2002 Fiscal Year Final Research Report Summary
Cell Biological Dissection of GLUT4 Trafficking Pathways
| Project/Area Number |
13671175
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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 |
Metabolomics
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| Research Institution | Gunma University, Institute For Molecular and Cellular Regulation |
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Principal Investigator |
SHIBATA Hiroshi Gunma University, Institute for Molecular and Cellular Regulation, Associate Professor, 生体調節研究所, 助教授 (20235584)
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| Project Period (FY) |
2001 – 2002
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| Keywords | insulin / glucose transporter / GTP-binding protein / Rab4 / syntaxin4 / SNARE / microtubules / 微小管 |
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| Research Abstract |
Our previous studies indicated that the exocytotic and endocytotic pathways for GLUT4 are regulated by GTP-binding proteins, Rab4 and dynamin, respectively. In the present study, we investigated the interaction of Rab4 with syntaxin4, a t-SNARE protein implicated in the insulin-induced exocytic fusion of the GLUT4-containing vesicle with the plasma membrane. Rab4 and syntaxin4 were co-immunoprecipitated from the lysates of rat adipocytes. The interaction of the two proteins were attenuated by pretreatment of the cells with insulin but enhanced with GTPγS. A GTPase deficient mutant of Rab4, but not a GTP-binding defective mutant was bound to syntaxin 4, suggesting that the interaction between the two proteins were regulated by the guanine-nucleotide-binding state of Rab4. In addition, we found that the presence of munc-18c, a negative regulator of the SNARE complex formation, displaced Rab4 from syntaxin 4, indicating that the dissociation of munc-18c from syntaxin4 is required for the Rab4-syntaxin 4 interaction.
We also found that insulin recruits GLUT4 from distinct compartments via distinct traffic pathways with differential microtubule dependence in rat primary adipocytes. Disruption of the microtubules with nocodazole revealed that insulinstimulated glucose transport and GLUT4 translocation were inhibited by about 50%. In addition, the time-course of the insulin stimulation of glucose transport was significantly delayed with nocodazole. Nocodazole partially inhibited insulin-induced translocation of IRAP and VAMP-2 but without effect on GLUT1 and VAMP-3. Thus, the microtubule-independent GLUT4 subpopulation seems to localize to the endosomal recycling compartment, and the independent subpopulation are likely to localize to more specialized compartment.
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[Publications] Suzuki, J., Ohnishi, H., Shibata, H., Iiri, T., Wada, A., Hirayama, T., Ueda, N., Kanamatsu, C., Tsuchida, T., Mashima, H., Yasuda, H. and Fujita, T.: "Dynamin is involved jin the vacuolation induced by Helicobacter Pylori vacuolating cytotoxin (VacA)"J. Clin. Invest.. 107. 363-370 (2001)
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