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Fig. 6. PAK4-induced rounding and PAK4wt localisation is prevented by PI 3K inhibitors (A) Cells were microinjected with a plasmid encoding PAK4{Delta}GBD and incubated for 2.5 hours in 0.2% FCS. Cells were then incubated for 30 minutes with LY294002. Cells were then incubated for 20 hours in the presence of HGF and LY294002. (B) Cells were microinjected with a plasmid encoding PAK4{Delta} and incubated for 2.5 hours in 0.2% FCS. Cells were then incubated for 30 minutes with LY294002, prior to further incubation in 0.2% FCS plus LY294002 for 20 hours. (C) Cells were microinjected with a plasmid encoding PAK4{Delta}GBD and incubated for 2 hours in 0.2% FCS. Cells were then incubated for 30 minutes with LY294002 prior to a further incubation in 0.2% FCS plus LY294002 for 2.5 hours. (D) Cells were microinjected with a plasmid encoding PAK4wt and incubated for 2 hours in 0.2% FCS. Cells were then incubated for 30 minutes with LY294002 prior to stimulation for 2.5 hours with HGF and LY294002. All the cells were fixed and stained for HA-tagged PAK4 and F-actin (E-H). Basal sections (at the level of stress fibres) of PAK4{Delta}GBD-expressing cells are shown, while apical sections (at the level of F-actin-containing microvilli) of PAK4{Delta}-expressing cells are shown in order to visualise clearly the rounded-up cells (arrows). Bar, 10 µm.