Fertilization: Beginning a New Organism
In sea urchin eggs, IP3 is formed initially at the site of sperm entry and can be detected within seconds of sperm-egg attachment. Inhibiting IP3 synthesis prevents Ca2+ release (Lee and Shen 1998; Carroll et al. 2000), whereas injected IP3 can release sequestered Ca2+ and lead to cortical granule exocytosis (Whitaker and Irvine 1984; Busa et al. 1985). Moreover, these IP3-mediated effects can be thwarted by pre-injecting the egg with calcium-chelating agents (Turner et al. 1986).
IP3-responsive calcium channels have been found in the egg endoplasmic reticulum. The IP3 formed at the site of sperm entry is thought to bind to IP3 receptors in these calcium channels, effecting a local release of Ca2+ (Ferris et al. 1989; Furuichi et al. 1989). Once released, Ca2+ can diffuse directly, or it can facilitate the release of more Ca2+ by binding to calcium-triggered calcium-release receptors, also located in the cortical endoplasmic reticulum (McPherson et al. 1992). These receptors release stored Ca2+ when they bind Ca2+, so binding Ca2+ releases more Ca2+, which binds to more receptors, and so on. The resulting wave of calcium release is propagated throughout the cell, starting at the point of sperm entry. The cortical granules, which fuse with the cell membrane in the presence of high Ca2+ concentrations, respond with a wave of exocytosis that follows the calcium wave. Mohri and colleagues (1995) have shown that IP3-released Ca2+ is both necessary and sufficient for initiating the wave of calcium release.
PHOSPHOLIPASE C: THE GENERATOR OF IP3 If IP3 is necessary for Ca2+ release and phospholipase C is required in order to generate IP3, the question then becomes, what activates PLC? This question has not been easy to address since (1) there are numerous types of PLC that (2) can be activated through different pathways, and (3) different species use different mechanisms to activate PLC. Results from studies of sea urchin eggs suggest that the active PLC in echinoderms is a member of the γ (gamma) family of PLCs (Carroll et al. 1997, 1999; Shearer et al. 1999). Inhibitors that specifically block PLCγ inhibit IP3 production as well as Ca2+ release. Moreover, these inhibitors can be circumvented by microinjecting IP3 into the egg. How PLCγ is activated by sperm is still a matter of controversy, although inhibitor studies have shown that membrane-bound kinases (Src kinases) and GTP-binding proteins play critical roles; Kinsey and Shen 2000; Giusti et al. 2003; Voronina and Wessel 2003, 2004; Townley et al. 2009). One possibility is that NAADP brought in by the sperm to initiate electrical depolarization also activates the enzyme cascade leading to IP3 production and calcium release (Churchill et al. 2003; Morgan and Galione 2007).