Helix-loop-helix-type transcription factor (HES-1) is expressed in osteoblastic cells, suppressed by 1,25(OH)2 vitamin D3, and modulates 1,25(OH)2 vitamin D3 enhancement of osteopontin gene expression.

Abstract

The active form of vitamin D, 1,25(OH)2 vitamin D3 (D3), is a potent modulator of osteoblastic function. In this study, we examined, the expression of a negative-type basic helix-loop-helix transcription factor, HES-1, in osteoblastic cells and the regulation of its expression by D3. We found that HES-1 is expressed as a 1.7 kb mRNA in rat osteoblastic osteosarcoma ROS17/2.8 cells. Treatment with D3 suppressed HES-1 mRNA levels by about 50%. This suppression was observed within 24 h and lasted for at least 48 h. The suppressive effect was dose-dependent starting at 10(-9) mol/L and saturated at 10(-8) mol/L. The vitamin D3 suppression of HES-1 mRNA level was blocked by actinomycin D as well as cycloheximide, suggesting the involvement of transcriptional control, which requires new protein synthesis. Proteins in the crude nuclear extracts prepared from ROS17/2.8 cells bound to the N-box sequence (CACNAG). To examine the function of HES-1 in osteoblasts, HES-1 was overexpressed in ROS17/2.8 cells. Overexpression of HES-1 suppressed the vitamin D-dependent upregulation of osteopontin gene expression in these cells. Vitamin D suppression of HES-1 gene expression was also observed in normal rat calvaria-derived osteoblast-enriched cells. These results indicate that HES-1 is expressed in osteoblastic cells and is involved in vitamin D3 regulation of osteoblastic gene expression.