DNA methylation mapping identifies gene regulatory effects in patients with systemic lupus erythematosus

Abstract

Objectives: Systemic lupus erythematosus (SLE) is a chronic autoimmune condition with heterogeneous presentation and complex aetiology where DNA methylation changes are emerging as a contributing factor. In order to discover novel epigenetic associations and investigate their relationship to genetic risk for SLE, we analysed DNA methylation profiles in a large collection of patients with SLE and healthy individuals.

Methods: DNA extracted from blood from 548 patients with SLE and 587 healthy controls were analysed on the Illumina HumanMethylation 450 k BeadChip, which targets 485 000 CpG sites across the genome. Single nucleotide polymorphism (SNP) genotype data for 196 524 SNPs on the Illumina ImmunoChip from the same individuals were utilised for methylation quantitative trait loci (cis-meQTLs) analyses.

Results: We identified and replicated differentially methylated CpGs (DMCs) in SLE at 7245 CpG sites in the genome. The largest methylation differences were observed at type I interferon-regulated genes which exhibited decreased methylation in SLE. We mapped cis-meQTLs and identified genetic regulation of methylation levels at 466 of the DMCs in SLE. The meQTLs for DMCs in SLE were enriched for genetic association to SLE, and included seven SLE genome-wide association study (GWAS) loci: PTPRC (CD45), MHC-class III, UHRF1BP1, IRF5, IRF7, IKZF3 and UBE2L3. In addition, we observed association between genotype and variance of methylation at 20 DMCs in SLE, including at the HLA-DQB2 locus.

Conclusions: Our results suggest that several of the genetic risk variants for SLE may exert their influence on the phenotype through alteration of DNA methylation levels at regulatory regions of target genes.

Keywords: gene polymorphism; systemic lupus erythematosus.

Figure 1

Results from the case–control epigenome-wide association study (EWAS) in systemic lupus erythematosus (SLE) in the discovery cohort. Inner circle: circular Manhattan plot of the results of the SLE case–control association analysis. P values are presented on the −log10 scale where the innermost scale line represents 10^–214. Middle circle: average methylation difference (∆β) between patients with SLE and controls for the top 100 differentially methylated CpG sites in the EWAS (scale −0.4 to 0.4). Green bars indicate decreased methylation and orange bars represent increased methylation levels in patients compared with controls. The outer circle represents chromosomes 1–22.

Figure 2

Violin plot of the DNA methylation levels at the CXCR6 gene in patients with systemic lupus erythematosus (SLE) and control individuals. Methylation levels at the CpG site cg08450017 in CXCR6 were increased in patients with SLE compared with controls in both the discovery and replication cohorts (P discovery=1.4×10^–130 and P replication=1.8×10⁻⁵¹). Median methylation beta values are represented by the white horizontal lines in the violin plots.

Figure 3

Enrichment of associated genetic variants in systemic lupus erythematosus (SLE) to methylation quantitative trait loci (meQTL) single nucleotide polymorphisms (SNPs) for differentially methylated CpG sites (DMCs) in SLE. The x-axis represents bins of P values from an SLE case–control genetic association analysis including 1135 Swedish patients with SLE and 2931 control individuals. Light grey bars represent SNPs which are significant meQTLs for CpG sites differentially methylated in SLE (466 CpG sites; 5307 SNPs). Bars in darker grey represent all SNPs on the ImmunoChip (133 838 quality controlled SNPs).

Figure 4

Illustration of genetic regulation of DNA methylation at the UBE2L3 genetic susceptibility locus for systemic lupus erythematosus (SLE) from a genome-wide association study (GWAS). The UBE2L3 locus on chromosome 22 with the differentially methylated CpG site (DMC) cg06850285 from the epigenome-wide association study is indicated at the top panel. The middle panel represents significant methylation quantitative trait loci (meQTLs) in controls only (illustrated by blue lines) or shared in both cases and controls (illustrated by black lines). The GWAS index single nucleotide polymorphism (SNP) is indicated in green. The bottom panel illustrates the RefSeq genes in the region.