PURPOSE: To develop and characterize a new drug-regulated gene expression system based on the nuclear receptor constitutive androstane receptor (CAR). METHODS: Both transient and stable transfection into HEK293 cells of luciferase plasmids under the control of either drug- and steroid-responsive nuclear receptor CAR or the tetracycline-sensitive transactivator tTA were used in development of stable cell lines. RESULTS: A stable first-generation cell line that expresses luciferase gene under the control of nuclear receptor CAR was developed. The luciferase expression in CAR-producing cells could be suppressed by androstanes and reactivated by structurally unrelated drugs chlorpromazine, metyrapone, phenobarbital, and clotrimazole. The kinetics of luciferase expression in CAR-producing cells and the tTA system were comparable. The overall regulation of CAR system was improved by modifications to the DNA binding domain and site. CONCLUSIONS: Because of its wide ligand selectivity and transferable ligand binding domain, CAR expands the repertoire of regulated gene expression systems.
PURPOSE: To develop and characterize a new drug-regulated gene expression system based on the nuclear receptor constitutive androstane receptor (CAR). METHODS: Both transient and stable transfection into HEK293 cells of luciferase plasmids under the control of either drug- and steroid-responsive nuclear receptor CAR or the tetracycline-sensitive transactivator tTA were used in development of stable cell lines. RESULTS: A stable first-generation cell line that expresses luciferase gene under the control of nuclear receptor CAR was developed. The luciferase expression in CAR-producing cells could be suppressed by androstanes and reactivated by structurally unrelated drugs chlorpromazine, metyrapone, phenobarbital, and clotrimazole. The kinetics of luciferase expression in CAR-producing cells and the tTA system were comparable. The overall regulation of CAR system was improved by modifications to the DNA binding domain and site. CONCLUSIONS: Because of its wide ligand selectivity and transferable ligand binding domain, CAR expands the repertoire of regulated gene expression systems.
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