PTP10D-mediated cell competition is not obligately required for elimination of polarity-deficient clones

Abstract

Animal organs maintain tissue integrity and ensure removal of aberrant cells through several types of surveillance mechanisms. One prominent example is the elimination of polarity-deficient mutant cells within developing Drosophila imaginal discs. This has been proposed to require heterotypic cell competition dependent on the receptor tyrosine phosphatase PTP10D within the mutant cells. We report here experiments to test this requirement in various contexts and find that PTP10D is not obligately required for the removal of scribble (scrib) mutant and similar polarity-deficient cells. Our experiments used identical stocks with which another group can detect the PTP10D requirement, and our results do not vary under several husbandry conditions including high and low protein food diets. Although we are unable to identify the source of the discrepant results, we suggest that the role of PTP10D in polarity-deficient cell elimination may not be absolute.

Keywords: Drosophila; Cell competition; Dlg; Epithelial polarity; PTP10D; Scrib.

Fig. 1.

PTP10D depletion does not alter removal of Dlg-deficient cells. (A-D) ptc-Gal4-driven dlg-KD causes apoptosis along the A-P boundary (A; anti-DCP-1 in red) and additional PTP10D-KD does not alter apoptosis (B). Expression of Dcr2 along with dlg-KD triggers additional apoptosis due to enhancement of the long-inverted repeat RNA targeting dlg, but additional PTP10D-KD does not alter apoptosis (C). Quantitation in D (mean±s.d., one-way ANOVA test, n=9 for WT, n=23 for dlg-KD, n=15 for dlg-KD+PTP10D-KD, n=9 for Dcr2+dlg-KD, n=17 for Dcr2+dlg-KD+PTP10D-KD). (E-H) PTP10D-KD along the A-P boundary leads to strong reduction of PTP10D protein (F; control in E; anti-PTP10D in gray). Additional expression of Dcr2 does not lead to a stronger depletion of PTP10D (G). Quantitation in H (mean±s.d., one-way ANOVA test, n=8 for WT, n=19 for PTP10D-KD, n=11 for Dc2+PTP10D-KD). Scale bars: 100 µm in A, and E, 10 µm in E’. Statistical significance is indicated with *P≤0.05, **P≤0.01, ***P≤0.001, and ****P≤0.0001.

Fig. 2.

PTP10D depletion does not rescue scrib clone removal. (A-G) eyFLP1-generated scrib clones are eliminated from the eye disc (B; control in A) and PTP10D-KD does not rescue scrib clone elimination (C). Quantitation of clone area in D (mean±s.d., one-way ANOVA test, n=23 for WT, n=23 for scrib, n=24 for scrib+PTP10D-KD) and apoptosis along the clone boundary in E (mean±s.d., one-way ANOVA test, n=15 for WT, n=15 for scrib, n=15 for scrib+PTP10D-KD). Adult eyes of scrib and scrib+PTP10D-KD flies show a rough eye phenotype that is not enhanced with PTP10D depletion (F,G). (H-N) Additional expression of Dcr2 in eyFLP5-generated scrib clones shows clone elimination (I; control in H) that is not enhanced by PTP10D-KD (J). Quantitation of clone area in K (mean±s.d., one-way ANOVA test, n=27 for WT, n=26 for scrib, n=29 for scrib+PTP10D-KD) and apoptosis along the clone boundary in L (mean±s.d., one-way ANOVA test, n=13 for WT, n=12 for scrib, n=14 for scrib+PTP10D-KD). Adult eyes of scrib and scrib+PTP10D-KD flies show similar rough eye phenotypes (M,N). Scale bars: 100 µm in A and H. Statistical significance is indicated with *P≤0.05, **P≤0.01, ***P≤0.001, and ****P≤0.0001.

Fig. 3.

