FIGURE 4:

Insulin-dependent GLUT4 translocation is prevented by silencing MICAL-L2 or expressing MICAL-L2-CT. (A) L6-GLUT4myc cells were transfected with GFP-coexpressing vectors containing shRNA interference to rat MICAL-L2 (sh-MICAL-L2) or unrelated shRNA. Cells were replated on coverslips for 48 h, serum starved, and stimulated with insulin (15 min) or not. Surface GLUT4myc was detected with anti-myc and Alexa 555–secondary antibodies (red), imaged by confocal microscopy, and quantified as in Materials and Methods. Fold changes relative to shRNA control from three independent experiments, >25 cells/experiment (mean ± SE, ***p < 0.001 for n = 3). (B) L6-GLUT4myc cells transfected with GFP-MICAL-L2-CT or GFP were stimulated with insulin, and surface GLUT4myc was detected as in A. Results from four experiments, >25 cells/experiment (mean ± SE, **p < 0.01).