Ge Xin Quan, Jackson Dean A, Blow J Julian
Wellcome Trust Biocentre, University of Dundee, Dundee DD1 5EH, United Kingdom.
Genes Dev. 2007 Dec 15;21(24):3331-41. doi: 10.1101/gad.457807.
In late mitosis and early G1, Mcm2-7 complexes are loaded onto DNA to license replication origins for use in the upcoming S phase. However, the amount of Mcm2-7 loaded is in significant excess over the number of origins normally used. We show here that in human cells, excess chromatin-bound Mcm2-7 license dormant replication origins that do not fire during normal DNA replication, in part due to checkpoint activity. Dormant origins were activated within active replicon clusters if replication fork progression was inhibited, despite the activation of S-phase checkpoints. After lowering levels of chromatin-bound Mcm2-7 in human cells by RNA interference (RNAi), the use of dormant origins was suppressed in response to replicative stress. Although cells with lowered chromatin-bound Mcm2-7 replicated at normal rates, when challenged with replication inhibitors they had dramatically reduced rates of DNA synthesis and reduced viability. These results suggest that the use of dormant origins licensed by excess Mcm2-7 is a new and physiologically important mechanism that cells utilize to maintain DNA replication rates under conditions of replicative stress. We propose that checkpoint kinase activity can preferentially suppress initiation within inactive replicon clusters, thereby directing new initiation events toward active clusters that are experiencing replication problems.
在有丝分裂后期和G1期早期，Mcm2 - 7复合物加载到DNA上，为即将到来的S期许可复制起点。然而，加载的Mcm2 - 7量远远超过正常使用的起点数量。我们在此表明，在人类细胞中，过量的与染色质结合的Mcm2 - 7许可了在正常DNA复制期间不启动的休眠复制起点，部分原因是检查点活性。如果复制叉进展受到抑制，休眠起点会在活跃复制子簇内被激活，尽管S期检查点被激活。通过RNA干扰（RNAi）降低人类细胞中与染色质结合的Mcm2 - 7水平后，对复制应激的反应中休眠起点的使用受到抑制。尽管与染色质结合的Mcm2 - 7水平降低的细胞以正常速率复制，但当受到复制抑制剂挑战时，它们的DNA合成速率显著降低且活力下降。这些结果表明，利用由过量Mcm2 - 7许可的休眠起点是细胞在复制应激条件下维持DNA复制速率所利用的一种新的且具有生理重要性的机制。我们提出，检查点激酶活性可以优先抑制非活跃复制子簇内的起始，从而将新的起始事件导向正在经历复制问题的活跃簇。
