FIGURE 7.
EGF-stimulated activation of ERK-1/2 occurs via a divergent signaling pathway compared with S1P in MDA-MB-453 cells. MDA-MB-453 cells were pretreated with AG 1478 (0.1 or 1 μm, 15 min) or siRNA EGF receptor (100 nm, 48 h) prior to stimulation with and without S1P (5 μm) or EGF (50 ng/ml) for 10 min. a and c, Western blot showing the effect of AG 1478 (a) and siRNA (c) knockdown of EGF receptor on the EGF- and S1P-stimulated activation of ERK-1/2. Also shown in c is the siRNA knockdown of EGF receptor (Mr = 170 kDa) detected on Western blots probed with anti-EGF receptor antibody. Bar graphs showing quantification of the effect of AG 1478 (b) and siRNA (d) knockdown of EGF receptors on EGF- and S1P-stimulated ERK-1/2 activation. Results in b and d are expressed as P-ERK-1/2:ERK-2 ratios (mean ± S.D.) for n = 3 experiments. In a and c, phosphorylated ERK-1/2 was detected on Western blots probed with anti-phosphorylated ERK-1/2 antibody. Blots were also probed with anti-ERK-1/2 antibody to ensure equal protein loading.