Spy1 interacts with p27Kip1 to allow G1/S progression
Spy1 interacts with p27Kip1 to allow G1/S progression
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DOI：
10.1091/mbc.e02-12-0820
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发表时间：
2003-09-01
影响因子：
3.3
通讯作者：
Donoghue, DJ
中科院分区：
文献类型：
--
作者：
Porter, LA;Kong-Beltran, M;Donoghue, DJ
通过G(1)/S的转变，细胞需要合成脱氧核糖核酸。细胞周期蛋白依赖性激酶2(CDK2)是细胞周期通过G(1)/S期及随后的复制活动的主要蛋白。P27是一种CDK抑制物(CKI)，可以与CDK2结合以防止该激酶的过早激活。Spy1是一种新的细胞周期调控蛋白，已被发现在显微注射到非洲爪哇卵母细胞和在哺乳动物细胞中表达时，会过早激活CDK2。为了确定Spy1在哺乳动物细胞周期调控中诱导增殖的机制，我们以人Spy1为诱饵，在酵母双杂交筛选中鉴定相互作用蛋白。其中一个分离的蛋白质是p27；这种新的相互作用在体外得到证实，使用细菌表达的和体外翻译的蛋白质，在体内，通过检查哺乳动物细胞中的内源性和转基因蛋白质。我们证明Spy1的表达可以克服p27诱导的细胞周期停滞，从而允许DNA合成和CDK2组蛋白HI激酶活性。此外，我们利用p27缺失细胞来证明Spy1的增殖效应依赖于内源性p27的存在。我们的数据表明Spy1与p27结合，通过G1/S转换促进细胞周期进程。
Progression through the G(1)/S transition commits cells to synthesize DNA. Cyclin dependent kinase 2 (CDK2) is the major kinase that allows progression through G(1)/S phase and subsequent replication events. p27 is a CDK inhibitor (CKI) that binds to CDK2 to prevent premature activation of this kinase. Speedy (Spy1), a novel cell cycle regulatory protein, has been found to prematurely activate CDK2 when microinjected into Xenopus oocytes and when expressed in mammalian cells. To determine the mechanism underlying Spy1-induced proliferation in mammalian cell cycle regulation, we used human Spy1 as bait in a yeast two-hybrid screen to identify interacting proteins. One of the proteins isolated was p27; this novel interaction was confirmed both in vitro, using bacterially expressed and in vitro translated proteins, and in vivo, through the examination of endogenous and transfected proteins in mammalian cells. We demonstrate that Spy1 expression can overcome a p27-induced cell cycle arrest to allow for DNA synthesis and CDK2 histone HI kinase activity. In addition, we utilized p27-null cells to demonstrate that the proliferative effect of Spy1 depends on the presence of endogenous p27. Our data suggest that Spy1 associates with p27 to promote cell cycle progression through the G1/S transition.