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生物時計分子の核内移行と概日リズム

Research Project

Project/Area Number 11237205
Research Category

Grant-in-Aid for Scientific Research on Priority Areas

Allocation TypeSingle-year Grants
Review Section Biological Sciences
Research InstitutionOsaka Bioscience Institute (2001-2003)
Kobe University (1999-2000)

Principal Investigator

内匠 透 (内近 透)  (財)大阪バイオサイエンス研究所, 神経科学部門, 研究室長 (00222092)

Project Period (FY) 1999 – 2003
Project Status Completed (Fiscal Year 2003)
Budget Amount *help
¥64,400,000 (Direct Cost: ¥64,400,000)
Fiscal Year 2003: ¥12,800,000 (Direct Cost: ¥12,800,000)
Fiscal Year 2002: ¥12,800,000 (Direct Cost: ¥12,800,000)
Fiscal Year 2001: ¥12,300,000 (Direct Cost: ¥12,300,000)
Fiscal Year 2000: ¥12,900,000 (Direct Cost: ¥12,900,000)
Fiscal Year 1999: ¥13,600,000 (Direct Cost: ¥13,600,000)
Keywords時計蛋白 / 概日リズム / 核移行 / 遺伝子 / 神経科学 / 生理学 / 脳・神経 / 時計分子 / 時計遺伝子 / 視交又上核
Research Abstract

培養細胞を用いた実験等から、CRYとPERとの核移行が、転写のフィードバック機構の本体を調節し、時間の決定機構となるという仮説をたて、時計分子の核移行の詳細な分子機構を検討した。CRY2には典型的な核移行シグナルNLSがカルボキシル端側に存在するが、CRY1には定型的NLSが存在しない。CRY1,CRY2をタグ付きGST融合蛋白として大腸菌にて発現、精製した蛋白をMH3T3培養細胞に注入することによりその核移行を検討した。CRY1,CRY2蛋白の細胞質内注入後、両者とも核内への移行がみられたが、WGA, Ran-GTP, G19V Ran-GTP等との共注入実験を行った所、同じく両者ともRan依存性に核移行することが明らかになった。さらに、CRY1,CRY2をそれぞれN末側C末側に二分割したCRY1N, CRY1C, CRY2N, CRY2Cを、同様に大腸菌を用いて蛋白を発現、精製し、細胞注入した結果、いずれもN末端側蛋白(CRY1N, CRY2N)は核移行を示さず、C端側(CRY1C, CRY2C)は核移行を観察した。セミインタクト細胞を用いたin vitro系では、CRY1C, CRY2Cともにimportinαβを介する核移行を行っていることが示された。NLS配列をもつCRY2Cの核移行はin vitro結合実験の結果からもimportinαβを介する核移行であることを明らかにした。さらに、CRY2のNLSの変異体では核移行が阻害されたことから、CRY2の核移行に必須であることを明らかにした。

Report

(5 results)
  • 2003 Annual Research Report
  • 2002 Annual Research Report
  • 2001 Annual Research Report
  • 2000 Annual Research Report
  • 1999 Annual Research Report
  • Research Products

    (19 results)

All Other

All Publications (19 results)

  • [Publications] Y.Kajimoto, O.Shirakawa, X.-H.Lin, T.Hashimoto, N.Kitamura, N.Murakami, T.Takumi, K.Maeda: "Synapse-associated protein 90/postsynaptic density-95-asssociated protein (SAPAP) is expressed differentially in phencyclidine-treated rats and is increased in the nucleus accumbens of patients with schizophrenia."Neuropsychopharamacology. 8. 1831-1839 (2003)

    • Related Report
      2003 Annual Research Report
  • [Publications] Y.Shigeyoshi, E.Meyer-Bernstein, K.Yagita, W.Fu, Y.Chen, T.Takumi, P.Schotland, A.Sehgal, H.Okamura: "Restoration of circadian behavioural rhythms in a period null Drosophila mutant(per01) by mammalian period homologues mPer1 and mPer2"Genes Cells. 7・2. 163-171 (2002)

    • Related Report
      2002 Annual Research Report
  • [Publications] S.Bouret, V.Prevot, T.Takumi, JC.Beauvillain, V.Mitchell: "Regulation by gonadal steroids of the mRNA encoding for a type I receptor for TGF-beta in the female rat hypothalamus"Neuroendocrinology. 76・1. 1-7 (2002)

