An inhibitor of p38 mitogen-activated protein kinase prevents insulin-stimulated glucose transport but not glucose transporter translocation in 3T3-L1 adipocytes and L6 myotubes.
The precise mechanisms underlying insulin-stimulated glucose transport still require investigation. Here we assessed the effect of SB203580, an inhibitor of the p38 MAP kinase family, on insulin-stimulated glucose transport in 3T3-L1 adipocytes and L6 myotubes. We found that SB203580, but not its inactive analogue (SB202474), prevented insulin-stimulated glucose transport in both cell types with an IC50 similar to that for inhibition of p38 MAP kinase (0.6 microM). Basal glucose uptake was not affected. Moreover, SB203580 added only during the transport assay did not inhibit basal or insulin-stimulated transport. SB203580 did not inhibit insulin-stimulated translocation of the glucose transporters GLUT1 or GLUT4 in 3T3-L1 adipocytes as assessed by immunoblotting of subcellular fractions or by immunofluorescence of membrane lawns. L6 muscle cells expressing GLUT4 tagged on an extracellular domain with a Myc epitope (GLUT4myc) were used to assess the functional insertion of GLUT4 into the plasma membrane. SB203580 did not affect the insulin-induced gain in GLUT4myc exposure at the cell surface but largely reduced the stimulation of glucose uptake. SB203580 had no effect on insulin-dependent insulin receptor substrate-1 phosphorylation, association of the p85 subunit of phosphatidylinositol 3-kinase with insulin receptor substrate-1, nor on phosphatidylinositol 3-kinase, Akt1, Akt2, or Akt3 activities in 3T3-L1 adipocytes. In conclusion, in the presence of SB203580, insulin caused normal translocation and cell surface membrane insertion of glucose transporters without stimulating glucose transport. We propose that insulin stimulates two independent signals contributing to stimulation of glucose transport: phosphatidylinositol 3-kinase leads to glucose transporter translocation and a pathway involving p38 MAP kinase leads to activation of the recruited glucose transporter at the membrane.
Sweeney
G
G
Programme in Cell Biology, Hospital for Sick Children, University of Toronto, Toronto, Ontario, M5G 1X8 Canada.
Somwar
R
R
Ramlal
T
T
Volchuk
A
A
Ueyama
A
A
Klip
A
A
D019325:3-O-Methylglucose / Q000378:metabolism; D016475:3T3 Cells; D017667:Adipocytes / Q000187:drug effects / Q000378:metabolism*; D000818:Animals; D001692:Biological Transport / Q000187:drug effects; D017871:Calcium-Calmodulin-Dependent Protein Kinases / Q000037:antagonists & inhibitors*; D003847:Deoxyglucose / Q000378:metabolism; D004791:Enzyme Inhibitors / Q000737:chemistry / Q000494:pharmacology*; D005947:Glucose / Q000378:metabolism*; D051272:Glucose Transporter Type 1; D051274:Glucose Transporter Type 3; D051275:Glucose Transporter Type 4; D007093:Imidazoles / Q000737:chemistry / Q000494:pharmacology*; D007328:Insulin / Q000494:pharmacology*; D051379:Mice; D020928:Mitogen-Activated Protein Kinases; D009004:Monosaccharide Transport Proteins / Q000378:metabolism*; D009124:Muscle Proteins; D009132:Muscles / Q000187:drug effects / Q000378:metabolism*; D009419:Nerve Tissue Proteins; D010766:Phosphorylation; D017346:Protein Serine-Threonine Kinases / Q000378:metabolism; D011518:Proto-Oncogene Proteins; D051057:Proto-Oncogene Proteins c-akt; D011725:Pyridines / Q000737:chemistry / Q000494:pharmacology*; D048051:p38 Mitogen-Activated Protein Kinases
D004791:Enzyme Inhibitors; D051272:Glucose Transporter Type 1; D051274:Glucose Transporter Type 3; D051275:Glucose Transporter Type 4; D007093:Imidazoles; D007328:Insulin; D009004:Monosaccharide Transport Proteins; D009124:Muscle Proteins; D009419:Nerve Tissue Proteins; D011518:Proto-Oncogene Proteins; D011725:Pyridines; C495678:Slc2a1 protein, mouse; C495689:Slc2a3 protein, mouse; C495693:Slc2a4 protein, mouse; D019325:3-O-Methylglucose; D003847:Deoxyglucose; C494922:Akt2 protein, mouse; D017346:Protein Serine-Threonine Kinases; D051057:Proto-Oncogene Proteins c-akt; D017871:Calcium-Calmodulin-Dependent Protein Kinases; D020928:Mitogen-Activated Protein Kinases; D048051:p38 Mitogen-Activated Protein Kinases; D005947:Glucose; C093642:SB 203580