Figure 5

Inhibitors of MEK1/2, but not of PI3K, inhibits morphological transformation and restores E-cadherin expression in IEC-6 cells transformed by Tpr-Met, TM-Grb2, TM-Shc1 and TM-Shc2 oncoproteins. (A) Basal phosphorylation and expression levels of Erk1/2 and Akt were assessed by IB analyses of TCL prepared from the indicated serum-starved IEC-6 cells. (B) The indicated cells were treated with vehicle (DMSO), or 10 μM of MEK1/2 and/or PI3K inhibitors. Photographs show typical morphologies following 48 hours of treatment. Similar morphological changes were observed after 24 hours of treatment (Additional file 3). (C) E-cadherin and actin protein levels were assessed by IB analysis after 48 hours of treatments with DMSO or the indicated inhibitors. (D) Relative E-cadherin and actin protein levels were determined by densitometry analysis. The bar graph shows the mean ± S.D. fold-changes in E-cadherin protein levels normalized to that of actin relative to DMSO treated cells. The values were calculated from at least three independent experiments. (E) The MEK1/2 and PI3K inhibitors were validated to selectively block serum-induced Erk1/2 and Akt phosphorylation in Tpr-Met-IEC-6 and Control-IEC-6 cells. Serum-starved cells were treated for 1 hour with DMSO or indicated inhibitors, prior to 5 minutes of stimulation with 10% serum. Erk1/2 and Akt phosphorylation and total protein levels were heaevaluated as above. Actin levels were used as a loading control. The results are representative of at least 2 independent experiments.