Unoccupied nuclear receptors for estrogen in human endometrial tissue

Extracts of cell nuclei from normal, nonmalignant, and cancerous human endometrial tissues were found to contain material capable of binding estradiol at 0 C in vitro. Measurements at the same temperature imply that only binding sites unoccupied by endogenous estrogen are determined. The estrogenic receptor character of this binding was demonstrated by: 1) high affinity of binding to estradiol (K_d = ~0.5 nM as assayed by the Scatchard method); 2) specificity of binding, competition by diethylstilbestrol and estriol for estradiol binding, and the absence of competition by cortisol, progesterone, and 5α-dihydrotestosterone; and 3) sedimentation constant at about 4S in a sucrose density gradient. Both unoccupied and occupied nuclear receptors were determined in normal, nonmalignant, and cancerous endometrial tissues. Unoccupied receptors were measured at 0 C for 2 h, while the occupied receptors were calculated from the total binding measured at 30 C for 2 h. Receptors were measured by a single saturating dose of 7.5 nM [³H]estradiol with or without a 100-fold excess of diethylstilbestrol to estimate the amount of nonspecific binding. Unoccupied nuclear receptors were found in all specimens assayed. The unoccupied nuclear receptor comprises 9-37% of the total estradiol receptors (cytoplasmic plus nuclear). The finding that a substantial number of receptors in the nuclei of normal and pathological endometrial tissue are unoccupied may indicate that the unoccupied receptor is a necessary product in the normal mechanism of estradiol action.