Backtracking Determines the Force Sensitivity of Rnap Ii in a Factor-dependent Manner.
From: Physical Biosciences Division, Lawrence Berkeley National Laboratory, Berkeley, California 94720, USA.
Nature
- Publish Date: Apr 2007
- ISSN: 1476-4687
- Volume: 446
- Issue: 7137
- Pages: 820-3
- Medium: Internet
- Language: English
- Citation (JAMA): Galburt Eric A, Grill Stephan W, Wiedmann Anna, et al. Backtracking Determines the Force Sensitivity of Rnap Ii in a Factor-dependent Manner.. Nature Apr 2007;446:820-3
Abstract
RNA polymerase II (RNAP II) is responsible for transcribing all messenger RNAs in eukaryotic cells during a highly regulated process that is conserved from yeast to human, and that serves as a central control point for cellular function. Here we investigate the transcription dynamics of single RNAP II molecules from Saccharomyces cerevisiae against force and in the presence and absence of TFIIS, a transcription elongation factor known to increase transcription through nucleosomal barriers. Using a single-molecule dual-trap optical-tweezers assay combined with a novel method to enrich for active complexes, we found that the response of RNAP II to a hindering force is entirely determined by enzyme backtracking. Surprisingly, RNAP II molecules ceased to transcribe and were unable to recover from backtracks at a force of 7.5 +/- 2 pN, only one-third of the force determined for Escherichia coli RNAP. We show that backtrack pause durations follow a t(-3/2) power law, implying that during backtracking RNAP II diffuses in discrete base-pair steps, and indicating that backtracks may account for most of RNAP II pauses. Significantly, addition of TFIIS rescued backtracked enzymes and allowed transcription to proceed up to a force of 16.9 +/- 3.4 pN. Taken together, these results describe a regulatory mechanism of transcription elongation in eukaryotes by which transcription factors modify the mechanical performance of RNAP II, allowing it to operate against higher loads.
Mesh Headings (Keywords): Biomechanics, Escherichia coli, RNA Polymerase II, Saccharomyces cerevisiae, Templates, Genetic, Transcription, Genetic, Transcriptional Elongation Factors
Check for Full Text / PubMed Unique Identifier (PMID): 17361130
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