Role of GPI-Anchored CD52 Glycoprotein in Sperm Function

描述(申请人提供)：CD52是一种GPI锚定糖蛋白，仅限于男性生殖道和淋巴细胞。尽管CD52的结构已被广泛研究，淋巴细胞CD52的功能也已被研究，但CD52在精子成熟和功能中的作用尚不清楚。在淋巴细胞表面，CD52与T细胞受体相关，并存在于含有酪氨酸激酶活性的蛋白质复合体中；含有CD52的复合体的交联会触发蛋白质酪氨酸磷酸化，诱导T细胞激活。因此，CD52在与TCR依赖的淋巴细胞激活相关的信号转导事件中被指示。我们假设精子表面CD52具有与淋巴细胞相似的信号转导相关功能和复合体形成特性。在精子中，CD52的功能作用已经使用S19，一种针对精子CD52的单抗(MAb)获得。S19Fab片段能抑制人精子与透明带的紧密结合，提示CD52本身或含有CD52的复合体参与了精子与透明带的相互作用。初步结果表明，人精子与双价S19单抗孵育可促进顶体反应(AR)，而单价S19 Fab不能促进顶体反应。这些结果表明，CD52聚集在精子AR的诱导中起作用，这是一种发生在精子与透明带结合的信号转导事件。总的来说，来自精子和淋巴细胞的研究结果表明，CD52在精子成熟和/或功能中发挥着功能作用。为了验证我们的假设，这项拟议的研究项目的具体目的是研究CD52在人类精子AR中的功能，并识别与精子CD52相关的蛋白质。CD52诱导的AR的获能依赖性和钙依赖性将被确定。将检查对百日咳毒素抑制的敏感性，以确定观察到的AR是否通过G蛋白介导的途径发生。将测定精子表面CD52交联后蛋白质酪氨酸磷酸化的变化。此外，与CD52相关的精子蛋白将被分离并进行显微测序。识别出的CD52相关蛋白将代表参与精子功能和精子-小带相互作用的信号转导事件的候选分子。这些研究将对精子CD52的功能和AR涉及的信号转导机制提供深入的认识。

DESCRIPTION (provided by applicant): CD52 is a GPI-anchored glycoprotein restricted to the male reproductive tract and lymphocytes. Although the structure of CD52 has been extensively characterized and the function of lymphocyte CD52 has been investigated, the role of CD52 in sperm maturation and function is unknown. On the lymphocyte surface, CD52 is associated with the T cell receptor and is present in protein complexes containing tyrosine kinase activity; cross linking of CD52-containing complexes were shown to trigger protein tyrosine phosphorylation and induce T cell activation. Thus, CD52 is indicated in signal transduction events associated with TCR-dependent activation of lymphocytes. We hypothesize that sperm-surface CD52 exhibits similar signal transduction-related functions and complex-forming properties as those identified in the lymphocyte. In spermatozoa, insights into the functional role of CD52 have been obtained using S19, a monoclonal antibody (mAb) specific for sperm CD52. S19 Fab fragments were shown to inhibit human sperm-zona pellucida tight binding implicating CD52 itself or a CD52-containing complex in sperm-zona interactions. Preliminary results indicate that incubation of human spermatozoa with the bivalent S19 mAb promotes the acrosome reaction (AR) while monovalent S19 Fab does not. These results suggest that CD52 aggregation has a role in induction of the sperm AR, a signal transduction-mediated event that occurs upon sperm-zona pellucida binding. Collectively, results from sperm and lymphocyte studies implicate a functional role for CD52 in sperm maturation and/or function. To test our hypotheses, the specific aims of the proposed research project are to investigate the function of CD52 in the human sperm AR and to identify proteins associated with sperm CD52. The capacitation- and calcium-dependence of the CD52-induced AR will be determined. Sensitivity to inhibition by pertussis toxin will be examined to determine whether the observed AR occurs via a G protein-mediated pathway. Changes in protein tyrosine phosphorylation upon cross linking of sperm- surface CD52 will be determined. Furthermore, CD52-associated sperm proteins will be isolated and microsequenced. The identified CD52-associated proteins will represent candidate molecules for signal transduction events involved in sperm function and for sperm-zona interactions. These studies will provide insights into the function of sperm CD52 and into the signal transduction mechanisms involved in the AR.