mTOR hyperactivation in Down Syndrome underlies deficits in autophagy induction, autophagosome formation, and mitophagy

a Electron microscopy analysis of 2？N and DS cells; section indicated by the square is shown at higher magnification in the right panels. Black arrow points to example of an abnormal mitochondrion. Quantification shown as percentage of damaged mitochondria in respect to total number. Scale bar 500？nm. b Quantification of MitoSOXTM intensity fluorescence reported as fluorescence arbitrary units (A.U.) for three independent experiments. The analysis was done using the software ImageJ. c Quantification of mitochondrial mass using Mitotracker green intensity fluorescence, acquired through SpectraMax M5 Microplate Reader and reported as fluorescence arbitrary units (A.U.) from at least three independent experiments. d Gene set analysis barcode plot of transcriptomic changes of transcriptional factors (TFs) regulating mitochondrial biogenesis7,42,43 (for list of genes, see Supp. Table 1) in DS fibroblasts compared to 2？N fibroblasts. The vertical bars represent TFs (genes) and are ranked horizontally by moderated t-statistics (x-axis). Pink, blue and grey shaded rectangles mark upregulated, downregulated and genes that are unchanged, respectively. The black trace above the bars shows relative enrichment of genes using Limma’s ROAST function (see Methods section). The analysis shows significant downregulation of TFs regulating mitochondrial biogenesis in DS compared to 2？N fibroblasts. (p？<？0.001; FDR？<？0.001).. Statistical analysis in A, B and C was performed using Student’s T-test. (*p？<？0.05; **p？<？0.01; ***p？<？0.001