We will use gametes of the sea urchin, Stronglyocentrotus purpuratus, as a model system to analyze biochemical aspects of the cortical and acrosome reactions, processes that are common to many fertilization systems. In studying the cortical reaction and the modification of the egg coat that accompanies it we will extend our results on the role of a peroxidative system that hardens the fertilization membrane. We shall characterize the ovoperoxidase released from the cortical granules and to be inserted into the fertilization membrane, and we will study the specific assembly of that structure, as well as its crosslinking in vitro. We will explore the mechanism of generation of activated oxygen species produced in the oxidative burst that act as substrates for ovoperoxidase and how the burst is regulated. We shall see whether a vesicular system exists in eggs for uptake and release of Ca2 ion, analogous to the sarcoplasmic reticulum of muscle, and whether it is involved in the Ca2 ion-mediated cortical reaction. We will pursue the ionic mechanism of triggering the acrosome reaction of sperm, by examining whether elevations in intracellular pH and calcium are the two steps that are necessary and sufficient for triggering the reaction. We will examine the mechanism of interaction of individual jelly components with sperm, to see whether distinct components mediate specific ion fluxes. These experiments should provide insights into molecular mechanisms of several important events in fertilization and thereby indicate novel targets for the control of fertility.

National Institute of Health (NIH)
National Institute of General Medical Sciences (NIGMS)
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Reproductive Biology Study Section (REB)
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University of Washington
Schools of Medicine
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Quest, A F; Chadwick, J K; Wothe, D D et al. (1992) Myristoylation of flagellar creatine kinase in the sperm phosphocreatine shuttle is linked to its membrane association properties. J Biol Chem 267:15080-5
Somers, C E; Shapiro, B M (1991) Functional domains of proteoliaisin, the adhesive protein that orchestrates fertilization envelope assembly. J Biol Chem 266:16870-5
Wothe, D D; Charbonneau, H; Shapiro, B M (1990) The phosphocreatine shuttle of sea urchin sperm: flagellar creatine kinase resulted from a gene triplication. Proc Natl Acad Sci U S A 87:5203-7
Tombes, R M; Shapiro, B M (1989) Energy transport and cell polarity: relationship of phosphagen kinase activity to sperm function. J Exp Zool 251:82-90
Ratto, A; Shapiro, B M; Christen, R (1989) Phosphagen kinase evolution. Expression in echinoderms. Eur J Biochem 186:195-203
Somers, C E; Battaglia, D E; Shapiro, B M (1989) Localization and developmental fate of ovoperoxidase and proteoliaisin, two proteins involved in fertilization envelope assembly. Dev Biol 131:226-35
Battaglia, D E; Shapiro, B M (1988) Hierarchies of protein cross-linking in the extracellular matrix: involvement of an egg surface transglutaminase in early stages of fertilization envelope assembly. J Cell Biol 107:2447-54
Turner, E; Hager, L J; Shapiro, B M (1988) Ovothiol replaces glutathione peroxidase as a hydrogen peroxide scavenger in sea urchin eggs. Science 242:939-41
Toowicharanont, P; Shapiro, B M (1988) Regional differentiation of the sea urchin sperm plasma membrane. J Biol Chem 263:6877-83
Shapiro, B M; Turner, E (1988) Oxidative stress and the role of novel thiol compounds at fertilization. Biofactors 1:85-8

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