Fig. 5: Defective JAK2/STAT5 signaling in CTCF-deficient BMDCs leads to downregulation of NF-κB pathway component genes.

a Western blotting performed with the indicated antibodies. The data are representative of three independent experiments with similar results. b Relative signal intensities of proteins in a were measured using ImageJ software. Error bars represent mean ± standard error of the mean (s.e.m). Significance was calculated using an unpaired two-tailed t-test using n = 3 independent samples. c Histogram showing the average tag density of STAT5 ChIP-seq peaks called for WT and KO BMDCs. d Heatmap of ChIP-seq signals with ±2 kb of unique STAT5 peaks comparing STAT5 enrichment (fold change > 1.3; FDR < 0.05) between WT and KO BMDCs. The data are representative of two independent experiments with similar results. e A pie chart depicting the numbers of genes, assigned to “lost in KO” STAT5 peaks and categorized by the overlap feature of the STAT5 peaks with promoters, distal enhancers, or both. f RNA-seq MA plot for the genes assigned to “lost in KO” STAT5 peaks. The number of genes exhibiting >1.3-fold decreases in WT (red) or KO (blue) BMDCs with a false discovery rate <0.05 has been indicated. g Enrichment of pathway terms on the “Lost in KO” STAT5 target genes whose RNA expression was downregulated in KO. Significance was calculated by one-sided Fisher’s Exact test. h Changes in RNA expression of the genes associated with the pathway term of “Regulation of NF-κB activity”. Error bars represent mean ± standard error of the mean (s.e.m.). n = 3 biologically independent samples. i Snapshots displaying virtual 4C plots, ChIP-seq signal tracks, and significant loops (from top to bottom) at the Trim25 (left) and Irak2 (right) loci. Virtual 4C plots (V4C) shows normalized H3K27ac HiChIP loop strength (represented as -Log10(Q)) with the TSSs of Trim25 (left) and Irak2 (right) genes as the viewpoint. IGV browser shows ChIP-seq signal tracks for STAT5, CTCF, SMC1, H3K27me3, H3K4me3, H3K4me1, and H3K27ac. Arcs show significant interactions with –Log10(Q) ≥ 5. Only the loops interacting with the viewpoint have been displayed. Source data are provided as a Source Data file.