Degradation of the CDK inhibitor p27^<Kip1> by a novel ubiquitin ligase.

新型泛素连接酶降解 CDK 抑制剂 p27^<Kip1>。

基本信息

批准号：
17590243
负责人：
UCHIDA Chiharu
金额：
$ 2.24万
依托单位：
Hamamatsu University School of Medicine
依托单位国家：
日本
项目类别：
Grant-in-Aid for Scientific Research (C)
财政年份：
2005
资助国家：
日本
起止时间：
2005 至 2006
项目状态：
已结题

来源：
https://kaken.nii.ac.jp/en/grant/KAKENHI-PROJECT-17590243/
关键词：
p27kip1 Ubiquitin Proteasome Degradation Cell cycle p27<kip1>

项目摘要

Cell cycle progression is controlled by cyclin-dependent kinases (CDKs). The CDK activities are negatively regulated by CDK inhibitor proteins (CKIs). In the quiescent and early G1 phase, p27^<Kip1>, one of the Cip/Kip-type CKIs, exists in abundance in the cell nucleus to suppress cell cycle progression. p27^<Kip1> is degraded in late G1 phase by the ubiquitin-proteasome pathway, allowing cells to enter S phase. In this study, we identified p27NBP1 as a p27^<Kip1>-interacting protein. p27NBP1 physically interacted with p27^<Kip1> and directly ubiquitinated p27^<Kip1> in an intact RING finger domain-dependent manner. Since we isolated p27NBP1 as a protein that interacts with the amino-terminal portion of p27^<Kip1> containing Ser10, we next examined whether interaction between these two proteins was affected by Ser10 phosphorylation status. In spite of the substitution of Ser10, p27^<Kip1> mutants associated with p27NBP1 to an extent similar to that of wild-type p27^<Kip1>. This result suggested that p27NBP1 interacts with p27^<Kip1> regardless of the status of Ser10 phosphorylation. Ablation of endogenous p27NBP1 by small interfering RNA increased the steady-state level of p27^<Kip1> and decelerated p27^<Kip1> turnover. p27^<Kip1> was decreased in synchronization with accumulation of p27NBP1 in late G1 phase. Furthermore, depletion of p27NBP1 resulted in inhibition of cell cycle progression from G1 to S phase in a p53-independent manner. Overall, the results indicate that p27NBP1 acts as a negative regulator of p27^<Kip1> function by promoting ubiquitin-dependent proteasomal degradation.

细胞周期进程由细胞周期蛋白依赖性激酶 (CDK) 控制。 CDK 活性受到 CDK 抑制蛋白 (CKI) 的负向调节。在静止期和早期 G1 期，p27^<Kip1>（Cip/Kip 型 CKI 之一）在细胞核中大量存在，可抑制细胞周期进程。 p27^<Kip1> 在 G1 期晚期通过泛素-蛋白酶体途径降解，使细胞进入 S 期。在本研究中，我们将 p27NBP1 鉴定为 p27^<Kip1> 相互作用蛋白。 p27NBP1 与 p27^<Kip1> 发生物理相互作用，并以完整的环指结构域依赖性方式直接泛素化 p27^<Kip1>。由于我们将 p27NBP1 分离为与含有 Ser10 的 p27^<Kip1> 氨基末端部分相互作用的蛋白质，因此我们接下来检查了这两种蛋白质之间的相互作用是否受到 Ser10 磷酸化状态的影响。尽管有 Ser10 的取代，p27^<Kip1> 突变体与 p27NBP1 的关联程度与野生型 p27^<Kip1> 相似。该结果表明，无论 Ser10 磷酸化状态如何，p27NBP1 都会与 p27^<Kip1> 相互作用。小干扰 RNA 消除内源性 p27NBP1 会增加 p27^<Kip1> 的稳态水平，并降低 p27^<Kip1> 周转率。 p27^<Kip1> 与 G1 期晚期 p27NBP1 的积累同步减少。此外，p27NBP1 的耗竭导致以不依赖于 p53 的方式抑制细胞周期从 G1 期进展到 S 期。总体而言，结果表明 p27NBP1 通过促进泛素依赖性蛋白酶体降解而充当 p27^<Kip1> 功能的负调节因子。