Mechanisms of intracellelar signal transduction in sperm-induced mause egg activation

精子诱导的哺乳动物卵子激活与Ca^2+浓度的瞬时增加有关，而Ca^2+浓度的瞬时增加被认为是来自对肌醇1，4，5-三磷酸敏感和对肌醇1，4，5-三磷酸不敏感的细胞内储存。尽管对肌醇1，4，5-三磷酸盐敏感的钙库的重要性已经进行了评估，但对肌醇1，4，5-三磷酸盐不敏感的钙库的身份和作用了解甚少。为了探索兰尼碱敏感性钙库的作用，我首先使用逆转录-聚合酶链反应来鉴定卵子中兰尼碱受体的转录本，并确定存在2型和3型受体的转录本。用能识别2型和3型受体的抗体对放射性碘标记的鸡蛋提取物进行免疫沉淀，可特异性地检测到一条M_r= 520，000的带。用激光共聚焦显微镜对受体进行免疫定位，发现受体均匀分布于囊泡的皮层， ...更多信息 完整的卵母细胞，但成为不对称本地化的皮质中的一个区域，与减数分裂纺锤体在中期II-逮捕鸡蛋，这种不对称的本地化开发中期I. Ryanodine受体在小鼠卵子激活的作用进行了检查，通过确定显微注射的Ryanodine或环ADP核糖对卵子激活的终点，无论是未受精或受精卵的影响。Ryanodine诱导的透明质酸糖蛋白ZP 2的转换，其受精后的形式ZP2_f在一个双相的浓度依赖性的方式;纳摩尔浓度刺激这种转换，而微摩尔浓度没有刺激作用。环ADP核糖也促进ZP 2转化，但具有双曲线浓度依赖性。这些化合物均不诱导细胞周期恢复。最后，在阻断Ca^<2+>从鱼尼汀敏感的细胞库中释放的条件下处理卵子，对受精后卵子的任何激活事件都没有影响。这些实验的结果表明，尽管兰尼碱受体存在并发挥作用，但钙离子库中的Ca^2+释放对于精子诱导的小鼠卵子激活并不是必需的。少

Sperm-induced activation of mammalian eggs is associated with a transient increase in Ca^<2+> concentrations thought to be derived from inositol 1,4,5-trisphosphate-sensitive and -insensitive intracellular stores. whereas the importance of inositol 1,4,5-trisphosphate-sensitive Ca^<2+> stores has been evaluated, the identity and role of inositol 1,4,5-trisphosphate-insensitive stores are poorly understood. To explore the role of the ryanodine-sensitive Ca^<2+> store, I first used reverse transcription-polymerase chain reaction to identify transcripts of the ryanodine receptor in eggs and determined that transcripts for the type 2 and 3 receptor were present. Immunoprecipitation of radioiodinated egg extracts with an antibody that recognizes both type 2 and 3 receptors detected specifically a band of M_r=520,000. Immunolocalization of the receptor (s) using laser-scanning confocal microscopy revealed that the receptor (s) was uniformly distributed in the cortex of the germinal vesicle-i … More ntact oocyte, but became asymmetrically localized to the cortex in a region apposed to the meiotic spindle in the metaphase II-arrested egg ; this asymmetrical localization developed by metaphase I.The role of the ryanodine receptor in mouse egg activation was examined by determining the effects of microinjected ryanodine or cyclic ADP ribose on endpoints of egg activation in either uninseminated or inseminated eggs. Ryanodine induced the conversion of the zona pellucida glycoprotein ZP2 to its post-fertilization form ZP2_f in a biphasic concentration-dependent manner ; nanomolar concentrations stimulated this conversion, whereas micromolar concentrations had no stimulatory effect. Cyclic ADP ribose also promoted the ZP2 conversion, but with a hyperbolic concentration dependence. Neither of these compounds induced cell cycle resumption. Last, treatment of eggs under conditions that would block the release of Ca^<2+> from the ryanodine-sensitive store had no effect on any event of egg activation following fertilization. Results of these experiments suggest that although ryanodine receptors are present and functional, release of Ca^<2+> from this store is not essential for sperminduced mouse egg activation. Less