Expression of MicroRNA miR-122 Facilitates an Efficient Replication in Nonhepatic Cells upon Infection with Hepatitis C Virus

Department of Molecular Virology, Osaka University, Osaka, Japan.
Journal of Virology (Impact Factor: 4.65). 05/2012; 86(15):7918-33. DOI: 10.1128/JVI.00567-12
Source: PubMed

ABSTRACT Hepatitis C virus (HCV) is one of the most common etiologic agents of chronic liver diseases, including liver cirrhosis and hepatocellular carcinoma. In addition, HCV infection is often associated with extrahepatic manifestations (EHM), including mixed cryoglobulinemia and non-Hodgkin's lymphoma. However, the mechanisms of cell tropism of HCV and HCV-induced EHM remain elusive, because in vitro propagation of HCV has been limited in the combination of cell culture-adapted HCV (HCVcc) and several hepatic cell lines. Recently, a liver-specific microRNA called miR-122 was shown to facilitate the efficient propagation of HCVcc in several hepatic cell lines. In this study, we evaluated the importance of miR-122 on the replication of HCV in nonhepatic cells. Among the nonhepatic cell lines expressing functional HCV entry receptors, Hec1B cells derived from human uterus exhibited a low level of replication of the HCV genome upon infection with HCVcc. Exogenous expression of miR-122 in several cells facilitates efficient viral replication but not production of infectious particles, probably due to the lack of hepatocytic lipid metabolism. Furthermore, expression of mutant miR-122 carrying a substitution in a seed domain was required for efficient replication of mutant HCVcc carrying complementary substitutions in miR-122-binding sites, suggesting that specific interaction between miR-122 and HCV RNA is essential for the enhancement of viral replication. In conclusion, although miR-122 facilitates efficient viral replication in nonhepatic cells, factors other than miR-122, which are most likely specific to hepatocytes, are required for HCV assembly.

Download full-text


Available from: Daisuke Okuzaki, Jan 07, 2014
  • Source
    • "This enabled us to generate a sub-genomic mutant virus that can replicate independently of miR-122 in both Hep3B cells and Huh7- derived cells. The liver-specific microRNA, miR-122, augments replication of HCV RNA in Huh7-derived cells, and has been used to enhance replication of stable HCV replicons in non-liver cells such as HEK 293 cells and mouse MEFs (Chang et al., 2008; Fukuhara et al., 2012; Jopling et al., 2005; Lin et al., 2010). This led to the development of our hypothesis that miR-122 could be used to permit transient, unselected replication of HCV in other nonpermissive cells. "
    [Show abstract] [Hide abstract]
    ABSTRACT: The study of Hepatitis C Virus (HCV) has benefitted from the use of the Huh7 cell culture system, but until recently there were no other widely used alternatives to this cell line. Here we render another human hepatoma cell line, Hep3B, permissive to the complete virus life cycle by supplementation with the liver-specific microRNA miR-122, known to aid HCV RNA accumulation. When supplemented, Hep3B cells produce J6/JFH-1 virus titres indistinguishable from those produced by Huh7.5 cells. Interestingly, we were able to detect and characterize miR-122-independent replication of di-cistronic replicons in Hep3B cells. Further, we show that Argonaute-2 (Ago2) is required for miR-122-dependent replication, but dispensable for miR-122-independent replication, confirming Ago2's role in mediating the activity of miR-122. Thus Hep3B cells are a model system for the study of HCV, and miR-122 independent replication is a model to identify proteins involved in the function of miR-122.
    Virology 12/2012; 436(1). DOI:10.1016/j.virol.2012.11.007 · 3.28 Impact Factor
  • Source
    [Show abstract] [Hide abstract]
    ABSTRACT: Hepatitis C virus (HCV) exhibits a narrow host range and a specific tissue tropism. Mice expressing major entry receptors for HCV permit viral entry, and therefore the species tropism of HCV infection is considered to be reliant on the expression of the entry receptors. However, HCV receptor candidates are expressed and replication of HCV-RNA can be detected in several nonhepatic cell lines, suggesting that nonhepatic cells are also susceptible to HCV infection. Recently it was shown that the exogenous expression of a liver-specific microRNA, miR-122, facilitated the efficient replication of HCV not only in hepatic cell lines, including Hep3B and HepG2 cells, but also in nonhepatic cell lines, including Hec1B and HEK-293T cells, suggesting that miR-122 is required for the efficient replication of HCV in cultured cells. However, no infectious particle was detected in the nonhepatic cell lines, in spite of the efficient replication of HCV-RNA. In the nonhepatic cells, only small numbers of lipid droplets and low levels of very-low-density lipoprotein-associated proteins were observed compared with findings in the hepatic cell lines, suggesting that functional lipid metabolism participates in the assembly of HCV. Taken together, these findings indicate that miR-122 and functional lipid metabolism are involved in the tissue tropism of HCV infection. In this review, we would like to focus on the role of miR-122 and lipid metabolism in the cell tropism of HCV.
    Journal of Gastroenterology 09/2012; 48(2). DOI:10.1007/s00535-012-0661-5 · 4.02 Impact Factor
  • [Show abstract] [Hide abstract]
    ABSTRACT: As a relatively simple virus, hepatitis C virus (HCV) depends extensively on its host to infect, replicate and disseminate. HCV has evolved host interactions that result in a restricted tropism, both in terms of cell type and species. Efforts into identifying and validating HCV-host interactions have been hampered by a limited number of infectious virus clones and cell lines that support HCV infection. Despite these limitations, consensus HCV-host interactions have emerged that help define the entry, replication, assembly, and tropism of HCV. This has had important implications in expanding our in vitro and in vivo systems to study HCV replication and pathogenesis. Additionally, a number of these host factors are being targeted for therapeutic development. In this review, we focus on medically relevant pro-viral host factors, their role in HCV biology, and their importance in expanding our model systems.
    10/2012; 2(6). DOI:10.1016/j.coviro.2012.09.013