Intracellular location of unoccupied 1,25-dihydroxyvitamin D receptors: a nuclear-cytoplasmic equilibrium.
In order to investigate the subcellular distribution of unoccupied 1,25-dihydroxyvitamin D3 receptors, highly purified cytoplasts and nucleoplasts were prepared from two kidney cell lines (PK1 and MDBK). This was accomplished utilizing the technique of enucleation by cytochalasin B and density gradient centrifugation. Unoccupied 1,25-dihydroxyvitamin D3 receptors were found in both the nuclear and cytosolic compartments, with approximately 70% of the receptors localized in the cytoplasm. When cells were pretreated with 1,25-[3H]dihydroxyvitamin D, prior to enucleation, it was found that 90% of the receptor-hormone complex was associated with nucleoplasts, thus demonstrating that cytochalasin B treatment does not alter the high-affinity association of the receptor-hormone complex with the nucleus. The ratio of unoccupied receptor/protein was found to be the same in whole cells, cytoplasts, and nucleoplasts for both cell types. The ratio of unoccupied receptor/DNA was highest in cytoplasts and lowest in nucleoplasts. Taken together, these data indicate that the unoccupied 1,25-dihydroxyvitamin D receptor is generally associated with cell proteins and not specifically associated with cell DNA. We therefore propose, at least for these cells, that the unoccupied 1,25-dihydroxyvitamin D receptor exists in equilibrium between the nuclear and cytosolic compartments of the whole cell, and receptor-hormone binding shifts this equilibrium to favor nuclear localization.
D000818:Animals; D002417:Cattle; D002460:Cell Line; D002467:Cell Nucleus / Q000378:metabolism* / Q000648:ultrastructure; D002499:Centrifugation, Density Gradient; D003571:Cytochalasin B; D003593:Cytoplasm / Q000378:metabolism* / Q000648:ultrastructure; D007668:Kidney / Q000378:metabolism; D008854:Microscopy, Electron; D008954:Models, Biological; D018167:Receptors, Calcitriol; D011987:Receptors, Steroid / Q000378:metabolism*; D013552:Swine
D018167:Receptors, Calcitriol; D011987:Receptors, Steroid; D003571:Cytochalasin B