S1a).
We next tested for association of IL23R
markers in an independent ileal CD case-control
Richard H. Duerr,1,2 Kent D. Taylor,3,4 Steven R. Brant,5,6 John D. Rioux,7,8 Mark S. Silverberg,9 cohort, consisting of 401 patients and 433 con-
Mark J. Daly,8,10 A. Hillary Steinhart,9 Clara Abraham,11 Miguel Regueiro,1 Anne Griffiths,12 trols, all of Jewish ancestry (8). Significant as-
Themistocles Dassopoulos,5 Alain Bitton,13 Huiying Yang,3,4 Stephan Targan,4,14 sociations were observed for several of the same
Lisa Wu Datta,5 Emily O. Kistner,15 L. Philip Schumm,15 Annette T. Lee,16 Peter K. Gregersen,16 markers that were associated in the non-Jewish
M. Michael Barmada,2 Jerome I. Rotter,3,4 Dan L. Nicolae,11,17 Judy H. Cho18* cohort (Table 1 and table S1b). In a combined
analysis of the data from the two ileal CD case-
The inflammatory bowel diseases Crohn’s disease and ulcerative colitis are common, chronic control cohorts (8), nine markers had highly
disorders that cause abdominal pain, diarrhea, and gastrointestinal bleeding. To identify genetic significant association P-values ranging from
factors that might contribute to these disorders, we performed a genome-wide association study. 1.60 × 10−9 to 3.36 × 10−13 (Table 1 and table S1b).
We found a highly significant association between Crohn’s disease and the IL23R gene on We then extended the replication study by
chromosome 1p31, which encodes a subunit of the receptor for the proinflammatory cytokine performing family-based association testing of
interleukin-23. An uncommon coding variant (rs11209026, c.1142G>A, p.Arg381Gln) confers 27 IL23R region markers in an independent co-
strong protection against Crohn’s disease, and additional noncoding IL23R variants are hort of 883 nuclear families in which both par-
independently associated. Replication studies confirmed IL23R associations in independent cohorts
1
of patients with Crohn’s disease or ulcerative colitis. These results and previous studies on the Division of Gastroenterology, Hepatology and Nutrition,
proinflammatory role of IL-23 prioritize this signaling pathway as a therapeutic target in Department of Medicine, School of Medicine, University of
inflammatory bowel disease. Pittsburgh, University of Pittsburgh Medical Center Presbyte-
rian, Mezzanine Level, C-Wing, 200 Lothrop Street, Pittsburgh,
PA 15213, USA. 2Department of Human Genetics, Graduate
rohn’s disease (CD) and ulcerative coli- CD, so we performed a genome-wide associ- School of Public Health, University of Pittsburgh, Crabtree

C tis (UC), the two common forms of idio-

pathic inflammatory bowel disease (IBD),
are chronic, relapsing inflammatory disorders of
ation study testing 308,332 autosomal single
nucleotide polymorphisms (SNPs) on the Illu-
mina HumanHap300 Genotyping BeadChip (8).
A300, 130 Desoto Street, Pittsburgh, PA 15261, USA. 3Medical
Genetics Institute, Cedars-Sinai Medical Center, 8700 Beverly
Boulevard, Los Angeles, CA 90048, USA. 4IBD Center, Division
of Gastroenterology, Cedars-Sinai Medical Center, 8700
Beverly Boulevard, Los Angeles, CA 90048, USA. 5Harvey M.
the gastrointestinal tract. Each has a peak age of Our study population consisted of 567 non-
and Lyn P. Meyerhoff Inflammatory Bowel Disease Center,
onset in the second to fourth decades of life and Jewish, European ancestry patients with ileal CD Department of Medicine, Johns Hopkins University School of
prevalences in European ancestry populations and 571 non-Jewish controls. We initially Medicine, B136, 1503 East Jefferson Street, Baltimore, MD
that average about 100 to 150 per 100,000 (1, 2). focused on ileal CD, the most common location 21231, USA. 6Department of Epidemiology, Bloomberg School
Although the precise etiology of IBD remains to of CD, to minimize pathogenic heterogeneity. of Public Health, Johns Hopkins University, 615 North Wolfe
be elucidated, a widely accepted hypothesis is After exclusion of study subjects with genotype Street, Baltimore, MD 21205, USA. 7Université de Montréal
and the Montreal Heart Institute, S-6400, 5000 Belanger
that ubiquitous, commensal intestinal bacteria completion rates less than 94%, we included 547 Street, Montreal, Quebec H1T 1C8, Canada. 8Medical and
trigger an inappropriate, overactive, and on- cases and 548 controls in subsequent analyses Population Genetics Program, Broad Institute of MIT and
going mucosal immune response that mediates (8). Single-marker allelic tests were performed Harvard, 7 Cambridge Center, Cambridge, MA 02142, USA.
9
intestinal tissue damage in genetically suscepti- using c2 statistics for all autosomal markers. Mount Sinai Hospital IBD Centre, University of Toronto, 441–
600 University Avenue, Toronto, Ontario M5G 1X5, Canada.
ble individuals (1). Genetic factors play an im- Three SNPs had nearly two orders of magnitude 10
Massachusetts General Hospital, Harvard Medical School,
portant role in IBD pathogenesis, as evidenced greater significance compared to the next most 185 Cambridge Street, Boston, MA 02114, USA. 11Department
by the increased rates of IBD in Ashkenazi Jews, significant markers, and they are the only of Medicine, University of Chicago, 5841 South Maryland
familial aggregation of IBD, and increased markers that remain significant at the 0.05 level Avenue, Chicago, IL 60637, USA. 12Department of Pediatrics,
The Hospital for Sick Children, 555 University Avenue, Toronto,
concordance for IBD in monozygotic compared after Bonferroni correction. Two of the three Ontario M5G 1X8, Canada. 13Royal Victoria Hospital, McGill
to dizygotic twin pairs (3). Moreover, genetic markers, rs2066843 (P = 2.86 × 10−9, corrected University Health Centre, 687 Pine Avenue West, Montreal,
analyses have linked IBD to specific genetic P = 8.82 × 10−4) and rs2076756 (P = 5.12 × 10−10, Quebec H3A 1A1, Canada. 14Immunobiology Research
variants, especially CARD15 variants on chro- corrected P = 1.58 × 10−4), are in the known CD Institute, Cedars-Sinai Medical Center, Davis 4063, 8700
Beverly Boulevard, Los Angeles, CA 90048, USA. 15Department
mosome 16q12 and the IBD5 haplotype (span- susceptibility gene, CARD15 (4, 5). The third of Health Studies, University of Chicago, 5841 South Maryland
ning the organic cation transporters, SLC22A4 marker, rs11209026 (P = 5.05 × 10−9, corrected Avenue, Chicago, IL 60637, USA. 16The Feinstein Institute
and SLC22A5, and other genes) on chromosome P = 1.56 × 10−3), is a nonsynonymous SNP for Medical Research, 350 Community Drive, Manhasset, NY
5q31 (3–7). CD and UC are thought to be related (c.1142G>A, p.Arg381Gln) in the IL23R gene 11030, USA. 17Department of Statistics, University of Chicago,
disorders that share some genetic susceptibility (GenBank accession: NM_144701, GeneID: 5734 South University Avenue, Chicago, IL 60637, USA. 18IBD
Center, Section of Digestive Diseases, Departments of Medicine
loci but differ at others. 149233) on chromosome 1p31. This gene en- and Genetics, Yale University, S155A, 300 Cedar Street, New
The replicated associations between CD codes a subunit of the receptor for the proin- Haven, CT 06519, USA.
and variants in CARD15 and the IBD5 haplo- flammatory cytokine, interleukin-23 (IL-23), *To whom correspondence should be addressed. E-mail:
type do not fully explain the genetic risk for and is therefore an intriguing functional can- [email protected]

www.sciencemag.org SCIENCE VOL 314 1 DECEMBER 2006 1461

REPORTS
ents and their IBD (CD, UC, or indeterminate and part of the intergenic region between IL23R allelic frequency of 1.9% in the non-Jewish pa-
IBD)–affected offspring were available for and IL12RB2 have the strongest association tients with ileal CD and 7.0% in non-Jewish
genotyping (Table 2 and table S2) (8). For signals (Fig. 1). There is no significant associa- controls. The glutamine allele appears to protect
Arg381Gln and other IL23R markers, we ob- tion within IL12RB2 (Fig. 1), and we did not against development of CD in both non-Jewish
served significant departure from random allele identify a IL12RB2 SNP in the International [odds ratio (OR) = 0.26, 95% confidence
transmission to CD-affected offspring in both HapMap CEU data that is correlated with an interval (CI) (0.15 to 0.43)] and Jewish [OR =
non-Jewish and Jewish families, providing further IBD-associated, IL23R region variant (8). 0.45, 95% CI (0.27 to 0.73)] case-control
evidence for association between CD and IL23R. The IL23R protein contains an extracellular cohorts. The glutamine allele is also significant-
We also observed distortion of allele transmission domain (composed of a signal sequence, an ly undertransmitted from heterozygous parents
to non-Jewish, UC-affected offspring, providing N-terminal immunoglobulin-like domain, and to non-Jewish and Jewish CD-affected off-
evidence for association of IL23R with non- two cytokine receptor domains), a single transmem- spring, non-Jewish UC-affected offspring, and
Jewish UC. There was no evidence for associa- brane domain, and a 252–amino acid cytoplasmic all IBD-affected offspring (transmitted:non-
tion of Arg381Gln or other IL23R region markers domain (9). Arg-381, in the cytoplasmic domain, transmitted = 45:130, P = 1.32 × 10−10 for the
in the Jewish UC families. In a combined analysis is the fifth amino acid internal to the transmem- IBD phenotype in all 883 families) (Table 2 and
of the data from all 883 nuclear families and both brane domain and is highly conserved between table S2). Our discovery of an uncommon pro-
case-control cohorts (8), all 10 IL23R markers in species (fig. S1). In contrast, two other non- tective allele, or conversely, a very common pre-
Table 2 showed highly significant association synonymous IL23R SNPs, rs1884444 (His3Gln) disposing allele, reflects a major theme in
with IBD, with P-values ranging from 3.55 × 10−9 and rs7530511 (Pro310Leu), which are located complex genetics; namely, that functional genetic
to 6.62 × 10−19. within the extracellular domain, show no evi- variation exerts a continuum of susceptibility,
The IL23R gene is contained within two large dence for disease association (table S1, a and b). neutral, and protective effects. Furthermore, al-
blocks of linkage disequilibrium, and markers in The glutamine allele of Arg381Gln is much leles conferring protection against one disease
the centromeric block containing exons 5 to 11 less common than the arginine allele, with an may result in increased risk for another (10).

Table 1. Non-Jewish and Jewish ileal Crohn's disease (CD) case-control odds ratios (OR) with 95% confidence intervals (CI) are shown for each case-
association study results for IL23R region markers with P-values < 0.0001 in control cohort (8). The ORs shown are for the minor allele. Combined Cochran-
the non-Jewish cohort. Minor allele frequencies (MAF), allelic test P-values, and Mantel-Haenszel P-values are also shown (8). UTR, untranslated region.

Non-Jewish case-control cohort Jewish case-control cohort

Marker Location CD Control CD Control Combined
(n = (n = OR (n = (n = OR P-value
P-value P-value
547) 548) [95% CI] 401) 433) [95% CI]
MAF MAF MAF MAF
rs1004819 Intron 0.374 0.280 3.79 × 10−6 1.53 [1.27,1.84] 0.426 0.334 1.00 × 10−4 1.48 [1.21,1.82] 1.54 × 10−9
rs7517847 Intron 0.331 0.443 1.09 × 10−7 0.62 [0.52,0.74] 0.240 0.352 5.84 × 10−7 0.58 [0.47,0.72] 3.36 × 10−13
rs10489629 Intron 0.378 0.475 4.27 × 10−6 0.67 [0.56,0.80] 0.355 0.465 5.79 × 10−6 0.63 [0.52,0.77] 1.14 × 10−10
rs2201841 Intron 0.385 0.291 4.57 × 10−6 1.52 [1.27,1.83] 0.414 0.315 2.92 × 10−5 1.53 [1.25,1.89] 5.46 × 10−10
rs11465804 Intron 0.020 0.063 7.52 × 10−7 0.30 [0.18,0.51] 0.048 0.096 1.39 × 10−4 0.47 [0.31,0.71] 5.97 × 10−10
rs11209026 Arg381Gln 0.019 0.070 5.05 × 10−9 0.26 [0.15,0.43] 0.033 0.070 7.95 × 10−4 0.45 [0.27,0.73] 3.55 × 10−11
rs1343151 Intron 0.275 0.370 2.26 × 10−6 0.65 [0.54,0.78] 0.229 0.336 1.69 × 10−6 0.59 [0.47,0.73] 1.64 × 10−11
rs10889677 Exon-3'UTR 0.385 0.288 1.82 × 10−6 1.55 [1.29,1.86] 0.419 0.316 1.51 × 10−5 1.56 [1.27,1.91] 9.58 × 10−11
rs11209032 Intergenic 0.393 0.293 1.03 × 10−6 1.56 [1.30,1.87] 0.382 0.298 3.49 × 10−4 1.45 [1.18,1.79] 1.60 × 10−9
rs1495965 Intergenic 0.498 0.412 2.93 × 10−5 1.44 [1.21,1.71] 0.469 0.412 2.04 × 10–2 1.26 [1.03,1.53] 2.55 × 10−6

Table 2. Family-based and combined (case-control and family-based) software package (8). Combined Fisher P-values for all case-control
association results. Family-based association P-values were computed (Table 1) and nuclear family cohorts are also shown (8). UTR, untrans-
using the empirical variance estimator implemented in the FBAT lated region.

Non-Jewish CD Non-Jewish UC Jewish CD Jewish UC All IBD Combined

Marker Location (518 families, (215 families, (77 families, (80 families, (883 families, (family-based
651 affected 251 affected 99 affected 91 affected 1,119 affected and case-control)
offspring) offspring) offspring) offspring) offspring) P-value
P-value P-value P-value P-value P-value
−5 −3 −2 −1
rs1004819 Intron 3.60 × 10 1.20 × 10 1.24 × 10 5.47 × 10 6.06 × 10−8 1.78 × 10−14
rs7517847 Intron 2.30 × 10−5 2.71 × 10−1 3.50 × 10−2 5.00 × 10−1 1.80 × 10−5 9.99 × 10−16
rs10489629 Intron 1.87 × 10−3 2.70 × 10−1 4.33 × 10−1 8.21 × 10−1 1.27 × 10−3 1.62 × 10−11
rs2201841 Intron 5.80 × 10−4 3.21 × 10−4 3.50 × 10−2 5.69 × 10−1 1.04 × 10−7 1.10 × 10−14
rs11465804 Intron 1.32 × 10−4 2.70 × 10−3 8.90 × 10−5 3.71 × 10−1 3.46 × 10−9 3.33 × 10−16
rs11209026 Arg381Gln 8.00 × 10−6 2.97 × 10−4 9.41 × 10−4 4.91 × 10−1 1.32 × 10−10 6.62 × 10−19
rs1343151 Intron 9.63 × 10−2 8.51 × 10−2 3.30 × 10−2 1.89 × 10−1 1.24 × 10−3 2.74 × 10−12
rs10889677 Exon-3'UTR 2.60 × 10−3 3.35 × 10−4 5.88 × 10−2 7.32 × 10−1 1.65 × 10−6 3.40 × 10−14
rs11209032 Intergenic 2.68 × 10−3 3.57 × 10−4 3.48 × 10−2 7.50 × 10−1 2.41 × 10−6 5.50 × 10−13
rs1495965 Intergenic 4.07 × 10−4 1.74 × 10−2 3.93 × 10−2 9.21 × 10−1 1.72 × 10−5 3.55 × 10−9

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In addition to Arg381Gln, we found several Notably, we found no evidence for associa- activities, has produced promising results in a
other variants within the IL23R gene that are tion in our non-Jewish, ileal CD case-control clinical trial of Crohn’s disease (25). It has been
also associated with IBD (Tables 1 and 2 and cohort (table S4) with the IL12RB1 gene, which postulated that specific targeting of the IL23p19/
tables S1 and S2). Marker rs11465804 is an encodes the second subunit of the IL-23 receptor IL23R pathway may be particularly effective in
intronic variant in a nonconserved region and is (9), or the IL23A and IL12B genes, which encode blocking organ-specific inflammation, with less
in significant linkage disequilibrium with the p19 and p40 subunits, respectively, of the compromise of protective responses (26). How-
Arg381Gln (correlation coefficient r 2 = 0.84 in heterodimeric IL-23 cytokine (12). ever, at least one model of murine colitis is
the case-control data) and therefore is unlikely to Previous work with mouse models has docu- worsened in the absence of IL-23, implicating a
confer disease risk independent of the latter. mented a requirement for IL-23 in murine colitis role for IL-23 in the down-regulation of IL-12
However, other markers show evidence for (13), experimental autoimmune encephalitis (27). In addition, IL-23 function may be important
association that appears to be independent of (14), and collagen-induced arthritis (15). IL-23 for proper responses to mycobacterial (28, 29)
Arg381Gln. For example, rs7517847, which has activity is present in the terminal ileum (16) and and intestinal infections (22). In assessing ther-
the most significant association P-value (3.36 × colon (17), and the present study demonstrates apeutic approaches, the strong protective effect
10−13) in the combined analysis of both ileal CD that IL23R variants are associated with both small of the Arg381Gln allele could potentially be ex-
case-control cohorts (Table 1), is not in significant intestinal (ileal CD) and large intestinal (UC) ploited to define desired functional outcomes (10).
linkage disequilibrium with Arg381Gln (r 2 = inflammation. Furthermore, transgenic expres- The contribution of the IL23R pathway to IBD
0.03 in the case-control data). To identify variants sion of IL-23 subunit p19 results in severe sys- will likely involve more than simple gain- or
that are independent of the Arg381Gln signal, we temic inflammation, including in the small and loss-of-function IL23R variants, and therapeutic
performed conditional association testing of the large intestine (18), highlighting this pathway’s interventions will be improved by a better under-
combined case-control data by stratifying on the particular role in promoting strong activation of standing of the context and tissue-specific events
Arg381Gln genotypes. The P-values for these effector T cells and perpetuation of organ-specific associated with functional IL23R polymorphisms.
conditional tests (table S3) demonstrate multiple inflammatory responses. At least part of this ef- References and Notes
residual association signals throughout IL23R, fect is likely mediated via inflammatory, IL-17– 1. D. K. Podolsky, N. Engl. J. Med. 347, 417 (2002).
indicating that there are multiple risk variants in producing T cells (19–23), and elevated IL-17 2. E. V. Loftus Jr., Gastroenterology 126, 1504 (2004).
3. S. Vermeire, P. Rutgeerts, Genes Immun. 6, 637 (2005).
the region. The IL23R gene is expressed as at least levels have been observed in the colonic mucosa 4. J.-P. Hugot et al., Nature 411, 599 (2001).
six alternatively spliced mRNAs, which generate of both CD and UC patients (24). 5. Y. Ogura et al., Nature 411, 603 (2001).
diverse isoforms of the receptor protein (11). The Taken together, these findings suggest that 6. J. D. Rioux et al., Nat. Genet. 29, 223 (2001).
7. V. D. Peltekova et al., Nat. Genet. 36, 471 (2004).
most common splice variants result in the deletion blockade of the IL-23 signaling pathway would 8. Materials and methods are available as supporting
of exons 7 and/or 10. We therefore speculate that be a rational therapeutic strategy for IBD. In material on Science Online.
the multiple genetic association signals detected support of this, a monoclonal antibody directed 9. C. Parham et al., J. Immunol. 168, 5699 (2002).
in the centromeric portion of IL23R (Fig. 1) could against the p40 subunit of the receptor, which 10. J. H. Nadeau, E. J. Topol, Nat. Genet. 38, 1095 (2006).
11. X. Y. Zhang et al., Immunogenetics 57, 934 (2006).
exert their influence via differential splicing. blocks both IL-23 and IL-12 proinflammatory 12. B. Oppmann et al., Immunity 13, 715 (2000).
13. D. Yen et al., J. Clin. Invest. 116, 1310 (2006).
14. D. J. Cua et al., Nature 421, 744 (2003).
15. C. A. Murphy et al., J. Exp. Med. 198, 1951 (2003).
16. C. Becker et al., J. Clin. Invest. 112, 693 (2003).
17. I. J. Fuss et al., Inflamm. Bowel Dis. 12, 9 (2006).
18. M. T. Wiekowski et al., J. Immunol. 166, 7563 (2001).
19. S. Aggarwal, N. Ghilardi, M. H. Xie, F. J. de Sauvage,
A. L. Gurney, J. Biol. Chem. 278, 1910 (2003).
20. C. L. Langrish et al., J. Exp. Med. 201, 233 (2005).
21. E. Bettelli et al., Nature 441, 235 (2006).
22. P. R. Mangan et al., Nature 441, 231 (2006).
23. M. Veldhoen, R. J. Hocking, C. J. Atkins, R. M. Locksley,
B. Stockinger, Immunity 24, 179 (2006).
24. S. Fujino et al., Gut 52, 65 (2003).
25. P. J. Mannon et al., N. Engl. J. Med. 351, 2069 (2004).
26. B. S. McKenzie, R. A. Kastelein, D. J. Cua, Trends
Immunol. 27, 17 (2006).
27. C. Becker et al., J. Immunol. 177, 2760 (2006).
28. F. A. Verreck et al., Proc. Natl. Acad. Sci. U.S.A. 101,
4560 (2004).
29. A. M. Cooper et al., J. Immunol. 168, 1322 (2002).
30. We thank the patients and their families for participating
in the studies. The National Institute of Diabetes and
Digestive and Kidney Diseases IBD Genetics Consortium is
funded by the following grants: DK62431 (S.R.B.),
DK62422 (J.H.C.), DK62420 (R.H.D.), DK62432 (J.D.R.),
DK62423 (M.S.S.), DK62413 (K.D.T.), and DK62429
(J.H.C.). A.H.S. is on the Scientific Advisory Boards of
Shire Pharmaceuticals, Schering (Canada), and Procter &
Gamble Pharmaceuticals.
Fig. 1. Association signals in the IL23R gene region on chromosome 1p31. (A) Genomic locations Supporting Online Material
of genes on chromosome 1p31 between 67,260,000 and 67,580,000 base pairs (Build 35). (B) The www.sciencemag.org/cgi/content/full/1135245/DC1
negative log10 association P-values (Cochran-Mantel-Haenszel chi-square test) from the combined Materials and Methods
Fig. S1
Jewish and non-Jewish case-control cohorts are plotted for genotyped markers in the region. (C) Tables S1 to S4
Pairwise r2 plot for International HapMap CEU data. The intensity of the shading is proportional to References and Notes
r2. The IL23R gene is contained within two blocks of linkage disequilibrium, and the association
18 September 2006; accepted 18 October 2006
signals are strongest in the centromeric block, which contains exons 5 to 11 and extends into the Published online 26 October 2006;
intergenic region between IL23R and IL12RB2. Note that markers in the block encompassing the 10.1126/science.1135245
IL12RB2 gene do not demonstrate significant association. Include this information when citing this paper.

www.sciencemag.org SCIENCE VOL 314 1 DECEMBER 2006 1463

REPORTS
ly, because microfluidic digital PCR yields fluo-
Microfluidic Digital PCR Enables rescent signal upon amplification of a gene
regardless of the number of copies present in
Multigene Analysis of Individual the cell, this approach can yield estimates of the
fraction represented by a given species within

