Fig. 6

FTD patient-derived i³Neurons with mutant GRN reveal altered protein turnover of lysosomal enzymes and FTD-associated proteins. A Generation of a set of FTD patient fibroblast-derived i³Neurons. First, CRISPR-Cas9 was used to insert an inducible NGN2 cassette into the AAVS1 locus of a patient fibroblast-derived iPSC line (ptMut). Next, CRISPR-Cas9 was used to correct the GRN mutation in ptMut to create an isogenic control iPSC line (ptWT) and then to knockout GRN in pWT to create the ptKO iPSC line. These iPSC lines were then differentiated into i³Neurons and dSILAC proteomics was performed. B Volcano plot of protein half-life changes in ptKO vs. ptWT i³Neuron. C Volcano plot of protein half-life changes in ptMut vs. ptWT i³Neuron. D Principal component analysis using protein half-lives in GRN-KO, WT, ptKO, ptMut, and ptWT i³Neurons groups. E Hierarchical clustering of five i³Neurons groups with five biological replicates in each group. F Scatter plot of protein half-life changes in ptKO vs. ptWT and ptMut vs. ptWT comparisons showing the consistency and potential gene dosage effect of ptKO and ptMut i³Neurons. G Heatmap showing key overlapping protein turnover changes in GRN KO vs. WT, ptKO vs. ptWT, and ptMut vs. ptWT i³Neurons. Heatmap colors represent the absolute half-life differences in days between comparison groups. Key proteins from lysosomes and relevant to FTD/ALS are highlighted in red and blue, respectively. H Schematic of proposed lysosomal impairment in progranulin-deficient neurons caused by GRN mutations in FTD patients