Mammalian Gamete Surface Organization and Function

描述（由申请人提供）：精卵融合发生在受精的关键时刻，受精卵形成。在最近的进展中，配子表面的关键蛋白质与膜融合有关。目前的证据表明：（1）精子中的亚当斯可能在融合中起作用;（2）卵细胞中的四跨膜蛋白CD 9和卵细胞中的GPI锚定蛋白是融合所必需的。 ADAM蛋白可能具有细胞-细胞粘附活性或蛋白酶活性，我们认为这些都是正常受精所必需的。虽然基因敲除研究表明，亚当斯1，2和3不是融合所必需的，但其他ADAM家族成员可能也有作用。我们将测试不同的ADAM是否在精子和卵子质膜的融合中起作用。我们还将测试一种已知的蛋白酶，ADAM 24，是否能产生多精受精的膜阻滞。由于基因缺失的研究表明，卵CD 9是所需的融合，我们专注于CD 9的作用机制和其本地化的脂质微区。已知CD 9具有可溶性配体，我们将测试精子是否具有与CD 9结合的相关跨膜配体。在组织培养系中，CD 9与表达CD 9的细胞表面上的许多其他蛋白质缔合。我们将测试卵子CD 9相关蛋白是否与精子表面结合。基因敲除研究还表明，卵GPI锚定蛋白是融合所必需的。我们建议鉴定关键的GPI锚定蛋白并分析其作用模式。由于多个蛋白质似乎在融合中发挥作用，我们问他们的活动是如何协调的。我们建议，在融合中发挥作用的活性卵分子必须组织成脂质微区。因此，这些研究旨在将联合收割机分子和细胞水平结合起来，为配子膜融合过程带来新的见解。

DESCRIPTION (provided by applicant): Sperm-egg fusion occurs at a critical moment in fertilization at which the zygote is formed. In recent progress key proteins on the gamete surfaces have been implicated in membrane fusion. Current evidence indicates: (1) sperm ADAMs may have a role in fusion and (2) the egg tetraspanin CD9 and egg GPI-anchored proteins are required for fusion. ADAM proteins potentially have either cell-cell adhesion activity or protease activity and we propose that each of these is required for normal fertilization. Although gene knockout studies have shown that ADAMs 1, 2 and 3 are not required for fusion, it is possible that other ADAM family members have a role. We will test if a different ADAM acts in fusion of the sperm and egg plasma membranes. We will also test if a known protease, ADAM 24, acts to create the membrane block to polyspermy. Since gene deletion studies have shown that egg CD9 is required for fusion, we focus on CD9's mechanism of action and its localization to lipid microdomains. CD9 is known to have a soluble ligand and we will test if sperm have a related transmembrane ligand to which CD9 binds. In tissue culture lines, CD9 associates with many other proteins on the surface of a CD9-expressing cell. We will test if an egg CD9-associated protein binds to the sperm surface. Gene knockout studies have also shown that egg GPI-anchored proteins are required for fusion. We propose to identify the critical GPl-anchored protein(s) and analyze its mode of action. Since multiple proteins appear to play a role in fusion, we ask how their activities are coordinated. We propose that to function in fusion the active egg molecules must be organized into lipid microdomains. Thus, these studies are designed to combine molecular and cellular levels to bring new insight into the process of gamete membrane fusion.