Figure 2. Hallmarks of proteotoxic stress in RP-deficient cells.

(a, b) Wing discs with one half RPS23^R67K/+ or RPS26^KO/+, labelled to visualise puromycilated peptides following a 30 min incubation in 20 μM of OPP. (c, d) Wing primordia from wild type or RPS23^R67K/+ larvae expressing HTT25Q-Cerulean throughout the pouch (under the control of pdm2-GAL4). (e) Quantification of HTT25Q punctae in RPS23^R67K/+ tissue (red dots, genotype as in panel d, n = 7 discs), expressed as a fold change (FC) in surface area coverage relative to that in otherwise wild type tissue (black dots, genotype as in panel c, n = 7 discs). (f,g) Anti-ubiquitin staining of a mosaic wing disc with the anterior half RPS23^R67K/+. Increased punctae staining in the anterior compartment expressed as a ratio with the posterior signal (n = 13 discs). (h,i) Anti-p62 staining of the same genotype as in panel f and quantified as for panel g (n = 9 discs). (j, k) Wild type or RPS23^R67K/R67K HEK293 cells stained with anti-p62 (green) and DAPI (red). (l) Coverage of p62 immunoreactivity in RPS23^R67K/R67K (n = 13 images) relative to that in wild type HEK293 cells (n = 11 images). (m) Lysotracker staining in a wing disc with one half RPS23^R67K/+. (n) P-eIF2α immunoreactivity in a wing disc with one half RPS23^R67K/+. Statistics: all error bars denote standard deviation. For panels e, g, i and l, a two-tailed unpaired t-test was carried out. P-values in e, ***P = 5.81E-05; g, **P = 1.08E-03; i, ***P = 1.96E-04; l, **P = 1.07E-03. Scale bars: 50 μm in panels a-h and m, n, and 10 μm in panels j, k. Genotypes for each figure panel are available in Supplementary Table 1. Source data is available for this figure.