Eukaryotic Transcription and Gene Regulation Flashcards
Types of gene regulation
Transcription Translation RNA processing RNA degradation RNA export Protein degradation
What are the three classes of RNA polymerase
RNA polymerase I
RNA polymerase II
RNA polymerase III
Describe RNA polymerase I
for large, ribosomal RNA
- components of ribosomes
Describe RNA polymerase II
Transcribes mRNA
Describe RNA polymerase III
Transcribes 5S rNA, tRNA and other small RNAs
- Translation of mRNA into proteins
What is the largest subunit in RNA polymerase II
Carboxy-terminal domain (CTD) - consists of multiple repeats of heptamers
What does the 5’ untranslated region contain?
Signals for translation initiation
What does the 3’ untranslated region contain?
Signals for translation termination
What is the direction of transcription?
5’-3’
what are the 4 stages of transcription?
1) Template recognition
2) Initiation
3) Elongation
4) Termination
What happens in the template recognition phase?
RNA polymerase binds to duplex DNA
DNA is unwound at the promoter
What happens during initiation?
Chains of 2-9 bases are synthesised and released
What happens during elongation?
RNA polymerase synthesise RNA
Unwound region moves with RNA polymerase
RNA polymerase reaches the end of the gene
What happens during termination
The RNA polymerase and RNA is released
What are the requirements for transcription?
Coactivators - do not bind to DNA but required for activators to interact with basal transcription factors
Chromatin needs to be open before RNA polymerase can bind to the promoter
Basal transcription factors - required by RNA polymerase II to form initiation complex at RNA polymerase II promoters
What is the name of the transcription factors needed for initiation by RNA polymerases?
TFIIX
What is a TATA box
Common component of RNA polymerase II promoters
~25bp upstream of the startpoint
What is at the start of an RNA polymerase II promoter?
Initiator element (initiator Inr)
TATA binding protein (TBP)
Component of the positioning factor required for each type of RNA polymerase to bind to the promoter
What is the factor for RNA polymerase II?
TFIID
What does TFIID consist of?
TBP
Multiple TBP-associated factors (TAFs)
What needs to happen to allow elongation?
TFIIE and TFIIH melt DNA to allow polymerase movement
Phosphorylation of the CTD to promote clearance
What is upstream of the transcription start site
Enhancer region (enhancer factors) Upstream control element (Upstream factors and inducible factors) TATA box (general factors)
What do enhancers form complexes with?
Activators
What conformation do enhancers work in?
Cis
What is required for enhancer function?
DNA looping
What is p53?
A tumour suppressor transcription factor induced by stress
What is p68?
An RNA helicase which acts as a co-activator of several transcription factors including p53
Induces cell cycle arrest and apoptosis
p53 binds to DNA, p68 binds to p53
What happens to the 5’ end during transcription
A 5’ methylated cap is formed by adding a G to the terminal base
What is the function of the cap?
Recognised by protein factors and influences mRNA stability, splicing, export and translation
What are the splice site consensus’?
Major introns
Minor introns
Branch point
Explain the splice site consensus of the major introns
5’ splice site at the 5’ end of the intron which includes a consensus sequence GU
3’ splice site at the 3’ end of the intron which includes a consensus sequence AG
Process of splicing
5’ exon is cleaved off
Intron lariat formation
3’ exon is cleaved off
The two exons are ligated together
What are the different modes of alternative splicing
- Intron retention
- Alternative 5’ splice sites
- Alternative 3’ splice sites
- Exon inclusion/ skipping
- Mutually exclusive exons
- Combinational exon selection
- Alternative promoter/ splicing
- Alternative polyadenylation/ splicing
Types of silencing enhancers and silencers
Intronic splicing silencer
Exonic splicing enhancer
Exonic splicing silencer
Intronic splicing enhancer
What is the function of p68 in regards to H-Ras?
suppresses inclusion of the H-Ras IDX alternative exon -
What is the effect of suppressing inclusion of H-Ras IDX alternative exon?
Leads to formation of p19H-Ras rather than p21H-Ras
What is the function of p19H-Ras?
G1/S delay
What is RAS?
an oncogene
What does the protein complex contain which cleaves that RNA to generate a 3’ end?
Specificity factor
Endonuclease
poly(A) polymerase
What is the function of the specificity factor and the endonuclease?
cleave RNA downstream of AAUAAA
What is the function of the poly(A) tails?
Controls mRNA stability and influences translation
How can p68 effect introns?
Increase splicing of both random and specific introns to increase cell activity
What does the vast majority of the genome transcribe?
Short and long non-coding RNAs (ncRNAs)
Describe long non-coding RNAs (lncRNAs)
Spliced and have polyadenylated signals
Tissue specific and development stage specific
What is the function of lncRNAs?
Epigenetic regulators of protein-coding expression
- transcription
- subcellular trafficking
- protein degradation
Describe siRNA
Generated by DICER cleavage of dsRNA duplexes
Incorporated into RNA-induced silencing complexes (RISC)
Involved in gene regulation, transposon control and viral defence
Describe miRNA
Generated by DICER cleavage of imperfect RNA hairpins
Encoded into long primary transcripts, often into introns
Incorporated with argonaute proteins into RNA-induced silencing complex (RISC)
Involved in post-translational gene regulation
What happens if you increase onco-miRNAs?
proliferation and inhibition of apoptosis
What happens if you decrease oncosuppressor miRNAs?
Metastasis and migration
Define onco-miRNA
If an miRNA represses transcription of a tumour suppressor
Define oncosuppressor miRNA
If a miRNA represses expression of an oncogene
What is low survival rate of breast cancer patients associated with?
High levels of intron containing p68 RNA
