Figure 2

Activation of Akt and Erk pathways in cells expressing mutant YFP-EGFR-ICD Del746. A. Panels show representative images (400X) of MCF-7 cells expressing YFP-EGFR-ICD wild type, Del746 or V948R, analyzed by immunofluorescence to detect phosphorylated Akt (pAkt). Phosphorylation of endogenous Akt was only detected in cells expressing the Del746-bearing protein. B. A similar analysis was carried out to detect phosphorylated ERK (pERK). Only cells expressing YFP-EGFR-ICD Del746 contained detectable levels of endogenous pERK. Exposure time is indicated inside the panels. DNA was counterstained with Hoechst. C. Images (400X) illustrate two morphological characteristics of Akt phosphorylation in cells expressing YFP-EGFR-ICD Del746. On one hand, pAkt showed a preferential localization to membrane ruffles, and often accumulated at the tip of lamellipodial protrusions (arrowhead). On the other hand, cells expressing high (cell#1) or low (cell#2) levels of YFP-EGFR-ICD Del746, often contained similar levels of pAkt.