Intracellular localization and translocation of 1 alpha, 25-dihydroxyvitamin D3 receptor in osteoblasts.

Abstract

The intracellular localization and translocation of the 1 alpha,25-dihydroxyvitamin D3 receptor (1 alpha,25(OH)2D3 receptor) in osteoblasts of mouse parietal bone and MC3T3-E1 cells were examined immunocytochemically using a rat monoclonal antibody to 1 alpha,25(OH)2D3 receptor. In osteoblasts of parietal bones in vivo, immunoreactivity for 1 alpha,25(OH)2D3 receptor was detected not only in the nuclei but also in lysosomal structures, and also sparsely in the cytoplasmic matrix. The transport of 1 alpha,25(OH)2D3 receptor was investigated immunocytochemically after incubation with 1 alpha,25(OH)2D3. In osteoblasts of parietal bones, after 1 min incubation with 10(-8) M 1 alpha,25(OH)2D3, the perinuclear cytoplasm showed intense immunoreactivity for 1 alpha,25(OH)2D3 receptor. After 10 min incubation, immunoreactivity was intense in the nuclei, especially in the heterochromatin. In MC3T3-E1 cells, after 1 min incubation with 1 alpha,25(OH)2D3, immunoreactivity for 1 alpha,25(OH)2D3 receptor was found in the form of a fibrillar structure extending radially to the periphery of the cells. The immunostaining pattern of anti-1 alpha,25(OH)2D3 receptor was similar to the distribution of microtubules stained with anti-beta-tubulin antibody. After 10 min incubation, the nuclei showed intense immunoreactivity for 1 alpha,25(OH)2D3 receptor. Incubation with colchicine dissociated the fibrillar structures and inhibited the intranuclear localization of the 1 alpha,25(OH)2D3 receptor. These findings suggest that the 1 alpha,25(OH)2D3 receptor is located in the nuclei, in lysosomal structures and also sparsely in the cytoplasmic matrix of osteoblasts in vivo, and that the receptor is transported to the perinuclear cytoplasm via microtubules to be then translocated into the nucleus, especially into the heterochromatin.