PARTIAL CHARACTERIZATION OF A NOVEL ESTROGEN-INDUCED PROTEIN IN THE RAT ADENOHYPOPHYSIS

In order to detect putative markers of prolactin-secreting pituitary t umours, adult rats were subjected to long-term oestrogenization with o estradiol benzoate (OE2) on a monthly basis. At 6 months, anterior pit uitaries were dissected and incubated either as tissue fragments or as dispersed cells with a [S-35]methionine mix for labelling. Proteins r eleased into the incubation medium and from tissue extracts were furth er analysed by sodium dodecyl sulphate-polyacrylamide gel electrophore sis and fluorography. Oestrogen induced the appearance in the incubati on medium of a protein (OE2 band) with an M(r) of 38 000 under reducin g conditions, and high specific activity. Surprisingly, such a protein was not detected in tissue extracts. The OE2 band was detectable by 7 days after the first dose of oestrogen, and remained throughout 1 yea r of treatment. The tumour cell line GH3 showed a similar OE2 band whi ch was further enhanced by oestrogens. The protein was observed simila rly in both female and male pituitary donors, either intact or gonadec tomized, and also in rats of different strains, suggesting that its ap pearance was independent of the strain of rat and gonadal status. Furt hermore, the OE2 band was specific for pituitary cells and not produce d by other oestrogenized tissues. No alteration in the rate of generat ion or the electrophoretic pattern of the OE2 band was observed when p ituitary cells from oestrogenized rats were metabolically labelled whi le being incubated with tunicamycin. Furthermore, a system for glycan detection, adsorption to Concanavalin A or incubation with endoglycosi dase F also failed to show a clear amount of glycosylation of the oest rogen-induced protein. Both immunoprecipitation experiments and time-l imited proteolysis with V8 protease ruled out the possibility that the OE2 band could be structurally related to either GH or prolactin. In conclusion, oestrogens induce the generation of a new monocatenary pro tein with an apparent M(r) of 38 000, which has at least one intramole cular disulphide loop and is not glycosylated. The OE2 band was detect ed only in incubation medium and never in tissue extracts.