Fig. 5

Palmitate inhibits ER-phagy via upregulation of Bcl-2. a–f Cells were starved in EBSS for 3 h in the presence or absence of palmitate (0.1 mM). a and b Representative micrographs (a) and quantification (b) of the average number of mRFP-LC3 puncta in cells transiently expressing mRFP-LC3 (n = 12 per each group). Scale bar, 10 μm. Data are mean ± SEM; **p = 0.0032, ****p < 0.0001 vs. control, ^####p < 0.0001, ^†p = 0.0358. c and d Representative micrographs (c) and quantification (d) of the average number of GFP-DFCP1 puncta in cells transiently expressing GFP-DFCP1 (n = 14 per each group). Scale bar, 10 μm. Data are mean ± SEM; ****p < 0.0001 vs. control, ^####p < 0.0001, ^††††p < 0.0001. e and f Immunoblotting analysis (e) and quantification (f) of Bcl-2 and Beclin1 (Bcl-2; n = 6 per each group, Beclin1; n = 4 per each group). Data are mean ± SEM; Bcl-2; ****p < 0.0001 vs. control, ^####p < 0.0001, ^††††p < 0.0001, Beclin1; ***p = 0.0007 vs. control, ^###p = 0.0007, ^†††p = 0.0005. g–j Cells were transfected with si-scram or si-Bcl-2 were starved in EBSS for 3 h in the presence or absence of palmitate (0.1 mM). g and h Immunoblotting analysis (g) and quantification (h) of Bcl-2 (n = 6 per each group). Data are mean ± SEM; **p = 0.0089 at EBSS + PA in si-scram vs. EBSS in si-scram, **p = 0.0054 at EBSS in si-Bcl-2 vs. EBSS in si-scram, ^††††p < 0.0001. i and j Representative micrographs (i) and quantification (j) of the average number of GFP-DFCP1 puncta in cells transiently expressing GFP-DFCP1 (n = 11 per each group). Scale bar, 10 μm. Data are mean ± SEM; ****p < 0.0001 vs. EBSS in si-scram, ^††††p < 0.0001. n.s., no significant difference