mRNAの品質管理システム(mRNAサーベイランス)の分子機序:ヒトSMG1の構造と機能の解析

• 研究課題/領域番号: 13480211
• 研究種目: 基盤研究(B)
• 配分区分: 補助金
• 応募区分: 一般
• 研究分野: 機能生物化学
• 研究機関: 横浜市立大学
• 研究代表者: 大野 茂男 横浜市立大学, 医学部, 教授 (10142027)
• 研究分担者: 秋本 和憲 横浜市立大学, 医学部, 助手 (70285104) ; 平井 秀一 横浜市立大学, 医学部, 助教授 (80228759)
• 研究期間 (年度): 2001 – 2002
• 研究課題ステータス: 完了 (2002年度)
• 配分額: 15,200千円 (直接経費: 15,200千円); 2002年度: 6,600千円 (直接経費: 6,600千円); 2001年度: 8,600千円 (直接経費: 8,600千円)
• キーワード: ナンセンス変異 / mRNA分解 / mRNAサーベイランス / p53 / プロテインキナーゼ / リン脂質類似キナーゼ / RNAヘリカーゼ / ADPアーゼ / mRNA分離 / リン脂質キナーゼ / ATPアーゼ

研究概要

遺伝子の変異や転写スプライシングの誤りなどから生体を守る防御機構として、ナンセンス変異を持つ異常mRNAを選択的に分解する機構(mRNAサーベイランス或いはnonsense mediated mRNA decay, NMD)が存在する。本研究では、酵母と線虫の遺伝学で明らかとなったNMDに関わる遺伝子群のヒト版の解析を進め、線虫の7種のNMD関連遺伝子全てについてそれらのヒトホモログを遺伝子タンパク質レベルで同定すると同時にほ乳類細胞におけるNMDの分子機構と生理的な意義の解析を進めた。
その結果、hSMG-1というプロテインキナーゼによるhUPF1の特定部位のリン酸化が、NMDに必須の過程であることを見いだした。さらに、hSMG-1の阻害剤を見いだし、ほ乳類細胞を用いてNMDを選択的に阻害してトランケート型のタンパク質を誘導できることを示し、変異遺伝子検索の新手法を可能とした。また、hSMG-5/hSMG-7複合体とホスファターゼPP2AによるhUPF1の脱リン酸化が、NMDに必須の過程であることをも明らかとした。また、hUPF1などについて、一連の変異体を構築し、NMDに及ぼす効果やサーベイランス複合体の会合状態に与える影響などを詳細に解析し、hUPF1のリン酸化に続いてその脱リン酸化の過程でサーベイランス複合体の分子会合と再編が起きることを見いだした。これらと平行して、hSMG-1の精製と生化学的な性格付けの過程で、その結合分子(hSMG-1-BP)を同定し、その線虫ホモログが線虫で、実際にNMDの促進作用を有することを見いだした。これら、一連の成果により、多細胞生物におけるNMDのmRNAの識別を行うサーベイランスの分子的な実体がリン酸化と脱リン酸化を介したSMG分子群の集合と離散にあるという新たな概念を明確に提示することに成功した。

研究成果

• [文献書誌] Ohnishi, T., Ohno, S., et al.: "Phosphorylated hUPF1 selectively associates with hSMG5 and hSMG7 for PP2A-mediated dephosphorylation required for mammalian Nonsense-Mediated mRNA Decay"(投稿中).
  • 説明: 「研究成果報告書概要(和文)」より
• [文献書誌] Yamashita, A., Ohno, S., et al.: "Human SMG-1, a novel phosphatidylinositol 3-kinase-related protein kinase, associates with components of the mRNA surveillance complex and is involved in the regulation of nonsense-mediated mRNA decay"Genes & Development. 15. 2215-2228 (2001)
  • 説明: 「研究成果報告書概要(和文)」より
• [文献書誌] Ohnishi, T., Yamashita, A., Kashima, I., Schell, T., Anders, K. R. Grimson, A., Hachiya, T., Matthias W. Hentze, M. W., Anderson, P., and Ohno, S.: "Phosphorylated hUPF1 selectively associates with hSMG5 and hSMG7 for PP2A-mediated dephosphorylation required for mammalian Nonsense-Mediated mRNA Decay."submitted for publication.
  • 説明: 「研究成果報告書概要(欧文)」より
• [文献書誌] Yamashita, A., Ohnishi, T., Kashima, I., Taya, Y. and Ohno, S.: "Human SMG-1, a novel phosphatidylinositol 3-kinase-related protein kinas, associates with components of the mRNA surveillance complex and is involved in the regulation of nonsense-mediated mRNA decay."Genes & Development 15. 2215-2228 (2001)
  • 説明: 「研究成果報告書概要(欧文)」より
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• [文献書誌] Yamaji, S., Suzuki, A., Sugiyama, Y., Koide, Y., Yoshida, M., Kanamori.H., Mohri, H., Ohno, S., Ishigatsubo, Y.: "Anovel integrin-linked kinase-binding protein, affixin, is involved in the early stage of cell-substrate interaction"J. Cell. Biol.. 153. 1251-1264 (2001)
• [文献書誌] Takaki, Y., Hirai, S., Manabe, N., Izumi, Y., Hirose, T., Nakaya, M., Suzuki, A., Mizuno, K., Akimoto, K., Tsukita, S., Shuin, T., Ohno, S.: "Dynamic changes in protein components of the tight junction during liver regeneration"Cell Tissue Res.. 305. 399-409 (2001)
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