EXOCYTOTIC SIGNALING THROUGH SPERM RECEPTORS FOR EGGS

我们的长期目标是对受精的分子基础有一个更清晰的基本认识，以便控制受精过程，促进受精或阻止受精。ZP 3是哺乳动物卵被中的一种蛋白质，它结合精子并刺激顶体反应，顶体囊泡的释放是精子穿透卵被所必需的。 尽管取得了这一进展，但精子上ZP 3受体的身份一直难以捉摸。 一个强有力的候选者是β 1，4-半乳糖基转移酶，这种酶首先在体细胞的高尔基体中发现，但后来在精子表面发现。β 1，4-半乳糖基转移酶结合ZP 3，但不结合其他蛋壳蛋白。 β 1，4-半乳糖基转移酶除了在配子粘附中起作用外，还能触发精子的顶体反应。 顶体反应可以用β 1，4-半乳糖基转移酶的抗体刺激，所述抗体充当ZP 3的模拟物以结合和交联β 1，4-半乳糖基转移酶。 有证据表明，结合β 1，4-半乳糖基转移酶刺激精子内至少一些与ZP 3刺激相同的信号系统，从而提出β 1，4-半乳糖基转移酶可能是顶体反应的主要ZP 3受体信号。 β 1，4-半乳糖基转移酶基因被定向删除的小鼠，虽然能生育，但产生的精子顶体反应和穿透卵被的能力严重受损。 在这个提议中，信号系统被激活的β 1，4-半乳糖基转移酶将被研究。 目标是确定结合β 1，4-半乳糖基转移酶特异性地是否可以完全复制由ZP 3结合激活的所有已鉴定的信号传导系统，研究β 1，4-半乳糖基转移酶突变体以鉴定信号传导所需的结构特征，并鉴定与胞质溶胶中发现的β 1，4-半乳糖基转移酶部分相互作用的蛋白质。 这些研究将阐明顶体反应（受精过程中的一个必要步骤）中卵被的精子受体的作用。

Our long-range objective is to develop a clearer fundamental understanding of the molecular basis of fertilization so that the process can be controlled to either promote fertilization or block it. Much progress has been made to identify ZP3, the protein in the mammalian egg coat that binds sperm and stimulates the acrosome reaction, the release of the acrosomal vesicle that must occur for sperm to penetrate the egg coat. Despite this progress, the identity of the ZP3 receptor on sperm has been elusive. One strong candidate is beta1,4-galactosyltransferase, an enzyme first discovered in the Golgi apparatus of somatic cells but later found on the surface of sperm. Beta1,4-Galactosyltransferase binds ZP3 but not other egg coat proteins. In addition to its role in gamete adhesion, beta1,4- galactosyltransferase acts to trigger the acrosome reaction in sperm. The acrosome reaction can be stimulated with antibodies to beta1,4-galactosyltransferase that act as mimics of ZP3 to bind and crosslink beta1,4-galactosyltransferase. There is evidence that binding beta1,4-galactosyltransferase stimulates at least some of the same signaling systems inside sperm that ZP3 stimulates, leading to the proposal that beta1,4- galactosyltransferase may be the major ZP3 receptor signaling the acrosome reaction. Mice with a targeted deletion of the beta1,4-galactosyltransferase gene, although fertile, produce sperm with severely compromised ability to acrosome react and penetrate the egg coat. In this proposal, the signaling systems that are activated by beta1,4-galactosyltransferase will be studied. The goals are to determine if binding beta1,4-galactosyltransferase specifically can completely reproduce all of the identified signaling systems activated by ZP3 binding, to study beta1,4-galactosyltransferase mutants to identify structural features necessary for signaling, and to identify proteins that interact with the portion of beta1,4- galactosyltransferase found in the cytosol. These studies will clarify the role of a sperm receptor for the egg coat in the acrosome reaction, a necessary step in the process of fertilization.