肝炎ウイルス蛋白質の細胞内輸送、品質管理に機能する宿主因子の探索

2009 Fiscal Year Annual Research Report

• Project/Area Number: 20659121
• Research Institution: National Institute of Infectious Diseases
• Principal Investigator: 鈴木 哲朗 National Institute of Infectious Diseases, ウイルス第二部, 室長 (00250184)
• Co-Investigator(Kenkyū-buntansha): 鈴木 亮介 国立感染症研究所, ウイルス第二部, 主任研究官 (50342902)
• Keywords: ウイルス / 感染症 / 蛋白質 / プロテオーム

Research Abstract

昨年度、C型肝炎ウイルス(HCV)の産生制御に小胞体関連品質管理機構(ERAD)が関与することを示した。ERADの調節分子EDEM1の遺伝子サイレンシングによって感染性HCV産生が亢進することを見出した。本年度、HCV生活環におけるERADの役割について更に詳細な解析を行った。HCV感染細胞へのsiRNA導入によるウイルス産生の上昇はEDEM1, EDEM3では認められるがEDEM2では見られなかった。一方各EDEMを強制発現したところ、EDEM2によってHCV産生が低下することがわかった。N型糖鎖修飾を受けるHCVエンベロープ蛋白E1、E2と各EDEMとの相互作用を免疫沈降法で解析したところ、E1, E2はEDEM1、EDEM2、EDEM3それぞれと相互作用することが示された。さらに、ERAD系で基質ユビキチン化を担うユビキチンリガーゼ複合体の構成因子であるSEL1LとEDEMの相互作用をHCV E2蛋白存在下で解析したところ、EDEM1、EDEM3はSEL1との相互作用が認められたが、EDEM2-SEL1L複合体は認められなかった。また、小胞体でのマンノーストリミングをマンノシダーゼ阻害剤Kifunensine処理によって抑制すると、HCV産生はKifunensineの濃度依存的に亢進した。
ERADは小胞体でミスフォールドした蛋白がサイトゾルへ引き出されユビキチン-プロテアソーム系で分解されるシステムであるが、今回の成績から、HCVの生活環においては成熟化途中あるいは成熟化した糖蛋白の一部もERADで分解され、結果的にウイルス産生レベルの低減に繋がると考えられた。HCV E1、E2はマンノーストリミング後、EDEM1/EDEM3に認識されユビキチンリガーゼ複合体ヘリクルートされるが、EDEM2はこの過程に競合的に働く。正負両面のERAD調節機構がHCV粒子産生に関与しているものと結論づけた。

