Cloning of a Recombinant Complementary DNA to a Baboon (Papio anubis) Estradiol-Dependent Oviduct-Specific Glycoprotein

Abstract

The estradiol-dominated baboon oviduct synthesizes and secretes a glycoprotein (mol wt, 120,000) that binds to oocyte zona pellucida in vivo. This glycoprotein separates into two major isoelectric variants (pl 8.0 and 4.5) on two-dimensional electrophoretic gels. This study was undertaken to further characterize the steroid-controlled gene expression of this glycoprotein. A recombinant cDNA library was prepared to poly(A)+ RNA isolated from oviducts obtained from estradiol-treated baboons. The library was screened with a polyclonal antibody prepared against the acidic component (pl 4.5) of the glycoprotein. A single positive plaque was purified. Digestion of the recombinant plasmid with EcoRI revealed fragments of 230 and 1000 basepairs. The antibody used to screen the library was epitope selected against the fusion protein produced by the purified recombinant phage. The epitope-selected antibody, when incubated with Western blots of oviductal explant culture medium obtained from estradiol-treated baboons, recognized both the acidic and the basic variants of the glycoprotein. Northern blot hybridization with a 32P-labeled insert indicated that a single message of 2.8 kilobases was present in oviducts obtained from estradiol-treated baboons; no hybridization was detectable to RNA from oviducts obtained from progesterone-treated baboons. A mRNA of comparable size was also found in human, hamster, rabbit, and mouse oviduct tissue. In vitro translation of oviductal poly(A)+ RNA from an estradiol-treated baboon followed by immunoprecipitation with the polyclonal antibody against the acidic component indicated that the protein component of the oviductal glycoprotein had a mol wt of 66,000.(ABSTRACT TRUNCATED AT 250 WORDS)

Oviductins possess chitinase‐ and mucin‐like domains: A lead in the search for the biological function of these oviduct‐specific ZP‐associating glycoproteins

“…Indeed, hamster oviductin was shown to consist of at least two principal charge isoforms, the a and p forms, that possess different levels of glycosylation (Malette and Bleau, 1993). We have recently proposed (Malette and Bleau, 1993) that the heterogeneity of hamster oviductin, also reported in other species (Gandolfi et al, 1989;Buhi et al, 1990;Boice et al, 1990a;Donnelly et al, 1991;Rapisarda et al, 1993), might target different populations of the glycosylated molecules to distinct cellular localizations.…”

“…It is noteworthy that no hybridization could be detected for rat oviductal RNA using baboon (Donnelly et al, 1991) and human (Arias et al, 1994) probes that effectively recognize messages in other rodents, and this might have evolutionary significance.…”

“…The cyclical variations of oviductin synthesis and secretion in other mammals are more subtle, possibly because of the shorter duration of their ovarian cycles. Indeed, Donnelly et al (1991) reported that a partial cDNA of baboon oviductin hybridizes to oviductal mRNA of mice, hamsters, and rabbits isolated from either periovulatory (elevated estradiol) or pregnantipseudopregnant (elevated progesterone) animals; this is in sharp contrast to the virtual absence of oviductin mRNA in the progesteronedominated baboon, human, and cat. These results were recently extended using a full length human cDNA (Arias et al, 1994).…”

“…Comparative Northern blot analyses (Donnelly et al, 1991;Arias et al, 1994;DeSouza and Murray, 1994) Sequence similarity of oviductins and mammalian members of the chitinase protein family. The amino acid sequences of the hamster (GenbankTM Accession number U15048), human (GenbankTM Accession number U095501, bovine (GenbankTM Accession number D16639), and baboon (Genbankm Accession number M59903) oviductins were optimally aligned using the program Pileup with the sequences of human cartilage glycoprotein 39 (HC gp-391, a major secretory product of articular chondrocytes and synovial cells (GenbankTM Accession number M80927), and YM-1, a secretory protein from mouse macrophages (GenbankTM Accession number M94584).…”

“…Indeed, while it has been clearly demonstrated that some of these oviduct-specific secretory proteins are released in response to endogenous estrogen at estrus in the mouse (Horvat et al, 1992), sheep (Sutton et al, 1986;Murray, 19931, human (Verhage et al, 1988), cow (Malayer et al, 1988;Gerena and Killian, 1990; Abe et al, 1993b), baboon (Ver- , and gilt (Buhi et al, 1989(Buhi et al, , 1990 or in ovariectomized estrogen-treated animals (Oliphant et al, 1984b;Buhi et al, ,1992Murray, 1993;) the role of steroid sex hormones in the control of the secretion (Malette et al, 1994) or mRNA expression (Donnelly et al, 1991;Arias et al, 1994) of others seems to be different. Some of these oviduct-specific secretory glycoproteins possess the interesting properties of interaction with the zona pellucida (zp) and/or perivitelline (PV) space of the oocyte during its passage through the oviduct and remain associated with the embryo until implantation.…”