The role of the transcription bubble and TFIIB in promoter clearance by RNA polymerase II.
We have studied promoter clearance at a series of RNA polymerase II promoters with varying spacing of the TATA box and start site. We find that regardless of promoter spacing, the upstream edge of the transcription bubble forms 20 bp from TATA. The bubble expands downstream until 18 bases are unwound and the RNA is at least 7 nt long, at which point the upstream approximately 8 bases of the bubble abruptly reanneal (bubble collapse). If either bubble size or transcript length is insufficient, bubble collapse cannot occur. Bubble collapse coincides with the end of the requirement for the TFIIH helicase for efficient transcript elongation. We also provide evidence that bubble collapse suppresses pausing at +7 to +9 caused by the presence of the B finger segment of TFIIB within the complex. Our results indicate that bubble collapse defines the RNA polymerase II promoter clearance transition.
Pal
Mahadeb
M
Department of Molecular Genetics, Lerner Research Institute, Cleveland Clinic Foundation, Cleveland, Ohio 44195, USA.
Ponticelli
Alfred S
AS
Luse
Donal S
DS
D020029:Base Pairing; D001483:Base Sequence; D006367:HeLa Cells; D006801:Humans; D008954:Models, Biological; D008969:Molecular Sequence Data; D011196:Potassium Permanganate / Q000494:pharmacology; D011401:Promoter Regions, Genetic; D012313:RNA / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D012319:RNA Polymerase II / Q000235:genetics / Q000378:metabolism*; D011994:Recombinant Proteins / Q000378:metabolism; D016385:TATA Box; D013698:Templates, Genetic; D035581:Transcription Factor TFIIB / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism*; D051758:Transcription Factor TFIIH; D019473:Transcription Factors, TFII / Q000235:genetics / Q000378:metabolism; D024363:Transcription Initiation Site; D014158:Transcription, Genetic
D011994:Recombinant Proteins; D035581:Transcription Factor TFIIB; D019473:Transcription Factors, TFII; D011196:Potassium Permanganate; D051758:Transcription Factor TFIIH; D012313:RNA; D012319:RNA Polymerase II