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. 2012 Mar 29;7(3):e34310. doi: 10.1371/journal.pone.0034310

Figure 3. Vpu expression induces apoptosis correlated with basal extrusion of apoptotic cells in the wing disc.

Figure 3

(A) dpp>Vpu wing phenotypes (26°C) suppressed to varying degrees (medium to very weak) by diap1 overexpression (dpp>diap1, Vpu). (B) Distribution (% of wings) between the different phenotypic classes at 26°C. The distribution when diap1 is over-expressed (blue and red) is statistically different in a Chi2 test from the reference distribution (yellow, n = 282): p = 1.4 10−86 with UY1835 (n = 107) and p = 2.9 10−51 with UAS-diap1 (n = 74). (C–F) Acridine Orange (AO) staining of dpp-Gal4/+ (C), dpp-Gal4 UAS-Vpu/+ (D), dpp-Gal4 UAS-Vpu/Def(H99) (E) and dpp-Gal4 UAS-Vpu/UY1835 (F) wing discs. (G–L) XY confocal sections of wing discs. (G) dpp-Gal4 UAS-Vpu/+ showing Vpu immunostaining (G′; green), TUNEL staining (G″; red), merge (G′″) and F-actin network (G″″; phalloidin labeling, gray). (H) dpp-Gal4/+ showing TUNEL staining (H′; red) and F-actin (H″; gray). (I) Vpu2-6/+; dpp-Gal4/+ showing Vpu2-6 immunostaining (I′; green), TUNEL staining (I″; red) and merge (I′″) at 29°C. Asterisks in G′,G′″, I′,I′″: patches of Vpu-expressing cells displaced posteriorly. Below each image in G–I and L: transverse XZ section (g′-g″″, h′, i′-i″′ and I′-I′″) through the plane indicated by the line in G″″, H″ I′″, and L′″, respectively. Epithelial cells are oriented with basal towards the bottom. (g″″, h′): F-actin. Arrowhead in g″-g′″: TUNEL staining in Vpu-expressing cells not extruded from the epithelial sheet. Arrow in g″″: cells extruding from the epithelium. (J–L) Immunodetection of DIAP1 (red) and Vpu (green) in dpp-Gal4/+ (J), UAS diap1-IR/+; dpp-Gal4/+ (K) and dpp-Gal4 UAS-Vpu/+ (L) discs. Arrowheads in K and L″: DIAP1 reduction. Arrowheads and arrow in l″-l′″: DIAP1-downregulation in Vpu-expressing cells located in the dpp stripe and in those extruded from the epithelium, respectively. (M–N) dpp-lacZ expression (X-Gal staining) in C765-Gal4/+; UAS-Vpu/+ (M) and C765-Gal4/UAS-diap1; UAS-Vpu/+ (N). (O–R) rpr-lacZ expression in dpp-Gal4/+ (O), dpp-Gal4 UAS-Vpu/+ (P), en-Gal4/+ (Q), en-Gal4/+; UAS-Vpu/+ (R).