Hormonal regulation of 25-hydroxyvitamin D3-1α-hydroxylase and 24-hydroxylase gene transcription in opossum kidney cells

In the kidney, 25-hydroxyvitamin D3 (25(OH)D) is converted to 1,25-dihydroxyvitamin D3 (1,25(OH)2D) by the 25(OH)D3-1α-hydroxylase enzyme, which contains a terminal cytochrome P450 (CYP1α) (systematic name: CYP27B1). Likewise, the kidney also produces 24,25-dihydroxyvitamin D3 and 1,24,25-trihydroxyvitamin D3 via a 24-hydroxylase whose terminal cytochrome P450 is CYP24. The purpose of this study was to characterize the transcriptional regulation of the CYP1α and CYP24 genes by parathyroid hormone (PTH) and 1,25(OH)2D in the kidney. Promoter–reporter gene constructs were transfected into opossum kidney (OK) cells, a renal proximal tubular cell line with endogenous PTH and 1,25(OH)2D receptors. PTH and forskolin stimulated CYP1α promoter activity via a cAMP-dependent pathway acting through the phosphorylation of CREB (cAMP-dependent response element-binding protein). This stimulation did not require new protein synthesis but may be modulated by short-lived proteins. 1,25(OH)2D modestly inhibited basal and forskolin-stimulated CYP1α promoter activity. The stimulation of CYP1α promoter activity by PTH and forskolin can account for the effect of these hormones on renal CYP1α mRNA levels. CYP24 promoter activity in transfected cells was increased by both 1,25(OH)2D and PTH, but there was no interaction between the two. The modest effects of 1,25(OH)2D and PTH on promoter activity and their lack of interaction do not account for the effects of these hormones on renal CYP24 mRNA levels. This suggests that there may be important posttranscriptional regulation of CYP24 mRNA in the kidney.