1α,25-Dihydroxycholecalciferol Receptors in Intestine

Abstract

Abstract Specific binding of 1α,25-dihydroxycholecalciferol in the cytosol and purified chromatin fractions of chick small intestinal mucosa was studied to investigate the mechanism by which this hormone arrives at its chromatin binding site. The cytosol fraction of the mucosa contains a macromolecule with a high affinity (Kd = 2.2 x 10-9 m) for 1α,25-dihydroxycholecalciferol and has a limited number of binding sites which are saturated at low hormone concentrations (≃ 10-8 m). The association of the sterol with the cytoplasmic component is thermolabile, and is destroyed by treatment with pronase, but not by nucleases, and the receptor component is therefore presumed to be a protein. The protein-sterol complex was isolated and characterized by sucrose gradient centrifugation and sediments at 3 to 3.5 S in the presence of 0.15 m KCl. The interaction of 1α,25-dihydroxy[3H] cholecalciferol with the cytosol protein is reversed by unlabeled 1α,25-dihydroxycholecalciferol g 25-hydroxycholecalciferol g 1α,25-hydroxycholecalciferol g cholecalciferol ≃ estradiol-17β. The subcellular localization of 1α,25-dihydroxycholecalciferol is temperature-dependent. The hormone remains bound to the cytosol fraction at 0° in whole intestines in vitro, and a dramatic movement of 1α,25-dihydroxycholecalciferol to the nuclear chromatin occurs at 37°. The temperature dependence of the hormone localization is also evident in intestinal mucosa homogenates and reconstituted cytosol-chromatin systems. This observation suggests that the cell membrane and nuclear membrane are not involved in the temperature-dependent process. It is postulated that the specific cytosol receptor for 1α,25-dihydroxycholecalciferol functions to transport the hormone to its ultimate site of action in the nuclear chromatin.