Chromatin accessibility in GM-CSF–stimulated LKS⁺ and LKS⁻ CD34⁺ FcγR⁺ progenitors. (A) LKS⁻ CD34⁺ FcγR⁺, or LKS⁺ cells were isolated from the bone marrow of 10 C57BL/6 mice, cultured with M-CSF (50 ng/mL) + SCF (20 ng/mL) and stimulated with or without GM-CSF (50 ng/mL) for 24 hours, nuclei were extracted, and single-cell assay for transposase-accessible chromatin with high-throughput sequencing (scATAC-seq) was performed. (B) Uniform manifold approximation and projection (UMAP) of scATAC-seq data with 10 clusters identified by K-nearest neighbor approach. (C) Distribution of cells in each cluster within samples. (D) Bar graph for the LKS⁻ CD34⁺ FcγR⁺ myeloid progenitors showing the fraction of cells present in each cluster. (E) UMAP of scATAC-seq data separated by sample and treatment condition. (F) Volcano plot of regions that are differentially accessible between cluster 3 and cluster 0. (G) Gene ontology enrichment of 793 genes with 10 kb of 1269 regions with greater accessibility in cluster 3. (H) Top results from motif enrichment analysis within 1269 regions with increased accessibility in cluster 3 compared with cluster 0. (I) Principal component analysis of bulk ATAC-seq–analyzed macrophages derived from M- or M+GM–treated Lin⁻ CD34⁺ FcγR⁺ or LKS⁺ progenitors. (J) Volcano plot of bulk ATAC-seq showing regions that are differentially accessible between M+GM vs M in macrophages derived from LKS⁻ CD34⁺ FcγR⁺ or LKS⁺ cells; statistically significant regions (n = 1299) labeled in red. (K) Heat map of bulk ATAC-seq signal showing differentially accessible regions between macrophages derived from M+GM- vs M-treated Lin⁻ CD34⁺ FcγR⁺ cells. Data derived from 3 biological replicates per condition.