Impaired sperm fertilizing ability in mice lacking Cysteine-RIch Secretory Protein 1 (CRISP1)

DA ROS VG, MALDERA JA, WILLIS WD, COHEN DJ, GOULDING EH, GELMAN DM, RUBINSTEIN M, EDDY EM, CUASNICU PS

Albuquerque, Estados Unidos

Congreso; 33rd Annual American Society of Andrology Meeting; 2008

American Society of Andrology

Mammalian fertilization is a complex multi-step process mediated by different molecules present on both gametes. Epididymal protein CRISP1, the first described member of the Cysteine RIch Secretory Protein (CRISP) family, was identified by our laboratory and postulated to participate in sperm-zona pellucida (ZP) interaction and gamete fusion by binding to egg-complementary sites. To elucidate the functional role of CRISP1 in vivo, we disrupted the Crisp1 gene and evaluated the effect on animal fertility and several sperm parameters. In vivo breeding revealed that male and female Crisp1-/- animals exhibited no differences in fertility compared to controls. In vitro analysis revealed that sperm motility and the ability to undergo a spontaneous or progesterone-induced acrosome reaction were not affected in Crisp1-/- mice whereas the level of capacitation-induced tyrosine phosphorylation was lower than in controls. In vitro fertilization assays showed that Crisp1-/- sperm exhibited a significantly reduced ability to penetrate both ZP-intact (p=0.0002) and ZP-free eggs (p=0.0001). Furthermore, when ZP-free eggs were inseminated with both Crisp1+/+ and Crisp1-/- sperm in a competitive fertilization assay, the mutant sperm exhibited a greater disadvantage in their fusion ability compared to that observed in a non-competitive situation. Finally, the finding that the fusion ability of Crisp1-/- sperm was further inhibited by the presence of CRISP1 or CRISP2 but not CRISP4 during gamete co-incubation, suggests that CRISP2 cooperates with CRISP1 during fertilization and might compensate for its lack in the mutant mice. Together, these results revealed that although not essential for animal fertility, CRISP1 is definitively a player in the fertilization system. To our knowledge, this is the first knockout mice generated for a CRISP protein. This information will contribute to a better understanding of the molecular mechanisms underlying mammalian fertilization and might have important functional implications for other members of the widely distributed and evolutionarily conserved CRISP protein family.