Foster J S, Wimalasena J
Department of Obstetrics and Gynecology, University of Tennessee Medical Center, Knoxville 37920, USA.
Mol Endocrinol. 1996 May;10(5):488-98. doi: 10.1210/mend.10.5.8732680.
Cyclin-dependent kinases (Cdk) act to regulate G1- to S-phase transition in mammalian cells. We have studied the effects of estradiol and the steroidal antiestrogen ICI 182, 780 on induction of Cdk activity and the consequent phosphorylation of retinoblastoma protein (Rb) in estrogen-responsive MCF-7 breast cancer cells. Treatment of growth-arrested MCF-7 cells with physiological concentrations of estradiol led to a time-dependent increase in Cdk2-associated and cyclin E-dependent kinase activity, which was accompanied by hyperphosphorylation of Rb and S-phase entry. Induction of both Cdk2 activity and DNA synthesis by estradiol was dose dependent and was inhibited by coadministration of ICI 182,780. Elicitation of Cdk2 activity was found to require prolonged (> 8h) estradiol exposure. Levels of cyclins E and A were unchanged in MCF-7 cells undergoing G1- to S-transit; however, synthesis and steady state levels of cyclin D1 protein were increased by estradiol. Cdk4-associated Rb kinase activity was evident in MCF-7 cells by 6 h after estradiol exposure and was inhibited by antiestrogen. Cdk2 and Cdk4 protein levels were not altered by estrogen treatment; however, faster migrating, phosphorylated Cdk2 forms increased in estradiol-treated MCF-7 cells by 12 after release from growth arrest. Cdtk-inhibitory activities, associated with p27kip-1, were eliminated from growth-arrested MCF-7 cells after treatment with estradiol but were not eliminated from cells cotreated with estradiol and ICI 182,780. These findings suggest that estradiol regulates G1 progression in MCF-7 cells through direct effects upon Cdk activation, Rb phosphorylation, and by inducing elimination of Cdk inhibitors.
细胞周期蛋白依赖性激酶（Cdk）在哺乳动物细胞中发挥作用，调节从G1期到S期的转变。我们研究了雌二醇和甾体类抗雌激素ICI 182,780对雌激素反应性MCF-7乳腺癌细胞中Cdk活性诱导以及视网膜母细胞瘤蛋白（Rb）随后磷酸化的影响。用生理浓度的雌二醇处理生长停滞的MCF-7细胞，导致与Cdk2相关的、细胞周期蛋白E依赖性激酶活性随时间增加，同时伴有Rb的过度磷酸化和进入S期。雌二醇诱导的Cdk2活性和DNA合成均呈剂量依赖性，并被同时给予的ICI 182,780所抑制。发现诱导Cdk2活性需要长时间（>8小时）暴露于雌二醇。在经历从G1期到S期转变的MCF-7细胞中，细胞周期蛋白E和A的水平没有变化；然而，雌二醇增加了细胞周期蛋白D1蛋白的合成和稳态水平。雌二醇暴露6小时后，MCF-7细胞中与Cdk4相关的Rb激酶活性明显，且被抗雌激素抑制。雌激素处理未改变Cdk2和Cdk4蛋白水平；然而，从生长停滞状态释放12小时后，在经雌二醇处理的MCF-7细胞中，迁移速度更快的磷酸化Cdk2形式增加。用雌二醇处理生长停滞的MCF-7细胞后，与p27kip-1相关的Cdtk抑制活性被消除，但在用雌二醇和ICI 182,780共同处理的细胞中未被消除。这些发现表明，雌二醇通过直接影响Cdk激活、Rb磷酸化以及诱导消除Cdk抑制剂来调节MCF-7细胞中的G1期进程。
