Interplay between σ region 3.2 and secondary channel factors during promoter escape by bacterial RNA polymerase

Ivan Petushkov, Daria Esyunina, Vladimir Mekler, Konstantin Severinov, Danil Pupov, Andrey Kulbachinskiy

Research output: Contribution to journalArticlepeer-review

10 Citations (Scopus)

Abstract

In bacterial RNA polymerase (RNAP), conserved region 3.2 of the σ subunit was proposed to contribute to promoter escape by interacting with the 50-end of nascent RNA, thus facilitating σ dissociation. RNAP activity during transcription initiation can also be modulated by protein factors that bind within the secondary channel and reach the enzyme active site. To monitor the kinetics of promoter escape in real time, we used a molecular beacon assay with fluorescently labeled σ70 subunit of Escherichia coli RNAP. We show that substitutions and deletions in σ region 3.2 decrease the rate of promoter escape and lead to accumulation of inactive complexes during transcription initiation. Secondary channel factors differentially regulate this process depending on the promoter and mutations in σ region 3.2. GreA generally increase the rate of promoter escape; DksA also stimulates promoter escape on certain templates, while GreB either stimulates or inhibits this process depending on the template. When observed, the stimulation of promoter escape correlates with the accumulation of stressed transcription complexes with scrunched DNA, while changes in the RNA 50-end structure modulate promoter clearance. Thus, the initiation-To-elongation transition is controlled by a complex interplay between RNAP-binding protein factors and the growing RNA chain.

Original languageEnglish
Pages (from-to)4053-4064
Number of pages12
JournalBiochemical Journal
Volume474
Issue number24
DOIs
Publication statusPublished - 15 Dec 2017
Externally publishedYes

Fingerprint

Dive into the research topics of 'Interplay between σ region 3.2 and secondary channel factors during promoter escape by bacterial RNA polymerase'. Together they form a unique fingerprint.

Cite this