Article
- The EMBO Journal (2006) 25, 3100 - 3109
- doi:10.1038/sj.emboj.7601197
Published online: 15 June 2006
Subject Category:
An 8 nt RNA triggers a rate-limiting shift of RNA polymerase II complexes into elongation
Aaron R Hieb, Sean Baran, James A Goodrich and Jennifer F Kugel
- Department of Chemistry and Biochemistry, University of Colorado at Boulder, Boulder, CO, USA
Correspondence to:
Jennifer F Kugel, Department of Chemistry and Biochemistry, University of Colorado at Boulder, 215 UCB, Boulder, CO 80309-0215, USA. Tel.: +1 303 735 0955; Fax: +1 303 492 5894; E-mail: Jennifer.Kugel@colorado.edu
James A Goodrich, Department of Chemistry and Biochemistry, University of Colorado at Boulder, 215 UCB, Boulder, CO 80309-0215, USA. Tel.: +1 303 492 3273; Fax: +1 303 492 5894; E-mail: james.goodrich@colorado.edu
Received 16 January 2006; Accepted 22 May 2006
Abstract
To better understand the critical conversions that RNA polymerase II complexes undergo during promoter escape, we determined in vitro the precise positions of the rate-limiting step and the last step requiring negative superhelicity or TFIIE and TFIIH. We found that both steps occur after synthesis of an 8 nt RNA during the stage encompassing translocation of the polymerase active site to the ninth register. When added to reactions just before this step, TFIIE and TFIIH overcame the requirement for negative superhelicity. The positions at which both steps occur were strictly dependent on RNA length as opposed to the location of the polymerase relative to promoter elements, showing that the transcript itself controls transformations during promoter escape. We propose a model in which completion of promoter escape involves a rate-limiting conversion of early transcribing complexes into elongation complexes. This transformation is triggered by synthesis of an 8 nt RNA, occurs independent of the general transcription factors, and requires under-winding in the DNA template via negative superhelicity or the action of TFIIE and TFIIH.
Keywords:
- DNA conformation,
- kinetics,
- promoter escape,
- RNA polymerase II,
- transcription
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