Elongation-Competent Pauses Govern the Fidelity of a Viral RNA-Dependent RNA Polymerase

David Dulin; Igor D Vilfan; Bojk A Berghuis; Susanne Hage; Dennis H Bamford; Minna M Poranen; Martin Depken; Nynke H Dekker

doi:10.1016/j.celrep.2015.01.031

Elongation-Competent Pauses Govern the Fidelity of a Viral RNA-Dependent RNA Polymerase

Cell Rep. 2015 Feb 17;10(6):983-992. doi: 10.1016/j.celrep.2015.01.031. Epub 2015 Feb 12.

Authors

David Dulin¹, Igor D Vilfan¹, Bojk A Berghuis¹, Susanne Hage¹, Dennis H Bamford², Minna M Poranen³, Martin Depken⁴, Nynke H Dekker⁵

Affiliations

¹ Department of Bionanoscience, Kavli Institute of Nanoscience Delft, Delft University of Technology, Lorentzweg 1, 2628 CJ Delft, the Netherlands.
² Department of Biosciences, University of Helsinki, Viikki Biocenter 2, P.O. Box 56 (Viikinkaari 5), 00014 Helsinki, Finland; Institute of Biotechnology, University of Helsinki, Viikki Biocenter 2, P.O. Box 56 (Viikinkaari 5), 00014 Helsinki, Finland.
³ Department of Biosciences, University of Helsinki, Viikki Biocenter 2, P.O. Box 56 (Viikinkaari 5), 00014 Helsinki, Finland. Electronic address: minna.poranen@helsinki.fi.
⁴ Department of Bionanoscience, Kavli Institute of Nanoscience Delft, Delft University of Technology, Lorentzweg 1, 2628 CJ Delft, the Netherlands. Electronic address: s.m.depken@tudelft.nl.
⁵ Department of Bionanoscience, Kavli Institute of Nanoscience Delft, Delft University of Technology, Lorentzweg 1, 2628 CJ Delft, the Netherlands. Electronic address: n.h.dekker@tudelft.nl.

PMID: 25683720
DOI: 10.1016/j.celrep.2015.01.031

Abstract

RNA viruses have specific mutation rates that balance the conflicting needs of an evolutionary response to host antiviral defenses and avoidance of the error catastrophe. While most mutations are known to originate in replication errors, difficulties of capturing the underlying dynamics have left the mechanochemical basis of viral mutagenesis unresolved. Here, we use multiplexed magnetic tweezers to investigate error incorporation by the bacteriophage Φ6 RNA-dependent RNA polymerase. We extract large datasets fingerprinting real-time polymerase dynamics over four magnitudes in time, in the presence of nucleotide analogs, and under varying NTP and divalent cation concentrations and fork stability. Quantitative analysis reveals a new pause state that modulates polymerase fidelity and so ties viral polymerase pausing to the biological function of optimizing virulence. Adjusting the frequency of such pauses offers a target for therapeutics and may also reflect an evolutionary strategy for virus populations to track the gradual evolution of their hosts.