Abstract:
:Transcription initiation is a major step in gene regulation for all organisms. In bacteria, the promoter DNA is first recognized by RNA polymerase (RNAP) to yield an initial closed complex. This complex subsequently undergoes conformational changes resulting in DNA strand separation to form a transcription bubble and an RNAP-promoter open complex; however, the series and sequence of conformational changes, and the factors that influence them are unclear. To address the conformational landscape and transitions in transcription initiation, we applied single-molecule Förster resonance energy transfer (smFRET) on immobilized Escherichia coli transcription open complexes. Our results revealed the existence of two stable states within RNAP-DNA complexes in which the promoter DNA appears to adopt closed and partially open conformations, and we observed large-scale transitions in which the transcription bubble fluctuated between open and closed states; these transitions, which occur roughly on the 0.1 s timescale, are distinct from the millisecond-timescale dynamics previously observed within diffusing open complexes. Mutational studies indicated that the σ70 region 3.2 of the RNAP significantly affected the bubble dynamics. Our results have implications for many steps of transcription initiation, and support a bend-load-open model for the sequence of transitions leading to bubble opening during open complex formation.
journal_name
Nucleic Acids Resjournal_title
Nucleic acids researchauthors
Duchi D,Gryte K,Robb NC,Morichaud Z,Sheppard C,Brodolin K,Wigneshweraraj S,Kapanidis ANdoi
10.1093/nar/gkx1146subject
Has Abstractpub_date
2018-01-25 00:00:00pages
677-688issue
2eissn
0305-1048issn
1362-4962pii
4643373journal_volume
46pub_type
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