Differential effects of phosphatidylinositol 3-kinase inhibition on intracellular signals regulating GLUT4 translocation and glucose transport

Phosphatidylinositol (PI) 3-kinase is required for insulin-stimulated trans location of GLUT4 to the surface of muscle and fat cells. Recent evidence s uggests that the full stimulation of glucose uptake by insulin also require s activation of GLUT4, possibly via a p38 mitogen-activated protein kinase (p38 MAPK)-dependent pathway. Here we used L6 myotubes expressing Myc-tagge d GLUT4 to examine at what level the signals regulating GLUT4 translocation and activation bifurcate. We compared the sensitivity of each process, as well as of signals leading to GLUT4 translocation (Akt and atypical protein kinase C) to PI 3-kinase inhibition. Wortmannin inhibited insulin-stimulat ed glucose uptake with an IC50 of 3 nM. In contrast, GLUT4myc appearance at the cell surface was less sensitive to inhibition (IC50 = 43 nm). This dis sociation between insulin-stimulated glucose uptake and GLUT4myc translocat ion was not observed with LY294002 (IC50 = 8 and 10 mum, respectively). The sensitivity of insulin-stimulated activation of PKC xi/gimel, Akt1, Akt2, and Akt3 to wortmannin (IC50 = 24, 30, 35, and 60 nm, respectively) correla ted closely with inhibition of GLUT4 translocation. In contrast, insulin-de pendent p38 MAPK phosphorylation was efficiently reduced in cells pretreate d with wortmannin, with an IC50 of 7 nM. Insulin-dependent p38 alpha and p3 8 beta MAPK activities were also markedly reduced by wortmannin (IC50 = 6 a nd 2 nm, respectively). LY294002 or transient expression of a dominant inhi bitory PI 3-kinase construct (Delta p85), however, did not affect p38 MAPK phosphorylation. These results uncover a striking correlation between PI 3- kinase, Akt, PKC xi/gimel, and GLUT4 translocation on one hand and their se gregation from glucose uptake and p38 MAPK activation on the other, based o n their wortmannin sensitivity. We propose that a distinct, high affinity t arget of wortmannin, other than PI 3-kinase, may be necessary for activatio n of p38 MA and GLUT4 in response to insulin.