title: Studying transcription initiation by RNA polymerase with diffusion-based single-molecule fluorescence.
abstract: Over the past decade, fluorescence-based single-molecule studies significantly contributed to characterizing the mechanism of RNA polymerase at different steps in transcription, especially in transcription initiation. Transcription by bacterial DNA-dependent RNA polymerase is a multistep process that uses genomic DNA to synthesize complementary RNA molecules. Transcription initiation is a highly regulated step in E. coli, but it has been challenging to study its mechanism because of its stochasticity and complexity. In this review, we describe how single-molecule approaches have contributed to our understanding of transcription and have uncovered mechanistic details that were not observed in conventional assays because of ensemble averaging.
authors.affiliation: Interdepartmental Program in Molecular, Cellular, and Integrative Physiology, University of California, Los Angeles, California, 90095.
authors.forename: Yazan
authors.initials: Y
authors.lastname: Alhadid
authors.affiliation: Department of Chemistry & Biochemistry, University of California, Los Angeles, California, 90095.
authors.forename: SangYoon
authors.initials: S
authors.lastname: Chung
authors.affiliation: Department of Chemistry & Biochemistry, University of California, Los Angeles, California, 90095.
authors.forename: Eitan
authors.initials: E
authors.lastname: Lerner
authors.affiliation: Department of Chemistry & Biochemistry, University of Colorado, Boulder, Colorado, 80303.
authors.forename: Dylan J
authors.initials: DJ
authors.lastname: Taatjes
authors.affiliation: Rowan University School of Osteopathic Medicine, Stratford, New Jersey, 08084.
authors.forename: Sergei
authors.initials: S
authors.lastname: Borukhov
authors.affiliation: Interdepartmental Program in Molecular, Cellular, and Integrative Physiology, University of California, Los Angeles, California, 90095.|Department of Chemistry & Biochemistry, University of California, Los Angeles, California, 90095.|Molecular Biology Institute (MBI), University of California, Los Angeles, California, 90095.|California NanoSystems Institute, University of California, Los Angeles, California, 90095.|Department of Physiology, University of California, Los Angeles, California, 90095.
authors.forename: Shimon
authors.initials: S
authors.lastname: Weiss
mesh_terms: D004269:DNA, Bacterial / Q000235:genetics / Q000378:metabolism*; D004259:DNA-Directed DNA Polymerase / Q000235:genetics / Q000378:metabolism*; D004926:Escherichia coli / Q000235:genetics / Q000378:metabolism*; D029968:Escherichia coli Proteins / Q000235:genetics / Q000378:metabolism*; D005453:Fluorescence; D012329:RNA, Bacterial / Q000096:biosynthesis* / Q000235:genetics; D061785:Transcription Initiation, Genetic / Q000502:physiology*
chemical_list: D004269:DNA, Bacterial; D029968:Escherichia coli Proteins; D012329:RNA, Bacterial; D004259:DNA-Directed DNA Polymerase
keywords: FRET; RNA polymerase; alternating laser excitation; single molecule; transcription