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Lecture 4: DNA transcription
Reference:
Essential Cell Biology by Alberts et al
Central dogma of molecular
biology
What is the pathway from DNA to Protein?
Transcription- DNA directed RNA synthesis
Transcription is the mechanism by which a
template strand of DNA is utilized by specific
RNA polymerases to generate one of the three
different types of RNA (mRNA, tRNA and rRNA)
What is the biological significance?
Allows selective expression of genes
Regulation of transcription controls time, place
and level of protein expression
Where does transcription take place?
Transcription in eukaryotes
A) transcribing a primary RNA transcript
B) modification of this transcript into mRNA
(transcriptional processing)
Basic structure of a eukaryotic gene
Regulatory region
coding region
What are the key events during transcription?
A. Initiation RNA
polymerase binds to
promoter & opens
helix
B. Polymerisation De
novo synthesis using
rNTPs as substrate
Chain elongation in 5’-3’
direction
C. Termination stops at
termination signal
Question
In what direction are the
RNA polymerases moving in
this image?
A) Initiation by RNA polymerase holoenzyme
Prokaryotes:
single multisubunit
complex
Eukaryotes: 3 types
RNA pol I
RNA pol II
RNA pol III
Located in nucleoli
Located in
nucleoplasm
Synthesises mRNA
precursors
Located in
nucleoplasm
Synthesises 5S
rRNA, tRNA, snRNAs
Synthesises most
rRNA precursors
What are promoters and terminators?
Region on DNA where RNA polymerase binds to initiate
or terminate transcription
Sequence of promoter determines direction of
RNA polymerase action
Which strand carries the coding sequence?
Promoters
Prokaryotes


Near 5’ end of
operons
Pribnow box –
consensus
sequence TATAAT
Eukaryotes



Near 5’ end of genes
Recognised by RNA pol II
Consensus promoter sequence for
constitutive structural genes –
GGGCGG
Selective structural genes – TATA
Enhancers and silencers
Sequences associated with a promoter which
enhance/repress the activity of a promoter due to its
association with proteins called transcription factors
Mediate most selective gene expression in eukaryotes
B) Polymerisation




RNA polymerase binds to promoter & opens helix
RNA polymerase catalyses addition of rNTPs in the 5’-3’
direction
RNA polymerase generates hnRNAs (~70-1000 nt long) &
all other RNAs
Stops at termination signal
C) Termination
specific termination sequence
e.g E.coli needs 4-10A followed by a palindromic GC
rich region
Additional termination proteins
e.g. Rho factor in E.coli
B) Transcriptional processing
3 main steps
1) RNA capping,
2) polyadenylation
3) splicing
1) Capping
Addition of 7
methylguanosine at 5’
end
Mediated by
guanylyltransferase



Probably protects
against degradation
Serves as recognition
site for ribosomes
Transports hnRNA from
nucleus to cytoplasm
2) Tailing
Addition of poly(A) residues at 3’ end
 Transcript cleaved 15-20nt past AAUAAA
 Poly(A)polymerase and cleavage &
polyadenylation specificity factor (CPSF)
attach poly(A) generated from ATP
3) Splicing
Highly precise removal
of intron sequences
Performed by snRNPs and
spliceosomes (large RNAprotein complex made of
small nuclear
ribonucleoproteins)
Recognise exon-intron
boundaries and splice
exons together by
transesterification
reactions
Transcriptional processing
of ovalbumin gene
Is an mRNA always processed in the same way?
-- “alternative splicing”
Differential splicing in specific tissues
Question:
Will counting the number of promoters yield the
number of “genes” in the human genome?
How do mRNAs get to the cytosol?
Figure 15-8
Why do eukaryotes
have DNA within a
membrane bound
compartment and
prokaryotes do not?
Could eukaryotes
function without it?
During "RNA processing"
A. all of the exons are removed
B. the RNA molecule is made from a DNA
template.
C. introns are removed from the RNA and
the exons are spliced together.
D. the RNA molecule is translated into a
protein molecule.
Since the two strands of the DNA
molecule are complementary
A. The RNA polymerase can bind to either strand.
B. Only one strand actually carries the genetic code
for a particular gene.
C. Each gene possesses an exact replica that can be
used should a mutation occur.
D. A gene transcribed in the 5’ to 3’ direction on one
strand can be transcribed in the 3’ to 5’ direction
on the other strand.
The presence of a poly-A tail on a
RNA molecule indicates that
A. there are exons present that must be
removed.
B. this RNA molecule does not contain introns.
C. the transcript should be immediately
degraded.
D. this is a rRNA molecule.
E. None of the above answers is correct
. “Alternative splicing” refers to
A. the use of introns as exons, or vice versa, during
RNA processing
B. splicing out of damaged DNA by DNA repair
enzymes.
C. joining of RNA from two different genes to form
a new mRNA.
D. the use of alternative reading frames when
translating an mRNA.
E. a new dance for people with alternative life
styles.