Figure 3.

Proteomic analysis of MEK inhibition in breast cancer cells. A, heat map of protein and phosphoprotein expression profiles. MDAMB231 cells were treated with MEK inhibitor U0126 (10 µmol/L) as described in Materials and Methods, and 30 min later, they were stimulated with EGF (10 ng/mL). The protein lysates were collected at 1, 4, and 24 h after EGF addition and analyzed by RPPA. Values are expressed as log₂ fold difference from control (untreated) samples at each time point. B, relative expression changes of AKTpS473, EGFRpY1068, phosphorylated MAPK (p-MAPK), and Cyclin D1 as detected by RPPA. Gray columns, EGF treatment; black columns, EGF + U0126 treatment. Control = 0. Values for phosphoproteins were normalized by corresponding total protein levels. C, conventional Western blot analysis of p-AKT and p-ERK (p-MAPK) expression in response to U0126 treatment in T47D and MDAMB231 cell lines. Cells were treated with MEK inhibitor in the same conditions as for RPPA experiment and the protein lysates were collected at 1 and 4 h after EGF stimulation.