title: Generation of a recombinant reporter hepatitis C virus useful for the analyses of virus entry, intra-cellular replication and virion production.
abstract: The lack of a culture system that efficiently produces progeny virus has hampered hepatitis C virus (HCV) research. Recently, the discovery of a novel HCV isolate JFH1 and its chimeric derivative J6/JFH1 has led to the development of an efficient virus productive culture system. To construct an easy monitoring system for the viral life cycle of HCV, we generated bicistronic luciferase reporter virus genomes based on the JFH1 and J6/JFH1 isolates, respectively. Transfection of the J6/JFH1-based reporter genome to Huh7.5 cells produced significantly greater levels of progeny virus than transfection of the JFH1 genome. Furthermore, the expression of dominant-negative Vps4, a key molecule of the endosomal sorting complex required for transport machinery, inhibited the virus production of JFH1, but not that of J6/JFH1. These results may account for the different abilities to produce progeny virus between JFH1 and J6/JFH1. Using the J6/JFH1/Luc system, we showed that the two polyanions heparin and polyvinyl sulfate decreased the infectivity of J6/JFH1/Luc virus in a dose-dependent manner. We also analyzed the function of microRNA on HCV replication and found that miR-34b could affect the replication of HCV. The reporter virus generated in this study will be useful for investigating the nature of the HCV life cycle and for identification of HCV inhibitors.
authors.affiliation: Division of Microbiology, Kobe University, Graduate School of Medicine, 7-5-1 Kusunoki-cho, Cyuo-ku, Kobe, Hyogo 650-0017, Japan.
authors.forename: Kazuya
authors.initials: K
authors.lastname: Kamada
authors.forename: Ikuo
authors.initials: I
authors.lastname: Shoji
authors.forename: Lin
authors.initials: L
authors.lastname: Deng
authors.forename: Chie
authors.initials: C
authors.lastname: Aoki
authors.forename: Suratno Lulut
authors.initials: SL
authors.lastname: Ratnoglik
authors.forename: Takaji
authors.initials: T
authors.lastname: Wakita
authors.forename: Hak
authors.initials: H
authors.lastname: Hotta
mesh_terms: D002460:Cell Line; D017930:Genes, Reporter; D016679:Genome, Viral; D016174:Hepacivirus / Q000235:genetics / Q000502:physiology*; D006526:Hepatitis C / Q000821:virology*; D006801:Humans; D008156:Luciferases / Q000096:biosynthesis / Q000235:genetics; D014162:Transfection; D014771:Virion / Q000235:genetics / Q000502:physiology*; D014776:Virus Cultivation / Q000379:methods; D014779:Virus Replication / Q000502:physiology*
chemical_list: D008156:Luciferases