1996 Fiscal Year Final Research Report Summary
Functional analysis of phosphorylation/dephosphorylation of cytoskeletal proteins in differentiation and growth of hematopoieic stem cell.
| Project/Area Number |
07671181
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| Research Category |
Grant-in-Aid for Scientific Research (C)
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| Allocation Type | Single-year Grants |
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| Section | 一般 |
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| Research Field |
Hematology
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| Research Institution | 1st Department of Internal Medicine, Faculy of Medicine University of Thokyo. |
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Principal Investigator |
HIGASHIHARA Masaaki Tokyo University Medicine lecturer, 医学部(病), 講師 (80165084)
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| 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
YONEYAMA Akiko TokyoUniversity Medicine lecturer, 医学部(病), 講師 (50175684)
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| Project Period (FY) |
1995 – 1996
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| Keywords | Hematopoietic stem cells / Differentiation / Growth / Cytoskeletal protein / Myosin / Phosphorylation / Dephosphorylation |
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| 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).
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