Activation of T lymphocytes , the key cells of the adaptive immune system, requires simultaneous engagement of the T-cell receptor by MHC class II peptides and costimulatory molecules. The cytotoxic T-lymphocyte-associated antigen 4 (CTLA4) gene located on chromosome 2q33 encodes CD152, a costimulatory molecule which acts as a vital negative regulator of T-cell activation and proliferation [106]. CTLA4 competes with costimulator CD28 for binding to B7 on antigen-presenting cells. The critical role of this molecule is demonstrated in CTLA4 knockout mice which die prematurely, at an age of 2–3 weeks, due to severe lymphoproliferation, lymphocytic infiltration and destruction of major organs [107]. In humans, mutations in the CTLA4 gene result in an immune dysregulation syndrome [108]. CTLA4 gene polymorphisms have been linked with susceptibility to a number of autoimmune diseases including autoimmune thyroid disease [109, 110], type 1 diabetes mellitus [111, 112], rheumatoid arthritis [113] and coeliac disease [114]. The association between CTLA4 gene variants and AAD has been reported in a number of studies in different European populations [83–85, 115, 116]. The most commonly described CTLA4 polymorphisms in AAD populations are non-synonymous polymorphism in exon 1 region of CTLA4 gene +49 A → G (Ala17Thr) [83, 84, 115], (AT)n dinucleotide repeat polymorphism within the 3′ untranslated region [116] and G or A alleles of the JO30 SNP downstream of this gene [85]. Polymorphisms in the latter are postulated to be the causative variants affecting the relative amount of the soluble and membrane-bound CTLA4 and therefore enabling CD28 to access more of its ligand, resulting in T-cell activation [117]. In contrast to this hypothesis, a number of studies have demonstrated that individuals with autoimmune conditions have, in fact, increased serum levels of soluble CTLA4 isoforms [118, 119] suggesting that the complexity of CTLA4-CD28 interaction and signalling is incompletely understood. Another mechanism of negative immune regulation by CTLA4-positive cells is the ability of CTLA4 to capture and degrade CD80 and CD86 (ligands shared with the stimulatory receptor CD28) from antigen-presenting cells [120].

Another key autoimmunity gene, PTPN22 on chromosome 1p13, encodes a tyrosine phosphatase which is a crucial regulator of immune homeostasis, inhibiting T-cell receptor (TCR) signalling. The association of a missense SNP (1858C > T) in PTPN22, encoding an arginine to tryptophan substitution at amino acid 620, has been identified in type 1 diabetes [121], rheumatoid arthritis [122], systemic lupus erythematosus [123] and Graves’ disease [79]. For a number of autoimmune diseases, this allelic variant ranks as the most important non-MHC single gene contributor to disease susceptibility. A number of stu dies have implicated this variant in susceptibility to AAD [79, 81, 82]. The functional effect of 1858 C > T polymorphism has not been fully elucidated. Some studies suggest that this variant results in increased suppression of TCR signalling [124, 125], while others suggest the opposite [126, 127]. One study suggested that proteolytic binding and cleavage of Arg620Trp is increased with consequent reduction in LYP levels in T and B cells leading to lymphocyte and dendritic cell hyperresponsiveness and autoimmunity [127].

Programmed death ligand-1 (PD-L1, CD274, B7-H1) is a costimulatory molecule binding the PD-1 moiety of T cells, leading to downregulation of cytokine production and T-cell activation, thereby inducing immune tolerance. PD-L1 variants have been implicated in susceptibility to AAD, although the effect on risk is very modest (OR 1.34 for the allele with the strongest association) [88].

