title: Insulin-triggered repositioning of munc18c on syntaxin-4 in GLUT4 signalling.
abstract: One of the most important actions of insulin is the stimulation of the uptake of glucose into fat and muscle cells. Crucial to this response is the translocation of GLUT4 (glucose transporter-4) to the plasma membrane. The insulin-stimulated GLUT4 vesicle docking at the plasma membrane requires an interaction between VAMP-2 (vesicle-associated membrane protein-2) on the GLUT4 vesicle and syntaxin-4 in the plasma membrane. In the basal state, munc18c is thought to preclude GLUT4 vesicle docking by inhibiting this interaction. Here, we have used FCS (fluorescence correlation spectroscopy) in single living cells to show that munc18c binds to syntaxin-4 in both the basal and insulin-stimulated states. We show that munc18c contains two binding sites for syntaxin-4, one of which is disrupted by insulin, while the other is activated by insulin. Insulin-triggered repositioning of munc18c on syntaxin-4 in this way in turn allows syntaxin-4 to adopt its 'open' conformation and bind VAMP-2, resulting in the docking of the GLUT4 vesicle at the cell surface. The results also demonstrate the utility of using FCS in intact single living cells to elucidate cell signalling events.
authors.affiliation: School of Biological Sciences, University of Southampton, Biomedical Sciences Building, Bassett Crescent East, Southampton SO16 7PX, U.K.
authors.forename: Natalie P
authors.initials: NP
authors.lastname: Smithers
authors.forename: Conrad P
authors.initials: CP
authors.lastname: Hodgkinson
authors.forename: Matt
authors.initials: M
authors.lastname: Cuttle
authors.forename: Graham J
authors.initials: GJ
authors.lastname: Sale
mesh_terms: D016475:3T3 Cells; D017667:Adipocytes / Q000502:physiology; D000818:Animals; D016466:CHO Cells; D006224:Cricetinae; D003412:Cricetulus; D018274:Electroporation; D051275:Glucose Transporter Type 4 / Q000502:physiology*; D005982:Glutathione Transferase / Q000235:genetics / Q000378:metabolism; D007328:Insulin / Q000494:pharmacology*; D051379:Mice; D018613:Microscopy, Confocal; D051938:Munc18 Proteins / Q000502:physiology*; D050765:Qa-SNARE Proteins / Q000502:physiology*; D011994:Recombinant Proteins / Q000378:metabolism; D015398:Signal Transduction; D014162:Transfection
chemical_list: D051275:Glucose Transporter Type 4; D007328:Insulin; D051938:Munc18 Proteins; D050765:Qa-SNARE Proteins; D011994:Recombinant Proteins; D005982:Glutathione Transferase