Expression of 25(OH)D3 24-hydroxylase in distal nephron: coordinate regulation by 1,25(OH)2D3 and cAMP or PTH.
Previous studies using microdissected nephron segments reported that the exclusive site of renal 25-hydroxyvitamin D3-24-hydroxylase (24OHase) activity is the renal proximal convoluted tubule (PCT). We now report the presence of 24OHase mRNA, protein, and activity in cells that are devoid of markers of proximal tubules but express characteristics highly specific for the distal 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 D3 [1,25(OH)2D3]. 24OHase protein and 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)2D3 on 24OHase mRNA. 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)2D3, a four- to sixfold induction in vitamin D receptor (VDR) mRNA was observed. VDR protein, as determined by Western blot analysis, was also enhanced in the presence of cAMP. Transient transfection analysis in DCT cells with rat 24OHase promoter deletion constructs demonstrated that cAMP enhanced 1, 25(OH)2D3-induced 24OHase transcription 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 be localized in the distal nephron; 2) both PTH and cAMP modulate the induction of 24OHase expression by 1,25(OH)2D3 in DCT cells in a manner different 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 mechanism involved in the cAMP-mediated modulation of 24OHase transcription.
Yang
W
W
Department of Biochemistry and Molecular Biology, University of Medicine and Dentistry of New Jersey-New Jersey Medical School and Graduate School of Biomedical Sciences, Newark, New Jersey 07103, USA.
Friedman
P A
PA
Kumar
R
R
Omdahl
J L
JL
May
B K
BK
Siu-Caldera
M L
ML
Reddy
G S
GS
Christakos
S
S
D015124:8-Bromo Cyclic Adenosine Monophosphate / Q000494:pharmacology; D000818:Animals; D001483:Base Sequence; D002117:Calcitriol / Q000494:pharmacology*; D002478:Cells, Cultured; D000242:Cyclic AMP / Q000494:pharmacology / Q000502:physiology*; D003577:Cytochrome P-450 Enzyme System; D004790:Enzyme Induction; D015971:Gene Expression Regulation, Enzymologic / Q000187:drug effects / Q000502:physiology*; D007684:Kidney Tubules / Q000201:enzymology*; D007686:Kidney Tubules, Distal / Q000201:enzymology; D051379:Mice; D008969:Molecular Sequence Data; D009399:Nephrons / Q000201:enzymology*; D010281:Parathyroid Hormone / Q000502:physiology*; D012333:RNA, Messenger / Q000235:genetics; D051381:Rats; D011993:Recombinant Fusion Proteins / Q000096:biosynthesis; D012045:Regulatory Sequences, Nucleic Acid; D013250:Steroid Hydroxylases / Q000096:biosynthesis / Q000235:genetics*; D019379:Teriparatide / Q000494:pharmacology; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology; D014158:Transcription, Genetic / Q000187:drug effects; D014162:Transfection; D065668:Vitamin D3 24-Hydroxylase
D010281:Parathyroid Hormone; D012333:RNA, Messenger; D011993:Recombinant Fusion Proteins; D019379:Teriparatide; D015124:8-Bromo Cyclic Adenosine Monophosphate; D003577:Cytochrome P-450 Enzyme System; D000242:Cyclic AMP; D013250:Steroid Hydroxylases; D065668:Vitamin D3 24-Hydroxylase; D002117:Calcitriol; D013755:Tetradecanoylphorbol Acetate