Bridge helix bending promotes RNA polymerase II backtracking through a critical and conserved threonine residue

Lin-Tai Da, Fátima Pardo-Avila, Liang Xu, Daniel-Adriano Silva, Lu Zhang, Xin Gao, Dong Wang, Xuhui Huang

Research output: Contribution to journalArticlepeer-review

38 Scopus citations

Abstract

The dynamics of the RNA polymerase II (Pol II) backtracking process is poorly understood. We built a Markov State Model from extensive molecular dynamics simulations to identify metastable intermediate states and the dynamics of backtracking at atomistic detail. Our results reveal that Pol II backtracking occurs in a stepwise mode where two intermediate states are involved. We find that the continuous bending motion of the Bridge helix (BH) serves as a critical checkpoint, using the highly conserved BH residue T831 as a sensing probe for the 3′-terminal base paring of RNA:DNA hybrid. If the base pair is mismatched, BH bending can promote the RNA 3′-end nucleotide into a frayed state that further leads to the backtracked state. These computational observations are validated by site-directed mutagenesis and transcript cleavage assays, and provide insights into the key factors that regulate the preferences of the backward translocation.
Original languageEnglish (US)
JournalNature Communications
Volume7
Issue number1
DOIs
StatePublished - Apr 19 2016

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