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A network of histone-chaperones involved in chromatin replication.

参与染色质复制的组蛋白伴侣网络。

基本信息

批准号：
18370068
负责人：
OHSUMI Keita
金额：
$ 11.08万
依托单位：
Tokyo Institute of Technology
依托单位国家：
日本
项目类别：
Grant-in-Aid for Scientific Research (B)
财政年份：
2006
资助国家：
日本
起止时间：
2006 至 2007
项目状态：
已结题

来源：
https://kaken.nii.ac.jp/grant/KAKENHI-PROJECT-18370068/
关键词：
genome chromatin histone proteins DNA replication nucleosome

项目摘要

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 … More 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

在本研究中，我们研究了非洲爪蟾卵中细胞周期控制的机制和组蛋白H3-H4的每个组蛋白-伴侣蛋白的作用，并发现：使用非洲爪蟾卵及其无细胞提取物，我们发现在非洲爪蟾卵中APC/C抑制剂蛋白Erp 1是激酶Mos的下游底物，Rsk对Erp 1的Mos依赖性磷酸化对稳定Erp 1和增强其抑制APC/C的活性至关重要，从而建立减数分裂-II停滞。我们还发现，在CSF阻滞的卵提取物中加入Ca^2+后，Ca ^2 +/钙调素依赖性蛋白磷酸酶钙调磷酸酶立即被瞬时激活，这与CaMK II无关，CaMK II的活性是通过Erp 1降解来取消减数分裂-II阻滞所必需的。因此，在非洲爪蟾卵中，减数分裂-II停滞是由Erp 1建立和维持的，Erp 1已被Mos-Rsk激酶级联激活，并被Erp 1降解所取消，Erp 1降解依赖于两种激酶级联。 ...更多信息我们还研究了组蛋白H3-H4伴侣CAF-1和Asf 1在组装基本和高阶染色质结构中的作用，使用非洲爪蟾卵提取物，其支持S期染色体DNA复制和M期染色体凝聚。我们发现，通过免疫耗竭提取物中的CAF-1，只有DNA复制后立即发生的染色质组装受到干扰。相反，Asf 1的耗竭严重损害染色质组装到新生的DNA在S期，不仅导致复制的染色质与核小体密度减少一半，但也减少凝聚，延长染色体在随后的M期。这些结果有力地表明，在非洲爪蟾卵CAF-1介导的核小体组装耦合到染色体DNA复制，而Asf-1是必要的从头核小体组装到复制的染色体DNA，和组蛋白伴侣介导的从头核小体组装在染色质复制过程中确保染色体的忠实传输，通过有丝分裂染色体凝聚。少