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Activation of cytosolic protein-tyrosine kinase and analysis of its substrate proteins

Research Project

Project/Area Number 03454158
Research Category

Grant-in-Aid for General Scientific Research (B)

Allocation TypeSingle-year Grants
Research Field Pathological medical chemistry
Research InstitutionFukui Medical School

Principal Investigator

YAMAMURA Hirohei  Fukui Medical School Professor, 医学部, 教授 (90030882)

Co-Investigator(Kenkyū-buntansha) TANIGUCHI Takanobu  Fukui Medical School Research Associate, 医学部, 助手 (60217130)
中村 俊一  福井医科大学, 医学部, 助手 (40155833)
Project Period (FY) 1991 – 1992
Project Status Completed (Fiscal Year 1992)
Budget Amount *help
¥7,000,000 (Direct Cost: ¥7,000,000)
Fiscal Year 1992: ¥2,400,000 (Direct Cost: ¥2,400,000)
Fiscal Year 1991: ¥4,600,000 (Direct Cost: ¥4,600,000)
KeywordsP72syk / Protein-tyrosine kinase / Platelets / Calmodulin / Ca^<2+> / Thrombin / Myosin light chain kinase / Wheat germ agglutinin / Ca^<2+> / 蛋白質チロシン残基燐酸化酵素 / 細胞質チロシンキナ-ゼ / 蛋白質分解酵素 / 40Kチロシンキナ-ゼ / 自己燐酸化 / ミエリン塩基性蛋白質
Research Abstract

We have reported a porcine gene syk encoding a non-receptor type 72-kDa PTK (p72^<syk>) which has a distinct structural character, that is the presence of the second src homology region 2 (SH2) instead of SH3, from other members of this group of PTK. The expression of p72^<syk> has been detected in porcine splenocytes, platelets, peripheral blood lymphocytes, polymorphonuclear leukocytes and tonsilocytes on immunoblot. In the cases of splenocytes and platelets, the activation of p72^<syk> was seen by the stimulation of wheat germ agglutinin. When washed porcine platelets was stimulated with thrombin and the activation of p72^<syk> was assessed in immunoprecipitation kinase assay, employing anti-CPTK40 antibodies, the activity of p72^<syk> increased within 5s, reached maximum at 10s and decreased to basal level within 60 s after 0.5 U/ml thrombin stimulation. This activation was enhanced dose dependently and was completely cancelled by the pretreatment of platelet suspension with hirudin, a specific antagonist of thrombin. In the Ca^<2+>-depleted condition both extra- and intracellularly, the activation of p72^<syk> was not affected at all but the deactivation was completely abrogated even at 120s after thrombin stimulation. In addition, the replenishment of Ca^<2+> resulted in a similar deactivation pattern as seen in Ca^<2+>-rich condition. Furthermore, this deactivation was also cancelled by the pretreatment of platelets with W7, a calmodulin antagonist, as well as ML9, a myosin-light-chain kinase inhibitor. These results have suggested that p72^<syk> is rapidly activated following thrombin stimulation with a peak at 10s and is negatively regulated mainly through a calmodulin-dependent protein kinase, myosin- light-chain kinase, in thrombin-stimulated platelets.

Report

(3 results)
  • 1992 Annual Research Report   Final Research Report Summary
  • 1991 Annual Research Report
  • Research Products

    (47 results)

All Other

All Publications (47 results)

  • [Publications] Hashimoto,E.: "Studies on protein kinase C tightly-bound to rat liver plasma membrane and its protease-avtivated form." Int.J.Biochem.23(4). 395-403 (1991)

    • Description
      「研究成果報告書概要(和文)」より
    • Related Report
      1992 Final Research Report Summary
  • [Publications] Hashimoto,E.: "Protease-activated protein kinase C in rat liver." Int.J.Biochem.23(5,6). 507-512 (1991)

    • Description
      「研究成果報告書概要(和文)」より
    • Related Report
      1992 Final Research Report Summary
  • [Publications] Shimomura,R.: "Possible endogenous substrate proteins for cytosolic protein-tyrosine kinase from porcine spleen." Int.J.Biochem.23(7,8). 72-731 (1991)

