Fig. 3. Significant reduction in cell numbers of Src*/CasSD expressing cells.

A: MCF-7 and TAM-R cells stably and constitutively expressing Src*/CasSD or control vector were plated at density of 10,000 cells/well. Cell numbers in TAM-R cell populations were determined by measuring ATP at day 1 and 3 after plating (left panel). TAM-R cell populations stably and constitutively expressing Src*, Src ^KM/CasSD or Src*/CasSD were plated at a density of 10,000 cells/well. Cell numbers in all cell populations were determined at days 2 and 4 after plating (right panel). Results indicated are representative of three independent experiments performed in triplicate ± standard deviation. P values were calculated using Student’s t-test. *** indicates p<0.0008 and ** indicates p<0.03, vector vs. Src*/CasSD. B: Expression of Src*/CasSD influences the protein expression of EGFR, ERBB2. Serum-starved MCF-7 and TAM-R cells stably and constitutively expressing the indicated constructs were subjected to WB analyses for ERBB family proteins (50 μg WCE), and tubulin for loading (representative of three independent experiments). C: Src*Cas/SD expression attenuates ERK activation. Serum-starved MCF-7 and TAM-R Src*/CasSD and vector stable cell populations were stimulated with 10 ng/ml of epidermal growth factor (EGF). WCE (20 μg) were analyzed by WB with pERK1/2, ERK1/2 and actin antibodies. Data are representative of two independent experiments. D: Expression of Src*/CasSD influences the phosphorylation of endogenous p130^Cas. WCE of TAM-R Src*/CasSD and vector control cell populations were analyzed by WB for expression of endogenous p130^Cas. Arrows indicate positions of phosphorylated and unphosphorylated p130^Cas (upper and lower, respectively). Densitometric analysis of three independent WBs was used to calculate the ratio of phosphorylated vs. unphosphorylated p130^Cas (right panel).