Fig. 2: Phosphoinositol metabolism is deregulated in BMI1^High;CHD7^Low G4 MB and activation of the mTOR pathway is observed specifically in MB cells modelling this subgroup.

a Schematic representation of the comparative analysis between BMI1^High;CHD7^Low MB cell lines and G4 patients with and without the signature performed to identify genes differentially expressed (DE) and differentially methylated probes (DMP) and commonly deregulated pathways. b Pie charts representing canonical pathways differentially enriched between MB cells or patients with or without BMI1^High;CHD7^Low signature identified in RNA-Seq or DNA methylation analysis. Canonical pathways are classified based on IPA categories list and colour-coded accordingly. c Venn diagram of canonical pathways enriched for genes differentially expressed (blue and light-green diagrams as coloured in a and b) or methylated (olive green and red as coloured in a and b) in BMI1^High;CHD7^Low cell lines and GR4 MBs with the matching signature. Highlighted the superpathway of phosphoinositol compounds (phosphoinositols), which is common to all the datasets analysed. d Bubble plot showing significant Reactome pathways obtained in phosphoproteomic analysis of BMI1^High;CHD7^Low MB lines. Bubbles are coloured based on FDR values, and size is proportional to the number of genes of specific pathways. e, f Cell viability assays of MB cells (e) or hNSC (f) with (BMI1^High;CHD7^Low) or without the signature (CTRL) upon 72 h of treatment with increasing concentrations of FLT3 inhibitor (TCS). Histograms represent the percentages of viable cells relative to non-treated cells (top). Measurement of Area Under Curve (AUC) (bottom) to compare the overall response to treatment. e n = 6 biologically independent experiments, (f) n = 4 biologically independent experiments, two-way ANOVA. g, h Cell viability assays of MB cells (g) or hNSC (h) with (BMI1^High;CHD7^Low) or without the signature (CTRL) upon 72 h of treatment with increasing concentrations of MET inhibitor (PHA). Histograms represent the percentages of viable cells relative to non-treated cells (top). Measurement of the area under the curve (AUC) (bottom) to compare the overall response to treatment. g n = 6 biologically independent experiments, h n = 4 biologically independent experiments, two-way ANOVA. i Western blot and quantification of phosphorylated/total RPS6 (pRPS6, Ser240/244) in MB cells. GAPDH immunoreactivity was used to normalise protein loading. n = 4 biologically independent experiments, one-way ANOVA. j Cell viability assays of MB cells with (BMI1^High;CHD7^Low, red) or without the signature (CTRL, green) upon 72 h of treatment with increasing concentrations of mTOR pathway inhibitors (rapamycin and torin). Histograms represent the percentages of viable cells relative to non-treated cells (top). Measurement of Area Under Curve (AUC) (bottom) to compare the overall response to treatment. n = 5 biologically independent experiments, two-way ANOVA. k Western blot and quantification of phosphorylated/total RPS6 (pRPS6, Ser240/244) after 24 h of treatment with increasing concentrations of IP6 in control (green) or BMI1^High;CHD7^Low (red) MB cells. GAPDH immunoreactivity was used to normalise protein loading. n = 4 biologically independent experiments, two-way ANOVA. All graphs report mean ± SEM. P values: *P < 0.05, **P < 0.01, ***P < 0.001 or ****P < 0.0001. Source data are provided as a Source Data file.