RFWD3-Mdm2 ubiquitin ligase complex positively regulates p53 stability in response to DNA damage
RFWD3-Mdm2 ubiquitin ligase complex positively regulates p53 stability in response to DNA damage
复制标题
DOI：
10.1073/pnas.0912094107
复制
发表时间：
2010-03-09
影响因子：
11.1
通讯作者：
Wang, Yi
中科院分区：
文献类型：
--
作者：
Fu, Xiaoyong;Yucer, Nur;Wang, Yi
在未受应激的细胞中，肿瘤抑制因子 p53 主要通过 Mdm2 通过泛素介导的蛋白水解作用维持在低水平。为了应对 DNA 损伤，p53 变得稳定并被激活，以开启细胞周期停滞和细胞凋亡所必需的转录程序。 p53 的激活导致 Mdm2 蛋白的积累，Mdm2 蛋白是 p53 的直接转录靶标。目前尚不清楚当 Mdm2 上调时如何保护 p53 免遭降解。在这里，我们报告电离辐射后后期 p53 的稳定与活性泛素化相关。我们发现 E3 泛素连接酶 RFWD3 (RNF201/FLJ10520) 与 Mdm2 和 p53 形成复合物，协同泛素化 p53，并且需要在 DNA 损伤的后期反应中稳定 p53。该过程受到 DNA 损伤检查点的调节，因为 RFWD3 被 ATM/ATR 激酶磷酸化，并且磷酸化突变体无法刺激 p53 泛素化。体外实验表明 RFWD3 是一种 p53 E3 泛素连接酶，RFWD3-Mdm2 复合物限制 Mdm2 对 p53 的多泛素化。我们的研究确定，当 G(1) 细胞周期检查点激活时，RFWD3 是 p53 稳定性的正调节因子，并解释了在高水平 Mdm2 存在的情况下如何保护 p53 免遭降解。
In unstressed cells, the tumor suppressor p53 is maintained at low levels by ubiquitin-mediated proteolysis mainly through Mdm2. In response to DNA damage, p53 is stabilized and becomes activated to turn on transcriptional programs that are essential for cell cycle arrest and apoptosis. Activation of p53 leads to accumulation of Mdm2 protein, a direct transcriptional target of p53. It is not understood how p53 is protected from degradation when Mdm2 is upregulated. Here we report that p53 stabilization in the late phase after ionizing radiation correlates with active ubiquitination. We found that an E3 ubiquitin ligase RFWD3 (RNF201/FLJ10520) forms a complex with Mdm2 and p53 to synergistically ubiquitinate p53 and is required to stabilize p53 in the late response to DNA damage. This process is regulated by the DNA damage checkpoint, because RFWD3 is phosphorylated by ATM/ATR kinases and the phosphorylation mutant fails to stimulate p53 ubiquitination. In vitro experiments suggest that RFWD3 is a p53 E3 ubiquitin ligase and that RFWD3-Mdm2 complex restricts the polyubiquitination of p53 by Mdm2. Our study identifies RFWD3 as a positive regulator of p53 stability when the G(1) cell cycle checkpoint is activated and provides an explanation for how p53 is protected from degradation in the presence of high levels of Mdm2.