Furuya S, Endo Y, Osumi K, Oba M, Nozawa S, Suzuki S
Department of Obstetrics and Gynecology, Keio University School of Medicine, Tokyo, Japan.
Fertil Steril. 1993 Jan;59(1):216-22. doi: 10.1016/s0015-0282(16)55642-8.
To examine the effects of protein phosphatase inhibitors on capacitation and protein phosphorylation in human sperm.
The chlortetracycline (CTC) fluorescence assay was used to monitor capacitated sperm treated with or without protein phosphatase inhibitors. Capacitation was confirmed by the ability of sperm to undergo the acrosome reaction in response to Ca++ ionophore A23187 or mouse zonae pellucidae. 32P-labeled sperm phosphoproteins were analyzed by one-dimensional gel electrophoresis to detect the effects of protein phosphatase inhibitors on protein phosphorylation.
The treatment of sperm with calyculin A resulted in the following: [1] the rapid appearance of the clear perimeter pattern, specifically, distribution of fluorescence over the entire head exhibiting a bright perimeter and bright midpiece, in a dose-dependent manner in the 1 to 100 nM range; [2] an accelerated ability to undergo the acrosome reaction; and [3] an enhanced phosphorylation of sperm phosphoproteins in a dose-related fashion in the 1 to 100 nM range. A similar stimulatory effect was observed only with a 100-fold higher concentrations of okadaic acid, another protein phosphatase inhibitor.
Our results strongly suggest that protein phosphorylation and dephosphorylation may be involved in the regulation of human sperm capacitation.
研究蛋白磷酸酶抑制剂对人精子获能及蛋白磷酸化的影响。
采用金霉素（CTC）荧光分析法监测经蛋白磷酸酶抑制剂处理或未处理的获能精子。通过精子对钙离子载体A23187或小鼠透明带发生顶体反应的能力来确认获能。用一维凝胶电泳分析32P标记的精子磷蛋白，以检测蛋白磷酸酶抑制剂对蛋白磷酸化的影响。
用花萼海绵诱癌素A处理精子导致以下结果：[1]清晰周边模式迅速出现，具体而言，荧光在整个头部的分布呈现明亮的周边和明亮的中段，在1至100 nM范围内呈剂量依赖性；[2]顶体反应能力加快；[3]在1至100 nM范围内，精子磷蛋白磷酸化以剂量相关方式增强。仅在浓度高100倍的另一种蛋白磷酸酶抑制剂冈田酸作用下观察到类似的刺激作用。
我们的结果有力地表明，蛋白磷酸化和去磷酸化可能参与人精子获能的调节。
