2007 Fiscal Year Final Research Report Summary
A network of histone-chaperones involved in chromatin replication.
| Project/Area Number |
18370068
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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 |
Molecular biology
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| Research Institution | Tokyo Institute of Technology |
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Principal Investigator |
OHSUMI Keita Tokyo Institute of Technology, Graduate School of Bioscience & Biotechnology, Associate Professor (20221822)
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| Co-Investigator(Kenkyū-buntansha) |
IWABUCHI Mari Tokyo Institute of Technology, Graduate School of Bioscience & Biotechnology, Assistant Professor(COE) (40275350)
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| Project Period (FY) |
2006 – 2007
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| Keywords | genome / chromatin / histone proteins / DNA replication / nucleosome |
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| Research Abstract |
In the present research, we investigated the mechanisms for the cell-cycle control and the role of each histone-chaperone for histone H3-H4 in Xenopue eggs, and found the followings.
Using Xenopus eggs and their cell-free extracts, we found that in Xenopus eggs the APC/C inhibitor protein Erp1 was a downstream substrate of the kinase Mos, and that Mos-dependent phosphorylation of Erp1 by Rsk was crucial for both stabilizing Erp1 and enhancing its activity to suppress the APC/C, thereby establishing meiosis-II arrest. We also found that the Ca^<2+>/calmodulin-dependent protein phosphatase calcineurin was transiently activated immediately after Ca^<2+> addition to CSF-arrested egg extracts, independently of CaMKII, whose activity is essential for cancellation of meiosis-II arrest via Erp1 degradation. Thus, in Xenopus eggs, meiosis-II arrest is established and maintained by Erp1 that has been activated by the Mos-Rsk kinase cascade and is canceled by Erp1 degradation that depends on both
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calcineurin and CaMKII.
We have also examined roles of the histone H3-H4 chaperones CAF-1 and Asf1 in assembling both basic and higher-order chromatin structures, using Xenopus egg extracts that support chromosomal DNA replication in S phase and chromosome condensation in M phase. We found that, by immunodepletion of CAF-1 from extracts, only chromatin assembly instantly after DNA replication was disturbed. In contrast, depletion of Asf1 severely impaired chromatin assembly onto nascent DNA during S phase, resulting in not only replicated chromatin with half-reduced nucleosome density but also less condensed, elongated chromosomes in the subsequent M phase. These results strongly suggested that in Xenopus eggs CAF-1 mediates nucleosome assembly coupled to chromosomal DNA replication while Asf-1 is essential for de novo nucleosome assembly onto replicated chromosomal DNA, and that the histone chaperone-mediated de novo nucleosome assembly during chromatin replication ensures the faithful transmission of chromosomes through contributing to mitotic chromosome condensation. Less
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