Identification of a starfish egg PLC-gamma that regulates Ca2+ release at fertilization.
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-gamma (PLC-gamma) 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-gamma was isolated from the starfish Asterina miniata. This cDNA, designated AmPLC-gamma, encodes a protein with 49% identity to mammalian PLC-gamma1. A 58-kDa Src family kinase interacted with recombinant AmPLC-gamma Src homology 2 (SH2) domains in a specific, fertilization-responsive manner. Immunoprecipitations of sea urchin egg PLC-gamma using an affinity-purified antibody directed against AmPLC-gamma revealed fertilization-dependent phosphorylation of PLC-gamma. Injecting starfish eggs with the tandem SH2 domains of AmPLC-gamma (which inhibits PLC-gamma activation) specifically inhibited Ca2+ release at fertilization. These results indicate that an endogenous starfish egg PLC-gamma interacts with an egg SFK and mediates Ca2+ release at fertilization via a PLC-gamma SH2 domain-mediated mechanism.
Runft
Linda L
LL
Department of Molecular, Cellular & Developmental Biology and the Marine Science Institute, University of California-Santa Barbara, Santa Barbara, CA 93106-9610, USA.
Carroll
David J
DJ
Gillett
Jessica
J
Giusti
Andrew F
AF
O'Neill
Forest J
FJ
Foltz
Kathy R
KR
D000595:Amino Acid Sequence; D000818:Animals; D002118:Calcium / Q000378:metabolism*; D005306:Fertilization / Q000502:physiology*; D008969:Molecular Sequence Data; D010063:Ovum / Q000378:metabolism*; D051966:Phospholipase C gamma; D013215:Starfish / Q000378:metabolism*; D010738:Type C Phospholipases / Q000302:isolation & purification / Q000378:metabolism*; D018909:src Homology Domains
D010738:Type C Phospholipases; D051966:Phospholipase C gamma; D002118:Calcium