Han Seung Jin, Chen Ruby, Paronetto Maria Paola, Conti Marco
Division of Reproductive Biology, Department of Obstetrics and Gynecology, Stanford University, Stanford, California 94305, USA.
Curr Biol. 2005 Sep 20;15(18):1670-6. doi: 10.1016/j.cub.2005.07.056.
In most species, the meiotic cell cycle is arrested at the transition between prophase and metaphase through unclear somatic signals. Activation of the Cdc2-kinase component of maturation promoting factor (MPF) triggers germinal vesicle breakdown after the luteinizing hormone (LH) surge and reentry into the meiotic cell cycle. Although high levels of cAMP and activation of protein kinase A (PKA) play a critical role in maintaining an inactive Cdc2, the steps downstream of PKA in the oocyte remain unknown. Using a small-pool expression-screening strategy, we have isolated several putative PKA substrates from a mouse oocyte cDNA library. One of these clones encodes a Wee1-like kinase that prevents progesterone-induced oocyte maturation when expressed in Xenopus oocytes. Unlike the widely expressed Wee1 and Myt1, mWee1B mRNA and its protein are expressed only in oocytes, and mRNA downregulation by RNAi injection in vitro or transgenic overexpression of RNAi in vivo causes a leaky meiotic arrest. Ser15 residue of mWee1B is the major PKA phosphorylation site in vitro, and the inhibitory effects of the kinase are enhanced when this residue is phosphorylated. Thus, mWee1B is a key MPF inhibitory kinase in mouse oocytes, functions downstream of PKA, and is required for maintaining meiotic arrest.
在大多数物种中，减数分裂细胞周期通过不明的体细胞信号在前期和中期的转换阶段被阻滞。促成熟因子（MPF）的Cdc2激酶成分被激活后，在促黄体生成素（LH）峰后触发生发泡破裂并重新进入减数分裂细胞周期。尽管高水平的cAMP和蛋白激酶A（PKA）的激活在维持Cdc2的无活性状态中起关键作用，但PKA在卵母细胞中的下游步骤仍不清楚。利用小池表达筛选策略，我们从小鼠卵母细胞cDNA文库中分离出了几种假定的PKA底物。其中一个克隆编码一种类似Wee1的激酶，当在非洲爪蟾卵母细胞中表达时，它能阻止孕酮诱导的卵母细胞成熟。与广泛表达的Wee1和Myt1不同，mWee1B mRNA及其蛋白仅在卵母细胞中表达，体外RNAi注射下调mRNA或体内RNAi转基因过表达会导致减数分裂阻滞渗漏。mWee1B的Ser15残基是体外主要的PKA磷酸化位点，当该残基被磷酸化时，激酶的抑制作用增强。因此，mWee1B是小鼠卵母细胞中关键的MPF抑制激酶，在PKA下游发挥作用，是维持减数分裂阻滞所必需的。
