Fig. 8

Endosome maturation, but not autophagy is impaired in C9 KO MNs. a Representative Western blot of iPSC-derived MNs treated with either DMSO, the autophagy inhibitor Bafilomycin A1 (Baf A1), the autophagy inducer Torin 1 or a combination of the latter, immunostained for LC3-I and LC3-II, p62, LAMP1, EEA1, Rab7 and β-actin/GAPDH, used to measure levels of autophagosomes, aggregates, lysosomes, early endosomes, late endosomes or for protein normalization respectively. b, c Quantifications of the Western blot shown in (a), measuring the relative levels of p62 (b) and the ratio of LC3-II/LC3-I (c); each dot represents one biological replicate. d, e Quantifications of the Western blot shown in (a), measuring the levels of p62 (d) or LC3-II (e) after treatment with Baf A1 and dividing those with the levels of either p62 or LC3-II without treatment; each dot represents one biological replicate. f–h Quantifications of the Western blot shown in (a), measuring the basal relative levels of LAMP1 (f), Rab7 (g) and EEA1 (h); each dot represents one biological replicate. i–k Quantification of the survival rate after 24 h (i), 48 h (j) or 72 h (k) treatment with Baf A1. The survival rate of iPSC-derived MNs is calculated by normalizing the CellTiter-Glo® signal of Baf A1-treated cells to DMSO control-treated wells; each dot represents the average of one biological replicate. Data represent mean ± SEM; data are pooled from six (b–h) or four (i–k) independent differentiations. Statistical significance was assessed by unpaired t-test (b–h) or one-way ANOVA and Tukey’s multiple comparison test (i–k); *p < 0.05, **p < 0.01