Abstract. The porcine zona pellucida (ZP) undergoes biochemical changes during the final phase of maturation prior to fertilization. The present study was conducted to elucidate whether the acidification of ZP glycoproteins during porcine oocyte maturation influences sperm-ZP interactions. Two-dimensional gel electrophoresis clearly demonstrated that ZP acidification occurred in accordance with the sialylation and sulfation of ZP glycoproteins in oocytes matured for 44 h. The increases in the incidences of sperm penetration and polyspermy with the progress of the IVM culture period were significantly suppressed by ZP desialylation on treatment with neuraminidase as a consequence of reductions in the number of sperm bound to ZPs and the acrosome reaction (AR) in ZP-bound sperm (P<0.05). In contrast, the blocking of ZP sulfation by NaClO 3 treatment during IVM markedly reduced the incidence of polyspermy with no inhibitory effect on penetration, but the number of sperm bound to ZPs and the rate of AR-inducing sperm were decreased to the same level as in desialylated oocytes. The results indicate that ZP sulfation influences sperm-ZP interactions in a ZP sialylationindependent manner. Moreover, sialylation and sulfation were not associated with a protective proteolytic modification of the ZP matrix before fertilization. These findings suggest that ZP acidification elicited by the sialylation and sulfation of ZP glycoproteins during oocyte maturation contributes to the porcine ZP acquiring the capacity to accept sperm.Key words: Acidification, Porcine oocytes, Sialylation, Sulfation, Zona pellucida (J. Reprod. Dev. 57: [744][745][746][747][748][749][750][751] 2011) A ll mammalian eggs are surrounded by a relatively thick, insoluble extracellular coat called the zona pellucida (ZP) [1][2][3]. During fertilization, the ZP mediates species-selective recognition between the oocyte and spermatozoon [4]. There is evidence that the sperm-ZP interaction is a carbohydrate-mediated event [5][6][7][8] that triggers a signal transduction pathway that results in the fenestration and fusion of the sperm plasma membrane and the outer acrosomal membrane (acrosome reaction, AR) [3].The porcine ZP is composed of three glycoprotein families, ZP1 (ZPA; 92 kDa), ZP3α (ZPB; 55 kDa) and ZP3β (ZPC; 55 kDa) [9,10]. ZP1 is split into two smaller molecules, ZP2 (69 kDa) and ZP4 (23 kDa), under reducing conditions [11]. Very recently, we reported that the increase in the amount of terminal N-acetylglucosamine (GlcNAc) residues in porcine ZP3 glycoproteins through new Nglycosylation for periods in excess of 20-24 h after meiotic maturation played a critical role in sperm-ZP interactions [12]. This new N-glycosylation is responsible for significant increases in the sperm penetration rate, the polyspermic fertilization rate, the number of sperm bound to ZPs and the number of AR-inducing sperm. These findings clearly imply that ZP glycoproteins undergo biochemical changes during oocyte maturation prior to fertilization. In this context, it is o...

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