Molecular characteristics of horse phospholipase C zeta (PLC)

A sperm-specific phospholipase C (PLC), PLCzeta (PLC), is thought to underlie the initiation of calcium ([Ca2+]i) oscillations that induce egg activation in mammals. In large domestic species, only bovine, porcine and recently equine PLC have been cloned, and the physiological functions of these molecules have not been fully characterized. Here, we evaluated the physiological functions of equine PLC (ePLC) in mouse oocytes. ePLC was cloned from testis using RT-PCR. The expression of ePLC messenger RNA was confirmed in testis but not in other tissues. Microinjection of ePLC complementary RNA (cRNA) into mouse oocytes induced long-lasting [Ca2+]i oscillations, and most of the injected oocytes formed pronuclei (PN). The injection of cRNAs encoding horse, mouse, human and cow PLC into mouse oocytes showed that ePLC had the highest [Ca2+]i oscillation-inducing activity among the species tested. Mutation of D202R, which renders the protein inactive, abrogated the activity of ePLC. The nuclear translocation ability of ePLC was defective when expressed in mouse oocytes. Taken together, our findings show for the first time that ePLC has highest activity of the mammalian species studied to date. Our findings will be useful for the improvement of reproductive technologies in the horse.