Novel Insights into the Autoimmunity from the Genetic Approach of the Human Disease
Pärt Peterson
Abstract
Autoimmune-polyendocrinopathycandidiasis-ectodermal dystrophy (APECED) is a monogenic inborn error of autoimmunity that is caused by damaging germline variants in the AIRE gene and clinically manifests with multiple autoimmune diseases in patients. Studies on the function of the AIRE gene, discovered in 1997, have contributed to fundamental aspects of human immunology as they have been important in understanding the basic mechanism of immune balance between self and non-self. This chapter looks back to the discovery of the AIRE gene, reviews its main properties, and discusses the key fndings of its function in the thymus. However, more recent autoantibody proflings in APECED patients have highlighted a gap in our knowledge of the disease pathology and point to the need to revisit the current paradigm of AIRE function. The chapter reviews these new fndings in APECED patients, which potentially trigger new thoughts on the mechanism of immune tolerance.
P. Peterson (*)
Institute of Biomedical and Translational Medicine, University of Tartu, Tartu, Estonia e-mail: part.peterson@ut.ee
Keywords
APECED · AIRE gene · Thymus · Immune tolerance · Autoimmunity
1.1 Introduction
The immune system protects us from harmful agents while keeping body cells and tissues intact and functional. To maintain this delicate balance, it needs to be controlled and regulated. During the last decades, studies of human monogenic diseases and their parallel animal models have revealed an important understanding of the pathogenesis of these diseases.
In addition, studies on certain so-called experiments of nature, have provided broader insight into the basic functioning of the human immune system and have underscored the fundamental roles of individual genes in keeping an immune balance between self and non-self. Their associated functional and regulatory pathways are central to the development and maintenance of immune cells, contributing to immune tolerance and homeostasis. Consequently, monogenic autoimmune diseases are highly informative for our understanding of immune balance as they link defned monogenic defects with clinical phenotypes.
© The Author(s), under exclusive license to Springer Nature Singapore Pte Ltd. 2024 M. Matsumoto (ed.), Basic Immunology and Its Clinical Application, Advances in Experimental Medicine and Biology 1444, https://doi.org/10.1007/978-981-99-9781-7_1
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Monogenic inborn errors of autoimmunity typically manifest at an early age and have a more severe disease course than multifactorial autoimmune diseases. Another characteristic feature is that they often involve multiple organ systems with variable severity. While the cases of monogenic autoimmunity are individually rare, the prevalence of individual symptoms may vary considerably. This chapter discusses autoimmunepolyendocrinopathy-candidiasis-ectodermal dystrophy (APECED) or autoimmune polyendocrine syndrome (APS1) as one of the most well-known examples of monogenic autoimmune disease. I will give a retrospective on the AIRE gene identifcation by our team in the 1990s and then an overview of the AIRE gene that is mutated in APECED, and discuss the clinical and immunological features of this disease.
1.2 The Identifcation of the AIRE Gene
The monogenic basis of APECED disease was known already for a long time. In the 1960s, reports pointed to the clinical and genetic heterogeneity among patients with Addison’s disease and that Addison’s disease could be part of two separate and distinct clinical syndromes, depending on the type of associated disorders [1, 2]. This led later to the distinction between APS1 and APS2 [3]. Finnish physician Jaakko Perheentupa gave APS1 the alternative name APECED to highlight its clinical picture and monogenic etiology.
My frst encounter with APECED disease was when I started working at the University of Tampere, Finland, in the early 1990s. My supervisor was Professor Kai Krohn, who had already worked with APECED patient-derived autoantibodies in the 1970s [4]. These autoantibodies reacted with protein fractions extracted from the adrenal cortex. The topic of my PhD thesis became the cloning and characterization of these autoantigens, using lambda phage cDNA library screening. As a result of this, we were able to identify three target proteins belonging to the P450 cytochrome family—steroid 17 hydroxy-
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lase (P450c17), steroid 21-hydroxylase (P450c21), and side chain cleavage enzyme (P450scc) [5–7], associated with the Addison’s disease in APECED.