    • Description
      「研究成果報告書概要(和文)」より
    • Related Report
      1992 Final Research Report Summary
  • [Publications] Takayama,H.: "Ionophore A23187-induced protein-tyrosine phosphorylation of human platelets:Possible synergism between Ca^<2+> mobilization and protein kinase C activation." Biochem.Biophys.Res.Commun.174(2). 992-997 (1991)

    • Description
      「研究成果報告書概要(和文)」より
    • Related Report
      1992 Final Research Report Summary
  • [Publications] Inazu,T.: "The lectin wheat germ agglutinin induces rapid protein-tyrosine phosphorylation in human platelets." Biochem.Biohys.Res.Commun.174(3). 1154-1158 (1991)

    • Description
      「研究成果報告書概要(和文)」より
    • Related Report
      1992 Final Research Report Summary
  • [Publications] Taniguchi,T.: "Molecular cloning of a porcine gene syk that encodes a 72-kDa protein-tyrosine kinase showing high susceptibility to proteolysis." J.Biol.Chem.266(24). 15790-15796 (1991)

    • Description
      「研究成果報告書概要(和文)」より
    • Related Report
      1992 Final Research Report Summary
  • [Publications] Yamada,T.: "The lectin wheat germ agglutinin stimulates a protein-tyrosine kinase activity of p72^<syk> in porcine splenocytes." Biochem.Biophys.Res.Commun.180(3). 1325-1329 (1991)

    • Description
      「研究成果報告書概要(和文)」より
    • Related Report
      1992 Final Research Report Summary
  • [Publications] Sakai,K.: "Identification of the phosphorylation sites of H2B histone by a catalytic fragment of p72^<syk> from porcine spleen." FEBS Letters. 294(1,2). 104-108 (1991)

    • Description
      「研究成果報告書概要(和文)」より
    • Related Report
      1992 Final Research Report Summary
  • [Publications] Takeuchi,F.: "Phosphorylation of histone H2A by protein kinase C and Identification of the phosphorylation site." J.Biochem.111(6). 788-792 (1992)

    • Description
      「研究成果報告書概要(和文)」より
    • Related Report
      1992 Final Research Report Summary
  • [Publications] Ohta,S.: "Protein-tyrosine phosphorylations induced by concanavalin A and N-formyl-methionyl-leucyl-phenylalanine in human neutrophils." Eur.J.Biochem.206. 895-900 (1992)

    • Description
      「研究成果報告書概要(和文)」より
    • Related Report
      1992 Final Research Report Summary
  • [Publications] Ohta,S.: "Protein-tyrosine kinase p72syk is activated by wheat germ agglutinin in platelets." Biochem.Biophys.Res.Commun.185(3). 1128-1132 (1992)

    • Description
      「研究成果報告書概要(和文)」より
    • Related Report
      1992 Final Research Report Summary
  • [Publications] Asahi,M.: "Thrombin-induced human platelet aggregation if inhibited by protein-tyrosine kinase inhibitors,ST638 and genistein." FEBS lett.309(1). 10-14 (1992)

    • Description
      「研究成果報告書概要(和文)」より
    • Related Report
      1992 Final Research Report Summary
  • [Publications] Takeuchi,N.: "Inhibitory effect of mitoxantrone on activity of protein kinase C and growth of HL60 cells." J.Biochem.112. 762-767 (1992)

    • Description
      「研究成果報告書概要(和文)」より
    • Related Report
      1992 Final Research Report Summary
  • [Publications] Taniguchi,T.: "Protein-tyrosine kinase p72syk is activated by thrombin and is negatively regulated through Ca^<2+> Mobilization in platelets." J.Biol.Chem.

    • Description
      「研究成果報告書概要(和文)」より
    • Related Report
      1992 Final Research Report Summary
  • [Publications] Hashimoto,E.: "Studies on protein kinase C tightly-bound to rat liver plasma membrane and its protease-activated form." Int.J.Biochem.23(4). 395-403 (1991)

    • Description
      「研究成果報告書概要(欧文)」より
    • Related Report
      1992 Final Research Report Summary
  • [Publications] Hashimoto,E.: "Protease-activated protein kinase C in rat liver." Int.J.Biochem.23(5,6). 507-512 (1991)

    • Description
      「研究成果報告書概要(欧文)」より
    • Related Report
      1992 Final Research Report Summary
  • [Publications] Shimomura,R.: "Possible endogenous substrate proteins for cytosolic protein-tyrosine kinase from porcine spleen." Int.J.Biochem.23(7,8). 727-731 (1991)

