title: Activation of GLUT1 by metabolic and osmotic stress: potential involvement of AMP-activated protein kinase (AMPK).
abstract: In the rat liver epithelial cell line Clone 9, the V(max) for glucose uptake is acutely increased by inhibition of oxidative phosphorylation and by osmotic stress. By using a membrane-impermeant photoaffinity labelling reagent together with an isoform-specific antibody, we have, for the first time, provided direct evidence for the involvement of the GLUT1 glucose transporter isoform in this response. Transport stimulation was found to be associated with enhanced accessibility of GLUT1 to its substrate and with photolabelling of formerly 'cryptic' exofacial substrate binding sites in GLUT1 molecules. The total amount of cell surface GLUT1 remained constant. The precise mechanism for this binding site 'unmasking' is unclear but appears to involve AMP-activated protein kinase: in the current study, osmotic and metabolic stresses were found to result in activation of the alpha 1 isoform of AMP-activated protein kinase, and transport stimulation could be mimicked both by 5-aminoimidazole-4-carboxamide ribonucleoside and by infection of cells with a recombinant adenovirus encoding constitutively active AMP-activated protein kinase. The effect of 5-aminoimidazole-4-carboxamide ribonucleoside, as for metabolic stress, was on the V(max) rather than on the K(m) for transport and did not affect the cell-surface concentration of GLUT1. The relevant downstream target(s) of AMP-activated protein kinase have not yet been identified, but stimulation of transport by inhibition of oxidative phosphorylation or by 5-aminoimidazole-4-carboxamide ribonucleoside was not prevented by either inhibitors of conventional and novel protein kinase C isoforms or inhibitors of nitric oxide synthase. These enzymes, which have been implicated in stress-regulated pathways in other cell types, are therefore unlikely to play a role in transport regulation by stress in Clone 9 cells.
authors: School of Biochemistry and Molecular Biology, University of Leeds, Leeds LS2 9JT, UK. K.Barnes@leeds.ac.uk
authors: Kay
authors: K
authors: Barnes
authors: Jean C
authors: JC
authors: Ingram
authors: Omar H
authors: OH
authors: Porras
authors: L Felipe
authors: LF
authors: Barros
authors: Emma R
authors: ER
authors: Hudson
authors: Lee G D
authors: LG
authors: Fryer
authors: Fabienne
authors: F
authors: Foufelle
authors: David
authors: D
authors: Carling
authors: D Grahame
authors: DG
authors: Hardie
authors: Stephen A
authors: SA
authors: Baldwin
mesh_terms: D055372:AMP-Activated Protein Kinases; D000620:Aminoimidazole Carboxamide / Q000031:analogs & derivatives* / Q000494:pharmacology; D000818:Animals; D002478:Cells, Cultured; D004791:Enzyme Inhibitors / Q000494:pharmacology; D005947:Glucose / Q000378:metabolism*; D051272:Glucose Transporter Type 1; D022781:Hepatocytes / Q000166:cytology / Q000201:enzymology*; D007004:Hypoglycemic Agents / Q000494:pharmacology; D009004:Monosaccharide Transport Proteins / Q000187:drug effects / Q000378:metabolism*; D009097:Multienzyme Complexes / Q000378:metabolism*; D019001:Nitric Oxide Synthase / Q000037:antagonists & inhibitors / Q000378:metabolism; D009997:Osmotic Pressure; D010085:Oxidative Phosphorylation / Q000187:drug effects; D020033:Protein Isoforms / Q000037:antagonists & inhibitors / Q000378:metabolism; D011493:Protein Kinase C / Q000037:antagonists & inhibitors / Q000378:metabolism; D017346:Protein Serine-Threonine Kinases / Q000378:metabolism*; D051381:Rats; D012265:Ribonucleotides / Q000494:pharmacology; D013312:Stress, Physiological / Q000378:metabolism*
chemical_list: D004791:Enzyme Inhibitors; D051272:Glucose Transporter Type 1; D007004:Hypoglycemic Agents; D009004:Monosaccharide Transport Proteins; D009097:Multienzyme Complexes; D020033:Protein Isoforms; D012265:Ribonucleotides; C495679:Slc2a1 protein, rat; D000620:Aminoimidazole Carboxamide; D019001:Nitric Oxide Synthase; D017346:Protein Serine-Threonine Kinases; D011493:Protein Kinase C; D055372:AMP-Activated Protein Kinases; C031143:AICA ribonucleotide; D005947:Glucose