PTP10D-null larvae efficiently eliminate scrib clones. (A-H) Animals devoid of PTP10D eliminate scrib¹ clones as efficiently as animals carrying WT PTP10D (B, C; control in A). This is the case for scrib² clones in animals with WT PTP10D or null for PTP10D as well (E, F; control in D). Quantitation of clone area in G (mean±s.d., one-way ANOVA test, n=19 for WT, n=31 for scrib¹, n=24 for PTP10D¹+scrib¹, n=17 for scrib², n=13 for PTP10D¹+scrib²) and apoptosis along the clone boundary in H (mean±s.d., one-way ANOVA test, n=11 for WT, n=14 for scrib¹, n=13 for PTP10D¹+scrib¹, n=14 for scrib², n=14 for PTP10D¹+scrib²). Scale bars: 100 µm in A, and D. Statistical significance is indicated with *P≤0.05, **P≤0.01, ***P≤0.001, and ****P≤0.0001.

Fig. 4.

Varying husbandry conditions do not change outcome of PTP10D depletion. (A-D) Larvae raised on corn-syrup-based food show efficient elimination of scrib clones (B; control in A) and PTP10D-KD does not rescue clone elimination (C). Quantitation in D (mean±s.d., one-way ANOVA test, n=12 for WT, n=18 for scrib, n=20 for scrib+PTP10D-KD). (E-L) Larvae raised on 1X yeast food [E-G; quantitation in H (mean±s.d., one-way ANOVA test, n=15 for WT, n=15 for scrib, n=13 for scrib+PTP10D-KD)] as well as on 4X yeast food [I-K; quantitation in L (mean±s.d., one-way ANOVA test, n=8 for WT, n=12 for scrib, n=15 for scrib+PTP10D-KD)] show comparable elimination of scrib clones (F,J) and this is not rescued by PTP10D-KD (G,K). (M-T) Larvae raised on 0.1X yeast food [Q-S; quantitation in T (mean±s.d., one-way ANOVA test, n=8 for WT, n=6 for scrib, n=11 for scrib+PTP10D-KD)] show smaller scrib clones compared to standard yeast food [M-O; quantitation in P (mean±s.d., one-way ANOVA test, n=11 for WT, n=12 for scrib, n=12 for scrib+PTP10D-KD)], but PTP10D-KD does not rescue scrib clone elimination on either food (O,S). (U-X) Larvae raised on standard yeast food supplemented with anti-fungal reagents show efficient elimination of scrib clones (V; control in U) and PTP10D-KD does not rescue clone elimination (W). Quantitation in X (mean±s.d., one-way ANOVA test, n=12 for WT, n=12 for scrib, n=12 for scrib+PTP10D-KD). Scale bars: 100 µm in A, E, I, M, Q, and U. Statistical significance is indicated with *P≤0.05, **P≤0.01, ***P≤0.001, and ****P≤0.0001.

Fig. 5.

Varying crowding conditions do not change outcome of PTP10D depletion. This set of experiments used standard yeast food to raise larvae and was carried out in an independent incubator set to 25°C, 70% humidity control and 12 h light/dark cycles. (A-L) Crosses raised in wide vials under medium larval density (egg laying of 15 females) show removal of scrib clones that is not changed by PTP10D-KD [A-C; quantitation in D (mean±s.d., one-way ANOVA test, n=11 for WT, n=15 for scrib, n=20 for scrib+PTP10D-KD)]. Larvae raised in narrow vials under high-density conditions [E-G; quantitation in H (mean±s.d., one-way ANOVA test, n=9 for WT, n=12 for scrib, n=9 for scrib+PTP10D-KD); egg laying of 40 females] as well as very high-density conditions [I-K; quantitation in L (mean±s.d., one-way ANOVA test, n=12 for WT, n=13 for scrib, n=11 for scrib+PTP10D-KD); egg laying of 60 females] show removal of scrib clones (F,J) that is not rescued by PTP10D-KD (G,K). Scale bars: 100 µm in A, E, and I. Statistical significance is indicated with *P≤0.05, **P≤0.01, ***P≤0.001, and ****P≤0.0001.