The overall goal of this proposal is to examine the role of Src family protein tyrosine kinases in egg activation during fertilization, using the starfish egg as a model system. The mechanism of Src function during fertilization will be investigated by monitoring Ca release in eggs that are microinjected with full length Src protein. If injection of Src stimulates Ca release, then the Ca release pathway that has been stimulated will be determined by injecting reagents that inhibit phospholipase Cgamma activation. The next phase of this proposal will use molecular cloning to id4entify Src kinase family members in starfish eggs. Once cDNAs encoding Src kinases are confirmed by sequence analysis, we will use a functional assay to determine which protein products are involved in egg activation and develop specific antibodies to any candidate proteins. These antibodies will then be used to determine if a Src family kinase is activated at fertilization, and if there is a fertilization-dependent interaction between the Src kinase and phospholipase C. These studies are significant because they will provide insight into the basic biology of signal transduction during fertilization, which may eventually lead to application in contraception of the treatment of infertility.
