Machida Yuichi J, Teer Jamie K, Dutta Anindya
Department of Biochemistry and Molecular Genetics, University of Virginia School of Medicine, Charlottesville, Virginia 22908, USA.
J Biol Chem. 2005 Jul 29;280(30):27624-30. doi: 10.1074/jbc.M502615200. Epub 2005 Jun 7.
The origin recognition complex (ORC) is involved in formation of prereplicative complexes (pre-RCs) on replication origins in the G1 phase. At the G1/S transition, elevated cyclin E-CDK2 activity triggers 1DNA replication to enter S phase. The CDK cycle works as an engine that drives progression of cell cycle events by successive activation of different types of cyclin-CDK. However, how the CDK cycle is coordinated with replication initiation remains elusive. Here we report that acute depletion of ORC2 by RNA interference (RNAi) arrests cells with low cyclin E-CDK2 activity. This result suggests that loss of a replication initiation protein prevents progression of the CDK cycle in G1. p27 and p21 proteins accumulate following ORC2 RNAi and are required for the CDK2 inhibition. Restoration of CDK activity by co-depletion of p27 and p21 allows many ORC2-depleted cells to enter S phase and go on to mitosis. However, in some cells the release of the CDK2 block caused catastrophic events like apoptosis. Therefore, the CDK2 inhibition observed following ORC2 RNAi seems to protect cells from premature S phase entry and crisis in DNA replication. These results demonstrate an unexpected role of ORC2 in CDK2 activation, a linkage that could be important for maintaining genomic stability.
起始识别复合物（ORC）参与在G1期复制起点上形成复制前复合物（pre-RC）。在G1/S期转换时，细胞周期蛋白E-CDK2活性升高触发DNA复制进入S期。CDK循环就像一个引擎，通过相继激活不同类型的细胞周期蛋白-CDK来驱动细胞周期事件的进程。然而，CDK循环如何与复制起始协调仍不清楚。在此我们报道，通过RNA干扰（RNAi）急性耗尽ORC2会使细胞因细胞周期蛋白E-CDK2活性低而停滞。这一结果表明，复制起始蛋白的缺失会阻止G1期CDK循环的进程。在ORC2 RNAi后，p27和p21蛋白积累，并且是CDK2抑制所必需的。通过共同耗尽p27和p21来恢复CDK活性，可使许多ORC2耗尽的细胞进入S期并继续进入有丝分裂。然而，在一些细胞中，CDK2阻滞的解除会导致诸如凋亡等灾难性事件。因此，在ORC2 RNAi后观察到的CDK2抑制似乎可保护细胞避免过早进入S期和DNA复制危机。这些结果证明了ORC2在CDK2激活中具有意想不到的作用，这种联系对于维持基因组稳定性可能很重要。