    • Description
      「研究成果報告書概要(欧文)」より
    • Related Report
      1992 Final Research Report Summary
  • [Publications] Takayama,H.: "Ionophore A23187-induced protein-tyrosine phosphorylation of human platelets: Possible synergism between Ca^<2+> mobilization and protein kinase C activation." Biochem.Biophys.Res.Commun.174(2). 992-997 (1991)

    • Description
      「研究成果報告書概要(欧文)」より
    • Related Report
      1992 Final Research Report Summary
  • [Publications] Inazu,T.: "The lectin wheat germ agglutinin induces rapid protein-tyrosine phosphorylation in human platelets." Biochem.Biophys.Res.Commun.174(3). 1154-1158 (1991)

    • Description
      「研究成果報告書概要(欧文)」より
    • Related Report
      1992 Final Research Report Summary
  • [Publications] Taniguchi,T.: "Molecular cloning of a porcine gene syk that encodes a 72-kDa protein-tyrosine kinase showing high susceptibility to proteolysis." J.Biol.Chem.266(24). 15790-15796 (1991)

    • Description
      「研究成果報告書概要(欧文)」より
    • Related Report
      1992 Final Research Report Summary
  • [Publications] Yamada,T.: "The lectin wheat germ agglutinin stimulates a protein-tyrosine kinase activity of p72^<syk> in porcine splenocytes." Biochem.Biophys.Res.Commun.180(3). 1325-1329 (1991)

    • Description
      「研究成果報告書概要(欧文)」より
    • Related Report
      1992 Final Research Report Summary
  • [Publications] Sakai,K.: "Identification of the phosphorylation sites of H2B histone by a catalytic fragment of p72^<syk> from porcine spleen." FEBS Lett.294(1,2). 104-108 (1991)

    • Description
      「研究成果報告書概要(欧文)」より
    • Related Report
      1992 Final Research Report Summary
  • [Publications] Takeuchi,F.: "Phosphorylation of histone H2A by protein kinase C and Identification of the phosphorylation site." J.Biochem.111(6). 788-792 (1992)

    • Description
      「研究成果報告書概要(欧文)」より
    • Related Report
      1992 Final Research Report Summary
  • [Publications] Ohta,S.: "Protein-tyrosine phosphorylations induced by concanavalin A and N-formyl-methionyl-leucyl-phenylalanine in human neutrophils." Eur.J.Biochem.206. 895-900 (1992)

    • Description
      「研究成果報告書概要(欧文)」より
    • Related Report
      1992 Final Research Report Summary
  • [Publications] Ohta,S.: "Protein-tyrosine kinase p72syk is activated by wheat germ agglutinin in platelets." Biochem.Biophys.Res.Commun.185(3). 1128-1132 (1992)

    • Description
      「研究成果報告書概要(欧文)」より
    • Related Report
      1992 Final Research Report Summary
  • [Publications] Asahi,M.: "Thrombin-induced human platelet aggregation is inhibited by protein-tyrosine kinase inhibitors, ST638 and genistein." FEBS lett.309(1). 10-14 (1992)

    • Description
      「研究成果報告書概要(欧文)」より
    • Related Report
      1992 Final Research Report Summary
  • [Publications] Takeuchi,N.: "Inhibitory effect of mitoxantrone on activity of protein kinase C and growth of HL60 cells." J.Biochem.112(6). 762-767 (1992)

    • Description
      「研究成果報告書概要(欧文)」より
    • Related Report
      1992 Final Research Report Summary
  • [Publications] Taniguchi,T.: "Protein-tyrosine kinase p72syk is activated by thrombin and is negatively regulated through Ca^<2+> Mobilization in platelets." J.Biol.Chem.

