1994 Fiscal Year Final Research Report Summary
Cell-Cycle Regulation of Microtubule Formation
Project/Area Number |
05680628
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Research Category |
Grant-in-Aid for General Scientific Research (C)
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Allocation Type | Single-year Grants |
Research Field |
Cell biology
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Research Institution | The Institute of Physical and Chemical Research (RIKEN) |
Principal Investigator |
MASUDA Hirohisa The Institute of physical and Chemical Research (RIKEN), BioDesign Research Group, Special Researcher, バイオデザイン研究グループ, 研究員 (30260219)
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Project Period (FY) |
1993 – 1994
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Keywords | Microtubule / Cell cycle / Mitosis / Gamma-tubulin / Schizosaccharo-myces Pombe / Spindle pole body |
Research Abstract |
The ability of the Schizosacchromyces pombe spindle pole body to nucleate microtubules is activated at the onset of mitosis for mitotic spindle formation, but it is inactivated during interphase. We have developed an in vitro assay for studying the molecular mechanism of spindle pole body activation using permeabilized interphase S.pombe cells and Xenopus mitotic extracts. We have shown that interphase spindle pole bodies are activated indirectly by cdc2 kinase in Xenopus mitotic extracts. In this study we examined a role of gamma-tubulin, a component of both interphase and mitotic spindle pole body, in formation of the microtubule nucleating complex at the mitotic spindle pole body. A polyclonal antibody specific to S.pombe gamma-tubulin inhibited microtubule nucleation from the activated spindle pole bodies. Addition of bacterially expressed S.pombe gamma-tubulin or its amino-terminal fragments to Xenopus mitotic extracts inhibited activation of spindle pole body. Using affinity chromatography of Xenopus mitotic extracts with the amino-terminal fragment, both fraction bound and unbound to the fragment were found required for the activation. The activation of spindle pole body was not due to recruitment of Xenopus gamma-tubulin to the spindle pole body, since partially fractionated Xenopus extracts containing no gamma-tubulin supported the activation. These results suggest that S.pombe gamma-tubulin is a component of the microtubule nucleating complex, and that mitosis-specific modification of or binding of a protein to the amino-terminal region of gamma-tubulin is necessary, but not sufficient for the activation of spindle pole body.
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Research Products
(10 results)