Environmental Bacteria the general microbial community. The number

of rrn operons present in a genome can vary
widely, ranging from 1 [e.g., Rickettsia prowazekii
Elizabeth A. Ottesen,1 Jong Wook Hong,2 Stephen R. Quake,3 Jared R. Leadbetter4* (19)] to 15 [e.g., Clostridium paradoxum (20)],
confounding the interpretation of traditional en-
Gene inventory and metagenomic techniques have allowed rapid exploration of bacterial diversity vironmental gene inventories. Moreover, the use
and the potential physiologies present within microbial communities. However, it remains of single-cell PCR to prepare clone libraries
nontrivial to discover the identities of environmental bacteria carrying two or more genes of avoids complications and PCR artifacts such as
interest. We have used microfluidic digital polymerase chain reaction (PCR) to amplify and analyze amplification biases and unresolvable chimeric
multiple, different genes obtained from single bacterial cells harvested from nature. A gene products (21).
encoding a key enzyme involved in the mutualistic symbiosis occurring between termites and their We used this technique to examine a com-
gut microbiota was used as an experimental hook to discover the previously unknown ribosomal plex, species-rich environment: the lignocellulose-
RNA–based species identity of several symbionts. The ability to systematically identify bacteria decomposing microbial community resident in the
carrying a particular gene and to link any two or more genes of interest to single species residing hindguts of wood-feeding termites. Therein, the
in complex ecosystems opens up new opportunities for research on the environment. bacterial metabolism known as CO2-reductive
homoacetogenesis is one of the major sources
major challenge of environmental sci- situ hybridization (FISH) and variants (13, 14)] of the bacterial fermentation product acetate

A ence is the identification of microbial

species capable of catalyzing important
activities in situ (1). PCR-based techniques that
allow colocalization of sequences through probe
hybridization, but these methods require that
both genes be actively transcribed, that their
(22). Acetogenic bacteria must compete for
hydrogen with Archaea that generate methane,
a potent greenhouse gas for which termites are
use single genes as proxies for organisms or key sequences be known in advance, and that their considered a small yet significant source.
microbial activities continue to provide valuable difference from related, nontarget genes is Because of their high rates of bacterially me-
insights into microbial community diversity sufficient to enable effective probe design and diated homoacetogenesis, many termites con-
(2–4). However, it has been difficult to interre- implementation. Single-cell whole-genome am- tribute less to the global methane budget than
late gene inventories to derive correspondences plification has recently been reported for a highly they might otherwise (23). Additionally, ace-
between any two or more specific genes of in- abundant, culturable marine microbial species, tate serves as the insect host’s major carbon and
terest, or to determine the phylogenetic species but has not yet been shown to be scalable to in- energy source, literally fueling a large propor-
identity of organisms carrying particular genetic terrogating multitudes of diverse, co-resident tion of this mutualistic symbiosis (22, 24, 25).
capabilities. Metagenomic (5) analyses of com- microbes (15). Here, we applied commercially A key gene of the homoacetogenesis pathway
plex communities are dominated by genome available microfluidic devices to perform a encodes formyl-tetrahydrofolate synthetase
“shrapnel”; unless the microbial community is variant of “digital PCR” (16), separating and (FTHFS) (26). Although a diverse inventory
dominated by one or a few species (6, 7), resident interrogating hundreds of individual environ- of termite hindgut community FTHFS variants
genomes are not reliably reconstructed via com- mental bacteria in parallel. already existed (27), the identities of the orga-
putation (8, 9). A gene of interest can be at- Microfluidic devices allow control and ma- nisms dominating homoacetogenesis in ter-
tributed to a specific organism only if it is linked nipulation of small volumes of liquid (17, 18), in mites had remained uncertain. Here, with the
to an unambiguous phylogenetic marker (i.e., on this case allowing for rapid separation and par- use of microfluidics, we discovered the iden-
the same genome fragment) (5, 10). Both PCR titioning of single cells from a complex parent tities of a multitude of FTHFS-encoding orga-
and metagenomic studies are typically carried out sample. Single, partitioned cells served as tem- nisms by determining their specific 16S rRNA
on homogenized, whole-community genomic plates for individual multiplex PCR reactions gene sequences.
DNA preparations. Thus, the cell as a distinct using primers and probes for simultaneous The “clone H group” of FTHFS genotypes
informational entity is almost entirely lost. amplification of both small-subunit ribosomal corresponds to a large fraction of the sequences
Outside of traditional culture-based isolation, RNA (rRNA) and metabolic genes of interest. collected during an inventory of FTHFS genes
few approaches can attribute multiple genes to a Primers and probes with broad target specific- present in the termite hindgut environment (27).
single species or cell type. Microautoradiogra- ities were used, with subsequent resolution of We designed a specific primer set and a
phy (11) and stable isotope probing (12) allow exact gene sequences after successful amplifi- fluorescein-labeled probe capable of on-chip
detection of cells or retrieval of genetic material cation and retrieval. This technique operates in- detection and amplification of the genotypes
from organisms that use a substrate of interest, dependent of gene expression, position on the comprising this FTHFS group. We also re-
but these techniques require active cellular in- genome, or physiological state of the cell at designed broad-specificity “all-bacterial” 16S
corporation of that substrate. Microscopy-based the time of harvest. The result was rapid colo- rRNA gene primers and used a previously pub-
in situ hybridization techniques [fluorescence in calization of two genes (encoding 16S rRNA lished probe (28) to amplify and detect bacterial
and a key metabolic enzyme) to single-genome rRNA genes. Both the all-bacterial 16S rRNA
1 templates, along with the determination of the gene and clone H group FTHFS primer-probe
Division of Biology, California Institute of Technology,
Pasadena, CA 91125, USA. 2Materials Research and Education fraction of cells within the community that en- sets showed single-molecule sensitivity in
Center, Samuel Ginn College of Engineering, Auburn Uni- coded them. Subsequent retrieval of PCR multiplex on-chip reactions using purified plas-
versity, Auburn, AL 36849, USA. 3Department of Bioengineer- products from individual chambers allowed se- mid or termite gut community DNA. The ob-
ing and Howard Hughes Medical Institute, Stanford University, quence analysis of both genes. served success rate for the amplification of
Stanford, CA 94305, USA. 4Environmental Science and
Engineering Program, California Institute of Technology, Phylogenetic analysis of the rRNA gene al- individual genes from single-molecule templates
Pasadena, CA 91125, USA. lows classification of the host bacterium, and the was 48% (fig. S1) (29); thus, the success rate
*To whom correspondence should be addressed. E-mail: metabolic gene is sequenced to confirm that the for coamplification of two genes from single-
[email protected] cell carried the genotype of interest. Additional- molecule templates is estimated to be about 1 in 5.

1464 1 DECEMBER 2006 VOL 314 SCIENCE www.sciencemag.org

REPORTS
Freshly collected termite hindgut luminal con- reamplified, cloned, sequenced, and analyzed
tents were suspended in a PCR reaction buffer using standard methods. Twenty randomly se-
and loaded into a microfluidic device (29). Each lected chambers that had amplified only a 16S
microfluidic panel uses micromechanical valves rRNA gene (and not FTHFS) yielded a diversity
to randomly partition a single PCR mixture into of Endomicrobia, Firmicutes, Bacteroidetes,
1176 independent 6.25-nl reaction chambers Proteobacteria, and Spirochaetes ribotypes, as
(Fig. 1). We considered single-cell separation to be expected on the basis of prior 16S rRNA gene
achieved when fewer than one-third of chambers clone libraries (32) (figs. S2 and S3). Two-thirds
showed rRNA gene amplification. Assuming a of chambers positive for FTHFS genes did not
Poisson distribution of cells, under such conditions amplify 16S rRNA genes when either all-bacterial
6% of chambers should have contained multiple or termite treponeme–specific rRNA gene primers
cells or cell aggregates (30). PCR was carried out were used. This amplification success rate is
on a conventional flat-block thermocycler. comparable to that observed when purified,
Amplification was monitored using 5′ nuclease single-molecule templates were used (e.g., fig.
probes to generate a fluorescent signal detected S1) and remains a target for refinement and
with a modified microarray scanner. improvement in the future.
Multiplex PCR amplifications from single PCR products were retrieved and analyzed
cells or cell aggregates were successfully per- from 28 reaction chambers that coamplified both
formed using diluted gut contents that had been FTHFS and 16S rRNA genes. In 10 of those
partitioned on-chip (Fig. 2, left). We found reactions, sequence analyses revealed that the Fig. 1. Microfluidic digital PCR chip. Top:
global averages of 1.2 (±0.8) × 108 total bacterial FTHFS gene had coamplified with a clade of Schematic diagram showing many parallel
16S rRNA gene encoding units and 1.5 (±1.0) × closely related 16S rRNA gene sequences af- chambers (blue) connected by channels to a
106 total clone H group FTHFS gene encoding filiating with the “termite spirochete cluster” single input. When pneumatic or hydraulic
pressure is applied to the control channel net-
units per Zootermopsis nevadensis termite (31). (33) of the genus Treponema. Members of this
work (red), the membranes between the red
This suggests that, in Z. nevadensis, these par- novel clade were never observed in chambers and blue channels are deflected upward,
ticular FTHFS genes are carried by a minority that lacked FTHFS gene amplification. An creating micromechanical valves. When the
population representing ~1% of gut symbionts. additional three chambers contained a single valves are closed, the continuous blue network
The observed variability of these measurements FTHFS type and multiple 16S rRNA genotypes, is partitioned into independent PCR reactors.
was not surprising, as the Z. nevadensis speci- one of which in each affiliated with the above- Bottom: Schematic showing how a single valve
mens examined were collected from different mentioned group [Zootermopsis environmental connection can be used to partition thousands
colonies and locations and had been maintained genomovar (ZEG) 11.4, 10.2, and 10.1]. These of chambers. In the device used, each exper-
in captivity for varying periods of time. latter reactions also contained two additional imental sample could be partitioned into 1176
Amplification products were retrieved from other Spirochaetes (Zn-FG7A and B in Fig. 3) chambers, and each device contained 12 such
reaction chambers via syringe needle and were in one chamber, a single g-Proteobacterium sample panels.

Bacterial” 16S rDNA ⋅ “Clone H Group” FTHFS

“All Bacterial” “Treponema” 16S rDNA ⋅ “Clone H Group” FTHFS
FTHFS
5 ⋅ 10-3 Dilution
Zootermopsis Hindgut Luminal Contents

5 ⋅ 10-4 Dilution
5 ⋅ 10-5 Dilution

Fig. 2. Multiplex microfluidic digital PCR of single cells in environmental primers and probes. Reaction chambers that contained both genes in 1/500,000
samples. Six panels from a representative experiment show microfluidic digital dilutions from this and other on-chip experiments were sampled and the PCR
PCR on diluted hindgut contents harvested from a single Z. nevadensis products were analyzed (see Fig. 3). Right: The same, except that 16S rRNA
individual. Left: Multiplex PCR using “all-bacterial” 16S rRNA gene (red primers specifically targeted members of the “termite cluster” (33) of the
fluorescence) and “clone H group” (27) FTHFS gene (green fluorescence) spirochetal genus Treponema.

www.sciencemag.org SCIENCE VOL 314 1 DECEMBER 2006 1465

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16s rRNA Genes Formyl-THF Synthetase Genes

ZEG 10.1 ZEG-10.1 FTHFS “Clone H Group”
93
ZEG 10.2 ZEG-10.2
80
ZEG 10.3 ZA-Gut Clone G2
ZEG 10.4 ZEG-10.4
ZEG-13.1 ZEG-10.3
86 ZEG-11.1
ZEG 11.1 ZEG-11.2
95 ZEG 11.2 ZEG-11.4
ZEG 11.3 ZA-Gut Clone I
97
ZEG 11.4 ZA Gut R 82
ZEG 12.1 ZEG-13.1
90
ZEG 12.2 ZEG-11.3
99 ZA-Gut Clone H
ZEG 12.3
ZA-Gut Clone G 94
ZEG 12.4 89
ZA-Gut Clone Z
T. primitia ZAS-2 ZEG 12.3
88 90
ZEG 12.4
90 Clone Zn-FG7A
ZEG 12.1 92
T. primitia ZAS-1 FTHFS “Clone P Group”
ZA-Gut Clone P
Clone Zn-FG7B ZEG 12.2 95
Clone Zn-FG15C T. primitia ZAS-2
97 89
Clone Rs-E64 T. primitia ZAS-1
Clone Zn-G2 ZA-Gut Clone A 100 98
0.01 T. azotonutricium ZAS-9 0.1

Fig. 3. “Clone H group” and “clone P group” FTHFS genes are encoded by of FTHFS genes via lateral gene transfer and can be observed in both
not yet cultivated termite gut treponemes. Left: Phylogenetic tree of 16S isolated species (Treponema primitia ZAS-1) and proposed “environmental
rRNA genes cloned from cultivated strain isolates (orange) and from genomovars” (ZEG 12.2). Scale bars represent substitutions per alignment
hindgut community microbiota. Right: Phylogenetic tree of FTHFS genes position. The trees were constructed using TreePuzzle (39); 630 (16S rDNA)
from the termite hindgut. Dotted lines connect genes believed to originate and 249 (FTHFS) nucleotide positions were used. Citations for all sequences
from the same genome. Incongruent gene phylogenies implicate acquisition are listed in table S1.

sequence (Zn-FG12) in the second, and a vealed that all such 16S rRNA gene sequences 16S rRNA gene primers (Fig. 2, left) and those
Firmicutes sequence (Zn-FG1) in the third. The affiliated within the termite gut treponeme clus- using the clone H primers against purified single-
remaining 15 chambers analyzed (which coam- ter of Spirochaetes. These 16S rRNA genes molecule templates (fig. S1), about one-third of
plified FTHFS and rRNA genes) yielded 16S group into four distinct ribotype clusters (Fig. 3). FTHFS-positive reaction chambers also ampli-
rDNA sequences in proportions that corre- These four sequence types share within-group fied detectable levels of 16S rRNA genes. Trep-
sponded well with the ribotype diversity en- sequence identity of >99% and between-group onemal cells were deduced to constitute 10 to
countered in the general non-FTHFS encoding identities of 95 to 99%. We propose the term 12% of the bacterial community of Z. nevadensis
population. On the basis of this evidence, we “environmental genomovar” (genome variant) (comparing amplification frequencies in the left
conclude that the unique cluster of termite gut to describe not-yet-cultivated organisms shown and right panels of Fig. 3).
treponeme rRNA gene sequences that were to encode two or more known genes of inter- Our results show that specific, not yet cul-
repeatedly identified in FTHFS-containing cham- est. Here, we label the four 16S ribotypes identi- tivated Treponema species encode variants of a
bers represents the ribotype of the FTHFS- fied as ZEG 10 through 13; three genomovars key gene underlying the dominant bacterial
encoding cells. We attribute the instances of (ZEG 10, 11, and 13) encode clone H group metabolism known to affect the energy needs
FTHFS colocalization with other rRNA gene FTHFS sequences, whereas one genomovar of their termite hosts. The microfluidic, mul-
sequences to cell-cell aggregations. The latter is (ZEG 12) encodes a clone P group FTHFS se- tiplex digital PCR approach taken here can be
not to be unexpected in a complex, wood quence. Previously, nine termite gut treponemes extended to expand our understanding of the
particle–filled, sticky environment such as the had been isolated and assigned the strain epithet genetic capacities of not-yet-cultivated species,
termite hindgut (34, 35). Such aggregations ap- ZAS (Zootermopsis acetogenic spirochete) and to collect and collate genetic information in
pear to be largely random, although there may 1 through 9 (37, 38). a manner that builds conceptual genomovars
be a slight enrichment of proteobacterial se- To build additional support for a spirochetal that directly represent the organisms catalyzing
quences relative to the general population (figs. origin of clone H group FTHFS genotypes, we important activities in various environments of
S2 and S3). Our results show that FTHFS se- designed and used a termite treponeme–specific global relevance.
quences present in ~1% of bacterial cells were, 16S rRNA gene primer set and gene probe, with
in 13 of 28 trials, found in association with a the aim of reducing nonspirochetal background References and Notes
16S rRNA sequence type not identified in 20 (Fig. 2, right). The frequency with which clone H 1. N. D. Gray, I. M. Head, Environ. Microbiol. 3, 481
random samplings of the bacterial popula- group FTHFS genes were recovered increased (2001).
tion (16S rRNA–only chambers) at large. The from 1 in 175 cells of the general bacterial pop- 2. E. Zuckerkandl, L. Pauling, J. Theor. Biol. 8, 357 (1965).
3. P. Hugenholtz, B. M. Goebel, N. R. Pace, J. Bacteriol.
probability of a 16S rRNA gene sequence type ulation, to 1 in 16 treponemal cells [several ter- 180, 4765 (1998).
that is present in less than 5% of the population mite gut treponemes are already known or 4. S. J. Sogin, M. L. Sogin, C. R. Woese, J. Mol. Evol. 1, 173
randomly colocalizing with FTHFS in 13 of 28 suspected to encode FTHFS genotypic variants (1971).
trials is low, on the order of 10−10 (36). that would not amplify with the clone H group 5. C. S. Riesenfeld, P. D. Schloss, J. Handelsman, Annu. Rev.
Genet. 38, 525 (2004).
Refined phylogenetic analysis of 16S rRNA FTHFS primer and probe set (27) (fig. S1)]. 6. G. W. Tyson et al., Nature 428, 37 (2004).
gene sequences that were repeatedly isolated Similar to the amplification success rates ob- 7. M. Strous et al., Nature 440, 790 (2006).
from FTHFS-containing reaction chambers reserved in experiments using the “all-bacterial” 8. S. G. Tringe et al., Science 308, 554 (2005).

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9. J. C. Venter et al., Science 304, 66 (2004); published 22. J. A. Breznak, J. M. Switzer, Appl. Environ. Microbiol. 52, 33. T. G. Lilburn, T. M. Schmidt, J. A. Breznak, Environ.
online 4 March 2004 (10.1126/science.1093857). 623 (1986). Microbiol. 1, 331 (1999).
10. O. Béjà et al., Science 289, 1902 (2000). 23. A. Brauman, M. D. Kane, M. Labat, J. A. Breznak, Science 34. J. A. Breznak, H. S. Pankratz, Appl. Environ. Microbiol.
11. J. L. Nielsen, D. Christensen, M. Kloppenborg, P. H. Nielsen, 257, 1384 (1992). 33, 406 (1977).
Environ. Microbiol. 5, 202 (2003). 24. D. A. Odelson, J. A. Breznak, Appl. Environ. Microbiol. 35. J. R. Leadbetter, J. A. Breznak, Appl. Environ. Microbiol.
12. M. Manefield, A. S. Whiteley, R. I. Griffiths, M. J. Bailey, 45, 1602 (1983). 62, 3620 (1996).
Appl. Environ. Microbiol. 68, 2002 (2002). 25. A. Tholen, A. Brune, Environ. Microbiol. 2, 436 (2000). 36. The binomial distribution function was used to calculate the
13. R. I. Amann, W. Ludwig, K. H. Schleifer, Microbiol. Rev. 26. L. G. Ljungdahl, Annu. Rev. Microbiol. 40, 415 (1986). probability that, in 13 of 28 trials, a sequence that is present
59, 143 (1995). 27. T. M. Salmassi, J. R. Leadbetter, Microbiology 149, 2529 in 5% of chambers (0 of 20 16S-only chambers) would
14. K. Zwirglmaier, W. Ludwig, K. H. Schleifer, Mol. Microbiol. (2003). randomly colocalize with clone H group FTHFS sequences.
51, 89 (2004). 28. M. T. Suzuki, L. T. Taylor, E. F. DeLong, Appl. Environ. 37. J. R. Leadbetter, T. M. Schmidt, J. R. Graber, J. A. Breznak,
15. K. Zhang et al., Nat. Biotechnol. 24, 680 (2006). Microbiol. 66, 4605 (2000). Science 283, 686 (1999).
16. B. Vogelstein, K. W. Kinzler, Proc. Natl. Acad. Sci. U.S.A. 29. See supporting material on Science Online. 38. T. G. Lilburn et al., Science 292, 2495 (2001).
96, 9236 (1999). 30. Assuming a Poisson distribution, if 67% of chambers are 39. H. A. Schmidt, K. Strimmer, M. Vingron, A. von Haeseler,
17. T. Thorsen, S. J. Maerkl, S. R. Quake, Science 298, 580 empty, then the expected number of cells per chamber is –ln Bioinformatics 18, 502 (2002).
(2002); published online 26 September 2002 (10.1126/ 0.67 or 0.40. The probability that a chamber contains more 40. We thank M. Unger, A. Daridon, and L. Warren for advice
science.1076996). than one cell is 1 – 0.67 – ((e−0.40)*(0.401))/(1!) or 6.2%. and discussions. Supported by NIH grant 1RO1
18. J. W. Hong, S. R. Quake, Nat. Biotechnol. 21, 1179 31. Value ± 1 standard deviation; 13 termites served as HG002644-01A1, NIH National Research Service Award
(2003). source of cells for n = 32 sample panels. All sample grant 5 T32 GM07616, an NIH Director's Pioneer Award,
19. H. Pang, H. H. Winkler, J. Bacteriol. 175, 3893 (1993). panels that met our conditions for single-cell separation and NSF grant DEB-0321753. S.R.Q. is a founder,
20. F. A. Rainey, N. L. Ward-Rainey, P. H. Janssen, H. Hippe, and contained at least one FTHFS-positive chamber were shareholder, and consultant for Fluidigm Corporation.
E. Stackebrandt, Microbiology 142, 2087 (1996). used in calculation of gut bacterial loads.
21. S. G. Acinas, R. Sarma-Rupavtarm, V. Klepac-Ceraj, 32. M. Ohkuma, T. Kudo, Appl. Environ. Microbiol. 62, 461 16 June 2006; accepted 30 October 2006
M. F. Polz, Appl. Environ. Microbiol. 71, 8966 (2005). (1996). 10.1126/science.1131370

Prevention of Brca1-Mediated development and ductal hyperplasia (20). These

results are consistent with the hypothesis that PRs
play a role in breast carcinogenesis.
Mammary Tumorigenesis in Mice by a To address the specific roles of ER and PRs
in Brca1-mediated tumorigenesis, we studied
Progesterone Antagonist p53 f5&6/f5&6Crec and Brca1 f11/f11p53 f5&6/f5&6Crec
mice (fig. S1A) (21, 22). Inactivation of both
Brca1 and p53 genes in the mouse mammary
Aleksandra Jovanovic Poole,1,2* Ying Li,1,2* Yoon Kim,1,2 Suh-Chin J. Lin,1,2† gland mimics the majority of human BRCA1-
Wen-Hwa Lee,1 Eva Y.-H. P. Lee1,2‡ associated tumors, which also harbor p53 muta-
f11/f11
tions (3, 4). Brca1 p53 f5&6/f5&6Crec mammary
Women with mutations in the breast cancer susceptibility gene BRCA1 are predisposed to breast and glands from nulliparous mice at 2.5 months of age
ovarian cancers. Why the BRCA1 protein suppresses tumor development specifically in ovarian showed about 4.5-fold more branching points
hormone–sensitive tissues remains unclear. We demonstrate that mammary glands of nulliparous compared with wild-type or p53 f5&6/f5&6Crec
Brca1/p53-deficient mice accumulate lateral branches and undergo extensive alveologenesis, a glands (Fig. 1A and fig. S1B). By 4 months of
phenotype that occurs only during pregnancy in wild-type mice. Progesterone receptors, but not age, the nulliparous Brca1 f11/f11p53 f5&6/f5&6Crec
estrogen receptors, are overexpressed in the mutant mammary epithelial cells because of a defect in mammary gland showed further accumulation of
their degradation by the proteasome pathway. Treatment of Brca1/p53-deficient mice with the side branches and extensive alveolar formation
progesterone antagonist mifepristone (RU 486) prevented mammary tumorigenesis. These findings (Fig. 1B). The mammary gland morphology of
reveal a tissue-specific function for the BRCA1 protein and raise the possibility that antiprogesterone mature, nulliparous Brca1 f11/f11p53 f5&6/f5&6Crec
treatment may be useful for breast cancer prevention in individuals with BRCA1 mutations. was similar to that of wild-type pregnant mice,
suggesting that proliferation of mammary epi-
thelial cells (MECs) was altered. Proliferation of
utations in the breast cancer suscep- carcinogenesis by the pleiotropic BRCA1 tumor MECs is regulated by ovarian hormones (23).

