Gene expression, DNA replication and genome maintenance are all initiated by proteins that must recognize specific targets from among a vast excess of nonspecific DNA. For example, to initiate transcription, Escherichia coli RNA polymerase (RNAP) must locate promoter sequences, which compose <2% of the bacterial genome. This search problem remains one of the least understood aspects of gene expression, largely owing to the transient nature of search intermediates. Here we visualize RNAP in real time as it searches for promoters, and we develop a theoretical framework for analyzing target searches at the submicroscopic scale on the basis of single-molecule target-association rates. We demonstrate that, contrary to long-held assumptions, the promoter search is dominated by three-dimensional diffusion at both the microscopic and submicroscopic scales in vitro, which has direct implications for understanding how promoters are located within physiological settings.
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We thank M. Gottesman, R. Gonzalez and the Greene laboratory for assistance and discussion throughout this work. We thank M. Gottesman, D. Duzdevich and members of our laboratories for carefully reading the manuscript. We thank R. Landick (University of Wisconsin–Madison, Madison, Wisconsin, USA) for providing RNAP expression constructs. This work was supported by US National Institutes of Health grants GM074739 (E.C.G.) and F32GM80864 (I.J.F.) and training grant T32GM00879807 (J.G.), a US National Science Foundation Award (E.C.G.) and by the Howard Hughes Medical Institute.
The authors declare no competing financial interests.
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Wang, F., Redding, S., Finkelstein, I. et al. The promoter-search mechanism of Escherichia coli RNA polymerase is dominated by three-dimensional diffusion. Nat Struct Mol Biol 20, 174–181 (2013). https://doi.org/10.1038/nsmb.2472
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