title: RORα and ROR γ are expressed in human skin and serve as receptors for endogenously produced noncalcemic 20-hydroxy- and 20,23-dihydroxyvitamin D.
abstract: RORα and RORγ are expressed in human skin cells that produce the noncalcemic 20-hydroxyvitamin D3 [20(OH)D3] and 20,23-dihydroxyvitamin D3 [20,23(OH)2D3]. Chinese hamster ovary (CHO) cells stably expressing a Tet-on RORα or RORγ expression vector and a ROR-responsive element (RORE)-LUC reporter, and a mammalian 2-hybrid model examining the interaction between the ligand binding domain (LBD) of RORα or RORγ with an LBD-interacting LXXLL-peptide, were used to study ROR-antagonist activities. These assays revealed that 20(OH)D3 and 20,23(OH)2D3 function as antagonists of RORα and RORγ. Moreover, 20(OH)D3 inhibited the activation of the promoter of the Bmal1 and G6pase genes, targets of RORα, and 20(OH)D3 and 20,23(OH)2D3 inhibited Il17 promoter activity in Jurkat cells overexpressing RORα or RORγ. Molecular modeling using crystal structures of the LBDs of RORα and RORγ revealed docking scores for 20(OH)D3, 20,23(OH)2D3 and 1,25(OH)2D3 similar to those of the natural ligands, predicting good binding to the receptor. Notably, 20(OH)D3, 20,23(OH)2D3, and 1,25(OH)2D3 inhibited RORE-mediated activation of a reporter in keratinocytes and melanoma cells and inhibited IL-17 production by immune cells. Our study identifies a novel signaling pathway, in which 20(OH)D3 and 20,23(OH)2D3 act as antagonists or inverse agonists of RORα and RORγ, that opens new possibilities for local (skin) or systemic regulation.-Slominski, A. T., Kim, T.-K., Takeda, Y., Janjetovic, Z., Broz˙yna, A. A., Skobowiat, C., Wang, J., Postlethwaite, A., Li, W., Tuckey, R. C., Jetten, A. M. RORα and ROR γ are expressed in human skin and serve as receptors for endogenously produced noncalcemic 20-hydroxy- and 20,23-dihydroxyvitamin D.
authors.affiliation: Department of Pathology and Laboratory Medicine, Department of Medicine, and aslominski@uthsc.edu jetten@niehs.nih.gov.
authors.forename: Andrzej T
authors.initials: AT
authors.lastname: Slominski
authors.affiliation: Department of Pathology and Laboratory Medicine.
authors.forename: Tae-Kang
authors.initials: TK
authors.lastname: Kim
authors.affiliation: Cell Biology Section, Division of Intramural Research, National Institute of Environmental Health Sciences, National Institutes of Health, Research Triangle Park, North Carolina, USA;
authors.forename: Yukimasa
authors.initials: Y
authors.lastname: Takeda
authors.affiliation: Department of Pathology and Laboratory Medicine.
authors.forename: Zorica
authors.initials: Z
authors.lastname: Janjetovic
authors.affiliation: Department of Tumor Pathology and Pathomorphology, Oncology Center, Professor Franciszek Łukaszczyk Memorial Hospital, Ludwik Rydygier Collegium Medicum, Nicolaus Copernicus University, Bydgoszcz, Poland;
authors.forename: Anna A
authors.initials: AA
authors.lastname: Brozyna
authors.affiliation: Department of Pathology and Laboratory Medicine.
authors.forename: Cezary
authors.initials: C
authors.lastname: Skobowiat
authors.affiliation: Department of Pharmaceutical Sciences, University of Tennessee Health Science Center, Memphis, Tennessee, USA;
authors.forename: Jin
authors.initials: J
authors.lastname: Wang
authors.affiliation: Department of Medicine, and Department of Veterans Affairs Medical Center, Memphis, Tennessee, USA; and.
authors.forename: Arnold
authors.initials: A
authors.lastname: Postlethwaite
authors.affiliation: Department of Pharmaceutical Sciences, University of Tennessee Health Science Center, Memphis, Tennessee, USA;
authors.forename: Wei
authors.initials: W
authors.lastname: Li
authors.affiliation: School of Chemistry and Biochemistry, University of Western Australia, Crawley, Western Australia, Australia.
authors.forename: Robert C
authors.initials: RC
authors.lastname: Tuckey
authors.affiliation: Cell Biology Section, Division of Intramural Research, National Institute of Environmental Health Sciences, National Institutes of Health, Research Triangle Park, North Carolina, USA; aslominski@uthsc.edu jetten@niehs.nih.gov.
authors.forename: Anton M
authors.initials: AM
authors.lastname: Jetten
mesh_terms: D056930:ARNTL Transcription Factors / Q000235:genetics / Q000378:metabolism; D000818:Animals; D016466:CHO Cells; D002112:Calcifediol / Q000031:analogs & derivatives* / Q000378:metabolism; D045744:Cell Line, Tumor; D002478:Cells, Cultured; D003412:Cricetulus; D004100:Dihydroxycholecalciferols / Q000378:metabolism*; D005260:Female; D005952:Glucose-6-Phosphatase / Q000037:antagonists & inhibitors / Q000235:genetics; D006801:Humans; D020381:Interleukin-17 / Q000235:genetics / Q000378:metabolism; D019169:Jurkat Cells; D015603:Keratinocytes / Q000378:metabolism; D008545:Melanoma / Q000235:genetics / Q000378:metabolism; D051379:Mice; D008811:Mice, Inbred DBA; D057094:Nuclear Receptor Subfamily 1, Group F, Member 1 / Q000235:genetics* / Q000378:metabolism*; D057132:Nuclear Receptor Subfamily 1, Group F, Member 3 / Q000235:genetics* / Q000378:metabolism*; D011401:Promoter Regions, Genetic / Q000235:genetics; D012867:Skin / Q000378:metabolism*
chemical_list: C547379:20,23-dihydroxyvitamin D3; C480634:20-hydroxyvitamin D3; D056930:ARNTL Transcription Factors; C538960:BMAL1 protein, human; D004100:Dihydroxycholecalciferols; D020381:Interleukin-17; D057094:Nuclear Receptor Subfamily 1, Group F, Member 1; D057132:Nuclear Receptor Subfamily 1, Group F, Member 3; C539738:RORA protein, human; C539900:RORC protein, human; D005952:Glucose-6-Phosphatase; D002112:Calcifediol
keywords: CYP11A1; keratinocytes; melanoma cells; retinoic acid-related nuclear receptors