Panel A: Confocal microscopy image showing wing disc with f z r overexpression clones labeled by G F P and stained with c D c p 1 and D A P I. Panel A prime: Confocal microscopy image showing magnified view of f z r overexpression clones region with c D c p 1 and D A P I staining. Panel B: Confocal microscopy image showing wing disc with r u x overexpression clones labeled by G F P and stained with c D c p 1 and D A P I. Panel B prime: Confocal microscopy image showing magnified view of r u x overexpression clones region with c D c p 1 and D A P I staining. Panel C: Confocal microscopy image showing 18 G y irradiated wing disc with f z r overexpression clones labeled by G F P and stained with c D c p 1 and D A P I. Panel C prime: Confocal microscopy image showing magnified irradiated region with G F P positive clones and apoptotic signal. Panel D: Confocal microscopy image showing 18 G y irradiated wing disc with r u x overexpression clones labeled by G F P and stained with c D c p 1 and D A P I. Panel D prime: Confocal microscopy image showing magnified irradiated region with clustered G F P positive cells. Panel E: Confocal microscopy image showing control wing disc with G F P expressing clones in s c r i b background stained with c D c p 1 and D A P I. Panel E prime: Confocal microscopy image showing magnified control region with dense G F P positive clones. Panel F: Confocal microscopy image showing irradiated s c r i b tumor wing disc with G F P clones stained with c D c p 1 and D A P I. Panel F prime: Confocal microscopy image showing magnified tumor region with G F P clones and protrusive structures. Panel G: Confocal microscopy image showing 35 G y irradiated tumor wing disc with f z r overexpression clones labeled by G F P and stained with c D c p 1 and D A P I. Panel G prime: Confocal microscopy image showing magnified view of G F P positive polyploid clones with protrusions and engulfment events. Panel H: Vertical bar graph quantifying number of engulfment events per mosaic clone across experimental conditions. Panel I: Vertical bar graph quantifying c D c p 1 positive apoptotic cells per engulfed cell within G F P positive clones.
Induced polyploid cells exhibit engulfment activity. (A and B) Representative confocal images of fzr-OE (A, wing disc) and rux-OE (B, eye disc) clones stained for cDcp1 (red) and DAPI (blue). GFP marks polyploid clones (green). Panels A′ and B′ show magnified views of the boxed regions in A and B, respectively. (C and D) Confocal images of fzr-OE (C) and rux-OE (D) clones following exposure to 18 Gy γ-irradiation. Clones were stained with cDcp1 (red) and DAPI (blue); GFP marks polyploid cells (green). Panels C′ and D′ show magnified views of the boxed regions. White arrowheads indicate apoptotic (cDcp1 positive) cells engulfed by neighboring polyploid cells. (E and G) Tumorigenic wing discs from scrib1/scribj7B3 trans-heterozygous mutant larvae containing mosaic GFP-positive clones. (E and E′) Control clones expressing GFP only. (F and F′) Clones expressing fzr (polyploid-inducing) with GFP. (G and G′)fzr-OE clones in a tumor disc 24 h after 35 Gy γ-irradiation. The second to fifth columns show magnified views of the boxed areas in the first column. cDcp1 (red) marks apoptotic cells; DAPI (blue) labels nuclei. White arrowheads indicate protrusions from fzr-OE polyploid clones; white arrows mark GFP-negative (non-polyploid) tumor cells engulfed by adjacent polyploid cells. (H) Quantification of engulfment in scrib1/scribj7B3 trans-heterozygous tumorigenic wing discs. The number of GFP-negative cells that are completely engulfed by GFP-expressing mosaic clones was quantified per clone cluster. (I) Quantification of apoptotic cells. The number of apoptotic cells (labeled by anti-cDcp1 antibody) per completely engulfed GFP-negative cell within GFP-positive mosaic clones was quantified. Data are mean ± SD from three independent experiments. **** represents P < 0.001 (unpaired two-tailed Student’s t test); n = 30 GFP-expressing clones from each indicated experiment. Scale bars: 20 μm (A–D) and 50 μm (E–G).