M tibility gene BRCA1 are associated

with an increased risk of breast and
ovarian cancers (1). Reduced BRCA1 expres-
suppressor has been attributed to its regulation of
estrogen receptor a (ERa) and two progesterone
receptor isoforms (PRs) (5–8), which play im-
In the estrous phase, MEC proliferation as mea-
sured by 5-bromo-2-deoxyuridine (BrdU) in-
corporation was about five times higher in the
sion due to promoter methylation may con- portant roles in breast development (9–15). Brca1 f11/f11p53 f5&6/f5&6Crec mice than it was
tribute to breast cancer progression (2). The BRCA1 interacts with ER and PRs directly and in wild-type or p53 f5&6/f5&6Crec mice (Fig. 1C
BRCA1 protein has been implicated in DNA modulates ligand-dependent and -independent and fig. S1C). Increased MEC proliferation in
damage repair, cell cycle checkpoint control, and transcription activities of ERa and PR, as well Brca1 f11/f11p53 f5&6/f5&6Crec mice was also seen
transcriptional regulation [reviewed in (3, 4)]. as nongenomic functions of ERa (5–8). How- in the diestrous phase (Fig. 1C and fig. S1C).
The specific suppression of breast and ovarian ever, the mechanisms by which the ER and PRs Previous studies have shown that progesterone
contribute to BRCA1-mediated carcinogenesis exerts its functional effects through paracrine
1 remain unclear. action (24). Indeed, BrdU-positive MECs were
Department of Biological Chemistry, University of Cali-
fornia, Irvine, CA 92697–4037, USA. 2Department of Hormone replacement therapy with profound adjacent to PR-positive cells; there were
Developmental and Cell Biology, University of California, gesterone and estrogen, but not estrogen alone, also BrdU and PR double-positive MECs in the
Irvine, CA 92697–4037, USA. has been associated with an elevation in breast hyperplastic Brca1 f11/f11p53 f5&6/f5&6Crec mam-
*These authors contributed equally to this work. cancer risk in postmenopausal women (16–18). mary gland (fig. S2), indicating that the paracrine
†Current address: Division of Developmental Biology, In mice, the long isoform of PR, PR-B, is action of PR was maintained, at least in most
Cincinnati Children's Hospital Research Foundation, Cin-
cinnati, OH 45229, USA.
required for full development of mammary gland cases.
‡To whom correspondence should be addressed. E-mail: (15, 19), and overexpression of the short isoform, To assess the contribution of circulating
[email protected] PR-A, leads to abnormal mammary gland estrogen and progesterone on MEC proliferation,

www.sciencemag.org SCIENCE VOL 314 1 DECEMBER 2006 1467

REPORTS
we treated ovariectomized mice with progesterone p53 f5&6/f5&6Crec mammary glands, but ducts ly, the number of BrdU-positive MECs in
daily for 3 days, followed by BrdU pulse-labeling containing one or two BrdU-positive cells were Brca1 f11/f11p53 f5&6/f5&6Crec mammary glands
2 weeks after surgery. In vehicle-treated mice, no detected in Brca1 f11/f11p53 f5&6/f5&6Crec mam- increased significantly upon exposure either to
BrdU-positive MECs were found in wild-type and mary glands (Fig. 2A and fig. S3A). Important- estradiol or progesterone alone or to a combina-
tion of both hormones (Fig. 2A and fig. S3A).
The strong mitogenic effect of estradiol and
progesterone on Brca1 f11/f11p53 f5&6/f5&6Crec
MECs prompted us to examine the expression
of ER and PRs by immunostaining. No dif-
ference in ER expression was detected during
diestrous or estrous phase.
We next evaluated PR protein expression. In
diestrous phase, PR was detected in the nuclei of
a scattered subset of epithelial cells in mice of all
genotypes (Fig. 2B and fig. S3B). In estrous
phase, 86.8 ± 4% of Brca1 f11/f11p53 f5&6/f5&6Crec
Fig. 1. Mutation in Brca1/p53 leads to increased mammary ductal branching, alveologenesis, and MECs were PR positive, compared with 27.8 ±
proliferation. (A) Number of branching points in mammary glands of 2.5-month-old wild-type (wt), 3.4% and 28.2 ± 2.4% of MECs in wild-type
p53 f5&6/f5&6Crec, and Brca1 f11/f11p53 f5&6/f5&6Crec mice was determined. The data represent averages of and p53 f5&6/f5&6Crec mammary glands, re-
branch points in five randomly selected areas ± SD. (*P ≤ 0.05) (B) Alveolar development in 4-month- spectively. Elevated expression of PR-A in
old p53 f5&6/f5&6Crec and Brca1 f11/f11p53 f5&6/f5&6Crec mice. Arrows indicate alveoli. (C) Proliferation of Brca1 f11/f11p53 f5&6/f5&6Crec mammary gland
mammary epithelial cells was determined at different estrous phases in wt, p53 f5&6/f5&6Crec, and
in the estrous phase was confirmed by Western
Brca1 f11/f11p53 f5&6/f5&6Crec mice by BrdU incorporation. Histogram shows the average number of BrdU-
blotting (Fig. 2C). Consistent with a previous
labeled cells per duct ± SD (*P ≤ 0.05). At least 15 mammary ducts per animal were evaluated (a
minimum of three mice per genotype). report, only a low amount of PR-B was de-
tected in nulliparous mice (Fig. 2C) (25). The
staining pattern of PR in Brca1+/− MECs was
similar to that of Brca1 f11/f11p53 f5&6/f5&6Crec
MECs (fig. S3B), indicating that Brca1 defi-
ciency correlates with PR accumulation. No
difference in PR transcript amounts was found
between different genotypes (fig. S3C) or in
T47D cells with or without BRCA1 suppres-
sion (fig. S4A). The elevated PR protein quan-
tity in Brca1-deficient MECs was accompanied
by overexpression of the PR target gene Bcl-xl
(fig. S4B). Interestingly, PR expression is in-
creased in normal MECs of breast cancer
patients with a germline mutation of the BRCA1
gene (26). Thus, BRCA1 may regulate PR at
the posttranscriptional level.
To explore this possibility, we established
MEC cultures from the mammary glands of
Brca1 f11/f11p53 f5&6/f5&6Crec and p53 f5&6/f5&6Crec
mice at 2 months of age. Ligand treatment
induced pronounced degradation of PR-A in
p53 f5&6/f5&6Crec MECs (Fig. 2D, lanes 1 and 2)
compared with Brca1 f11/f11p53 f5&6/f5&6Crec
Fig. 2. Mitogenic effect of progesterone on Brca1 f11/f11p53 f5&6/f5&6Crec mammary gland and MECs (Fig. 2D, lanes 5 and 6). PR becomes
stabilization of progesterone receptor in Brca1 f11/f11p53 f5&6/f5&6Crec mammary epithelial cells. (A) polyubiquitinated upon exposure to proges-
Ovariectomized mice (14 to 20 weeks old) were treated with vehicle and 1 mg of E2, 1 mg of P4, or terone and is subsequently degraded by the
E2 and P4 (E2+P4) for 3 days, and BrdU was injected 2 hours before sacrifice. BrdU-positive MECs proteasome (27). Thus, we found that treat-
were detected by immunostaining and quantified in 15 mammary ducts. Average number of BrdU- ment with the proteasome inhibitor, MG132,
positive cells in ovariectomized, E2-, P4-, and E2+P4-treated mice is shown in the histogram (error led to accumulation of PR-A in p53 f5&6/f5&6Crec
bars indicate SE,*P ≤ 0.05). (B and C) Expression of PR protein in wt, p53 f5&6/f5&6Crec, and
MECs (Fig. 2D, lanes 2 and 4) but not in
Brca1 f11/f11p53 f5&6/f5&6Crec mice. Mammary gland tissues harvested at the diestrous or estrous
Brca1 f11/f11p53 f5&6/f5&6Crec MECs (Fig. 2D, lanes
phase were subjected to immunohistochemical staining by using PR antibody (anti-PR). Histogram
represents the average percentage of PR-expressing cells per duct (error bars indicate SD). A 6 and 8). In the presence of cyclohexamide, 90%
minimum of five ducts per animal was evaluated (B). Whole-cell extracts from the mammary gland of PR-A remained in Brca1 f11/f11p53 f5&6/f5&6Crec
tissues as in (B) were used for immunoprecipitation by using PR antibody followed by Western MECs, whereas only 40% of PR-A was detected
blotting analyses (C). Arrow indicates PR-A; arrowhead indicates PR-B. (D) Effect of proteasome in p53 f5&6/f5&6Crec MECs 6 hours after the ad-
inhibitor, MG132, on PR protein quantities in p53 f5&6/f5&6Crec and Brca1 f11/f11p53 f5&6/f5&6Crec dition of the synthetic progesterone R5020
MECs. Cells were starved for 4 hours and then treated with or without MG132 (10 mM) and R5020 (Fig. 2E). These data suggest that Brca1 regulates
(10 nM) for 6 hours as indicated. Western blotting for PR was performed. b-actin served as a PR stability.
loading control. (E) Half-life of PR. MECs were treated with 100 mg/ml cyclohexamide and 10 nM To confirm that BRCA1 exerts a similar
R5020. Cells were harvested at the indicated time points. p53 f5&6/f5&6Crec MECs, open circles; regulatory role in human breast cancer cells, we
Brca1 f11/f11p53 f5&6/f5&6Crec MECs, solid circles. depleted BRCA1 with small interfering RNA

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REPORTS
(siRNA) in T47D cells. This treatment led to a
3.5-fold increase in PR-A protein as well as an
increase in PR-B, but it did not affect the ERa
protein concentration (Fig. 3A, lanes 1 and 2).
Treatment with MG132 stabilized both PR
isoforms (PR-A and PR-B) in control cells but
not in siBRCA1-treated cells (Fig. 3A, lanes
1 and 3 versus 2 and 4). In the absence of
ligand, PR-A and PR-B were stable, and only a
slight increase was detected in siBRCA1-
treated cells (Fig. 3A, lanes 5 and 6). Ligand-
induced phosphorylation of PR-B and PR-A,
as demonstrated by the mobility shift, was
not affected by BRCA1 suppression (Fig. 3A).
Previous studies have shown that phospho-
rylation of Ser294 of PR-B is required for
ligand-dependent proteasome degradation (27).
However, BRCA1 suppression did not decrease Fig. 3. Effect of BRCA1 on PR stability,
PR-B phosphorylation at Ser294 (Fig. 3A). phosphorylation, and polyubiquitination in
We next evaluated ligand-induced poly- human breast epithelial cells. (A) Effect of
ubiquitination of PR in control and BRCA1- BRCA1 inhibition on ERa and PR quantities and
PR phosphorylation. Human breast cancer
depleted normal human MCF10A MECs. PR
T47D cells were infected with SiLuc or SiBRCA1 adenovirus. After being starved overnight, cells were
ubiquitination was reduced in BRCA1-depleted
treated with cycloheximide (80 mg/ml), R5020 (10 nM), and/or MG132 (10 mM) for 6 hours as indicated.
cells. Conversely, overexpression of wild-type Quantities of ERa and PR protein were compared by using Western blotting. Phosphorylation of PR-A and
BRCA1 increased the amount of ubiquitinated PR-B upon R5020 treatment was determined by mobility shift and change of protein mobility upon
PR (Fig. 3B). A protein complex consisting of phosphatase treatment. PR phosphorylation at Ser294 was evaluated with Ser294-phospho-specific
BRCA1 and BARD1 displays E3 ubiquitin antibody. a-tubulin served as a loading control. (B) BRCA1 regulates PR ubiquitination. MCF10A cells
ligase activity (28). To test whether E3 ligase were transiently co-transfected with hemagluttinin (HA)-tagged ubiquitin, PR-A, and BRCA1 constructs (wt
activity of BRCA1 is required for PR poly- or mutants) as indicated, followed by infection with adenovirus expressing siRNA targeting BRCA1 or
ubiquitination, we introduced wild-type, ubiquitin luciferase. Before harvest, cells were treated with 10 mM MG132 for 2 hours, followed by incubation with
ligase–defective ring-domain mutant BRCA1C61G 10 nM R5020 for 2 hours. Anti-PR immunoprecipitates (IP) were analyzed by immunoblotting (IB) by
(28), and ZBRK1 interaction–defective mutant using HA (top) and PR (middle) antibodies. Anti-BRCA1 immunoblotting shows efficiency of siBRCA1
BRCA1Q356R (29, 30) into MCF10A cells. PR-A (bottom). PP1, protein phosphatase 1.

Fig. 4. Antiprogesterone
treatment inhibits mam-
mary tumorigenesis by
decreasing ductal branch-
ing and alveolar pro-
liferation in Brca1 f11/f11
p53 f5&6/f5&6Crec mice.
(A) Nulliparous adult
female Brca1 f 1 1 / f 1 1
p53 f5&6/f5&6Crec mice,
ages 3 to 4 months,
were implanted with ei-
ther a pellet containing
35 mg/60 day constant-
release mifepristone (n =
14) or a placebo pellet (n = 4). Mice were monitored weekly for tumor formation.
(B) Mammary gland branching in control pellet (left) or mifepristone-treated (right)
Brca1 f11/f11p53 f5&6/f5&6Crec mice. Mammary glands were removed 5 weeks after
pellet implantation. (C) Whole mounts of mammary glands from age-matched
Brca1 f11/f11p53 f5&6/f5&6Crec mice without (a and c) or with (b and d) mifepristone
pellet implantation. Boxed areas in a and b were enlarged (c and d, respectively).
Mammary glands were removed 60 days after pellet implantation. Positive staining
with X-galactosidase for LacZ expression marks the cells with an active Cre transgene.

www.sciencemag.org SCIENCE VOL 314 1 DECEMBER 2006 1469

REPORTS
ubiquitination increased in cells overexpressing in Brca1/p53 conditional knockout mice. In 13. S. Mallepell, A. Krust, P. Chambon, C. Brisken, Proc. Natl.
wild type and BRCA1Q356R, but not BRCA1C61G contrast, previous work has shown that treatment Acad. Sci. U.S.A. 103, 2196 (2006).
14. W. P. Bocchinfuso et al., Endocrinology 141, 2982 (2000).
(fig. S5), indicating that E3 ligase activity of of Brca1 f11/f11p53 f5&6/f5&6 MMTV-Cre mice with 15. B. Mulac-Jericevic, J. P. Lydon, F. J. DeMayo,
BRCA1 is required for PR polyubiquitination. the selective estrogen receptor modifier, tamox- O. M. Conneely, Proc. Natl. Acad. Sci. U.S.A. 100, 9744
However, PR failed to be ubiquitinated by ifen, can increase mammary tumor incidence, an (2003).
BRCA1 and its interacting protein, BARD1, as effect attributed to the estrogenic activities of 16. J. E. Rossouw et al., JAMA 288, 321 (2002).
17. V. Beral, E. Banks, G. Reeves, Lancet 360, 942 (2002).
assessed by an in vitro assay (fig. S6). tamoxifen in Brca1-deficient cells (32). 18. S. Lee et al., Int. J. Cancer 118, 1285 (2006).
Because progesterone is a potent mitogen for In a recent study, exposure to a progesterone 19. J. P. Lydon et al., Genes Dev. 9, 2266 (1995).
Brca1 f11/f11p53 f5&6/f5&6Crec MECs, we next pellet was found to dramatically increase mam- 20. G. Shyamala, X. Yang, G. Silberstein, M. H. Barcellos-Hoff,
tested whether blockade of PR activity by a pro- mary gland volume in Brca1 conditional E. Dale, Proc. Natl. Acad. Sci. U.S.A. 95, 696 (1998).
21. X. Xu et al., Nat. Genet. 22, 37 (1999).
gesterone antagonist could prevent or delay mam- knockout mice but had little effect in wild-type
22. S. C. Lin et al., Cancer Res. 64, 3525 (2004).
mary carcinogenesis in Brca1 f11/f11p53 f5&6/f5&6Crec mice (8). Our findings of deregulated PR 23. F. Bresciani, Science 146, 653 (1964).
conditional knockout mice. Mice were treated turnover and mitogenic effect of progesterone 24. C. Brisken et al., Proc. Natl. Acad. Sci. U.S.A. 95, 5076
with a placebo pellet or with a pellet containing in Brca1-deficient MECs are consistent with (1998).
the antiprogesterone mifepristone (RU 486). these results (8). Importantly, the mifepristone- 25. M. D. Aupperlee, K. T. Smith, A. Kariagina, S. Z. Haslam,
Endocrinology 146, 3577 (2005).
The pellet released the drug over a 60-day mediated inhibition of mammary tumorigenesis 26. T. A. King et al., Cancer Res. 64, 5051 (2004).
period, and the mice were monitored weekly for in our Brca1/p53-deficient model provides a 27. C. A. Lange, T. Shen, K. B. Horwitz, Proc. Natl. Acad. Sci.
tumor formation. The median tumor latency of molecular framework for future clinical evalua- U.S.A. 97, 1032 (2000).
Brca1 f11/f11p53 f5&6/f5&6Crec mice was 6.6 months tion of antiprogesterones as a potential chemo- 28. R. Hashizume et al., J. Biol. Chem. 276, 14537
(2001).
(n = 25) with complete penetrance (Fig. 4A). All preventive strategy in women who carry BRCA1 29. L. Zheng et al., Mol. Cell 6, 757 (2000).
the control untreated mice as well as the placebo- mutations. 30. S. Furuta et al., Proc. Natl. Acad. Sci. U.S.A. 102, 9176
treated mice (n = 4) developed palpable tumors (2005).
by 8.7 and 5.2 months of age, respectively. In References and Notes 31. P. Soriano, Nat. Genet. 21, 70 (1999).
1. Y. Miki et al., Science 266, 66 (1994). 32. L. P. Jones et al., Oncogene 24, 3554 (2005).
contrast, no palpable tumors were detected in the 2. M. E. Thompson, R. A. Jensen, P. S. Obermiller, 33. The authors thank Baylor College of Medicine Core Facility
mifepristone-treated mice (n = 14) at 12 months D. L. Page, J. T. Holt, Nat. Genet. 9, 444 (1995). for providing extracts of insect cells overexpressing PR-A and
of age. Five weeks of mifepristone treatment 3. N. S. Ting, W. H. Lee, DNA Repair (Amsterdam) 3, 935 PR-B, C. X. Deng for Brca1 floxed mice, P.-L. Chen for siRNA
substantially reduced branching and suppressed (2004). adenoviruses, S. Furuta and C. Smith for technical advice,
4. N. Turner, A. Tutt, A. Ashworth, Nat. Rev. Cancer 4, 814 and the R. Sprague family for their generosity. This study was
alveologenesis in the mammary glands of (2004). supported by National Cancer Institute (NCI) grant
Brca1 f11/f11p53 f5&6/f5&6Crec mice (Fig. 4B). By 5. S. Fan et al., Science 284, 1354 (1999). CA049649, BCRF-35127 fund, U.S. Department of Defense
using R26R reporter mice to monitor Cre 6. L. Zheng, L. A. Annab, C. A. Afshari, W. H. Lee, T. G. Boyer, grant DAMD17-02-1-0694 (E.L.), and NCI postdoc training
activity (31), we found LacZ-positive normal Proc. Natl. Acad. Sci. U.S.A. 98, 9587 (2001). fellowship (A.J.P.).
7. M. Razandi, A. Pedram, E. M. Rosen, E. R. Levin
Brca1 f11/f11p53 f5&6/f5&6 MECs as well as hy- Mol. Cell. Biol. 24, 5900 (2004).
perplastic foci in the mifepristone-treated mice 8. Y. Ma et al., Mol. Endocrinol. 20, 14 (2006).
Supporting Online Material
www.sciencemag.org/cgi/content/full/314/5804/1467/DC1
(Fig. 4C). These foci did not progress to tumors, 9. S. Nandi, J. Natl. Cancer Inst. 21, 1039 (1958).
Materials and Methods
however. These results suggest that PR function 10. L. Hennighausen, G. W. Robinson, Nat. Rev. Mol. Cell
Figs. S1 to S6
Biol. 6, 715 (2005).
is critical for Brca1-mediated mammary carci- 11. X. Li, D. M. Lonard, B. W. O'Malley, Mech. Ageing Dev.
References
nogenesis and that antiprogesterone treatment 125, 669 (2004). 25 May 2006; accepted 20 October 2006
can prevent or delay mammary carcinogenesis 12. K. S. Korach, Science 266, 1524 (1994). 10.1126/science.1130471

1470 1 DECEMBER 2006 VOL 314 SCIENCE www.sciencemag.org

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Cold Spring Harbor 2007 Meetings & Courses
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Cell Death
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AWARDS

“Bernstein Award” 2007

Young Scientists Research Award in
Computational Neuroscience

The German Federal Ministry of Education and

Research (BMBF) has established the “National
Network for Computational Neuroscience”
with four high-performing “Bernstein Centers
for Computational Neuroscience” as the major
structural elements.

The “Bernstein Award” is equipped with up

meetings and announcements

to 1.25 Mio Euros in the form of a grant over

a period of five years. It will be awarded to a
highly qualified young researcher, considering
the candidates’ verifiable research profile in the
field of Computational Neuroscience and the
scientific concept for a future young research
group. Young researchers can apply for their
own position and group. The group funded by
the “Bernstein Award” will become an integral
part of the National Network for Computational
Neuroscience. Future announcements of the
“Bernstein-Award” are in the scope of the
Ministry’s planning.

The grant is provided for a scientiﬁc project of

a young research group headed by a postdoc
regardless of nationality. The project will be
conducted at a German university or research
institution – within or outside the Bernstein
Centers. It is a prerequisite for funding that
the university or research institution concerned
employs the young researcher during the
funding period and supports him/her with the
basic equipment in terms of laboratory space
and other infrastructure. A statement made to
that effect by the receiving institution must
be included with the project outline to be
submitted.

Deadline for applications is May 31st, 2007.

For more detailed information about the

“Bernstein Award” including application
conditions please visit:

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The Wellcome Trust Scientific Conference Programme is hosted at the dedicated conference facilities on the
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and bioinformatics, including the Wellcome Trust Sanger Institute and the European Bioinformatics Institute.

