Fig. 4: LN location defines the epigenomic landscape of non-endothelial SCs.

CD45^-CD24^-CD31^-Pdpn⁺ non-endothelial SCs were isolated from mLNs and pLNs of 6–10 weeks old GF or 7–12 weeks old SPF mice, and WGBS (A–C), ATAC-seq (D–G), or RNA-seq (F–H) analyses were performed. A–C DMRs were identified in colonization- (SPF vs. GF) and location-dependent (mLN vs. pLN) pairwise comparisons (2–3 replicates per condition). Scale bar depicts the extent of methylation with 100% being fully methylated and 0% being non-methylated. A The heatmap represents the mean methylation of significant DMRs within the promotor region of 286 genes. B Heatmaps represent CpG methylation of exemplary DMRs. The distance from the TSS is indicated and the size of genomic loci denoted in base-pairs (bp). C The heatmap represents the mean methylation of significant DMRs within the promotor region of TFs. D, E DARs were identified in colonization- (SPF vs. GF) and location-dependent (mLN vs. pLN) pairwise comparisons (3 replicates per condition). D Volcano plots of mean ATAC-seq FPKM comparing indicated samples. The number of DARs (top) and genes (bottom) is indicated per comparison. E GO analysis of biological processes of genes with location-dependent DARs (enrichment analysis of GO terms calculated with Fisher´s exact test). The numbers denote GO identifiers. F–H DEGs (3 replicates per condition) and DARs were identified in mLN vs. pLN pairwise comparisons. F Colored numbers in the scatterplot represent the number of genes with DAR and/or differential expression. Only genes with accessible loci within the promotor region were included in the analysis. On the x-axis, log₂(FC) of accessibility per DAR is plotted and on the y-axis, the log₂(FC) of gene expression for the comparison of mLN vs. pLN. G GO analysis of biological processes of DEGs and/or genes with at least one DAR (enrichment analysis of GO terms calculated with Fisher´s exact test). H Heatmaps represent the expression of all DEGs within the GO groups highlighted in (G). The numbers denote GO identifiers. ATAC-seq assay for transposase accessible chromatin sequencing, DAR differentially accessible region, DEG differentially expressed gene, det detection, DMR differentially methylated region, FC fold change, FPKM fragments per kilobase of peak per million reads, GF germ-free, GO gene ontology, mLN mesenteric lymph node, met metabolism, neg negative, pLN peripheral skin-draining lymph node, pos positive, reg regulation, SC stromal cell, sig signaling, SPF specific pathogen-free, TF transcription factor, TSS transcription start site, WGBS whole-genome bisulfite sequencing. Source data are provided in a Source data file.