The long-term goal of this new R15 (AREA) proposal is to understand the mechanism(s) by which egg and sperm recognize and interact with each other in the tunicate Ascidia cevatodes and how this interaction leads to fertilization. During fertilization or ionophore-induced activation, ascidian eggs release N-acetylglucosaminidase activity in the surrounding seawater which prevent polyspermy. The working hypotheses are that there is a link between ascidian sperm-egg binding and subsequent events of egg activation including the release of N-acetylglucosaminidase. The P.I. proposes to identify and characterize the molecular components involved in sperm-egg interaction, and to investigate the mechanism(s) by which these molecules lead to egg activation. In particular, N-acetylglucosaminidase from egg, sperm and follicle cells will be isolated and characterized using kinetic analysis, physical characterization and competitive binding assays. In addition, Dr. Goode proposes to explore the putative role of tyrosine kinase and an ecto-phospholipase in the release of the N-acetylglucosaminidase using immunoblotting techniques. The overall aim of the studies proposed in this AREA application is to gain information regarding the mechanism by which ascidians prevent polyspermy, a process which occurs in all organisms including mammals.