Along with the screening of autoantibodies, we had several contacts with the patients and their family members, which enabled us to collect peripheral blood material for cells and serum, and other tissue samples from the Finnish patients. Therefore, quite soon after completing my thesis in 1996, we became interested in identifying the gene mutated in APECED. From the beginning, it was obvious that the gene should somehow function in the regulation of the immune system and impact the mechanism of immune tolerance, differentiating between self and non-self responses.
At the same time, Professor Leena Peltonen’s research group at the University of Helsinki was working on gene identifcation, linking the disease mutations to 21q22.3, the terminal end of the longer arm of chromosome 21 [8]. The chromosomal region 21q22.3 was problematic as markers on this region showed an exceptionally high recombination rate, but their linkage disequilibrium analysis narrowed the critical genomic region to around 500 K base pairs.
In the mid-1990s, the sequence of the human genome was not yet identifed, but there was an ongoing global effort, the Human Genome Project, to sequence all the genes of the human genome. We soon understood that we needed to cooperate with experts in human genome analysis and gene cloning. Kai contacted Prof Stylianos Antonarakis from the University of Geneva and Dr. Hamish Scott, who did his postdoc in Geneva. They were a perfect team to cooperate on the identifcation and characterization of novel, human disease-associated genes, which were at that time mapped to fragments of the human genome sequence. They also had been collecting DNA samples from Swiss and North Italian APECED patients, which, along with Finnish samples, made up a cohort of patients from different ethnic origins.
At that time, only a handful of genes were known in the 21q22.3 genomic region. One known gene localized to the APECED region was
encoding liver-type phosphofructokinase (PFKL), which later turned out as a neighboring gene to AIRE. PFKL, however, was soon excluded as an APECED gene as it had a very distinct function and was expressed outside of immune tissues. Another candidate gene that was located at the end of chromosome 21 and sharing amino acid sequence similarity with P450 steroid hydroxylase autoantigens, lanosterol synthase (LSS), proved to be quickly wrong too.
The sequences of the large fragments of genomic DNA from chromosomal regions became available in GenBank as bacterial artifcial chromosomes (BACs). The genome was mapped by markers, usually specifc nucleotide repeats, whose relative positions or links to each other were known. However, one of the challenges in the search for the genes was that their exact position and order on BAC and other large segments of the genome were unknown. It was also not clear how to map the correct exons and introns to the genomic sequence, and the main key to this was to use a large collection of expressed sequence tags (ESTs) and exon prediction programs. ESTs were fragments of mRNA sequences derived through sequencing reactions performed on randomly selected clones from cDNA libraries of human tissues. They were usually short, less than a few hundred nucleotides, but were produced and made available in GenBank in large batches. Nevertheless, fnding a gene was more like seeking a needle in a haystack. Most of the work was using a Blast program to pull together the EST fragments and align them to the genomic BAC regions. The hope was to get an intact structure of the new gene with proper exons and introns, and their junction regions using Grail and Genscan programs, which looked for open reading frames and exon-intron boundaries in the genomic sequence. The next task was to align the translated sequence to known proteins to see whether it has meaningful similarity to protein domains. Then we sequenced the exon regions in the patients and healthy controls to identify possible mutations that disrupt the reading frame, repeating this procedure over again. We went through a number of exons of candidate genes, including CFPA410
then known as C21ORF2, which is 30 K base pairs away from the AIRE gene and causes inherited retinopathies [9].
Finding novel genes from the human genome was a tedious task. Stylianos Antonarakis suggested that we should team up with Professor Nobuyoshi Shimizu from Keio University, Tokyo, whose research group worked on sequencing the terminal region of chromosome 21 as part of the Human Genome Project. Human chromosome 21 was considered a model chromosome for the Human Genome Project because of its small size and its association with Down syndrome. Prof Shimizu had a large lab that worked on sequencing human chromosomal fragments and on the identifcation of novel disease genes, which also included APECED. They had constructed a human genomic bacterial artifcial chromosome (BAC) library consisting of tens of thousands of clones, each containing an average DNA insert size of 100 K base pairs, and covering the human genome three times. Dr. Jun Kudoh, who was the key investigator on the APECED project from Shimizu’s lab, and his colleague Dr. Kentaro Nagamine, studied the BAC contigs from the 21q22.3 region consisting of the sequences of DNA segments that overlapped in a way that provided a contiguous representation of this region [10]. One of these contigs included a 450 K base pair region covering the critical region identifed by Peltonen’s team. They were immediately eager to work together on this joint effort. The collaboration between the three research groups consisted of frequent emails and phone calls, and rapid sharing of the data when one or another team discovered a small additional piece of this larger mosaic. The entire collaboration was highly inspiring and stimulating, and at times, it felt like an obsession as thinking about the APECED gene took up most of my awake brain capacity.
