Gamete Recognition in Mice Depends on the Cleavage Status of an Egg’s Zona Pellucida Protein

Abstract

At fertilization, mouse sperm bind to the zona pellucida (which consists of glycoproteins ZP1, ZP2, and ZP3) that surrounds eggs. A ZP2 cleavage model of gamete recognition requires intact ZP2, and a glycan release model postulates that zona glycans are ligands for sperm. These two models were tested by replacing endogenous protein with ZP2 that cannot be cleaved (Zp2Mut) or with ZP3 lacking implicated O glycans (Zp3Mut). Sperm bound to two-cell Zp2Mut embryos despite fertilization and cortical granule exocytosis. Contrary to prediction, sperm fertilized Zp3Mut eggs. Sperm at the surface of the zona pellucida remained acrosome-intact for more than 2 hours and were displaced by additional sperm. These data indicate that sperm-egg recognition depends on the cleavage status of ZP2 and that binding at the surface of the zona is not sufficient to induce sperm acrosome exocytosis.

Protease activity involvement in the passage of mammalian sperm through the zona pellucida

“…This model comes from results where mouse ZP1 and ZP3 has been replaced by their human homologus in transgenic mice. Nonetheless, the human proteins were able to form a stable and well-organized zona pellucida at the macroscopic level; the zona is not functionally active, as with the mouse proteins (Gahlay et al, 2010;Rankin et al, 2003). Thus, the simplest explanation for our results is that acrosin treatment disrupts and/or disorganizes the supramolecular structure of hamster zona pellucida, (mainly the carbohydrates moieties), making the binding sites for spermatozoa unavailable.…”

“…However, the fertilization rate decreased under this condition. Recently it has been proposed that a specifi c supramolecular structure of mammalian zona pellucida is important to achieve fertilization and polyspermy block in mammals (Gahlay et al, 2010;Rankin et al, 2003). This model comes from results where mouse ZP1 and ZP3 has been replaced by their human homologus in transgenic mice.…”

“…Acrosome-intact spermatozoa bear a non-active (pro) acrosin, which is activated upon acrosome reaction (Barros et al, 1996;Codelia et al, 2005;Cortes et al, 2006;Moreno and Alvarado, 2006). Different studies have shown that acrosin can digest zona pellucida glycoproteins and it is believed that its enzyme activity is involved in the cleavage of ZP2 during fertilization (Florman and Wassarman, 1985;Gahlay et al, 2010). However, there is no experimental evidence that zonaintact eggs treated with acrosin, or another serine-protease, are impervious to fertilization.…”

“…In order to penetrate the zona pellucida, as one hypothesis states, acrosome-intact sperm recognizes and binds with a single O-linked oligosaccharides chain, which is probably ZP3 (Florman and Wassarman, 1985;Wassarman et al, 2001). On the other hand, a ZP2 cleavage model has recently been proposed that argues that rather than sperm binding to a single ligand, sperm binding is supported by a three-dimensional zona structure (Gahlay et al, 2010;Rankin et al, 2003). This model is predicated on the cleavage status of ZP2, rendering the zona pellucida either permissive (uncleaved ZP2) or non-permissive (cleaved ZP2) to account for sperm binding to the zona pellucida surrounding eggs, but not to those surrounding two-cell embryos (Gahlay et al, 2010;Rankin et al, 2003).…”

“…On the other hand, a ZP2 cleavage model has recently been proposed that argues that rather than sperm binding to a single ligand, sperm binding is supported by a three-dimensional zona structure (Gahlay et al, 2010;Rankin et al, 2003). This model is predicated on the cleavage status of ZP2, rendering the zona pellucida either permissive (uncleaved ZP2) or non-permissive (cleaved ZP2) to account for sperm binding to the zona pellucida surrounding eggs, but not to those surrounding two-cell embryos (Gahlay et al, 2010;Rankin et al, 2003). ZP2 has also been proposed as a binding partner of acrosome-reacted sperm (Bleil et al, 1988;Howes and Jones, 2002).…”