Flt3^ITD/Runx1^DELand Flt3^ITD/NHD13 induce distinct cooperative changes in gene expression and enhancer activation. (A) Principal component analysis for MPPs for the indicated genotypes at P14, based on bulk RNA-seq. n = 4 per genotype. (B) Heatmap displaying the expression of genes that were induced in either Flt3^ITD/NHD13 or Flt3^ITD/Runx1^DEL MPPs, relative to wild-type (FDR < 0.05, positive fold change ≥2). (C) Venn diagram showing minimal overlap between Flt3^ITD/NHD13 and Flt3^ITD/Runx1^DEL cooperative target genes, defined as genes from panel B with significantly higher expression in compound mutant MPPs relative to both single-mutant MPPs (FDR < 0.05). (D) Heatmap showing H3K27ac levels for enhancers (defined as overlapping ATAC-seq and H3K4me1 peaks) that were more highly active in Flt3^ITD/NHD13 MPPs relative to wild-type MPPs, and one or both single mutants (based on H3K27ac peak heights, FDR < 0.1 by DiffBind analysis). Peaks are clustered based on whether H3K27ac levels in the compound mutant are elevated relative to Flt3^ITD or NHD13 single mutant MPPs, or both (cooperative target enhancers). (E) A similar heatmap as in panel D, except showing enhancers that were more highly active in Flt3^ITD/Runx1^DEL MPPs. (F-G) Aggregate peak heights for cooperative target enhancers from panels D-E, respectively. (H) Venn diagram showing the minimal overlap between Flt3^ITD/NHD13 and Flt3^ITD/Runx1^DEL cooperative target enhancers.