Localization of human Mcm10 is spatially and temporally regulated during the S phase
Localization of human Mcm10 is spatially and temporally regulated during the S phase
复制标题
DOI：
10.1074/jbc.m314017200
复制
发表时间：
2004-07-30
影响因子：
4.8
通讯作者：
Hanaoka, F
中科院分区：
文献类型：
--
作者：
Izumi, M;Yatagai, F;Hanaoka, F
Mcm10 （Dna43）是酿酒酵母DNA复制起始的重要蛋白。最近，我们发现了一个人类Mcm10同源物，发现它以细胞周期依赖的方式受蛋白水解和磷酸化调节，并且在细胞周期的S期只与染色质结合。然而，Mcm10的确切作用仍然未知。为了研究人Mcm10的定位动力学，我们建立了表达绿色荧光蛋白（GFP）标记Mcm10的HeLa细胞系。从s期早期到中期，GFP-Mcm10出现在离散的核灶中。S期早期，整个细胞核出现数百个病灶。s期中期，病灶出现在核外周和核仁区。在S期晚期和G(2)期，GFP-Mcm10定位于核仁。虽然GFP-Mcm10在S期的分布与复制位点相似，但在大多数情况下，GFP-Mcm10的位点并不与DNA合成位点共定位。此外，GFP-Mcm10分布模式的转变比复制灶模式或增殖细胞核抗原灶模式的变化早30-60分钟。这些结果表明，人类Mcm10在复制前30-60分钟被暂时招募到复制位点，并在复制前复合体激活后与染色质分离。
Mcm10 (Dna43) is an essential protein for the initiation of DNA replication in Saccharomyces cerevisiae. Recently, we identified a human Mcm10 homolog and found that it is regulated by proteolysis and phosphorylation in a cell cycle-dependent manner and that it binds chromatin exclusively during the S phase of the cell cycle. However, the precise roles that Mcm10 plays are still unknown. To study the localization dynamics of human Mcm10, we established HeLa cell lines expressing green fluorescent protein (GFP)-tagged Mcm10. From early to mid-S phase, GFP-Mcm10 appeared in discrete nuclear foci. In early S phase, several hundred foci appeared throughout the nucleus. In mid-S phase, the foci appeared at the nuclear periphery and nucleolar regions. In the late S and G(2) phases, GFP-Mcm10 was localized to nucleoli. Although the distributions of GFP-Mcm10 during the S phase resembled those of replication foci, GFP-Mcm10 foci did not colocalize with sites of DNA synthesis in most cases. Furthermore, the transition of GFP-Mcm10 distribution patterns preceded changes in replication foci patterns or proliferating cell nuclear antigen foci patterns by 30-60 min. These results suggest that human Mcm10 is temporarily recruited to the replication sites 30-60 min before they replicate and that it dissociates from chromatin after the activation of the prereplication complex.