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Functional analysis of phosphorylation/dephosphorylation of cytoskeletal proteins in differentiation and growth of hematopoieic stem cell.

Research Project

Project/Area Number 07671181
Research Category

Grant-in-Aid for Scientific Research (C)

Allocation TypeSingle-year Grants
Section一般
Research Field Hematology
Research Institution1st Department of Internal Medicine, Faculy of Medicine University of Thokyo.

Principal Investigator

HIGASHIHARA Masaaki  Tokyo University Medicine lecturer, 医学部(病), 講師 (80165084)

Co-Investigator(Kenkyū-buntansha) INAGAKI Masaki  TokyoMetropolitan Institute of Gerontology Neurophysiology Chief, 神経生理部, 部門長
TSUJINO Shiho  TokyoUniversity Medicine medical staff, 医学部, 医員
AOKI Katsunori  TokyoUniversity Medicine medical staff, 医学部(病), 医員
MIYAZAKI Kouji  TokyoUniversity Medicine assistant, 医学部(病), 助手 (90261966)
YONEYAMA Akiko  TokyoUniversity Medicine lecturer, 医学部(病), 講師 (50175684)
Project Period (FY) 1995 – 1996
Project Status Completed (Fiscal Year 1996)
Budget Amount *help
¥2,200,000 (Direct Cost: ¥2,200,000)
Fiscal Year 1996: ¥1,000,000 (Direct Cost: ¥1,000,000)
Fiscal Year 1995: ¥1,200,000 (Direct Cost: ¥1,200,000)
KeywordsHematopoietic stem cells / Differentiation / Growth / Cytoskeletal protein / Myosin / Phosphorylation / Dephosphorylation / モノクロナル抗体 / myosin I / 白血病細胞 / 細胞分裂 / ビメンチン
Research Abstract

Among the four projects in these two years, only one project was successfully achieved. That is, we could clarify the functional domain of light meromyosin of smooth muscle myosin. Now we are going to determine the amino-acid sequences of antibody-recognizing sites. Monoclonal antibodies (McAbs) against light meromyosin (LMM) were produced and characteried. Anti-LMM antibodies, but not anti-S-1 or anti-S-2 antibody, almost completely inhibited filament formation in a equimolar concentration by the analyzes of turvidity and SDS-PAGE.McAbs against LMM showed no effects on Ca2+-ATPase and acto-ATPase activities of gizzard myoshi. These results suggest that LMM plays a crucial role in the formation of filament, but not ATPase activities.
Immunoelectroscopical analysis showed that the centroal region is more important for the filamentformation than c-terminal regions. By the analyzes with immunoblotting and ultracentrifugation, McAbs against thiophosphorylated myosin also detected unphsophorylated myosin. McAbs against phosphorylated vimentin produced by Dr.M.Inagaki showed several cytochemical findings : phosphorylated vimentins were detected in cytoplasm, showing diffuse patterns. To be of interest, in mitotic phase more intense signals were detected in perinuclear regions. These findings suggst that phosphorylated vimentin plays an important role in mitotic phase. We regret that the experiments of mutant gene of myosin light and heavy chain could not be performed, although we already received such genes form Dr.M.Ikebe (USA).

Report

(3 results)
  • 1996 Annual Research Report   Final Research Report Summary
  • 1995 Annual Research Report
  • Research Products

    (5 results)

All Other

All Publications (5 results)

  • [Publications] Tujino S, et al: "Primary structure of light and heavy chain variable regions of antioodies recognizaing phosphory lated vimentins" Biochem. Biophys. Res. Commun.219. 633-637 (1996)

    • Description
      「研究成果報告書概要(和文)」より
    • Related Report
      1996 Final Research Report Summary
  • [Publications] Tsujino S., Sekimata M., Inagaki N., Kamei Y., Higashihara M., Kurokawa K., Imajoh-Ohmi S., Inagaki M.: "Primary structure of light and heavy chain variable regions of antibodies recognizing phosphorylated vimentins" Biochem.Biophys.Res.Commun. 219. 633-637 (1996)

    • Description
      「研究成果報告書概要(欧文)」より
    • Related Report
      1996 Final Research Report Summary
  • [Publications] Higashihara M,et al.: "Inhibition of 20-KDa myosin light chain exchange by monoclonal antibodies against 17-KDa myosin light chain" FEBS Letters. 363. 57-60 (1995)

    • Related Report
      1996 Annual Research Report
  • [Publications] Tange T et al.: "Establishment and characterization of a new human mesothelioma cell line(T-85)" Pathul.International. 45. 791-800 (1995)

    • Related Report
      1996 Annual Research Report
  • [Publications] Horie R et al.: "A variant CD30 protein lacking extracellular and transmembrane domains in induced in HL60" BLood. 88. 2422-2432 (1996)

    • Related Report
      1996 Annual Research Report

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Published: 1995-04-01   Modified: 2016-04-21  

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