GABARAPs participate in mTOR inactivation but not in eIF2α phosphorylation in response to lysosomal damage. (A) Immunoblot analysis of the phosphorylation ULK1 (S757), S6K (T389), 4EBP1 (S65), and eIF2α (S51) in parental HeLa (WT), LC3^TKO, GBRP^TKO, and hexa^KO cells treated with 4 mM LLOMe for 30 min. (B ⅰ–ⅳ) Quantification of phosphorylation of ULK1 (S757; i), S6K (T389; ii), 4EBP1 (S65; iii), and eIF2α (S51; iv) in A; Quantification, n = 3. (C) Quantification by HCM and confocal microscopy analysis of overlaps between mTOR and LAMP2 in parental HeLa (WT), LC3^TKO, GBRP^TKO, and hexa^KO cells treated with 4 mM LLOMe for 30 min. HCM images in Fig. S4 M. Scale bar, 5 µm. (D) Quantification by HCM of G3BP1 puncta. Parental HeLa (WT), LC3^TKO, GBRP^TKO, and hexa^KO cells were treated with 4 mM LLOMe for 30 min. Red masks, computer-identified G3BP1 puncta. Data, means ± SEM; HCM: n ≥ 3 (each experiment: 500 valid primary objects/cells per well, ≥5 wells/sample). †, P ≥ 0.05 (not significant); **, P < 0.01, ANOVA. See also Fig. S4. Source data are available for this figure: SourceData F8.