Visualisation and analysis of hepatitis C virus non-structural proteins using super-resolution microscopy

Abstract

Hepatitis C virus (HCV) RNA replication occurs in the cytosol of infected cells within a specialised membranous compartment. How the viral non-structural (NS) proteins are associated and organised within these structures remains poorly defined. We employed a super-resolution microscopy approach to visualise NS3 and NS5A in HCV infected cells. Using single molecule localisation microscopy, both NS proteins were resolved as clusters of localisations smaller than the diffraction-limited volume observed by wide-field. Analysis of the protein clusters identified a significant difference in size between the NS proteins. We also observed a reduction in NS5A cluster size following inhibition of RNA replication using daclatasvir, a phenotype which was maintained in the presence of the Y93H resistance associated substitution and not observed for NS3 clusters. These results provide insight into the NS protein organisation within hepatitis C virus RNA replication complexes and the mode of action of NS5A inhibitors.

Metadata

Authors/Creators:

Bartlett, C
Curd, A https://orcid.org/0000-0002-3949-7523
Peckham, M https://orcid.org/0000-0002-3754-2028
Harris, M https://orcid.org/0000-0002-9821-1003

© The Author(s) 2018. This article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons license, and indicate if changes were made. The images or other third party material in this article are included in the article’s Creative Commons license, unless indicated otherwise in a credit line to the material. If material is not included in the article’s Creative Commons license and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this license, visit http://creativecommons.org/licenses/by/4.0/.

Dates: