Fig. 5.

Insensitivity to erlotinib in PIK3R2-depleted cells correlates with p38 MAPK nuclear translocation and p38 MAPK-mediated DNA repair.

(A-B) SKOV3 cells transfected with or without PIK3R2 siRNA were treated with erlotinib alone (10 μM) or in combination with losmapimod (10 μM) for 72 h. (A) An anti-p38 MAPK antibody (green) and DAPI (blue) were used for immunofluorescence staining. Scale bar, 20 μm. (B) Cells were subjected to subcellular fractionation. Representative blots of 3 independent experiments are shown. The numbers below the blots indicate the mean densitometry values normalized to those of GAPDH (cytoplasmic marker) or lamin A/C (nuclear marker). (C-D) SKOV3 cells stably expressing PIK3R2 shRNA or vector control were treated with erlotinib alone (10 μM) or in combination with losmapimod (10 μM) for (C) 24 h or (D) 72 h. DNA damage was visualized by comet assay (Left). Scale bar, 100 μm. The amount of damaged DNA was expressed as percentage of DNA in comet tail (% tail DNA) (right). The box and whisker plot was plotted by Graphpad Prism software using the Tukey method. *, P < 0.05; **, P < 0.005; ***, P < 0.001; ^#, P < 0.0001 using ordinary one-way ANOVA for analysis within group (vector or PIK3R2 shRNA) and 2-way ANOVA for comparison between groups (vector vs PIK3R2 shRNA) with Sidak's multiple comparison test.