Control of transcription and chromatin Flashcards
What is gene expression?
Process by which information in genes (DNA) is decoded into protein, it is a multistep process which is highly regulated.
What is gene expression regulated by?
Mostly transcription.
What is the function of a promoter?
RNA polymerase binds here.
It dictates how efficient transcription will be.
What is the promoter in prokaryotes (bacteria)?
Sigma 70 subunit/factor
What is a promoter?
Cis acting DNA regulatory element through which transcription is initiated and controlled.
What is the structure of the promoter in prokaryotes?
Hexametric
35 sequence long upstream
A simple molecule
What are the regions within the eukaryotic promoters?
Regulatory region
Core (basal) region
ORE
What is within the regulatory region in prokaryotic promoters?
Enhancer
x2 UAS
What is within the core (basal) region in prokaryotic promoters?
TATA box
Initiator (Inr)
DPE
BRE
What is the DPE?
Downstrean core promoter element
What is BRE?
TFIIB recognition element
What is the location of the TATA box?
-31 to -26
Why are some gene locations negative?
Upstream of the transcription site.
What is the location of the initiator?
-2 to +4
What is the location of DPE?
+28 to +32
What is the location of BRE?
-37 to -32
What is a TATA box?
The TATA box is a sequence of DNA, consisting of nucleobases TATAAA, located in the promoter region about 25 base pairs before the site of transcription.
What are the eukaryotic promoters?
Associated with regions with a high frequency of CG sequences: “CpG islands”
What are the specifics of CpG islands within acting as a promoter?
In mammals most C residues followed by G a are methylated (to 5-methyl C).
However generally C residues in CpG islands escape methylation (hypomethylated).
What is the function of CpG islands within silencing?
Methylation of CpG islands is associated with silencing (txn “switched off”).
What within the eukaryotic promoters is the regulatory region?
Silencer
Enhancer
URS
X2 UAS
What are the activator binding sites within the eukaryotic promoters?
UAS & Enhancer
What are the repressor binding sites within the eukaryotic promoters?
URS & Silencer
What within the eukaryotic promoter is inside the core (basal) region?
Txn Start-site
What are the Tools for Identifying Promoter Elements?
- Sequence comparison
- Reporter Analysis
What is reporter analysis?
Fuse promoter to reporter.
Reporter genes encode proteins whose levels can easily be measured.
Allows for a measure of gene expression.
What can reporter analysis identify?
When a gene is expressed?
Where it is expressed?
What signals it responds to?
What factors and sequences control its expression?
What are the target genes for RNA polymerase I?
rRNA
What is the location of RNA polymerase I?
Nucleolus
What are the target genes for RNA polymerase II?
mRNA
snRNAs
miRNAs
What is the location of RNA polymerase II?
Nucleus
What are the target genes for RNA polymerase III?
tRNA
5S RNAs
U6 RNA
7S RNA
What is the location of RNA polymerase III?
Nucleus
What is the structure of Bacterial RNA polymerase?
ß subunit
ß’ subunit
α subunit (x2)
ω subunit
What is the structure of Yeast RNA polymerase II (Eukaryotic)?
12 subunits
What are the similarities between RNA polymerases?
All DNA dependent
What are General Transcription Factors (GTF’s)?
Bacterial RNA polymerase requires σ-factor to recognise promoter DNA.
What are the functions of General Transcription Factors (GTF’s)?
(1) RNA pol specific
(2) Multi component factors
(3) Form a complex on TATA box
(4) Recruit RNA pol II to the promoter
(5) Direct initiation at start-site
What is the order of addition for Pre Initiation Complex (PIC) Assembly?
Recognition and binding TF2 d to TATA.
Then TF2A followed by TF2B.
TFIF already attached to RNA polymerase II.
TF2E the TF2H.
Not the only order.
How is transcription initiated by RNA polymerase II?
Helicase activity of TFIIH separates the strands at the start site.
As Pol II begins transcribing it is phosphorylated on the C-terminal domain (CTD)
The CTD is a series of repeats located at the C-terminal end
of the largest β’ homologous subunit of Pol II.
What happens to the rest of the transcription factors when transcription begins?
TFIID and TFIIA may stay behind
TFIIB, TFIIE and TFIIH are released
TFIIF moves down the template with Pol II
How many subunits does the TFIID have?
13
How many subunits does the TFIIA have?
3
How many subunits does the TFIIB have?
1
How many subunits does the TFIIF have?
2
How many subunits does the TFIIE have?
2
How many subunits does the TFIIH have?
9
What is the function of TFIID?
BINDS TO THE TATA BOX (CORE PROMOTER).
RECRUITS TFIIB.
What is the function of TFIIA?
STABILIZES TFIID BINDING
ANTI REPRESSION FUNCTION
What is the function of TFIIB?
RECRUITS RNA pol II-TFIIF
IMPT FOR START SITE SELECTION
What is the function of TFIIF?
STIMULATES ELONGATION
DESTABILIZES NON SPECIFIC RNA pol II-DNA INTERACTIONS
What is the function of TFIIE?
RECRUITS TFIIH AND MODULATES TFIIH ACTIVITY
What is the function of TFIIH?
PROMOTER MELTING AND CLEARANCE
CTD KINASE ACTIVITY
DNA REPAIR COUPLING
What is the structure of TFIIH?
TFIIH is composed of 9-10 subunits
It can be divided into two parts, CORE + CAK
Contains ATPase.
What is the CAK molecule?
The CAK module contains one of the kinases that phosphorylates the CTD of RNAP II
What is the function of the ATPase within the TFIIH?
TFIIH contains a ATPase called XPB (or Ssl2) that plays a key role in PROMOTER MELTING
What is TFIID?
TFIID is the Central RNA pol II Transcription Factor
What is the structure of the TFIID?
TATA binding protein (TBP)
TBP associated factors (TAFs)
What is TBP?
Molecular saddle
What is the advantage function of TBP?
TBP can direct the assembly of the PIC on a TATA-containing
promoter (in vitro).