Vitamin D₃ supports osteoclastogenesis via functional vitamin D response element of human RANKL gene promoter

Abstract

Receptor activator of NF-kappaB ligand (RANKL) has been identified as requisite for osteoclastogenesis. To elucidate the molecular mechanism that conducts its catabolic action on bone, the effect of 1alpha,25 dihydroxyvitamin D(3) (1alpha,25(OH)(2)D(3)) on osteoclastogenesis and RANKL mRNA expression was examined by coculture, RT-PCR and nuclear run-on studies. By accelerating the transcription rate of the RANKL gene in SaOS2 osteoblastic cells, 1alpha,25(OH)(2)D(3) enhanced in vitro osteoclast formation from peripheral monocytes. Cloning and characterization of the 5'-flanking region of the human RANKL gene revealed that the basic promoter comprises inverted TATA- and CAAT-boxes flanked by RUNX2 binding sites. Both electrophoresis mobility shift assay (EMSA) and transfection studies demonstrated that 1alpha,25(OH)(2)D(3) activated human RANKL promoter through vitamin D responsive elements (VDRE) located at -1584/-1570 by binding VDR and RXRalpha heterodimers in a ligand-dependent manner. The results provide direct evidence that 1alpha,25(OH)(2)D(3) augments osteoclastogenesis by transactivating the human RANKL gene in osteoblastic cells through VDRE.