My laboratory is studying the molecular basis of fertilization in the mouse. Using a variety of biochemical, immunological and genetic probes, we have recently shown that initial gamete recognition between the sperm surface and the egg zona pellucida is mediated, at least in part, by the binding of a sperm surface enzyme, galactosyltransferase (GalTase) to its complementary substrate in the zona pellucida. GalTase is present on the cell surface throughout all stages of spermatogenesis, during which time it becomes redistributed from an initially diffuse and uniform distribution on primary spermatocytes to its final location on mature sperm overlying a discrete portion of the intact acrosome. Before sperm are able to bind the zona pellucida, they must first release a large molecular weight competitive GalTase substrate, which is occupying the zona binding site. Sperm binding to the zona pellucida induces the completion of the acrosome reaction, and recent findings show that despite the loss of the relevant plasma membrane domain, the GalTase is redistributed to the lateral sperm surface, presumably for some subsequent function. In this proposal we will more rigorously define the association of the GalTase with the sperm surface, purify the enzyme to apparent homogeneity and characterize its substrate specificity. The complementary substrate in the zona pellucida for the sperm GalTase will be identified and compared to the zona receptor for sperm identified by other workers. We will determine if fertilization inactivates the zona substrate for the sperm GalTase, leading to the zona block to polyspermy. Experiments will determine if the GalTase, leading to the zona block to polyspermy. Experiments will determine if the GalTase on the surface of early spermatogenic cells participates during germ cell adhesion to the Sertoli epithelium. Finally, we will examine some possible functions of the GalTase redistributed on acrosome- reacted sperm following initial sperm-zona binding. These experiments will give us more insight into the molecular nature of mouse gamete receptors, how they are modified during fertilization, and what other possible functions they may serve both before and after initial sperm-egg binding.
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