Nuclear receptor 4A2 and C/EBPβ regulate the parathyroid hormone-mediated transcriptional regulation of the 25-hydroxyvitamin D3-1α-hydroxylase

1α-Hydroxylase is the enzyme responsible for the production of the active form of vitamin D, 1,25-dihydroxyvitamin D3. 1α-Hydroxylase, found largely in the kidney, is known to be up-regulated by parathyroid hormone (PTH), however the mechanism of action of PTH and any required transcription factors have not been clearly identified. During gene array analysis we observed that NR4A2, a nuclear orphan receptor, is markedly up-regulated in a porcine kidney cell line (AOK-B50) following PTH stimulation. NR4A2 over-expression increases the endogenous induction of 1α-hydroxylase mRNA in the absence of PTH, however optimal stimulation is achieved when both NR4A2 and PTH are present. An unconventional site of action of NR4A2 was localized to a fragment comprising the sequence from −35/+22 of the 1α-hydroxylase promoter at a C/EBP consensus site. Study of the involvement of C/EBPβ in the 1α-hydroxylase regulation revealed that the transcriptional enhancement by NR4A2 on the 1α-hydroxylase promoter is inhibited by C/EBPβ. In addition, C/EBPβ over-expression decreases the endogenous levels of both NR4A2 and 1α-hydroxylase mRNA.