Meanwhile, we continued working to characterize the expression of the putative transcripts in immune tissues. We used PCR-amplifed cDNA probes from predicted exons to perform northern blotting with RNA from immune organs. Most of the probes gave no signal, but one probe, with the name GR1, gave strong positive bands identify-
ing several transcripts in the thymus tissue and lower expression in lymph nodes. However, the spleen, peripheral blood leukocytes, bone marrow, and appendix were all negative, as well as the autoimmune target tissues of APECED (adrenal gland, liver, and pancreas). Considering our main hypothesis that APECED is caused by a defect in immune tolerance, the fnding of strong expression in the thymus was very promising.
In the summer of 1997, the sequencing at Shimizu’s laboratory revealed APECED mutations in the gene, which overlapped with the GR1 probe that hybridized to the thymic RNA in the northern blot. This was the breakthrough in the hunt for the gene. The novel gene was located just proximal to the earlier known PFKL gene, which we have excluded as the APECED gene. The gene had 14 exons, and the mutation in exon 6, causing a stop codon at position 257 (R257X), was present in several patients. Two Swiss patients from one family and four Finnish patients were homozygous for the R257X allele, while other Finnish patients were heterozygous. The sequencing at Shimizu’s lab, Keio University, soon identifed other mutations in the same gene. Further rapid amplifcation of cDNA ends (RACE) showed a new transcript that consisted of 1635 base pairs and also identifed minor splice variants.
The initial working name was the APECED gene. However, using the disease name for a gene was considered misleading as there were examples where defects in several genes caused a disease or vice versa; various mutations in a gene could cause different forms of diseases like muscular dystrophy. Therefore, we initially proposed to name AIR for Autoimmune Regulator. When the paper by Nagamine et al. was submitted to Nature Genetics, the editors notifed us that another transcript is called AIR and we should change the name. Indeed, the noncoding antisense transcript from the IGF2R gene, involved in genetic imprinting, was called AIR in the literature (now AIRN). To make the distinction, we renamed the gene AIRE.
From its protein sequence, it was obvious that AIRE was related to transcriptional control as it had a nuclear localization signal and two PHD-
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type zinc fnger motifs. Further cloning and transfection of the cDNA in fusion with GFP showed its location in nuclear dot-like bodies [11]. The mRNA in situ hybridization and antibodies to AIRE revealed its expression in rare epithelial cells in the thymic medulla and in vivo locations in similar nuclear bodies as after the transfection. The fnding that medullary epithelial cells in the thymus express AIRE was compatible with the defect in immune tolerance seen in APECED, and this was something that we expected as we knew that the negative selection of potentially self-reacting lymphocytes is taking place in the thymic medulla [12].
In parallel with our group, the team led by Prof Leena Peltonen identifed the gene and came to very similar conclusions. Their gene mapping approach was also based on studying BAC and cosmids clones, and their fuorescence in situ hybridization confrmed the chromosomal localization. By large-scale sequencing, they ended up with the APECED gene and its exon-intron structure in the vicinity of PFKL. Because R257X was present in most of the Finnish patients, they named it as a Finnish major mutation and calculated its carrier frequency as 1:250 in Finland. Both papers were published in Nature Genetics in the December 1997 issue back-to-back [13, 14].