Winter 2006–07 Humanising Model Genomics of Autumn 2007

Organisms to Understand Common Diseases
Animal Health Research: the Pathogenesis of 8 –10 July Mouse Molecular Genetics
Recent development Human Disease* The availability of whole-genome 5–9 September
and future directions 1– 4 May association studies has started to New to Hinxton, this conference
24 – 26 January The first conference on redefine the genetic architecture brings together researchers of
understanding how the model of genetically complex disorders, multiple disciplines studying various
The first conference focusing on
system works for human disease, and over the next few years will molecular and genetic aspects
livestock diseases in developing
including cardiovascular, metabolic, reveal new susceptibility genes of mammalian development
countries, and the impact of
infectious, neuromuscular, for a wide range of common and disease, primarily using the
those diseases on human health
neurological and cancer diseases. human diseases. mouse model.
and wellbeing.
Molecular Biology of Evolution of
Spring 2007 Summer 2007 Hearing and Deafness Brain and Behaviour
11–14 July 12–16 September
Genomic Disorders Biotechnology: The first conference at Hinxton
20 – 24 March In relation to human addressing the genetics of deafness, Integrated Approaches
This meeting aims to present and and animal health* developmental biology, molecular to Brain Complexity†
basis of sensory function, cochlear
discuss the latest findings relating 13 –17 June damage, repair and regeneration, 26 – 28 September
to the genomic basis of human
This meeting will cover: the use expression analysis, molecular The third conference on
variation, congenital disorders
of molecular tools in biomedical diagnostics, otologic disease and understanding the structural
and acquired diseases.
applications, including animal health approaches to treatments. and functional complexity of the
and breeding, cloning and stem
Microbial Genomes 2007 vertebrate nervous system.
cells; and genetic modification, Interactome Networks†
11–14 April ethics and regulatory issues.
Pharmacogenomics†
The second conference at
29 August– 2 September
Hinxton on genome analysis, Vaccine Development The third genomes to systems 18– 21 October
microbial genome diversity Technologies conference with topics including Focusing on the opportunities of
and evolution, computational ORFeome, binary interaction emerging genomic information and
and functional microbial genomics,
4–7 July maps, DNA/protein networks, technologies to interdisciplinary
and host–pathogen interactions. This series of conferences is aimed assembly, annotation, visualisation, approaches in the study of variable
at accelerating vaccine availability, data integrations and interactome responses of humans to drugs and
particularly for diseases against modelling. toxic agents and how research may
which we do not yet have vaccines benefit the individual.
or for which available vaccines
are inadequate.

* Joint conference with European Science Foundation † Joint conference with Cold Spring Harbor Laboratory

More information and registration details will be Venue

available at www.wellcome.ac.uk/conferences Wellcome Trust Conference Centre, Wellcome Trust Genome
or contact the Wellcome Trust Meetings Programme Campus, Hinxton, Cambridgeshire, CB10 1RQ, UK
team at [email protected] www.wtconference.org.uk

The Wellcome Trust Conference Centre is operated through two companies: Hinxton Hall Limited, a charity registered in England (no. 1048066) and a company registered in England
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AWARDS
UNCF•MERCK SCIENCE INITIATIVE
“Amind is a terrible thing to waste”
U N D E R G R A D U AT E G R A D U AT E POSTDOCTORAL
SCIENCE RESEARCH SCIENCE RESEARCH SCIENCE RESEARCH
SCHOLARSHIP AWARDS DISSERTATION FELLOWSHIPS FELLOWSHIPS
I 15 Awards Annually I 12 Fellowships Annually I 10 Fellowships Annually
I Scholarships up to $25,000 I Fellowship Stipends up to $42,000 I Fellowship Stipends up to $70,000
I Two Summer Internships at I Department Grants of $10,000 I Department Grants of $15,000
a Merck Research Facility I Support for 12-24 months I Support for 12-24 months
An applicant must: An applicant must: An applicant must:
• Be a full-time student at any • Be enrolled full-time in a Ph.D. • Hold a Ph.D. or equivalent degree in
four-year college or university or equivalent doctoral a biomedical life or physical science
• Have junior year academic status program in a biomedical life or • Be appointed as a new or continuing
• Major in a life or physical science physical science postdoctoral fellow by the end of 2007
(first professional degrees excluded) • Be engaged in and within 1-3 years at an academic or non-academic
meetings and announcements

• Have a minimum cumulative GPA of completing dissertation research research institution (private industrial
of 3.3 (4.0 point scale) laboratories are excluded)

Applicants must be African American (Black), U.S. citizens or permanent residents, and attending an
institution in the U.S.A. Applications must be submitted online at www.uncf.org/merck/ or postmarked by
December 15, 2006
For more information, please contact your department chairperson or Jerry L. Bryant, Ph.D., at the United Negro
College Fund, Inc., 8260 Willow Oaks Corporate Drive, P.O. Box 10444, Fairfax, VA 22031-4511, by fax (703)
205-3574, by e-mail at [email protected].

COURSES

DREW UNIVERSITY
RESIDENTIAL SCHOOL ON MEDICINAL CHEMISTRY:
CHEMISTRY AND BIOLOGY IN DRUG DISCOVERY
Madison, New Jersey - June 11-15, 2007

The Residential School on Medicinal Chemistry is a weeklong

graduate level course organized to provide an accelerated
program for medicinal chemists and biologists who wish to
broaden their knowledge of small molecule drug discovery
and development. Attendance is limited to 200 selected
participants with preference given to applicants having five
years or less of drug discovery experience. The School’s aim
is to concentrate on the fundamentals that are useful in drug
discovery spanning initial target validation through clinical
development. Several case histories of recent successful drug
development programs will also be presented.

The five-day program consists of lectures, seminars, case

histories and discussions covering the following topics:
Strategic issues in drug discovery Structure-based drug design
Target validation Drug-like properties
Receptor binding Patents
Enzyme inhibition Plasma protein binding
Ion channels Pharmacokinetics & ADME
High throughput screening Drug metabolism
Hit to lead progression Preclinical toxicology
Lead discovery & modification Clinical development

More information and application forms can be obtained at

www.depts.drew.edu/resmed or by contacting the School’s
office at Drew University, Hall of Sciences, Room 317A,
Madison, NJ 07949, USA; Phone: 973/408-3787;
Fax: 973/408-3504 or E-mail: [email protected]
COURSES

MBL
ou ded 888 as t e a e o og ca abo ato y

2007 Courses
Substantial ﬁnancial assistance is available for many of our courses!

Analytical & Quantitative Light Microscopy Molecular Mycology: Current Approaches

May 9 - May 18 to Fungal Pathogenesis
August 7 - August 23
Biology of Parasitism: Modern Approaches

meetings and announcements

June 7 - August 4 Neural Development & Genetics of Zebraﬁsh
August 9 - August 22
BioMedical Informatics
1st Session: May 27 - June 3 Neural Systems & Behavior
2nd Session: September 23 - September 30 June 9 - August 5

Embryology: Concepts & Techniques Neurobiology

in Modern Developmental Biology June 2 - July 29
June 9 - July 22
Neuroinformatics
Frontiers in Reproduction: Molecular August 11 - August 26
& Cellular Concepts & Applications
May 5 - June 17 Optical Microscopy & Imaging in the
Biomedical Sciences
Pathogenesis of Neuroimmunologic October 9 - October 18
Diseases
August 12 - August 25 Physiology: Cell and Computational Biology
June 9 - July 29
Methods in Computational Neuroscience
July 29 - August 26 Summer Program in Neuroscience, Ethics,
& Survival (SPINES)
Microbial Diversity June 16 - July 14
June 16 - August 2
Workshop on Molecular Evolution
Molecular Biology of Aging July 22 - August 3
July 29- August 18

FOR MORE INFORMATION CONTACT: Admissions Coordinator,

[email protected], (508) 289-7401, MBL, 7 MBL Street, Woods Hole,
MA 02543
Applications are encouraged from women and members of underrepresented minorities.
The MBL is an Equal Opportunity/Afﬁrmative Action Employer.

www.MBL.edu/education
University of Maryland Biotechnology Institute - Shady Grove
Tenure Track Faculty Positions
Over the next several years, up to ten new tenure track faculty positions will be available to outstanding investigators at the University of Maryland Bio-
technology Institute’s Shady Grove campus. An ambitious expansion of the research programs at Shady Grove will build on world class scientiﬁc research
at UMBI’s Center for Advanced Research in Biotechnology (CARB, http://carb.umbi.umd.edu, a partnership with the National Institute of Standards
and Technology (NIST)) and the Center for Biosystems Research (CBR, http://www1.umbi.umd.edu/~cbr). Areas of ongoing research at UMBI-Shady
Grove include chemical biology, mass spectrometry, structural biology, bioinformatics, experimental and computational biophysics, systems modeling,
plant and insect biology, and are supported by a highly collaborative research environment.

The campus has undergone a major, $60M expansion, opening a new, 140,000 ft2 research building. The new facility is being equipped with a state-of-
the-art greenhouse with a plant transformation facility, an insect transformation suite with biosafety level 3 laboratories and animal facilities, genomics
and proteomics laboratories including instrumentation for microarray analysis of gene expression and mass spectrometry for protein identiﬁcation and
analysis, state-of-the-art capabilities in GMP biomanufacturing, and expanded space for the W.M. Keck Structural Biology Laboratory, a leading center
for the study of protein structures using X-ray crystallography and nuclear magnetic resonance (NMR) spectroscopy. UMBI-Shady Grove is located in
Maryland’s thriving biotechnology corridor, with easy access to the NIH, NCI, NIST and the U. S. Department of Agriculture campuses, is close to major
universities in College Park and Baltimore, and is surrounded by small, mid-size and large biotechnology companies.

Applications are currently invited for three tenure track faculty positions:
• Metabolomics: Applicants in the ﬁeld of metabolomics using advanced analytical methods will be considered, especially those with interests that
include metabolite changes in response to disease or environmental stress, applications in functional genomics, metabolic networks, medicinal plant
metabolism, and development of metabolomic databases (Position# 300879).
• Pathobiology: Applicants in areas broadly related to pathobiology will be considered. Candidates with interests in plant or animal pathogens, parasites,
vectors or the responses of hosts to infection are encouraged to apply, especially those that may interact with existing programs in structural biology,
molecular interactions, genomics and proteomics and computational biology (Position# 300880).
• Structural Biology (X-ray crystallography or NMR spectroscopy): Applicants will be considered who have research interests in any area of con-
temporary structural biology, including biomedical, plant or insect biology (Position# 300881).

Successful applicants will be expected to develop a very competitive and externally funded research program. Applicants should submit their curriculum
vitae, a summary of future research plans, and names of three references electronically to [email protected] with reference to the position number
or by mail to the appropriate Search Committee: SEARCH COMMITTEE, UMBI Shady Grove, 9600 Gudelsky Drive, Rockville, MD 20850. Review
of candidates will begin January 1, 2007 and continue until the position is ﬁlled.

UMBI is an EEO/ADA/AA Employer.

SYMPOSIA
A NATIONAL SYMPOSIUM:

Office of the Science and Technology

PREDICTIVE HEALTH
Adviser to the Secretary of State AND SOCIETY
Jefferson Science Fellowships
The National Academies is pleased to announce a call for nominations and applications for
the 2007 Jefferson Science Fellows program. This program establishes a new model for
engaging the American academic science, technology and engineering communities in the
formulation and implementation of U.S. foreign policy. Jefferson Science Fellows will spend
one year at the U.S. Department of State in Washington, D.C. and may periodically travel to
U.S. foreign embassies and/or missions. Following the fellowship year, the Jefferson
Science Fellow will return to his/her academic career, but will remain available to the U.S.
Department of State for short-term projects over the following five years.
Jefferson Science Fellow awards are open to tenured academic scientists, technologists and December 18-19, 2006
engineers from U.S. institutions of higher learning. Nominees/applicants must be U.S. Emory Conference Center, Atlanta, Georgia
citizens and will be required to obtain a security clearance.
Speakers include
Detailed information on the Jefferson Science Fellows program is available on the Web: Elias Zerhouni, NIH Director;
www.national-academies.org/jsf. The deadline for nominations and applications for Kári Stefánsson, deCODE;
the 2007 program year is December 31, 2006. David Schwartz, NIEHS Director.
The Jefferson Science Fellows program is co-sponsored by the MacArthur Foundation and Sponsored by the
the Carnegie Corporation. Women and minorities are especially encouraged to apply. Emory/Georgia Tech Predictive Health Initiative
For more information, or to register, visit the website,
www.emory.edu/CME. Or contact Jennifer Vazquez
at 404-712-2660, [email protected]
Department of Health and Human Services
National Institutes of Health
Director, National Center for Research Resources and
Associate Director for Clinical Research (Extramural)

The Office of the Director, National Institutes of Health (NIH) in Bethesda, Maryland, is seeking applications from exceptional candidates for
the position of Director, National Center for Research Resources (NCRR). The Director, NCRR, will also serve as the NIH Associate Director
for Clinical Research (Extramural). NCRR, with a staff of approximately 100 employees and a $1 billion budget, is the focal point at NIH
for biomedical, clinical and translational research resources. The incumbent serves as a principal advisor to the Director, NIH; participates in
discussions relative to the development of major policy decisions affecting biomedical, clinical and translational research resources; provides
advice and consultation to NIH components, advisory councils and grantee organizations and institutions; and assures that effective administrative
procedures are established so that program operations and obligations of government funds and other resources are rendered consistent with
statutory and regulatory requirements and within limitations imposed by the Department of Health and Human Services (DHHS) and Executive
Branch policies. As Associate Director for Clinical Research (Extramural), the incumbent is expected to provide leadership for clinical research
activities across the NIH. This leadership will involve the coordination of clinical research activities to enhance the integration of basic and
clinical research. The Associate Director for Clinical Research will work closely with the other Institute and Center Directors to enhance the
efficiency and effectiveness of clinical research supported by the NIH. Applicants must possess a Ph.D., M.D., or a comparable doctorate degree
in the health sciences field plus senior level scientific experience and knowledge of biomedical, clinical and/or translational research programs in
one or more health science areas. Salary is commensurate with experience and a full package of benefits (including retirement, health, life, long
term care insurance, Thrift Savings Plan participation, etc.) is available. A detailed vacancy announcement, along with mandatory qualifications
and application procedures, can be obtained via the NIH Home Page at: http://www.jobs.nih.gov under the Senior Job Openings section. Dr.
Stephen Katz, Director, National Institute of Arthritis and Musculoskeletal and Skin Diseases, and Dr. David Schwartz, Director, National Institute
of Environmental Health Sciences, will be serving as co-chairs of the search committee. Questions on application procedures may be addressed
to Ms. Regina Reiter at [email protected] or discussed with Ms. Reiter by calling 301-402-1130. Applications must be received by
November 27, 2006.

Department of Health and Human Services Department of Health and Human Services
National Institutes of Health National Institutes of Health
Tenure-Track Position Clinical Center

The Division of Intramural Research, National Institute on Deafness and Tenure-track Physician
Other Communication Disorders (NIDCD), located in Bethesda, MD, is Clinical Center/Nuclear Medicine Department
seeking a tenure-track scientist to establish an independent research program This position is located in The Warren G. Magnuson Clinical Center, Nuclear
to study molecular and/or cellular mechanisms of hearing and balance. We Medicine Department (NMD).
welcome applications from candidates with a wide range of expertise. Prefer-
ence will be given to candidates whose experimental approaches complement We are seeking a research-oriented physician for a possible tenure-track position. An
those of our existing strong programs in the genetics, development and cell M.D. or M.D. /PhD with U.S. Nuclear Medicine Board certification and CT training
biology of hearing. The successful candidate will join a dynamic group of is needed to provide diagnostic and therapeutic nuclear medicine procedures as well
scientists in a growing intramural program that is at the forefront of research as to participate in clinical research protocols of the NIH Intramural Program. U.S.
on communication disorders. citizenship or permanent residency status is required.
Please submit your curriculum vitae, bibliography, and a letter describing your
The NIDCD offers an exceptional working environment including well- clinical, research, and management experience to: Mrs. Veronica Olaaje, HR
equipped research laboratories and numerous opportunities for collaboration. Specialist, DHHS, NIH, OD/CSD-E, 2115 E. Jefferson Street, Rm. 2B209
Candidates for this position must possess a Ph.D. and/or M.D., post-doctoral MSC-8503, Bethesda, MD 20892-8503. Phone: 301-435-4748. Email: volaaje@
experience, and an outstanding publication record. Salary is commensurate mail.nih.gov.
with education and experience.
Salary is commensurate with experience. This appointment offers a full benefits
Please submit a curriculum vitae including bibliography, three reprints of package (including retirement, health, life and long term care insurance, Thrift
recent relevant publications, statement of research interests, an outline of Savings Plan participation, etc.). Application packages should be submitted as early
your proposed research, and the names and addresses of three references to: as possible, but no later than December 31, 2006.
Ms. Trudy Joiner, Office of the Scientific Director, NIDCD, 5 Research Selection for this position will be based solely on merit, without discrimination for
Court, Room 2B28, Rockville, MD 20850 ([email protected]). non-merit reasons such as race, color, religion, sex, national origin, politics, marital
Applications will be accepted until December 15, 2006. status, sexual orientation, physical or mental handicap, age or membership or non-
membership in an employee organization.
BIOENGINEERING FACULTY POSITION
University of California San Francisco
The Program in Bioengineering in the School of Medicine at the University of California San Francisco, seeks to hire a tenure track faculty
member to mount an exciting research program and to teach graduate, professional, and postdoctoral students.

The unique opportunities for interactions between basic and clinical scientists at UCSF have enabled the development of new medical treatment
strategies, including novel methods for delivering and evaluating cell and drug-based therapies. We seek candidates who will advance therapeutic
bioengineering at UCSF in the following research areas: the development of new molecular probes for imaging and tissue targeting, manipula-
tion of progenitor cells, design of biological activity sensors for normal and abnormal physiology, fabrication of tissue replacements and drug
delivery devices, and computational modeling of disease processes.

Faculty participate in the Program in Quantitative Biology (PQB) at UCSF, the Joint UCSF/UCB Graduate Group in Bioengineering and the
California Institute for Quantitative Biomedical Research (QB3). Applicants should have a doctoral degree or equivalent in biological, engineer-
ing or physical sciences, with a major focus on applications to biomedical problems. Priority will be given to an appointment at the Assistant
or Associate Professor level.

Review of applications will commence January 2007. Applicants should send, by post or email: curriculum vitae, electronic ﬁles or reprints
of one or two key publications, and a two-page summary of past research and future goals. Applicants must arrange for three letters of recom-
mendation to be sent by post or email. All materials should be addressed to:

Tejal Desai, Ph.D., Bioengineering Faculty Search Committee

Byers Hall, Suite 216, MC 0775, University of California San Francisco
San Francisco, California 94143

Electronic submission of all materials is encouraged and should be emailed to:

[email protected]

Questions may be addressed to Hillie Cousart, Ph.D. Manager - Program in Bioengineering, (415) 514-9242.

UCSF is an Afﬁrmative Action/Equal Opportunity Employer. The University undertakes afﬁrmative action to assure equal employment oppor-
tunity for underutilized minorities and women, for persons with disabilities, and for Vietnam-era veterans and special, disabled veterans.

Dunedin, New Zealand

Stuart Chair of Science Fellowships for Postdoctoral Scholars at

Communication Woods Hole Oceanographic Institution
Centre for Science Communication New or recent doctoral graduates with research interests associated
with the following are encouraged to submit fellowship applications
Applications are invited for an appointment to the Stuart Chair of prior to January 15, 2007.
Science Communication, the endowed foundation Professor of a Departments - Three awards related to the following areas are anticipated:
newly formed Centre for Science Communication at the University of Otago. The Centre is Applied Ocean Physics & Engineering; Biology; Marine Chemistry &
administratively located in the Division of Sciences but will have strong links to the other Geochemistry; Geology & Geophysics; Physical Oceanography
three academic divisions. The Chair will be the senior academic leader and Director of the Institutes - Each of the following Institutes, which foster interdisciplinary
Centre, charged with its national and international development and with ensuring that the research addressing critical issues, will award a fellowship to support
Centre is committed to excellence in all its activities. related research: Coastal Ocean Institute; Deep Ocean Exploration
The successful appointee will be passionate about communicating science and its Institute; Ocean and Climate Change Institute; Ocean Life Institute
promotion and popularisation in the community. They will have an exemplary international The NOAA-WHOI Cooperative Institute for Climate & Ocean Research
research reputation, a capacity to provide academic vision and strategic leadership, (CICOR) will award a Fellowship in one of three theme areas: Coastal
and the ability to work with staff from diverse academic and cultural backgrounds. It is Processes; Climate; Marine Ecosystems.
expected that links with other academic units throughout the University will be initiated The Beacon Institute for Rivers and Estuaries will award a Fellowship
and fostered, and that strong associations with the appointee’s primary research area will related to estuaries, rivers, and/or nearshore coastal ocean research.
be maintained to promote continued scholarship there as well as publications directed at The National Ocean Sciences Accelerator Mass Spectrometer Facility
science communication. will award a fellowship in the development and implementation of new
Further information about the University of Otago can be found at http://www.otago.ac.nz techniques in Radiocarbon Studies in Marine Science.
Speciﬁc enquiries may be directed to Professor Vernon Squire, Pro-Vice-Chancellor, Sciences, Awards are competitive, with primary emphasis placed on research
Tel 64 3 479 7977, Email [email protected] promise. Fellowships are for 18-months, with an annual stipend of
Reference Number: A06/214. Closing Date: Wednesday 31 January 2007. $54,000, a modest research budget and eligibility for group health
insurance. Recipients are encouraged to pursue their own research
APPLICATION INFORMATION interest in association with Resident Scientific and Senior Technical
Staff. Communication with potential WHOI advisors prior to submitting
With each application you must include an application form, an EEO Information Statement, an application is encouraged.
a covering letter, contact details for three referees and one copy of your full curriculum
vitae. For an application form, EEO Information Statement and a full job description Further information, application forms, and links to Individual Departments,
Institutes and Centers and their research themes
go to: www.otago.ac.nz/jobs Alternatively, contact the Human Resources Division,
may be obtained at:
Tel 64 3 479 8269, Fax 64 3 479 8279, Email [email protected] http://www.whoi.edu/apo/postdoctoral,
Equal opportunity in employment is University policy. or by contacting the Postdoctoral Fellowship
Committee at: (508) 289-2219, or [email protected].
An Equal Opportunity/Affirmative Action Employer
www.otago.ac.nz/jobs
DEAN, College of Natural What will I find in a career at Lilly?
and Agricultural Sciences
A unique opportunity for decanal leadership exists at The
Answers.
University of California, Riverside in the College of Natural and For more than 130 years, Eli Lilly and Company has been dedicated to
meeting the health care needs of people around the world. We address
Agricultural Sciences. these needs primarily by developing innovative medicines—investing a
higher percentage of our sales in research and development than any
other major pharmaceutical company.
A doubling of faculty positions is projected over the next
decade, adding to a rich infusion of new appointments over the
past decade. This opportunity for growth occurs in the context
BIOMEDICAL INFORMATICS
The advent of post-genomic technologies such as gene expression,
of our distinctive, internationally renowned base of physical, proteomics, functional genomics, genetic association studies, etc., and
their application towards the goal of personalized medicine requires
biological, biomedical and agricultural sciences, combined with development of new informatics methods for integration and analysis of
a new Health Sciences Research Initiative. An individual who preclinical and clinical data. The applications developed by the candidate
can recognize the value of this existing diversity will find will provide decision enabling data and analyses in support of Lilly’s
pharmacogenomics and tailored therapeutics programs. The candidate
unparalleled opportunities to exercise a vision for its future. In will develop software to support discovery, translation, validation of
undergraduate diversity, UC Riverside is the flagship campus of clinical diagnostics and biomarkers, and enable integrative analysis of
genomic, proteomic and clinical phenotypic data. The developed systems
the UC system. will be utilized by both Biological Research Scientists and Clinical
Research Physicians. This position will directly support clinical trial
research performed by the Diagnostics and Exploratory Medicine group
UCR seeks a distinguished scientist and leader for this position. at Lilly.
The Dean is expected to provide the strong leadership and
Candidates must have a Ph.D. in biomedical informatics, bioinformatics,
administrative direction in research, curriculum, fundraising computer science, computational biology, or a related field; or M.S. in
and outreach necessary to carry the College to preeminence. biomedical informatics, bioinformatics, computer science, computational
biology, or a related field, and 2 or more years of work experience.

