Figure 7.

ATG9A protein regulates adhesion dynamics. (A–D) ATG9A depletion reduces intrinsic adhesion dynamics. (A) Representative live-cell TIRF images of U87 MG cells expressing PXN-EGFP together with the indicated siRNAs. Image sequences show adhesion changes over a 40-min period. Righthand panels: Color scale output generated from FAAS, representing early (blue) to late (red) adhesions. Scale bar, 20 µm. (B–D) FAAS output of the number of dynamic adhesions (B), assembling adhesions (C), and disassembling adhesions (D). Data represent means and SEM (siCont, n = 13 cells; siATG9A #2, n = 13 cells; from five independent experiments). (E) ATG9A depletion inhibits EGF-induced formation of adhesion complexes. U87 MG cells were transfected with the indicated siRNAs. After starvation (1 h) in serum-free DMEM, cells were treated (1 h) with or without EGF (50 ng/ml), fixed, and labeled with an anti-PXN antibody (green) and DAPI (nuclei, blue). The number of adhesion complexes was quantified for each cell. Data represent means and SEM (n = 144–158 cells per group; cells from independent experiments were color-coded). Scale bar, 20 µm; inset magnifications, 10 µm. Statistical significance was evaluated using Mann–Whitney U test (B–D) or one-way ANOVA followed by Sidak post hoc test (E). **, P < 0.01; ***, P < 0.001.