Journal: iScience
Article Title: Chromatin interaction-based annotation of regulatory elements reveals dynamic promoter-enhancer interactions in lymphocyte development
doi: 10.1016/j.isci.2025.112855
Figure Lengend Snippet: EBF1 interacts with the regulatory elements of broadly expressed genes (A) Heatmap with the z -scores of normalized gene expression of Ebf1-bound elements in early lymphoid progenitors from the Immgen dataset (GEO: GSE100738 ). Percentage of genes in each cluster is presented. (B) Gene ontology (GO) analysis of enriched biologic processes in genes bound by EBF1 at promoters or distal regions. Gene sets are categorized based on differential expression between Wt and Ebf1 −/− FL pro-B cells [Up-/Down-regulated: padj 0.05 & |log2FC| >1, common: log2FC < log2(1.5)]. (C) Diagram displaying the relative number of generated CD19 + cells after 11 days of in vitro incubation of KIT + cells on OP9 stroma cells compared to R26 control. The red to blue color scale represents log2FC Wt vs. Ebf1 −/− from RNA-seq. Data are based on at least 3 separate experiments, and samples (n = 3–12) with a single guide or pool of individual guides are aggregated per gene. Statistical analysis is based on Student’s t test using the R26 control guide as reference: ∗ p < 0.05, ∗∗ p < 0.01, ∗∗∗ p < 0.001, ∗∗∗∗ p < 0.0001. (D) Promoter and distal elements associated to the gene sets from 2B are identified by ICE-A interaction-based annotation with H3K4me3 PLAC-seq data from FL Wt pro-B cells. Fraction of elements with a significant change (padj <0.05 & abs(log2FC) > 1) in accessibility between Wt and Ebf1 −/− FL pro-B cells are visualized in a stacked barplot, split into categories based on gene category and EBF1 occupancy. (E) The Myc locus is shown as an example of a gene that shows EBF1-addicted activation, visualized in the WashU genome Epigenome Browser. Selected EBF1-bound elements located in a distal enhancer region ∼1.6 Mb from the Myc TSS are highlighted in gray. The epigenome browser tracks include EBF1 ChIP-seq (GEO: GSE159957 ), ATAC-seq data from Wt/Ebf1 −/− FL pro-B cells and H3K4me3 PLAC-seq interactions from Wt FL pro-B cells. (F) Venn diagram displaying the overlap between common genes (defined as in B) and genes with differential expression at EBF1 degradation (|log2FC| > log2(1.5)), from ( GSE201141 ). Genes that show the strongest EBF1-addicted activation or repressive expression are listed.
Article Snippet: At day 11, the cells were stained with an antibody cocktail containing CD45 (30-F11, AF700, BioLegend, 103128), CD19 (1D3, PECy7, eBioscience, 15360900), Thy1.2 (53-2.1, Pacific Blue, BioLegend, 140306), Gr1 (RB6-8C5, APCCy7, BioLegend, 108424), CD11b (M1/70, APCCy7, BioLegend, 101226), NK1.1 (PK136, PE, BioLegned, 108708) and CD11c (N418, APC, BioLegned, 117310).
Techniques: Gene Expression, Quantitative Proteomics, Generated, In Vitro, Incubation, Control, RNA Sequencing, Activation Assay, ChIP-sequencing, Expressing
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![EBF1 interacts with the regulatory elements of broadly expressed genes (A) Heatmap with the z -scores of normalized gene expression of Ebf1-bound elements in early lymphoid progenitors from the Immgen dataset (GEO: GSE100738 ). Percentage of genes in each cluster is presented. (B) Gene ontology (GO) analysis of enriched biologic processes in genes bound by EBF1 at promoters or distal regions. Gene sets are categorized based on differential expression between Wt and Ebf1 −/− FL pro-B cells [Up-/Down-regulated: padj 0.05 & |log2FC| >1, common: log2FC < log2(1.5)]. (C) Diagram displaying the relative number of generated <t>CD19</t> + cells after 11 days of in vitro incubation of KIT + cells on OP9 stroma cells compared to R26 control. The red to blue color scale represents log2FC Wt vs. Ebf1 −/− from RNA-seq. Data are based on at least 3 separate experiments, and samples (n = 3–12) with a single guide or pool of individual guides are aggregated per gene. Statistical analysis is based on Student’s t test using the R26 control guide as reference: ∗ p < 0.05, ∗∗ p < 0.01, ∗∗∗ p < 0.001, ∗∗∗∗ p < 0.0001. (D) Promoter and distal elements associated to the gene sets from 2B are identified by ICE-A interaction-based annotation with H3K4me3 PLAC-seq data from FL Wt pro-B cells. Fraction of elements with a significant change (padj <0.05 & abs(log2FC) > 1) in accessibility between Wt and Ebf1 −/− FL pro-B cells are visualized in a stacked barplot, split into categories based on gene category and EBF1 occupancy. (E) The Myc locus is shown as an example of a gene that shows EBF1-addicted activation, visualized in the WashU genome Epigenome Browser. Selected EBF1-bound elements located in a distal enhancer region ∼1.6 Mb from the Myc TSS are highlighted in gray. The epigenome browser tracks include EBF1 ChIP-seq (GEO: GSE159957 ), ATAC-seq data from Wt/Ebf1 −/− FL pro-B cells and H3K4me3 PLAC-seq interactions from Wt FL pro-B cells. (F) Venn diagram displaying the overlap between common genes (defined as in B) and genes with differential expression at EBF1 degradation (|log2FC| > log2(1.5)), from ( GSE201141 ). Genes that show the strongest EBF1-addicted activation or repressive expression are listed.](https://pub-med-central-images-cdn.bioz.com/pub_med_central_ids_ending_with_5072/pmc12275072/pmc12275072__gr3.jpg)
