Evidence for the Regulation of Glycosylation of Golden Hamster (Mesocricetus auratus) Oviductin During the Estrous Cycle1

Abstract

The oviduct contributes to the reproductive environment by secreting various factors, including a family of glycoproteins termed oviductins. Although many studies have demonstrated that ovarian hormones modulate oviductin gene expression in several mammalian species, there has been controversy surrounding the regulation of golden hamster oviductin. The current study was undertaken to investigate the transcriptional and translational modifications of hamster oviductin during the estrous cycle. First, we verified that hamster oviductin mRNA expression remains constant throughout the estrous cycle by semiquantitative reverse transcription polymerase chain reaction. We then developed a polyclonal antibody against recombinant hamster oviductin (rhaOvm). The anti-rhaOvm antibody was subsequently used in conjunction with quantitative immunocytochemistry to investigate the oviductin levels in the hamster oviduct during the estrous cycle. Quantification of immunolabeling revealed a high, consistent level of glycoprotein throughout the estrous cycle. Therefore, it appears that the production of oviductin is not regulated differentially during the estrous cycle. Size variations in hamster oviductin expression were also investigated by Western blot analysis. The oviduct contains several forms of oviductin at each stage of the estrous cycle, the native glycosylated form(s) of 160-350 kDa, and several precursor forms of 70-100 kDa. Although variations in the intensities of the polydispersed band were not evident during the estrous cycle, additional bands ranging from 90 to 100 kDa were detected in the estrus, metestrus, and diestrus 1 stages. The results from the present investigations suggest that whereas ovarian hormones do not appear to influence the hamster oviductin mRNA and protein expressions, glycosylation of hamster oviductin appears to be differentially regulated during the estrous cycle.

The Common and Species-Specific Roles of Oviductal Proteins in Mammalian Fertilization and Embryo Development

“…At least in ungulates, oviductin increases the ZP's resistance to proteolysis, and this ZP hardening might be of crucial importance to warrant monospermic fertilization or sperm selection. The presence of different isoforms of oviductin and different levels of glycosylation of oviductin has been reported (McBride et al 2004). Studies using recombinant oviductins in different forms would disclose the molecular basis of such physiological differences.…”

What controls polyspermy in mammals, the oviduct or the oocyte?

“…Most of the carbohydrates are present in O‐linked chains but N‐linked chains have also been detected (Malette & Bleau, 1993). Glycosylation differences detected over the oestrous cycle could be responsible for a different biological role of oviductal‐secreted glycoproteins (McBride et al , 2004; McBride Brockhausen, & Kan, 2005). Third, OVGP1 activity could be affected by ZP composition.…”

Detection of Nascent and/or Mature Forms of Oviductin in the Female Reproductive Tract and Post-ovulatory Oocytes by Use of a Polyclonal Antibody Against Recombinant Hamster Oviductin

“…The nucleotide sequence of the haOv m fragment is unique to hamster oviductin and does not resemble any other glycoproteins. The specificity of the affinity-purified anti-haOv m antibody has been established by Western blotting analysis (McBride et al 2004). …”

“…Because mammalian oviductins share a common property of being highly glycosylated macromolecules, it has been suggested that different forms of oviductins, varying in number and distribution of oligosaccharide side chains, may be responsible for their diverse biological functions (Buhi 2002). Recently, we have produced a polyclonal antibody against recombinant hamster oviductin that recognizes a peptide portion of hamster oviductin (McBride et al 2004). The newly developed anti-rhaOv m antibody differs from the previously used monoclonal antibodies, which recognize glycosidic epitopes attached to the protein backbone (Araki et al 1987;Kan et al 1988Kan et al ,1989Kan et al ,1993St-Jacques and Bleau 1988;Abe and Oikawa 1990;Martoglio and Kan 1996).…”