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Journal of Cell Science, Vol 92, Issue 4 713-722, Copyright © 1989 by Company of Biologists

JOURNAL ARTICLES

Interaction of mouse sperm with purified sperm receptors covalently linked to silica beads

MH Vazquez, DM Phillips and PM Wassarman
Department of Cell and Developmental Biology, Roche Institute of Molecular Biology, Roche Research Center, Nutley, NJ 07110.

We describe a solid-phase assay that has permitted further analysis of zona pellucida glycoprotein, ZP3, as sperm receptor and acrosome reaction-inducer during fertilization in mice. The assay employs silica beads that contain epoxy groups to which purified, mouse oocyte ZP3 is covalently linked (ZP3-beads). ZP3-beads were characterized, e.g. by whole-mount autoradiography and flow cytofluorometry, incubated with capacitated mouse sperm under a variety of conditions, and the extent of sperm binding determined by light microscopy. Results of experiments presented suggest the following: (1) sperm bind specifically to ZP3-beads, but not to silica beads either exposed to 2-aminoethanol or derivatized with oocyte ZP2, fetuin or bovine serum albumin. (2) In nearly all cases, only one sperm binds per ZP3-bead and binding occurs via the sperm head. (3) The extent of sperm binding to ZP3-beads is dependent on ZP3 and sperm concentrations, as well as on incubation time and temperature. (4) Sperm binding to ZP3-beads is unaffected by antibodies directed against ZP3, but is inhibited in a reversible manner by treatment of ZP3-beads with galactose oxidase. (5) Only acrosome-intact sperm bind to ZP3-beads but, once bound, sperm can undergo the acrosome reaction, which results in their release from ZP3-beads. (6) Islet-activating protein and 3-quinuclidinyl benzilate, two inhibitors of the zona pellucida-induced acrosome reaction, prevent sperm bound to ZP3-beads from undergoing the acrosome reaction. These results confirm and extend previous studies of sperm-egg interaction in mice, and suggest that the solid-phase assay will be useful for both cellular and biochemical analyses of mammalian fertilization.


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© The Company of Biologists Ltd 1989