Fig. 6

Intestinal α-syn triggers the DUOX–ROS–JNK pathway. a, b Levels of reactive oxygen species (ROS). Fluorescence staining for peroxidase activity using dihydroethidium (DHE) in fly midgut (a). The quantification of fluorescence intensity (b). Scale bars, 100 μm. c The level of H₂O₂ was elevated by intestinal α-syn. ROS activity was detected by the H₂O₂ assay. d Survival rates of intestinal PD flies challenged with rotenone, with or without ROS-scavenging N-acetylcysteine treatment. e, f Activation of DUOX and JNK signal by the overexpression of α-syn in guts. The levels of Duox and upd3 of the middle intestine were assessed in 28-day-old flies with RT-PCR. g An increase of phospho-JNK level in esg^TS > Syn intestine. Green, GFP; Red, anti-phospo-JNK; blue, DNA; asterisk, esg-negative and p-JNK-positive cells; scale bars, 0.2 μm. h JNK signaling is required for the attenuation of lifespan of esg^TS > Syn flies treated with rotenone. A dominant-negative form of Jun (Jun^bZIP) and a wild-type form of Basket (bsk) were used to inhibit and activate the JNK signaling in midguts, respectively. R: rotenone. i JNK-mediated proliferation of intestinal stem cells in esg^TS > Syn intestines. Mean ± SEM; the P-values for survival curves were calculated using log-rank tests (using total fly numbers), and for category graphs using one-way ANOVA with Bonferroni multiple-comparison test. *P < 0.05; **P < 0.01; ***P < 0.001; ns, not significant