Format

Send to

Choose Destination
Mol Cell. 2016 Sep 15;63(6):939-50. doi: 10.1016/j.molcel.2016.08.011. Epub 2016 Sep 8.

RNA Polymerase Pausing during Initial Transcription.

Author information

1
Biological Physics Research Group, Clarendon Laboratory, Department of Physics, University of Oxford, Oxford OX1 3PU, UK.
2
CNRS FRE 3689, Centre d'études d'agents Pathogénes et Biotechnologies pour la Santé (CPBS), 1919 route de Mende, 34293 Montpellier, France.
3
Biological Physics Research Group, Clarendon Laboratory, Department of Physics, University of Oxford, Oxford OX1 3PU, UK. Electronic address: kapanidis@physics.ox.ac.uk.

Abstract

In bacteria, RNA polymerase (RNAP) initiates transcription by synthesizing short transcripts that are either released or extended to allow RNAP to escape from the promoter. The mechanism of initial transcription is unclear due to the presence of transient intermediates and molecular heterogeneity. Here, we studied initial transcription on a lac promoter using single-molecule fluorescence observations of DNA scrunching on immobilized transcription complexes. Our work revealed a long pause ("initiation pause," ∼20 s) after synthesis of a 6-mer RNA; such pauses can serve as regulatory checkpoints. Region sigma 3.2, which contains a loop blocking the RNA exit channel, was a major pausing determinant. We also obtained evidence for RNA backtracking during abortive initial transcription and for additional pausing prior to escape. We summarized our work in a model for initial transcription, in which pausing is controlled by a complex set of determinants that modulate the transition from a 6- to a 7-nt RNA.

KEYWORDS:

DNA scrunching; RNA polymerase; initial transcription; promoter escape; single-molecule FRET; transcriptional pausing

PMID:
27618490
PMCID:
PMC5031556
DOI:
10.1016/j.molcel.2016.08.011
[Indexed for MEDLINE]
Free PMC Article

Supplemental Content

Full text links

Icon for Elsevier Science Icon for PubMed Central
Loading ...
Support Center