Figure 4
Expression of RhoA-F suppresses the ability of P61A6 to inhibit proliferation. A. Western blotting with anti-HA antibody showing expression of stably transfected 3xHA-RhoA and 3xHA-RhoA-F constructs in H358 cells. B. Stably transfected H358 cells were treated with DMSO or 10 μM P61A6 for 48 hours. Membrane and cytosolic fractions were prepared and examined for RhoA (Left Panel) and RhoA-F (Right Panel) localization on membrane and cytosolic fractions by Western blotting. Na+/K + ATPase and GAPDH were used as control for membrane and cytosolic fractions, respectively. C. H358 RhoA and E. H358 RhoA-F cells were treated with DMSO or P61A6 at different concentrations (2.5 μM, 5.0 μM and 10 μM) under low serum conditions (medium with 0.5% FBS) for 10 days. Cell proliferation assay was performed in triplicate every 2nd day for 10 days. D. Differences in proliferation rate between cells treated with different concentrations of P61A6 and DMSO-treated control is characterized by increasing statistical significance.