Etv6^R355X/+ HSCs are reduced and have increased expression of inflammatory response genes when examined 6 weeks after transplantation. (A) Uniform manifold approximation and projection visualization of 22 cell populations identified from the posttransplant Etv6^+/+ and Etv6^R355X/+ LSK-enriched BM, colored by cell population. (B) Proportions of cells in each of the 22 clusters (left). Box plots depicting the proportions of Etv6^R355X/+ or Etv6^+/+ cells within clusters that show significant reductions across 3 replicates at 6 weeks after transplantation (right). ∗P value < .05, t test. (C) Summary of significant differentially expressed genes from all Etv6^R355X/+ clusters compared with Etv6^+/+ from scRNA-seq data (false discovery rate < 0.05) (left). Upregulated (red) and downregulated (blue) genes marked by |log2 fold-change (FC)| >1. Volcano plots for HSC, MPP, and ELP clusters are shown on the right: each dot represents a gene, colored by P value < 10e−6 (blue), log2 FC > 1 (green), or both (red). (D) Gene-set enrichment analysis (GSEA) analysis using custom gene sets of 813 genes, exhibiting significant depletion of ETV6 binding from anti-ETV6 CUT&RUN in Etv6^R355X/+ LSK compared with Etv6^+/+ LSK cells in HSC (top) and MPP (bottom) clusters. (E) Top 10 significant gene sets from GSEA of Etv6^R355X/+ vs Etv6^+/+ HSC, MPP, and ELP cell clusters using Hallmark gene sets (MSigDb version 7.5.1).