SPERM FUNCTION IN FERTILIZATION EVENTS

受精事件中的精子功能

• 批准号: 2888905
• 负责人: GARY E OLSON
• 金额: $ 23.48万
• 依托单位: VANDERBILT UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 1985
• 资助国家: 美国
• 起止时间: 1985-07-01 至 2002-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/2888905
• 关键词: acrosome; affinity chromatography; fertilization; guinea pigs; hamsters; immunoelectron microscopy; immunofluorescence technique; immunoprecipitation; intermolecular interaction; intracellular membranes; laboratory rabbit; ligands; membrane biogenesis; membrane fusion; membrane proteins; membrane structure; molecular site; protein binding; protein isoforms; protein localization; protein structure function; sperm capacitation; syntaxin; western blottings

The sperm plasma and acrosomal membranes are partitioned into domains of distinct molecular composition which perform specific functions required for mammalian fertilization. The long range goals of this proposal are to define the molecular processes which assemble discrete membrane domains and to identify the biochemical mechanisms regulating the fusion between the periacrosomal plasma membrane and the outer acrosomal membrane during the acrosome reaction. Four aims are proposed addressing these goals. Aim one is to identify the function of the periacrosomal plasma membrane protein PM52 (sperad) in sperm capacitation and/or the acrosome reaction. Immunoprecipitation and blot overlay analyses will be performed to identify ligands for the cytoplasmic domain of PM52 and permeant peptide mimetics of the proline- rich domain of PM52 will be tested for effects on capacitation and the acrosome reaction. Aim 2 is to determine if docking interactions between the periacrosomal plasma membrane and the outer acrosomal membrane generate domain-specific protein localization. Immunoblotting and blot overlays will identify PM52 binding proteins in the outer acrosomal membrane and immunofluorescence experiments will test if disruption of PM52-outer acrosomal membrane interaction results in protein redistribution. Aim 3 is to determine if germ cell isoforms of syntaxin and synaptophysin function in the membrane fusion events of the acrosome reaction. Immunolocalization and immunoprecipitation analyses will be performed to determine if these proteins assemble into a membrane fusion complex required for acrosomal exocytosis. Aim 4 is to identify protein-protein interactions regulating the assembly of the outer acrosomal membrane-acrosomal matrix complex. Biochemical and ultrastructural experiments will be performed to identify proteins of the outer acrosomal membrane which bind to, and regulate the distribution of, acrosomal matrix assemblies within the acrosome. Completion of these studies will provide new insights into the assembly of the acrosomal segment and of its functions during mammalian fertilization and the findings have potential application to strategies for fertility regulation and/or improvement.

精子等离子体和粘膜膜分为域 独特的分子组成，执行特定功能 哺乳动物受精所必需的。 远距离目标 建议是定义组装离散的分子过程 膜域并确定调节的生化机制 周围质膜和外部的融合 在ACROSOM反应过程中，粘膜膜膜。 提出了四个目标 解决这些目标。 目的是确定 细胞体质膜蛋白PM52（Sperad）中的精子 电容和/或ACROSOM反应。 免疫沉淀和印迹 将进行覆盖分析以识别 PM52的细胞质结构域和脯氨酸的pereant肽模拟物 PM52的丰富域将测试是否对电容的影响和 ACROSOM反应。 AIM 2是确定是否对接互动 在周围质膜和外科体的外周围质膜之间 膜产生域特异性蛋白质定位。 免疫印迹 和印迹覆盖物将识别外部的PM52结合蛋白 AROSOMAL膜和免疫荧光实验将测试是否 PM52-OUTER粘膜膜相互作用的破坏导致 蛋白质再分配。 AIM 3是确定生殖细胞同工型是否 在膜融合事件中的语法和突触素功能 ACROSOM反应。 免疫定位和免疫沉淀分析 将执行以确定这些蛋白质是否组装成 诊断胞吐作用所需的膜融合复合物。 目标4是 鉴定调节组装的蛋白质 - 蛋白质相互作用 外脊肉膜 - 杂色基质复合物。 生化和 将进行超微结构实验以识别 与结合并调节的外科体膜 Acrosos中的Arosomal基质组件的分布。 这些研究的完成将为大会提供新的见解 在哺乳动物期间的教科体段及其功能 受精和发现可能适用于策略 用于生育和/或改善。