A Functional Genomics Pipeline to Identify High-Value Asthma and Allergy CpGs in the Human Methylome

BackgroundDNA methylation of cytosines at CpG dinucleotides is a widespread epigenetic mark; but genome-wide variation has been relatively unexplored due to the limited representation of variable CpGs on commercial high-throughput arrays. ObjectiveTo explore this hidden portion of the epigenome, we combined whole-genome bisulfite sequencing (WGBS) with in silico evidence of gene regulatory regions to design a custom array of high-value CpGs. We focused these studies in airway epithelial cells from children with and without allergic asthma because these cells mediate the effects of inhaled microbes, pollution, and allergens on asthma and allergic disease risk. MethodsWe identified differentially methylated regions (DMRs) from WGBS in nasal epithelial cell (NEC) DNA from 39 children with and without allergic asthma of both European and African ancestries. We selected CpGs from DMRs, previous allergy or asthma Epigenome-Wide Association Studies (EWAS), or Genome-Wide Association Study (GWAS) loci, and overlapped them with functional annotations for inclusion on a custom Asthma&Allergy array. Using both the Custom and EPIC arrays, we performed EWAS of allergic sensitization (AS) in NEC DNA from children in the URECA birth cohort and using the Custom array in the INSPIRE birth cohort. We assigned each CpG on the arrays to its nearest gene and its promotor capture Hi-C interacting gene and performed expression quantitative trait methylation (eQTM) studies for both sets of genes. ResultsCustom array CpGs were enriched for intermediate methylation (IM) levels compared to EPIC CpGs. IM CpGs were further enriched among those associated with AS and for eQTMs on both arrays. ConclusionsOur study revealed signature features of high-value CpGs and evidence for epigenetic regulation of genes at AS EWAS loci that are robust to race/ethnicity, ascertainment, age, and geography. Clinical ImplicationsThese studies identified allergic sensitization-associated differentially methylated CpGs and their target genes in airway epithelium, providing potential epigenetic mechanisms in the development of allergic diseases and suggesting novel drug targets. Capsule SummaryThis study of previously unexplored regions of the airway epithelial methylome revealed novel epigenetic mechanisms regulating genes previously implicated in the pathogenesis of asthma and allergic diseases.