Sperm surface galactosyltransferase activities during in vitro capacitation

Abstract

Studies using genetic and biochemical probes have suggested that mouse sperm surface galactosyltransferase may participate during fertilization by binding N-acetylglucosamine (GlcNAc) residues in the egg zona pellucida. In light of these results, [mouse] sperm surface galactosyltransferase activity during in vitro capacitation was examined to determine whether changes in enzymatic activity correlated with fertilizing ability. Surface galactosyltransferases on uncapacitated sperm are preferentially loaded with polyN-acetyllactosamine substrates. As a consequence of capacitation in Ca2+-containing medium, these polylactosaminyl substrates are spontaneously released from the sperm surface, thereby exposing the sperm galactosyltransferase for binding to the zona pellucida. Sperm capacitation can be mimicked, in the absence of Ca2+, either by washing sperm in Ca2+-free medium or by pretreating sperm with antiserum that reacts with the galactosyltransferase substrate. In both instances, sperm galactosylation of endogenous polylactosaminyl substrates is reduced, coincident with increased galactosylation of exogenous GlcNAc, and increased binding to the zona pellucida. Binding of capacitated sperm to the egg can be inhibited by pronase-digested high-MW polylactosaminyl glycosides extracted from epididymal fluids or from undifferentiated F9 embryonal carcinoma cells. These glycosides function as decapacitation factors when added back to in vitro fertilization assays. These glycoside decapacitation factors inhibit sperm-egg binding by competing for the sperm surface galactosyltransferase, since they are galactosylated by sperm in the presence of UDP[3H]galactose, and enzymatic removal of terminal GlcNAc residues reduces decapacitation factor competition. Conventional low-MW glycosides, isolated from either epididymal fluid or differentiated F9 cells, fail to inhibit capacitated sperm binding to the zona pellucida. A molecular mechanism for one aspect of sperm capacitation is defined, and why the removal of decapacitation factors is a necessary prerequisite for sperm binding to the zona pellucida is explained.