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Proc Natl Acad Sci U S A. 2008 Mar 18;105(11):4370-5. doi: 10.1073/pnas.0800422105. Epub 2008 Mar 11.

Advantages of a single-cycle production assay to study cell culture-adaptive mutations of hepatitis C virus.

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  • 1Hepatitis Viruses and Molecular Hepatitis Sections, Laboratory of Infectious Diseases, National Institute of Allergy and Infectious Diseases, National Institutes of Health, Bethesda, MD 20892, USA.

Abstract

The JFH1 strain of hepatitis C virus (HCV) is unique among HCV isolates, in that the wild-type virus can traverse the entire replication cycle in cultured cells. However, without adaptive mutations, only low levels of infectious virus are produced. In the present study, the effects of five mutations that were selected during serial passage in Huh-7.5 cells were studied. Recombinant genomes containing all five mutations produced 3-4 logs more infectious virions than did wild type. Neither a coding mutation in NS5A nor a silent mutation in E2 was adaptive, whereas coding mutations in E2, p7, and NS2 all increased virus production. A single-cycle replication assay in CD81-deficient cells was developed to study more precisely the effect of the adaptive mutations. The E2 mutation had minimal effect on the amount of infectious virus released but probably enhanced entry into cells. In contrast, both the p7 and NS2 mutations independently increased the amount of virus released.

PMID:
18334634
PMCID:
PMC2393785
DOI:
10.1073/pnas.0800422105
[PubMed - indexed for MEDLINE]
Free PMC Article
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