Abstract
During mammalian fertilization, sperm adhere to the extracellular coat of the egg, or zona pellucida, in a species-specific manner. In mouse, evidence suggests that sperm recognize and bind to specific oligosaccharide ligands within the zona pellucida glycoprotein, ZP3, via beta1,4-galactosyltransferase I (GalT I), a lectin-like receptor on the sperm surface. Although in vitro experiments using isolated gametes lend support to this model, recent in vivo studies of genetically altered mice question whether ZP3 and/or GalT I are solely responsible for sperm-egg binding. In this regard, sperm from GalT I-null mice bind poorly to ZP3 and fail to undergo a zona-induced acrosome reaction; however, they still bind to the ovulated egg coat in vitro. In this report, we characterize a novel ZP3- and GalT I-independent mechanism for sperm adhesion to the egg coat. Results show that the ovulated zona pellucida contains at least two distinct ligands for sperm binding: a ZP3-independent ligand that is peripherally associated with the egg coat and facilitates gamete adhesion; and a ZP3-dependent ligand that is present in the insoluble zona matrix and is recognized by sperm GalT I to facilitate acrosomal exocytosis. The ZP3-independent ligand is not a result of contamination by egg cortical granules, nor is it the mouse homolog of oviduct-specific glycoprotein. It behaves as a 250 kDa, WGA-reactive glycoprotein with a basic isoelectric point, distinguishing it from the acidic glycoproteins that form the insoluble matrix of the egg coat. When eluted from isoelectric focusing gels, the acidic matrix glycoproteins possess sperm-binding activity for wild-type sperm, but not for GalT I-null sperm, whereas the basic glycoprotein retains sperm-binding activity for both wild-type and GalT I-null sperm. Thus, GalT I-null sperm are able to resolve gamete recognition into at least two distinct binding events, leading to the characterization of a novel, peripherally associated, sperm-binding ligand on the ovulated zona pellucida.
Highlights
Gamete interaction is an obligatory event during mammalian fertilization when sperm bind to the egg in a species-specific manner
Results show that the ovulated zona pellucida contains at least two distinct ligands for sperm binding: a ZP3-independent ligand that is peripherally associated with the egg coat and facilitates gamete adhesion; and a
The ZP3-independent ligand is not a result of contamination by egg cortical granules, nor is it the mouse homolog of oviduct-specific glycoprotein. It behaves as a 250 kDa, wheat germ agglutinin (WGA)-reactive glycoprotein with a basic isoelectric point, distinguishing it from the acidic glycoproteins that form the insoluble matrix of the egg coat
Summary
Gamete interaction is an obligatory event during mammalian fertilization when sperm bind to the egg in a species-specific manner. Pioneering studies by Wassarman and colleagues suggest that in mouse, sperm bind to a specific class of oligosaccharides on ZP3 ( referred to as ZPC) (Spargo and Hope, 2003), one of the three glycoproteins that constitute the extracellular coat of the mouse egg, or zona pellucida. This was demonstrated by the ability of purified soluble ZP3, as well as its oligosaccharide chains, to competitively inhibit sperm-egg binding in vitro (Bleil and Wassarman, 1980a; Florman and Wassarman, 1985). Binding of ZP3 leads to sperm activation of both pertussis toxin (PTx)-sensitive heterotrimeric Gproteins as well as voltage-independent and -dependent cation channels that result in elevated pHi and Ca2+i, triggering acrosomal exocytosis (Arnoult et al, 1996; Endo et al, 1987; Endo et al, 1988; O’Toole et al, 2000)
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