| Project/Area Number |
13680786
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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 |
Cell biology
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| Research Institution | NARA INSTITUTE OF SCIENCE AND TECHNOLOGY |
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Principal Investigator |
OBUSE Chikasi Nara Institute of Science and Technology, Biological Science, Assistant Professor, バイオサイエンス研究科, 助手 (00273855)
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| Project Period (FY) |
2001 – 2002
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| Project Status |
Completed (Fiscal Year 2002)
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| Budget Amount *help |
¥3,600,000 (Direct Cost: ¥3,600,000)
Fiscal Year 2002: ¥1,700,000 (Direct Cost: ¥1,700,000)
Fiscal Year 2001: ¥1,900,000 (Direct Cost: ¥1,900,000)
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| Keywords | ORC / heterochromatin / chromosome / mass spectrography / functional regulation / affinity putification / replication / proteomics / 複合体 / 抗体 |
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| Research Abstract |
Origin recognition complex (ORC) plays a central role in regulating initiation of DNA replication in eukaryotes. We have observed cell cycle dependent oscillation of ORC1 (the ORC1 cycle), in which it accumulates in G1 and degrades in S phase, although other ORC subunits (ORCs2-5) exist at almost constant levels throughout the cell cycle. We studied their behavior in human cell nuclei in relation with the ORC1oscilliation. The results demonstrated that ORCs2-5 form a complex throughout the cell cycle, which further associates with ORC1 temporarily according to accumulation of ORC1 in G1 nuclei. ORCs2-5 exist both in nuclease insoluble and soluble fractions. The former population appears in parallel with the accumulation of ORC1 associating with nuclease resistant, non-chromatin nuclear structures. Thus, ORCs2-5 will be temporarily recruited to the nuclease resistant structures by formation of ORC1-5 complex. Indeed, artificial reduction of ORC1 level by transfection of its siRNA in human cells resulted in a shift of ORC2 to the nuclease sensitive population. Furthermore, we observed that association of MCM to chromatin fractions is also blocked by this treatment. These data propose that oscillation of ORC1 actively regulates the status of ORC complex in human cell nuclei by tethering ORCs2-5 to the nuclear structures. This dynamic shift further drives loading of MCMs on chromatin. Thus, it is suggested that the pre-replication complex in human cells will be regulated by temporal accumulation of ORC1 in G1 nuclei.
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