Linda L. Runft (a), 1, David J. Carroll (b), 1, Jessica Gillett (a), 1, Andrew F. Giusti (c), Forest J. O'Neill (a) and Kathy R. Foltz (a)
1 These three authors contributed equally to this collaborative project.
Developmental Biology, 2004, 269: 220-236
Abstract
At fertilization, eggs undergo a cytoplasmic free Ca2+ rise, which is necessary for stimulating embryogenesis. In starfish eggs, studies using inhibitors designed against vertebrate proteins have shown that this Ca2+ rise requires an egg Src family kinase (SFK) that directly or indirectly activates phospholipase C- (PLC-) to produce IP3, which triggers Ca2+ release from the egg's endoplasmic reticulum (ER) [reviewed in Semin. Cell Dev. Biol. 12 (2001) 45]. To examine in more detail the endogenous factors in starfish eggs that are required for Ca2+ release at fertilization, an oocyte cDNA encoding PLC- was isolated from the starfish Asterina miniata. This cDNA, designated AmPLC-, encodes a protein with 49% identity to mammalian PLC-1. A 58-kDa Src family kinase interacted with recombinant AmPLC- Src homology 2 (SH2) domains in a specific, fertilization-responsive manner. Immunoprecipitations of sea urchin egg PLC- using an affinity-purified antibody directed against AmPLC- revealed fertilization-dependent phosphorylation of PLC-. Injecting starfish eggs with the tandem SH2 domains of AmPLC- (which inhibits PLC- activation) specifically inhibited Ca2+ release at fertilization. These results indicate that an endogenous starfish egg PLC- interacts with an egg SFK and mediates Ca2+ release at fertilization via a PLC- SH2 domain-mediated mechanism.
Author Keywords: Calcium; Fertilization; Phospholipase C-; Echinoderms
Corresponding author. Department of Molecular, Cellular and Developmental Biology and the Marine Science Institute, University of California-Santa Barbara, Bren 4L, Santa Barbara, CA 93106-9610. Fax: +1-805-893-8062.
(a) Department of Molecular, Cellular & Developmental Biology and the Marine Science Institute, University of California-Santa Barbara, Santa Barbara, CA 93106-9610, USA
(b) Department of Biological Sciences, Florida Tech, Melbourne, FL 32901, USA
(c) Gen-Probe, Inc., San Diego, CA 92121, USA | 
