| Project/Area Number |
16590056
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| Research Category |
Grant-in-Aid for Scientific Research (C)
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| Allocation Type | Single-year Grants |
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| Section | 一般 |
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| Research Field |
Biological pharmacy
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| Research Institution | University of Shizuoka |
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Principal Investigator |
SUGATANI Junko University of Shizuoka, School of Pharmaceutical Sciences, Associate Professor, 薬学部, 助教授 (30098131)
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| Co-Investigator(Kenkyū-buntansha) |
YOSHINARI Kouichi University of Shizuoka, School of Pharmaceutical Sciences, Assistant Professor, 薬学部, 講師 (60343399)
MIWA Masao University of Shizuoka, School of Pharmaceutics Sciences, Professor, 薬学部, 教授 (10046287)
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| Project Period (FY) |
2004 – 2005
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| Project Status |
Completed (Fiscal Year 2005)
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| Budget Amount *help |
¥3,500,000 (Direct Cost: ¥3,500,000)
Fiscal Year 2005: ¥1,400,000 (Direct Cost: ¥1,400,000)
Fiscal Year 2004: ¥2,100,000 (Direct Cost: ¥2,100,000)
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| Keywords | Nuclear receptor / CAR / Cell cycle / Drug-metabolizing enzyme / UGT1A1 / Transcriptional cofactor / Growth factor / constitutive androstane receptor(CAR) / グルクロン酸抱合酵素UGT / A1 / コファクター / CYP2B6 / グルクロン酸抱合酵素 |
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| Research Abstract |
UDP-glucuronosyltransferase (UGT) 1A1 glucuronidates endogenous metabolites, such as bilirubin, and exogenous substances, and plays a critical role in their detoxification and excretion. We found the phenobarbital response activity to a 290-bp distal enhancer sequence (-3499/-3210) of the human UGT1A1 gene that is activated by the constitutive androstane receptor (CAR). In addition, we show that dexamethasone at submicromolar concentrations enhances the pregnane X receptor (PXR) activator-mediated expression of the UGT1A1 gene and protein in HepG2 cells. We investigated the molecular mechanism of UGT1A1 induction by glucocorticoids at submicromolar concentrations and PXR activators and the functional cross-talk between the glucocorticoid receptor (GR) and CAR/PXR. The glucocorticoid-response element (GRE) was characterized by cotransfection experiments, site-directed mutagenesis and electrophoretic mobility shift assays. Analysis of the human UGT1A1 promoter revealed GREs at -3404/-338
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9 and -3251/-3236 close to the CAR/PXR response element gtNR1 (-3382/-3367). Furthermore, in an in vitro reporter gene assay, dexamethasone effectively enhanced CAR/PXR-mediated transactivation of the 290-bp distal enhancer module in HepG2 cells and CV-1 cells in the presence of exogenously expressed GR and glucocorticoid receptor-interacting protein 1 (GRIP1). Taken together, these results demonstrate a rational mechanistic basis for UGT1A1 induction by glucocorticoids and PXR activators, showing that activated GR enhances CAR/PXR-mediated UGT1A1 regulation with the transcriptional cofactor GRIP1.
In this study, we demonstrate that induction of CAR and the nuclear localization are promoted in HepG2 hepatoma cells arrested at G_1 phase of cell cycle. Addition of epidermal growth factor to the cells decreased the expression of CAR. Moreover inhibition of CAR expression by RNA interference suppressed gene expression of UGT1A1 and Gadd45β. CAR may exert the effects by influencing the expression of gene involved in not only the metabolism of endogenous and exogenous substances but also the control of cell proliferation. Less
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