Characterization of a Cytoplasmic Receptor for 5±–Dihydrotestosterone in the Caput Epididymidis of Intact Rabbits1

Abstract

We have demonstrated the existence of a highly specific binding component for 17²– hydroxy–5±–androstane–3–one (5±DHT) in cytosol obtained from the caput epididymidis of sexually mature intact rabbits. Using sucrose gradient analysis and charcoal assay methods we were not able to obtain evidence that free receptor activity exists in the corpus or cauda epididymidis. All three epididymal segments contain a component that binds cortisol. The cortisol binding molecule is distinct from the 5±DHT receptor as shown by competition studies and sucrose gradient analysis. Unlabeled testosterone is able to compete with [³H] 5±DHT for sites on the receptor molecule, but is approximately 1/5 as effective as unlabeled 5±DHT. I7²estradiol, 53DHT, cortisol, and progesterone are weak or ineffective competitors. The 5±DHT receptor sediments as a single symmetrical peak with a sedimentation coefficient of approximately 4S on linear 5–20% sucrose gradients. At the concentration of labeled hormone used, only a single high affinity binding component exists in caput cytosol. This component has a K_d of approximately 5.6 X 10^-9M.The concentration of binding sites is approximately 6.0 X 10^-8 moles/g of tissue. The receptor is thermolabile with complete inactivation and/or denaturation occurring upon incubation at 37 C or 65 C. Studies using hydrolytic enzymes and the sulfhydryl blocking agent N—ethylmaleimide indicate that the receptor is at least part protein in nature. (Endocrinology92: 310, 1973)