Interleukin-2 (Il-2) and its receptor are important determinants of the immune response. Il-2 is a potent T-cell growth stimulator and influences T-cell differentiation, in particular formation of the regulatory T cell (T_reg) lineage. T_reg cells are crucial in maintaining self-tolerance due to their ability to suppress autoreactive T cells which escape negative selection in the thymus. IL2RA gene encodes the alpha subunit (CD25) of the IL2 receptor, a unique subunit conferring high affinity to IL2. Polymorphic variants of IL2 (4q27) and IL2RA (10p15.1) genes have been associated with type 1 diabetes and rheumatoid arthritis [128–130]. The C minor allele in IL2 conferred protection from AAD in a Polish cohort. However, there was no association found between IL2 polymorphisms and Norwegian patients with AAD [73]. Similarly, an association between IL2RA polymorphisms and susceptibility to AAD was found in Norwegian subjects, but this finding has not been replicated in British or Polish series [73, 90]. Although there is known genetic heterogeneity between various AAD populations, the association between AAD and IL2 or IL2RA needs further replication in larger cohorts.

STAT4 (signal transducer and activator of transcription 4 ) on chromosome 2q32 encodes a transcription factor which is implicated in Th-1 and Th-17 differentiation and activation. Polymorphisms of STAT4 have been shown to be associated with rheumatoid arthritis [131–133], systemic lupus erythematosus [122] and type 1 diabetes [134]. A meta-analysis by Mitchell et al. revealed a significant association between STAT4 polymorphisms and AAD in European populations [76]. The allelic variability identified in AAD maps to intron 3 of the STAT4 gene and is in moderate-to-strong linkage disequilibrium with STAT4 polymorphisms identified in the other autoimmune conditions. The exact mechanism by which polymorphisms in this gene lead to autoimmune disease remains unknown.

The GATA3 gene on chromosome 10p14 encodes a C2C2-type zinc finger transcription factor which regulates a number of steps in T-cell development and differentiation. In particular, this transcription factor has been shown to be the Th2 lineage master regulator [135] and could therefore contribute to T-cell dysregulation present in autoimmune disease. A recent meta-analysis demonstrated an association between GATA3 polymorphisms and AAD in European cohorts [76]. The minor G allele at rs3802604 was protective for AAD (OR 0.9). This finding is, however, in contrast to the known association of the same allele with susceptibility to rheumatoid arthritis [136] possibly reflecting immunopathogenic differences between these two conditions.

The BACH2 gene on chromosome 6q15 plays a vital role in CD⁴⁺ T-cell differentiation; in particular it is crucial in the formation of T_reg, which are the key cells in maintaining immune tolerance. Polymorphisms at the BACH2 locus have been associated with type 1 diabetes, generalised vitiligo, autoimmune thyroid disease, Crohn’s disease and coeliac disease [137–141]. Recently, an association between the minor T allele at rs3757247 in the BACH2 locus and susceptibility to AAD was described in UK and Norwegian cohorts [91].

The first linkage study in multiplex AAD families implicated regions on chromosomes 6 (corresponding to the HLA region), 7, 9 and 18 in the susceptibility to AAD [92]. A follow-on study, looking at 64 SNPs underlying the linkage peaks on chromosomes 9 and 18 conducted in case-control cohorts from the UK, Norway and Sweden, revealed nominal association with three independent SNPs in chromosome 18 and AAD. One of these encodes a non-synonymous variant (pCys751Gly) in exon 9 of the NFATC1 (nuclear factor of activated T cells, cytoplasmic, calcineurin-dependent 1) gene. Upon T-cell activation, family members of NFAT translocate to the nucleus where they can activate target genes and as such play a central role in gene transcription during the immune response [142]. NFATC1 has been shown to play a role in the regulation of PD-1 expression, a cell surface receptor functioning as an immune checkpoint and reducing T-cell activation [143].

The GPR174 (G protein-coupled receptor 174) gene at Xq21.2 consists of one exon encoding a protein which belongs to the G protein-coupled receptor superfamily and is involved in immune cell signal transduction. Recently a lysophosphatidylserine (LysoPS) was found to be a ligand for GPR174, and the interaction between the two has been s hown to stimulate an increase in the intracellular cyclic adenosine monophosphate (cAMP) [144]. cAMP has been previously demonstrated to be a potent negative regulator of T-cell immune function [145]. These findings offer a plausible link between GPR174 polymorphisms and autoimmunity. Polymorphisms in GPR174 have been found to be associated with Graves’ disease [146] [147]. A serine to proline non-synonymous variant in GPR174 has been associated with AAD [89]. The localisation of GPR174 on chromosome X and its role in AAD autoimmunity are particularly interesting given the gender bias observed in this disorder, although it is unlikely that a single gene is responsible for the higher susceptibility of females to develop AAD (Fig. 4.3).

