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. 2021 Jan 28;17(1):e1009300. doi: 10.1371/journal.pgen.1009300

Fig 2. Epithelial cell-turnover is essential for robust wing development in M/+ animals.

Fig 2

(A-E) Adult wings of wild-type (A), RpS3/+ (B), RpS3/+, nub-Gal4, UAS-p35 (C), nub-Gal4, UAS-p35 (D), or RpS3/+, nub-Gal4, UAS-p35, UAS-RpS3 (E) fly. Scale bar, 500 μm. (F) The rate of defective wings in the genotypes shown in (A) (n = 150), (B) (n = 271), (C) (n = 128), (D) (n = 125), and (E) (n = 44). (G and H) Dying cells were visualized by anti-cleaved PARP staining (white) in the wing discs of RpS3/+, H99/+ (G) or RpS3/+, nub-Gal4, UAS-DroncDN (H). Undead cells were not generated in these situations. Scale bar, 100 μm. (I) Boxplot with dots representing cleaved-PARP-positive dying cells per pouch in genotypes shown in (Fig 1B) (n = 12, number of wing pouches), (G) (n = 16), and (H) (n = 33). Error bars, SEM; ***, p<0.001; non-parametric Mann-Whitney U-test. (J and K) Wing disc of RpS3/+, H99/+ (J) or RpS3/+, nub-Gal4, UAS-DroncDN (K) were stained with anti-phospho-histone H3 (pH3) (Ser10) antibody (white). Wing pouches are marked in pale green. Scale bar, 100 μm. (L) Boxplot with dots representing pH3 positive cells per pouch in genotypes shown in (Fig 1A) (n = 34, number of wing pouches), (Fig 1B) (n = 31), (J) (n = 19), and (K) (n = 27). Error bars, SEM; ***, p<0.001; non-parametric Mann-Whitney U-test. (M) The rate of defective wings in the genotypes of wild-type (n = 70), RpS3/+ (n = 86), RpS3/+, H99/+ (n = 83), H99/+ (n = 100), RpS3/+, nub-Gal4, UAS-DroncDN (n = 63), and nub-Gal4, UAS-DroncDN (n = 109). (N) Adult wing phenotypes were classified as following five types: (a) normal, (b) vein (weak) (weakly having a brunched vein in the point of “landmark 14” (arrow) [69]), (c) vein (strong) (having other additional vein phenotypes (arrowheads) in addition to “landmark 14”), and (d) severe. Genotypes are as follows: w1118 (A), FRT82B, Ubi-GFP, RpS3Plac92/+ (B), nub-Gal4/+; FRT82B, Ubi-GFP, RpS3Plac92/UAS-p35 (C), nub-Gal4/+; UAS-p35/+ (D), nub-Gal4/UAS-RpS3; UAS-p35/FRT82B, Ubi-GFP, RpS3Plac92 (E), nub-Gal4/+; UAS-CD8-PARP-Venus, RpS3Plac92/Df(3L)H99 (G, J), nub-Gal4/+; UAS-CD8-PARP-Venus, RpS3Plac92/UAS-DroncDN (H, K), nub-Gal4/+; UAS-CD8-PARP-Venus/+ (wild-type), nub-Gal4/+; UAS-CD8-PARP-Venus, RpS3Plac92/+ (RpS3-/+), nub-Gal4/+; UAS-CD8-PARP-Venus, RpS3Plac92/Df(3L)H99 (RpS3-/+, H99/+), nub-Gal4/+; UAS-CD8-PARP-Venus/Df(3L)H99 (H99/+), nub-Gal4/+; UAS-CD8-PARP-Venus, RpS3Plac92/UAS-DroncDN (RpS3-/+, nub>DroncDN), and nub-Gal4/+; UAS-CD8-PARP-Venus/UAS-DroncDN (nub>DroncDN) in (M).