Fig. 1

The glaucoma associated OPTN(E50K) mutation alters endogenous protein modification and aggregates in hPSC-RGCs. A Real-time RT-PCR quantification of OPTN mRNA levels (n = 3 for each WT and E50K; t-test, p = 0.181). B–C Western blot and the relative protein expression of OPTN and p62 to β-actin in hPSC-RGCs (n = 5 for each WT and E50K; t-test, OPTN ****p < 0.0001, p62 ***p = 0.0002). D Proteomics analysis demonstrated changes in the expression of autophagy-associated proteins in OPTN(E50K) hPSC-RGCs. E–F Immunostaining displayed the expression of OPTN puncta in BRN3:tdTomato hPSC-RGCs from WT and E50K under steady state (control) and chloroquine (CQ) treatment. Arrows indicate the OPTN puncta. Scale bar: 10 μm. G–H Quantification of OPTN puncta in hPSC-RGCs. I–J Immunostaining displayed the expression of p62 puncta in BRN3:tdTomato hPSC-RGCs from WT and E50K under control and CQ treatment. K–L Quantification of p62 puncta in hPSC-RGCs. G–H and K–L: n = 3 biological replicates using Ctrl-WT n = 60, Ctrl-E50K n = 60, CQ-WT n = 57 and CQ-E50K n = 48 technical replicates; One-way ANOVA, Tukey post hoc test. ****p < 0.0001, ***p < 0.001, **p < 0.01, *p < 0.05, ns = not significant, p > 0.05). Data are represented as mean values ± SEM