Vitamin D has been recognised for its effects on the immune system. On a molecular level, the active form of vitamin D, 1,25-dihydroxyvitamin D, leads to reduced expression of HLA class II molecules on endocrine cells and inhibits T-cell proliferation [148, 149]. 1,25-Dihydroxyvitamin D is also implicated in innate immunity by inhibiting differentiation of dendritic cells which are potent antigen-presenting cells [150]. Polymorphisms in the vitamin D receptor (VDR) gene, located on chromosome 12q12-14, have been studied in a number of autoimmune conditions including type 1 diabetes and autoimmune thyroid disease with conflicting results [151–154]. Although specific genotypes in VDR have been associated with AAD risk in a relatively small German cohort, no such association was found for an individual VDR allele [77]. Additional studies are required to confirm VDR as a susceptibility locus for AAD. In contrast to this, a promoter polymorphism in the CYP27B1 gene (−1260C > A) has been shown to be associated with AAD in two independent cohorts [74, 75]. More recently, an intronic SNP, rs4646536, in CYP27B1, was associated with AAD in a large meta-analysis of several European cohorts [76]. CYP27B1 hydro xylase catalyses the conversion of 25-hydroxyvitamin D to its active form, 1,25-dihydroxyvitamin D. A promoter polymorphism in CYP27B1 might affect enzyme transcription and thus the rate of final hydroxylation of 1,25-dihydroxyvitamin D.

The Fc receptor-like 3 gene (FCRL3) located on chromosome 1q21 encodes an orphan cell surface receptor belonging to the immunoglobulin receptor superfamily, expressed predominantly on B lymphocytes. A polymorphism in the FCRL3 promoter region (FCRL3_3*C) has been implicated in susceptibility to rheumatoid arthritis, autoimmune thyroid disease and systemic lupus erythematosus in Asian cohorts [155]. Contrary to the findings in other autoimmune conditions, a study of a UK AAD cohort found that the FCRL3_3*C variant confers protection from the disease [78]. The allele most associated with disease risk in this cohort was found to be FCRL3_3*T (OR = 1.61). Based on functional studies of this locus, FCRL3_3*T is predicted to result in lower promoter activity. Such contradictory findings for one haplotype conferring both protection and disease susceptibility in different populations illustrate the complexity of the genetic underpinning of polygenic autoimmune diseases including AAD.

NLRP1 (nuclear localisation leucine-rich-repeat protein 1 ) is a regulator of the innate immune response. NLRP1 belongs to the NOD-like receptor family and participates in recognising microbial products, such as lipopolysaccharide, and assembly of inflammasomes, cytoplasmic protein complexes mediating pro-inflammatory responses via cytokine activation [156]. Polymorphisms of the NLRP gene have been reported to confer risk for a number of autoimmune conditions including vitiligo [157], type 1 diabetes [86], coeliac disease [158] and rheumatoid arthritis [159]. A coding variant of NLRP1 (Leu155His) has been associated with AAD in two European cohorts [86, 87]. Surprisingly, different alleles were found to confer risk of AAD in Polish (minor allele A) and Norwegian (major allele T) populations.

The CLEC16A (C-type lectin domain family 16) gene encodes a protein of unknown function but which is almost exclusively expressed in immune cells such as dendritic cells, B lymphocytes and natural killer cells. This makes it a plausible susceptibility gene for autoimmunity. A polymorphism in the CLEC16A gene (intronic SNP rs12917716) was found to be associated with AAD in a Norwegian cohort with an OR of 0.71. Comparable effects of CLEC16A SNPs were previously demonstrated in cohorts of subjects with type 1 diabetes (OR 0.65 to 0.83) [160, 161].