Depletion of Coro1B promotes autophagy and V-ATPase lysosomal localization. (A) Confocal images of late L2 larval fat body cells expressing GFP-mCherry-Atg8a in control (Ctrl) and Coro knockdown larvae. Scale bar, 20 μm. (B) Quantification of the ratio of autolysosomes (GFP⁻ mCherry⁺) to total Atg8a puncta per cell in A; data are shown as mean ± S EM, n = 3, ≥ 20 cells. (C) Confocal images of wL3 larval fat body cells from flies expressing mCherry-Atg8a and indicated transgenes driven by Cg-Gal4. Scale bar, 20 μm. (D) Quantification of Atg8a puncta number and size shown in C. Data are shown as mean ± SEM, n = 3, ≥ 15 cells. (E) Immunofluorescence analysis of colocalization of V-ATPase (V1H) and lysosome (LAMP1) in control and Coro1B knockdown HeLa cells. Scale bar, 20 μm (original) and insets, 3x zoom. (F) Quantification of the colocalization of V1H and LAMP1 in E. Pearson’s correlation coefficient and colocalization rate were analyzed by ImageJ. Data are shown as mean ± SEM, n = 3, ≥ 20 cells. (G) Immunofluorescence analysis of colocalization of V-ATPase (V1H) and lysosome (LAMP1) in control and Coro1B knockdown cells treated with LatA (200 nM) or Arp2/3 inhibitor CK666 (200 μM) for 2 h. Scale bar, 20 μm (original) and insets, 2x zoom. (H) Quantification of the colocalization of V1H and LAMP1 in G. Data are shown as mean ± SEM, n = 3, ≥ 25 cells. Significance was determined using one-way ANOVA and Tukey’s multiple comparisons test (D and H), and Student’s t test (B and F); ****P < 0.0001; NS not significant.