After the gene identifcation, we had several meetings at Keio University hosted by Prof Shimizu. At that time, it was fascinating to see the huge sequencing capacity of their facilities, having a row of multi-capillary ABI3700 machines, the main DNA sequencer of that time, whereas we had only one. Shimizu had many projects ongoing in parallel including the identifcation of Parkin (PRKN) gene causing juvenile Parkinson’s disease and the sequencing of medaka, a small freshwater fsh living in rice paddies in Japan. Through this international collaboration, we later cloned and characterized another AIRE-neighboring gene, DNMT3L, which interacts with DNMT3A and DNMT3B in de novo DNA methylation [15, 16]. Over the subsequent 2 years, our team spent considerable time identifying APECED-causing mutations in the AIRE gene [17–19].
The discovery of AIRE contributed to a new period of understanding thymic immune tolerance. At the same period with AIRE cloning, the seminal studies on the expression of tissuespecifc antigens in the medullary thymic epithelial cells (mTECs), called “ectopic” or “promiscuous” gene expression, were made by Bruno Kyewski and Ludger Klein, and others [12, 20, 21]. Their experiments demonstrated that quantitative as well as qualitative differences in transcriptional regulation of intrathymic gene expression can set the threshold for tolerization and lead to the exclusion of self-antigens from central tolerance and thus predispose to autoimmunity. A critical proof demonstrating Aire role in immune tolerance came from studies on Airedefcient mice by Mark Anderson, Christophe Benoist, and Diane Mathis and colleagues at Harvard University in 2002 [22].
1.3 AIRE Has Domains Characteristic of the Transcriptional Regulator
AIRE protein sequence and domain structure are well conserved among mammalians [23]. Most of the placental mammalian species have substantial similarity not only in protein sequence but also in upstream regulatory elements of the gene. Expectedly, the primates share the highest sequence similarity throughout the chromosomal region of the gene. The structure of Aire is also conserved in two other mammalian groups; marsupials (opossum) and monotremes (platypus), which both are evolutionally conserved with placental mammals in their immunoglobulin and T cell receptor loci [24]. Birds, amphibians, and fsh seem to have a gene that has similar domains with Aire in the N-terminal region but not in C-terminus, and its role in immune mechanisms in lower animal classes remains unknown.
AIRE domains highlight its role in transcriptional regulation: a conserved nuclear localization signal, the HSR and SAND domains in its N-terminus, and two PHD-type zinc fngers. Close to the HSR domain AIRE has a bipartite
nuclear localization signal for the transport into the nucleus, where it forms dot-like structures, which resemble PML bodies [25–27]. The SAND is a conserved domain found in a variety of proteins involved in transcriptional regulation and chromatin remodeling [28]. SAND was originally suggested to function as a DNA binding domain; however, instead of the critical KDWK amino acid motif needed for DNA binding, AIRE has a KNKA sequence. Within the N-terminal HSR and SAND regions, it has a similarity with the Sp100 protein, an autoantigen in primary biliary cirrhosis. The Sp100 protein family contains other members, Sp110 and Sp140 that, interestingly, also localize subcellularly to the nuclear bodies and have one PHD fnger but have a bromodomain instead of the second PHD fnger [29–31]. The structural similarity of Sp100 protein family members with AIRE provokes interesting parallels as they too have a role in immune regulation.
AIRE contains two PHD-type zinc fngers, which are the structurally conserved chromatinbinding domain of approximately 65 amino acids and are found in about 150 human proteins [32]. Many PHD fngers act as nucleosome interaction determinants and selectively bind unmodifed or methylated at lysine 4 histone H3 tails. Indeed, the frst PHD fnger of AIRE interacts with unmethylated histone H3 at lysine 4 (H3K4me3) and mediates AIRE binding to chromatin [33–37]. AIRE-PHD fngers are needed for its transcriptional activity, and pathological missense or truncation mutations in the patients disrupt their structures. The second AIRE PHD fnger, however, does not interact directly with histone H3 even though it is important for the transactivation capacity. The two AIRE-PHD fngers have independent non-interacting folds connected by an unstructured proline-rich linker, and the second PHD fnger likely acts in the interaction of other proteins via so far unknown chromatin-associated nuclear partners [38].
