M phase phosphorylation of the epigenetic regulator UHRF1 regulates its physical association with the deubiquitylase USP7 and stability
M phase phosphorylation of the epigenetic regulator UHRF1 regulates its physical association with the deubiquitylase USP7 and stability
复制标题
表观遗传调节因子 UHRF1 的 M 期磷酸化调节其与去泛素化酶 USP7 的物理关联和稳定性
DOI：
10.1073/pnas.1116349109
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发表时间：
2012-03-27
影响因子：
11.1
通讯作者：
Shi, Yang
中科院分区：
文献类型：
--
作者：
Ma, Honghui;Chen, Hao;Shi, Yang
UHRF1（具有泛素样，具有PHD和环手指结构域1）在DNA CpG甲基化，异染色质功能和基因表达中起重要作用。已经建议对UHRF1的过表达有助于肿瘤发生。但是，UHRF1的调节在很大程度上是未知的。在这里，我们证明了去偶联性usp7与UHRF1相互作用。使用USP7的相互作用缺陷和催化突变体进行互补实验，我们证明了USP7的物理相互作用和催化活性对于UHRF1泛素化和稳定性调节都是必需的。质谱分析鉴定了UHRF1 USP7相互作用结构域内丝氨酸652的磷酸化，并由UHRF1 S652磷酸（S652PH）特定抗体进一步证实。重要的是，S652PH抗体在有丝分裂细胞中鉴定了磷酸化的UHRF1，并且S652可以通过M相特异性激酶CDK1-CYCLIN B在体外磷酸化。 UHRF1 S652磷酸化显着降低了在体外和体内与USP7的相互作用，这与细胞周期的M相中的UHRF1稳定性降低相关。相反，携带S652A突变的UHRF1更稳定，该突变使UHRF1具有抗磷酸化的UHRF1。重要的是，携带S652A突变体的细胞生长得更慢，表明保持适当水平的UHRF1水平对于细胞增殖调节很重要。综上所述，我们的发现发现了一个细胞周期特异性的信号传导事件，该事件使UHRF1与USP7的相互作用相互作用，从而将UHRF1暴露于蛋白酶体介导的降解中。这些发现确定了通过调节细胞UHRF1水平的分子机制，这可能会影响细胞增殖。
UHRF1 (Ubiquitin-like, with PHD and RING finger domains 1) plays an important role in DNA CpG methylation, heterochromatin function and gene expression. Overexpression of UHRF1 has been suggested to contribute to tumorigenesis. However, regulation of UHRF1 is largely unknown. Here we show that the deubiquitylase USP7 interacts with UHRF1. Using interaction-defective and catalytic mutants of USP7 for complementation experiments, we demonstrate that both physical interaction and catalytic activity of USP7 are necessary for UHRF1 ubiquitylation and stability regulation. Mass spectrometry analysis identified phosphorylation of serine (S) 652 within the USP7-interacting domain of UHRF1, which was further confirmed by a UHRF1 S652 phosphor (S652ph)-specific antibody. Importantly, the S652ph antibody identifies phosphorylated UHRF1 in mitotic cells and consistently S652 can be phosphorylated by the M phase-specific kinase CDK1-cyclin B in vitro. UHRF1 S652 phosphorylation significantly reduces UHRF1 interaction with USP7 in vitro and in vivo, which is correlated with a decreased UHRF1 stability in the M phase of the cell cycle. In contrast, UHRF1 carrying the S652A mutation, which renders UHRF1 resistant to phosphorylation at S652, is more stable. Importantly, cells carrying the S652A mutant grow more slowly suggesting that maintaining an appropriate level of UHRF1 is important for cell proliferation regulation. Taken together, our findings uncovered a cell cycle-specific signaling event that relieves UHRF1 from its interaction with USP7, thus exposing UHRF1 to proteasome-mediated degradation. These findings identify a molecular mechanism by which cellular UHRF1 level is regulated, which may impact cell proliferation.