| Project/Area Number |
09671047
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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 |
内分泌・代謝学
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| Research Institution | KYOTO UNIVERSITY |
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Principal Investigator |
YOSHIMASA Yasunao Kyoto Universitiy Graduate School of Medicine, Associate Professor, 医学研究科, 講師 (00252437)
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| Co-Investigator(Kenkyū-buntansha) |
HOSODA Kiminori Kyoto University Graduate School of Human and Assistant Professor, 医学研究科, 助手 (40271598)
NISHIMURA Haruo Kyoto Universitiy Graduate School of Medicine, Euviromental Studies, Assistant P, 人間環境学研究科, 助手 (40281729)
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| Project Period (FY) |
1997 – 1998
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| Project Status |
Completed (Fiscal Year 1998)
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| Budget Amount *help |
¥3,000,000 (Direct Cost: ¥3,000,000)
Fiscal Year 1998: ¥1,000,000 (Direct Cost: ¥1,000,000)
Fiscal Year 1997: ¥2,000,000 (Direct Cost: ¥2,000,000)
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| Keywords | 3T3-L1 adipocyte / MAP kinase / MAP kinase kinase / Glucose transporter / Insulin resistance / Adipocyte differentiation / MAPキナーゼ・キナーゼ / インスリン |
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| Research Abstract |
To address a role of MAP kinase in the regulation of glucose transport, we made a constitutively active mutant of MAP kinase kinase and introduced it into 3T3-L1 preadipocytes using a retrovirus-mediated transfection procedure. The deletion of 20 amino acids (32-5 1) in the amino terminal region of Xenopus MAP kinase and the replacement of serine residues on the 218 and 222 positions by glutamic acid (dSESE-MAPKK) let Xenopus MAP kinase constitutively active. The isolated cell clones differently expressing dSESE-MAPKK (Clone 219 ; higher, Clone 233 lower expression) efficiently differentiated to adipocytes by a standard differentiation cocktail. Accordingly, the increased expression of dSESE-MAPKK protein during differentiation resulted in the increased basal MAP kinase activity in Clone 219 adipocytes but to a lesser extent in Clone 233 adipocytes. In contrast to Clone 233 and parental adipocytes, basal 2-deoxyglucose uptake was enhanced 4-fold in Clone 219 adipocytes, in accordance w
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ith increased expression of GLUT 1 mRNA and protein. Whereas GLUT4 mRNA was similarly expressed in all the adipocytes, GLUT4 protein appeared to decrease in Clone 219 adipocytes. More importantly, subcellular fractionation studies showed that the localization of both GLUT1 and GLUT4 in the plasma membranes was markedly increased in the basal state in Clone 219 adipocytes compared with that in Clone 233 and parental adipocytes, in which both glucose transporters were preferentially located in intracellular compartments. Consequently, insulin-induced translocation of GLUT 1 was abolished in Clone 219 adipocytes, although the remaining intracellular GLUT4 was still responsive to insulin stimulation leading to the movement to the plasma membrane. As combined effects on the situation of GLUT 1 and GLUT4, the foldness of insulin stimulation of glucose transport based on the basal activity was reduced in cells expressing constitutively active MAP kinase. These results imply that chronic activation of MAP kinase could be one of the mechanisms for insulin resistance. Less
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