The zona pellucida is an extracellular coat that surrounds all mammalian eggs. Sperm must penetrate the zona pellucida in order to reach and fuse with the plasma membrane of unfertilized eggs. Penetration is accomplished by a sequence of events involving both egg and sperm. First, sperm must bind to the outer margin of the zona pellucida. Such binding is mediated in a relatively species-specific manner by "sperm receptors" in the zona pellucida. Second, sperm must undergo the "acrosome reaction", a membrane fusion event, in order to traverse the zona pellucida. Here we review results from our own laboratory which demonstrate that, during the course of sperm-egg interaction in mice, zona pellucida glycoprotein ZP3 serves as both receptor for sperm and inducer of the acrosome reaction. Furthermore, we review evidence from our laboratory indicating that the sperm receptor activity of ZP3 is dependent only on its 0-linked carbohydrate components, whereas acrosome reaction-inducing activity is dependent on the polypeptide portion of ZP3 as well.

Fertilization is the process by which sperm and eggs unite to form a zygote, the true beginning of a new individual. In mammals, sperm first make contact with eggs at the surface of the egg's extracellular coat, or zona pellucida (Fig. 1a). This contact can lead to binding of sperm to eggs via species-specific sperm receptors present in the zona pellucida. Bound sperm then undergo changes that enable them to penetrate the zona pellucida and to fuse with the egg's plasma membrane. This fusion results in activation of eggs, and development of the organism ensues (for review, see Gwatkin 1977; Yanagimachi 1981; Bedford 1982; Wassarman 1983a; Wassarman et al. 1984a).

During their growth phase, mouse oocytes synthesize and secrete three different glycoproteins, called ZP1, 2 and 3, that constitute the extracellular coat, or zona pellucida, of the oocyte. One of these glycoproteins, ZP3, exhibits properties expected for a sperm receptor. We have now used rabbit antisera that recognize ZP3 to immunoprecipitate [35S]methionine‐labeled, intracellular precursors of this glycoprotein from growing oocytes cultured in vitro in the presence or absence of tunicamycin, a drug that prevents addition of N‐linked oligosaccharides to nascent polypeptide chains. Electrophoretic analyses of these immunoprecipitates, as well as of immunoprecipitates digested with endo‐beta‐N‐acetylglucosaminidase H (Endo H), indicate that ZP3 is synthesized as a 44,000 mol. wt. polypeptide chain to which either three or four high‐mannose‐type oligosaccharides are added, resulting in 53,000 and 56,000 mol. wt. ZP3 precursors, respectively. The latter species are converted to mature ZP3 (mol. wt. approximately 80,000) by processing of the high‐mannose‐type oligosaccharides (Endo H‐sensitive) to complex‐type oligosaccharides (Endo H‐insensitive) prior to ZP3 secretion. The evidence presented reveals that the extreme heterogeneity of mature ZP3, with respect to both mol. wt. and isoelectric point, is partly a consequence of the N‐linked oligosaccharides and not the polypeptide chain itself.

An antiserum directed specifically against ZP2, the major glycoprotein of the mouse egg's extracellular coat (zona pellucida), has been used to immunoprecipitate intracellular precursors of ZP2 that were synthesized by growing mouse oocytes cultured in vitro. Pulse-chase experiments revealed that the immediate precursor of mature, 120 kilodalton (kd) ZP2 is a 91 kd species that unlike mature ZP2, is sensitive to digestion by endo-β-N-acetylglucosaminidase H (Endo H) and is converted by the endoglycosidase into an 81 kd species. An 81 kd species is only found intracellularly when growing oocytes are cultured in the presence of tunicamycin. These results suggest that ZP2 is synthesized as an 81 kd polypeptide chain that is first “core”-glycosylated at asparagine residues with high-mannose-type oligosaccharides, giving rise to a 91 kd intermediate (Endo H-sensitive), and then processed to complextype oligosaccharides prior to secretion as mature, 120 kd ZP2 (Endo H-insensitive). Furthermore, electrophoretic analyses of mature ZP2, ZP2 precursor (91 kd) and Endo H-treated ZP2 precursor (81 kd) suggest that there are six N-linked oligosac-charides per molecule and that the extreme heterogeneity of mature ZP2 is a consequence of the oligosaccharides and not the polypeptide chain itself.