MECHANISMS OF MAMMALIAN FERTILIZATION

透明带(ZP)是一种厚厚的细胞外覆盖物，包裹在所有哺乳动物的卵子周围。这个 小鼠卵子ZP由三种糖蛋白组成，称为mZP1-3。编码糖蛋白的基因是 仅在2-3周内生长的小鼠卵母细胞和新生小鼠卵母细胞表达 糖蛋白被组装成组成ZP的交联丝。其中一个 糖蛋白mZP3是通过自由游动、顶体完整的物种特异性方式识别的 当精子与未受精卵结合时。与mZP3结合诱导精子经历顶体 反应，一种细胞胞吐作用。只有顶体反应的精子才能穿透ZP并融合 与卵质膜形成合子。在本申请中，描述了实验 继续解决有关ZP糖蛋白和鸡蛋ZP的重大问题，具体目标是 这项建议的目的是评估mZP2和mZP3多肽一级结构之间的关系 并利用生长中的小鼠卵母细胞将这些糖蛋白分泌和组装成细丝， 转基因小鼠，和转基因哺乳动物细胞。ZP的分泌和组装被认为是 糖蛋白受rnZP2和MZL&GT；3多肽的特定结构域调控。许多人 提出的实验是可能的，因为在实验期间获得的结果和发展的方法 上一个授权期。这些实验包括使用突变的和野生型的mZP基因， 不同表位标记的mZP基因构建物显微注射到生长中的小鼠卵母细胞， 转基因小鼠携带突变的转基因基因，并转基因哺乳动物细胞。这些措施的结果 实验应该提供重要的分子信息，说明 ZP糖蛋白一级结构，生长中卵母细胞分泌ZP糖蛋白，以及 将ZP糖蛋白组装成长丝。鉴于鼠标之间有很大的相似性 和人类ZP糖蛋白，在小鼠和人类受精之间，很可能是 这里获得的信息也适用于人类。此外，这项研究可能会有 开发预防ZP的新型可逆避孕方法的重要后果 人类卵子发生过程中的形成。

The zona peltucida (ZP) is a thick extracellular c0at that surrounds all mammalian eggs. The mouse egg ZP consists of three glycoproteins, called mZP1-3. Genes encoding the glycoproteins are expressed exclusively by growing mouse oocytes during a 2-3 week period and the nascent glycoproteins are assembled into crosslinked filaments that constitute the ZP. One of the glycoproteins, mZP3, is recognized in a species-specific manner by free-swimming, acrosome-intact sperm when they bind to unfertilized eggs. Binding to mZP3 induces sperm to undergo the acrosome reaction, a form of cellular exocytosis. Only acrosome-reacted sperm can penetrate the ZP and fuse with egg plasma membrane to form a zygote. In this application, experiments are described that continue to address significant questions about ZP glycoproteins and the egg ZP, The Specific Aim of this proposal is to evaluate relationships between mZP2 and mZP3 polypeptide primary structures and secretion and assembly of these glycoproteins into filaments using growing mouse oocytes, transgenic mice, and transfected mammalian cells. It is proposed that secretion and assembly of ZP glycoproteins is regulated by specific domains of rnZP2 and mZl>3 polypeptides. Many of the experiments proposed are possible because of results obtained and methodolgy developed during the previous grant period. The experiments include the use of mutated and wild-type mZP genes, a variety of epitope-tagged mZP cDNA constructs microinjected into growing mouse oocytes, transgenic mice carrying mutated transgenes, and transfected mammalian cells. Results of these experiments should provide important molecular information about possible relationships between ZP glycoprotein primary structures, secretion of ZP glycoproteins from growing oocytes, and assembly of ZP glycoproteins into long filaments. In view of the great similarities between mouse and human ZP glycoproteins and between fertilization in mice and humans, it is very likely that information obtained here will apply to humans as well. Furthermore, the research could have important consequences for development of novel reversible contraceptive methods that prevent ZP formation during oogenesis in humans.