Evidence That Fertilization Activates Starfish Eggs by Sequential Activation of a Src-like Kinase and Phospholipase Cγ

doi:10.1074/jbc.M001091200

Evidence That Fertilization Activates Starfish Eggs by Sequential Activation of a Src-like Kinase and Phospholipase Cγ*

From the ^‡Marine Biological Laboratory, Woods Hole, Massachusetts 02543, the ^§Department of Physiology, University of Connecticut Health Center, Farmington, Connecticut 06032, and the ^¶Department of Biological Structure, Biomolecular Structure Center, University of Washington, Seattle, Washington 98195

Abstract

Recent evidence has indicated a requirement for a Src family kinase in initiating Ca²⁺ release at fertilization in starfish eggs (Giusti, A. F., Carroll, D. J., Abassi, Y. A., Terasaki, M., Foltz, K. R., and Jaffe, L. A. (1999) J. Biol. Chem. 274, 29318–29322). We now show that injection of Src protein into starfish eggs initiates Ca²⁺ release and DNA synthesis, as occur at fertilization. These responses depend on the phosphorylation state of the Src protein; only the kinase active form is effective. Like Ca²⁺ release at fertilization, the Ca²⁺ release in response to Src protein injection is inhibited by prior injection of the SH2 domains of phospholipase Cγ. These findings support the conclusion that in starfish, sperm-egg interaction causes egg activation by sequential activation of a Src-like kinase and phospholipase Cγ. Injection of the SH2 domain of Src, which inhibits Ca²⁺ release at fertilization, does not inhibit Ca²⁺ release caused by Src protein injection. This indicates that the requirement for a Src SH2 domain interaction is upstream of Src activation in the pathway leading to Ca²⁺ release at fertilization.

Footnotes

↵* This work was supported by grants from the National Institutes of Health (to A. G. 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.
Published, JBC Papers in Press, March 22, 2000, DOI 10.1074/jbc.M001091200
‖ To whom correspondence should be addressed: Dept. of Physiology L5004, University of Connecticut Health Center, 263 Farmington Ave., Farmington, CT 06032. Tel.: 860-679-2661; Fax: 860-679-1661; E-mail: ljaffe@neuron.uchc.edu.
↵2 W. Xu, unpublished results.
Abbreviations:

IP₃

inositol 1,4,5-trisphosphate

PLCγ

phospholipase Cγ

SH2

Src homology 2

P⁻Src

Src protein that is completely unphosphorylated

PY530Src

Src that is phosphorylated at Tyr-530 and not at other sites
- Received February 9, 2000.
- Revision received March 20, 2000.
The American Society for Biochemistry and Molecular Biology, Inc.