The Human Peroxisome Proliferator-activated Receptor dGene is a Primary Target of 1a,25-Dihydroxyvitamin D3 and its Nuclear Receptor

0022-2836/$ see front matter q 2005 E Abbreviations used: 1a,25(OH)2D dihydroxyvitamin D3; AcH4, acetyl atrial natriuretic factor; ARP0, acidic CREB binding protein; ChIP, chrom precipitation; CYP24, vitamin D3 24 direct repeat spaced by three nucleo polymerase II; PPAR, peroxisome p receptor; RXR, retinoid X receptor; T TSS, transcription start site; VDR, v VDRE, vitamin D sterol response el E-mail address of the correspond [email protected] Peroxisome proliferator-activated receptor (PPAR) d is the most widely expressed member of the PPAR family of nuclear receptor fatty acid sensors. Real-time PCR analysis of breast and prostate cancer cell lines demonstrated that PPARd expression was increased 1.5 to 3.2-fold after three hours stimulation with the natural vitamin D receptor (VDR) agonist, 1a,25-dihydroxyvitamin D3 (1a,25(OH)2D3). In silico analysis of the 20 kb of the human PPARd promoter revealed a DR3-type 1a,25(OH)2D3 response element approximately 350 bp upstream of the transcription start site, which was able to bind VDR-retinoid X receptor (RXR) heterodimers and mediate a 1a,25(OH)2D3-dependent upregulation of reporter gene activity. Chromatin immuno-precipitation assays demonstrated that a number of proteins representative for 1a,25(OH)2D3-mediated gene activation, such as VDR, RXR and RNA polymerase II, displayed a 1a,25(OH)2D3dependent association with a region of the proximal PPARd promoter that contained the putative DR3-type VDRE. This was also true for other proteins that are involved in or are the subject of chromatin modification, such as the histone acetyltransferase CBP and histone 4, which displayed ligand-dependent association and acetylation, respectively. Finally, realtime PCR analysis demonstrated that 1a,25(OH)2D3 and the synthetic PPARd ligand L783483 show a cell and time-dependent interference in each other’s effects on VDR mRNA expression, so that their combined application shows complex effects on the induction of VDR target genes, such as CYP24. Taken together, we conclude that PPARd is a primary 1a,25(OH)2D3-responding gene and that VDR and PPARd signaling pathways are interconnected at the level of cross-regulation of their respective transcription factor mRNA levels. q 2005 Elsevier Ltd. All rights reserved.