Eukaryotic Transcription 5 Flashcards
SRBs
- bind to the CTD of RNA pol II large subunit in yeast
- suppressor of RNA pol B
Mutation of SRB4: how severe?
mutations in these genes overcome mutations in the CTD, which implies that SRBs interact with the CTD
- a mutation in SRB4 eliminates 93% of transcription in both basal and activated
SRB4: which module?
head of the mediator
Sin4 deletion: tail-less mutant; consequence
are defective in activated transcription
what is the role of Cdk8/CycC in inhibiting action of TFIIH?
- Srb10/Srb22 module inhibits TFIIH by phosphorylating the cycH subunit of H
- the cdk8 module is dissociable
what are the two ways to build affinity column to capture Mediator
- can be purified using an affinity column with immobilized CTD
- can also purify using VP16 acidic activator column
Model for how Mediator modulates amplitude of transcription
- Transactivators determine the Mediator conformation, which in turn, controls the rate of RNA polymerase entry onto the promoter. Thus Mediator controls the rates of initiation and reinitiation.
- Specific Mediator conformation determined by transactivator contact.
What controls Mediator conformation?
x
mediator
- large complex of proteins known as the mediator is also apart of the preinitiation complex
- contains 24 subunits
- purify with RNA pol 2 and binds to CTD
Mediator complex activities
- 5 - 10 fild increase in basal transcription
- 30 - 50 fold stimulation of TFIIH dependent phosphorylation of the CTD
- integrator of positive and negative signals
head of the mediator complex
- attachment sites to RNA pol 2 and the CTD of RNA pol 2
- Srb4
middle of the mediator
- Med9/10
- specific function unclear
Tail of mediator
- Gal 11 module
- contacts with transactivators and repressor proteins
- Sin 4 mutants
Rbp4/7
- mediator contacts Rbp4/7 subunits and triggers the opening of the clamp
- this provides better access for DNA to enter the active site of the polymerase
SWI/SNF proteins
- disrupts nucleosome structure resulting in 30 fold increase in binding of transactivator proteins
CTCF
- recruits cohesion, which actively protrudes DNA to form loops. The DNA loops bring distal enhancers in proximity with the core promoter
what other proteins participate in DNA looping and are sometimes referred to as super enhancers?
LIM and LDB1
super enhancers
coordinate the expression of the many olfactory receptor genes
- marks on the histone H3 lysine also play a prominent role in regulations
