Nuclear progesterone receptor in hamster uterus: measurement by [3H]progesterone exchange during the estrous cycle.

Abstract

Previous studies with hamster uterine cytosol demonstrated a progesterone (P) receptor (Rp) that was responsive to estrogen and P action. This report describes a [3H]P exchange procedure which permits the reliable assay of nuclear Rp during the hamster estrous cycle and other experimental conditions. Nuclear Rp was extracted from uterine nuclei at 2 C using 0.5 M KC1 in Tris buffer containing glycerol. [3H]P exchange was achieved by overnight incubation of nuclear KC1 extract at 2 C. Nuclear Rp levels were obtained by Scatchard plot analysis of specific [3H]P-binding results. Nuclear Rp sedimented at 3.5S compared to 4.6S for salt-dissociated cytosol Rp in sucrose-glycerol gradients. The steroid-binding specificity of nuclear Rp was identical to that of cytosol Rp under the same ionic conditions. In vivo experiments revealed that nuclear Rp translocation was target tissue specific, hormone specific, and dependent on P dosage. A 6-h retention of nuclear Rp wasobserved in response to varying doses of P (0.1–4 mg/100 g BW). During the estrous cycle, nuclear Rp levels remained relatively constant (5–10 pmol/g tissue) on days 1–3 and increased markedly (28 pmol/g tissue) at 1600 h on day 4, coincident with preovulatory P secretion and cytosol Rp depletion. Cytosol Rp was translocated to the nuclear fraction by exogenous P treatment on all days of the cycle, but the magnitude of the nuclear Rp response was correlated with cytosol Rp availability. These results demonstrate the applicability of the [3H]P exchange assay for monitoring nuclear Rp translocation under physiological conditions.