Fig. 3 | Nature Communications

Fig. 3

From: Deregulated Gab2 phosphorylation mediates aberrant AKT and STAT3 signaling upon PIK3R1 loss in ovarian cancer

Fig. 3

Gab2 is required in PIK3R1 loss-induced signaling and oncogenic phenotypes. a Total cell lysate of SKOV3 cells transfected with either PIK3R1 siRNA, GAB2 siRNA or in combination for 72 h were harvested for western blotting. b SKOV3 cells were transfected with PIK3R1 siRNA or co-transfected with GAB2 siRNA. After 24 h, cells were transfected with 4xM67 pTATA TK-Luc (containing STAT3-binding site) with Renilla luciferase vector for another 24 h. Dual-luciferase reporter assay was performed. c Total RNA of SKOV3 cell transfected with PIK3R1 siRNA or in combination with GAB2 siRNA for 72 h were harvested for real-time PCR. d SKOV3 cells were transfected with PIK3R1 siRNA or co-transfected with GAB2 siRNA, cell viability was measured over 7 days. e Representative images of migrated or invaded SKOV3 cells (upper) and means with SD of migrated or invaded cells of five fields at magnification of 100 × (lower). Scale bar, 200 μm. f Total lysates of transfected SKOV3 cells were immunoprecipitated with Gab2 antibody prior to western blotting. g SKOV3 cells were transfected with PIK3R1 siRNA prior to proximity ligation assay with the indicated pairs of antibodies. Representative images are presented (upper) with numbers of averaged spots per nucleus (lower). Scale bar, 50 μm. The numbers below the blots represent the mean values from densitometry readings of three independent experiments. Error bars represent SD. **p < 0.005; ***p < 0.001; ns, no significant difference compared with mock using t-test

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