Through binding to critical chromosomal factors, AIRE enhances transcriptional elongation and promotes the upregulation of hundreds of genes in thymic epithelial cells [39–41]. The top interaction partners of AIRE are DNA-dependent
protein kinase (DNA-PK) and topoisomerase 2-alpha (TOP2A), which work together in relaxing torsional tensions of the DNA double helix [42–45]. Abramson et al. described in detail the AIRE interacting complex that contains DNA-PK, TOP2 but also poly ADP ribose polymerase 1 (PARP-1), and Ku proteins [42]. The interaction of AIRE with DNA-PK and TOP2A appears to be a key event in AIRE target gene expression. AIRE is an atypical transcriptional regulator with an unusual capacity to enhance gene expression without promoter and RNA processing elements [46]. It has been shown to interact with chromatin at super-enhancer regions where it regulates the expression of multiple genes, often located in clusters or larger chromosomal domains [47, 48].
1.4 AIRE Has a Unique Role in Guarding Self-Tolerance
AIRE has a central and unique role in the immune system as it promotes the expression of a large number of self-antigens in the thymus [49, 50]. The expression in mTECs matches with the high expression of MHC class II and CD80 to ensure effcient antigen presentation and is tightly controlled. AIRE is regulated by TNF family member RANK-RANKL and CD40-CD40L signaling, both activating the NF-κB pathway [51–53]. This upstream enhancer region is shared across mammals and is indispensable for RANK-induced Aire expression as a defect in these NF-κB sites completely abolishes Aire expression in the thymus [54, 55]. In the more proximal promoter region, the gene has a CpG island and its demethylation is necessary for the AIRE expression [56]. Also, a gene locus-insulating chromatin modifer CTCF and several transcriptional regulators such as Irf4 and Irf8 [57] and Ets factors, common for epithelial-specifc expression [58], add to the tight regulation of the gene. In addition to AIRE expression in mTECs, multiple studies have shown extrathymic expression of AIRE in secondary lymphoid organs, where it may support peripheral immune tolerance with various functions [59–61]. However, the peripheral
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expression level is substantially lower compared to the thymus, and its contribution to the peripheral immune tolerance beyond its role in the thymus would need more studies.
Aire-defcient mouse studies have provided insight into understanding the mechanism of Aire. Studies on Aire-defcient mouse models of APECED revealed that Aire plays an important role in the generation of T cell tolerance in the thymus, mainly by inducing the expression of a large repertoire of transcripts, many of which encode proteins normally restricted to organs residing in the periphery [49]. The lack of AIRE expression in the thymus results in a defect in the clonal deletion of autoreactive T cells and ultimately to autoimmunity [22, 62, 63]. Airedefcient mice exhibit a defective negative selection of thymic CD4+ T cells, leading to the presence of autoreactive T cells in the periphery. Aire defciency is closely associated with decreased production of thymic regulatory T cells (Tregs), which play a key role in suppressing autoreactive responses in the periphery. Airedefcient mice show decreased numbers of Tregs already in the perinatal period with a specifc subtype of Tregs lacking very early in life [64]. Most of the Treg specifcities are independent of Aire, whereas some Treg-specifc clones are Aire-dependent and likely drive the autoimmunity [65, 66]. Several studies have confrmed lower numbers of Tregs and/or impaired suppressive capacity in APECED patients [67–70] and fewer Treg clones with common TCRβ sequences, which instead were found among conventional T cells [71].
Single-cell studies have revealed remarkable heterogeneity in mTECs [47, 72, 73]. A recent study by Michelson et al. divided mTECs into various subpopulations based on their resemblance to peripheral tissues, which were called “mimetic cells” and suggested that Aire is partially and variably required for the development of these subpopulations [74]. They showed that Aire engages lineage-defning transcription factors to instigate peripheral cell types, in the context of earlier studies proposing the thymic medulla to function as a mixture of epithelial cells [75, 76]. Mouse studies have shown that
Aire is also necessary for the maturation program of mTECs and that the mTEC compartment does not mature normally in the absence of Aire [77, 78]. As its defciency in mice leads to impaired differentiation of the mTEC-high population in the thymus, it may affect the self-antigen expression via a scarcity of these various epithelial cell populations by developmental block [79].
