Hepatitis B virus (HBV) entry has been analyzed using infection-susceptible cells, including primary human hepatocytes, primary tupaia hepatocytes, and HepaRG cells. Recently, the sodium taurocholate cotransporting polypeptide (NTCP) membrane transporter was reported as an HBV entry receptor. In this study, we established a strain of HepG2 cells engineered to overexpress the human NTCP gene (HepG2-hNTCP-C4 cells). HepG2-hNTCP-C4 cells were shown to be susceptible to infection by blood-borne and cell culture-derived HBV. HBV infection was facilitated by pretreating cells with 3% dimethyl sulfoxide permitting nearly 50% of the cells to be infected with HBV. Knockdown analysis suggested that HBV infection of HepG2-hNTCP-C4 cells was mediated by NTCP. HBV infection was blocked by an anti-HBV surface protein neutralizing antibody, by compounds known to inhibit NTCP transporter activity, and by cyclosporin A and its derivatives. The infection assay suggested that cyclosporin B was a more potent inhibitor of HBV entry than was cyclosporin A. Further chemical screening identified oxysterols, oxidized derivatives of cholesterol, as inhibitors of HBV infection. Thus, the HepG2-hNTCP-C4 cell line established in this study is a useful tool for the identification of inhibitors of HBV infection as well as for the analysis of the molecular mechanisms of HBV infection.

Hepatitis C virus (HCV) is a major causative agent of hepatocellular carcinoma. HCV genome replication occurs in the replication complex (RC) around the endoplasmic reticulum membrane. However, the mechanisms regulating the HCV RC remain widely unknown. Here, we used a chemical biology approach to show that estrogen receptor (ESR) is functionally associated with HCV replication. We found that tamoxifen suppressed HCV genome replication. Part of ESRα resided on the endoplasmic reticulum membranes and interacted with HCV RNA polymerase NS5B. RNA interference-mediated knockdown of endogenous ESRα reduced HCV replication. Mechanistic analysis suggested that ESRα promoted NS5B association with the RC and that tamoxifen abrogated NS5B-RC association. Thus, ESRα regulated the presence of NS5B in the RC and stimulated HCV replication. Moreover, the ability of ESRα to regulate NS5B was suggested to serve as a potential novel target for anti-HCV therapeutics.

Interferon-stimulated gene of 20 kDa protein (ISG20) degrades RNA of hepatitis B virus to impede the replication of HBV in vitro and in vivo Interferon-stimulated gene of 20 kDa protein (ISG20) degrades RNA of Hepatitis B virus to impede the replication of HBV in vitro and in vivo