Cloning of a recombinant complementary DNA to a baboon (Papio anubis) estradiol-dependent oviduct-specific glycoprotein.
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)
Donnelly
K M
KM
Department of Obstetrics and Gynecology, University of Illinois College of Medicine, Chicago 60680.
Fazleabas
A T
AT
Verhage
H G
HG
Mavrogianis
P A
PA
Jaffe
R C
RC
D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D015152:Blotting, Northern; D002415:Cats; D003001:Cloning, Molecular; D006224:Cricetinae; D004274:DNA, Recombinant / Q000737:chemistry; D000939:Epitopes; D004958:Estradiol / Q000494:pharmacology*; D005187:Fallopian Tubes / Q000187:drug effects / Q000378:metabolism*; D005260:Female; D015698:Genomic Library; D006023:Glycoproteins / Q000737:chemistry / Q000235:genetics*; D006801:Humans; D066298:In Vitro Techniques; D051379:Mice; D008969:Molecular Sequence Data; D008970:Molecular Weight; D010215:Papio; D011374:Progesterone / Q000494:pharmacology; D012333:RNA, Messenger / Q000378:metabolism; D011817:Rabbits; D012689:Sequence Homology, Nucleic Acid
D004274:DNA, Recombinant; D000939:Epitopes; D006023:Glycoproteins; C091468:OVGP1 protein, human; D012333:RNA, Messenger; D011374:Progesterone; D004958:Estradiol