PLCζ causes Ca2+ oscillations in mouse eggs by targeting intracellular and not plasma membrane PI(4,5)P2

Sperm-specific phospholipase C zeta (PLC zeta) activates embryo development by triggering intracellular Ca2+ oscillations in mammalian eggs indistinguishable from those at fertilization. Somatic PLC isozymes generate inositol 1,4,5-trisphophate-mediated Ca2+ release by hydrolyzing phosphatidylinositol 4,5-bisphosphate (PI(4,5)P-2) in the plasma membrane. Here we examine the subcellular source of PI(4,5)P-2 targeted by sperm PLC zeta in mouse eggs. By monitoring egg plasma membrane PI(4,5)P-2 with a green fluorescent protein-tagged PH domain, we show that PLC zeta effects minimal loss of PI(4,5)P-2 from the oolemma in contrast to control PLC delta 1, despite the much higher potency of PLC zeta in eliciting Ca2+ oscillations. Specific depletion of this PI(4,5)P-2 pool by plasma membrane targeting of an inositol polyphosphate-5-phosphatase (Inp54p) blocked PLC delta 1-mediated Ca2+ oscillations but not those stimulated by PLC zeta or sperm. Immunolocalization of PI(4,5)P-2, PLC zeta, and catalytically inactive PLC zeta (ciPLC zeta) revealed their colocalization to distinct vesicular structures inside the egg cortex. These vesicles displayed decreased PI(4,5)P-2 after PLC zeta injection. Targeted depletion of vesicular PI(4,5)P-2 by expression of ciPLC zeta-fused Inp54p inhibited the Ca2+ oscillations triggered by PLC zeta or sperm but failed to affect those mediated by PLC delta 1. In contrast to somatic PLCs, our data indicate that sperm PLC zeta induces Ca2+ mobilization by hydrolyzing internal PI(4,5)P-2 stores, suggesting that the mechanism of mammalian fertilization comprises a novel phosphoinositide signaling pathway.