4. Epidemiology of mutations: Describe age of father effect.

5. Compare and contrast repair mechanisms employed by cells and identify in which cell
cycle stage each occurs.

6. Recognize diseases and syndromes caused by faulty repair, explain mechanisms, and
causes.

Mutations and DNA Repair lecture notes

DNA lesions that cause changes and are not corrected in the DNA base sequence = mutation
·when a high level of damage is detected, cells stop their division and attempt DNA repair. If
there is too much damage already done then cells will undergo apoptosis (cell death).
·when cells undergo many cell divisions they are more likely to have errors/mutations

Genetic damage (2 types)

·exogenous damage: influences from your environment (UV light, ionizing radiation,
carcinogens (cancer causing)
·endogenous damage: accidental consequences of metabolic processes (oxidation, nitrosylation,
hydrolysis of DNA strands, errors in DNA replication

EX: UV light: pyrimidines dimers can absorb the the UV light making abnormal covalent bonds
called pyrimidine dimers. These dimers then distort the DNA structure by bonding to their
neighboring pyrimidine friends instead of the normal counterparts from other strands.
Pyrimidine dimers alter replication and transcription. Made from UV light!

2. List and explain causes of mutations: where and when?

Substitution mutation: change in one DNA base.

Ex: if guanine were methylated this would block a hydrogen bond causing the cells to see only 2
hydrogen bonds instead of 3 and will insert the wrong base during replication.

Q: (which part of the body are prone to the effects of UV light?)

A: Eyes & skin

Hydrolysis: involves hydrolysis between a purine (usually guanine) and deoxyribose which
leads to an apurinic base or deamination of cytosine because forms into uracil.
-deamination of cytosine
 -deamination of cytosine is due to the change in base pairing: cytosine pairs with guanine
and uracil with adenine. Deamination of cytosine causes problems b/c the deamination
 base is thymine and thymine is a normal base in DNA so the repair system doesn’t know
which strand is wrong. This makes cytosine a hotspot!

Nitrous acid (HNO2) is used in meat as a preservative and anti-bacterial agent. It reacts with
adenine & cytosine and causes deamination: adenine becomes hypoxanthine & cytosine becomes
uracil.

Clinical correlation: Aflatoxin B1

-considered a potent naturally occurring carcinogen causing liver tumors. Aflatoxin B1 is
produced from Aspergillus mold. When aflatoxin mixes with p-450 in liver it can lead to an
activated form of the molecule and adds itself to guanine

1. Compare and contrast different mutation types at nucleotide, gene, protein, and genome
levels.
·Transitions, transversions, indels, larger deletions and duplications
·Silent, missense, nonsense, frameshift, splice site
·Loss of function, gain of function, genomic instability.

Mutation types:
 Transition: purine to purine (A-G) or pyrimidine to pyrimidine (C-T)
 Transversion: purine to pyrimidine (G-T) or (A-C)
 Indel: insertion/deletion of a single nucleotide pair
 Loss of function: mutations leading to proteins that are quickly degraded ( due to
premature stop codons, nonfunctional)
 Gain of function: mutation is due to protein binding to something it shouldn’t bind to
 Point mutation: alternation in single nucleotide
o Silent mutation: point mutation, codes for same amino acid (often tRNA wobble
position/3rd position)

 Ex:
 Still the same amino acid but different code. No effect on protein structure

o Missense mutation: point mutation, leads to different amino acid

 Ex: sickle cell


o Nonsense mutation: incorrect nucleotide = early STOP codon (UGA, UAA,
UAG)

 Ex:
  Frameshift mutation: insertion/deletion of a number of nucleotides. Not a multiple of 3.
Most dramatic mutation b/c it misleading of all codons following a frameshift
“downwards” meaning the entire downstream codon sequence is significantly altered.

o Ex:
o Ex: Proflavin (molecule used as antiseptic against Gram positive bacteria).
Proflavine is a carcinogen b/c it induces frameshift mutations by having an
aromatic structure which makes the molecule flat and this is perfect for insertion
between bases.

 Splice Site mutation: introns are removed and exon sequences splice together. Deleting/
creating new splice sites = consequences to the final codon sequence of mRNA. Meaning
if “AG” (splice acceptor) gets altered the spliceosome will scan for the next “end
sequence” in the pre-mRNA and results in an ENTIRE EXON BEING REMOVED from
the final mRNA. If “GU” (splice donor) gets altered results in an ENTIRE INTRON
BEING INCORRECTLY INCLUDED.

Q: A mutation replaces a Guanine on a DNA strand w/ a Thymine. What is this called?

