Xu Shuichan, Abbasian Mahan, Patel Palka, Jensen-Pergakes Kristen, Lombardo Christian R, Cathers Brian E, Xie Weilin, Mercurio Frank, Pagano Michele, Giegel David, Cox Sarah
Department of Biochemistry and Biomarker Development, Signal Pharmaceuticals, LLC, San Diego, California 92121, USA.
J Biol Chem. 2007 May 25;282(21):15462-70. doi: 10.1074/jbc.M610758200. Epub 2007 Apr 4.
p27, an important cell cycle regulator, blocks the G(1)/S transition in cells by binding and inhibiting Cdk2/cyclin A and Cdk2/cyclin E complexes (Cdk2/E). Ubiquitination and subsequent degradation play a critical role in regulating the levels of p27 during cell cycle progression. Here we provide evidence suggesting that both Cdk2/E and phosphorylation of Thr(187) on p27 are essential for the recognition of p27 by the SCF(Skp2/Cks1) complex, the ubiquitin-protein isopeptide ligase (E3). Cdk2/E provides a high affinity binding site, whereas the phosphorylated Thr(187) provides a low affinity binding site for the Skp2/Cks1 complex. Furthermore, binding of phosphorylated p27/Cdk2/E to the E3 complex showed positive cooperativity. Consistently, p27 is also ubiquitinated in a similarly cooperative manner. In the absence of p27, Cdk2/E and Cks1 increase Skp2 phosphorylation. This phosphorylation enhances Skp2 auto-ubiquitination, whereas p27 inhibits both phosphorylation and auto-ubiquitination of Skp2.
p27是一种重要的细胞周期调节因子，它通过结合并抑制细胞周期蛋白依赖性激酶2/细胞周期蛋白A（Cdk2/cyclin A）和细胞周期蛋白依赖性激酶2/细胞周期蛋白E复合物（Cdk2/E）来阻断细胞中的G(1)/S期转换。泛素化及随后的降解在细胞周期进程中调节p27水平方面起着关键作用。在此，我们提供证据表明，Cdk2/E以及p27上苏氨酸187（Thr(187)）的磷酸化对于泛素蛋白异肽连接酶（E3）即SCF(Skp2/Cks1)复合物识别p27而言都是必不可少的。Cdk2/E提供一个高亲和力结合位点，而磷酸化的Thr(187)为Skp2/Cks1复合物提供一个低亲和力结合位点。此外，磷酸化的p27/Cdk2/E与E3复合物的结合表现出正协同性。同样，p27也以类似的协同方式被泛素化。在缺乏p27的情况下，Cdk2/E和Cks1会增加Skp2的磷酸化。这种磷酸化增强了Skp2的自身泛素化，而p27则抑制Skp2的磷酸化和自身泛素化。
