TATA-binding protein and transcription factor IIB induce transcript slipping during early transcription by RNA polymerase II.
To better understand the mechanism of steps in early transcription by RNA polymerase II (pol II), we investigated the molecular determinants of transcript slipping within complexes assembled on promoters containing a pre-melted transcription bubble from -9 to +3. Transcript slippage occurs when an RNA transcript contains a repetitive sequence that allows the transcript to slip back and pair with the template strand of the DNA at a new register before transcription continues. We established the contributions of individual transcription factors, DNA elements, and RNA length to slipping on a heteroduplex template using a highly purified human pol II transcription system. We found that transcripts slip at a very defined point in the transcription reaction, after pol II completes phosphodiester bond synthesis at register +5. This point is set by the position of the polymerase active site on the DNA template, as opposed to the length of the transcript, as well as by a repetitive CUCU sequence that must occur from +2 to +5. Interestingly, slipping at this juncture is induced by TATA-binding protein and transcription factor IIB and requires a TATA box but not a transcription factor IIB recognition sequence. We propose a model in which transcribing complexes, upon completing phosphodiester bond synthesis at register +5, enter one of two branches in which they either complete productive synthesis of the transcript or undergo multiple rounds of transcript slipping.
Gilman
Benjamin
B
Department of Chemistry, University of Colorado, Boulder, Colorado 80309-0215, USA.
Drullinger
Linda F
LF
Kugel
Jennifer F
JF
Goodrich
James A
JA
D001483:Base Sequence; D020134:Catalytic Domain; D004247:DNA / Q000235:genetics / Q000378:metabolism; D004951:Esterification; D006801:Humans; D008969:Molecular Sequence Data; D012319:RNA Polymerase II / Q000235:genetics / Q000378:metabolism*; D035181:TATA-Box Binding Protein / Q000235:genetics / Q000378:metabolism*; D013698:Templates, Genetic; D013997:Time Factors; D035581:Transcription Factor TFIIB / Q000235:genetics / Q000378:metabolism*; D014158:Transcription, Genetic / Q000235:genetics*
D035181:TATA-Box Binding Protein; D035581:Transcription Factor TFIIB; D004247:DNA; D012319:RNA Polymerase II