PDF(Pubmed)
Abstract：
The human Origin Recognition Complex (ORC) is required not only for the initiation of DNA replication, but is also implicated in diverse cellular functions, including chromatin organization, centrosome biology, and cytokinesis. The smallest subunit of ORC, Orc6, is poorly conserved amongst eukaryotes. Recent studies from our laboratory have suggested that human Orc6 is not required for replication licensing, but is needed for S-phase progression. Further, ATR-dependent phosphorylation of Orc6 at T229 is implicated in DNA damage response during S-phase. In this study, we demonstrate that the CDK-dependent phosphorylation of Orc6 at T195 occurs during mitosis. While the phosphorylation at T195 does not seem to be required to exit mitosis, cells expressing the phosphomimetic T195E mutant of Orc6 impede S-phase progression. Moreover, the phosphorylated form of Orc6 associates with ORC more robustly, and Orc6 shows enhanced association with the ORC outside of G1, supporting the view that Orc6 may prevent the role of Orc1-5 in licensing outside of G1. Finally, Orc6 and the phosphorylated Orc6 localize to the nucleolar organizing centers and regulate ribosome biogenesis. Our results suggest that phosphorylated Orc6 at T195 prevents replication.
摘要：
人类起源识别复合物（ORC）不仅是DNA复制的起始所必需的，但也牵涉到不同的细胞功能，包括染色质组织，中心体生物学,和胞质分裂。ORC最小的亚单位,Orc6在真核生物中保守性差。我们实验室最近的研究表明，复制许可不需要人Orc6，但S期进展是必需的。Further,在T229的Orc6的ATR依赖性磷酸化与S期的DNA损伤反应有关。在这项研究中,我们证明了在有丝分裂期间发生T195处的CDK依赖性Orc6磷酸化。虽然在T195的磷酸化似乎不需要退出有丝分裂，表达Orc6的磷模拟T195E突变体的细胞阻碍S期进展。此外，Orc6的磷酸化形式与ORC的结合更强烈，和Orc6显示与G1以外的ORC的增强关联，支持Orc6可能阻止Orc1-5在G1以外的许可中的作用的观点。最后,Orc6和磷酸化的Orc6定位于核仁组织中心并调节核糖体生物发生。我们的结果表明，在T195磷酸化的Orc6阻止了复制。
