A soluble phospholipase C (PLC) from boar sperm generates InsP3 and hence causes Ca2+ release when added to sea urchin egg homogenate. This PLC activity is associated with the ability of sperm extracts to cause Ca2+ oscillations in mammalian eggs following fractionation. A sperm PLC may, therefore, be responsible for causing the observed Ca2+ oscillations at fertilization. In the present study we have further characterized this boar sperm PLC activity using sea urchin egg homogenate. Consistent with a sperm PLC acting on egg PtdIns(4,5)P2, the ability of sperm extracts to release Ca2+ was blocked by preincubation with the PLC inhibitor U73122 or by the addition of neomycin to the homogenate. The Ca2+-releasing activity was also detectable in sperm from other species and in whole testis extracts. However, activity was not observed in extracts from other tissues. Moreover recombinant PLCβ1, -γ1, -γ2, -∆1, all of which had higher specific activities than boar sperm extracts, were not able to release Ca2+ in the sea urchin egg homogenate. In addition these PLCs were not able to cause Ca2+ oscillations following microinjection into mouse eggs. These results imply that the sperm PLC possesses distinct properties that allow it to hydrolyse PtdIns(4,5)P2 in eggs.
A screening campaign in sea urchin egg homogenate as a platform for discovering modulators of NAADP-dependent Ca2+ signaling in human cells
