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Vps4 and the ESCRT-III complex are required for the release of infectious hepatitis C virus particles

¹ University of Leeds;
² Department of Pathology, University of Cambridge

³ E-mail: m.harris{at}leeds.ac.uk

The mechanisms by which infectious hepatitis C virus (HCV) particles are assembled and released from infected cells remain poorly characterised. In this regard, many other enveloped viruses, notably HIV-1, have been shown to utilise the host vacuolar protein sorting machinery (also known as the endosomal sorting complex required for transport or ESCRT) to traffic through the cell and effect the membrane rearrangements required for the formation of enveloped particles. We postulated that this might also apply to HCV. To test this hypothesis we established a method of conditional virus-like particle assembly involving trans-complementation of an envelope-deleted JFH-1 genome using plasmid transfection. This system reliably produced virus particles that were infectious and could be easily enumerated by focus-forming assay in Huh7 cells. Following co-transfection with plasmids expressing various dominant-negative forms of either components of the ESCRT-III complex, or Vps4 (the AAA ATPase that recycles the ESCRT complexes), a reduction in particle production was seen. No significant effect was observed after co-transfection of dominant-negative ESCRT-I or Alix, an ESCRT associated protein. Dominant-negative Vps4 or ESCRT-III components had no effect on either virus genome replication, or the accumulation of intracellular infectious particles. These data were confirmed using cell culture infectious HCV and we conclude, therefore, that HCV requires late components of the ESCRT pathway for release of infectious virus particles.

Received 1 October 2009; accepted 8 October 2009.