1α,25 Dihydroxyvitamin D3 Rapidly Regulates the Mouse Osteoprotegerin Gene Through Dual Pathways

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Abstract
1α,25(OH)2D3 rapidly and transiently suppressed OPG gene expression both by accelerating the degradation of mRNA and by suppressing promoter activity. The latter process was mediated through the AP‐1 binding site by a reduction in the proportion of phospho‐c‐Jun in a JNK‐independent manner. Introduction: Osteoclastogenesis is regulated by an integrated network of numerous bone metabolic factors, among which 1α,25‐dihydroxyvitamin D3 [1α,25(OH)2D3] promotes osteoclastogenesis by reciprocally upregulating the expression of RANKL and downregulating that of osteoprotegerin (OPG). Materials and Methods: To analyze the mechanism by which 1α,25(OH)2D3 suppresses OPG, we characterized cis‐acting elements of the mouse OPG gene and assessed the post‐transcriptional modifications by actinomycin D assays. Results: 1α,25(OH)2D3 rapidly and transiently suppressed OPG expression and shortened the half‐life of OPG mRNA; additionally, the c‐Jun homodimer bound to the AP‐1 binding site (TGACTGA, −293/−287) and maintained steady‐state transcription of the OPG gene. Furthermore, mutation of the AP‐1 site negated 1α,25(OH)2D3‐driven OPG suppression. Moreover, 1α,25(OH)2D3 treatment of ST2 cells decreased the amount of phosphorylated c‐Jun protein (phospho‐c‐Jun), while the total amount of c‐Jun remained constant; however, the amount of phosphorylated Jun N‐terminal kinase (JNK) was nearly unchanged by 1α,25(OH)2D3 treatment. Conclusion: Taken together with the observation that the OPG promoter has no consensus negative vitamin D‐responsive elements, these data suggest that 1α,25(OH)2D3 transrepresses mouse OPG by reducing the proportion of phospho‐c‐Jun in a JNK‐independent manner. Our data indicated that short‐term treatment with 1α,25(OH)2D3 effectively downregulated OPG expression both by accelerating the degradation of OPG mRNA and by transrepressing the OPG gene through its AP‐1 binding site in the catabolic phase. The OPG gene became insensitive to 1α,25(OH)2D3 treatment, however, and reverted to its steady‐state expression level over time, leading to the anabolic phase of the effect of 1α,25(OH)2D3 on bone.

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