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A more recent version of this article appeared on March 1, 2003
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Submitted on June 3, 2002
Revised on November 7, 2002
Accepted on November 22, 2002
1 Institute for Molecular Biosciences and Department of Physiology and Pharmacology, University of Queensland, St. Lucia, Brisbane, QLD 4072, Australia
2 Institute for Molecular Biosciences, University of Queensland, St. Lucia, Brisbane, QLD 4072, Australia; and Garvan Institute of Medical Research, St. Vincent's Hospital, 384 Victoria St. Darlinghurst, 2010, NSW, Australia
3 Membrane Biology Laboratory, Institute of Molecular and Cell Biology, National University of Singapore, 30 Medical Drive, Singapore 117609
4 Garvan Institute of Medical Research, St. Vincent's Hospital, 384 Victoria St. Darlinghurst, 2010, NSW, Australia
* Corresponding author. E-mail address: D.James{at}garvan.org.au.
Insulin stimulates glucose transport in fat and muscle cells by triggering exocytosis of the glucose transporter GLUT4. In order to define the intracellular trafficking of GLUT4, we have studied the internalisation of an epitope tagged version of GLUT4 from the cell surface. GLUT4 rapidly traversed the endosomal system en route to a perinuclear location. This perinuclear GLUT4 compartment did not colocalise with endosomal markers (EEA1, transferrin) or TGN38, but showed significant overlap with the TGN t-SNAREs Syntaxins 6 and 16. These results were confirmed by vesicle immunoisolation. Consistent with a role for Syntaxins 6 and 16 in GLUT4 trafficking we found that their expression was up-regulated significantly during adipocyte differentiation and insulin stimulated their movement to the cell surface. GLUT4 trafficking between endosomes and TGN was regulated via an acidic targeting motif in the carboxyl terminus of GLUT4, as a mutant lacking this motif was retained in endosomes. We conclude that GLUT4 is rapidly transported from the cell surface to a subdomain of the TGN that is enriched in the t-SNAREs Syntaxins 6 and 16 and that an acidic targeting motif in the C-terminal tail of GLUT4 plays an important role in this process.
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