Mammalian Gamete Surface Organization and Function

描述(申请人提供)：精子-卵子融合发生在受精的关键时刻，也就是合子形成的时刻。在最近的研究中，配子表面的关键蛋白与膜融合有关。目前的证据表明：(1)精子ADAMS可能在融合中起作用；(2)融合需要卵子Tetraspanin CD9和卵子GPI锚定蛋白。 ADAM蛋白可能具有细胞-细胞黏附活性或蛋白酶活性，我们认为这两种活性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，但其他亚当家族成员可能也有作用。我们将测试是否有不同的亚当参与精子和卵子质膜的融合。我们还将测试一种已知的蛋白酶ADAM 24是否作用于创建多精受精的膜块。由于基因缺失的研究表明，鸡蛋CD9的融合是必需的，所以我们重点研究CD9的S作用机制及其在脂质微区的定位。CD9已知有一个可溶的配体，我们将测试精子是否有一个与CD9结合的相关跨膜配体。在组织培养系中，CD9与表达CD9的细胞表面的许多其他蛋白质结合。我们将测试卵子CD9相关蛋白是否与精子表面结合。基因敲除研究也表明，鸡蛋GPI锚定蛋白是融合所必需的。我们建议鉴定关键的GPL锚定蛋白(S)，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，我们询问它们的活动是如何协调的。我们认为，为了在融合中发挥作用，活跃的鸡蛋分子必须组织成脂质微域。因此，这些研究旨在结合分子和细胞水平，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.