1.5 APECED Starts Early in Childhood and Is Heterogenic
APECED is a rare disease but more frequent in some populations that have been isolated in the past, such as Iranian Jews (1:9000), Sardinians (1:14,000), and Finns (1:25,000) [80]. Although it can be also underdiagnosed, the prevalence in other countries is very low and varies from 1:43,000 in Slovenia to 1:500,000 in France [81, 82]. APECED usually starts in early childhood and is known for its classical triad of chronic mucocutaneous candidiasis (CMC), hypoparathyroidism, and Addison’s disease, the onset following the aforementioned order. However, multiple other diseases, and the majority of these are autoimmune, can manifest in the patients [80, 81, 83].
CMC usually affects the skin and mucosal tissues. Hypoparathyroidism, which is relatively rare alone, is the most frequent and sometimes the only endocrine component in APECED. Sporadic Addison’s disease is also rare, but common in APECED patients. Overall, the number and severity of clinical symptoms vary greatly between patients, even among those with identical mutations. The symptoms tend to increase with age with the median age of onset for the classical triad of the disease (CMC, hypoparathyroidism, and Addison’s disease) being 11 years [84]. The incidence at a later age is quite rare. The reasons why the parathyroid gland and adrenal cortex are the main autoimmune targets in APECED remain unknown, and would deserve investigation, but are challenging to study as animal models do not follow the same disease pattern. However, the selective destruc-
tion of the adrenal cortex should prompt future studies addressing the pathological etiology of Addison’s disease in APECED.
Other endocrine and nonendocrine autoimmune disorders may occur in APECED patients. The most common are type 1 diabetes (T1D), autoimmune thyroid disease, and hypophysitis. APECED (also known as APS1) is similar to APS2 in that patients with both autoimmune polyendocrinopathies develop Addison’s disease and have an increased risk to have T1D and/or autoimmune thyroid disease. However, APS2 is multifactorial and is associated with HLA, while APECED is not. Gonadal insuffciency is more common in females. Patients often suffer from gastrointestinal autoimmunity, enteropathy with malabsorption, hepatitis, autoimmune gastritis with or without vitamin B12 and iron defciencies, and exocrine pancreatitis. The patients have other lesions affecting ectodermal structures namely enamel dysplasia, vitiligo, alopecia, keratoconjunctivitis, and nail dystrophy. In addition, urticaria-like erythema and hyposplenism or splenic atrophy have been reported [84, 85], which increase the susceptibility to bacterial infections.
Interestingly, APECED appears to exhibit substantial clinical heterogeneity between ethnic populations, although systematic analyses are rare due to the low prevalence. For example, American patients do not seem to develop the full classical triad of candidiasis, hypoparathyroidism, and Addison’s disease as the frst manifestations. A systematic analysis of over 150 APECED patients from the US revealed enrichment of nonendocrine autoimmune manifestations relative to other APECED cohorts [83]. They developed a hexad of nonendocrine disease manifestations, including urticarial eruption, autoimmune gastritis, intestinal malabsorption, autoimmune pneumonitis, autoimmune hepatitis, and Sjögren’s-like syndrome, with much higher frequency (40–80%) compared to European APECED patients (<20%). Additionally, only 20% of the patients developed their frst two consecutive manifestations out of the classic triad. In contrast, around 80% of the patients developed three non-triad manifestations before eventually
reaching a classic diagnostic dyad, resulting in signifcant delays in establishing a clinical diagnosis. Some of the non-triad manifestations before the development of a classic diagnostic dyad in American APECED patients were urticarial eruption (which was the most common initial disease manifestation together with CMC), enamel hypoplasia, and intestinal malabsorption [83]. This has led to the redefned clinical features and diagnostic criteria for APECED, where the criteria for clinical diagnosis are fulflled if a patient develops 2 of the 6 manifestations, which include nonendocrine urticarial eruption, intestinal dysfunction, and enamel hypoplasia, along with classic components of CMC, hypoparathyroidism, and Addison’s disease.
