| Project/Area Number |
12470228
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| Research Category |
Grant-in-Aid for Scientific Research (B)
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| Allocation Type | Single-year Grants |
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| Section | 一般 |
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| Research Field |
Metabolomics
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| Research Institution | KYOTO UNIVERSITY |
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Principal Investigator |
SEINO Yutaka Kyoto University Graduate School of Medicine Professor, 医学研究科, 教授 (40030986)
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| Co-Investigator(Kenkyū-buntansha) |
YAMADA Yuichiro Kyoto University Graduate School of Medicine Associate Professor, 医学研究科, 助教授 (60283610)
渡辺 理江 京都大学, 医学研究科, 日本学術振興会特別研究員
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| Project Period (FY) |
2000 – 2002
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| Project Status |
Completed (Fiscal Year 2002)
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| Budget Amount *help |
¥14,300,000 (Direct Cost: ¥14,300,000)
Fiscal Year 2002: ¥2,200,000 (Direct Cost: ¥2,200,000)
Fiscal Year 2001: ¥4,100,000 (Direct Cost: ¥4,100,000)
Fiscal Year 2000: ¥8,000,000 (Direct Cost: ¥8,000,000)
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| Keywords | pancreatic β-cell line / Tet expression system / SAGE / Transcription factor / Target gene / GLUT2 / Proliferation and differentiation / 膵Β細胞株 / 膵β細胞の発生と機能 / Tet Expression System |
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| Research Abstract |
Mutations in the hepatocyte nuclear factor (HNF)-1α gene have been linked to subtype 3 of maturity-onset diabetes of the young (MODY), a disease characterized by a primary defect in insulin secretion. Here we show that the human GLUT2 gene is closely regulated by HNF-1α via sequences downstream of the transcriptional start site by interaction with transcriptional coactivator p300. The promoter region of the human GLUT2 gene was subcloned into luciferase expression plasmids that were transfected together with HNF-1α expression plasmid into a pancreatic β-cell line, HIT-T15, to evaluate transcriptional activities. HNF-1α enhanced human GLUT2 promoter activity sixfold. Site-direct mutagenesis and footprint analyses showed that the HNF-1α binding site (+200 to +218) is critical in human GLUT2 gene expression. Furthermore, mammalian two hybrid and immunoprecipitation studies revealed the transactivation domain of HNF-1α (amino acids 391-540) to interact with both the NH_2-terminal region (amino acids 180-662) and the COOH-terminal region (amino acids 1,818-2,079) of p300. These findings demonstrated that HNF-1α binds to the 5'-untranslated region of GLUT2 and that p300 acts as a transcriptional coactivator for HNF-1α. In addition, these results provided new insight into the regulatory function of HNF-1α or by suggesting a molecular basis for human GLUT2 gene expression.
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