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. 2022 Sep 1;219(11):e20212437. doi: 10.1084/jem.20212437

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© 2022 Lenaerts et al.

This article is available under a Creative Commons License (Attribution 4.0 International, as described at https://creativecommons.org/licenses/by/4.0/).

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Figure 4. — EBF1-dependent chromatin sites in MPP3 cells are associated with expression in CLPs. (A) Selected terms are represented from enrichment analysis of DE genes (FDR < 0.1) between Ebf1^WT and Ebf1^KO MPP3 (left) and MPP4 (right) cells. Enrichr datasets (GO_Biological_Process_2021, PanglaoDB_Augmented_2021, Gene_Perturbations_from_GEO_down) were used for enrichment analysis. Biological replicates n = 4. (B) Expression analysis of genes annotated to KO Reduced MPP3 peaks identified in ATAC analysis, in Ebf1^WT and Ebf1^KO MPP3 (upper panel), and MPP4 (lower panel) cells. The x axis represents log₁₀ mean normalized counts of all genes. The y axis represents shrunken |log₂ fold change| values of all genes in Ebf1^WT versus Ebf1^KO MPP3 (upper panel), and MPP4 (lower panel) cells, and is capped at 2. Genes in orange indicate the genes annotated to EBF1-dependent accessibility sites in MPP3 cells. (C) Heatmap displaying average ChIP signal for chromatin marks at DA peaks. Original data from Lara-Astiaso et al. (2014). (D and E) Comparison of chromatin accessibility changes in MPP3 cells (D) and MPP4 (E) cells to gene expression changes in GMP versus CLP cells. The y axis represents shrunken log₂ fold changes of gene expression in GMP versus CLP cells. The x axis represents log₂ fold changes of peak read counts in Ebf1^KO versus Ebf1^WT MPP3 (D) and Ebf1^KO versus Ebf1^WT MPP4 (E) cells. Highlighted box shows positive correlation of KO Reduced MPP3 peaks and CLP gene expression. (D and E) Left: Dots represent DA peaks in MPP3 and MPP4 cells. Right: Dots represent DE genes and ATAC peaks that overlap their TSS (TSS peaks). (F and G) GSEA results of all MPP3 (F) and MPP4 (G) ATAC peaks predicted to be enhancers (CRUP probability >0.8), ranked by log₂ fold change, against a CLP-specific gene set (left) and a GMP-specific gene set (right). Positive enrichment scores reflect enrichment of the gene set in Ebf1^KO cells, negative enrichment scores reflect enrichment of the gene set in Ebf1^WT cells. CLP and GMP RNA-seq data analyzed in D–G was retrieved from GEO dataset GSE162662.