NS5A inhibitors unmask differences in functional replicase complex half-life between different hepatitis C virus strains

Public Deposited
Analytics

Downloadable Content

Download PDF
Request Version for Screen Reader
Creator
  • Benzine, Tiffany
    • Affiliation: University of North Carolina at Chapel Hill
    • Other Affiliation: Department of Medicine
  • Brandt, Ryan
    • Other Affiliation: Department of Mathematics; North Carolina State University
  • Lovell, William C.
    • ORCID: 0000-0001-8625-6581
    • Affiliation: University of North Carolina at Chapel Hill
  • Yamane, Daisuke
    • Affiliation: University of North Carolina at Chapel Hill
    • Other Affiliation: Department of Microbiology and Cell Biology; Tokyo Metropolitan Institute of Medical Science
  • Neddermann, Petra
    • Other Affiliation: INGM -Istituto Nazionale di Genetica Molecolare Romeo ed Enrica Invernizzi
  • De Francesco, Raffaele
    • Other Affiliation: INGM -Istituto Nazionale di Genetica Molecolare Romeo ed Enrica Invernizzi
  • Lemon, Stanley M.
    • Affiliation: University of North Carolina at Chapel Hill
    • Other Affiliation: Department of Medicine
  • Perelson, Alan S.
    • ORCID: 0000-0002-2455-0002
    • Other Affiliation: Theoretical Biology and Biophysics; Los Alamos National Laboratory
  • Ke, Ruian
    • Other Affiliation: Department of Mathematics; North Carolina State University
  • McGivern, David R.
    • Affiliation: University of North Carolina at Chapel Hill
    • Other Affiliation: Department of Medicine
Abstract
  • Hepatitis C virus (HCV) RNA is synthesized by the replicase complex (RC), a macromolecular assembly composed of viral non-structural proteins and cellular co-factors. Inhibitors of the HCV NS5A protein block formation of new RCs but do not affect RNA synthesis by pre-formed RCs. Without new RC formation, existing RCs turn over and are eventually lost from the cell. We aimed to use NS5A inhibitors to estimate the half-life of the functional RC of HCV. We compared different cell culture-infectious strains of HCV that may be grouped based on their sensitivity to lipid peroxidation: robustly replicating, lipid peroxidation resistant (LPOR) viruses (e.g. JFH-1 or H77D) and more slowly replicating, lipid peroxidation sensitive (LPOS) viruses (e.g. H77S.3 and N.2). In luciferase assays, LPOS HCV strains declined under NS5A inhibitor therapy with much slower kinetics compared to LPOR HCV strains. This difference in rate of decline was not observed for inhibitors of the NS5B RNA-dependent RNA polymerase suggesting that the difference was not simply a consequence of differences in RNA stability. In further analyses, we compared two isoclonal HCV variants: the LPOS H77S.3 and the LPOR H77D that differ only by 12 amino acids. Differences in rate of decline between H77S.3 and H77D following NS5A inhibitor addition were not due to amino acid sequences in NS5A but rather due to a combination of amino acid differences in the non-structural proteins that make up the HCV RC. Mathematical modeling of intracellular HCV RNA dynamics suggested that differences in RC stability (half-lives of 3.5 and 9.9 hours, for H77D and H77S.3, respectively) are responsible for the different kinetics of antiviral suppression between LPOS and LPOR viruses. In nascent RNA capture assays, the rate of RNA synthesis decline following NS5A inhibitor addition was significantly faster for H77D compared to H77S.3 indicating different half-lives of functional RCs.
Date of publication
Keyword
DOI
Identifier
Resource type
  • Article
Rights statement
  • In Copyright
Journal title
  • PLoS Pathogens
Journal volume
  • 13
Journal issue
  • 6
Page start
  • e1006343
Language
  • English
ISSN
  • 1553-7374
  • 1553-7366

Relations

Parents:

This work has no parents.

In Collection:

Items

Thumbnail Title Date Uploaded Visibility Actions
file details PubMedCentral-PMC5464671.pdf 2017 Public Download