Fig. 3

Constitutive activation of ERK/MAPK pathway, but not PI3K pathway, prevents autophagy induced by FGFR1 inhibition. a and b ERK and AKT phosphorylation by western blot in H1581 and H520 cells stably transduced by two independent FGFR1 shRNA constructs or a non-targeting control vector. FGFR1 protein expression levels were included as a control of knockdown efficiency. The filters were stripped and reprobed for total ERK and AKT to ensure equal loading of cell protein in each lane. c H1581 cells were treated with 1 μM AZD4547 for 24 h. FGF2 (25 ng/ml) was added for 2 h before harvesting. d H520 cells were treated with AZD4547 (1 μM) for 24 h. Cells were treated with FGF2 for 2 h before harvesting and then subjected to western blotting analysis. e Quantification of LC3-II levels in (c); mean ± SD, n = 3, ***p < 0.001. f Quantification of LC3-II levels in (d); mean ± SD, n = 3, ***p < 0.001. g Western blot showing that the expression of a caAKT1 construct failed to prevent increase in LC3-II induced by shFGFR1 in H1581 cells. h Western blot showing that the expression of a caMEK1 prevented increase in LC3-II induced by shFGFR1 in H1581 cells. i Quantification of LC3-II levels in (g); mean ± SEM, n = 3, NS, not significant. j Quantification of LC3-II levels in (h); mean ± SEM, n = 3, ***p < 0.001