The College has 237 permanent, ladder-rank faculty positions in For more information or to apply, visit www.lilly.com/careers. Eli Lilly
and Company is an equal opportunity employer.
thirteen departments and nine centers, with doctoral programs in
a full array of scientific disciplines, as well as interdisciplinary www.lilly.com/careers
programs. Research funding in the College for fiscal 2006 was
over 53 million dollars. The campus anticipates growth in
enrollment from the present 17,000 students to 25,000 by 2015.

The Dean is responsible for faculty appointments and

promotions, developing public and private funding sources,
fiscal management, allocation of resources, and coordination of
College programs within the overall campus academic plan.
The Dean reports to the Executive Vice Chancellor/Provost.
Salary is commensurate with experience and qualifications. The
Assistant/Associate Professor of Bioinformatics
appointment will be effective July 1, 2007 or whenever a Computational Biology Department
suitable candidate is identified. of Biostatistics
A comprehensive, land-grant research institution, UC Riverside Harvard School of Public Health
is a vibrant, expanding campus, located in inland Southern
The Department of Biostatistics at Harvard School of Public Health
California, 60 miles southeast of Los Angeles. It lies within an (HSPH) is seeking an outstanding candidate for a tenure-track faculty
hours drive of Palm Springs, the San Gabriel and San position in Bioinformatics/Computational Biology at the level of
Bernardino Mountains, the Mojave and Coachella Deserts and Assistant or Associate Professor. The successful candidate would join
Pacific Ocean beaches. an active group at HSPH developing novel computational and statisti-
cal methods, and conduct collaborative research with clinical and basic
Nomination and Application Procedures: Candidates should scientists at Harvard University and its afﬁliated medical centers. She/he
is expected to play a vital leadership role in expanding the quantitative
include a letter of application with statement of interests, a
science research and educational programs at HSPH in bioinformatics
current CV, and contact information for at least five references. and computational biology and its related ﬁelds. Candidates should have
All inquiries, nominations and applications will be held in doctoral degree and a demonstrated record of achievement; candidates in
confidence. Review of applications will begin January 2, 2007 all areas of computational biology, bioinformatics and statistical science
and continue until the position is filled. Inquiries and questions are encouraged to apply.
should be directed to Professor Christopher Reed
([email protected]), Chair of Search Committee, Department Please send a letter of application, including a statement of current and
of Chemistry, UC Riverside, CA 92521; phone (951) 827 5197. future research interests, a curriculum vitae, sample publications, and
the names of three referees to the address below. Applicants should ask
Nominations and applications should be sent by E-mail to:
their three referees to write independently to this address:
Professor Raymond L. Williams ([email protected]),
Director of Executive Searches, 3144 Hinderaker Hall, Computational Biology Junior Faculty Search Committee
University of California, Riverside, CA 92521. UCR is an Department of Biostatistics
affirmative action/equal opportunity employer. Harvard School of Public Health
655 Huntington Avenue, 4th Floor
Boston, MA 02115

Harvard School of Public Health is strongly committed to increasing

the representation of women and minority members among its faculty
and particularly encourages applications from such candidates.
The Christian-Albrechts University in Kiel invites applications for the positions of
14 Professorships (W1 and W2)
as part of the Excellence Cluster „The Future Ocean“.
The Christian-Albrechts University in Kiel (CAU) jointly with the Leibniz-Institute of Marine Sciences (IFM-GEOMAR) is establishing a cross-faculty,
multi-disciplinary research focus with the title “The Future Ocean” for the multi-facetted study of human-induced ocean change, marine hazards and marine
resources (www.uni-kiel.de/future-ocean/). As part of this initiative, 14 new Professorships (W1 and W2) are being established which we wish to fill with
exceptional young researchers from the fields of natural sciences, law, economics and medicine. Salary will normally be at grade W1, salary at grade W2 is
however possible dependent on fulfilling legal and personal qualification requirements.
The W2-Professorships are initially for 5 years. The W1 Junior Professors will initially be appointed for 3 years („Beamtenverhältnis auf Zeit“); dependent
on performance of the Junior Professor after these three years, the position can be extended by up to 3 additional years. The willingness to learn German is
expected. The positions also bring with them funding for additional personnel and research materials so that the new Professors should quickly be able to set
up excellent research groups. Multidisciplinary marine research is an important focus of the CAU. As a consequence we wish to open up long-term career
opportunities at the University for successful junior faculty. Based upon performance of the individual appointees, a proportion of these initially non-tenured
positions will be changed into tenured W2/W3-positions. We are seeking for each post an exceptional scientist.
Mathematics-Natural Sciences Faculty
Ocean acidification (www.uni-kiel.de/future-ocean/a1): to study the influence of ocean acidification on marine organisms. Research foci could range from
the cellular to the ecosystem level and from molecular studies to field experiments. The position will be hosted at IFM-GEOMAR.
Seafloor warming (www.uni-kiel.de/future-ocean/a2): to study the influence of seafloor warming on sedimentary gas hydrates and/or benthic fauna with the
help of a variety of methods (e.g. field studies and experiments, laboratory studies, numerical models). The position will be hosted at IFM-GEOMAR.
Ocean circulation (www.uni-kiel.de/future-ocean/a4): to study the relationships between changes in ocean circulation and the hydrological cycle with the
use of Earth System models and their further development by implementation of palaeoenvironmental parameters. The candidate should have experience in
the transient climate modelling of long time periods.
CO2-Sequestration (www.uni-kiel.de/future-ocean/a5): to study various scenarios of CO2-sequestration from an integrative stand-point, using molecular
dynamics modelling, advanced spectroscopic methods and experimental high-pressure simulations of oceanic in-situ conditions. The candidate should be a
demonstrated expert in one of these fields and build a group to cover the other fields, working closely with experts in adjacent fields already present in the
Cluster in Kiel.
Chemistry of the ocean surface (www.uni-kiel.de/future-ocean/a6): to study the chemical structures and heterogeneous reactions on aerosols, in clusters
or on air/ice or air/water interfaces with modern methods. The candidate should have demonstrated experience in the fields of reaction kinetics and/or
spectroscopy.
Seafloor resources (www.uni-kiel.de/future-ocean/b3): to study the temporal and spatial influence of fluid movement on the formation of seafloor resources
using a combination of numerical modelling with field information on physical and chemical boundary conditions. The position will be hosted at IFM-
GEOMAR.
Natural hazards (www.uni-kiel.de/future-ocean/b4): to study geological hazards at plate boundaries, particularly earthquakes and slope instabilities. The
position will be hosted at IFM-GEOMAR.
Sea level rise and coastal erosion (www.uni-kiel.de/future-ocean/b5): to study processes of hydrodynamics and sediment dynamics in the coastal zone.
The candidate should have demonstrated experience in marine geophysics, especially in the field of high resolution hydro-acoustics at the sediment-water
interface.
Risk management in the coastal zone (www.uni-kiel.de/future-ocean/b5): with demonstrated experience in geographic hazard research, preferably in
coastal regions. The work should focus on the (further) development of qualitative and quantitative methods of risk assessment (particularly remote sensing)
as well as the development of decision support systems for risk management.
Economics and Social Sciences Faculty
Living resources and overfishing (www.uni-kiel.de/future-ocean/b1): with demonstrated experience in economics, especially resource economics, property
rights and decisions under uncertainty. The appointee will be expected to work in the area of “Living Resources and Overfishing” and if possible also “Coasts at
Risks”. This implies a willingness to carry out interdisciplinary research with other Cluster disciplines (esp. Marine Biology, Geography, Law). The appointee
should also teach courses in the Bachelor/Master of Science in Economics and supervise doctoral students.
Ocean economics (www.uni-kiel.de/future-ocean/a7): with specialization in environmental and resource economics to work on the subject “Valuing the
Oceans“ within the Excellence-Cluster, investigating economic aspects of the role of the oceans in the global carbon cycle. We expect a willingness to carry
out interdisciplinary studies both within the subject area and within the Cluster as a whole. The appointee should also teach courses in the Bachelor/Master
of Science in Economics and supervise doctoral students.
Technical Faculty
CO2-take-up in the ocean (www.uni-kiel.de/future-ocean/a3): with expertise in computer science, particularly algorithm design, numerical mathematics
or optimization. The appointee is expected to study the optimization of the modeling of oceanic CO2-uptake through the assimilation of both physical and
biogeochemical data. Participation in the teaching programs in computer science (Diplom, Bachelor/Master of Science) is expected.
Medical Faculty
W2 - Molecular marine medicine (www.uni-kiel.de/future-ocean/b2): with experience in molecular and cell biology, in comparative genetics/genomics,
in in silico analysis and in the molecular pathophysiology of human diseases. This position forms part of the molecular medicine focus of the CAU. Central
to the work will be an evolutionary understanding of complex barrier break-downs in humans, a presently unsolved medical problem. Marine organisms with
simple barrier functions should be used as models.
Faculty of Law
W2 - Law of the Sea (www.uni-kiel.de/future-ocean/b6): for research and teaching in the law of the sea, ideally in combination with both, domestic
and international environmental and economic law. Additionally, participation in the teaching of public law would be welcome. Applicants must possess a
“Habilitation” or demonstrate equivalent scientific achievements.
In addition to the necessary formal qualifications (as set out in §99 of the University Law of Schleswig-Holstein), a pre-requisite for a Junior Professorship
is an excellent dissertation. The CAU would like to fill at least 50% of these new positions with female applicants. Peer-Monitoring, career counselling and
special support will be provided if requested. The CAU offers a family-friendly working environment and is pro-active with respect to double-career families
(www.uni-kiel.de/audit-fgh). The State Government and the University support the employment of disabled persons. Persons with disabilities will, with
appropriate qualifications and aptitudes, be employed preferentially.
Selection will take place according to the normal procedure for university Professors. Applications, including a curriculum vitae, qualifying documentation
(publications, evidence of external funding, teaching experience) and a research and financial plan for the establishment of the research group can be submitted
until 8 January 2007 to: The Rector, Prof. Dr. Thomas Bauer, Rektor der CAU, Christian-Albrechts-Platz 4, D-24118 Kiel, Germany.
School of Medicine
Chair, Department of Biochemistry
West Virginia University School of Medicine is seeking an accomplished
academic leader and scientist to serve as Professor and Chair, Depart-
CHIEF, DIVISION OF
ment of Biochemistry. The successful candidate will have the leadership
skills and vision required to achieve the research, education and service
REGENERATIVE MEDICINE
missions of a basic science department in a multidisciplinary research Department of Medicine
and graduate training environment. An essential responsibility for the
new Chair will be to integrate the traditional Departmental missions $JG %2K:G67K8[ 30 &K61K2K+ G4+68OG28 30 /G.K-K2G K7 7GGMK21
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research in a multidisciplinary environment. The SRP established six core 44NK-+287 O978 J3N. / +2.36 J

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Neuroscience, and Diabetes and Obesity. New research directions in -966G28 & 83
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new faculty.
As part of the commitment to research expansion in the Health Sciences
Center, a new research building is under construction. Supporting core
research facilities in proteomics and protein sequencing, microscopy
and cellular imaging, functional MRI and brain imaging, and mouse
transgenics is an institutional priority. These facilities complement the Department of Cell Biology
current expansion of the Health Sciences Library and the new Blanchette
Rockefeller Neurosciences Institute building. West Virginia University Tenure Track Faculty Position in Cell Biology
is a comprehensive, land grant, and a “Carnegie-designated Research The Department of Cell Biology at the University of Alabama at Bir-
University with high research activity.” There are approximately 26,000 mingham (www.uab.edu/cellbio) invites applications for faculty posi-
undergraduate and 5,500 graduate students across the entire campus. The tions at the Assistant, Associate or Full Professor level in the broad area
WVU Health Sciences Center includes the Schools of Medicine, Phar- of molecular cell biology. Rank and tenure status commensurate with
macy, Dentistry and Nursing. Each school has both health professional qualifications and experience. Preference will be for investigators whose
and graduate training programs. Faculty in the School of Medicine are research expertise complement existing strengths in the department, which
involved in well-established PhD training programs in the biomedical include developmental biology, cancer biology, signal transduction, cell-
sciences, an MD/PhD Scholars program, and interdisciplinary graduate matrix interactions and membrane trafficking. Successful candidates will
training in Public Health. Morgantown has 55,000 residents and is rated be expected to develop a strong extramurally funded research program
as one of the best small towns in the U.S., with affordable housing, excel- and to contribute to departmental responsibilities associated with gradu-
lent schools, a picturesque countryside and many outdoor activities. ate and professional student training. UAB offers a highly interactive
Qualifications: PhD and/or MD degree with a record of excellence scientific environment with state-of-the-art research facilities. The pres-
in research, the ability to attract and develop extramurally funded, ence of numerous multidisciplinary research centers fosters collaboration
multidisciplinary research programs, and experience in graduate and among the basic science and clinical faculty. The Medical School and the
professional student education. It is very important that the candidate Cell Biology Department are each ranked in the top 20 for NIH research
be able to bridge the boundaries of traditional disciplines, including the funding. The Department provides excellent laboratory facilities, highly
promotion of collaborative, translational research efforts between basic competitive salaries and start-up funds, and access to numerous core facili-
and clinical faculty. Review of applications will commence immediately ties. Candidates should have a Ph.D. or equivalent degree, postdoctoral
and continue until the position is filled. E-mailed applications are pre- experience, and clear evidence of research productivity.
ferred with attachments for a curriculum vitae, a cover letter indicating Applicants should send a current curriculum vitae, a brief summary of
your interest in the position addressed to the Chair of the Search Com- past accomplishments and future research plans, and three letters of
mittee (below), and the addresses including email for three references reference to:
(in confidence), and should be sent to the Administrative Assistant, Cell Biology Search Committee
Carol Smith ([email protected]). If necessary, applications sent c/o Maxine Rudolph
by standard mail and all other communications should be addressed to
1918 University Boulevard - MCLM 660
the Chair of the search committee: Richard D. Dey, Ph.D., Professor
The University of Alabama at Birmingham
and Chair, Department of Neurobiology and Anatomy, P.O. Box
Birmingham, AL 35294-0005
9128, West Virginia University, Morgantown, WV 26506-9128 (304
293-5979; [email protected]). Email: [email protected]
West Virginia University is an Affirmative Action/ The University of Alabama at Birmingham is an Equal Opportunity/
Equal Opportunity Employer. Affirmative Action Employer.
Bioinformatics Faculty Positions
The University of Louisville is seeking applicants
to fill 10 new tenure-track faculty positions as
part of a major multidisciplinary expansion of the
university’s Bioinformatics and Computational
Biology infrastructure.

Excellence in Research Position Description

and Education The University of Louisville Bioinformatics Task Force invites
applications for five tenure-track positions scheduled to
The Departments of Bio-
begin in fall 2007. The remaining five positions will be filled
chemistry and Molecular
in fall 2009.
Biology (School of Medicine),
Bioinformatics and Biostatis-
Depending on the experience of the applicant, available po-
tics (School of Public Health
sitions are at the assistant, associate or full professor level.
and Information Sciences),
Responsibilities will include research and teaching with high
Computer Engineering and
levels of collaborative and interdisciplinary activities.
Computer Science (J.B.
Speed School of Engineer- Ideal candidates will be highly motivated scientists with
ing), Mathematics (College excellent communication, presentation and interpersonal
of Arts and Sciences) and skills and a bioinformatics research focus to complement
basic science departments the expansion of biomedical research programs. Research
within the School of Dentistry experience may include structural bioinformatics, metabolo-
have partnered to coordinate mics, proteomics and genetic data analyses, high-through-
efforts to bring bioinformat- put analyses, systems biology, visualization techniques and
ics to the forefront in shap- algorithms development.
ing the future of research at
the University of Louisville. Candidate Requirements
Come join our team!
bioinformatics.louisville.edu The positions require a Ph.D. in biochemistry, computer
science, engineering or bioinformatics. Strong English lan-
The University of Louisville is guage skills and a willingness to collaborate with research-
an exciting academic commu- ers with diverse backgrounds are also necessary. In addition,
nity of people whose ideas applicants should have a demonstrated record of excellence
and work make outstanding in collaborative research activity and teaching, including
contributions to Kentucky experience with extramural funding agencies.
and the nation. For more
information, go to: To Apply
For an application and additional information go to:
www.louisville.edu bioinformatics.louisville.edu/jobs.html

The University of Louisville is an

Equal Opportunity Employer.
National Exposure Research Laboratory
Post-Doctoral Program
• The National Exposure Research Laboratory (NERL) of the United States Environmental Protection
Agency is accepting applications beginning November 20, 2006 through January 31, 2007 for approxi- Cancer Immunobiology Center
Research Non-Tenure Track
mately 16 federal, three-year post-doctoral research positions.
• Candidates will engage in research in areas such as environmental monitoring and characterization; com-
puter modeling of the transport, transformation, and fate of pollutants in multiple media and at multiple
scales; human and ecological exposure analysis; remote sensing applications; and landscape ecology.
Faculty Member
The Cancer Immunobiology Center at the University
• Specific research opportunities are posted on the NERL website at http://www.epa.gov/nerl. of Texas Southwestern Medical Center at Dallas, TX
• Post-doctoral positions will be in one or more of the following locations: Research Triangle Park, North is seeking a Research Non-Tenure Track Faculty
Carolina; Cincinnati, Ohio; Las Vegas, Nevada; Athens, Georgia; or Washington DC metropolitan area. Member with a strong background in biochemistry
and experience in running a GMP laboratory. The
FULL FEDERAL EMPLOYMENT BENEFITS:
faculty candidate will both supervise and work in
• Full three-year appointments • Paid relocation to EPA duty location
this facility. The applicant will interact with a
• Travel to professional and scientific meetings • Vacation and sick leave research team, a clinical trials director and a
• Federal health benefits, life insurance, and retirement program technical group. Experience in producing proteins in
• Salary range of $51,972 - $84,559 (subject to increase in January 2007) mammalian cells or bacteria, monoclonal antibodies
• Flexible start date beginning in March and no later than September 2007 and immunconjugates is desirable. The reagents
APPLICATION PROCESS – Consult the NERL website at http://www.epa.gov/nerl for instructions generated in this facility will be used in Phase I and
on how to apply. Note – online applications from journal websites are not accepted. Applicants must II clinical trials. Salary and start-up packages are
provide: highly competitive. All applicants must have a PhD
• Up-to-date Curriculum Vitae with minimum of 3 years of biotech experience in a
• Letter of recommendation from your research advisor or comparable official GMP facility. Applicants must have experience in
• Cover letter indicating: positions and locations of interest, your email address, U.S. Citizenship status, writing protocols, data sheets, batch records and
AND How you learned of this program oversight of FDA compliance.
• DD-214, if claiming veteran’s preference Please send cover letter, letters of reference, curriculum
Applicants must be United States citizens or permanent residents. Only in the absence of qualified U.S. vitae, and summary of professional goals to:
citizens will permanent residents who are citizens of countries specified as exceptions to the appropriations
act ban on paying non-U.S. citizens be considered.
Cancer Immunobiology Center
c/o Linda Berry
Specific job information is posted on the NERL Internet site at http://www.epa.gov/nerl. EPA provides UT Southwestern Medical Center
reasonable accommodations to applicants with disabilities. If you need a reasonable accommodation for any 5323 Harry Hines Blvd.
part of the application and hiring process, please notify the Agency. The decision on granting reasonable Dallas, Texas 75390-8576
accommodation will be on a case-by-case basis. Email: [email protected]
The U.S. EPA is an Equal Opportunity Employer. EOE

OTOLARYNGOLOGIST
The Section of Otolaryngology - Head and Neck Surgery at
Dartmouth-Hitchcock Medical Center seeks a board certified Endowed Eminent Scholar in
or board eligible Otolaryngologist for a full-time faculty posi- Molecular Cancer Pharmacology
tion.The candidate should possess an interest in an academic Tulane Cancer Center and the Louisiana Cancer Research
career and in the education of medical students and resi- Consortium (LCRC) seek an outstanding cancer scientist to become
dents.This position will combine a general otolaryngology an Endowed Eminent Scholar, The Joe W. and Dorothy Dorsett
Brown Foundation Distinguished Chair in Molecular Cancer
with a subspecialty practice in otology or pediatric otolaryn- Pharmacology. The eminent scholar holding this tenure track posi-
gology. Fellowship training in otology/ neurotology or pedi- tion will be responsible for coordinating basic research leading to the
atric otolaryngology is desirable. Research interests will be discovery and pre-clinical development of cancer therapeutics.
encouraged. Academic rank will be commensurate with qual- Tulane University Health Sciences Center and Louisiana State
ifications and experience. University Health Sciences Center in New Orleans have joined
together to develop the LCRC, with the goal of achieving NCI desig-
Interested applicants are encourage to send letters of nation as a Comprehensive Cancer Center. Continuing funding from
inquiry and CV to: a new state tax on cigarettes and significant financial commitment by
both Tulane and LSU provide generous resources for the suc-
Daniel Morrison, MD, Chairman cessful candidate to recruit additional faculty members in both
Section of Otolaryngology - Head & Neck Surgery basic and clinical sciences. The goal is to establish a world-
class program bringing basic research toward clinical testing.
Dartmouth-Hitchcock Medical Center
The successful candidate will enjoy modern laboratory space,
One Medical Center Drive access to shared core resources, and the opportunity to devel-
Lebanon, NH 03756 op further the LCRC Cores.
Telephone: 603-650-8123 Qualified candidates should forward CV and three letters of ref-
erence to: Roy S. Weiner, M.D., Director, Tulane Cancer Center,
Tulane University Health Sciences Center, 1430 Tulane Ave., SL-68,
New Orleans, LA 70112, [email protected] or Krishna C. Agrawal,
Ph.D.,Chairman, Department of Pharmacology, Tulane University
Health Sciences Center, 1430 Tulane Ave., SL-83, New Orleans, LA
70112, [email protected].
Dartmouth-Hitchcock Medical Center is an affirmative action/equal opportunity
employer and is especially interested in identifying female and minority candidates. The position will remain open until a suitable /
qualified applicant has been identified.
An affirmative action / equal opportunity employer.
www.DHMC.org
FELLOWSHIPS

MBL
ou ded 888 as t e a e o og ca abo ato y

2007 Summer Research Fellowships

• Funding Available for Summer Research
APPLICATION DEADLINE: JANUARY 16, 2007

The MBL is pleased to announce the availability of funding for the following summer research
programs in 2007 for junior or senior investigators holding a Ph.D., M.D., or equivalent degree.
These prestigious awards provide funds for research and housing. Proposals for fellowship support
will be considered in, but are not limited to, the following ﬁelds of investigation:

Cellular & Molecular Physiology Neurobiology Parasitology

Molecular Biology Innate Immunity Microbiology
Developmental Biology Ecology

• Funding Available for Summer Research in Neuroscience

APPLICATION DEADLINE: JANUARY 16, 2007

The MBL is pleased to announce the availability of funding for the following summer research
programs in Neuroscience in 2007. These programs will provide up to $50,000/year/award with a
possibility for renewal for 3 years. As participants in the MBL’s new Neuroscience Institute, scholars
in these programs will beneﬁt from the rich intellectual and interactive environment of the scientiﬁc
community at the MBL.

