High Ca²⁺-sensitivity and Ca²⁺ Oscillation-inducing Activity of PLC-zeta

Abstract

Sperm-specific phospholipase C-zeta (PLCζ) is known to induce intracellular Ca²⁺ oscillations and subsequent early embryonic development, when expressed in mouse eggs by injection of RNA encoding PLCζ. We addressed characteristics of recombinant mouse PLCζ protein generated by baculovirus/Sf9-cell expression system. PLCζ protein injected into mouse eggs induced repetitive Ca²⁺ spikes quite similar to those produced by injection of sperm extract, probably via produced InsP₃ and Ca²⁺ release from the ER. Recombinant PLCδ1 also induced Ca²⁺ oscillations, but 20-fold higher concentration was required. In the enzymatic assay, PLCζ exhibited a significant PtdInsP₂-hydrolyzing activity at [Ca²⁺] as low as 10 nM and had 70% maximal activity at 100 nM [Ca²⁺] that is usually the basal [Ca²⁺]i level of cells. On the other hand, the activity of PLCδ1 increased at [Ca²⁺] between 1 and 30 μM. EC₅₀ was 52 nM for PLCζ and 5.7 μM for PLCδ1: PLCζ has approximately 100-fold higher Ca²⁺-sensitivity than PLCδ1. Such high Ca²⁺-sensitivity of PLC activity as PLCζ that can be active in cells at the resting state is thought to be an appropriate characteristic as the sperm factor which is introduced into the egg cytoplasm upon sperm-egg fusion, first triggers Ca²⁺ release, and maintains Ca²⁺ oscillations. [Jpn J Physiol 54 Suppl:S70 (2004)]