Particulate pollutant-induced trained immunity is mediated by H3K27ac in circulating monocytes
Next, we asked whether pollution-induced trained immunity is specifically mediated by H3K27ac modifications. Epigenomic ChIP Seq profiling of pollutant (SRM-1648a) versus media trained circulating monocytes, freshly purified from a healthy donor, revealed H3K27 acetylation patterns, depending on the training type, throughout the genomic regions that include promoter-transcription start sites (TSS) (Fig. 5A-B). In promoters, operationally defined as regions within 3 kb of a TSS, we identified H3K27ac marks that specifically associate with pollutant exposure, 6 days post training (Fig. 5C). In line with the inflammatory signature (Fig. 3-4), we detected an enhanced H3K27ac in the promoter-TSS of the genes encoding inflammatory mediators such as IL-1β, IL-6, IL-8, and MIP-1α upon pollutant training, arguing that this training drives a pro-inflammatory transcriptional program in monocytes (Fig. 5D). Training with pollutant enhanced H3K27ac marks in the promoter-TSS of IRF 8 and AHR , which encode essential transcription factors, Interferon Regulatory Factor 8 and Aryl Hydrocarbon Receptor. Furthermore, the strongest H3K27ac signal among transcription factors was detected upstream of TCF3 , encoding transcription factor 3. Finally, pollution-induced training of monocytes increased H3K27ac in promoter-TSS of SETD1A encoding SET Domain Containing 1A, a dedicated H3K4 methyltransferase (Fig. 5D). To understand the biological pathways affected during pollutant-induced trained immunity, we performed a gene ontology (GO) and KEGG pathway analysis using clusterProfiler R package (21 ). As shown in Fig. 5E, H3K27ac was highly enriched in the promoter-TSS of genes involved in inflammatory responses such as chemokine and TNF signaling, consistent with increased secretion of inflammatory mediators such as IL-8, and TNF. Interestingly, other inflammatory pathways relevant to asthma immunopathogenesis such as MAPK signaling, TCR activation, Fcγ receptor-mediated phagocytosis, autophagy, NOD-like receptor signaling were found among the pathways with highest H3K27ac mark specifically activated by particulate pollutants in circulating monocytes. The comprehensive list of genes, peaks, and pathways specific to pollutant-induced training have been summarized in Supplementary Table 2-4, respectively. In addition, untrained monocytes, baseline control group, possess a specific H3K27ac pattern of specific genes and peaks as well as biological pathways, e.g. tight junctions, cell adhesion, and insulin secretion as shown in Fig. 5E and Supplementary Table 5-7. Finally, monocytes underwent training with β-glucan, served as positive control, for 6 days (Table 8). Collectively, pollution training leaves H3K27ac scars in specific genome regions that are not affected in the control untrained monocytes.