Pausing controls branching between productive and non-productive pathways during initial transcription in bacteria

David Dulin (), David L. V. Bauer, Anssi M. Malinen, Jacob J. W. Bakermans, Martin Kaller, Zakia Morichaud, Ivan Petushkov, Martin Depken, Konstantin Brodolin, Andrey Kulbachinskiy and Achillefs N. Kapanidis ()
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David Dulin: University of Oxford
David L. V. Bauer: University of Oxford
Anssi M. Malinen: University of Oxford
Jacob J. W. Bakermans: University of Oxford
Martin Kaller: University of Oxford
Zakia Morichaud: Institut de Recherche en Infectiologie de Montpellier (IRIM) UMR9004 CNRS-Université de Montpellier
Ivan Petushkov: Russian Academy of Sciences
Martin Depken: Kavli Institute of Nanoscience, Delft University of Technology
Konstantin Brodolin: Institut de Recherche en Infectiologie de Montpellier (IRIM) UMR9004 CNRS-Université de Montpellier
Andrey Kulbachinskiy: Russian Academy of Sciences
Achillefs N. Kapanidis: University of Oxford

Nature Communications, 2018, vol. 9, issue 1, 1-12

Abstract: Abstract Transcription in bacteria is controlled by multiple molecular mechanisms that precisely regulate gene expression. It has been recently shown that initial RNA synthesis by the bacterial RNA polymerase (RNAP) is interrupted by pauses; however, the pausing determinants and the relationship of pausing with productive and abortive RNA synthesis remain poorly understood. Using single-molecule FRET and biochemical analysis, here we show that the pause encountered by RNAP after the synthesis of a 6-nt RNA (ITC6) renders the promoter escape strongly dependent on the NTP concentration. Mechanistically, the paused ITC6 acts as a checkpoint that directs RNAP to one of three competing pathways: productive transcription, abortive RNA release, or a new unscrunching/scrunching pathway. The cyclic unscrunching/scrunching of the promoter generates a long-lived, RNA-bound paused state; the abortive RNA release and DNA unscrunching are thus not as tightly linked as previously thought. Finally, our new model couples the pausing with the abortive and productive outcomes of initial transcription.

Date: 2018
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DOI: 10.1038/s41467-018-03902-9

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