Extended Data Fig. 4: Proteasome defects are linked to the prospective loser status but not to translation inhibition.
From: Proteotoxic stress is a driver of the loser status and cell competition
(a-b) Apoptosis as detected by anti-cleaved caspase-3 reactivity (green), in Prosβ2+/− (a, left), RpS3+/− (a, middle), or Prosβ2+/−, RpS3+/− transheterozygote (a, right) wing discs and corresponding quantification (n = 10, 10, and 10 respectively, two-sided two sample Kolmgorov-Smirnov test without p-adjustment for multiple comparisons) (b). (c-d) Apoptotic cell death as detected by cleaved-dcp1 (red) in Prosβ2+/− (c, left), a RpL27A+/− (c, middle), or a RpL27A+/−, prosβ2+/− transheterozygote (c, right) wing discs, and corresponding quantification (n = 8, 13, and 10 respectively, two-sided Mann-Whitney U test without p-adjustment for multiple comparisons) (d). (e-g) ProteoFLUX CL1-GFP signal (green) in wing discs expressing mahj-RNAi in the P compartment (RFP-positive), immediately after heat shock (e) or two hours later (f) and corresponding signal quantifications (n = 9 and 7 respectively, two-sided two sample Kolmgorov-Smirnov test) (g). (h-j) ProteoFLUX CL1-GFP signal (green) in wing discs harboring wild-type A cells and 4E-BPTA-expressing P cells (labelled by the absence of Ci, magenta), immediately after heat shock (h) or two hours later (i), and corresponding signal quantifications relative to wing discs containing an RpS3+/− A compartment and wildtype P compartment (images not shown) (n = 9 and 10 for 0 and 2 hour 4E-BPTA, and n = 7 and 7 for 0 and 2 hour RpS3+/−, respectively; two-sided two-sample Kolmgorov-Smirnov test without p-adjustment for multiple comparisons) (j). (k) Transmission Electron microscopy images of a wing disc with wildtype P (left panel) and RpS3+/− A compartments (right panel). Red arrows indicate phago-lysosomal structures containing ribosomes. The scale bar is 500 nm. (l-m) Phospho-eIF2α staining (red) in wing discs harboring RpS3+/− A cells (GFP-positive) and wild-type P cells (GFP-negative) (l) and corresponding signal quantifications (n = 6, two-sided Wilcoxon ranked-sum test) (m). (n-o) A wing disc harboring RpS3+/− clones (GFP-positive) and stained for phospho-eIF2α (red) (n) and corresponding signal quantification (n = 9, two-sided paired t-test) (o). For all micrographs other than those in (k), scale bars correspond to 50 µm. For all quantifications, the horizontal line represents the mean and whiskers indicate 95% confidence intervals. All n numbers refer to the number of individual wing discs.
