Zonadhesin: Characterization, localization, and zona pellucida binding

Zonadhesin is a multiple-domain transmembrane protein that is believed to f unction as a sperm-zona pellucida binding protein. In this study we sequenc ed zonadhesin from rabbit testis and analyzed its processing, expression, l ocalization, and zona pellucida binding. We show that the precursor protein occurs exclusively in the testis and that proteolytic processing results i n the formation of three fragments: p43 (D1 domain), p97 (D2-D4 domains), a nd p58 (D4 domain-C-terminal). In mature spermatozoa the p43 and p97 fragme nts exist as disulfide-bonded dimers. During spermatogenesis, synthesis of zonadhesin mRNA chiefly occurs in primary spermatocytes, whereas the protei n is abundant in both Sertoli cells and spermatids. In spermatozoa the prot ein is localized exclusively to the anterior acrosome but is not available for binding antibody on live spermatozoa. Once the acrosome reaction is ind uced, zonadhesin is lost from the spermatozoon, but remains with the acroso mal shroud. We show that recombinant D4 domain can bind zona pellucida, and we propose that zonadhesin functions after the acrosome reaction has been initiated to bind the acrosomal shroud to the zona pellucida.