title: Transcriptional activity of a fluorinated vitamin D analog on VDR-RXR-mediated gene expression.
abstract: The transcriptional activity of the hexafluorinated derivative of 1,25-dihydroxyvitamin D3 [1,25-(OH)2D3], 26,26,26,27,27,27-hexafluoro-1,25-dihydroxyvitamin D3 [F6-1,25-(OH)2D3], was examined in cultured cells by a transient expression assay (CAT assay) using expression vectors for the rat nuclear vitamin D3 receptor (VDR) and the rat 9-cis-retinoic acid receptor (RXR beta), and a reporter plasmid containing a consensus vitamin D3 response element (VDRE) consisting of two directly repeated AGGTCA motifs spaced by 3 bp (DR3). At physiological concentrations, the transcriptional activity of F6-1,25-(OH)2D3 was 2-4 times more potent than that of 1,25-(OH)2D3 in both nontarget (HeLa) and target (UMR106) cells for 1,25-(OH)2D3. The transcriptional activity of F6-1,25-(OH)2D3 was also higher when the endogenous target gene (osteopontin), which has a VDRE related to the DR3 in its promoter, was induced. A gel-shift assay using DR3 as a probe and in vitro synthesized receptors showed that the ligand-induced DNA binding of VDR required RXR to form a heterodimer. Moreover, in this assay we found that F6-1,25-(OH)2D3 induced the receptor-DNA complex at a 10-fold lower concentration than 1,25-(OH)2D3 without influencing the dissociation kinetics. However, the binding affinity of F6-1,25-(OH)2D3 for VDR was slightly lower than that of 1,25-(OH)2D3. The increased DNA binding of ligand-bound VDR by introducing hexafluorines into 1,25-(OH)2D3 may potentiate the transcriptional activity. Thus, the higher biological activity of F6-1,25-(OH)2D3 may be exerted at least in part by enhanced transcriptional activity.
authors: Department of Agricultural Chemistry, Faculty of Agriculture, Tokyo University of Agriculture, Japan.
authors: H
authors: H
authors: Sasaki
authors: H
authors: H
authors: Harada
authors: Y
authors: Y
authors: Handa
authors: H
authors: H
authors: Morino
authors: M
authors: M
authors: Suzawa
authors: E
authors: E
authors: Shimpo
authors: T
authors: T
authors: Katsumata
authors: Y
authors: Y
authors: Masuhiro
authors: K
authors: K
authors: Matsuda
authors: K
authors: K
authors: Ebihara
mesh_terms: D000818:Animals; D001483:Base Sequence; D002117:Calcitriol / Q000031:analogs & derivatives* / Q000378:metabolism / Q000494:pharmacology; D015500:Chloramphenicol O-Acetyltransferase / Q000235:genetics; D004268:DNA-Binding Proteins / Q000502:physiology*; D005786:Gene Expression Regulation / Q000187:drug effects; D006367:HeLa Cells; D006801:Humans; D008969:Molecular Sequence Data; D051381:Rats; D018167:Receptors, Calcitriol / Q000378:metabolism / Q000502:physiology*; D018168:Receptors, Retinoic Acid / Q000502:physiology*; D012045:Regulatory Sequences, Nucleic Acid / Q000502:physiology; D047488:Retinoid X Receptors; D014157:Transcription Factors / Q000502:physiology*; D014158:Transcription, Genetic / Q000187:drug effects*; D014162:Transfection; D005466:Fluorocarbons
chemical_list: D002117:Calcitriol; D015500:Chloramphenicol O-Acetyltransferase; D004268:DNA-Binding Proteins; D018167:Receptors, Calcitriol; D018168:Receptors, Retinoic Acid; D047488:Retinoid X Receptors; D014157:Transcription Factors; C040488:26,26,26,27,27,27-hexafluoro-1,25-dihydroxyvitamin D3; D005466:Fluorocarbons