Jak2^V617F/+Trp53^−rp and Jak2^V617F/+Trp53^R172H/– MEPs are characterized by aberrant gene expression. (A) Principal component analysis (PCA) based on bulk RNA-seq data from flow cytometry–sorted samples. The top 2 components (PC1 and PC2) of the PCA results were used for plot in the plane. (B) Heat map showing representative MEP-priming transcription factor (TFs) expression in WT MEPs and LSKs from WT, J^VF, J^VFP^+/−, J^VFP^−/−, J^VFP^172/+, and J^VFP^172/–. (C) Gene set enrichment analysis (GSEA) for comparing gene expression of J^VFP^−/− and J^VFP^172/– MPN MEPs vs previously described signatures related to leukemia stem cells or hematopoietic stem cells. (D) Methylcellulose replating assay showing the enhanced replating capability of J^VFP^−/− and J^VFP^172/– MPN stage MEPs, relative to J^VF MEPs (n = 4 for each genotype). A total of 5000 MEPs were plated per well in the first platting, and 5000 cells from each prior platting were used for subsequent replating. Data are represented as mean ± standard error of the mean (SEM). The unpaired t test was used to compare the mean of 2 groups. (E) Bar graphs showing the top pathways enriched in J^VFP^−/− and J^VFP^172/– PEL MEPs. (F) Heat map of upregulated genes common to J^VFP^−/− and J^VFP^172/– PEL MEPs compared with respective controls. (G) Volcano plot from RNA-seq data comparing J^VFP^−/− and J^VFP^172/– PEL MEPs vs controls highlighting BMP pathway–related gene expression. *P ≤ .05, **P ≤ .01, ***P ≤ .001.