Epigenetic control of innate and adaptive immune memory

Abstract

Clonal expansion and immunological memory are hallmark features of the mammalian adaptive immune response and essential for prolonged host control of pathogens. Recent work demonstrates that natural killer (NK) cells of the innate immune system also exhibit these adaptive traits during infection. Here we demonstrate that differentiating and 'memory' NK cells possess distinct chromatin accessibility states and that their epigenetic profiles reveal a 'poised' regulatory program at the memory stage. Furthermore, we elucidate how individual STAT transcription factors differentially control epigenetic and transcriptional states early during infection. Finally, concurrent chromatin profiling of the canonical CD8⁺ T cell response against the same infection demonstrated parallel and distinct epigenetic signatures defining NK cells and CD8⁺ T cells. Overall, our study reveals the dynamic nature of epigenetic modifications during the generation of innate and adaptive lymphocyte memory.

Fig. 1 |. NK cells undergo dynamic chromatin changes during viral infection.

a, Schematic of timepoints assayed for time course of ATAC-seq (n = 3 or 4 samples per d) and RNA-seq profiling (n = 2 samples per d). b, Number of DA (false discovery rate (FDR) < 0.05) regions that either gain (red) or lose (blue) chromatin accessibility at indicated transition timepoints. c, Absolute numbers and proportions of all DA regions versus high-fold change (FC; absolute log₂(FC) > 1) regions. d, Shown are line graphs (left) and heatmap (right) of high-FC peaks. Line plots showing mean (red line) and s.d. (gray ribbon) of mean-centered normalized log₂ values for each high-FC cluster. Heatmap is hierarchically clustered based on all high-FC log₂ peak counts (see Supplementary Fig. 1e) and shows the top 10% most variable regions within each cluster, with stable and transient clusters as indicated. e, Heatmap of top 20 most enriched pathways of any high-FC cluster shown as −log₁₀ raw binomial P values, as calculated by Genomic Regions Enrichment of Annotations Tool (GREAT). TCR, T cell antigen receptor.

Fig. 2 |. NK cells show epigenetic and transcriptional coordination during early stages of viral infection.

a, PCA of ATAC-seq and RNA-seq data on all atlas peaks and all genes. b, Hierarchical clustering on scaled Euclidean distances of normalized log₂ values of ATAC-seq and RNA-seq data. c, Heatmap of significant Spearman correlations (FDR < 0.05) for indicated gene sets (y axis). All coefficients shown are greater than 0.4. d, Scatter plots of average ATAC-seq and RNA-seq log₂(FC) for the top six gene sets ranked by Spearman coefficient, categorized by ATAC-seq peak type. ATAC-seq FC reflects a summarized value calculated from peak types and transition days (shown as numerator/denominator), as indicated within parentheses.

Fig. 3 |. STAT4 and STAT1 mediate chromatin remodeling in distinct manners.

a, Bar plot showing proportions of STAT-occupied regions categorized by peak type. b, Number of STAT-occupied DA regions (FDR < 0.05) that either gain (red) or lose (blue) accessibility at indicated transition timepoints. c, MA plots of DA regions categorized by peak type comparing Stat4^−/− (blue) or Stat1^−/− (green) to wild-type (WT) 2 d postinfection (n = 2 or 3 per condition). d, Scatter plot of log₂(FC) of differentially expressed (DE) genes (FDR < 0.05) comparing Stat4^−/− (blue) or Stat1^−/− (green) to WT 2 d postinfection (n = 2 or 3 per condition). e, Venn diagram of overlapping gene loci that are occupied (top) by indicated STAT protein, and DE (left) and DA (right) upon STAT deficiency. Darker center circle indicates number of DE genes with overlapping DA and occupied peak regions. PI, postinfection. f, Heatmap of gene expression from overlapping genes highlighted in e. g, Genomic tracks of mapped STAT4 ChIP-seq and Stat4^−/− vs. WT ATAC-seq reads, zoomed in on regions of interest. Y axis depicts normalized counts, while x axis displays genomic axis with scale bar. Bar plots quantify number of reads within highlighted peak regions (dashed lines), shown as mean ± s.d.

Fig. 4 |. Memory NK cells are epigenetically distinct from naive NK cells.

a, Enrichment of pathway terms on filtered DA regions (FDR < 0.05, absolute log₂(FC) > 0.5, assigned to genes that show significant changes by RNA-seq at any transition timepoint) between memory and naive NK cells shown as bar plots of −log₁₀ binomial raw P value. b, Select gene loci from reported pathways shown in a. Left: genomic tracks, with dashed lines indicating DA peak regions. Y axis depicts normalized counts, while x axis displays genomic axis with scale bar. Right: gene expression as normalized log₂ counts. Line represents the mean. c, Enriched de novo motifs found by Hypergeometric Optimization of Motif Enrichment (HOMER) among filtered DA regions, as described in a. ISRE data from GSE23622; TCF-LEF data from SRA012054; NF-κB data from MA0778.1 of JASPAR database (http://jaspar.genereg.net/); ISRE, interferon-stimulated response element; IRF, interferon-regulatory factor; HMG, high-mobility group transcription. d, Metacoverage of DA median normalized tag counts surrounding de novo motif regions listed in c found among all filtered DA regions. e, Select DA gene loci that contain motif instances, with actual sequence in black and de novo motif sequence below. Y axis depicts normalized counts, while x axis displays genomic axis with scale bar. BCR, B cell antigen receptor.

Fig. 5 |. Naive NK cells resemble memory CD8⁺ T cells.

a, PCA of ATAC-seq on all common cell-normalized log₂ values between Ly49H⁺ NK cells and m45-specific CD8⁺ T-cells (n = 2 or 3 per condition) during MCMV infection. b, Hierarchical clustering of Euclidean distances calculated from cell-normalized log₂ values. c, Heatmap of CD8⁺ T cells and NK cells depicting differentially accessible regions based on CD8⁺ T cell comparisons.

Fig. 6 |. Memory NK cells and CD8⁺ T cells possess common epigenetic signatures.

a, Heatmap of log₂(FC) comparing chromatin accessibility (left) and transcription (right) of memory and naive timepoints for NK cells and CD8⁺ T cells. b, Fold change of transcription and accessibility from commonly regulated peaks comparing memory and naive cell states in both NK cells and CD8⁺ T cells. DA regions (FDR < 0.05, absolute log₂(FC)> 0.5) are represented by number of dots, with colors showing magnitude and direction of FC. Extreme outliers of accessibility FC were capped at the 75th percentile±3× the interquartile range. Position of bottom circle represents transcriptional FC of differentially expressed genes (FDR < 0.05, absolute log₂(FC) > 1). Shown are genes with average transcripts-per-million (TPM) counts greater than 5 across naive, effector, and memory timepoints. c, Table of top enriched motifs from all common DA regions shown in a that are either higher in memory or lower in memory compared to naive. AP-1 data from GSE36099; TCF-LEF data from MA0769.1 of JASPAR database; bZIP, basic leucine zipper domain. d, Metacoverage of normalized tag counts surrounding de novo motif regions listed in c among all common DA regions. e, Select DA gene loci that contain motif instances with genomic sequence in black and de novo motif sequence below. Y axis depicts normalized counts, while x axis displays genomic axis with scale bar. Bar plots quantify number of reads overlapping the motif center (dashed line), shown as mean ± s.d.