Figure 5. Characteristic patterns of chromatin accessibility distinguish between LCH cell subsets.

A) Scatterplots contrasting differential gene expression with differential chromatin accessibility between progenitor-like LCH-S1 cells and the more differentiated LCH subsets LCH-S12 (top) and LCH-S11 (bottom). The x-axis denotes differences in gene expression (mean normalized UMI count based on scRNA-seq) and the y-axis displays the mean difference in chromatin accessibility over all regulatory regions linked to the respective gene (mean of two replicates, mean normalized reads per million [RPM]). Colored points denote LCH-subset-specific marker genes (from Fig. 3D).

B) Heatmap showing ATAC-seq signal intensity for 1,964 differentially accessible regions identified in pair-wise comparisons of one LCH cell subset against the two other subsets. Regions are grouped into six modules based on differential chromatin accessibility. ATAC-seq signal intensity scores are scaled by column (ATAC-seq peaks) for better visualization of differences.

C) Enrichment analysis with Enrichr for genes linked to each of the six modules of LCH-subset-specific chromatin accessible regions, showing the top-3 most enriched terms ordered by the Enrichr combined score⁵⁸. FDR-adjusted p-value: *, p < 0.1; **, p < 0.05. Further details are provided in Supplementary Table S3.

D) Heatmap of transcription factor binding enrichment for LCH-subset-specific modules, based on LOLA³¹ analysis using a large collection of ChIP-seq peak profiles. Top: LOLA enrichments colored by the log₂ odds ratio of enriched overlap between the region modules and ChIP-seq peaks for the corresponding transcription factor, compared to the background of all regulatory regions in the ATAC-seq dataset. Bottom: Mean expression (normalized UMI count) of the gene encoding each transcription factor in different LCH subsets (showing gene expression in LCH_E, which matches the ATAC-seq data, as well as the mean expression across all patient samples). Further details are provided in Supplementary Table S3.

E) Enrichment of transcription factor binding motifs for LCH-subset-specific modules, based on HOCOMOCO³² motif occurrences identified using FIMO⁵⁹. Top: Motif enrichments colored by the log₂ odds ratio of enriched overlap between the module regions and DNA motif hits for the corresponding transcription factor, compared to the background of all regulatory regions identified in the ATAC-seq dataset. Bottom: Expression levels of the genes encoding the corresponding transcription factors (as in panel D). Right: Consensus DNA motif for selected transcription factors. FDR-adjusted p-value, Fisher’s exact test: *, p < 0.1; **, p < 0.05, ***, p < 0.005.