    • Related Report
      2002 Annual Research Report
  • [Publications] T.Takumi: "Rapid cDNA cloning by PCR screening(RC-PCR)"Methods Mol Biol. 192. 385-389 (2002)

    • Related Report
      2002 Annual Research Report
  • [Publications] H.Ohyama, H.Kajita, K.Omori, T.Takumi, N.Hiramoto, T.Iwasaka, H.Matsuda: "Inhibition of cardiac delayed rectifier K^+ currents by an antisense oligodeoxynucleotide against lsk(MinK) and over-expression of lsk mutant D77N in neonatal mouse hearts"Pflugers Arch.. 442. 329-335 (2001)

    • Related Report
      2001 Annual Research Report
  • [Publications] S.Bouret, M.T.V.Chuoi-Mariot, V.Prevot, D.Croix, T.Takumi, S.Jegou, H.Vaudry, J.-C.Beauvilain, V.Mitchell: "Evidence that TGF-b may directly modulate POMC mRNA expression in the female rat arcuate nucleus"Endocrinology. 142. 4055-4065 (2001)

    • Related Report
      2001 Annual Research Report
  • [Publications] Y.Shigeyoshi, W.Fu, Y.Chen, K.Yagita, T.Takumi, P.Schotland, A.Sehgal, H.Okamura: "Restoration of circadian behavioral rhythms in a period null Drosophila mutant(per0) by mammalian Period homologues mPer1 and mPer2"Genes Cells. (in press).

    • Related Report
      2001 Annual Research Report
  • [Publications] T.Takumi: "A rapid cDNA cloning in the post-genome era"Methods Mol.Biol.. (in press).

    • Related Report
      2001 Annual Research Report
  • [Publications] Prevot,V. et al.: "Evidence that members of the TGFβ superfamily play a role in regulation of the GnRH neuroendocrine Axis"J.Nueroendocrinol.. 12. 665-670 (2000)

    • Related Report
      2000 Annual Research Report
  • [Publications] Ohyama,H. et al: "Inhibition of cardiac delayed rectifier K^+ currents by an antisense oligodesxynucleotide against 1sk and overexpression of 1sk mutant"Eur.J.Physiol.. (in press).

    • Related Report
      2000 Annual Research Report
  • [Publications] Takumi,T.: "Rapid cDNA cloning by PCR screening"Methods Mol.Biol.. (in press).

    • Related Report
      2000 Annual Research Report
  • [Publications] Takumi T. et al.: "A mammalian ortholog of Drosophila timeless, highly expressed in SCN and retina, forms a complex with mPER1"Genes to Cells. 4. 67-75 (1999)

    • Related Report
      1999 Annual Research Report
  • [Publications] Maebayashi Y. et al.: "A putative transcription Factor with seven zinc-finger motifs identified in the developing suprachiasmatic nucleus by the differential display PCR method"The Journal of Neuroscience. 19・22. 10176-10183 (1999)

    • Related Report
      1999 Annual Research Report
  • [Publications] Prevot V. et al.: "Evidence that members of the TGF-β superfamily play a Keyrole in the regulation of the GnRH neuroendocrine axis: expression of a type1 serine-threonine kinase receptor"Journal of Neuroendocrinology. (in press). (2000)

    • Related Report
      1999 Annual Research Report
  • [Publications] 内匠透 他: "転写因子による生物時計の制御"実験医学. 17・3. 198-204 (1999)

    • Related Report
      1999 Annual Research Report
  • [Publications] 内匠透 他: "生物時計の分子機構"神緑会学術雑誌. 15. 93-96 (1999)

    • Related Report
      1999 Annual Research Report
  • [Publications] 岡村均 他: "哺乳類時計遺伝子 mPer"Molecular medicine. 36. 315-324 (1999)

    • Related Report
      1999 Annual Research Report
  • [Publications] 久野宗 他: "脳を知る"秀潤社. 205 (1999)

    • Related Report
      1999 Annual Research Report
  • [Publications] 村松正實 他: "新遺伝子工学ハンドブック"羊工社. 329 (1999)

    • Related Report
      1999 Annual Research Report

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Published: 1999-04-01   Modified: 2018-03-28  

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