Within the first minute after sperm contact with a sea urchin or frog egg, a remarkable series of signals passes back and forth between sperm and egg plasma membranes. Contact with the fertilizing sperm causes ion channels to open in the egg plasma membrane, causing it to depolarize. This deplorization prevents other sperm from fertilizng the egg. Meanwhile, contact of the fertilizing sperm has set in motion a sequence of biochemical events which causes the egg to undergo exocytosis of its cortical vesicles and to begin development.
One specific aim of this proposal involves examining the pathway coupling sperm-egg interaction to ion channel opening and cortical vesicle exocytosis. We will examine the hypothesis that the fertilizing sperm activates a receptor in the egg plasma membrane, and that the receptor activates a G-protein, which stimulates PIP2 phosphodiesterase, leading to InsP3 production and Ca2+ release. Ca2+ release is proposed to cause ion channel opening and exocytosis by a pathway involving a Ca2+/calmodulin- dependent phosphatase. The methods will involve microinjection as well as biochemical approaches, including the use of monoclonal antibodies. The other specific aim involves investigating how a change in the egg's membrane potential regulates sperm-egg fusion. We will examine two hypotheses: first, we will examine the possibility that the sperm membrane contains a positively charged fusion protein which accounts for the voltage-dependence of sperm-egg fusion. Second, we will examine an alternate possibility, that the egg membrane contains a sperm receptor whose exposure to sperm on the external surface of the egg depends on the egg's membrane potential. The methods will involve cross-species fertilization, lipid bilayer techniques, and application of monoclonal antibodies and proteases to eggs at various voltages. The significance of this work to the study of human development is that the cell biological prinicples we discover in our work invertebrates and amphibians will form a basis for future studies of mammals. The basic science background may ultimately find applications to practical problems such as contraception and infertility.
Showing the most recent 10 out of 39 publications
