title: Physical and functional interaction between hepatitis C virus NS5A protein and ovarian tumor protein deubiquitinase 7B.
abstract: Hepatitis C virus (HCV) NS5A protein plays crucial roles in viral RNA replication, virus assembly, and viral pathogenesis. Although NS5A has no known enzymatic activity, it modulates various cellular pathways through interaction with cellular proteins. HCV NS5A (and other HCV proteins) are reportedly degraded through the ubiquitin-proteasome pathway; however, the physiological roles of ubiquitylation and deubiquitylation in HCV infection are largely unknown. To elucidate the role of deubiquitylation in HCV infection, an attempt was made to identify a deubiquitinase (DUB) that can interact with NS5A protein. An ovarian tumor protein (OTU), deubiquitinase 7B (OTUD7B), was identified as a novel NS5A-binding protein. Co-immunoprecipitation analyses showed that NS5A interacts with OTUD7B in both Huh-7 and HCV RNA replicon cells. Immunofluorescence staining revealed that HCV NS5A protein colocalizes with OTUD7B in the cytoplasm. Moreover, HCV infection was found to enhance the nuclear localization of OTUD7B. The OTUD7B-binding domain on NS5A was mapped using a series of NS5A deletion mutants. The present findings suggest that the domain I of NS5A is important and the region from amino acid 121 to 126 of NS5A essential for the interaction. Either V121A or V124A mutation in NS5A disrupts the NS5A-OTUD7B interaction. The results of this in vivo ubiquitylation assay suggest that HCV NS5A enhances OTUD7B DUB activity. Taken together, these results suggest that HCV NS5A protein interacts with OTUD7B, thereby modulating its DUB activity.
authors.affiliation: Division of Microbiology, Center for Infectious Diseases, Kobe University Graduate School of Medicine, Kobe, Hyogo, Japan.|Department of Physiology, Faculty of Medicine, Universitas Indonesia, Jakarta, Indonesia.
authors.forename: Imelda Rosalyn
authors.initials: IR
authors.lastname: Sianipar
authors.affiliation: Division of Microbiology, Center for Infectious Diseases, Kobe University Graduate School of Medicine, Kobe, Hyogo, Japan.
authors.forename: Chieko
authors.initials: C
authors.lastname: Matsui
authors.affiliation: Division of Microbiology, Center for Infectious Diseases, Kobe University Graduate School of Medicine, Kobe, Hyogo, Japan.
authors.forename: Nanae
authors.initials: N
authors.lastname: Minami
authors.affiliation: Institute of Biochemistry and Molecular Biology, Hubei University, Wuhan, China.
authors.forename: Xiang
authors.initials: X
authors.lastname: Gan
authors.affiliation: Division of Microbiology, Center for Infectious Diseases, Kobe University Graduate School of Medicine, Kobe, Hyogo, Japan.
authors.forename: Lin
authors.initials: L
authors.lastname: Deng
authors.affiliation: Division of Microbiology, Center for Infectious Diseases, Kobe University Graduate School of Medicine, Kobe, Hyogo, Japan.
authors.forename: Hak
authors.initials: H
authors.lastname: Hotta
authors.affiliation: Division of Microbiology, Center for Infectious Diseases, Kobe University Graduate School of Medicine, Kobe, Hyogo, Japan.
authors.forename: Ikuo
authors.initials: I
authors.lastname: Shoji
mesh_terms: D002460:Cell Line; D003593:Cytoplasm / Q000737:chemistry; D004252:DNA Mutational Analysis; D010450:Endopeptidases / Q000378:metabolism*; D016174:Hepacivirus / Q000502:physiology*; D022781:Hepatocytes / Q000821:virology; D054884:Host-Pathogen Interactions; D006801:Humans; D047468:Immunoprecipitation; D008856:Microscopy, Fluorescence; D016297:Mutagenesis, Site-Directed; D011485:Protein Binding; D054730:Protein Interaction Domains and Motifs; D025941:Protein Interaction Mapping; D017361:Viral Nonstructural Proteins / Q000378:metabolism*; D012324:RNA-Dependent RNA Polymerase
chemical_list: D010450:Endopeptidases; D017361:Viral Nonstructural Proteins; C087036:NS-5 protein, hepatitis C virus; D012324:RNA-Dependent RNA Polymerase; C476458:OTUD7B protein, human
keywords: NS5A; deubiquitinase; hepatitis C virus; ovarian tumor protein deubiquitinase 7B