A conserved proline between domains II and III of hepatitis C virus NS5A influences both RNA replication and virus assembly

Hughes, M., Gretton, S., Shelton, H., Brown, D.D., McCormick, C.J., Angus, A.G.N., Patel, A.H., Griffin, S. and Harris, M. (2009) A conserved proline between domains II and III of hepatitis C virus NS5A influences both RNA replication and virus assembly. Journal of Virology, 83(20), pp. 10788-10796. (doi: 10.1128/JVI.02406-08) (PMID:19656877) (PMCID:PMC2753128)

Abstract

We previously demonstrated that two closely spaced polyproline motifs, with the consensus sequence Pro-X-X-Pro-X-Lys/Arg, located between residues 343-356 of NS5A, mediated interactions with cellular SH3 domains. The N-terminal motif (termed PP2.1) is only conserved in genotype 1 isolates whereas the C-terminal motif (PP2.2) is conserved throughout all HCV isolates although this motif was shown to be dispensable for replication of the genotype 1b sub-genomic replicon. In order to investigate the potential role of these motifs in the viral lifecycle we have undertaken a detailed mutagenic analysis of these proline residues in the context of both genotype 1b (FK5.1) or 2a sub-genomic replicons and the genotype 2a infectious clone, JFH-1. We show that the PP2.2 motif is dispensable for RNA replication of all sub-genomic replicons and furthermore is not required for virus production in JFH-1. In contrast, the PP2.1 motif is only required for genotype 1b RNA replication. Mutation of proline 346 within PP2.1 to alanine dramatically attenuated genotype 1b replicon replication in three distinct genetic backgrounds, but the corresponding proline 342 was not required for replication of the JFH-1 subgenomic replicon. However, the P342A mutation resulted in both a delay to virus release and a modest (up to 10-fold) reduction in virus production. These data point to critical roles for these proline residues at multiple stages in the HCV lifecycle, however they also caution against extrapolation of data from culture adapted replicons to infectious virus

• Item Type: Articles
• Status: Published
• Refereed: Yes
• Glasgow Author(s) Enlighten ID: Angus, Dr Allan and Patel, Professor Arvind
• Authors: Hughes, M., Gretton, S., Shelton, H., Brown, D.D., McCormick, C.J., Angus, A.G.N., Patel, A.H., Griffin, S., and Harris, M.
• Subjects: Q Science > QR Microbiology > QR355 Virology
• College/School: College of Medical Veterinary and Life Sciences; College of Medical Veterinary and Life Sciences > School of Infection & Immunity > Centre for Virus Research
• Journal Name: Journal of Virology
• Journal Abbr.: J. Virol.
• Publisher: American Society for Microbiology
• ISSN: 0022-538X
• ISSN (Online): 1098-5514
• Published Online: 05 August 2009