CYP24A1 is a multifunctional, P450 mitochondrial 24-hydroxylase enzyme that is responsible for catabolism of the most active vitamin D hormone (calcitriol, 1,25(OH)D), its precursor (calcifediol, 25(OH)D), and numerous other vitamin D metabolites at the 23- and 24-carbon positions. In the kidney, is induced by 1,25(OH)D, induced by FGF23, and potently suppressed by PTH to tightly control the circulating blood levels of 1,25(OH)D. This gene is believed to be under the control of a pair of classic promoter proximal (PRO) vitamin D response elements (VDREs) that are aided by distal, downstream (DS) containing enhancers that we identified more recently. The DS1 enhancer cluster was found to respond to PTH and FGF23 actions in a kidney-specific manner. The DS2 enhancer cluster was found to assist in the response of 1,25(OH)D in kidney, as well as other target tissues. Despite this knowledge, the contribution of the PRO VDREs to gene expression, what drives basal expression in the kidney, how FGF23 activates , and importantly, how PTH suppresses , all remain unknown. Here in this study, we utilize homology directed CRISPR to mutate one or both VDREs in the PRO region of the gene in the mouse to address these questions. We found that the VDRE (VDRE1) more proximal to the to the transcriptional start site (TSS) is the dominant VDRE of the pair and mutation of both VDREs leads to a dramatic loss of VDR, a reduction of gene expression in the kidney, and a near elimination of 1,25(OH)D induction in the intestine. FGF23 induction of was reduced with mutation of the PRO VDREs, however, co-treatment of 1,25(OH)D and FGF23 synergistically increased expression even with the loss of the PRO VDREs. PTH suppression of gene expression was unchanged with PRO VDRE mutations, despite a minor reduction in total pCREB occupancy. Finally, VDR occupancy was dramatically reduced across the DS enhancers in the locus after the PRO VDREs mutation. Taken together, our data suggest a cooperative relationship between the DS and PRO enhancers in the regulation of by 1,25(OH)D and FGF23, and despite the overall reduction of CREB on the genome it appeared that suppression either does not rely on CREB or that the PRO VDREs are unconnected to PTH suppression altogether. These studies point to the DS1 region as a basal switch for expression and help further define the interconnected genomic control of these hormones on vitamin D catabolism.
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http://www.ncbi.nlm.nih.gov/pmc/articles/PMC11370538 | PMC |
http://dx.doi.org/10.1101/2024.08.23.609393 | DOI Listing |
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