| Project/Area Number |
59440094
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| Research Category |
Grant-in-Aid for General Scientific Research (A)
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| Allocation Type | Single-year Grants |
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| Research Field |
分子遺伝学・分子生理学
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| Research Institution | Nagoya University |
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Principal Investigator |
ASAKURA Sho Institute of Molecular Biology, Faculty of Science, Nagoya University, 理学部, 教授 (80022531)
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| Co-Investigator(Kenkyū-buntansha) |
KAMIYA Ritsu Institute of Molecular Biology, Faculty of Science, Nagoya University, 理学部, 助手 (10124314)
OWARIBE Katsushi Institute of Molecular Biology, Faculty of Science, Nagoya University, 理学部, 助手 (90109257)
HIGASHI-FUJIME Sugie Institute of Molecular Biology, Faculty of Science, Nagoya University, 理学部, 助手 (60022662)
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| Project Period (FY) |
1984 – 1986
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| Project Status |
Completed (Fiscal Year 1986)
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| Budget Amount *help |
¥16,500,000 (Direct Cost: ¥16,500,000)
Fiscal Year 1986: ¥1,500,000 (Direct Cost: ¥1,500,000)
Fiscal Year 1985: ¥4,000,000 (Direct Cost: ¥4,000,000)
Fiscal Year 1984: ¥11,000,000 (Direct Cost: ¥11,000,000)
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| Keywords | Actin / Myosin / In vitro motility / Dark-field microscopy / Fluorescence microscopy / Fluorescence label / カルシウム / アクチン繊維 / ミオシン繊維 / アクチン・ミオシン間の滑り運動 / 滑り運動の再構成 |
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| Research Abstract |
To clarify the molecular mechanism of cell motility, we have been trying to construct model systems that show motility in vitro, using purified proteins such as actin and myosin. In one type of experiment, single myosin filaments were visualized with a dark-field microscope with a high-powered illumination. In the presence of Mg-ATP, myosin filaments moved along actin filaments at a maximal speed of 5 um/sec. There appeared to be no obvious correlation between the speed and the length of a filament. In another type of experiment, actin was labeled with a fluorescent dye, fluorescein isothiocyanate (FITC), and observed with a fluorescence microscope, a sensitive video camera, and a digital image processor. The FITC-actin filaments displayed only a vigorous flexural movement in the presence of myosin and Mg-ATP. However, when the concentration of actin was increased by adding unlabeled actin filaments, the FITC-flaments became to move in the direction of their lengths. This movement was uni-directional and temperature-dependent; at 27 C, the speed was about 5 um/s. When FITC-actin was complexed with tropomyosin and troponin, the in vitro movement of this kind became steeply dependent on the calcium concentration in the medium; directional movement was observed at calcium concentrations higher than <10^(-5.6)> M, but no movement was observed in lower than <10^(-5.8)> M calcium ion. This indicates that the Ca-regulation of the actin/myosin motile systems in vitro is a cooperative process. Other experiments have demonstrated that a similar directional movement of FITC-actin filaments could be caused by heavy meromyosin, which lacked the tail portion of myosin and existed in a dispersed state. This finding indicates that the sliding movement between myosin and actin filaments does not require assembly of myosin. It will be an interesting future problem to examine whether a single-headed myosin also can cause actin filaments to move directionally.
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