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Id gene expression and its suppression by 1,25-dihydroxyvitamin D3 in rat osteoblastic osteosarcoma cells.
Abstract
Id is one of the helix-loop-helix family of proteins that regulate differentiation in several types of cells, including myoblasts. We found that Id mRNA was constitutively expressed in ROS17/2.8 rat osteoblastic osteosarcoma cells and that the level of Id message in these cells was suppressed by 1,25-dihydroxyvitamin D3 (vitamin D), a calcitropic hormone known to enhance expression of differentiation-related phenotypes in these cells. The vitamin D suppression was dose-dependent, starting at 10(-11) M and saturating at 10(-8) M. This vitamin D effect was seen within 6 hr after the initiation of the treatment and lasted at least 48 hr. Cycloheximide did not block the vitamin D suppression of Id message level. Vitamin D reduced the rate of Id gene transcription by approximately 80% as estimated by nuclear run-on assay. Electrophoretic mobility-shift assay indicated the specific binding of nuclear factors from ROS17/2.8 cells to an E-box DNA sequence, and the binding signal was enhanced in nuclear extracts of the cells treated with vitamin D. The suppressive effect was specific to vitamin D, since another calcitropic hormone, the synthetic compound dexamethasone, did not suppress Id message expression. These observations indicate the presence of helix-loop-helix proteins in osteoblastic cells and indicate that vitamin D, a potent regulator of differentiation in several types of cells, controls the expression of the helix-loop-helix molecules.
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