Department of Biological Sciences, Seoul National University, Seoul 151-742, Korea.
J Biol Chem. 2012 Apr 6;287(15):11891-8. doi: 10.1074/jbc.M111.338467. Epub 2012 Feb 13.
During the late M to the G(1) phase of the cell cycle, the origin recognition complex (ORC) binds to the replication origin, leading to the assembly of the prereplicative complex for subsequent initiation of eukaryotic chromosome replication. We found that the cell cycle-dependent phosphorylation of human ORC2, one of the six subunits of ORC, dissociates ORC2, -3, -4, and -5 (ORC2-5) subunits from chromatin and replication origins. Phosphorylation at Thr-116 and Thr-226 of ORC2 occurs by cyclin-dependent kinase during the S phase and is maintained until the M phase. Phosphorylation of ORC2 at Thr-116 and Thr-226 dissociated the ORC2-5 from chromatin. Consistent with this, the phosphomimetic ORC2 protein exhibited defective binding to replication origins as well as to chromatin, whereas the phosphodefective protein persisted in binding throughout the cell cycle. These results suggest that the phosphorylation of ORC2 dissociates ORC from chromatin and replication origins and inhibits binding of ORC to newly replicated DNA.
在细胞周期的 M 期到 G1 期晚期，起始识别复合物（ORC）与复制起始点结合，导致前复制复合物的组装，随后启动真核染色体复制。我们发现，人类 ORC 的六个亚基之一 ORC2 的细胞周期依赖性磷酸化，将 ORC2、-3、-4 和 -5（ORC2-5）亚基从染色质和复制起始点上解离下来。ORC2 的 Thr-116 和 Thr-226 的磷酸化是由细胞周期蛋白依赖性激酶在 S 期进行的，并一直维持到 M 期。ORC2 的 Thr-116 和 Thr-226 的磷酸化将 ORC2-5 从染色质上解离下来。与此一致的是，磷酸模拟的 ORC2 蛋白表现出与复制起始点以及染色质结合的缺陷，而磷酸缺陷的蛋白在整个细胞周期中持续结合。这些结果表明，ORC2 的磷酸化将 ORC 从染色质和复制起始点上解离下来，并抑制 ORC 与新复制的 DNA 的结合。
