title: Negative regulation of intracellular hepatitis C virus replication by interferon regulatory factor 3.
abstract: BACKGROUND
  Interferon regulatory factor (IRF)-3 plays an important role in initiating cellular interferon-stimulated gene-mediated antiviral responses. In the present study, we evaluated the effects of IRF-3 expression and activation on intracellular hepatitis C virus (HCV) replication using an HCV replicon system.
  
  
  METHODS
  An HCV replicon was constructed that expressed a neomycin-selectable chimeric firefly luciferase reporter protein. A small interfering (si) RNA oligonucleotide directed against IRF-3 mRNA was designed and synthesized. A eukaryote expression plasmid vector was constructed that expressed IRF-3 mRNA under control of the cytomegalovirus early promoter/enhancer. To evaluate transcriptional activity of the interferon-stimulated genes, a reporter vector was used that expressed firefly luciferase under control of the interferon-stimulated response element (ISRE).
  
  
  RESULTS
  The baseline expression of IRF-3 did not significantly differ between cells with and without expression of the replicon. Transfection of an IRF-3 expression plasmid into the cells raised the ISRE-luciferase activities. The increase of ISRE activity was significantly more potent in the replicon-expressing cells than in cells without replicon expression. Concomitantly, the overexpression of IRF-3 suppressed HCV replication levels. In contrast, siRNA knockdown of IRF-3 suppressed ISRE activity by 38% +/- 2%. Interestingly, the suppression of IRF-3 resulted in a significant increase of HCV replication, by up to twofold, depending on the IRF-3 suppression levels.
  
  
  CONCLUSIONS
  IRF-3 negatively regulated intracellular HCV replication, and was partially activated in cells that expressed the HCV replicon. Thus, IRF-3 is a key molecule controlling HCV replication through modulation of host interferon gene responses.
authors.affiliation: Department of Gastroenterology and Hepatology, Tokyo Medical and Dental University, Bunkyo-ku, Tokyo 113-8519, Japan.
authors.forename: Tsuyoshi
authors.initials: T
authors.lastname: Yamashiro
authors.forename: Naoya
authors.initials: N
authors.lastname: Sakamoto
authors.forename: Masayuki
authors.initials: M
authors.lastname: Kurosaki
authors.forename: Nobuhiko
authors.initials: N
authors.lastname: Kanazawa
authors.forename: Yoko
authors.initials: Y
authors.lastname: Tanabe
authors.forename: Mina
authors.initials: M
authors.lastname: Nakagawa
authors.forename: Cheng-Hsin
authors.initials: CH
authors.lastname: Chen
authors.forename: Yasuhiro
authors.initials: Y
authors.lastname: Itsui
authors.forename: Tomoyuki
authors.initials: T
authors.lastname: Koyama
authors.forename: Yoshie
authors.initials: Y
authors.lastname: Takeda
authors.forename: Shinya
authors.initials: S
authors.lastname: Maekawa
authors.forename: Nobuyuki
authors.initials: N
authors.lastname: Enomoto
authors.forename: Hiroshi
authors.initials: H
authors.lastname: Sakugawa
authors.forename: Mamoru
authors.initials: M
authors.lastname: Watanabe
mesh_terms: D015153:Blotting, Western; D045744:Cell Line, Tumor; D015870:Gene Expression; D015967:Gene Expression Regulation, Viral / Q000502:physiology*; D016174:Hepacivirus / Q000502:physiology*; D006801:Humans; D007150:Immunohistochemistry; D050838:Interferon Regulatory Factor-3 / Q000502:physiology*; D010957:Plasmids; D034741:RNA, Small Interfering; D012093:Replicon; D014162:Transfection; D014779:Virus Replication / Q000502:physiology*
chemical_list: D050838:Interferon Regulatory Factor-3; D034741:RNA, Small Interfering