Overview of transcription
Transcription is the first step in gene
expression, in which information from a gene
is used to construct a functional product such
as a protein. The goal of transcription is to
make a RNA copy of a gene’s DNA sequence.
For a protein-coding gene, the RNA copy, or
transcript, carries the information needed to
build a polypeptide (protein or protein
subunit). Eukaryotic transcripts need to go
through some processing steps before
translation into proteins.
RNA polymerase
The main enzyme involved in transcription is
RNA polymerase, which uses a single-stranded
DNA template to synthesize a complementary
strand of RNA. Specifically, RNA polymerase
builds an RNA strand in the 5’ to 3’ direction,
adding each new nucleotide to the 3’ end of
the strand.
Stages of transcription in bacteria
Transcription of a gene takes place in three stages: initiation, elongation, and termination.
Initiation. RNA polymerase binds to a
sequence of DNA called the promoter,
found near the beginning of a gene.
Each gene (or group of co-transcribed
genes, in bacteria) has its own
promoter. Once bound, RNA
polymerase separates the DNA
strands, providing the single-stranded
template needed for transcription.
Elongation. One strand of DNA,
the template strand, acts as a
template for RNA polymerase.
As it “reads” this template one
base at a time, the polymerase
builds an RNA molecule out of
complementary nucleotides,
making a chain that grows from
5’ to 3’. The RNA transcript
carries the same information as
the non-template (coding)
strand of DNA, but it contains
the base uracil (U) instead of
thymine (T).
Termination. Sequences called terminators signal that the RNA transcript is complete. Once they are
transcribed, they cause the transcript to be released from the RNA polymerase. An example of a
termination mechanism involving formation of a hairpin in the RNA is shown below.
Post-transcriptional Processing or RNA processing
In prokaryotes, no RNA processing is necessary: – the nascent RNA is usually the mRNA.
In eukaryotes, the nascent RNA is called primary transcript-RNA – needs to be processed – and
transported to the cytoplasm for translation to occur.
In eukaryotes, the transcript of a protein-coding gene is called a pre-mRNA and must go through
extra processing before it can direct translation.
The processing steps are:
– Addition of a 5’ 7-methyl guanosine cap (capping)
– Addition of a poly-A tail at the 3’ end (polyadenylation)
– RNA splicing to remove intervening sequences (remove introns)
Eukaryotic pre-mRNAs must have their ends modified, by addition of a 5’ cap (at the beginning) and
3’ poly-A tail (at the end).
Many eukaryotic pre-mRNAs undergo
splicing. In this process, parts of the
pre-mRNA (called introns) are
chopped out, and the remaining
pieces (called exons) are stuck back
together.
End modifications increase the
stability of the mRNA, while splicing
gives the mRNA its correct sequence. (If the introns are not removed, they’ll be translated along with
the exons, producing a “gibberish” polypeptide.)
Translating Genetic Information
The process of translation can be seen as the decoding of instructions for making proteins, involving
mRNA in transcription as well as tRNA. The genes in DNA encode protein molecules, which are the
“workhorses” of the cell, carrying out all the functions necessary for life.
Initiation
▪ During initiation, a group of proteins called initiation factors assist in assembling the ribosome
around the mRNA.
▪ The initiation factors temporarily recognize specific sequences in the mRNA.
▪ The small ribosomal subunit then recognizes the initiation factors, followed by the large
ribosomal subunit.
▪ The ribosome is assembled around the mRNA, much like a series of toy plastic blocks.
▪ Near the beginning of the mRNA is a codon called the start codon (AUG). This codes for an
amino acid called methionine.
▪ Three regions are important as the ribosome is assembled around the mRNA. They are
commonly called the A, P, and E sites.
▪ Each site will fit a single tRNA.
▪ The only tRNA that can effectively enter the site is the one whose anticodon complements the
codon of the mRNA revealed within the site.
▪ In initiation, the assembly of the ribosome occurs with the AUG start codon within the P site.
ThisEnds the initiation stage.
Elongation
▪ The elongation stage involves the assembly of specified amino acids into a polypeptide chain.
▪ The key to elongation are the E, P, and A sites within the ribosome.
▪ Following initiation, the first tRNA (for methionine) is located within the P site.
▪ A second codon in the mRNA is exposed in the A site.
▪ Only a tRNA with an anticodon complementary to the mRNA codon exposed in the A site will
correctly fit. At this point there are two tRNAs in the ribosome.
▪ By an enzymatic reaction, the amino acids between the P and A chains are joined together
by aPeptide bond.
▪ As the peptide bond forms, the amino acid is released from the tRNA in the P site. The
ribosome then moves one codon down the mRNA (in the 3′ direction).
▪ As it does so, the tRNA that was in the P site enters into the E site and leaves the ribosome.
▪ The tRNA that was in the A site, which still has the polypeptide chain attached, moves into
theP site.
▪ A new mRNA codon is then revealed in the A site.
▪ A tRNA with an anticodon complementary to the exposed mRNA codon then enters the A
site, and the process repeats itself.
Termination
▪ The process of termination begins once the end of the mRNA is reached by the ribosome.
▪ In place of tRNAs, proteins called release factors enter into the A site.
▪ Since the release factors do not contain amino acids, the process of translation is stopped at
this point.
▪ The release factors also promote the disassembly of the ribosome and its interaction with
the mRNA.
▪ The end result of translation is a polypeptide chain. This polypeptide chain must undergo a
series of folds in order to produce a functional protein.
