IRF5 haplotypes demonstrate diverse serological associations which predict serum interferon alpha activity and explain the majority of the genetic association with systemic lupus erythematosus

Abstract

Objective: High serum interferon α (IFNα) activity is a heritable risk factor for systemic lupus erythematosus (SLE). Auto-antibodies found in SLE form immune complexes which can stimulate IFNα production by activating endosomal Toll-like receptors and interferon regulatory factors (IRFs), including IRF5. Genetic variation in IRF5 is associated with SLE susceptibility; however, it is unclear how IRF5 functional genetic elements contribute to human disease.

Methods: 1034 patients with SLE and 989 controls of European ancestry, 555 patients with SLE and 679 controls of African-American ancestry, and 73 patients with SLE of South African ancestry were genotyped at IRF5 polymorphisms, which define major haplotypes. Serum IFNα activity was measured using a functional assay.

Results: In European ancestry subjects, anti-double-stranded DNA (dsDNA) and anti-Ro antibodies were each associated with different haplotypes characterised by a different combination of functional genetic elements (OR>2.56, p<1.9×10(-14) for both). These IRF5 haplotype-auto-antibody associations strongly predicted higher serum IFNα in patients with SLE and explained >70% of the genetic risk of SLE due to IRF5. In African-American patients with SLE a similar relationship between serology and IFNα was observed, although the previously described European ancestry-risk haplotype was present at admixture proportions in African-American subjects and absent in African patients with SLE.

Conclusions: The authors define a novel risk haplotype of IRF5 that is associated with anti-dsDNA antibodies and show that risk of SLE due to IRF5 genotype is largely dependent upon particular auto-antibodies. This suggests that auto-antibodies are directly pathogenic in human SLE, resulting in increased IFNα in cooperation with particular combinations of IRF5 functional genetic elements. SLE is a systemic autoimmune disorder affecting multiple organ systems including the skin, musculoskeletal, renal and haematopoietic systems. Humoral autoimmunity is a hallmark of SLE, and patients frequently have circulating auto-antibodies directed against dsDNA, as well as RNA binding proteins (RBP). Anti-RBP autoantibodies include antibodies which recognize Ro, La, Smith (anti-Sm), and ribonucleoprotein (anti-nRNP), collectively referred to as anti-retinol-binding protein). Anti-retinol-binding protein and anti-dsDNA auto-antibodies are rare in the healthy population. These auto-antibodies can be present in sera for years preceding the onset of clinical SLE illness and are likely pathogenic in SLE.

Figure 1

(A) Diagram of the IRF5 gene indicating the location of previously described functional elements in European ancestry. The first three darker boxes indicate alternately spliced first exons (exons 1a, 1b and 1c), lighter boxes indicate subsequent exons (2–9), and the black box indicates the 3′ UTR. Arrows show the location of the SNPs included in this study. Haplotype diagrams are shown for European ancestry (B.) and African–American (C.) patients with SLE, constructed using Haploview 4.2 using the solid spine of linkage disequilibrium (LD) method. Pairwise r² values are shown in the boxes, and darker shading indicates increasing r² values. IRF, interferon regulatory factors; SLE, systemic lupus erythematosus; SNPs, single nucleotide polymorphisms; UTR, untranslated region.

Figure 2

Serum IFNα activity in patients with SLE and healthy first-degree relatives. A. Serum IFNα activity in European ancestry patients with SLE (n=312) stratified by the number of TACA and TATA haplotypes and the presence or absence of anti-dsDNA antibodies. B. European ancestry patients with SLE (n=312) stratified by the number of TACA haplotypes and the presence or absence of positive test for anti-Ro antibodies (anti-Ro+ antibodies). C. African–American patients with SLE (n=206) stratified by presence or absence of the European-derived IRF5 SLE-risk haplotype (TACA) and presence of anti-Ro auto-antibodies. Units on the Y-axis represent a summed IFNα-induced gene expression score as measured by our functional assay. Bars indicate the median, error bars show the interquartile range, p-values calculated from the Mann–Whitney U test. dsDNA, double-stranded DNA; IFN, interferon; SLE, systemic lupus erythematosus.

Figure 3

Schematic diagram of the proposed relationships between SLE-specific auto-antibodies, IRF5 genotype and IFNα in SLE pathogenesis. 1. IRF5 haplotypes could influence susceptibility to form particular auto-antibodies (arrow indicated by a?). 2. Immune complexes containing SLE-specific auto-antibodies are taken into cells, and the nucleic acid component triggers endosomal TLR7 and TLR9. 3. Particular IRF5 SLE-risk variants augment downstream IFNα production in the setting of different auto-antibodies, resulting in high serum IFNα (4.) and subsequent risk of SLE (5.). IFN, interferon; IRF, interferon regulatory factors; SLE, systemic lupus erythematosus; TLR, toll-like receptors.