Phosphorylation of human Fen1 by cyclin-dependent kinase modulates its role in replication fork regulation
Phosphorylation of human Fen1 by cyclin-dependent kinase modulates its role in replication fork regulation
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DOI：
10.1038/sj.onc.1206606
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发表时间：
2003-07-10
期刊：
影响因子：
8
通讯作者：
Hübscher, U
中科院分区：
文献类型：
--
作者：
Henneke, G;Koundrioukoff, S;Hübscher, U
细胞周期蛋白依赖性激酶（Cdk） Cdk1-Cyclin A可在S期后期磷酸化DNA复制机制的关键酶Flap内切酶1 （Fen1）。Cdk1-cyclin A在体外和体内与Fen1形成复合物。此外，体内检测到Fen1磷酸化，并依赖于Cdks活性。作为体外Cdk1-Cyclin a磷酸化的功能结果，Fen1的内切酶和外切酶活性降低，但其DNA结合不受影响。此外，Cdk1-Cyclin A对Fen1的磷酸化使其与增殖细胞核抗原（PCNA）的结合消失，从而阻止了PCNA对Fen1的刺激。同时，表达Cdk1-Cyclin A磷酸化缺陷的S187A突变体的人类细胞在S期积累，这与通过DNA复制调节细胞周期的失败一致。我们的研究结果表明，Cdks通过直接靶向参与DNA复制的关键酶，在S期结束时发挥了新的调节作用。
Cyclin-dependent kinase (Cdk) Cdk1-Cyclin A can phosphorylate Flap endonuclease 1 (Fen1), a key-enzyme of the DNA replication machinery, in late S phase. Cdk1-cyclin A forms a complex in vitro and in vivo with Fen1. Furthermore, Fen1 phosphorylation is detected in vivo and depends upon Cdks activity. As a functional consequence of phosphorylation by Cdk1-Cyclin A in vitro, endo- and exonuclease activities of Fen1 are reduced whereas its DNA binding is not affected. Moreover, phosphorylation of Fen1 by Cdk1-Cyclin A abrogates its proliferating cell nuclear antigen (PCNA) binding thus preventing stimulation of Fen1 by PCNA. Concomitantly, human cells expressing the S187A mutant defective for Cdk1-Cyclin A phosphorylation accumulate in S phase consistent with a failure in cell cycle regulation through DNA replication. Our results suggest a novel regulatory role of Cdks onto the end of S phase by targeting directly a key enzyme involved in DNA replication.