Subcellular localization of beta1,4-galactosyltransferase on bull sperm and its function during sperm-egg...

The process of sperm-oocyte recognition is a complex interaction between the plasma membrane of sperm and the extracellular matrix of the oocyte. The best studied mammalian system is the mouse, in which sperm plasma membrane receptors recognize specific oligosaccharides on the egg coat glycoprotein ZP3. A well-defined ZP3 receptor on mouse sperm is beta1,4-galactosyltransferase (GalT). In this study, we investigated the possibility that GalT is present on bull sperm, and that it may participate during bovine sperm-oocyte binding. Using Western immunoblotting, bull sperm were found to have a protein of molecular weight similar to mouse GalT at approximately 60 kDa. Immunogold low voltage scanning electron microscopy reveals that GalT epitopes are confined to the anterior cap of fresh or capacitated bull sperm. To investigate the function of bovine sperm GalT, fresh bull sperm were pretreated with either preimmune or anti-GalT antibody and added to in vitro-matured bovine oocytes. Sperm exposed to preimmune serum fertilized 82.7% (153 of 185) of the oocytes, whereas sperm exposed to anti-GalT antiserum fertilized only 42.3% (202 of 478) of the oocytes. We determined whether the inhibition of fertilization resulted from a direct inhibition of sperm-oocyte binding. The number of sperm bound to eggs was determined by low voltage scanning electron microscopy following pretreatment with preimmune or anti-GalT antibody. An average of 25.3+/-2.2 (mean +/- SEM) sperm bound per half-oocyte when treated with preimmune serum. In contrast, exposure of sperm to anti-GalT antiserum significantly lowered (P<0.001) the frequency of sperm binding to 9.9+/-0.8 bound per half-oocyte. These results show that GalT is present on the anterior cap of the bovine sperm head, where it participates in fertilization by facilitating sperm-oocyte binding. The function of GalT in both the murine and bovine systems suggests that it may serve as a generalized gamete receptor in mammals.