Functional Cooperation between CCAAT/Enhancer-Binding Proteins and the Vitamin D Receptor in Regulation of 25-Hydroxyvitamin D₃ 24-Hydroxylase

Abstract

1,25-Dihydroxyvitamin D₃ [1,25(OH)₂D₃] induces the synthesis of 25-hydroxyvitamin D₃ 24-hydroxylase [24(OH)ase], an enzyme involved in its catabolism, thereby regulating its own metabolism. Here we demonstrate that CCAAT enhancer binding protein β (C/EBPβ) is induced by 1,25(OH)₂D₃ in kidney and in osteoblastic cells and is a potent enhancer of vitamin D receptor (VDR)-mediated 24(OH)ase transcription. Transfection studies indicate that 1,25(OH)₂D₃ induction of 24(OH)ase transcription is enhanced a maximum of 10-fold by C/EBPβ. Suppression of 1,25(OH)₂D₃-induced 24(OH)ase transcription was observed with dominant negative C/EBP or osteoblastic cells from C/EBPβ^−/− mice. A C/EBP site was identified at positions −395 to −388 (−395/−388) in the rat 24(OH)ase promoter. Mutation of this site inhibited C/EBPβ binding and markedly attenuated the transcriptional response to C/EBPβ. We also report the cooperation of CBP/p300 with C/EBPβ in regulating VDR-mediated 24(OH)ase transcription. We found that not only 1,25(OH)₂D₃ but also parathyroid hormone (PTH) can induce C/EBPβ expression in osteoblastic cells. PTH potentiated the induction of C/EBPβ and 24(OH)ase expression in response to 1,25(OH)₂D₃ in osteoblastic cells. Data with the human VDR promoter (which contains two putative C/EBP sites) indicate a role for C/EBPβ in the protein kinase A-mediated induction of VDR transcription. From this study a fundamental role has been established for the first time for cooperative effects and cross talk between the C/EBP family of transcription factors and VDR in 1,25(OH)₂D₃-induced transcription. These findings also indicate a novel role for C/EBPβ in the cross talk between PTH and 1,25(OH)₂D₃ that involves the regulation of VDR transcription.

ACKNOWLEDGMENTS

We are indebted to the investigators who contributed reagents to this study (see Materials and Methods). We thank Michael Huening for his help and advice.

This work was supported by grants from the National Institutes of Health (grant DK38961 to S.C., grant DK53980 to P.N.M., and grant DK56310 to T.L.M.). A.L.M.S. was supported by a predoctoral fellowship from the Pharmaceutical Research Manufacturer's Association.