Extended Data Fig. 1: Transient PRC1 depletion generates neoplastic tumours that persist after PH protein recovery.

a- GFP staining (in green) used as a readout of the conditional ph knockdown (ph-KD) system described in Fig. 1a after no ph-KD (control), constant or transient ph-KD. The tissues were counterstained with DAPI (blue). Two independent experiments were performed with similar results. b- gfp mRNA fold change RT-qPCR measurement after no ph-KD (control), constant or transient ph-KD. Bars correspond to the mean ± standard deviation (whiskers) inferred from three biological replicates (grey dots). c- PH immunostaining (in red) after no ph-KD (control), constant or transient ph-KD. The tissues were counterstained with DAPI (blue). Two independent experiments were performed with similar results. d- Quantification of the western blot illustrated in Fig. 1b and of two other biological replicates. Bars correspond to the mean ratio ±standard deviation (whiskers) of the signal of PH over that of TUBULIN (PH/TUB) calculated from three biological replicates (grey dots). Two-sided unpaired t.test: *pval < 0.05, ns = pval > 0.05 (not significant). Error bars represent the standard error of the mean for three biological replicates. Dunnet’s test: ns = not significant, **pval < 0.01. e- Western blot showing the PH protein after early L3 EDs were subjected to 24 h of white-KD (w-KD, control) or ph-KD followed by 0 h, 24 h, 48 h and 72 h of recovery at 18 °C (see bottom axis). This time course illustrates acute depletion and allows visualization of the kinetics of PH recovery after ph-KD. f- Hatching rate after constant or transient ph-KD. g-i- DAPI (in grey, g), F-actin (in red, h) and ELAV (in magenta, i) stainings of L3 EDs after no ph-KD (control), ph-KD throughout the three larval stages (L1, L2, L3) or transient (24 h) ph-KD during the first (L1), second (L2), early (Early L3) or late (Late L3) of the L3 stage, respectively. DAPI staining is used to assess ED growth, F-actin for apico-basal polarity, and the neuronal marker ELAV for differentiation. Note that late L3 tissues look normal immediately after the end of the ph-KD. Nevertheless, their cells are reprogrammed into a malignant state, as indicated by the fact that allografts of these tissues induce tumours, as shown in Extended Data Fig. 7i, j. Two independent experiments were performed with similar results. Scale bars: 10 μm (a, c, h, i), 100 μm (g). j-l- DAPI (in gray, j), F-actin (in red, k) and ELAV (in magenta, l) stainings of EDs after no Psc/Suz(2)-KD (control, left), constant (middle) or transient Psc/Suz(2)-KD (right), respectively. DAPI staining is used to assess growth, F-actin for apico-basal polarity, and the neuronal marker ELAV for differentiation. Two independent experiments were performed with similar results. m- ED sizes quantified as overall area of DAPI staining after no Psc/Suz(2)-KD (control), constant or transient Psc/Suz(2)-KD conditions. n = 30 for each condition. Two-sided Wilcoxon test: *pval < 5e-2, ****pval<1e-5. Box plots show the median (line), upper and lower quartiles (box) ±1.5x interquartile range (whiskers), outliers are not shown. Scale bars: 100 μm (j), 10 μm (k, l).