Figure 1
The PERK/ATF4 pathway is responsible for Psn overexpression-induced loss of DIAP1 and apoptosis. (a) Third-instar imaginal wing discs with the Psn-expressing domain detected thanks to GFP (green) and TUNEL staining (red) to detect apoptosis. Genotypes are vg-GAL4/+; UAS-EGFP/+ (top), vg-GAL4, UAS-Psn, UAS-Psn/+; UAS-EGFP/+ (middle) and UAS-p35/+; vg-GAL4, UAS-Psn, UAS-Psn/+; UAS-EGFP/+ (bottom). (b) Third-instar imaginal wing discs carrying an xbp1::EGFP reporter transgene (green) to detect ER stress and immunostained with an anti-activated Caspase 3 staining (red) to detect apoptosis. Genotypes are vg-GAL4/+; xbp1::EGFP/+ (top row) and vg-GAL4, UAS-Psn, UAS-Psn/+; xbp1::EGFP/+ (middle row). Bottom row images are enlargement of boxed regions. Colocalization between anti-activated Caspase 3 staining and Ire1 activity reporter is highlighted in white thanks to the ImageJ Colocalization plugin in the merge enlargement. Note that developmental apoptosis is only observed in the notum (top). (c) TUNEL staining of Psn-expressing third-instar imaginal wing discs in the absence or presence of dronc mutations. Genotypes are vg-GAL4, UAS-Psn, UAS-Psn/+ (left) and vg-GAL4, UAS-Psn, UAS-Psn/+; droncI24/ droncI29(right). (d, e) Third-instar imaginal wing discs with the Psn-expressing domain detected thanks to GFP (green), TUNEL stained (red) to detect apoptosis and immunostained with an anti-DIAP1 antibody (blue). (d) Genotypes are vg-GAL4/+; UAS-EGFP/+ (top), vg-GAL4, UAS-Psn, UAS-Psn/+; UAS-EGFP/+ (middle) and vg-GAL4, UAS-Psn, UAS-Psn/+; UAS-EGFP/UAS-atf4-RNAi (bottom). (e) Genotypes are vg-GAL4, UAS-Psn, UAS-Psn /UAS-nGFP; UAS-EGFP/+ (top), vg-GAL4, UAS-Psn, UAS-Psn/UAS-nGFP; UAS-p35/+ (bottom) (f) Anti-β-Galactosidase staining to detect diap1J5C8 reporter expression in vg-GAL4/+; diap1J5C8/+ (left column) vg-GAL4, UAS-Psn, UAS-Psn/+; diap1J5C8/+ (center); and vg-GAL4, UAS-Psn, UAS-Psn/+; diap1J5C8/UAS-atf4-RNAi (right column) third-instar wing imaginal discs. The black lines correspond to the projection of the corresponding enlarged transverse XZ sections shown under disk images. Enlarge areas are oriented with their basal side to the left. The asterisk indicates the diap1 expression reduction (center)
