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1991 Fiscal Year Final Research Report Summary

Unification and reconstruction of myosin phosphorylation theory on contractile response of smooth muscle and nonmuscle cells.

Research Project

Project/Area Number 01044066
Research Category

Grant-in-Aid for international Scientific Research

Allocation TypeSingle-year Grants
SectionJoint Research
Research InstitutionNagoya University

Principal Investigator

HIDAKA Hiroyoshi  Nagoya University, Professor, 医学部, 教授 (80100171)

Co-Investigator(Kenkyū-buntansha) HARTSHORNE David J.  Arizona University, Professor, 生化学栄養食品化学部門, 教授
TOKUMITSU Hiroshi  Nagoya University, Assistant professor, 医学部, 助手 (20237077)
WATANABE Masato  Nagoya University, Assistant professor, 医学部, 助手 (30220924)
KOBAYASHI Ryoji  Nagoya University, Associate professor, 医学部, 助教授 (00020917)
Project Period (FY) 1989 – 1991
KeywordsMyosin light chain phosphorylation / Protein phosphorylation / Ca^<2+> binding protein (s) / Protein Kinase / KN-62
Research Abstract

Myosin phosphorylation-dephosphorylation is the primary Ca^2-mediated regulatory process in smooth muscle. However, recent physiological studies showed that the tension in intact smooth muscle fiber is maintained in spite of the dephosphorylation of myosin, and have suggested that other control mechanisms may exist which modulate the contractile state of the muscle. Can contraction be regulated by protein kinase (s) other than myosin light chain kinase (MLCK), and by Ca2^2-binding proteins other than calmodulin? In this international scientific research program, we have attempted to unify and reconstruct the myosin phosphorylation theory on contractile response of smooth muscle and non-muscle cells, and obtained the following results, according to the schedule.
1) We prepared monoclonal antibodies directed against chicken gizzard MLCK. One of the monoclonal antibody, MM-7 inhibited the kinase activity and the superprecipitation of bovine aortic smooth muscle actomyosin. We also demonstr … More ated the existence of it least 4 subspecies of MLCK in chicken tissues and the heterogeneity of tissue- and species-specific isozyme forms.
2) Caldesmon, an actin and calmodulin binding protein, was phosphorylated by PK-C and calmodulin-dependent protein kinase.
3) The calmodulin-dependent caldesmon kinase was an isozyme of the brain-rich calmodulin-dependent protein kinase II (CaM KII).
4) CaM KII phosphorylated purified myosin light chain at same sites, as MLCK did. Our original CaM KII specific inhibitor, KN-62 inhibited the various agonist-induced contraction in rabbit common carotid arterial strips. CaM KII may be involved in smooth muscle contraction.
5) We detected and purified three new Ca^<2+> binding proteins, using our original compounds affinity chromatography. One was calcyclin and the others were novel Ca^<2+>-binding proteins (tentatively designated calgizzarin and calvasculin). The presence of these Ca^<2+>binding proteins in smooth muscle cells show that novel intracellular Ca^<2+> messenger system (s) may exist. Less

  • Research Products

    (30 results)

All Other

All Publications (30 results)

  • [Publications] M.Hagiwara et al.: "Selective binding of L-Thyroxine by myosin light chain kinase." J.Biol.Chem.264. 40-44 (1989)

    • Description
      「研究成果報告書概要(和文)」より
  • [Publications] T.Ishikawa et al.: "Thyroid hormones directly interact with vascular smooth muscle strips." Mol.Pharmacol.35. 760-765 (1989)

    • Description
      「研究成果報告書概要(和文)」より
  • [Publications] S.Mamiya et al.: "Thyroid hormones inhibit platelet function and myosin light chain kinase." J.Biol.Chem.264. 8575-8579 (1989)

    • Description
      「研究成果報告書概要(和文)」より
  • [Publications] M.Hagiwara et al.: "Monoclonal antibody assessment of tissue- and species-specific myosin light chain kinase isozymes." J.Biochem.106. 71-75 (1989)

    • Description
      「研究成果報告書概要(和文)」より
  • [Publications] H.Tokumitsu et al.: "Anti-gizzard MLCK monoclonal antibody MM13 inhibits superprecipitation and phosphorylation of bovine aortic smooth muscle actomyosin." J.Biochem.106. 511-514 (1989)

    • Description
      「研究成果報告書概要(和文)」より
  • [Publications] J.C.Abougou et al.: "Phosphorylation of caldesmon." FEBS Lett.257. 408-410 (1989)

    • Description
      「研究成果報告書概要(和文)」より
  • [Publications] T.Tanaka et al.: "Calcium signaling of calcium-binding proteins and drug actions." Calcium Protein Signaling. 255. 165-171 (1989)

    • Description
      「研究成果報告書概要(和文)」より
  • [Publications] T.Tanaka et al.: "Phosphorylation of high-M_r caldesmon by protein kinase C modulates the regulatory function of this protein on the interaction between actin and myosin." Eur.J.Biochem.188. 495-500 (1990)

    • Description
      「研究成果報告書概要(和文)」より
  • [Publications] R.Kobayashi et al.: "Purification,characterization and partial sequence analysis of 32-kDa calcimedin from chicken gizzard." Arch.Biochem.Biophys.277. 203-210 (1990)

    • Description
      「研究成果報告書概要(和文)」より
  • [Publications] T.Ishikawa et al.: "Molecular pharmacology of calcium,calmodulin-dependent myosin phosphorylation in vascular smooth muscle." American J.Hypertension. 3. 231s-234s (1990)

