Role of sialic acid in bovine sperm–zona pellucida binding

Abstract

Sperm binding activity has been detected in zona pellucida (ZP) glycoproteins and it is generally accepted that this activity resides in the carbohydrate moieties. In the present study we aim to identify some of the specific carbohydrate molecules involved in the bovine sperm-ZP interaction. We performed sperm binding competition assays, in vitro fecundation (IVF) in combination with different lectins, antibodies and neuraminidase digestion, and chemical and cytochemical analysis of the bovine ZP. Both MAA lectin recognising alpha-2,3-linked sialic acid and neuraminidase from Salmonella typhimurium with catalytic activity for alpha-2,3-linked sialic acid, demonstrated a high inhibitory effect on the sperm-ZP binding and oocyte penetration. These results suggest that bovine sperm-ZP binding is mediated by alpha-2,3-linked sialic acid. Experiments with trisaccharides (sialyllactose, 3'-sialyllactosamine and 6'-sialyllactosamine) and glycoproteins (fetuin and asialofetuin) corroborated this and suggest that at least the sequence Neu5Ac(alpha2-3)Gal(beta1-4)GlcNAc is involved in the sperm-ZP interaction. Moreover, these results indicate the presence of a sperm plasma membrane specific protein for the sialic acid. Chemical analysis revealed that bovine ZP glycoproteins contain mainly Neu5Ac (84.5%) and Neu5GC (15.5%). These two types of sialic acid residues are probably linked to Galbeta1,4GlcNAc and GalNAc by alpha-2,3- and alpha-2,6-linkages, respectively, as demonstrated by lectin cytochemical analysis. The use of a neuraminidase inhibitor resulted in an increased number of spermatozoa bound to the ZP and penetrating the oocyte. From this last result we hypothesize that a neuraminidase from cortical granules would probably participate in the block to polyspermy by removing sialic acid from the ZP.

Role of Acidification Elicited by Sialylation and Sulfation of Zona Glycoproteins During Oocyte Maturation in Porcine Sperm-zona Pellucida Interactions

“…These findings indicate that the sialylation of ZP glycoproteins during oocyte maturation is associated with sperm-ZP interactions. A similar phenomenon is observed in bovine oocytes, and the number of sperm bound to ZPs and the rate of penetration were remarkably decreased in oocytes treated with neuraminidase compared with untreated oocytes [19]. In our preliminary study, SSA (recognizing the α-2,6-linked sialic acid) and MAL II (maackia amurensis lectin II; recognizing the α-2,3-linked sialic acid residues) lectins added to IVF medium strongly diminished the number of sperm bound to ZPs in matured oocytes (30.9 ± 1.4 and 35.6 ± 2.0, respectively) compared with the control (63.7 ± 1.7; unpublished data).…”

“…Porcine ZP3 glycoproteins are highly heterogeneous mainly as a results of the differences in the amount of sulfated N-acetyllactosamine and sialic acid in the acidic chains [16] and have been shown to be required for the binding of sperm to the intact ZP leading to completion of AR [17]. It is reported that the sialic acid and sulfate of ZP glycoproteins are physiologically involved in regulating the binding of sperm to ZPs and the partial activation of proacrosin to form α-and β-acrosin in mouse [18], bovine [19] and porcine [20] oocytes. Contrary to these reports, Liu et al [21] suggest that neither the sulfate nor sialic acid present on mouse ZP3 oligosaccharides is directly involved in sperm receptor or AR-inducing activity.…”

Determination of glycosidase activity in porcine oviductal fluid at the different phases of the estrous cycle

“…Therefore, the presence of active glycosidases at a neutral pH in the OF could have potential roles in different events related to sperm reservoir formation, sperm capacitation, final oocyte maturation, gamete interaction, and early embryo development. It has already been proposed that glycosidases could be involved in the control of polyspermy (Miller et al 1993, Velasquez et al 2007, sperm-oviductal epithelial cells interaction (Lefebvre et al 1997), sperm-zona pellucida binding (Miller et al 1993, Matsumoto et al 2002, Venditti et al 2007, sperm capacitation (Taitzoglou et al 2007), or dispersion of cumulus cells (Takada et al 1994). All these events take place in the oviduct and have been the aim of studies to increase our knowledge about the molecular basis of reproductive functions, but, surprisingly, we lack of precise studies about the activities of the different glycosidases in the OF.…”

Improving bovine semen diluents: insights from the male and female reproductive tracts, and the potential relevance of cervical mucins

“…However, release of this glycoprotein occurs during capacitation and this could expose binding sites required for other functionally relevant interactions at the appropriate time and location. This is a plausible scenario for bovines given that sialic acid recognition seems to be important for BOGP binding, and is also involved in ZP binding (Velasquez et al, 2007). Sperm must still negotiate a mucin-rich environment in the oviduct before reaching these sites.…”

“…To the best of our knowledge, a similar influence of cervical mucin on bovine sperm has not yet been demonstrated. However, it is plausible that bovine sperm could interact specifically with mucin glycans as bovine sperm have been shown to have affinity for such sugars (Lefebvre et al, 1997;Velasquez et al, 2007). Satoh et al (1995) deduced that mucin-type oligosaccharides with terminally exposed sialic acid residues could exist on BOGP, as affinity for WGA was high.…”