Figure 5. EGFR-V665M is transforming in NIH3T3 cells.
A. NIH3T3 cells stably expressing the indicated wild-type or mutated forms of EGFR (or pBabe-puro vector only) were seeded at a density of 8×103 cells per well in a standard colony forming assay (see Experimental Procedures). Cells were incubated for 3 weeks in the absence or presence of 50ng/ml EGF, and colonies were then counted. The results are presented as the means of 3 experiments ± standard deviation. To control for possible differences in expression level of the mutants, NIH3T3 stably expressing each mutant were lysed and analyzed by SDS-PAGE and immunoblotting with anti-EGFR. B. Upper panel: Example of tumors isolated from nude mice 3 months after s.c. injection with NIH3T3 cells stably expressing wild-type or V665M EGFR or the pBabe-puro vector. Lower panel: Volumes of tumors derived from the cells indicated above were evaluated 3 months after s.c. injection. Open circles represent single mice with a total of 4 mice per cell type analyzed.