| Project/Area Number |
10440225
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| Research Category |
Grant-in-Aid for Scientific Research (B).
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| Allocation Type | Single-year Grants |
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| Section | 一般 |
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| Research Field |
遺伝
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| Research Institution | The University of Tokyo |
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Principal Investigator |
TOH-E Akio The University of Tokyo, Department of Biological Sciences, Professor, 大学院・理学系研究科, 教授 (90029249)
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| Co-Investigator(Kenkyū-buntansha) |
UESONO Yukifumi The University of Tokyo, Department of Biological Sciences, Assistant Professor, 大学院・理学系研究科, 助手 (30251408)
KIKUCHI Yoshiko The University of Tokyo, Department of Biological Sciences, Associated, 大学院・理学系研究科, 助教授 (00138124)
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| Project Period (FY) |
1998 – 2000
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| Project Status |
Completed (Fiscal Year 2000)
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| Budget Amount *help |
¥13,400,000 (Direct Cost: ¥13,400,000)
Fiscal Year 2000: ¥1,700,000 (Direct Cost: ¥1,700,000)
Fiscal Year 1999: ¥3,600,000 (Direct Cost: ¥3,600,000)
Fiscal Year 1998: ¥8,100,000 (Direct Cost: ¥8,100,000)
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| Keywords | 26S proteasome / cell cycle / mitosis / metaphase-anaphase transition / RPN9 / Saccharomyces cerevisiae / two-hybrid screening / プロテアソーム / 蛋白質分解 / Rpn9 / Rpn10 / 蛋白質の集合 / M期中期アレスト / ユビキチン化 |
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| Research Abstract |
We have isolated the RPN9 gene by two hybrid screening using RPN10 (formerly SUN1) as bait, which encodes a multiubiquitin chain receptor residing in the 26S proteasome. A yeast strain carrying a disrupted allele of RPN9 was found to be temperature sensitive for its growth. At the restrictive temperature, the Δrpn9 strain accumulated multiubiquitinated proteins. When the proteasome fractions separated by the glycerol gradient centrifugation were analysed by native PAGE, we found that the 26S proteasome of the Δrpn9 cells was shifted to lighter fractions than expected and that there was a larger amount of the incomplete proteasome complexes in the fractions. Furthermore, Rpn10 was not detected in the 26S proteasome of Δrpn9 cells. These results indicate that Rpn9 is needed for incorporating Rpn10 into the 26S proteasome and that Rpn9 participates in assembly and/or stability of the 26S proteasome.
Δrpn9 cells were found to be arrested at metaphase ; dumbbell shaped, single nucleus at the isthmuth, unseparated sister chromatids. When a Δrpn9 Δpds1 strain was shifted to a restrictive temperature, the cells stopped growing. No anaphase cells were observed, but sister chrmomatids were separated. Judging from these phenotypes, we anticipate the presence of some proteins which should be degraded by the 26S proteasome during anaphase progression.
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