CDK Inhibitor p21 Is Degraded by a Proliferating Cell Nuclear Antigen-coupled Cul4-DDB1Cdt2 Pathway during S Phase and after UV Irradiation
CDK Inhibitor p21 Is Degraded by a Proliferating Cell Nuclear Antigen-coupled Cul4-DDB1Cdt2 Pathway during S Phase and after UV Irradiation
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DOI：
10.1074/jbc.m806045200
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发表时间：
2008-10-24
影响因子：
4.8
通讯作者：
Tsurimoto, Toshiki
中科院分区：
文献类型：
--
作者：
Nishitani, Hideo;Shiomi, Yasushi;Tsurimoto, Toshiki
以前的报告表明，染色质相关的PCNA夫妇的DNA复制与Cul 4-DDB 1（Cdt 2）依赖的蛋白水解的许可因子Cdt 1。CDK抑制剂p21，另一种PCNA结合蛋白，也在S期和UV照射后降解。在这里，我们表明，p21是由相同的泛素-蛋白酶体途径Cdt 1在HeLa细胞中降解。当PCNA或Cul 4-DDB 1（Cdt 2）的组分被沉默或当p21上的PCNA结合位点被突变时，在S期和UV照射后p21的降解都被阻止。p21在细胞中表达时与Cul 4A和DDB 1蛋白共免疫沉淀。纯化的Cul 4A-DDB 1（Cdt 2）复合物在体外泛素化p21。p21蛋白在S期表达较低，在G（2）期表达升高。突变分析表明，除了PCNA结合结构域，其侧翼区也是重要的识别Cul 4-DDB 1（Cdt 2）。我们的研究结果提供了一个新的方面的蛋白水解控制p21在细胞周期。
Previous reports showed that chromatin-associated PCNA couples DNA replication with Cul4-DDB1(Cdt2)-dependent proteolysis of the licensing factor Cdt1. The CDK inhibitor p21, another PCNA-binding protein, is also degraded both in S phase and after UV irradiation. Here we show that p21 is degraded by the same ubiquitin-proteasome pathway as Cdt1 in HeLa cells. When PCNA or components of Cul4-DDB1(Cdt2) were silenced or when the PCNA binding site on p21 was mutated, degradation of p21 was prevented both in S phase and after UV irradiation. p21 was co-immunoprecipitated with Cul4A and DDB1 proteins when expressed in cells. The purified Cul4A-DDB1(Cdt2) complex ubiquitinated p21 in vitro. Consistently, p21 protein levels are low during S phase and increase around G(2) phase. Mutational analysis suggested that in addition to the PCNA binding domain, its flanking regions are also important for recognition by Cul4-DDB1(Cdt2). Our findings provide a new aspect of proteolytic control of p21 during the cell cycle.