Expression of 25(OH)D-3 24-hydroxylase in distal nephron: coordinate regulation by 1,25(OH)(2)D-3 and cAMP or PTH

Previous studies using microdissected nephron segments reported that the ex clusive site of renal 25-hydroxyvitamin D-3-24-hydroxylase (24OHase) activi ty is the renal proximal convoluted tubule (PCT). We now report the presenc e of 24OHase mRNA, protein, and activity in cells that are devoid of marker s of proximal tubules but express characteristics highly specific for the d istal tubule. 24OHase mRNA was undetectable in vehicle-treated mouse distal convoluted tubule (DCT) cells but was markedly induced when DCT cells were treated with 1,25 dihydroxyvitamin D-3 [1,25(OH)(2)D-3]. 24OHase protein a nd activity were also identified in DCT cells by Western blot analysis and HPLC, respectively. 8-Bromo-cAMP (1 mM) or parathyroid hormone [PTH-(1-34); 10 nM] was found to potentiate the effect of 1,25(OH)(2)D-3 on 24OHase mRN A. The stimulatory effect of cAMP or PTH on 24OHase expression in DCT cells suggests differential regulation of 24OHase expression in the PCT and DCT. In the presence of cAMP and 1,25(OH)(2)D-3, a four- to sixfold induction i n vitamin D receptor (VDR) mRNA was observed. VDR protein, as determined by Western blot analysis, was also enhanced in the presence of cAMP. Transien t transfection analysis in DCT cells with rat 24OHase promoter deletion con structs demonstrated that cAMP enhanced 1,25(OH)(2)D-3-induced 24OHase tran scription but this enhancement was not mediated by cAMP response elements ( CREs) in the 24OHase promoter. We conclude that 1) although the PCT is the major site of localization of 24OHase, 24OHase mRNA and activity can also b e localized in the distal nephron; 2) both PTH and cAMP modulate the induct ion of 24OHase expression by 1,25(OH)(2)D-3 in DCT cells in a manner differ ent from that reported in the PCT; and 3)in DCT cells, upregulation of VDR levels by cAMP, and not an effect on CREs in the 24OHase promoter, is one m echanism involved in the cAMP-mediated modulation of 24OHase transcription.