Research Products

• [Journal Article] Identification of hepatitis C virus genotype 2a replicon variants with reduced susceptibility to ribavirin (2010)
  • Author(s): Hmwe SS, Aizaki H, Date T, Murakami K, Ishii K, Miyamura T, Koike K, Wakita T, Suzuki T
  • Journal Title: Antiviral Res 85 Pages: 520-524
  • Peer Reviewed
• [Journal Article] Production of infectious hepatitis C virus by using RNA polymerase I-mediated transcription (2010)
  • Author(s): Masaki T, Suzuki R, Saeed M, Mori K, Matsuda M, Aizaki H, Ishii K, Maki N, Miyamura T, Matsuura Y, Wakita T, Suzuki T
  • Journal Title: J Virol (In press)
  • Peer Reviewed
• [Journal Article] Hepatitis C virus proteins modulate microRNA expression and chemosensitivity in malignant hepatocytes (2010)
  • Author(s): Braconi C, Valeri N, Gasparini P, Huang N, Taccioli C, Nuovo G, Suzuki T, Croce CM, Patel T
  • Journal Title: Clin Cancer Res 16 Pages: 957-966
  • Peer Reviewed
• [Journal Article] Establishment of an indicator cell system for hepatitis C virus (2010)
  • Author(s): Tanaka Y, Mori Y, Tani H, Abe T, Moriishi K, Kojima H, Nagano T, Okabe T, Suzuki T, Tatsumi M, Matsuura Y
  • Journal Title: Microbiol Immunol 54 Pages: 206-220
  • Peer Reviewed
• [Journal Article] Biological properties of purified recombinant HCV particles with anepitope-tagged envelope (2010)
  • Author(s): Takahashi H, Akazawa D, Kato T, Date T, Shirakura M, Nakamura N, Mochizuki H, Tanaka-Kaneko K, Sata T, Tanaka Y, Mizokami M, Suzuki T, Wakita T
  • Journal Title: Biochem Biophys Res Commun (In press)
  • Peer Reviewed
• [Journal Article] Enhanced expression of lymphomagenesis-related genes in peripheral blood B cells of chronic hepatitis C patients (2010)
  • Author(s): Ito M, Murakami K, Suzuki T, Mochida K, Suzuki M, Ikebuchi K, Yamaguchi K, Mizuochi T
  • Journal Title: Clin Immunol (In press)
  • Peer Reviewed
• [Journal Article] Involvement of creatine kinase B in hepatitis C virus genome replication through interaction with the viral NS4A protein (2009)
  • Author(s): Hara, H., Aizaki, H., Matsuda, M., Shinkai-Ouchi, F., Inoue, Y., Murakami, K., Shoji, I., Kawakami, H., Matsuura, Y., Lai, M.M.C., Miyamura, T., Wakita, T., Suzuki, T.
  • Journal Title: J Virol 83 Pages: 5137-5147
  • Peer Reviewed
• [Journal Article] Human VAP-C negatively regulates hepatitis C virus propagation (2009)
  • Author(s): Kukihara H, Moriishi K, Taguwa S, Tani H, Abe T, Mori Y, Suzuki T, Fukuhara T, Taketomi A, Maehara Y, Matsuura Y
  • Journal Title: J Virol 83 Pages: 7959-7969
  • Peer Reviewed
• [Journal Article] Cellular vimentin content regulates the protein level of Hepatitis C viruscore protein and the Hepatitis C virus production in cultured cells (2009)
  • Author(s): Tsutsumi T, Matsuda M, Aizaki H, Moriya K, Miyoshi H, Fujie H, Shintani Y, Yotsuyanagi H, Miyamura T, Suzuki T, Koike K
  • Journal Title: Hepatology 50 Pages: 378-386
  • Peer Reviewed
• [Journal Article] Identification of Annexin Al as a novel substrate for E6AP-mediated ubiquitylation (2009)
  • Author(s): Shimoji T, Murakami K, Sugiyama Y, Matsuda M, Inubushi S, Nasu J, Shirakura M, Suzuki T, Wakita T, Kishino T, Hotta H, Miyamura T, Shoji I
  • Journal Title: J Cell Biochem 16 Pages: 1123-1135
  • Peer Reviewed
• [Journal Article] Tacrolimus ameliorates metabolic disturbance and oxidative stress caused by hepatitis C virus core protein : analysis using mouse model and cultured cells (2009)
  • Author(s): Moriya K, Miyoshi H, Tsutsumi T, Shinzawa S, Fujie H, Shintani Y, Yotsuyanagi H, Moriishi K, Matsuura Y, Suzuki T, Miyamura T, Koike K
  • Journal Title: Am J Pathol 175 Pages: 1515-1524
  • Peer Reviewed
• [Journal Article] Identification of bisindolylmaleimides and indolocarbazoles as inhibitors of HCV replication by tube-capture-RT-PCR (2009)
  • Author(s): Murakami Y, Noguchi K, Yamagoe S, Suzuki T, Wakita T, Fukasawa H
  • Journal Title: Antiviral Res 83 Pages: 112-117
  • Peer Reviewed
• [Journal Article] Co-chaperone activity of human butyrate-induced transcript 1 facilitates hepatitis C virus replication through an Hsp90-dependent pathway (2009)
  • Author(s): Taguwa S, Kambara H, Omori H, Tani H, Abe T, Mori Y, Suzuki T, Yoshimori T, Moriishi K, Matsuura Y
  • Journal Title: J Virol 83 Pages: 10427-10436
  • Peer Reviewed
• [Journal Article] Evaluation of hepatitis C virus core antigen assays in detecting recombinant viral antigens of various genotypes (2009)
  • Author(s): Saeed M, Suzuki R, Kondo M, Aizaki H, Kato T, Mizuochi T, Wakita T, Watanabe H, Suzuki T
  • Journal Title: J Clin Microbiol 47 Pages: 4141-4143
  • Peer Reviewed
• [Presentation] Plasmid-based production of trans-complemented HCV particles : its use for functional analysis of NS2 (2009)
  • Author(s): Suzuki R, Saito K, Ando T, Ishii K, Matsuura Y, Miyamura T, Wakita T, Suzuki T
  • Organizer: 16th International Symposium on Hepatitis C Virus and Related Viruses
  • Place of Presentation: Nice, France
  • Year and Date: 20091003-20091007
• [Presentation] Identification of novel serine/threonine protein kinases responsible for HCV NS5A phosphorylation (2009)
  • Author(s): Masaki T, Matsunaga S, Takahashi H, Sawasaki T, Wakita T, Suzuki T.
  • Organizer: 16th International Symposium on Hepatitis C Virus and Related Viruses
  • Place of Presentation: Nice, France
  • Year and Date: 20091003-20091007
• [Presentation] Structural requirements of virion-associated cholesterol for HCV morphogenesis and infectivity (2009)
  • Author(s): Yamamoto M, Aizaki H, Goto K, Hamano K, Miyamura T, Wakita T, Suzuki T.
  • Organizer: 16th International Symposium on Hepatitis C Virus and Related Viruses
  • Place of Presentation: Nice, France
  • Year and Date: 20091003-20091007
• [Book] HCVの分子生物学 (2009)
  • Author(s): 政木隆博、鈴木哲朗
  • Total Pages: 660(担当分:4)
  • Publisher: 日本臨床