    • Description
      「研究成果報告書概要(欧文)」より
    • Related Report
      1992 Final Research Report Summary
  • [Publications] Takeuchi,F.: "Phosphorylation of histone H2A by protein kinase C and Identification of the phosphorylation site." J.Biochem.111(6). 788-792 (1992)

    • Related Report
      1992 Annual Research Report
  • [Publications] Ohta,S.: "Protein‐tyrosine phosphorylations induced by concanavalin A and N‐formyl‐methionyl‐leucyl‐phenylalanine in human neutrophils." Eur.J.Biochem.206. 895-900 (1992)

    • Related Report
      1992 Annual Research Report
  • [Publications] Ohta,S.: "Protein‐tyrosine kinase p72syk is activated by wheat germ agglutinin in platelets." Biochem.Biophys.Res.Commun.185(3). 1128-1132 (1992)

    • Related Report
      1992 Annual Research Report
  • [Publications] Asahi,M.: "Thrombin‐induced human platelet aggregation is inhibited by protein‐tyrosine kinase inhibitors,ST638 and genistein." FEBS lett.309(1). 10-14 (1992)

    • Related Report
      1992 Annual Research Report
  • [Publications] Takeuchi,N.: "Inhibitory effect of mitoxantrone on activity of protein kinase C and growth of HL60 cells." J.Biochem.112. 762-767 (1992)

    • Related Report
      1992 Annual Research Report
  • [Publications] Taniguchi,T.: "Protein‐tyrosine kinase p72syk is activated by thrombin and is negatively regulated through Ca^<2+> Mobilization in platelets." J.Biol.Chem.

    • Related Report
      1992 Annual Research Report
  • [Publications] Eikichi Hashimoto: "Studies on protein kinase C tightlyーbound to rat liver plasma membrane and its proteaseーactivated form." Int.J.Biochem.23(4). 395-403 (1991)

    • Related Report
      1991 Annual Research Report
  • [Publications] Eikichi Hashimoto: "Proteaseーactivated protein kinase C in rat liver." Int.J.Biochem.23(5,6). 507-512 (1991)

    • Related Report
      1991 Annual Research Report
  • [Publications] Reiko Shimomura: "Possible endogenous substrate proteins for cytosolic proteinーtyrosine kinase from porcine spleen." Int.J.Biochem.23(7,8). 727-731 (1991)

    • Related Report
      1991 Annual Research Report
  • [Publications] Hiroshi Takayama: "Ionophore A23187ーinduced proteinーtyrosine phosphorylation of human platelets:Possible synergism between Ca^<2+> mobilization and protein kinase C activation." Biochem.Biophys.Res.Commun.174(2). 992-997 (1991)

    • Related Report
      1991 Annual Research Report
  • [Publications] Tetsuya Inazu: "The lectin wheat germ agglutinin induces rapid proteinーtyrosine phosphorylation in human platelets." Biochem.Biophys.Res.Commun.174(3). 1154-1158 (1991)

    • Related Report
      1991 Annual Research Report
  • [Publications] Takanobu Taniguchi: "Molecular cloning of a porcine gene syk that encodes a 72ーkDa proteinーtyrosine kinase showing high susceptibility to proteolysis." J.Biol.chem.266(24). 15790-15796 (1991)

    • Related Report
      1991 Annual Research Report
  • [Publications] Takechiyo Yamada: "The lectin wheat germ agglutinin stimulates a proteinーtyrosine kinase activity of p72^<syk> in porcine splenocytes." Biochem.Biophys.Res.Commun.180(3). 1325-1329 (1991)

    • Related Report
      1991 Annual Research Report
  • [Publications] Keiko Sakai: "Identification of the phosphorylation sites of H2B histone by a catalytic fragment of p72^<syk> from porcine spleen." FEBS Letters. 294(1,2). 104-108 (1991)

    • Related Report
      1991 Annual Research Report
  • [Publications] Fumito Takeuchi: "Phosphorylation of Histone H2A by protein kinase C and Identification of the Phosphorylation Site." J.Biochem.

    • Related Report
      1991 Annual Research Report
  • [Publications] 山村 博平: "病理学キ-ワ-ド" 文光堂, 1 (1991)

    • Related Report
      1991 Annual Research Report
  • [Publications] 山村 博平: "病理学キ-ワ-ド" 文光堂, 1 (1991)

    • Related Report
      1991 Annual Research Report
  • [Publications] 稲津 哲也: "代謝" 中山書店, 5 (1991)

    • Related Report
      1991 Annual Research Report
  • [Publications] 谷口 隆信: "注目の臨床実験検査法" 中山書店, 9 (1991)

    • Related Report
      1991 Annual Research Report

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Published: 1991-04-01   Modified: 2016-04-21  

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