Fig. 1: Oxidative phosphorylation downstream of NOTCH1 is essential for preleukemic stem cells function.

a Gene set enrichment analysis of NOTCH1-bound genes in Molecular Signatures Database. The list of genes in which NOTCH1 binding was observed within 2 kb of transcription start sites in murine T-ALL cell lines was extracted as described in Materials and Methods. False discovery rates (FDRs) are shown for overlaps computed with Hallmark gene sets or canonical pathways (Reactome and Kyoto Encyclopedia of Genes and Genomes). TCA: tricarboxylic acid; ETC, electron transport chain. b Dynamics of NOTCH1 target gene expression during thymocyte differentiation. The numbers of NOTCH1 target genes that increased >1.3-fold at each transition (gray bars, left axis). Gene expression data are from the Immunological Genome Project. GSEA was conducted on the gene sets whose expression increased at each transitional stage. FDR values overlap with the OxPhos gene set (Hallmark) were computed as above (green bars, right axis). ETP, early T-lineage precursor; DN3a, (CD4⁻CD8⁻CD25⁺CD44⁻CD28⁻), DN3b, (CD4⁻CD8⁻CD25⁺CD44⁻CD28⁺). c Heatmap of expression of NOTCH1-bound genes within the OxPhos pathway. Ultralow input (ULI) RNA-seq data were obtained from IMMGEN. ETP, early thymocyte progenitor (Lin^lowCD25⁻CD44⁺Kit⁺), DN2, double negative 2 (Lin^lowCD25⁺CD44⁺Kit⁺), DN3 (CD4⁻CD8⁻CD25⁺CD44⁻), DN4 (CD4⁻CD8⁻CD25⁻CD44⁻CD28⁺), ISP, immature single positive (CD4⁻CD8⁺CD24^hiTCR^lo), DP (CD4⁺CD8⁺TCR^loCD24^hi), T4 (CD4⁺CD8⁻ − TCR^hi), T8 (CD4⁻CD8⁺TCR^hi). d GSEA of the TARGET gene set (Therapeutically Applicable Research to Generate Effective Treatments) (N = 265), indicated enrichment of genes related to glutamine metabolism, mitochondrial metabolism as well as translation and downregulation of apoptosis-related genes in patients with NOTCH1-mutations (each vertical bar in x-axis is gene rank in the pathway list and y-axis represents running enrichment score). e The OxPhos gene signature in T-ALL patients from TARGET cohort. f Enrichment of mitochondrial translation genes in T-ALL patients from TARGET cohort. g Enrichment of apoptosis-related genes in T-ALL patients from the TARGET cohort; for (e), (f) and (g): two-sided t-test; The center line represents the median and whiskers represents maximum (Q3 + 1.5*IQR) and minimum value (Q1+1.5*IQR).