Figure 5.

Insulin-stimulated phosphorylation of Akt and of GSK3 are decreased in nelfinavir-treated-, p110-CAAX-, and myc-GLUT4-GFP-expressing adipocytes. 3T3-L1 adipocytes expressing a membrane-targeted catalytic subunit of PI 3-kinase (p110-CAAX) and myc-GLUT4-GFP were generated and treated or not with nelfinavir (Nelf) as in the legend for Fig. 4. After immunostaining surface myc in nonpermeabilized cells, cells were permeabilized and incubated with anti-pSer473-PKB/Akt (A) or pSer9-GSK3 (B), followed by an alexa 633-conjugated secondary antibody. Confocal microscopy was performed, with acquisition in three channels. These experiments were repeated three times, and quantification of the intensity of the pGSK signal in nelfinavir treated and control cells (n = 30 in each experiment) is shown in C in arbitrary units. *, P < 0.05.