Structural basis of RNA polymerase II backtracking, arrest and reactivation
- 23 February 2011
- journal article
- research article
- Published by Springer Science and Business Media LLC in Nature
- Vol. 471 (7337), 249-253
- https://doi.org/10.1038/nature09785
Abstract
RNA polymerase II (RNA pol II) moves forwards along the DNA strand during gene transcription, synthesizing messenger RNA as it goes. It can also move backwards and stall — a useful property for regulatory purposes or if it hits an obstacle such as a nucleosome. This arrested state is reactivated by transcription factor IIS (TFIIS). Now, the crystal structure of a backtracked yeast RNA pol II complex containing observable backtracked RNA has been determined at 3.3 Å resolution, as well as the structure of a backtracked complex containing TFIIS. The structures reveal possible mechanisms of transcriptional stalling and transcription reactivation. During gene transcription, RNA polymerase (Pol) II moves forward along DNA and synthesizes mRNA. However, Pol II can also move backwards and stall, which is important for regulatory purposes or when the polymerase hits an obstacle such as a nucleosome. This arrested state is reactivated by the transcription factor TFIIS. Here, a crystal structure is presented of a backtracked yeast Pol II complex in which the backtracked RNA can be observed, plus a structure of a backtracked complex that contains TFIIS. A model is presented for Pol II backtracking, arrest and reactivation during transcription elongation. During gene transcription, RNA polymerase (Pol) II moves forwards along DNA and synthesizes messenger RNA. However, at certain DNA sequences, Pol II moves backwards, and such backtracking can arrest transcription. Arrested Pol II is reactivated by transcription factor IIS (TFIIS), which induces RNA cleavage that is required for cell viability1. Pol II arrest and reactivation are involved in transcription through nucleosomes2,3 and in promoter-proximal gene regulation4,5,6. Here we present X-ray structures at 3.3 Å resolution of an arrested Saccharomyces cerevisiae Pol II complex with DNA and RNA, and of a reactivation intermediate that additionally contains TFIIS. In the arrested complex, eight nucleotides of backtracked RNA bind a conserved ‘backtrack site’ in the Pol II pore and funnel, trapping the active centre trigger loop and inhibiting mRNA elongation. In the reactivation intermediate, TFIIS locks the trigger loop away from backtracked RNA, displaces RNA from the backtrack site, and complements the polymerase active site with a basic and two acidic residues that may catalyse proton transfers during RNA cleavage. The active site is demarcated from the backtrack site by a ‘gating tyrosine’ residue that probably delimits backtracking. These results establish the structural basis of Pol II backtracking, arrest and reactivation, and provide a framework for analysing gene regulation during transcription elongation.Keywords
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