Transcription
Gene Expression Part 1
Gene Expression
• There are 4 major events that occur
durin the process of gene expression
– Transcription
– RNA processing
– Translation
– Protein processing
A Gene is a Transcription Unit
Promoter
Regulatory
sequence
Terminator
DNA
Transcription
3
mRNA 5
Start
codon
Ribosomalbinding site
Many
codons
Stop
codon
Prokaryotic Gene Structure
DNA
+1
Cis-Regulatory
Elements
USE/Promoter
/Operator
ShineDelgarno
box
Stop Codon
TAA, TAG,
TGA
ATG
Coding Sequence = ORF 1
5’ UTR = Leader
sequence
Protein A
Cistron 1
Regulatory Sequences
ShineDelgarno
box
ATG
Stop Codon
Coding Sequence = ORF 2
Spacer = 5’UTR
of 2nd cistron
Protein B
Cistron 2
Structural or Coding Sequences
Regulatory and Coding Sequence Unit = Operon
Termination
sequence
Prokaryotic Gene Structure
DNA
Shine-Delgarno
Stop Codon
Shine-Delgarno
Stop Codon
box
TAA,
TAG, TGA
+1
box
ATG
ATG
TAA, TAG, TGA
Cis-Regulatory
Coding Sequence= ORF
Coding Sequence= ORF
Elements
USE/Promoter
/Operator
Protein A
5’ UTR = Leader
sequence
Cistron 1
Terminator
sequence
Cistron 2
5’ UTR = Leader
sequence
Polycistronic
mRNA
Protein B
Spacer = 5’UTR
of 2nd cistron
Spacer
ORF Protein A
+1
AUG
Shine-Delgarno
box
AUG
Stop Codon
Shine-Delgarno
UAA, UAG, UGA
box
ORF Protein B
Stop Codon
UAA, UAG, UGA
Eukaryotic Gene Structure
CisRegulatory
Elements
Promoter/
Enhancer
Stop Codon
TAA, TAG, TGA
Start Codon
ATG
Exon1
Exon2
Exon3
5’ UTR
3’ UTR
Exon1
Exon2
Start Codon
Exon1
5’ UTR
Exon3
Stop Codon
polyA tail
Exon2 Exon3 AAAAAAAA
3’ UTR
Transcription Proceeds Through 3 Steps
Initiation
•
Transcription factors & RNA
polymerase recognize & bind
the promoter
•
DNA adjacent to the promoter
is denatured forming the open
promoter complex
Elongation
•
RNA polymerase moves along
the DNA in synthesizing a RNA
transcript. Synthesis is 5’3’ –
Only 1 strand of DNA is read as a
template.
Termination
•
A termination signal is reached
causing RNA polymerase to
dissociated from the DNA
Figure 12.2
A Prokaryotic Promoter
Coding strand
Promoter
–35 sequence
5
T TG AC A
A AC T G T
3
Template strand
Transcriptional
start site
–10 sequence
+1
3
T AT AAT
A
A TAT TA
T
5
A
5
RNA
3
Transcription
Reaching A Consensus
+1 Transcribed
–35 region –10 region
lac operon TTTACA N17 TATGTT N6 A
lacI
GCGCAAN17 CATGAT N7 A
trp operon
TTGACA N17 TTAACT N7 A
rrnX
TTGTCT N16 TAATAT N7 A
recA
TTGATA N16 TATAAT N7 A
lexA
TTCCAA N17 TATACT N6 A
tRNAtyr
TTTACA N16 TATGAT N7 A
Consensus TTGACA
TATAAT
RNA Polymerases
Structure of a bacterial
RNA polymerase
Structure of a eukaryotic
RNA polymerase II
RNA Polymerases
• Differences between eukaryotes & prokaryotes
• Prokaryotes
– 1 enzyme with 4 subunits
• 2 α’s, 1 , & 1 ’
• actual polymerase function
– Sigma factors (σ )
• recognize & bind promoter DNA sequence
• Eukaryotes
– 3 separate holoenzymes – each has ~12 subunits
• RNA Pol I – 28S, 18S, 5.8S rRNA
• RNA Pol II – mRNA, snRNA
• RNA Pol III – tRNA, 5S rRNA
– 3 sets of basal transcription factors
• recognize promoter DNA sequences
The Process of Transcription
• Initiation
– Where/when most regulation of gene
expression occurs
– Different between proks & euks
• Elongation
– Essentially same between prokaryotes &
eukaryotes
– Some regulation, more in proks than euks
• Termination
– Different between proks & euks
– Some regulation
Prokaryotic Transcription Initiation
Figure 12.6
Rho Dependent Termination in Prokaryotes
rho utilization site
Rho protein is
a helicase
Figure 12.8
r-dependent termination
Rho Dependent Termination in Prokaryotes
Figure 12.8
r-dependent termination
Rho Independent Termination in Prokaryotes
 r-independent termination requires two sequences in the RNA
– A stem-loop structure upstream of 7-9 U residues
NusA
Stabilizes
the RNA pol
pausing
Figure 12.9
r-independent termination
Eukaryotic Promoters
Pol I
• RNA Pol I
– rRNA
precursor
UBF
SL1
UCE
Core
45S rRNA
precursor
Pol II
• RNA Pol II
– mRNAs,
U6 snRNA
• RNA Pol III
– tRNA, 5S
rRNA,
GSTFs TFIID
UAS
TATA
box
Pol III
pre-mRNA
TFIIIB
TFIIIC
AB
tRNA
precursor
The Pol II Transcription
Initiation Complex
Figure 12.12
Figure 12.12
Pol II Initiation Complex
• TFIIH regulates
formation of the open
complex
•
•
Has a helicase function that promotes melting
of DNA
Phosphorylates the carboxyl terminal domain
domain of RNA pol II
– RNA pol II releases TFIIB
– Promoter clearance occurs
RNA pol II proceeds to
the elongation stage
Chromatin Structure Affects Promoter
Access
Core histone
proteins
Histone
acetyltransferase
Histone
deacetylase
COCH3
COCH3
COCH3
COCH3 COCH3
COCH3 COCH
3
COCH3
COCH3
COCH3
COCH3
Acetyl
group
Histone acetylation
ATP-dependent
chromatin remodeling
complex
Change in the relative positions
of a few nucleosomes
Change in the spacing of
nucleosomes over a long distance
Chromatin remodeling