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Nat Commun. 2016 Apr 19;7:11244. doi: 10.1038/ncomms11244.

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

Author information

1
Department of Chemistry, School of Science and Institute for Advance Study, Hong Kong University of Science and Technology, Clear Water Bay, Kowloon, Hong Kong.
2
Department of Cellular and Molecular Medicine, School of Medicine; Skaggs School of Pharmacy and Pharmaceutical Sciences, University of California San Diego, La Jolla, California 92093, USA.
3
Department of Biochemistry, University of Washington, Seattle, Washington 98195, USA.
4
King Abdullah University of Science and Technology, Computational Bioscience Research Center, Computer, Electrical and Mathematical Sciences and Engineering Division, Thuwal 23955-6900, Saudi Arabia.
5
Division of Biomedical Engineering, School of Science and Institute for Advance Study, Hong Kong University of Science and Technology, Clear Water Bay, Kowloon, Hong Kong.
6
Center of Systems Biology and Human Health, School of Science and Institute for Advance Study, Hong Kong University of Science and Technology, Clear Water Bay, Kowloon, Hong Kong.

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.

PMID:
27091704
PMCID:
PMC4838855
DOI:
10.1038/ncomms11244
[Indexed for MEDLINE]
Free PMC Article

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