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Author:
Date:
2014-07-20
[Abstract] The protein recruitment onto chromatin is a critical process for DNA metabolism, including DNA replication, DNA repair and DNA recombination. Especially DNA modification enzymes and checkpoint proteins are loaded onto DNA damage sites in a context-dependent manner. In our recent study (Kunoh and Habu, 2014), the chromatin association of Pcf1, a large subunit of Chromatin Assembly Factor-1 (CAF-1), was monitored after exposure of cells to hydroxyurea which slowed down the DNA replication. Results of the chromatin fractionation assay provided evidence that Pcf1 was recruited to chromatin upon DNA replication stress. A similar procedure enabled to reveal the chromatin association of Orp1, Mcm proteins, and Swi6 (Sadaie et al., 2008; Ogawa et al., 1999). This assay allows us ...
[摘要] 染色质上的蛋白质募集是DNA代谢的关键过程，包括DNA复制，DNA修复和DNA重组。特别是DNA修饰酶和检查点蛋白以上下文依赖性方式装载到DNA损伤位点。在我们最近的研究中（Kunoh和Habu，2014），在将细胞暴露于羟基脲（其减慢DNA复制）后，监测Pcf1的染色质缔合，其为染色质装配因子-1（CAF-1）的大亚基。染色质分馏测定的结果提供了Pcf1在DNA复制应激时募集到染色质的证据。类似的程序能够揭示Orp1，Mcm蛋白和Swi6的染色质缔合（Sadaie等人，2008; Ogawa等人，1999）。该测定允许我们从活细胞分离染色质结合和非结合蛋白。以下各部分的免疫印迹提供了关于我们的靶蛋白的染色质结合状态的信息。
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