Requirement of a Src Family Kinase for Initiating Calcium Release at Fertilization in Starfish Eggs ,

doi:10.1074/jbc.274.41.29318

Requirement of a Src Family Kinase for Initiating Calcium Release at Fertilization in Starfish Eggs*,210

Abstract

Signal transduction leading to calcium release in echinoderm eggs at fertilization requires phospholipase Cγ-mediated production of inositol trisphosphate (IP₃), indicating that a tyrosine kinase is a likely upstream regulator. Because previous work has shown a fertilization-dependent association between the Src homology 2 (SH2) domains of phospholipase Cγ and a Src family kinase, we examined whether a Src family kinase was required for Ca²⁺ release at fertilization. To inhibit the function of kinases in this family, we injected starfish eggs with the SH2 domains of Src and Fyn kinases. This inhibited Ca²⁺ release in response to fertilization but not in response to injection of IP₃. We further established the specificity of the inhibition by showing that the SH2 domains of several other tyrosine kinases (Abl, Syk, and ZAP-70), and the SH3 domain of Src, were not inhibitory. Also, a point-mutated Src SH2 domain, which has reduced affinity for phosphotyrosine, was a correspondingly less effective inhibitor of fertilization-induced Ca²⁺ release. These results indicate that a Src family kinase, by way of its SH2 domain, links sperm-egg interaction to IP₃-mediated Ca²⁺ release at fertilization in starfish eggs.

Footnotes

↵* This work was supported by a summer research fellowship from the Marine Biological Laboratory, Woods Hole, MA (to A. F. G.), a grant from the Patrick and Catherine Weldon Donaghue Medical Research Foundation (to M. T.), a grant from the National Science Foundation (to K. R. F.), and grants from the National Institutes of Health (to D. J. C., K. R. F., and L. A. J.).The costs of publication of this article were defrayed in part by the payment of page charges. The article must therefore be hereby marked “advertisement” in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.
↵210 The on-line version of this article (available athttp://www.jbc.org) contains two Quicktime movies.
↵‖ To whom correspondence should be addressed: Dept. of Physiology, MC3505, University of Connecticut Health Center, Farmington, CT 06032. Tel.: 860-679-2661; Fax: 860-679-1661; E-mail: ljaffe@neuron.uchc.edu.
↵2 S. Roche, personal communication.
↵3 W. H. Kinsey, personal communication.
Abbreviations:

IP₃

inositol trisphosphate

SH2

Src homology 2

PLCγ

phospholipase Cγ
- Received April 28, 1999.
- Revision received June 8, 1999.
The American Society for Biochemistry and Molecular Biology, Inc.