Fig. 3: Impact of proteasome inhibition and lysosome acidification impairment on protein ubiquitylation in iNeurons.
From: Aging and diet alter the protein ubiquitylation landscape in the mouse brain
A Experimental scheme for iNeurons drug treatment and characterization (N = 4, biological replicates from two independent differentiation batches). Created in BioRender. Ori, A. (2025) https://BioRender.com/2947h3v. B PCA based on proteome data from iNeurons. The ellipses (drawn manually) highlight the differentiation between day groups. The percentage of variance explained by each principal component is indicated. C PCA based on ubiquitylated peptide abundances from iNeurons. The ellipses highlight the drug treatments. The percentage of variance explained by each principal component is indicated. D Barplots showing the number of identified ubiquitylated sites in different sample groups (N = 4 except N = 3 for 14 days DMSO and 7 days bortezomib, biological replicates, data shown as averages ± SD). Gene set enrichment analysis (GSEA) for ubiquitylated peptides affected by bortezomib (E) or bafilomycin (F) in 14-day iNeurons based on GO cellular component terms (Adj.P < 0.05, weighted Kolmogorov–Smirnov test). G Correlation between changes of ubiquitylation observed during mouse brain aging and 14-day iNeurons treated with bortezomib or bafilomycin (P < 0.05 in both datasets, two-sided Pearson’s correlation test). H Scatterplot comparing ubiquitylation changes observed in mouse brain aging and 14-day iNeurons treated with bortezomib (upper panel) or bafilomycin (lower panel) (P < 0.05 in both datasets, two-sided Pearson’s correlation test). I Heatmap of ubiquitylation changes occurring in neurodegeneration-associated proteins. Sites significant (P < 0.05, Spectronaut differential abundance test) in at least one of the compared datasets are shown. The lysine numbering refers to the mouse protein sequence. In this figure, mouse ubiquitylation changes were corrected for protein abundance changes. *P ≤ 0.05; **P ≤ 0.01, ***P ≤ 0.001, ****P ≤ 0.0001. Source data are provided as a Source Data file. Specific P values are listed in Supplementary Data 8. Related to Figs. S6–S8, and Supplementary Data 4 and 5.