ALBERT AND ELLEN GRASS FACULTY GRANTS PROGRAM

Proposals must describe collaborative research in any area of neuroscience by teams of two or more
investigators, with a minimum stay of six weeks at the MBL in Woods Hole. The intent of the program
is to attract new investigators to the MBL at the assistant, associate, or full professor levels. The collab-
orative research can be between new teams of investigators or between new investigators who wish
to collaborate with a more established investigator.

Requests will also be considered for collaborative research projects at the MBL during the off-season
period. Funds may be used for laboratory and equipment rentals, supplies, and incidental expenses,
housing and travel costs.

DART FOUNDATION SCHOLARS PROGRAM IN LEARNING & MEMORY

Proposals must be targeted to the study of learning and memory with a minimum stay of six weeks at
the MBL. Applications are encouraged from junior- or senior-level neuroscientists holding a Ph.D., M.D.
or equivalent degree. Awards provide funds for research and laboratory rental, and cover the costs of
housing and travel.

For application forms and information, contact: Fellowships Coordinator: [email protected]

or call Lenny Dawidowicz, 508.289.7268, MBL, 7 MBL Street, Woods Hole, MA 02543.

Applications are encouraged from women and members of underrepresented minorities. The MBL is an Equal Opportunity/
Afﬁrmative Action Employer.

www.MBL.edu/fellowships
University of Michigan
Computational Biology
CARDIOLOGIST
of Complex Systems
TRANSLATIONAL RESEARCH SCIENTIST
The Section of Cardiology at Dartmouth-Hitchcock Medical
Center/Dartmouth Medical School is seeking to recruit two new faculty
members (M.D or M.D./Ph.D.) board certified/eligible in
Cardiovascular Disease with a strong interest in basic or translational
research. The appointments will be made at the Assistant
Professor/Associate Professor/Professor level depending on qualifications
and experience. A position in the Angiogenesis Research Center is avail-
able to the qualified candidate. We seek individuals with a strong record
of academic productivity and the potential to establish or bring an inde-
pendent research program focusing on vascular biology and develop-
ment, genetics, molecular imaging or myocardial biology. An expertise in
zebrafish or Xenopus research is particularly welcome. The successful
The Center for Computational Medicine and candidate will be expected to participate in clinical activities of the
Section of Cardiology and to engage in teaching in the Experimental
Biology (CCMB) seeks to hire several Assistant/ Molecular Medicine graduate program and in the Angiogenesis Research
Associate Professors to develop independent Center. State-of-the-art facilities in the Section of Cardiology and
Angiogenesis Research Center include 3D echo, MR and CT imaging,
research programs involving the modeling of an advance microscopy core, and a mouse physiology and imaging cores.
complex biological systems. This work should Additionally, Dartmouth Medical School offers many facilities that
include micro-CT and micro-PET imaging, transgenics/knockout core
address multiple levels of organization that connect and genomics and proteomics cores.
molecules to function. We are interested in multi- Please e-mail your curriculum vitae, a description of your research
scale, integrative analyses of metabolic pathways, program, career goals and the names of three references to:
[email protected], Dr. Michael Simons, Director,
regulatory networks, signal transduction cascades, Angiogenesis Research Center, Dartmouth Medical School,
and physiological systems. Research may be strictly Lebanon, NH 03756.
computational or involve a wet lab component.
Candidates should have an earned doctoral degree,
at least two years post-doctoral experience, and a
passion for teaching graduate and/or professional
students. Successful candidates will have a tenure- www.dhmc.org
Dartmouth College is an Equal Opportunity/Affirmative Action employer and
track appointment in a Medical School Basic Sci- encourages applications from women and members of minority groups.

ence department and a research appointment in the

CCMB, with its Bioinformatics Graduate Program
and NIH National Center for Integrative Biomedi-
cal Informatics. You will have a generous start-up Lancaster University is a dynamic
package, access to our extensive campus-wide institution committed to building on
the reputation it has developed
computational environment, and encouragement
during its first forty years for
to collaborate with colleagues in interdisciplinary pioneering innovation and excellence
Centers on Type 1 and Type 2 Diabetes, Metabolo- in teaching and research.
mics and Obesity, Neurosciences, Organogenesis, To apply or receive further
Department of
Infectious Diseases, Cardiovascular Disorders, Environmental Science information online, please visit
Genetics, Cancer Biology, and Complex Systems. http://www.personnel.lancs.ac.uk/ or,
Professor/ telephone Personnel Services,
The postings and application process can be
viewed at http://www.umich.edu/~jobs/. Please Reader of quoting reference A763, on
answerphone (01524) 846549.
refer to posting numbers 4504, 4506, and 4507. Environmental Closing date: 12 January 2007.
You need apply for only one position; the screen-
ing committee will direct your materials to the Science In pursuance of the strategic growth
most appropriate basic science department. Please of this research-led Department
Reader (which is an integral part of the
submit electronically a cover letter, your research £41,133 - £46,295 p.a. Lancaster Environment Centre) we
plans, curriculum vitae, any current grant support, Professor by negotiation invite applications for the post of
and contact information for four references. min £50,064 p.a. Reader or Professor in either:

Please contact Gil Omenn or Violet Elder (violet • Earth System Atmospheric
Science or
@umich.edu) at the Center for Computational
Medicine and Biology if you have questions; feel • Environmental Radionuclide
Management
free to submit a second set of materials to this e-mail
address if you so desire. The anticipated start date for both
posts is 1 June 2007.
www.ccmb.med.umich.edu/jobs/EBS Informal enquiries may be addressed
to [email protected] or
The University of Michigan is an Aiming for Greater
Diversity [email protected]
Equal Opportunity/Afﬁrmative Action Employer.
HOWARD HUGHES MEDIC AL INSTITUTE

HHMI Investigator Competition

in Patient-Oriented Research

We invite applications from physician- Highly creative researchers who bridge the gap
between clinical medicine and basic science are in
scientists who have demonstrated
a unique position to exploit our knowledge of the
originality and productivity as patient- human genome and other recent advances to make
oriented researchers and who show discoveries that will improve human health.
exceptional promise for future The Howard Hughes Medical Institute seeks to appoint
contributions to the understanding approximately 15 outstanding physician-scientists
as HHMI investigators. This competition is open to
and treatment of human disease.
researchers with faculty appointments at 121 leading
institutions in the United States. Candidates should
Eligibility
apply directly to HHMI; prior institutional approval
M.D. or M.D./Ph.D. (or the equivalent) is not required.
Licensed to practice medicine in the The Howard Hughes Medical Institute, a nonprofit
United States medical research organization, plays a powerful role
Tenured or tenure-track position (or the in advancing biomedical research and education in
equivalent) at one of 121 eligible institutions the United States. HHMI’s investigator program rests
Four to 16 years of experience as an on the conviction that scientists of exceptional talent,
independent investigator commitment, and imagination will make fundamental
biological discoveries for the betterment of human
Principal investigator on an active NIH health if they receive the resources, time, and freedom
R01 grant or project leader on an active to pursue challenging questions. The Institute’s
NIH P01 grant investigators, selected through rigorous national
Outstanding patient-oriented research competitions, include 11 Nobel Prize winners and
program 115 members of the National Academy of Sciences.
The Howard Hughes Medical Institute is an equal
Application deadline: January 18, 2007 opportunity employer. Women and members of racial
and ethnic groups traditionally underrepresented in
Application information: the biomedical sciences are encouraged to apply.
www.hhmi.org/investigator_por/sci
VASCULAR BIOLOGIST
DEPARTMENT OF PATHOLOGY
BETH ISRAEL DEACONESS MEDICAL CENTER
HARVARD MEDICAL SCHOOL
The Department of Pathology at Beth Israel Deaconess Medical Center is seek-
ing a full-time biomedical scientist at the Assistant Professor level in the area of
vascular biology and angiogenesis. The Medical Center is a tertiary care facility NANOPHOTONICS/NANOPLASMONICS
and a major teaching hospital of Harvard Medical School. The Department of ATOMIC FORCE MICROSCOPY
Pathology is embarking on the most ambitious program of growth in its history
and is now actively expanding the strength and depth of both its clinical and The Center for Nanoscale Science and Technology at NIST in Gaith-
research faculty. Ground has been broken for a new, state-of-the-art research ersburg, MD is seeking two exceptional experimentalists with strong
building that will accommodate most of the research scientists within the Medi- records of creativity and achievement in the fields of (a) nanopho-
cal Center and all of the scientists within the Pathology Department.
tonics and/or nanoplasmonics, and (b) atomic force microscopy.
We are seeking candidates of exceptional promise who have strong records The applicants should possess the leadership abilities necessary to
of research creativity and productivity in basic or translational research in build a thriving research program, and should have a strong interest
vascular biology and angiogenesis. The research should involve fundamental
mechanisms for the regulation of angiogenesis. It may also involve the use of
in development of new instrumentation and measurement methods.
model organisms or the development of new technologies and strategies for It is important that the applicants be able to interact with multiple
the study of tumor angiogenesis. disciplines and present effectively their programs to a variety of
The successful candidate will receive a highly competitive start-up package,
audiences. The new research programs will interface with and build
appointment to the faculty of Harvard Medical School and full membership in upon extensive NIST programs for electrical, magnetic, chemical,
the Beth Israel Deaconess Vascular Biology Research Center. The Department of physical, optical, and biological nanoscale measurements and stan-
Pathology strongly encourages interactions among research and clinical faculty dards. For additional information about the Center for Nanoscale
and provides opportunities to access an extraordinary human tissue resource Science and Technology please visit http://cnst.nist.gov.
through its Divisions of Anatomic Pathology and Laboratory Medicine. We
also provide unparalleled opportunities for collaborative interactions within We will consider filling these positions at any appropriate level
the basic and applied vascular biology research community at Harvard Medical (payband III-V, salary $54,272-$139,774). Candidates must have
School and its affiliated teaching hospitals.
a PhD degree in physical science or engineering. Experience in
Applicants must hold a PhD and/or MD degree. Beth Israel Deaconess Medical nanophotonics and/or nanoplasmonics is required for one posi-
Center is committed to increasing the representation of women and members tion, and in atomic force microscopy for the other. CVs will be
of minority groups on its faculty, and we particularly encourage applications
accepted on a continuing basis and should be sent by e-mail to
from such candidates. Interested applicants should submit curriculum vitae, a
statement outlining existing and planned research activities and career goals, [email protected].
and the names of three professional references to: Dr. Jack Lawler, Direc-
tor, Division of Cancer Biology and Angiogenesis, Beth Israel Deaconess The Department of Commerce is an Equal Opportunity Employer.
Medical Center, Research North Room 270C, 99 Brookline Avenue, Boston, US citizenship is required.
MA 02215.

+-9N8[ 37K8K32 K2 3OR+6+8K:/ Tenure Track Position in Nanotechnology

/2/8K-73ORN/Z $6+K87 University of California San Francisco
The Department of Pharmaceutical Chemistry and the California Institute
QNN/1/ Q0 &/8/6K2+6[ //.K-K2/ for Quantitative Biomedical Research, QB3, seek highly qualified appli-
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.[2+OK- 8/+-JK21 +2. 6/7/+6-J 92K:/67K8[ ;J/6/ 78+000+-9N8[+2. 789./287 +NKM/ have strong research interests in nanotechnology. This position is part of a
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major initiative at the Mission Bay/China Basin Landing campus of UCSF
RRNK-+8K327 +6/ K2:K8/. 036 + 8/296/ 86+-M R37K8K32 +8 8J/ 3NN/1/ 30 &/8/6K2+6[ and will build on the University’s strengths in Chemistry and Quantitative
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,/J+:K36 +2. ./:/N3RO/28
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786+8/1K/7 83 /ZRN3K8 8J/ R38/28K+N 30 .31 +2. 38J/6 .3O/78K- +2KO+N Applicants should have a Ph.D., M.D. or advanced degree with considerable
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research experience and are expected to establish a dynamic research program.
+2.K.+8/7 7J39N. J+:/ + J

/+2.36 &/ 36 /59K:+N/28 ./16//7,/ Applicants are also expected to actively participate in graduate training in the
-3OOK88/. 83 ./:/N3RK21 + 78+8/308J/+68 /Z8/62+NN[ 092./. 6/7/+6-J Chemistry and Chemical Biology Program and in other Programs in Quantita-
R6316+O +2. ,/ O38K:+8/. 83 R+68K-KR+8/ K2 8J/ 92K:/67K8[¶7 8/+-JK21 +2. tive Biology, Biophysics or Bioengineering and in professional school teach-
R9,NK- 7/6:K-/ OK77K327
$J/ R37K8K32 K2-N9./7 + -3OR/8K8K:/ 78+689R R+-M+1/ ing. Applicants will be eligible for membership in the Comprehensive Cancer
+2. K7 300/6/. +8 8J/ 77K78+28 630/7736 N/:/N;K8J 6/:K7K32 +RR63R6K+8/ 83 8J/ Center, the Program in Biological Sciences (PIBS) and other graduate training
79--/7709N +RRNK-+28¶7 59+NK0K-+8K327
ORJ+7K7 ;KNN ,/ RN+-/. 32 K./28K0[K21 programs including the UCSF/UCB joint Program in Bioengineering.
+2 /Z-/R8K32+NN[ 8+N/28/. +2. R63OK7K21 K2.K:K.9+N ;J3 -+2 ,9KN. 32 8J/
/ZK78K21 +2. 3213K21 /ZR+27K327 30 R6316+O7 K2 -3OR98+8K32+N 1/23OK-7 UCSF seeks candidates whose experience, teaching, research, or community
-3OR+6+8K:/ O/.K-K2/
K2-N9.K21 + 2/; 1/2/8K- +6-JK:/ +2. O3N/-9N+6 service has prepared them to contribute to our commitment to diversity and
1/2/8K-7 8J+8 +6/ R+68 30 362/NN %2K:/67K8[¶7 .K0/ #-K/2-/7 +2K8K+8K:/ excellence.

NK0/7-K/2-/7
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/R+68O/28+N +00KNK+8K32 ;K8JK2 8J/ 3NN/1/ ;KNN ,/ Please send a curriculum vitae, three letters of reference, a summary of cur-
./8/6OK2/. ,+7/. 32 8J/ 79--/7709N +RRNK-+28¶7 ,+-M16392. +2. /ZR/68K7/

rent research (up to 3 pages), and a concise outline of future research (up to
N/+7/ 7/2. + -966K-9N9O :K8+/ 8J6// N/88/67 30 6/0/6/2-/ +2. + 6/7/+6-J 3 pages) by January 31, 2007, to the address below.
78+8/O/28 K2 + 036O+8 83 6Q0/77Q6 ,QJ2 #-JKO/28K J+K6 Barbara Raymond
3O4+6+8K:/ /2/8K-7 #/+6-J QOOK88//-Q O-,-Q62/NN
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Nanotechnology Search Committee
"/:K/; Q0 +44NK-+8KQ27 ;KNN ,/1K2 KOO/.K+8/N[ +2. ;KNN -Q28K29/ 928KN Department of Pharmaceutical Chemistry
8J/ 4Q7K8KQ2 K7 0KNN/.
'3O/2 +2. 92./66/R6/7/28/. University of California, San Francisco
OK236K8K/7 +6/ 786321N[ /2-396+1/. 83 +RRN[
600 16th Street, MC 2280
Genentech Hall, Room 518
San Francisco, CA 94158-2517
UCSF is an Affirmative Action/Equal Opportunity Employer. The
362/NN %2K:/67K8[ K7 +2 00K6O+8K:/ -8K32 University undertakes affirmative action to assure equal employment
59+N 4436892K8[ O4N3[/6 +2. .9-+836
opportunity for underutilized minorities and women, for persons with
J884-J6Q2K-N/
-QOLQ,746Q0KN/7
J8O disabilities and for covered veterans.
FACULTY
POSITIONS IN
CELLULAR AND
Eawag is the Swiss Federal Institute of Aquatic Science
MOLECULAR
IMMUNOLOGY
and Technology, a Swiss-based and internationally active re-
search institute within the ETH domain, committed to an
ecological, economical and socially responsible management
of water (Eawag; http://www.eawag.ch). We have an open
position for an outstanding individual as The Immunology Program at the H. Lee Moffitt Cancer Center &
Research Institute and the University of South Florida’s College of
Head of the Department of Medicine, Department of Interdisciplinary Oncology, are seeking
highly qualified (PhD or MD) applicants for tenure track positions
Systems Analysis, Integrated in Cellular and Molecular Immunology at Assistant, Associate
and Full Professor levels. While applicants in all areas of Cellular
Assessment, and Modelling and Molecular Immunology may apply, we are especially interested
in individuals with molecular approaches and signal transduction
The mission of the department is to develop and apply
models of natural, technical and social systems to improve
mechanisms related to studies of T cells, chemokines, and the
our understanding of these systems, of the consequences tumor microenvironment. Successful applicants will be expected to
of environmental policies, and to support environmental develop an outstanding research program in their area of interest.
decision making.
Assistant Professor must have at least four years of postdoctoral
The department supports 6 research groups with research experience in tumor immunology and high quality publications in
topics in systems analysis and surface water modelling; peer-reviewed journals. The Associate/Full Professor must have a
modelling of soil, groundwater and watersheds; material proven track record of independent research and demonstrated
flows in the anthroposphere; modelling of social systems; sustained extramural funding. In addition, the Associate Professor
water environment and food security; and integrative mo- rank requires at least five years of experience with continuing and
delling and decision analysis. The researchers in the depart- productive service as an Assistant Professor. The Professor rank
ment strongly collaborate with other departments of Ea- requires documentation of national recognition, leadership ability
wag and internationally. Many of them teach at ETH Zürich and at least five years of experience with continuing and productive
or the University of Zürich and are involved in continuing service as an Associate Professor. Salary is negotiable.
education programs for water managers.
The Moffitt Cancer Center and Research Institute provides an
The successful candidate is expected to head the depart-
exceptional environment for basic and translational research in
ment and establish a research group on integrative model-
ling and decision analysis. She or he should have
Immunology, Molecular Oncology and Drug Discoveries. Extensive
state-of-the-art core facilities are available for flow cytometry, gene
• An excellent scientific record in environmental systems profiling, proteomics, mouse model development, high throughput
analysis and modelling. screening/chemistry, and drug discovery. Successful applicants will
• Capability and willingness to lead a multidisciplinary be provided generous laboratory and office space in the new
research department in a stimulating environment Vincent A. Stabile Research Building.
of water research.
• A sound knowledge of decision analysis and statistics. Please reference position 11851. Send curriculum vitae
• Experience in leading a research group in these fields and and a brief statement of major academic interests in one
potential to attract external funding from competitive single pdf document to The Immunology Search Committee
sources. at [email protected]. Application review begins
• Experience in management of significant research- and December 1, 2006. Applications will be accepted and
consulting projects. continuously reviewed until the position is filled.
• Willingness to cooperate in inter- and multidisciplinary
projects. USF Health is committed to increasing its diversity and will give
• Experience in academic teaching and continuing education. individual consideration to qualified applicants for this position
• Strong and integrative organizational and communication with experience in ethnically diverse settings, who possess
skills.
varied language skills, or who have a record of research that
As a top research institute in aquatic sciences, Eawag pro-
supports/benefits diverse communities or teaching a diverse
vides excellent facilities for high quality research, provides student population.
a stimulating interdisciplinary research environment, and
operates a child care centre. It hosts over 100 PhD students
(mainly from ETH Zurich) which conduct research under the
direction of Eawag faculty. Strong collaboration with ETH
Zurich, which is one of the leading European universities,
contributes to the attractive work environment. Zurich
ranks among the cities with the highest quality of life world-
wide.

For questions please contact Prof. Peter Reichert

([email protected]). Your application letter, including CV,
publication list, research interests as well as names and
addresses of 5 referees, should be sent to:
Jadranka Vögelin, Human Resources, Eawag,
CH-8600, Dübendorf, Switzerland, or per e-mail
([email protected]). Applications should be The University of South Florida is an EO/EA/AA Employer.
For disability accommodations, contact Kathy Jordan at
submitted by Jan. 31 2007, (813) 632-1451 a minimum of five working days in advance.
interviews are planned for early March 2007. According to FL law, applications and meetings
regarding them are open to the public.

