Fig. 1: Mitophagic flux is impaired in Drosophila expressing human wild-type or mutant α-synuclein.

a Western blot for α-synuclein on extracts from thoraces of 4-week-old flies expressing wild-type or mutant human α-synuclein (w1118;;UAS-mito-Keima,mef-2-GAL4/UAS-SNCA X) and control flies (CTRL1: w1118;;UAS-mito-Keima,mef-2-GAL4/+). b Quantification of the α-synuclein/GAPDH ratio in (a) (n = 5 for all conditions, except for CTRL2 where n = 3; P = 0.94, one-way ANOVA with post hoc Tukey’s test). CTRL2 flies are w1118;;UAS-mito-Keima,mef-2-GAL4/UAS-smGdP. c Confocal images showing overlay of live mito-Keima emission at 458 nm (green) and 543 (red) excitation in indirect flight muscle of 4-week-old CTRL1, CTRL2, CTRL RNAi (w1118; UAS-Ctrl RNAi/+; UAS-mito-Keima, mef-2-GAL4), wild-type and mutant α-synuclein (w1118;;UAS-mito-Keima,mef-2-GAL4/UAS-SNCA X) and parkin RNAi (w1118; UAS-parkin RNAi/+; UAS-mito-Keima, mef-2-GAL4/+) flies. High 543/458 ratio signal puncta correspond to mito-Keima present in lysosomes, indicated by arrowheads. Scale bar, 10 µm. d High mito-Keima (543/458) ratio area/total mitochondrial area was quantified as an index of mitophagic flux (n = 5–6 flies per condition, from at least 3 different crosses). In each fly, 10 random 2500 μm² fields were analyzed. One-way ANOVA with post hoc Tukey’s test. *P < 0.0001 compared to all flies expressing α-synuclein and parkin RNAi flies. Error bars represent SEM.