Glucocorticoid binding in the hen oviduct.
[(3)H]Triamcinolone acetonide (15nm) was incubated with cytosol (150000g fraction) prepared from oviducts of egg-laying hens. The extent of steroid binding, as determined by charcoal assays, was greatest between 2-4h at 4 degrees C. A similar time curve was obtained when cytosol preparations were first fractionated with (NH(4))(2)SO(4) before labelling. The addition of 10mm-Na(2)MoO(4) or 10mm-ATP during the incubation of hen oviduct cytosol with [(3)H]triamcinolone acetonide lowered the extent of steroid binding. The presence of glycerol (20%), however, increased the extent of [(3)H]triamcinolone acetonide binding in cytosol fractions from chick (330%) and hen (160%) oviducts. The [(3)H]triamcinolone acetonide-receptor complex was stable for over 4h at 4 degrees C, but dissociated rapidly at 37 degrees C, exhibiting a half-life of about 10min. The presence of 10mm-Na(2)MoO(4) and 10mm-ATP or both had a small protective effect on the dissociation of [(3)H]triamcinolone acetonide-receptor complex. The receptor from hen oviduct showed significant affinity for unlabelled triamcinolone acetonide, cortisol, compound R5020 and dihydrotestosterone and, to a lesser extent, for oestradiol, oestrone and progesterone. Diethylstilboestrol treatment of immature chicks appeared to induce a more specific binder, which showed affinity for unlabelled triamcinolone acetonide, cortisol and compound R5020 only. Scatchard analysis of [(3)H]triamcinolone acetonide binding in hen oviduct cytosol revealed a K(d) value of 6.4nm. The steroid-receptor complex sedimented as a 7-8S and a 4S entity on low-salt (0.01m-KCl)- and high-salt (0.3m-KCl)-containing sucrose gradients respectively. The cytosol [(3)H]triamcinolone acetonide-receptor complex showed no affinity for ATP-Sepharose or DNA-cellulose, but acquired this ability on heat activation (23 degrees C, 40min). The data indicate the avian oviduct possesses a high-affinity binding molecule that fulfils the criteria of a glucocorticoid receptor.
D000255:Adenosine Triphosphate / Q000494:pharmacology; D000818:Animals; D002645:Chickens; D002846:Chromatography, Affinity; D005260:Female; D005990:Glycerol / Q000494:pharmacology; D066298:In Vitro Techniques; D007700:Kinetics; D008982:Molybdenum / Q000494:pharmacology; D010057:Oviducts / Q000187:drug effects / Q000378:metabolism*; D011965:Receptors, Glucocorticoid / Q000187:drug effects / Q000378:metabolism*; D011987:Receptors, Steroid / Q000378:metabolism*; D014222:Triamcinolone Acetonide / Q000378:metabolism*
D011965:Receptors, Glucocorticoid; D011987:Receptors, Steroid; D008982:Molybdenum; D000255:Adenosine Triphosphate; D014222:Triamcinolone Acetonide; D005990:Glycerol