title: Alternative endocytosis pathway for productive entry of hepatitis C virus.
abstract: Previous studies have shown that hepatitis C virus (HCV) enters human hepatic cells through interaction with a series of cellular receptors, followed by clathrin-mediated, pH-dependent endocytosis. Here, we investigated the mechanisms of HCV entry into multiple HCV-permissive human hepatocyte-derived cells using trans-complemented HCV particles (HCVtcp). Knockdown of CD81 and claudin-1, or treatment with bafilomycin A1, reduced infection in Huh-7 and Huh7.5.1 cells, suggesting that HCV entered both cell types via receptor-mediated, pH-dependent endocytosis. Interestingly, knockdown of the clathrin heavy chain or dynamin-2 (Dyn2), as well as expression of the dominant-negative form of Dyn2, reduced infection of Huh-7 cells with HCVtcp, whereas infectious entry of HCVtcp into Huh7.5.1 cells was not impaired. Infection of Huh7.5.1 cells with culture-derived HCV (HCVcc) via a clathrin-independent pathway was also observed. Knockdown of caveolin-1, ADP-ribosylation factor 6 (Arf6), flotillin, p21-activated kinase 1 (PAK1) and the PAK1 effector C-terminal binding protein 1 of E1A had no inhibitory effects on HCVtcp infection into Huh7.5.1 cells, thus suggesting that the infectious entry pathway of HCV into Huh7.5.1 cells was not caveolae-mediated, or Arf6- and flotillin-mediated endocytosis and macropinocytosis, but rather may have occurred via an undefined endocytic pathway. Further analysis revealed that HCV entry was clathrin- and dynamin-dependent in ORL8c and HepCD81/miR122 cells, but productive entry of HCV was clathrin- and dynamin-independent in Hep3B/miR122 cells. Collectively, these data indicated that HCV entered different target cells through different entry routes.
authors.affiliation: Department of Virology II, National Institute of Infectious Diseases, Tokyo, Japan.
authors.forename: Mami
authors.initials: M
authors.lastname: Matsuda
authors.affiliation: Department of Virology II, National Institute of Infectious Diseases, Tokyo, Japan.
authors.forename: Ryosuke
authors.initials: R
authors.lastname: Suzuki
authors.affiliation: Department of Virology II, National Institute of Infectious Diseases, Tokyo, Japan.
authors.forename: Chikako
authors.initials: C
authors.lastname: Kataoka
authors.affiliation: Department of Virology II, National Institute of Infectious Diseases, Tokyo, Japan.
authors.forename: Koichi
authors.initials: K
authors.lastname: Watashi
authors.affiliation: Department of Virology II, National Institute of Infectious Diseases, Tokyo, Japan.
authors.forename: Hideki
authors.initials: H
authors.lastname: Aizaki
authors.affiliation: Department of Tumor Virology, Okayama University Graduate School of Medicine, Dentistry, and Pharmaceutical Sciences, Okayama, Japan.
authors.forename: Nobuyuki
authors.initials: N
authors.lastname: Kato
authors.affiliation: Research Institute for Microbial Diseases, Osaka University, Osaka, Japan.
authors.forename: Yoshiharu
authors.initials: Y
authors.lastname: Matsuura
authors.affiliation: Department of Infectious Diseases, Hamamatsu University School of Medicine, Shizuoka, Japan.
authors.forename: Tetsuro
authors.initials: T
authors.lastname: Suzuki
authors.affiliation: Department of Virology II, National Institute of Infectious Diseases, Tokyo, Japan.
authors.forename: Takaji
authors.initials: T
authors.lastname: Wakita
mesh_terms: D000089982:ADP-Ribosylation Factor 6; D002460:Cell Line; D004705:Endocytosis / Q000502:physiology*; D005786:Gene Expression Regulation / Q000502:physiology; D016174:Hepacivirus / Q000502:physiology*; D022781:Hepatocytes / Q000378:metabolism / Q000502:physiology / Q000821:virology; D006801:Humans; D053586:Virus Internalization