    • Description
      「研究成果報告書概要(和文)」より
  • [Publications] H.Tokumitsu et al.: "A calcium-binding protein from rabbit lung cytosol identified as the product of growth-regulated gene(2A9)and its binding proteins." Arch.Biochem.Biophys.288. 202-207 (1991)

    • Description
      「研究成果報告書概要(和文)」より
  • [Publications] H.Todoroki et al.: "Purification,characterization and partial sequence analysis of a newly identified EF-hand type 13 kDa Ca^<2+>-binding protein from smooth muscle and nonmuscle tissues." J.Biol.Chem.266. 18668-18673 (1991)

    • Description
      「研究成果報告書概要(和文)」より
  • [Publications] Y.Watanabe et al.: "Isolation and characterization of a calcium-binding protein derived from mRNA termed p9Ka,pEL-98,18A2,or 42A by the newly synthesized vaso-relaxant,W-66 affinity chromatography." Arch.Biochem.Biophys.292. 563-569 (1992)

    • Description
      「研究成果報告書概要(和文)」より
  • [Publications] M.Watanabe et al.: "Molecular cloning and sequencing of a cDNA clone encoding a new calcium binding protein,named calgizzarin,from rabbit lung." Biochem.Biophys.Res.Commun.181. 644-649 (1991)

    • Description
      「研究成果報告書概要(和文)」より
  • [Publications] H.Tokumitsu,et al.: "A Calcyclin-associated protein is a newly identified member of the Ca^<2+>/phospholipid-binding proteins,annexin family." J.Biol.Chem.

    • Description
      「研究成果報告書概要(和文)」より
  • [Publications] M. Hagiwara et al.: "Selective binding of L-Thyroxine by myosin light chain kinase." J. Biol. Chem.264. 40-44 (1989)

    • Description
      「研究成果報告書概要(欧文)」より
  • [Publications] T. Ishikawa et al.: "Thyroid hormones directly interact with vascular smooth muscle strips." Mol. Pharmacol.35. 765-760 (1989)

    • Description
      「研究成果報告書概要(欧文)」より
  • [Publications] S. Mamiya et al.: "Thyroid hormones inhibit platelet function and myosin light chain kinase." J. Biol. Chem.264. 8575-8579 (1989)

    • Description
      「研究成果報告書概要(欧文)」より
  • [Publications] M. Hagiwara et al.: "Monoclonal antibody assessment of tissue-and species-specific myosin light chain kinase isozymes." J. Biochem.106. 71-75 (1989)

    • Description
      「研究成果報告書概要(欧文)」より
  • [Publications] H. Tokumitsu et al.: "Anti-gizzard MLCK monoclonal antibody MM13 inhibits superprecipitation and phosphorylation of bovine aortic smooth muscle actomyosin." J. Biochem.106. 511-514 (1989)

    • Description
      「研究成果報告書概要(欧文)」より
  • [Publications] J. C. Abougou et al.: "Phosphorylation of caldesmon." FEBS Lett.257. 408-410 (1989)

    • Description
      「研究成果報告書概要(欧文)」より
  • [Publications] T. Tanaka et al.: "Calcium signaling of calcium-binding proteins and drug actions." Calcium Protein Signaling. 255. 165-171 (1989)

    • Description
      「研究成果報告書概要(欧文)」より
  • [Publications] T. Tanaka et al.: "Phosphorylation of high-Mr caldesmon by protein kinase C modulates the regulatory function of this protein on the interaction between actin and myosin." Eur. J. Biochem.188. 495-500 (1990)

    • Description
      「研究成果報告書概要(欧文)」より
  • [Publications] R. Kobayashi et al.: "Purification, characterization and partial sequence analysis of 32-kDa calcimedin from chicken gizzard." Arch. Biochem. Biophys.277. 203-210 (1990)

    • Description
      「研究成果報告書概要(欧文)」より
  • [Publications] T. Ishikawa et al.: "Molecular pharmacology of calcium, calmodulin-dependent myosin phosphorylation in vascular smooth muscle." American J. Hypertension. 3. 231S-234S (1990)

    • Description
      「研究成果報告書概要(欧文)」より
  • [Publications] H. Tokumitsu et al.: "A calcium-binding protein from rabbit lung cytosol identified as the product of growth-regulated gene (2A9) and its binding proteins." Arch. Biochem. Biophys.288. 202-207 (1991)

    • Description
      「研究成果報告書概要(欧文)」より
  • [Publications] H. Todoroki et al.: "Purification, characterization and partial sequence analysis of a newly identified EF-hand type 13 kDa Ca^<2+>-binding protein from smooth muscle and nonmuscle tissues." J. Biol. Chem.266. 18668-18673 (1991)

    • Description
      「研究成果報告書概要(欧文)」より
  • [Publications] Y. Watanabe et al.: "Isolation and characterization of a calcium-binding protein derived from mRNA termed p9Ka, pEL-98, 18A2, or 42A by the newly synthesized vaso-relaxant, W-66, affinity chromatography." Arch. Biochem. Biophys.292. 563-569 (1992)

    • Description
      「研究成果報告書概要(欧文)」より
  • [Publications] M. Watanabe et al.: "Molecular cloning and sequencing of a cDNA clone encoding a new calcium binding protein, named calgizzarin, from rabbit lung." Biochem. Biophys. Res. Commun.181. 644-649 (1991)

    • Description
      「研究成果報告書概要(欧文)」より
  • [Publications] H. Tokumitsu, et al.: "A Calcyclin-associated protein is a newly identified member of the Ca^<2+>/phospholipid-binding proteins, annexin family." J. Biol. Chem.

    • Description
      「研究成果報告書概要(欧文)」より

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Published: 1993-03-16  

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