Transcription

Learning Outcomes
At the end of this topic, students should:
Understand DNA transcription
Understand RNA translation
Central Dogma
 How do DNA carry genetic codes?
The language of DNA only has four
alphabets A, T, C and G
These four alphabets can make up genetic
words but the words in genetics only has
three alphabets codons
These combination of three nucleotide
encodes for a specific amino acids.
The amino acids produced will form proteins
that will then be used for biological processes.
 DNA transcription
DNA transcription is the process where by the
RNA is formed from the DNA that carries
genetic codes.
Protein production
Formation of rRNA and tRNA
The RNA produced by transcription is what we
call the messenger RNA (mRNA)
Once messenger RNA is produced, it will then
exit the nucleus (in eukaryotes) and enter the
cytoplasm for translation process
Transcription in Prokaryotes
 Transcription is catalyzed by RNA polymerase
which makes RNA using DNA as a template.
E. coli RNA polymerase most well-studied
molecular weight about 500,000 Da
four different types of subunits: , , , and s
the core enzyme is 2
the holoenzyme is 2s
the role of the s subunit is recognition of the
promoter locus; the s subunit is released after
transcription begins
of the two DNA strands, the one that serves as the
template for RNA synthesis is called the template
strand or antisense strand; the other is called the
coding strand or sense strand
the holoenzyme binds to and transcribes only the
template strand
 Transcription of DNA occurs in four main stages:
1. binding of RNA polymerase to DNA at a promoter,
2. initiation of transcription on the template DNA
strand,
3. subsequent elongation of the RNA chain, and
4. eventual termination of transcription, accompanied
by the release of RNA polymerase and the completed
RNA product from the DNA template.
RNA polymerase moves along the template
strand of the DNA in the 3-prime to 5-prime
direction, and the RNA molecule grows in the 5-
prime to 3-prime direction.
How does RNA polymerase know
where to begin transcription?
 The DNA promoter region in prokaryotes is a
stretch of about 40 base pairs adjacent to and
including the transcription startpoint.

Upstream: 3 end Downstream: 5 end

 The essential features of the promoter are;
startpoint (designated +1 and usually an A),
the six-nucleotide -10 sequence,
the six-nucleotide -35 sequence.
The two key sequences are located approximately
10 nucleotides and 35 nucleotides upstream from
the startpoint.
The number of nucleotides separating the
consensus sequences from each other and from
the startpoint are important for promoter
function, but the identity of these nucleotides is
not.
 During elongation, RNA polymerase binds to about 30
base pairs of DNA
Termination of transcription requires a termination
sequence that triggers the end of transcription.
Two classes exist, rho dependent and rho
independent.
In rho independent termination, a short complementary
GC-rich sequence of 30-40 bp sequence followed by a
series of T (U in the transcribed RNA) will form a "brake"
that will help release the RNA polymerase from the
template.
In rho dependent termination, binding of rho to the mRNA
releases it from the template.
Rho independent termination
Overall process
 Regulation - prokaryotes
The control of transcription is largely responsible for
the control of expression of genes; done by
alternatives factors different factors for different
genes
Enhancers/silencers upstream of promoters
Transcription factors bind to it
Operons group of genes that are controlled
together
Inducer will determine if the gene will be
expressed or not i.e. no inducer gene not
expressed
transcription attenuation eg. Trp operon
Transcription in Eukaryotes
 Transcription in eukaryotes
Although transcription in eukaryotes is similar to that
in prokaryotes, the process appears to be complex.
Instead of one RNA polymerase, there are three (RNA
Polymerases I, II, and III) involved in eukaryotic
transcription.
1. RNA polymerase I (localized to the nucleolus) transcribes
the rRNA precursor molecules.
2. RNA polymerase II produces most mRNAs and snRNAs.
3. RNA polymerase III is responsible for the production of
pre-tRNAs, 5SrRNA and other small RNAs.
The mitochondia and chloroplasts have their own RNA
polymerases.
 Three classes of eukaryotic promoters
1. RNA polymerase I: The promoter for RNA
polymerase I has two components:
a core promoter (surrounding the startpoint)
an upstream control element. After the binding
of appropriate transcription factors to both
sites, RNA polymerase I binds to the core
promoter.
2. RNA polymerase II: The typical promoter for RNA
polymerase II has a short initiator sequence,
consisting mostly of pyrimidines and usually a TATA
box about 25 bases upstream from the startpoint.
- Require specific transcription factor (STF)
3. RNA polymerase III: The promoters for RNA
polymerase III vary in structure but the ones for
tRNA genes and 5S rRNA genes are located entirely
downstream of the startpoint, within the
transcribed sequence.
In tRNA genes, about 30-60 base-pairs of DNA
separate promoter elements;
In 5S rRNA genes, about 10-30 base-pairs
promoter elements.
 General transcription factors (GTF) and the
polymerase undergo a pattern of sequential
binding to initiate transcription of nuclear genes.
TFIID binds to the TATA box followed by
the binding of TFIIA and TFIIB.
The resulting complex is now bound by the
polymerase, to which TFIIF has already attached.
The initiation complex is completed by the addition of
TFIIE, TFIIJ, and TFIIH.
After an activation step requiring ATP-dependent
phosphorylation of the RNA polymerase molecule, the
polymerase can initiate transcription at the startpoint.
 Termination signals end the transcription of
RNA by RNA polymerase I and RNA
polymerase III without the activity of hairpin
structures as seen in prokaryotes.
mRNA is cleaved 10 to 35 base-pairs
downstream of a AAUAAA sequence (a poly-A
tail signal).
Post-transcriptional modification
 Messenger RNA in eukaryotes requires
excessive processing
Capping of 5 end
5 prime "cap" (a guanosine nucleotide
methylated at the 7th position) 1. to give
stability to mRNA. 2. protect from
nuclease
Polyadenylating - Adding poly-A
1. Protect from nuclease degradation
2. Assist in translation process
Splicing removal of introns in pre-mRNA
END OF LECTURE