Fig. 7. The effect of constitutively active PKB constructs on IRAP-GFP translocation in 3T3-L1 adipocytes. 3T3-L1 adipocytes were microinjected with plasmids encoding IRAP-GFP (A-C) and either constitutively active PKB (A; PKB[DD]), constitutively active PKB fused to the N-terminus of GLUT4 (B; PKB[DD]-Glut4) or a constitutively active myristoylated PKB (C; Myr-PKB). 16 hours later the cells were serum starved for 2 hours and then fixed and stained with anti-HA antibodies to visualise the PKB constructs (as indicated). Panels A-C show the distribution of the HA-tagged PKB constructs and GFP fluorescence resulting from the expression of the IRAP-GFP construct. Bars, 10 µm. In D, cells were scored for the presence of IRAP-GFP in the plasma membrane after incubation in the absence or presence of insulin for 30 minutes as discussed in the Materials and Methods. The results shown represent means±s.d. for three independent experiments with each condition being represented by a minimum of 50 cells. ^*P<0.01 versus the vector only injected cells incubated in the presence of insulin.