| Project/Area Number |
07044226
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| Research Category |
Grant-in-Aid for international Scientific Research
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| Allocation Type | Single-year Grants |
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| Section | Joint Research |
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| Research Institution | Yamaguchi University School of Medicine |
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Principal Investigator |
OKA Yoshitomo Yamaguchi University School of Medicine, Professor, 医学部, 教授 (70175256)
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| Co-Investigator(Kenkyū-buntansha) |
DAVID James E University of Queensland, Center for Molecular Biology and Biotechnology, 分子生物学遺伝子工学センター, 上級研究員
INUKAI Koichi Tokyo University Hospital, Resident, 医学部・附属病院, 医員
KATAGIRI Hideki Tokyo University Hospital, Resident, 医学部・附属病院, 医員
TANIZAWA Yukio Yamaguchi University Hospital, Assistant, 医学部・附属病院, 助手 (00217142)
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| Project Period (FY) |
1995 – 1996
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| Project Status |
Completed (Fiscal Year 1996)
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| Budget Amount *help |
¥4,400,000 (Direct Cost: ¥4,400,000)
Fiscal Year 1996: ¥2,200,000 (Direct Cost: ¥2,200,000)
Fiscal Year 1995: ¥2,200,000 (Direct Cost: ¥2,200,000)
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| Keywords | glucose itasnporter / GLUT1 / GLUT5 / Caco-2 cells / chimeric protein / sorting / MDCK細胞 / 糖輸送 / 促通拡散輸送 / キメラ蛋白 / 細胞内ソーティング |
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| Research Abstract |
Caco-2, a human differentiated intestinal epithelial cell line, is a promising model for investigating the mechanism of polarized targeting of apical and basolateral membrane proteins. We stably transgected rat GLUT5 cDNA and rabbit GLUT1 cDNA into Caco-2 cells with an expression vector. Immunohistochemical study revealed that the GLUT5 protein expressed was localized at apical membranes and that the GLUT1 expressed was present primarily in the basolateral membranes of cells grown on permeable support. Next, to investigate the domain responsible for determining apical vs basolateral sorting in glucose transporters, we prepared several GLUT1-GLUT5 chimeric cDNAs and transfected them into Caco-2 cells. A GLUT1 (N-terminus-6th transmembrane domain (TM6)) -GLUT5 (intracellular loop (IL) -C-terminus) chimera was observed exclusively at the apical membrane, while GLUT1 (N-terminus-IL) -GLUT5 (TM7-C-terminus) and GLUT1 (N-terminus-TM12) -GLUT5 (C-terminal domain) chimeras were observed mainly at the basolateral membrane, a localization similar to that of GLUT1. Based on these results, the C-terminal domain does not determine isoform-specific targeting of GLUT1 and GLUT5.Rather, it is the intracellular loop in glucose transporters that appears to play a pivotal role in apical-basolateral sorting signals in Caco-2 cells.
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