A: Transversion

3. Compare and contrast the effects of mutations in somatic versus germline cells.
Somatic vs. germline mutations

Somatic Cells Germline Cells

 Occur in single cell  Occur in germline which is special
 Embryo: mutations cannot be passed tissue in sex cells-ova & sperm
on to offspring  Passed down to offspring
 Organisms: occurs in single cell that  Results in diseases = genetically
 divides to produce clones with the inherited. Because they are inherited
same mutation = phenotypical mutated every cell of the individual contains
region of body. the mutation and expression of disease
 If mutation occurs early in  Mutation present in entire organism
development = larger  Ex: NF-1 gene (neurofibromatosis
 Is only seen in the individual & not in type 1): cond. Where patients have
the gametes or inherited by offspring changes in skin pigmentation = café
 Ex: McCune-Albright syndrome au lait spots, tumor growths along
nerves of skin + brain
Consequences of Mutations
·mutations in noncoding regions alter gene expression or induce alternative splice sites
·mutations in coding regions alter codon sequences of mRNAs = mutant proteins

5. Compare and contrast repair mechanisms employed by cells and identify in which cell
cycle stage each occurs.
 Direct Repair: cells attempt to repair damaged nucleotide bases w/o breaking the
phosphodiester bond that holds the nucleotides together. This repair is used to address
spontaneous methylation such as:
o O6-methylguanine methyltransferase (MGMT) -detects & removes anomalous
methyl groups
o Photolyase – an enzyme that brakes pyrimidine dimers in a process called
photorectivation. It cleaves the pyrimidine dimers apart. Humans do not have
photolyase but bacteria, fungi, and plants do!
 Single Stranded Repair: uses complementary strands to repair breaks
o EX: Base Excision Repair:
 Process: GEL Please: Glycosylase creates AP site Endonuclease
cleaves 5’ end & removes nucleotide Lyase cleaves 3’ end DNA
polymerase B adds new undamaged nucleotide to 3’ end Ligase seals
the nicks
 Occurs throughout the cell cycle (all phases: G1, G2, etc..), defective in
some cancers, neurological disorders
 Lesions it repairs: Alkylation, deamination, oxidation
 Disease: Familial Adenomatous Polyposis
 AD defect of APC gene
 ↓ BER (base excision repair) = polyps / colon cancer
 Lesions are endogenous
 

 Nucleotide Excision Repair: repair dimers

o Process : Endonucleases remove damaged bases (gets rid of thymine dimers)
DNA pol adds new bases  DNA ligase links the new bases to old chain (one
continuous strand)
o Occurs during G1 phase
o Repais buljky defects that distort the helix, commonly pyrimidine dimers (b/s
UVA / UVB b/w two thymine)
o Disease: Xeroderma Pigmentosum
 AR defect in NER (nucleotide excision repair)
 Can’t repair thymine dimers
 Leads to sunlight intolerance, skin cancer, actinic keratoses (children),
corneal ulcers

 Mismatch Repair: repairs incorrect nucleotide placement

o Repairs G/T or A/C pairing using MSH proteins, (MutS, MutH, MutL enzymes)
at S Phase checkpoint (b/c DNA replicate & create mismatches wrong bases)
o Process: Newly synthesized daughter strand  MSH proteins identify mismatch
basesendonuclease breaks strand  exonuclease removes incorrect bases 
DNA pol adds correct bases  DNA ligase links reseals DNA strand
o Disease: Lynch Syndrome
 AD defect in MLH1 or MSH2 genes
 ↓ mismatch repair = hereditary non-poolyosis colorectal cancer (HNPCC)
 Microsatellite instability (unstable number of repeating di/trinucleotides)
 

 Double Stranded Repair:

o Homologous Recombination:
 Uses sister chromatids as template  repair double stranded breaks
 No loss of DNA, more accurate repairs, occurs late S phase/G2
 Disease: BRCA1 mutation: defective homologous recombination 
breast/ovarian cancer (defect in ability to repair genome instability)
 Disease: BLM: defective homologous recombination  Bloom Syndrome
 Bloom Syndrome: short stature, narrow face, red rash, ↑ risk skin cancer
 Disease: Fanconi Anemia: Aplastic anemia = bone marrow failure,
skeletal abnormalities, AML (bone cancer).
 Features of Fanconi Anemia: Café-au-lait spots, kidney
abnormalities, intellectual disability, hydrocephalus, deformity in
thumb, short stature


o Nonhomologous End Joining:
 Less accurate
 Reanneals two DNA fragments, no template required, occurs
throughout cell cycle (all phases)
 No template strand → ↑ Errors
 Occurs when no sister chromatid available, does not require homology
 Repairs breaks from ionizing radiation/oxidative free radicals, radiation
 Desease: Ataxia Telangiectasis
 AR defect on ATM mutation
 Defective NHEJ (non-homologous end joining)  ↓ B/T cells
 Characteristics: Ataxia telangiectasis (loss of balance), spider
angiomas, sinopulmonary infections (IgA deficient) (body
secretions)
  ↑ AFP (alpha fetal protein), ↓ IgA, (eye + skin) ↓IgG, ↓IgE

Q: what is direct repair

A: attempts to repair damaged nucleotides w/o severing the polynucleotides phosphodiesterase
backbone

Q: what are common types of lesions BER corrects

A: Deamination, alkylation, oxidation

Q: How does NER differ from BER?

A: NER can recognize a broader range of single-strand lesions and removes an oligonucleotide
instead of excising a single nucleotide.

Q: Which mechanism repairs the effects of nitrous acid and aflatoxin B?

Answer: Nitrous acid leads to formation of hypoxanthine or uracil which are repaired using the
BER. Aflatoxin B is a bulky adduct which needs NER for repair.

Q: How do HR and NHEJ differ?

Answer: NHEJ is error-prone; HR restores nucleotide sequence. HR primarily occurs during S
and G2 phases; NHEJ can occur in any phase.