The Regulation of Transcription in Eukaryotes Flashcards
What determines the identity and capability of a cell
relative abundance of gene products
Describe transcription - the basics
- first step in converting DNA to gene products
- first step in genotype to phenotype
- primary regulatory gateway for gene expression
Number of genes per cell:
40,000
Number of transcripts per cell:
300,000
Number of proteins per cell:
1,000,000,000
Describe the amplification of transcripts in a cell
~5x
What is the average transcript number per gene?
~10
Describe the amplification of proteins in a cell
~130x
amplification
What is the average protein number per gene?
~1,300
Transcription in eukaryotes is regulated at…
three temporally distinct stages
When is transcription in eukaryotes regulated?
- initiation
- elongation
- termination
Describe transcription regulation at initation
- transcription factors
- promoters
- pre-initiation complex
Describe transcription regulation at elongation
- TFIIS
- nucleosome/chromatin remodeling
Describe transcription regulation at termination
Cleavage and polyadenylation
Describe transcription factors
- the first step transcriptional regulation
- composed of one or more DNA binding domains and protein-protein interaction domains.
- bind to short DNA sequence elements (6-20bp)
- recruit (or block recruitment) of other proteins
- act as activators or repressors
What the humans genes coding for transcription factors
- 1600 genes
- 8%
Describe transcription factor gene families
- ~20
- ~50% of transcription factors in human genome are C2H2 zinc finger transcription factors
Describe the mechanics of transcription factors
- DNA binding domain binds to short sequences in the nucleosome
- recruit enzymes that alter nucleosomes
- recruit the PIC
- GTFs
PIC
pre-initiation complex
General Transcription Factors
GTFs
List the GTFs
- TFIIA
- TFIIB
- TFIID
- TFIIE
- TFIIF
- TFIIH
Describe TFIID
- 13 subunit protein complex
- first recruited component of the pre-initiation complex
- binds to TBP
TBP
TATA binding protein
Describe the TFIID/TBP complex
- assembles without interacting with DNA
- TFIID/TBP complex binds DNA via TBP subunit.
- first stage of pre-initiation complex assembly.
- can be recruited without a TATA binding site for TBP
Describe the TATA box
- consensus sequence is TATAWAW
- W is either A or T
Describe TFIIA
- 3 subunit protein complex
- binds to TFIID/TBP
- stabilises TBP interaction with DNA
Describe TFIIB
- single protein
- facilitates recruitment of RNA polymerase II
Describe TFIIF
- TFIIF – tetramer
- Pol II recruited at same time.
- stabilises Pol II interaction with TFIIB and TFIID
Describe Pol II
- RNA polymerase II
- 12 subunit complex
tetrameric
two subunit complex
Describe TFIIE
- 2 subunit complex
- recruits TFIIH
Describe TFIIH
- 10 subunit complex
- acts as a DNA helicase
Describe the two main function of TFIIH
- opening the promoter to load the polymerase
- phosphorylating Pol II CTD – promoter escape
What happens on pre-initiation assembly?
- phosphorylated Pol II recruits capping enzyme
- first nucleotide that exits the polymerase gets capped
- cap protects the RNA from degradation
- polymerase escapes the promoter and proceeds to transcribe the gene
transcription factors act
in concert
If one transcription factor is mutated
All efforts are lost
The combinatorial action of transcription factors …
specifies cell identity and function
Describe elongating polymerases
- not under control of TFs or PIC
Describe polymerase gene transcription
- average gene ~20,000 bp: 4.5 mins @ 72 nucleotides sec-1
- longest gene ~2.2 million bp: 8.5 hours @ 72 nucleotides sec-1
What happens if transcription goes wrong?
- elongating polymerase backtracks along the DNA
- 3’ OH no longer available for extension
- backtracking cannot be recovered by polymerase
- RNA must be cleaved to produce new 3’OH
- once cleaved, polymerase can resume
Describe cleavage on elongation backtracking?
- TFIIS
- creates two domains
Describe the average gene
- ~20,000 bp
- ~90 nucleosomes
Describe the longest gene
- ~2.2 million bp
- 10,000 nucleosomes
How does Pol-II navigate nucleosomes
with the help of a large cohort of chromatin remodeling enzymes
Describe transcription termination
- PAS encoded in the DNA
- Polymerase transcribes past the PAS
- CAP complex binds Pol II and PAS in RNA
grabs the polymerase and RNA
PAS
polyadenylation signal
- AAUAAA
CAP
- Cleavage and polyadenylation
complex - 83 proteins
