Termination process

Question 1

How is the preinitiation complex formed

Answer 1

Core promoter elements assemble complex

TBP binds TATA box - TFIIB binds BRE

Remaining components bind pre-initiation complex

Question 2

Initiation

Answer 2

RNA polymerases open 14bp of duplex DNA

In Pol II - transcription bubble opened by helicase subunits of TFIIH - requires ATP

RNA polymerase fails to make full length RNA first attempt - ‘abortive initiation’ leads to release of short RNAs (2-9nts)

Question 3

Elongation

Answer 3

Elongation coupled to mRNA processing in eukaryotes

Phosphorylation on 5th serine in heptad repeat on CTD region of RPB1 subunit

Binding of negative elongation factors - leads to transcription pausing

Phosphorylation recruits RNA processing enzymes - add guanosine cap to 5’ end of mRNA

Capping causes phosphorylation of 2nd serine in CTD heptad repat

RNA pol II resumes elongation

Question 4

Termination in bacteria

Answer 4

Type I and II terminators

Type I - Rho independent - No additional factors required

Type II - Rho dependent - Rho factor required - uses ATP

Question 5

Termination in eukaryotes

Answer 5

RNA pol II trascribes by polyadenylation and 3’ end processing signals

RNA-processing complexes associate with both processing signals and phosphorylated CTD of large subunit

mRNA cleaved - transcription terminated

RNA pol II released from DNA

mRNA cleaved at poly(A) site

Nascent RNA downstream of Poly(A) cleavage site - digested by 5’ to 3’ exonuclease

Polymerization disrupted - RNA pol II disassociates from DNA template