Phosphorylation of MCM3 Protein by Cyclin E/Cyclin-dependent Kinase 2 (Cdk2) Regulates Its Function in Cell Cycle
Phosphorylation of MCM3 Protein by Cyclin E/Cyclin-dependent Kinase 2 (Cdk2) Regulates Its Function in Cell Cycle
复制标题
细胞周期蛋白 E/细胞周期蛋白依赖性激酶 2 (Cdk2) 磷酸化 MCM3 蛋白可调节其在细胞周期中的功能
DOI：
10.1074/jbc.m111.226464
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发表时间：
2011-11-18
影响因子：
4.8
通讯作者：
Ye, Xin
中科院分区：
文献类型：
--
作者：
Li, Junhui;Deng, Min;Ye, Xin
MCM 2 -7蛋白形成具有DNA解旋酶活性的稳定异六聚体，在真核细胞的DNA复制中起作用。MCM 2 -7复合物以细胞周期依赖性方式加载到染色质上。MCM 2 -7蛋白的磷酸化有助于MCM 2 -7复合物的形成。然而，特定MCM磷酸化的调节仍需要阐明。在这项研究中，我们证明了MCM 3是细胞周期蛋白E/Cdk 2的底物，并且可以被细胞周期蛋白E/Cdk 2在Thr-722磷酸化。我们发现，与野生型MCM 3相比，MCM 3 T722 A突变体结合染色质的效率低得多，这表明该磷酸化位点参与了MCM 3加载到染色质上。有趣的是，MCM 3的过表达，而不是MCM 3 T722 A突变体，抑制S期进入，而它不影响退出有丝分裂。MCM 3的敲除不影响S期进入和进展，表明一小部分MCM 3足以完成正常的S期。这些结果表明，MCM 3蛋白在染色质上的过量积累可以抑制DNA复制。其他研究表明，过量的MCM 3上调CHK 1 Ser-345和CDK 2 Thr-14的磷酸化。这些数据表明，MCM 3的磷酸化除了调节MCM 2 -7复合物的形成之外，还有助于其控制细胞周期的S期检查点的功能。
MCM2-7 proteins form a stable heterohexamer with DNA helicase activity functioning in the DNA replication of eukaryotic cells. The MCM2-7 complex is loaded onto chromatin in a cell cycle-dependent manner. The phosphorylation of MCM2-7 proteins contributes to the formation of the MCM2-7 complex. However, the regulation of specific MCM phosphorylation still needs to be elucidated. In this study, we demonstrate that MCM3 is a substrate of cyclin E/Cdk2 and can be phosphorylated by cyclin E/Cdk2 at Thr-722. We find that the MCM3 T722A mutant binds chromatin much less efficiently when compared with wild type MCM3, suggesting that this phosphorylation site is involved in MCM3 loading onto chromatin. Interestingly, overexpression of MCM3, but not MCM3 T722A mutant, inhibits the S phase entry, whereas it does not affect the exit from mitosis. Knockdown of MCM3 does not affect S phase entry and progression, indicating that a small fraction of MCM3 is sufficient for normal S phase completion. These results suggest that excess accumulation of MCM3 protein onto chromatin may inhibit DNA replication. Other studies indicate that excess of MCM3 up-regulates the phosphorylation of CHK1 Ser-345 and CDK2 Thr-14. These data reveal that the phosphorylation of MCM3 contributes to its function in controlling the S phase checkpoint of cell cycle in addition to the regulation of formation of the MCM2-7 complex.