Characterization of epigenetic deregulation in patients with CH with DNMT3A mutations. (A) Proportion of each cell type identified in the scRNA-seq data from the 10x multiome platform stratified by 5 conditions as shown in the y-axis. The healthy cohort is further stratified by age: <50 and >50 years, respectively. (B) Violin plots showing cell-type–specific changes in chromatin accessibility measured as the total number of cut sites (sum of TF-IDF–normalized cut site counts across all peaks; scATAC-seq data from multiome) in each cell type. Only cell types with >100 cells are shown. Black dots show the mean value. Not significant (ns), P > .05; ∗P ≤ .05; ∗∗P ≤ .01; ∗∗∗P ≤ .001; ∗∗∗∗P ≤ .0001 (Wilcoxon rank sum test). NK and CD4 T cells showed significant increase in chromatin accessibility in patients with DNMT3Amt CH compared with TET2mt CH in the context of COVID-19. (C) Coverage plot showing epigenomic dysregulation of IL-32 in DNMT3Amt CH compared to TET2mt CH, through multiomics analysis. The plot shows the coaccessible peaks with IL-32 transcription start site (TSS) in patients with TET2mt CH and those with DNMT3Amt CH, both with COVID-19 (coaccessibility score > 0.1; blue and red arcs), the chromatin accessibility signal per group of cells, IL-32 gene expression (violin plots; right), candidate cis-regulatory elements predicted by ENCODE (colored-coded bars), open chromatin peaks (gray bars), differentially accessible peaks that are more accessible in patients with DNMT3Amt CH than those with TET2mt CH in CD4 T cells (light blue bars), CpG sites hypomethylated in patients with DNMT3Amt CH compared with patients with TET2mt CH (dark blue bars), and CpG sites overlapping open chromatin regions (black bars) around the IL-32 gene locus. Labeled loci A and B are chr16:3123999-3124965 and chr16:3263558-3264913, respectively. These loci are regions in which patients with DNMT3Amt CH gained accessibility in CD4 T cells, overlapped with hypomethylated CpG sites and gained coaccessibility with IL-32 TSS. (D) Box plots showing methylation levels (β values) per patient in TET2mt CH and DNMT3Amt CH cohorts (both with COVID-19) at 3 hypomethylated CpG sites shown in panel C. The middle line represents the median; the lower and upper edges of the rectangle represent the first and third quartiles, respectively; and the lower and upper whiskers represent the interquartile range × 1.5. The groups were compared using Wilcoxon rank sum test. (E-F) Violin plots showing significant increase in chromatin accessibility in DNMT3Amt cells compared with DNMT3Awild-type cells as determined by GoTChA analysis. The data shown are the total number of cut sites (E) and the number of cut sites at loci A and B from panel C (F), in DNMT3A wild-type and DNMT3Amt cells from 2 samples (DNMT3Amt clonal cytopenias of undetermined significance) profiled using GoTChA. Red dots show the mean value. Mutation site in the DNMT3A gene is shown in the bottom of panel E. ns, P > .05; ∗P ≤ .05; ∗∗P ≤ .01; ∗∗∗P ≤ .001; ∗∗∗∗P ≤ .0001 (Wilcoxon rank sum test). cDC, classical dendritic cells; CTCF, CCCTC-binding factor; Dist., distal; gdT, γδ T cells; Int, intermediate; MAIT, mucosal-associated invariant T cells; Mono, monocytes; pDC, plasmacytoid dendritic cells; Prox., proximal; Treg, regulatory T cells.

Characterization of epigenetic deregulation in patients with CH with DNMT3A mutations. (A) Proportion of each cell type identified in the scRNA-seq data from the 10x multiome platform stratified by 5 conditions as shown in the y-axis. The healthy cohort is further stratified by age: <50 and >50 years, respectively. (B) Violin plots showing cell-type–specific changes in chromatin accessibility measured as the total number of cut sites (sum of TF-IDF–normalized cut site counts across all peaks; scATAC-seq data from multiome) in each cell type. Only cell types with >100 cells are shown. Black dots show the mean value. Not significant (ns), P > .05; ∗P ≤ .05; ∗∗P ≤ .01; ∗∗∗P ≤ .001; ∗∗∗∗P ≤ .0001 (Wilcoxon rank sum test). NK and CD4 T cells showed significant increase in chromatin accessibility in patients with DNMT3Amt CH compared with TET2mt CH in the context of COVID-19. (C) Coverage plot showing epigenomic dysregulation of IL-32 in DNMT3Amt CH compared to TET2mt CH, through multiomics analysis. The plot shows the coaccessible peaks with IL-32 transcription start site (TSS) in patients with TET2mt CH and those with DNMT3Amt CH, both with COVID-19 (coaccessibility score > 0.1; blue and red arcs), the chromatin accessibility signal per group of cells, IL-32 gene expression (violin plots; right), candidate cis-regulatory elements predicted by ENCODE (colored-coded bars), open chromatin peaks (gray bars), differentially accessible peaks that are more accessible in patients with DNMT3Amt CH than those with TET2mt CH in CD4 T cells (light blue bars), CpG sites hypomethylated in patients with DNMT3Amt CH compared with patients with TET2mt CH (dark blue bars), and CpG sites overlapping open chromatin regions (black bars) around the IL-32 gene locus. Labeled loci A and B are chr16:3123999-3124965 and chr16:3263558-3264913, respectively. These loci are regions in which patients with DNMT3Amt CH gained accessibility in CD4 T cells, overlapped with hypomethylated CpG sites and gained coaccessibility with IL-32 TSS. (D) Box plots showing methylation levels (β values) per patient in TET2mt CH and DNMT3Amt CH cohorts (both with COVID-19) at 3 hypomethylated CpG sites shown in panel C. The middle line represents the median; the lower and upper edges of the rectangle represent the first and third quartiles, respectively; and the lower and upper whiskers represent the interquartile range × 1.5. The groups were compared using Wilcoxon rank sum test. (E-F) Violin plots showing significant increase in chromatin accessibility in DNMT3Amt cells compared with DNMT3Awild-type cells as determined by GoTChA analysis. The data shown are the total number of cut sites (E) and the number of cut sites at loci A and B from panel C (F), in DNMT3A wild-type and DNMT3Amt cells from 2 samples (DNMT3Amt clonal cytopenias of undetermined significance) profiled using GoTChA. Red dots show the mean value. Mutation site in the DNMT3A gene is shown in the bottom of panel E. ns, P > .05; ∗P ≤ .05; ∗∗P ≤ .01; ∗∗∗P ≤ .001; ∗∗∗∗P ≤ .0001 (Wilcoxon rank sum test). cDC, classical dendritic cells; CTCF, CCCTC-binding factor; Dist., distal; gdT, γδ T cells; Int, intermediate; MAIT, mucosal-associated invariant T cells; Mono, monocytes; pDC, plasmacytoid dendritic cells; Prox., proximal; Treg, regulatory T cells.

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