Single-cell transcriptome analysis of SETBP1^LSL and SETBP1^G870S Lin⁻ BM cells. (A) Uniform manifold approximation and projection (UMAP) plot showing the integrated single-cell RNA (scRNA) analysis, color-coded by cell type. ETP, earliest thymic progenitors; T-DN2a, T-DN2a cells; CDP, common dendritic progenitors; MDP, monocyte-dendritic cell progenitors; Tem, T effector memory cells; DC, dendritic cells. (B) UMAP plot reporting the expression level of the artificial SETBP1-EGFP transcript in SETBP1^LSL (left) and SETBP1^G870S (right) 90-day-old littermates in a blue-to-yellow color scale. (C) UMAP plots reporting the expression level of Setbp1, Gata2, Hoxa9, Ezh2, and Gata1 in SETBP1^LSL (left) and SETBP1^G870S (right) mice in a blue-to-yellow color scale. (D) Violin plot showing the expression level of Setbp1, Gata2, Hoxa9, and Ezh2 in the HSC cluster and of Gata1 in the CMP cell cluster of SETBP1^LSL (green) and SETBP1^G870S (orange) mice. Differentially expressed genes were identified using a negative binomial generalized linear model; the false-positive discovery rate (FDR) was controlled using the Bonferroni procedure. (E-G) Enriched gene sets for TGF-beta signaling, Hoxa9 downregulated targets, and PRC2 targets identified in in SETBP1^G870S vs SETBP1^LSL HSC cells. FDR represents the Benjamini-Hochberg adjusted P value; normalized enrichment score (NES) is the normalized gene set enrichment analysis (GSEA) enrichment score. HOXA9 downreg. targets represents genes that are downregulated in HSCs expressing HOXA9.²⁶ (H) Heat map of the top GSEA leading edge genes of the TGF-beta signaling, Hoxa9 downregulated targets, and PRC2 targets pathways in SETBP1^LSL (left, green ribbon) and SETBP1^G870S (right, orange ribbon) mice. (I) Western blot analysis of PB samples obtained from SETBP1^LSL and SETBP1^G870S mice; transgenic SETBP1 is detected both by anti-SETBP1 and anti-V5 antibodies. Actin is shown as a loading control.