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. 2017 Aug 30;6:e26747. doi: 10.7554/eLife.26747

Figure 2. Caspases are required for ROS generation and neoplastic overgrowth in scrib−/− RasV12.

Figure 2.

(A–D’) Expression of the effector caspase inhibitor p35 (B), drICE RNAi (C) and dronc RNAi (D) suppresses scrib−/− RasV12 clone size (green) and ROS generation in scrib−/− RasV12 clones. The (‘) panels indicate the labeling of the ROS indicator DHE (grey). Scale bars: 50 μm. (E) DHE quantification reveals that ROS levels are significantly reduced in scrib−/− RasV12 mutant clones with reduced or inhibited caspase activity. Shown is the mean signal intensity ±SD of DHE labelings in clones, analyzed by one-way ANOVA with Holm-Sidak test for multiple comparisons. ****p<0.0001. Multiple clones from five to ten discs of each genotype were analyzed. (F–I) The growth and invasion of cephalic complexes of 11 day old scrib−/− RasV12 larvae (F) is strongly suppressed by p35 (G), drICE RNAi (H) and dronc RNAi (I). Clone size (green) in (F–I) is strongly reduced. DAPI labels the outline of the tissue. Scale bars: 200 μm. (J–M) Adult eyes of surviving scrib−/− RasV12 animals expressing p35 (K), drICE RNAi (L) and dronc RNAi (M). The percentage number in the top right of each panel indicates the adult survival rate relative to pupal survival. (N) Reduction or inhibition of caspase activity in scrib−/− RasV12 mutant clones significantly improves the pupariation rates of animals bearing scrib−/− RasV12 mosaic eye imaginal discs. Pupariation rates were determined as the ratio of late stage mutant pupae vs total pupae and were analyzed by one-way ANOVA with Holm-Sidak test for multiple comparisons. Error bars are SD. P values are relative to scrib−/− RasV12 results (left column) and are indicated above the experimental columns. ****p<0.0001. At least 100 pupae were counted per genotype. Experiments were performed three times. Genotypes: (A,F,J) yw ey-FLP/+; act>y+>Gal4, UAS-GFP56ST/+; FRT82B tub-Gal80/UAS-RasV12 FRT82B scrib2; (B–D,G–I,K–M) yw ey-FLP/+; act>y+>Gal4, UAS-GFP56ST/UAS-X; FRT82B tub-Gal80/UAS-RasV12 FRT82B scrib2 with UAS-X being UAS-p35 (B,G,K), UAS-drICERNAi (C,H,L) and UAS-droncRNAi (D,I,M).