Binding characteristics of [ 3H]Δ 5-androstene-3β,17β-diol to a nuclear protein in the rabbit vagina

Abstract

In this study, we investigated the binding characteristics of [ 3 H ]Δ 5-androstene-3β,17β-diol to rabbit vaginal cytosolic and nuclear extracts and in freshly excised intact tissue strips. [ 3 H ]Δ 5-Androstene-3β,17β-diol bound to a protein(s) in the vaginal nuclear extract with high affinity ( K d=3–5 nM) and limited capacity (50–100 fmol/mg protein). No specific binding was detected in the cytoplasmic extracts. Competitive binding studies showed that binding of [ 3 H ]Δ 5-androstene-3β,17β-diol was effectively displaced with unlabeled Δ 5-androstene-3β,17β-diol but not with dehydroepiandrosterone, testosterone, dihydrotestosterone, triamcinolone acetonide, or progesterone. However, estradiol at high concentrations partially displaced bound [ 3 H ]Δ 5-androstene-3β,17β-diol. Incubation of freshly excised vaginal tissue strips with [ 3 H ]Δ 5-androstene-3β,17β-diol in the absence or presence of excess unlabeled Δ 5-androstene-3β,17β-diol for 1 h at 37 °C resulted in specific binding to a soluble macromolecule in the nuclear KCl extracts. In addition, quantitative measurement of estrogen receptor, androgen receptor and Δ 5-androstene-3β,17β-diol binding protein was performed by equilibrium ligand binding assays using extracts of distal vaginal tissue from intact animals or ovariectomized animals treated for 2 weeks with vehicle, estradiol, testosterone, or estradiol plus testosterone. These changes in steroid hormone levels resulted in opposing trends between the estrogen receptor and Δ 5-androstene-3β,17β-diol binding protein, suggesting that Δ 5-androstene-3β,17β-diol binding protein is regulated differently by the hormonal milieu than the estrogen receptor. These data suggest that rabbit vaginal tissue expresses a novel binding protein which specifically binds Δ 5-androstene-3β,17β-diol and is distinct from the androgen and estrogen receptors.