IL2RA Methylation and Gene Expression in Relation to the Multiple Sclerosis-Associated Gene Variant rs2104286 and Soluble IL-2Rα in CD8+ T Cells

Abstract

CD8⁺ T cells are involved in the pathogenesis of multiple sclerosis (MS). The interleukin-2 receptor α (IL-2Rα) is important for CD8⁺ T cell function, and single nucleotide polymorphisms (SNPs) in the IL2RA gene encoding IL-2Rα increase the risk of MS. Therefore, in isolated CD8⁺ T cells we investigated IL2RA gene methylation and gene expression in relation to the MS-associated IL2RA SNP rs2104286 and soluble IL-2Rα (sIL-2Rα). We have identified allele specific methylation of the CpG-site located in intron 1 that is perturbed by the rs2104286 SNP in CD8⁺ T cells from genotype-selected healthy subjects (HS). However, methylation of selected CpG-sites in the promotor or 5'UTR region of the IL2RA gene was neither associated with the rs2104286 SNP nor significantly correlated with IL2RA gene expression in HS. In CD8⁺ T cells from HS, we explored expression of immune relevant genes but observed only few associations with the rs2104286 SNP. However, we found that sIL-2Rα correlated negatively with expression of 55 immune relevant genes, including the IL-7 receptor gene, with Spearman's rho between -0.49 and -0.32. Additionally, in HS by use of flow cytometry we observed that the IL-7 receptor on naïve CD8⁺ T cells correlated negatively with sIL-2Rα and was downregulated in carriers of the rs2104286 MS-associated risk genotype. Collectively, our study of resting CD8⁺ T cells indicates that the rs2104286 SNP has a minor effect and sIL-2Rα may negatively regulate the CD8⁺ T cell response.

Keywords: CD8+ T cells; IL-7 receptor alpha; IL2RA; allele specific methylation; interleukin-2 receptor alpha; multiple sclerosis; rs2104286; sCD25.

Figure 1

(A) Overview of the 6 CpG-sites in the IL2RA regulatory region and intron 1 that passed quality control and were methylated above background. Four CpG-sites were in the region corresponding to the 5’UTR and 1 CpG-site was located upstream of this in the promotor region. CpG10:6057082 was located in intron 1 at the site of rs2104286. (B) Methylation at the CpG10:6057082 that is perturbed by the MS-associated IL2RA rs2104286 SNP in healthy subjects (CC = 20; CT = 5; TT = 25) and MS patients (CC = 1; CT = 7; TT = 4). Of the 50 HS, 38 were from cohort 1 and 12 were from cohort 2. Data were analysed by one-way analysis of variance. Error-bars represent mean with SD.

Figure 2

Surface expression of the IL-7Rα on CD8⁺ T cells in relation to sIL-2Rα and the MS-associated SNP rs2104286. (A) Spearman correlation between sIL-2Rα and frequency (freq.) of IL-7Rα⁺CD8⁺ T cells in healthy subjects (N = 33). (B) The freq. of IL-7Rα⁺ cells on naïve, CM, EM and T_EMRA subsets. (C) Spearman correlation between sIL-2Rα and the freq. of IL-7Rα⁺ naïve (CD45RA^hiCCR7⁺) CD8⁺ T cells, freq. of IL-7Rα⁺ central memory (CM, CD45RA^-CCR7⁺) CD8⁺ T cells, freq. of IL-7Rα⁺ effector memory (EM, CD45RA^-CCR7^-) CD8⁺ T cells as well as freq. of IL-7Rα⁺ terminal effector memory (T_EMRA, CD45RA^hiCCR7^-) CD8⁺ T cells in healthy subjects (N = 31). Gating strategy is presented in Supplementary Figure 2. (D) The MS-associated SNP rs2104286 in relation to the freq. of IL-7Rα⁺CD8⁺ T cells (TT = 18, CC = 14) and the freq. of IL-7Rα⁺ naïve CD8⁺ T cells (TT = 18, CC = 13) in genotype-selected healthy subjects. Genotype comparisons were analysed by a non-parametric Mann-Whitney U test. Error-bars represent median with interquartile range. ns, non-significant.