Eawag: Das Wasserforschungs-Institut des ETH-Bereichs

www.moffitt.usf.edu
POSITIONS OPEN POSITIONS OPEN POSITIONS OPEN
ENDOWED CHAIR/HISTORY OF SCIENCE
Thomas Hart and Mary Jones Horning
Professorship
Oregon State University (OSU) invites nomina-
tions and applications for appointment to the Thomas
FACULTY POSITION IN BIOCHEMISTRY Hart and Mary Jones Horning Professorship in the The Yale University School of Medicine is seeking
AND MOLECULAR BIOLOGY Humanities. This endowed Chair is in the Depart- faculty at the ASSISTANT or ASSOCIATE PRO-
University of Nebraska Medical Center ment of History and will be filled by an Historian of FESSOR level with independent research programs
The Department of Biochemistry and Molecular Science in any specialty. The holder of the Chair is in basic research pertinent to all areas of kidney bi-
Biology invites applications for two tenure-leading expected to be a senior scholar with a distinguished ology or disease. Research interest in glomerular
positions at the rank of ASSISTANT/ASSOCI- record of publication and teaching in the history of disease, kidney development, or translational science
ATE PROFESSOR. Qualifications include a Ph.D. science. Appointment to the Chair will carry tenure are encouraged. Laboratory space, startup package,
or M.D. degree and relevant postdoctoral experi- and a position of Professor in the Department of and protected time commensurate with needs are
ence. Preference for both positions will be given to History. The estate of the late Benjamin Horning available in the newly opened Anlyan Center. Suc-
outstanding candidates who can utilize state-of-the- established an endowment at Oregon State Univer- cessful applicants should be Board-certified/Board-
art biochemical and molecular approaches to cancer sity for the purpose of supporting the Humanities. eligible in nephrology. Demonstrated success in
research, proteomics, and chromatin remodeling. Dr. Horning_s intent was to improve and extend the competing for extramural funding is desirable. Please
The ideal candidates will have research interests and teaching of humanities to students in the sciences reply with curriculum vitae, a description of the re-
experience that will supplement ongoing research in and technical areas offered by Oregon State Univer- search program, and the names of three references by
the Department (website: http://www.unmc.edu/ sity. Visit our website: http://oregonstate.edu/ February 1, 2007, to:
Biochemistry/). cla/history/. Stefan Somlo, M.D.
The successful applicant will be expected to have Nominations and applications should be sent to Chief, Section of Nephrology
or develop a funded, independent research pro- the: Chair of the Horning Search Committee, Paul Yale University School of Medicine
gram and to contribute significantly to the teach- Farber, Oregon State University, Department of P.O. Box 208029
ing programs of the Department. Submit your History, 306 Milam Hall, Corvallis OR 97331- 333 Cedar Street
curriculum vitae, a description of research interests 5104. Nominations and application should consist New Haven, CT 06520-8029
and teaching experience, and three letters of refer- of a letter of interest (or a letter describing the nom- E-mail: [email protected].
ence to: Surinder K. Batra, Ph.D. Chair, BMB inee), curriculum vitae, and the names of three to Yale University is an Affirmative Action/Equal Opportunity
Search Committee, Department of Biochemistry/ five references. References will not be contacted Employer.
Molecular Biology, 985870 Nebraska Medical until the Horning Search Committee has confirmed
Center, Omaha, NE 68198-5870. Also e-mail a interest from the nominee. To ensure full consider-
PDF or MSWord file of the above documents to ation, applications (or nominations) must be received
e-mail: [email protected]. University of Nebraska by January 31, 2007. OSU is an Affirmative Action/Equal ASSISTANT or ASSOCIATE PROFESSOR
Medical Center is an Equal Opportunity/Affirmative Action Opportunity Employer. of HORTICULTURE
Employer. Minorities and women are encouraged to apply.
The Department of Horticulture at the University
of Kentucky is seeking a creative and talented in-
FACULTY POSITION TWO FACULTY POSITIONS dividual to fill a 12-month, Tenure-Track position in
Department of Applied Science Dartmouth Medical School research (80 percent) and teaching (20 percent).
College of William and Mary The Angiogenesis Research Center at Dartmouth The Department is interested in filling the position
The Department of Applied Science at the Col- Medical School is seeking to recruit two new tenure- with an individual with demonstrated potential to
lege of William and Mary, an interdisciplinary Ph.D.- track faculty members (M.D., Ph.D., or M.D./Ph.D.) establish a nationally recognized research program in
focused Department established in 1995, invites The appointments will be made at the ASSISTANT horticulture. The person in this position will be ex-
applications for a tenure-track position at the ASSIST- PROFESSOR/ASSOCIATE PROFESSOR/ pected to secure extramural funding to support re-
ANT PROFESSOR level in biophysics, neuro- PROFESSOR level depending on qualifications search, to mentor graduate students, and to foster
physiology, biomedical engineering, biomaterials, and experience. We seek individuals with a strong research collaboration. This position will contribute
or a related field, emphasizing either computational record of academic productivity and the potential to to undergraduate education through academic stu-
or experimental approaches. The new faculty mem- establish or bring an independent research program dent advising and teaching a formal horticulture
ber will be expected to establish a vigorous, indepen- focusing on vascular development, immunology, course. Individuals with diverse horticulture and/or
dent, and well-funded graduate research program at genetics, or signaling. An expertise in zebrafish or plant science-related backgrounds are encouraged
the interface of the physical, mathematical, and bio- Xenopus research is particularly welcome. The suc- to apply. A Ph.D. in horticulture or related plant
logical sciences. Excellence and high commitment to cessful candidate will be expected to actively engage science is required. Candidates must be able to
the teaching of graduate and undergraduate students in teaching in both the Experimental Molecular demonstrate evidence of research excellence and the
is expected of all faculty at the College. Located two Medicine (PEMM) graduate program and within ability to communicate effectively. Apply online at
hours south of Washington, D.C. in Williamsburg, the Angiogenesis Research Center. State-of-the-art website: http://www.uky.edu/HR/UKjobs/,
Virginia, the College of William and Mary is the facilities within the Angiogenesis Research Center using the position title above. Review of applications
second oldest University in the United States and include an advance microscopy core and a core for will begin January 31, 2007, and continue until a
was recently named by the editors of Newsweek as the generation and analysis of new mouse models suitable applicant is identified. For more information
the Bhottest small state school[ in the nation. Can- and mouse imaging. Additionally, Dartmouth Med- contact Dr. Dewayne Ingram at telephone: 859-
didates should submit complete curriculum vitae, ical School offers many facilities that include micro- 257-1758 or e-mail: [email protected].
research statement, and copies of no more than five CT and micro-PET imaging, and genomics and
refereed publications to: Faculty Search Committee, proteomics cores. Please e-mail your curriculum vitae,
Department of Applied Science, The College of a description of your research program, career goals, FACULTY POSITION
William and Mary, P.O. Box 8795, Williamsburg, and the names of two to three references to e-mail: Cognitive Psychology
VA 23187-8795, and arrange to have three letters of [email protected] or mail to:
recommendation mailed to the same address. Review Dr. Michael Simons, Director, Angiogenesis Re- The Department of Psychology at Columbia Uni-
of materials is expected to begin January 1, 2007, and search Center, Dartmouth Medical School, Leb- versity seeks an ASSISTANT PROFESSOR in the
continue until the position is filled. For more in- anon, NH 03756. Dartmouth College is an Equal area of cognitive psychology to begin July 1, 2007.
formation see website: http://as.wm.edu. Opportunity/Affirmative Action Employer and is especially Candidates should provide evidence of excellence in
The College is an Equal Employment Opportunity/Affir- interested in identifying female and minority candidates. research and a strong commitment to both graduate
mative Action Employer. and undergraduate education. Ph.D. in psychology
or related field required at the time of appointment.
Applicants should submit their curriculum vitae, in-
POSTDOCTORAL RESEARCHER POSI- UNIVERSIDAD DE LOS ANDES cluding e-mail address, copies of relevant papers, and
TION to study E. coli AraC family transcription Faculty Positions arrange to have three letters of reference sent to the:
activators, including development of antibacterial The Department of Chemistry at the Universidad Cognitive Psychology Search Committee, Depart-
agents against AraC family virulence activators. de Los Andes in Bogot" D.C., Colombia, invites ap- ment of Psychology, Columbia University, 1190
Ph.D. required. Review of applications begins plications for full-time PROFESSORAL POSI- Amsterdam Avenue, MC 5501, 406 Schermerhorn
December 10, 2006, and continues until position TIONS and VISITING PROFESSOR in the areas Hall, New York, NY 10027. We will begin re-
filled. For more information and to apply go to of biochemistry, organic chemistry, and geochem- viewing applications on December 1, 2006, and will
website: https://jobs.ku.edu and search for po- istry. Applicants should have a Ph.D. degree in the continue until the position is filled. For more infor-
sition 00206132. Inquiries to: Dr. Susan Egan, area of interest. Candidates must be committed to mation see website: http://www.columbia.edu/
Department of Molecular Biosciences, University excellence in teaching and research. Applicants should cu/psychology.
Kansas, Lawrence, Kansas, at e-mail: [email protected]. submit detailed curriculum vitae and make arrange- Applications from minorities and women are encouraged.
University of Kansas is an Equal Opportunity/Affirmative ments to have recommendation letters sent to e-mail: Columbia University is an Affirmative Action/Equal Oppor-
Action Employer. [email protected]. tunity Employer.

1492 1 DECEMBER 2006 VOL 314 SCIENCE www.sciencecareers.org

Max-Planck-Gesellschaft
Max Planck Society
Call for Proposals
BMBF Competition GO-Bio
Group Leaders Biotechnology
Selbstständige
The German Federal Ministry of Education and Research (BMBF)
provides the opportunity to build up independent research groups
for outstanding scientists from Germany and abroad. The main
Nachwuchsgruppen
Independent Junior
objective is to work on innovative, applied research oriented topics
in the biosciences fields and to translate the inventive research
activities into new entrepreneurial initiatives .
Besides a convincing scientific concept for new approaches to
biosciences, candidates must present a promising strategy for Research Groups
application and commercialization of the outcomes. Additionally
applicants need a German research institution to host and support
their independent research group. Depending on the proposed
concept the research group may consist of 1 group leader,
The Max Planck Society invites applica-
6 scientific members and 2 technical assistants . tions from outstanding young scientists
Funded Projects are identified in a two-step procedure by a jury. in all fields of research pursued by the
Successful candidates and their teams will be funded by grants
for an initial period of up to 3 years. Depending on a successful Max Planck Society (Biology and
progress, the project can be extended for a maximum of 3 years.
Medicine; Chemistry, Physics and
The closing date for project outlines is January 15, 2007
Contact:
Technology; Human Sciences).
Dr. Ralf Jossek, e-mail: [email protected]
www.fz-juelich.de/ptj/go-bio
Successful applicants will have demon-
strated the ability to perform excellent
research. They will be offered an
Independent Junior Research
WORKING AT THE
UNIVERSITY OF GENEVA
Group Leader position
(W2; equivalent to associate professor
level without tenure) including a five-
The FACULTY OF SCIENCE seeks a
year grant (research positions, budget,
FULL or ASSOCIATE PROFESSOR investments) at a
in Molecular Biology
(Professeur-e ordinaire ou Professeur-e adjoint-e) Max Planck Institute of their
POST: Full-time research and teaching position in the general
choice.
area of molecular biology. Special consideration given to
scientists studying important biological problems using novel Applications should include a CV, a list of
chemical, genetic or biophysical approaches.
publications, copies of three publica-
REQUIREMENTS: Ph.D. degree or equivalent. Experience in tions, a one-page summary of scientific
teaching and leading an independent research project.
achievements, and a two-page research
er
STARTING DATE: 1 July 2007/at the earliest. plan. For further information and detailed
application instructions see
Candidates files must be addressed before February 1st,
2007 (extension of the previous deadline) to : Décanat de la
Faculté des sciences, 30, Quai Ernest-Ansermet, CH-1211 http://www.snwg.mpg.de
Genève 4, from whom additional information can be obtained
regarding the responsibilities of the post and other conditions.
The Max Planck Society is committed to
The University of Geneva is an equal opportunity employer equal opportunities and to employing
and encourages applications from female candidates. disabled persons.
The deadline for application is January
10, 2007.
POSITIONS OPEN POSITIONS OPEN POSITIONS OPEN
FACULTY POSITIONS IN ATMOSPHERIC The Biological Sciences Department at California
SCIENCE State Polytechnic University, Pomona, invites applica-
The MIT Department of Earth, Atmospheric and tions for a tenure-track, ASSISTANT or ASSOCI-
Planetary Sciences seeks applicants for two faculty ATE PROFESSOR position, in cell and molecular
POSTDOCTORAL FELLOWSHIPS biology, beginning September 2007. Candidates are
positions in atmospheric science. One position is in Institut Pasteur, Paris, France
atmospheric chemistry, the second is in other areas required to have research and development experience
of atmospheric science. Founded in 1887 by Louis Pasteur and located in in biotechnology or the pharmaceutical industry and
POSTDOCTORAL
the heart FELLOWSHIPS
of Paris, the Institut Pasteur is a world-
Atmospheric Chemistry: Areas of specific interest will be expected to participate in the undergraduate
include multiphase (gas, aerosol, cloud) chemical renowned Institut Pasteur,organization.
private research Paris, France The Pasteur biotechnology major program. A Ph.D. in biology,
Founded
Foundation in 1887outstanding
is seeking by Louis Pasteur and Appli-
Fellowship lo- molecular biology, or related fields is required and
and physical processes, and the multiple roles of cated in the heart
atmospheric chemistry in climate. Our preference is cants. Candidates mayofapply
Paris,tothe
anyInstitut Pasteur
laboratory within postdoctoral experience is preferred. We will consider
for a scientist with strong laboratory and/or field 10 is a world-renowned
departments: private
cell biology research develop-
and infection; orga- cell and molecular biology candidates with a broad
nization.
mental Thegenomes
biology; Pasteur Foundation
and genetics;is immunolo-
seeking range of interests, but applicants with stem cell re-
measurement experience but scientists with out-
standing theoretical and modeling experience ap- gy;outstanding
infection Fellowship Applicants. microbiology;
and epidemiology; Candidates search are especially encouraged to apply. The suc-
may apply to
neuroscience; any laboratory
parasitology within 10 depart-
and mycology; structural cessful candidate will combine excellence in teaching
plied to field measurements are also encouraged to ments: cellchemistry;
biology and infection; developmental
apply. Depending on accomplishments and experience, biology and and virology. See website for with an externally funded research program that will
biology; genomes and genetics; immunology;
details. involve undergraduate and Master_s students. Teach-
the appointment can be at ANY LEVEL, including
FULL PROFESSOR. The successful candidate will
infection andare
Fellowships /60,000 permicrobiology;
epidemiology; year for threeneu-
years ing responsibilities will include courses in cell and
have an outstanding record of accomplishment in
roscience;
(/45,000 stipend plus /15,000).
parasitology and mycology; struc-
U.S. citizenship molecular biology, and development of a graduate-
tural biology
required. andFebruary
Deadline: chemistry;2, and
2007. virology. See level course related to the individual_s area of exper-
their discipline, a strong commitment to teaching website [email protected].
for details.
and student advising, and an abiding interest in E-mail: tise. Cal Poly Pomona is a comprehensive Master_s
Fellowships
Website: are /60,000 per year for three
http://www.pasteurfoundation.org. level University with a diverse student body. The suc-
relating their work to complementary work in the years (/45,000 stipend plus /15,000). U.S. citi-
atmospheric and climate sciences at MIT. Joint cessful candidate will have demonstrated ability to be
zenship required. Deadline: February 2, 2007. responsive to the educational equity goals of the Uni-
appointments with other MIT departments are also E-mail: [email protected].
potentially negotiable where appropriate. versity and its increasing ethnic diversity and inter-
Website: http://www.pasteurfoundation. national character. Applicants should forward (1)
Atmospheric Science: We seek individuals with a org.
strong background and interest in atmospheric phys- curriculum vitae, (2) statement of teaching philoso-
ics, dynamics, synoptic meteorology, and/or climate phy, (3) proposed plan of research, (4) reprints of
science. Candidates should have a thorough under- three representative publications, and (5) the names
standing of theory and a desire to build a top-quality and contact information of three references to: Chair,
research program which can link to ongoing projects Cell and Molecular Biology Search Committee,
NEW FACULTY POSITIONS Biological Sciences Department, California State
in the Department. We are particularly interested in Department of Pharmacology, Toxicology,
individuals with a strong commitment to research, Polytechnic University, 3801 West Temple Avenue,
and Therapeutics Pomona, CA 91768-4132. Review of applications
teaching, and graduate advising. Strong preference University of Kansas Medical Center (KUMC)
will be given to candidates at the junior faculty level. will begin on January 22, 2007. Official transcripts
To apply to either of these positions, please send The Department of Pharmacology, Toxicology, and three letters of reference will be required of all
your curriculum vitae, a statement of your research and Therapeutics under the direction of Curtis finalists. For further information, visit the Depart-
and teaching objectives, and the names of five po- Klaassen, Professor and Chair (website: http:// ment website: http://www.csupomona.edu/
tential references to: Professor Maria Zuber, Head, www.kumc.edu/pharmacology/), is continuing its Èbiology. As required by the Clery Disclosure
Department of Earth, Atmospheric and Plane- expansion by inviting applications for two ASSIST- Act, the University_s annual security report is avail-
tary Sciences, MIT, Cambridge, MA 02139; and ANT PROFESSORS, tenure-track faculty positions able at website: http://www.csupomona.edu/
to e-mail: [email protected]. MIT is an Equal Opportunity/ to augment the strength of our eight recent hires. Èpublic_safety.
Affirmative Action Employer. Applications from women, mi- Preference will be given to candidates in areas such as California State Polytechnic University, Pomona, is an
norities, veterans, older workers, and individuals with disabil- nuclear receptors, toxicology, or xenobiotic dis- Equal Opportunity, Affirmative Action Employer. Cal Poly
ities are strongly encouraged. position (absorption, distribution, metabolism, ex- Pomona subscribes to all state and federal regulations and pro-
cretion) that complement existing strengths in the hibits discrimination based on gender, race, sexual orientation,
Department and the Medical Center. This expansion national origin, disability, marital status, age, religion, or vet-
is supported by a new Centers of Biomedical Re- eran status. The University hires only individuals lawfully au-
FACULTY POSITIONS
search Excellence (COBRE) grant entitled Nuclear thorized to work in the United States.
IN BIOLOGICAL SCIENCES
Receptors in Liver Function and Dysfunction, a re-
The Department of Biological Sciences at Mis- cently renewed training grant in environmental
sissippi State University (website: http://www. sciences, and a new research building. Broad areas
msstate.edu/dept/biosciences) invites applications ENVIRONMENTAL GEOPHYSICS
of strength at the Medical Center include cancer,
for ASSISTANT PROFESSOR tenure-track posi- neuroscience, reproductive biology, renal pathophys- The Vancouver Campus of Washington State
tions that begin August 16, 2007. These faculty mem- iology, and growing efforts in liver biology. A University (WSU) invites applications for a full-time,
bers will contribute to one of three focus areas: cell competitive startup package and appropriate space tenure-track ASSISTANT PROFESSOR in envi-
biology/genetics, ecology/evolution or microbiology/ will be offered. Standard support facilities are present, ronmental geophysics. Area of emphasis is open, but
immunology. The scientific infrastructure at Missis- including biotechnology, transgenics, proteomics, candidates with expertise in surface or shallow sub-
sippi State University includes focus areas in pro- and a state-of-the-art imaging center. The Depart- surface processes, and who complement the strengths
teomics, genomics and digital biology, along with ment also has excellent molecular biology (robot, real of existing science faculty, are strongly encouraged
these supporting facilities: the Life Sciences and time PCR, sequencer), and liquid chromatography/ to apply. Successful applicant will teach two courses
Biotechnology Institute (website: http://www. mass spectrometry facilities. Applications will be per year, advise both graduate and undergraduate
mafes.msstate.edu/biotech), the Electron Micro- reviewed as they are received until the positions are students, and establish a productive, externally funded
scope Center (website: http://www.msstate.edu/ filled. Anticipated appointment date is as early as research program. Excellence in research and in-
dept/emc) and the GeoResources Institute (website: July 1, 2007. Applicants must be proficient in the struction are the main criteria for selection. Mini-
http://www.gri.msstate.edu). Successful candi- use of the English language. Applicants should mum qualifications: Ph.D. in geophysics-related
dates will develop externally funded research pro- provide curriculum vitae, statement of research in- discipline by date of hire. Preferred candidates will
grams in any of the above-mentioned areas, direct terests, and names of three references. To review the demonstrate a commitment to working with diverse
graduate students, and contribute to the teaching position description and apply online go to website: student and community populations. WSU Vancou-
mission of the Department. Minimum requirements http://jobs.kumc.edu and search for position ver is located across the Columbia River from Port-
include a Ph.D. in a related biological sciences field, J0020073. Paid for by KUMC. The University of Kansas land, Oregon, and offers significant opportunities
but all-but-dissertation candidates will be consid- Medical Center is proud to be an Equal Opportunity/Affir- for research and an excellent quality of life. Ad-
ered. To apply, send curriculum vitae, reprints of mative Action Employer. ditional information is available at website: http://
three representative publications, a concise statement www.vancouver.wsu.edu/programs/sci/. Appli-
of current and future research interests (one page), cants should submit a cover letter, curriculum vitae,
and identify the position/area you are applying for, statement of research interests and accomplish-
plus relevant areas of teaching competence. Applicants POSTDOCTORAL FELLOW to work on a ments, statement of teaching philosophy and in-
should also arrange for at least three letters of palate development. Expertise in cell signaling, tissue terests, copies of two publications, and three letters
reference to be submitted on their behalf. Screening culture, biochemical, molecular biological, and mor- of reference to: Environmental Geophysics Search,
will begin January 15, 2007, and will continue until phological techniques required. Submit curriculum Washington State University Vancouver, 14204
the positions are filled. Send application packets to: vitae, current research activities, and career goals in N.E. Salmon Creek Avenue, Vancouver, WA
Dr. Nancy Reichert, Interim Head, Department of electronic format to: Human Resources, The Texas 98686-9600. Review of completed applications will
Biological Sciences, P.O. Box GY, Mississippi State A&M University System Health Science Center, begin on January 2, 2007. Washington State University is
University, Mississippi State, MS 39762 (e-mail: 3302 Gaston Avenue, Dallas, TX 75246; e-mail: an Equal Opportunity/Affirmative Action Educator and Em-
[email protected]). Mississippi State [email protected]. Baylor College of Dentistry is an ployer. Members of groups historically underrepresented in science
University is an Affirmative Action/Equal Opportunity Employer. Equal Opportunity Employer. are strongly encouraged to apply.

