MAMMALIAN SPERM SURFACE ORGANIZATION AND FUNCTION

本提案的总体目标是了解 精子-卵子质膜黏附和融合过程。第一步 是顶体反应精子顶端(内部)之间的接触 顶体膜)和卵细胞质膜。第二步 这些精子表面赤道/后头部之间的粘附力 区域和卵质膜。在粘合之后可能会有其他步骤 并导致膜融合。先前的工作涉及两个精子表面 蛋白质，受精素和胱氨酸蛋白，作用于赤道/后部 头部区域绑定步骤。缺乏受精素的突变(基因敲除)小鼠 制作。同源的、突变的雄性是不育的。目标1建议 利用这些小鼠来确定受精蛋白在配子中的确切作用 膜结合、融合和卵子激活。Cyritestin基因敲除 也在产生。目标2建议使用缺乏的雄性小鼠 Cyritestin用于确定Cyritestin在配子膜结合中的功能， 融合和卵子激活。目标3专注于配子的第一步 膜相互作用，即精子与卵子的接触 浮出水面。将测试这种接触是否会促进以后的粘合 步骤和/或吞噬类过程将IAM带入 卵子细胞质。Aim 4建议研究精子后可能发生的步骤 捆绑，但在他们融合之前。它将被测试是否有一个 融合前必须发挥作用的金属蛋白酶或表面ATP受体 发生。目标5解决了鸡蛋上有什么分子的问题 质膜参与与精子的相互作用，从而导致 核聚变。它将确定鸡蛋表面整合素，α-6-β-1， 受精素或胞二磷胆碱的粘附性伴侣。它还将 向我们提出一种独特的组织培养细胞系方法 与小鼠精子融合以鉴定另一种促进 膜融合。了解配子融合的分子基础 这一过程可能会使辅助生殖领域出现新的发展。 此外，鉴定出的功能性配子蛋白是潜在的 以药物和药物为基础的新型避孕方法的目标 免疫学方法。

The overall goal of this proposal is to understand each step in the process of sperm-egg plasma membrane adhesion and fusion. The first step is the contact between the tip of the acrosome-reacted sperm (the inner acrosomal membrane, IAM) and the egg plasma membrane. The second step is the adhesion between these sperm surface equatorial/posterior head region and the egg plasm membrane. Additional steps may follow adhesion and lead to membrane fusion. Previous work implicated two sperm surface proteins, fertilin and cyritestin, as acting in the equatorial/posterior head region binding step. Mutant (knockout) mice lacking fertilin were produced. The homologous, mutant males were infertile. Aim 1 proposes to use these mice to define the precise role of fertilin in gamete membrane binding, fusion and egg activation. A cyritestin gene knockout is also being generated. Aim 2 proposes to use male mice lacking cyritestin to determine cyritestin function in gamete membrane binding, fusion and egg activation. Aim 3 focuses on the first step in gamete membrane interaction, i.e. contact of the sperm IAM with the egg surface. It will be tested if this contact promotes later adhesion steps and/or the phagocytosis-like process that takes the IAM into the egg cytoplasm. Aim 4 proposes to study steps that may occur after sperm bind, but before they fuse. It will be tested if there is a metalloprotease or a surface ATP receptor that must act before fusion occurs. Aim 5 addresses the question of what molecules on the egg plasma membrane participate in interactions with the sperm that lead to fusion. It will determine if the egg surface integrin, alpha-6-beta-1, an adhesion partner for either fertilin or cyritestin. It will also propose to us a unique approach with a tissue culture cell line that fuses with mouse sperm to identify another egg protein that promotes membrane fusion. Understanding the molecular basis of the gamete fusion process may allow new developments in assisted reproduction. Furthermore, the identified, functional gamete proteins are potential targets for novel contraceptive methods based on pharmacological and immunological approaches.