IRF5 activation in monocytes of SLE patients is triggered by circulating autoantigens independent of type I IFN

Systemic lupus erythematosus (SLE) is a complex systemic autoimmune disorder characterized by multiple immunologic abnormalities that lead to a break in self-tolerance and the production of autoantibodies targeting nucleic acids and associated proteins. Patients display elevated type I IFN in their serum and IFNα-induced gene transcripts in their blood cells that correlates with disease activity and severity (reviewed in (1)). While the underlying etiology of SLE remains obscure, significant evidence documents the importance of genetic risk factors and environmental stressors. Genetic variants of IFN regulatory factor 5 (IRF5) have been strongly associated with the risk of SLE in large-scale genetic association studies of patients with different ancestral backgrounds (2–5). IRF5 is a transcription factor that controls inflammatory and immune responses. It regulates the expression of type I IFNs, IFN stimulated genes (ISGs), and inflammatory cytokines/chemokines in response to pathogenic stimuli (6–9); it is also a critical mediator of cellular apoptosis (10, 11). In recent work, we have shown that IRF5 expression and alternative splicing were significantly upregulated in primary blood cells of SLE patients (12). In particular, elevated IRF5 protein expression in monocytes of SLE patients was associated with the IRF5 SLE homozygous risk haplotype (4xCGGGG indel, T-allele of SNP rs2004640, A-allele of SNP rs10954213, and C-allele of SNP rs10488631) (12). Basal/constitutive IRF5 expression in immune cells of healthy donors differs significantly between immune cell subpopulations but is generally cytoplasmic and highest in monocytes and plasmacytoid dendritic cells (PDC) as compared to T, B and NK cells (13). Each of these immune cell subpopulations display aberrant activation and function in SLE that is likely to contribute to disease pathogenesis (14). Monocytes/macrophages (Mo/Mϕ) have been increasingly recognized to play a dynamic role in the initiation and perpetuation of SLE given their hallmark functions in phagocytosis and antigen presentation (15, 16). Mo are a key component of the innate immune system involved in the regulation of the adaptive immune response (17). The Mo/Mϕ system, including conventional DC, has versatile cellular functions aimed to defend, regulate inflammation, and induce immunity; consequently, it maintains a central role in initiating the immune response. Deficiencies in any of these cellular functions would be expected to contribute to systemic autoimmunity. Indeed, recent studies in lupus patients have identified numerous Mo/Mϕ defects involving aberrant activation as measured by surface protein expression, deregulation of cytokine production, and defective phagocytic capacity (16, 18, 19). The mechanisms underlying these abnormalities remain unclear. Given the association of elevated IRF5 expression in Mo of SLE patients with the IRF5 risk haplotype (12), we sought to examine the “activation” status of IRF5 in SLE Mo as compared to other immune cell subpopulations. IRF5 is constitutively localized to the cytoplasm in unstimulated cells (7) and generally requires a stimulus that leads to activation/nuclear localization in order for it to function as a transcription factor. We and others have demonstrated that IRF5 becomes post-translationally modified in response to a variety of stimuli leading to its translocation from the cytoplasm to the nucleus (6, 7, 9, 20–22). In this study, we assess the in vivo “activation” status of IRF5 in immune cells of genotyped SLE patients and healthy donors using Amnis ImageStream technology to detect IRF5 cellular localization and determine the impact of circulating serum factors on this dynamic process.