Lecture 3 - transcription in Eukaryotes Flashcards
Compared to bacterial RNA polymerase, how many subunits do eukaryotic RNA polymerases have?
they have 12 core subunits compared to bacterial which only have 5
What are focused promoters?
they initiate transcription at a single transcription start site
What are dispersed promoters?
they initiate transcription at one of many transcription start sites in a 50-100 nucleotide region, which is common in vertebrate organisms
What is the directionality of eukaryotic promoters usually?
bidirectional
What is the TATA box?
a binding site for TATA-binding protein
*they are uncommon in vertebrates
What does the initiator (Inr) encompass?
the transcription start site
Since the initiator complex is highly degenerate (lots of possibilities) in vertebrates, what does this enable?
the use of dispersed core promoters
What letter of the initiator is the start site? (YYANWYY)
A
What is basal transcription?
when the core promoter produces a low level of transcription
Name 2 examples of regulatory elements that are usually positioned near the promoter.
GC and CAAT boxes
What do regulatory factors do?
they bind to regulatory elements to activate or repress transcription
*this is a trans-acting mechanism
What do enhancers do?
they stimulate transcription over long distances (they are promoters!)
What do silencers do?
they inhibit transcription
What do enhancers help to do?
recruit RNA polymerase II to a promoter and to help stimulate the transition from transcription initiation to productive elongation
What does the Mediator Complex do?
it bridges the gap between gene promoters and enhancers
List the general steps of eukaryotic transcription initiation.
Pre-initiation complex (PIC) assembly, open complex formation, transcription start site scanning, promoter escape, promoter-proximal pausing, and pause release and productive elongation
What is TFIID and what does it do?
its composed of TATA-binding protein (TBP)
* it recognizes the TATA box and many other core promoter elements (including the initiator)
What doe TFIIB do?
it binds to TFIID and helps to recruit RNA polymerase II to the promoter, partially binding to TFIIF
What does TFIIF do?
TFIIF binds to RNA polymerase II and helps to mediate recruitment of Polymerase II to they promoter by interacting with TFIIB
What does TFIIE do?
recruits TFIIH to the pre-initiation complex and stimulates the helicase and CTD phosphorylation activities of TFIIH
What does TFIIH do?
Unwinds DNA to form an open complex using helicase activity and phosphorylates the RNA polymerase II CTD during the transition from initiation to elongation
What is the first step in the mechanism of transcription initiation?
the TFIID binds to the TATA box (or other core promoter elements)
Step 2 of the mechanism of transcription initiation?
TFIIB binds to TFIID
Step 3 of the mechanism of transcription initiation?
TFIIB promotes the binding of RNA polymerase II to the core promoter… here, TFIIF interacts wit TFIIB
Step 4 of the mechanism of transcription initiation?
TFIIE and TFIIH binds to RNA polymerase II to form the pre initiation complex
Step 5 of the mechanism of transcription initiation?
TFIIH uses its helicase activity to unwind DNA and form an open complex. TFIIH then phosphorylates the RNA polymerase II CTD, which releases Pol II TFIIB and TFIIH are released
What occurs during promoter escape?
rna polymerase breaks free of the promoter elements
what occurs during promoter-proximal pausing?
RNA polymerase pauses just downstream of the promoter
What occurs during the pause release and productive elongation stage?
RNA polymerase exits the pause and transcribes the gene
What protein causes pausing?
pausing is induced by Negative Elongation Factor
*NELF restrains RNA polymerase II
What is promoter-proximal pausing important for?
*for keeping promoters in an active states
*RNA quality control
*and gene regulation
Which protein releases RNA polymerase II from the pause by phosphorylating the CTD?
Positive Transcription Elongatio factor b (P-TEFb)
What does CTD phosphorylation coordinate?
the RNA polymerase II transcription cycle
Throughout the transcription cycle, CTD becomes what?
highly phosphorylated
At the beginning of genes, what is highly phosphorylated?
Ser5
What is the phosphorylation of Ser5 involved in?
recruiting capping enzyme
What two amino acids are involved in recruiting the spliceosome to mRNA?
Ser5 and Ser2
What is the polyadenylation signal sequence required for?
to add poly-A tail to mRNA