1494 1 DECEMBER 2006 VOL 314 SCIENCE www.sciencecareers.org

School of University of California, Irvine
Computational Science at DEAN, SCHOOL OF BIOLOGICAL SCIENCES
Florida State University The University of California, Irvine invites applications and nominations for the position of Dean, School of
Biological Sciences. The university seeks an independent thinker with skills to navigate within a complex/
Faculty Position in multi-constituent organization and who is decisive while fair and strategically focused. The successful
Computational Evolutionary candidate has a keen intellectual capacity and creativity, and is an open and persuasive communicator who
leads from values; a candidate with energy and vision to head and continue the mission of the school. Key
Biology selection criteria will include:
The Computational Evolutionary Biology • Demonstrated leadership in promoting the latest intellectual advances in the Biological Sciences
(CEB) group at the School of Computa- • Experience and demonstrated success with external relations/development - Demonstrated skill/ability to
work effectively with the business community and other constituents in resource development and advance-
tional Science (http://www.scs.fsu.edu)
ment of the School
at Florida State University seeks candi- • Faculty leadership and team building - Proven ability to inspire consensus and develop a culture that is
dates for a faculty position in computa- mutually supportive, goal-directed and ambitious. Ability to attract and recruit world-class faculty
tional evolutionary biology starting in • Administrative management - Demonstrated success as an administrator, including staff development,
Fall 2007. The successful applicant will facility/resource management and fiscal leadership
have joint appointments in the School of
Celebrating 40 years of innovation, the University of California, Irvine is a top-ranked public university dedi-
Computational Science and a secondary cated to teaching, scholarship and community service. Founded in 1965, UCI is among the fastest-growing
department best suited to their research campuses in the University of California system, with more than 24,000 students, 1,400 faculty members and
interests. 8,300 staff. The second-largest employer in dynamic Orange County, UCI contributes an annual economic
We seek candidates at the Assistant Profes- impact estimated at $3.3 billion. It is located on a 1500-acre site three miles from the ocean in the community
of Orange County. The School of Biological Sciences, one of ten schools at the University of California, Irvine,
sor level but exceptional candidates at more
currently has an operating budget of $27.5 million, and extramural grant funds of $38.8 million. The School
senior levels will also be considered. We has an enrollment of approximately 3500 undergraduate students and 400 graduate students, 105 full time
are especially interested in applicants who faculty and over 250 additional academic researchers, and 149 staff members. The School represents a premier
are applying computational phylogenetics center for biological education and research with four solid departments: Developmental and Cell Biology,
to model evolutionary processes. This Ecology and Evolutionary Biology, Molecular Biology and Biochemistry, and Neurobiology and Behavior.
includes those developing algorithms to
Candidates for this position should have a strong academic record that would justify a senior rank appointment
estimate ancestral values for molecular/ in one of the four departments with a sustained record of peer reviewed extramural funding and an interna-
morphological traits, population param- tional reputation in a biological sciences discipline. Review of applications will begin on February 1, 2007
eters, and selection/mutation models, and and the position will remain open until filled. We encourage electronic application submission; please click
those working to elucidate the evolution of the online application link to begin the submission process: http://www.evc.uci.edu/searches/index.html.
protein structure/function, gene-networks Paper applications and nominations may be sent to the address below: School of Biological Sciences Dean
and morphological development. We will Search Committee, C/O Heike Rau, 509 Administration, University of California, Irvine, Irvine, CA
also consider exceptional applicants whose 92697-1000; OR email: [email protected].
research centers around algorithm develop- UCI is an Equal Opportunity Employer committed to excellence through diversity and strongly encour-
ment for phylogenomics. ages applications from all qualified applicants, including women and minorities.
This hire builds on existing strengths in
theoretical phylogenetics and population
genetics at FSU and comes at a time when
the university is expanding research in CHAIRMAN, ASSISTANT/ASSOCIATE PROFESSOR
the life sciences through its Pathways of Department of Cell Biology BIOLOGY
Excellence Cluster hiring initiatives. The University of Oklahoma Health Compensation: $38,001 - $79,220
department of Biological Science is cur- Sciences Center Commensurate with qualifications
rently recruiting a new cluster of faculty to Applications/nominations are invited for Chair of and experience
explore the mappings between molecular the Department of Cell Biology at the University College Web Site: www.ccny.cuny.edu
processes and resulting phenotypes (http: of Oklahoma Health Sciences Center. This indi- Notice Number: FY12577
//pathways.fsu.edu/faculty/igp/). This vidual must be committed to the Department’s Closing Date: 12/27/06
expansion will provide excellent oppor- mission of biomedical research in cellular, POSITION DESCRIPTION AND DUTIES
tunities for new collaborations between developmental, and molecular biology, as well The Biology Department at City College invites applica-
the Computational Evolutionary Biology as medical education. The successful candidate tions for a tenure track position in Neuroscience. The
group and members of the Department of will have an M.D., Ph.D., or equivalent doc- candidate must use molecular, genetic or cellular
Biological Science. toral degree, qualify for tenured appointment as approaches to problems in neurobiology. The candidates'
Professor, and be an internationally recognized research should complement the research interests of
A Ph.D. in one of the sciences is required. leader in his/her area of research. Candidates existing faculty; these include cellular and molecular
Postdoctoral experience is highly desired. with interests in cancer biology and diabetes aspects of development and behavior, bird-song develop-
The new faculty member will be expected are particularly sought, given the Department’s ment, sensory motor integration and plasticity, neuro-
physiology and anatomy of the visual cortex, and visual
to participate in MS/PhD degree programs participation in the ongoing expansion of the control of eye growth. City College has an active and
in both Computational Science and their OU Cancer Institute and the Oklahoma Dia- expanding neuroscience community, which include the
secondary department, to have an active betes Center. Leadership positions in the OUCI departments of Biomedical Engineering, Psychology and
research program and to be involved in are available for qualified applicants. With over Physiology/Pharmacology.
teaching in both departments. $180 million in public and private support, OUCI QUALIFICATION REQUIREMENTS
represents the largest investment in biomedical Doctorate required. Candidate must demonstrate a
Those interested in being considered for research in the state’s history. Please send a letter strong interest and commitment to undergraduate teach-
the position should apply electronically to of application, curriculum vitae, the names and ing and the capability of developing and maintaining an
http://www.scs.fsu.edu/jobs.php. Appli- contact information for five references, plus a active research program supported by external funding.
cations received by January 1, 2007 are one-page summary that includes teaching phi- TO APPLY: PLEASE SEND COVER LETTER, CURRICU-
assured of full consideration. losophy and goals for maintaining and expand- LUM VITAE, NAMES OF THREE PROFESSIONAL REFER-
ing the Department to: Dr. Robert Foreman, ENCES, TO: Ofer Tchernichovski, Chairman,
Applications require electronic submis- Chair, Cell Biology Chair Search Committee, Neuroscience Search, Biology Dept., Room J526,
sion of a Curriculum Vitae, research and The City College of New York, 160 Convent Avenue,
The University of Oklahoma, BMSB 653, 940 New York, NY 10031
teaching statements (PDF files preferred) Stanton L. Young Blvd., Oklahoma City, OK,
The City University of New York is an Equal Employment
and the names of four references. Inquiries 73104 (http://w3.ouhsc.edu/cell%5Fbiology/). Opportunity/Affirmative Action/Immigration Reform and Control
concerning the position should be sent to: The review of applications will begin immedi- Act/ Americans with Disabilities Act Employer
[email protected]. ately and continue until the position is filled.
FSU is an EO/AA Employer committed to The University of Oklahoma is an Equal
diversity in hiring. Opportunity Institution.
POSITIONS OPEN POSITIONS OPEN POSITIONS OPEN
DIRECTOR
Stanley S. Scott Cancer Center
Louisiana State University Health Sciences Center
New Orleans FACULTY POSITION
The Louisiana State University Health Sciences Molecular and Cellular Pharmacology
PROFESSOR, PHARMACOGENOMICS Center (LSUHSC), School of Medicine in New The Department of Molecular and Cellular Phar-
The University of Miami Miller School of Medi- Orleans, Stanley S. Scott Cancer Center, is accepting macology at the UniversityPOSITION
FACULTY of Miami, Miller School
cine is fostering major expansion of its genetics applications for the Director of the Cancer Center. of Medicine is seeking
Molecular applications
and Cellular for a TENURE-
Pharmacology
research program. The Department of Psychiatry The position requires an M.D., Ph.D., or both, with TRACK FACULTY POSITION
The Department of Molecular (rank
andopen).
CellularCan-
and Behavioral Sciences is under new leadership combined M.D., Ph.D. degrees preferred. Prefer- didates
Pharmacology at the University of degree
must have a Ph.D. and/or M.D. Miami,and
(Julio Licinio, M.D., Chairman) and it is under- ence will be given to candidates with experience in have an established
Miller record ofisresearch
School of Medicine excellence.
seeking applica-
going significant growth and is opening a new Cen- working within or in developing a successful plan for Applicants
tions forfrom all areas of molecular/cellular
a TENURE-TRACK FACULTY bi-
ter on Pharmacogenomics. The Center is focused on an NCI designated cancer center. The ideal incum- ology and biomedical
POSITION researchCandidates
(rank open). are welcome.mustThe
understanding pharmacogenomics mechanisms bent must demonstrate scholarly experience as evi- newhavefaculty member
a Ph.D. and/or will
M.D.complement
degree andexisting
have an re-
underlying clinical response and identifying new denced by academic accomplishments, publications, search efforts in
established the Department.
record Rank andAp-
of research excellence. salary
targets for drug development. We are recruiting for service on national study sections, service on edito- willplicants
be commensurate
from all areas withof experience. Generous
molecular/cellular
a new faculty member at the PROFESSOR level rial boards, and a track record of extramural research laboratory
biology space and startupresearch
and biomedical funds are
areavailable.
welcome.
with the ability to develop research programs in funded through NCI or other NIH funding mech- Applicants
The new shouldfaculty send electronic
member will and hard copies
complement
molecular genetics, genomics, and in genetics re- anisms. Administrative experience with building either of existing
their curriculum vitae, statement of
research efforts in the Department. research in-
lated to antidepressant response. Outstanding back- cancer research programs or cores in an academic terests
Rankandanddirection,
salary will and contact information
be commensurate with ex- for
ground with publications, grant track record, and setting is preferred. The incumbent should qualify for three references,
perience. to e-mail:
Generous [email protected]
laboratory space and startup
mentoring experience is required. We seek a Senior appointment at the level of Professor. A track record (e-copies)
funds areand Dr. James D. Potter, Search Com-
available.
Investigator with a documented track record who in basic, translational, epidemiological, and/or clin- mitteeApplicants
Chair, Department
should send of Molecular and and
electronic Cellu-
can develop new and independent funded lines of ical research is required. Knowledge of current clinical lar hard
Pharmacology,
copies of their University
curriculum of vitae,
Miamistate-
Miller
research in pharmacogenomics applied to depression oncology operations in relation to academic medicine School
mentof of Medicine, P.O. Box
research interests and 016189,
direction, Miami,
and
and antidepressants. Competitive startup package will be viewed positively. The ideal candidate should FLcontact
33101.information
An Equal Opportunity/Affirmative
for three references, Action
to
available with excellent benefitsSend curriculum have demonstrated ability in developing translational Employer.
e-mail: [email protected] (e-copies) and
vitae, contact information for three references to: research from the basic scientific observations to clin- Dr. James D. Potter, Search Committee
Julio Licinio, M.D., Chairman, Department of ical trials or clinically relevant research. Chair, Department of Molecular and Cellular
Psychiatry and Behavioral Sciences, University of The incumbent will be appointed to the appro- Pharmacology, University of Miami Miller
Miami Miller School of Medicine, e-mail: licinio@ priate clinical department and will be a member of School of Medicine, P.O. Box 016189,
miami.edu. The University of Miami is an Equal Oppor- the Stanley S. Scott Cancer Center. Also the incum- Miami, FL 33101. An Equal Opportunity/Affir-
tunity/Affirmative Action Employer. bent will be Co-Director of the Louisiana Cancer mative Action Employer.
Research Consortium of LSU and Tulane. Please
send curriculum vitae including current grant fund-
ASSISTANT PROFESSOR
ing, a brief cover application letter detailing profes-
Molecular Microbiology/Immunology The Office of Science, Department of Energy is
sional interests and goals, and the names of three
The Department of Biological Sciences at Purdue seeking a motivated and highly qualified individual
references to: Bernard Wan, Dean_s Office, School
University Calumet invites applications for a tenure- to serve as the ASSOCIATE DIRECTOR, Office
of Medicine, 533 Bolivar Street, New Orleans,
track position to begin in August 2007. A Ph.D. of Biological and Environmental Research. As such,
LA 70112 with an e-mail: [email protected].
degree, postdoctoral experience, and excellent verbal/ you will provide leadership and direction in estab-
LSUHSC is an Affirmative Action/Equal Opportunity
written communication skills are required. Respon- lishing vision, strategic plans, goals, and objectives
Employer.
sibilities will include (1) teaching undergraduate and for the research activities supported. You may apply
graduate courses, which may include introductory through two different methods, one is for a SENIOR
biology, microbiology, immunology, and upper- DEAN, DIVISION OF NATURAL SCIENCES EXECUTIVE SERVICE appointment and the sec-
division courses in areas of expertise, (2) establishing AND MATHEMATICS ond is for an INTERGOVERNMENTAL PER-
a vigorous extramurally funded research program, University of Denver SONNEL ACT appointment. The announcement
involving graduate/undergraduate students, that Applications and nominations are invited for the number is SES-SC-HQ-005. The announcement opens
will complement and enhance current departmental position of Dean of Natural Sciences and Mathe- on November 6, 2006, and closes on December 21,
strengths in molecular biology and biotechnology, matics at the University of Denver (DU). The Dean 2006. Visit website: http://www.usajobs.opm.gov/
and (3) service to the University. Purdue University is the chief academic, administrative and budgetary for more information and for instructions concerning
Calumet, the Chicago-area campus of the Purdue officer of the Division, is responsible for leadership application procedures.
University system, is a M.S. Comprehensive Univer- of all internal programs, as well as external relation- TWO FACULTY POSITIONS IN MICROBIAL
sity with over 9,300 students, of which approxi- ships and constituencies, and reports to the Provost. PATHOGENESIS
mately 425 undergraduate majors and 25 graduate The Division (website: http://www.nsm.du.edu) University of Kansas Medical Center
students are enrolled in the Department of Biolog- consists of the Departments of Biological Sciences,
ical Sciences (website: http://www.calumet.purdue. Chemistry and Biochemistry, Geography, Mathemat- The Department of Microbiology, Molecular Ge-
edu/biology/). Review of applications will begin ics, Physics and Astronomy, the Eleanor Roosevelt netics and Immunology at the University of Kansas
January 15, 2007, and continue until the position is Institute, and the Denver Research Institute. The Medical Center (KUMC) invites applications for two
filled. Inquiries or letters of application stating teaching successful candidate should have: proven admin- tenure-track faculty positions: one at the ASSISTANT
philosophy and research interests, curriculum vitae, istrative experience; strong organizational, commu- PROFESSOR level and one at the ASSOCIATE
copies of graduate/undergraduate transcripts, and nication, leadership and interpersonal skills; the PROFESSOR level. We seek exceptional candidates
three original letters of recommendation with contact ability to interact effectively with all levels of aca- (Ph.D., M.D., or M.D./Ph.D. degree), with docu-
information should be sent to: Dr. W-T Evert Ting, demic administration, faculty, students, and the mented evidence of quality research and a commit-
Chair of Search Committee, Department of Bio- external community; experience in promoting grant- ment to research and teaching at a major research
logical Sciences, Purdue University Calumet, 2200 funded research and fundraising; a doctoral degree in University medical center.
169th Street, Hammond, IN 46323-2094; e-mail: a relevant field; a record of accomplishment in both The successful candidates will be expected to es-
[email protected]. Purdue University teaching and research to merit appointment as a tablish extramurally funded (Assistant Professor) or
Calumet is an Equal Access/Equal Opportunity/Affirmative tenured full Professor in one of the departments of have extramurally funded (Associate Professor) in-
Action Employer. the Division. For a detailed description of the po- dependent research programs focused on bacterial or
sition, and the application process see website: viral pathogenesis. Competitive salaries, startup pack-
http://portfolio.du.edu/NSMSearch. The Univer- ages, state of the art BSL-3 laboratories, animal care
ASSISTANT PROFESSOR, BIOLOGY facilities, and excellent core facilities are available.
Saint Francis University sity of Denver is strongly committed to enhancing diversity. It
encourages applications from women, minorities, people with Applicants apply online for Assistant Professor
Reach higher and go farther with the Department disabilities, and veterans. DU is an Equal Employment Oppor- position J0087656, or Associate Professor J0083042,
of Biology at Saint Francis University, tenure-track, tunity/Affirmative Action Employer. and attach a letter of interest, curriculum vitae, de-
Assistant Professor position opportunity in neuro- scription of research plans and contact information
biology to begin August 2007. For a complete job for at least three references at website: http://
description, application information, and University The Institute of Technology of the University of jobs.kumc.edu. Additional information describing
facts please visit our website: http://www.francis. Minnesota, Twin Cities, invites nominations and the Department can be found at website: http://
edu or refer to website: http://www.sciencemag. applications for HEAD OF THE DEPARTMENT www.kumc.edu/microbiology. To ensure full con-
org/. Saint Francis University is committed to diversity, and OF MECHANICAL ENGINEERING. Complete sideration, applications should be received by Janu-
we encourage candidates who will contribute to meeting that job description and application instructions can be ary 15, 2007, but applications will be accepted until
goal to apply. Applications and nominations of women and found at website: http://www.me.umn.edu/. The the positions are filled. Paid for by KUMC.
minorities are strongly encouraged. Affirmative Action/Equal University of Minnesota is an Equal Opportunity Educator The University of Kansas Medical Center is an Equal Em-
Opportunity Employer. and Employer. ployment Opportunity/Affirmative Action Employer.

1496 1 DECEMBER 2006 VOL 314 SCIENCE www.sciencecareers.org

Science
Careers Forum
Dave Jensen
Indusry
Recruiter

• How can you write a resume that stands out in a crowd?

• What do you need to transition from academia to industry?
• Should you do a postdoc in academia or in industry?

Let a trusted resource like ScienceCareers.org help you

answer these questions. ScienceCareers.org has partnered
with moderator Dave Jensen and four well-respected advisers
who, along with your peers, will ﬁeld career related questions.

Visit ScienceCareers.org and start an online dialogue.

Bring your career

concerns to the table. Dialogue
online with professional career
counselors and your peers.

C OLUMBIA U NIVERSITY
Biomechanics
2006–07
The Department of Biomedical Engineering in the Fu Foundation School
of Engineering and Applied Science at Columbia University is seeking to DEAN
fill a tenure-track faculty position at the Assistant or Associate Professor
level but exceptionally qualified candidates will be considered for a
COLLEGE OF MEDICINE
Burlington, VT
higher-level position. Applicants should have a doctoral degree in
The University of Vermont invites applications and nominations for a Dean
biomedical engineering or a closely related discipline, and should be of its College of Medicine. The committee seeks a Dean who will
prepared to establish a vigorous and independent research program in galvanize the strengths of the College and the University to build one of
any of the broadly defined areas of biomechanics: functional tissue the nation’s premier medical schools. Combining the ethos of a major
research university with the innovative, personalized education of a
engineering, biological systems modeling, molecular modeling, cellular smaller institution, the College excels in research (its $77.3 million in
or molecular biomechanics, biomechanics of growth and remodeling, external funding puts it in the top third of medical schools for NIH funding
per faculty member), teaching, and service. The College delivers patient
biofluid mechanics, tissue mechanics, computer and robot-assisted care to the state and region in partnership with Fletcher Allen Health
surgery, and/or bioMEMS. Care, Vermont’s only academic medical center. Located in Burlington,
Vermont, one of the nation’s “most livable” and beautiful small urban
Applicants should send a complete curriculum vitae, three publication environments, the College is personal, intimate, and independent minded;
reprints, a statement of research interests, and names of four an ideal mix for an intellectually rigorous community.
references by March 1, 2007 to: The Dean will lead in building the research mission of the College, extending
the College’s remarkable success in its teaching programs, partnering
Professor X. Edward Guo with the whole campus on both teaching and research innovations, and
Chair of Biomechanics Search allying seamlessly with Fletcher Allen Health Care to deliver the very
finest medical care to urban and rural Vermont. In short, the College seeks
Department of Biomedical Engineering a Dean who will play a central role in moving UVM toward its aspiration of
Columbia University becoming the nation’s premier small public research university.
351 Engineering Terrace, Mail Code 8904 Applications and nominations should be sent to Philip Jaeger,
Isaacson, Miller, 1875 Connecticut Avenue NW, Suite 710,
1210 Amsterdam Avenue Washington, DC 20009.
New York, NY 10027 Electronic submission of material is strongly encouraged:
[email protected].
The search will remain open until the position has been filled.
In employment as in education, the University of Vermont is committed
Columbia University is an affirmative action/equal opportunity employer. to equal opportunity and affirmative action and seeks candidates with
Women and minorities are encouraged to apply. a proven commitment to diversity. Women and members of underrepresented
groups are encouraged to apply.
POSITIONS OPEN POSITIONS OPEN POSITIONSOPEN
POSITIONS OPEN
TWO POSTDOCTORAL RESEARCH
ASSOCIATE POSITIONS IN
ENVIRONMENTAL NEUROTOXICOLOGY
Two Postdoctoral Research Associate positions
are available beginning December 1, 2006, in the POSTDOCTORAL
POSTDOCTORALPOSITIONS POSITIONS
Insecticide Toxicology Research Laboratory at Cor- The Center Thefor Infectious
Center Disease Dynamics
for Infectious
The laboratory of Dr. Katherine Ferrara at nell University_s New York State Agricultural Ex- (CIDD) Disease
The Pennsylvania State University
University of California, Davis, and the Center for Dynamics (CIDD)
periment Station campus in Geneva, New York. The TheCenterPennsylvania
for Infectious State Disease
University Dynamics
Molecular and Genomic Imaging are seeking a Successful applicants will participate in NIEHS- provides
highly motivated POSTDOCTORAL FELLOW The aCenter
highlyfor collaborative,
Infectious Disease interdisciplinary
Dynamics
funded research to define the mechanisms of in- environment to address challenges in infectious
or PROJECT SCIENTIST to develop radiochem- provides a highly collaborative, interdisciplinary
secticide action on rat and human voltage-sensitive disease research.to There
ical methods to characterize the biodistribution of environment address are immediate
challenges openings
in infectious
sodium channels, map the binding sites for insecti- available for applicants with expertise in the
lipid and polymer-shelled nanoparticles. Research in cides in relation to sites of action of other toxicants
disease research. There are immediate openings
following
our laboratory involves the development of methods availableareas:
for molecular
applicants virology; innateinimmuni-
with expertise the
and drugs, and identify the molecular basis of the ty; following protein
genomics;areas: structure/function;
molecular virology; innate mathe-
im-
to enhance local drug delivery and imaging methods differences in sensitivity to insecticides between mam- matical biology.
for the assessment of drug and vehicle biodistribu- munity; genomics; protein structure/function;
malian sodium channel isoforms and between mam- Please see website:
biology.http://www.cidd.psu.edu
tion and cell trafficking. Ideal candidates will have a malian and insect sodium channels. A Ph.D. degree in
mathematical
for detailed descriptions of these positions and
Ph.D. in organic chemistry or radiochemistry and Please see website: http://www.cidd.psu.
an appropriate biological discipline is required. Pref- application procedures.
experience in synthesizing, designing, and validating edu for detailed descriptions of these positions
erence will be given to applicants with prior training Pennsylvania Stateprocedures.
is committed to Affirmative Action,
probes for nuclear medicine. Experience with posi- and experience in at least one of the following areas:
and application
Equal Pennsylvania
Opportunity State
and the diversity of its workforce.
tron emission tomography (PET) probe synthesis is the development and use of transfected mammalian
is committed to Affirmative Action,
particularly desirable. Development of independent cell lines for the expression and assay of receptors and
Equal Opportunity, and the diversity of its workforce.
research program and funding will be encouraged ion channels; voltage or patch-clamp analysis of ion
and assistance provided. To apply, please send your currents; or, the development and validation of
curriculum vitae, a brief statement of research in- GRANTS
ligand-receptor docking models. Salary will be based
terests, and contact information for five references on NIH postdoctoral compensation guidelines; an BRAIN TUMOR SOCIETY RESEARCH GRANTS
to: Katherine W. Ferrara, Biomedical Engineer- attractive fringe benefits package is included. Send a One-Year $100,000 grants
ing, University of California, Davis, 451 E. letter of application with curriculum vitae and the Two-Year $200,000 grants
Health Sciences Drive, GBSF 2303, Davis, CA names of three professional references to: Professor Available in the United States and Canada
95616, or to e-mail: [email protected]. David M. Soderlund, Department of Entomolo- Letter of Intent Deadline: January 16, 2007
gy, New York State Agricultural Experiment The Brain Tumor Society (BTS) is awarding grants
FACULTY POSITION IN THE BIOLOGY OF Station, Cornell University, 630 West North to fund basic scientific and translational research
HEPATITIS C VIRUSES Street, Geneva, NY 14456 (e-mail: dms6@cornell. directed at finding a cure for brain tumors. Grants
Department of Microbiology-Immunology edu). Cornell University is an Equal Opportunity, Affirma- are awarded annually at a maximum of $100,000 per
Northwestern University tive Action Educator and Employer. year. Grants may be used for startup projects or
Feinberg School of Medicine supplementary funding. Funds cannot be used for
A tenure-track position is open for a full-time fac- indirect costs. Clinical projects will not be funded.
ulty researcher (Ph.D., M.D./Ph.D. or M.D.) study- Letter of intent packets available on website: http://
ing hepatitis C viruses. POSTDOCTORAL FELLOWSHIPS www.tbts.org.
Rank is open, and salary is negotiable. All ap- AVAILABLE
plicants should have substantial peer-reviewed pub- MARKETPLACE
lications that demonstrate research productivity and The Lombardi Comprehensive Cancer Center at
the ability to perform cutting-edge research. Can- Georgetown University, a multidisciplinary NCI-
didates for an ASSISTANT PROFESSOR posi- designated cancer research center with $70 million Diverse Small Molecules
tion should have postdoctoral research experience. in grant support, is currently recruiting POSTDOC- Ready for Screening
Persons seeking appointment as ASSOCIATE PRO- TORAL FELLOWS into positions funded by an
FESSOR should have substantial research produc- NCI training grant. The goal is to develop strong High Quality & ChemBridge
Drug-Like
tivity and a history of grant support and academic basic and translational scientists with an interest in Corporation
service. Candidates should have an interest in teach- cancer research. Successful applicants will choose a Pre-Plated in DMSO
ing graduate and medical students. Starting date is mentor from an interdisciplinary group of investiga-
Very Competitively
negotiable after September 1, 2007. Application ma- tors who are committed to cancer research. Research Priced
terials will be reviewed as received but, to receive full programs include: Development of novel anticancer
therapies; the genetic and molecular mechanisms of Upwards of 200,000 Website: www.chembridge.com
consideration, should be received by February 1, Compounds Email: [email protected]
2007. Please send complete curriculum vitae and the malignant progression; the role of growth factor sig-
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suitable candidate is selected. Applicants should send Association, 293: 90, 2005; Histology Histopathology,
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three to five references to: Joel Trexler, Search Com- Please forward curriculum vitae and three references
mittee Chair, Department of Biological Sciences, to: Kenneth Maiese, M.D., Neurology, 8C-1 UHC,
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