The cell-cycle regulatory protein Cks1 is required for SCFSkp2-mediated ubiquitinylation of p27
The cell-cycle regulatory protein Cks1 is required for SCFSkp2-mediated ubiquitinylation of p27
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DOI：
10.1038/35060126
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发表时间：
2001-03-01
影响因子：
21.3
通讯作者：
Hershko, A
中科院分区：
文献类型：
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作者：
Ganoth, D;Bornstein, G;Hershko, A
细胞周期蛋白依赖性激酶（CDK）抑制剂p27在G1期晚期通过泛素途径降解（1），使CDK活性驱动细胞进入S期（2）。p27的泛素化需要其在Thr 187处的磷酸化（参考文献3、4）和随后被S期激酶相关蛋白2（Skp 2;参考文献5-8）识别，S期激酶相关蛋白2是与Skp 1、Cul-1和ROC 1/Rbx 1缔合以形成scF泛素连接酶复合物的F盒蛋白家族的成员（9）。然而，在体外连接的p27泛素不能重建已知的纯化成分的SCFSkp 2复合物。在这里，我们发现，缺失的因子是CDK亚基1（Cks 1），它属于高度保守的Suc 1/Cks家族的蛋白质，结合一些CDK和磷酸化的蛋白质，是必不可少的细胞周期进程。人Cks 1，而不是家族的其他成员，在完全纯化的系统中重建p27的泛素连接，与Skp 2结合，并大大增加T187-磷酸化的p27与Skp 2的结合。我们的研究结果代表的第一个证据表明，SCF复合物需要一个辅助蛋白的活动，以及结合其磷酸化底物。
The cyclin-dependent kinase (CDK) inhibitor p27 is degraded in late G1 phase by the ubiquitin pathway(1), allowing CDK activity to drive cells into S phase(2). Ubiquitinylation of p27 requires its phosphorylation at Thr 187 (refs 3, 4) and subsequent recognition by S-phase kinase associated protein 2 (Skp2; refs 5-8), a member of the F-box family of proteins that associates with Skp1, Cul-1 and ROC1/Rbx1 to form an scF ubiquitin ligase complex(9). However, in vitro ligation of p27 to ubiquitin could not be reconstituted by known purified components of the SCFSkp2 complex. Here we show that the missing factor is CDK subunit 1 (Cks1), which belongs to the highly conserved Suc1/Cks family of proteins that bind to some CDKs and phosphorylated proteins and are essential for cell-cycle progression. Human Cks1, but not other members of the family, reconstitutes ubiquitin ligation of p27 in a completely purified system, binds to Skp2 and greatly increases binding of T187-phosphorylated p27 to Skp2. Our results represent the first evidence that an SCF complex requires an accessory protein for activity as well as for binding to its phosphorylated substrate.