1.6 AIRE Mutations: Expanding Universe
In addition to p.R257X in exon 6, another relatively common mutation found in patients from many countries is a 13 base-pair deletion (967–979del13bp) in exon 8. Certain mutations are population-specifc, especially those which have been isolated in the past. As mentioned earlier, the most common mutation in Finnish APECED patients is p.R257X. Other examples include the p.R139X mutation that is common among Sardinian [17], p.R203X among Sicilian [17–19, 86–88]. Many mutations are nonsense mutations or deletion/insertions causing the AIRE protein to lack distal functional domains. Missense mutations often occur in the HSR region responsible for the homodimerization and correct intracellular localization of the protein. Dominant AIRE mutations in one allele that increase susceptibility to autoimmune diseases have been reported. The dominant single AIRE point mutation G228W was reported in an Italian family with autoimmune thyroid disease. Overall, these patients had a milder phenotype than typical APECED [89]. This variant also caused a reduced expression of AIRE-dependent thymic gene expression in a mouse model [90].
Recent studies have revealed AIRE gene variants, which are heterozygous dominant-negative
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polymorphisms, mostly occurring in PHD fngers and causing a mild disease [91]. Genome-wide association studies have found an association of the AIRE variant (p.R471C) with Addison’s disease and pernicious anemia [92–94]. These variants are prevalent among healthy populations and occur in up to 1/1000 individuals, and their incomplete penetrance can be partly explained by other factors, like other modifer genes, epigenetic changes, or environmental factors. The classical phenotype is rarely seen in patients with heterozygous dominant variants although some have autoantibodies typical for APECED. Instead, the clinical picture is diverse, with enteropathy, immunodefciency, and vitiligo as common features. For now, more than 500 genetic variants are known, and many of them can potentially affect AIRE function. This remains an interesting topic and the mechanistic aspects of the new dominant AIRE mutations need to be identifed but suggest a dosage dependency of AIRE to maintain the balance of immune tolerance [94]. It should be noted that the relatives of APECED patients having defects in one allele seem not to have increased susceptibility to autoimmune diseases.
Despite considerable phenotypic variation, no strong correlation with genotype exists in APECED. Even siblings from the same families with identical gene mutations quite often present with different phenotypes, especially in the manifestation of those components that are outside of the classical clinical triad. Nevertheless, some genotype-phenotype associations seem to be more frequent than expected. For example, Iranian Jews with the p.Y85C mutation seem to have candidiasis and Addison’s disease at lower frequencies [95]. In mutation and phenotype analysis of large cohorts of patients, candidiasis has been found more often in patients with p.R139X and p.R257X mutations (stop codon before the SAND domain) than with the 967-979del13bp mutation [96, 97]. In the American APECED cohort with diverse genetic backgrounds, compound heterozygous AIRE mutations are frequent, with the two most common mutations being p.L323SfsX51 followed by p.R257X [98]. The p.L323SfsX51 mutation was
found to be associated with the development of nonendocrine autoimmune manifestations such as pneumonitis and hepatitis. However, most of the phenotypic variation of APECED remains unexplained and could be explained by the stochastic generation of T or B cell repertoire or variation in clonal deletion. APECED has no well-established association with the HLA genes, which distinguishes it from APS2 and sporadic Addison’s disease cases.
1.7 Autoantibodies to Intracellular Enzymes
The hallmark of APECED is the variety of hightiter autoantibodies to a large spectrum of autoantigens. A prominent set of APECED-associated autoantibodies are shared with multifactorial autoimmune endocrine diseases, and some of them are expressed tissue-specifcally. Addison’s disease in APECED is associated with autoantibodies to steroidogenic P450 cytochrome enzymes. These are steroid 21-hydroxylase (P450c21 or CYP21) expressed in the adrenal cortex, and steroid 17-alpha-hydroxylase (P450c17 or CYP17) and side chain cleavage enzyme (P450scc or CYP11A1), which are both expressed in adrenals and gonads [5, 6, 99, 100]. Autoantibodies directed against glutamic acid decarboxylases (GAD1 and GAD2) are characteristic of T1D and those against thyroid peroxidase (TPO) and thyroglobulin are frequently present in APECED patients with autoimmune thyroid disease [101, 102]. Other often found autoantibodies are against aromatic L-amino acid decarboxylase (AADC), tryptophan hydroxylase, tyrosine hydroxylase, and leucine-rich-repeat protein 5 (NALP5) [103–105]. The comprehensive analysis of autoantibodies in APECED offered systemic insight into the autoantibody targets, and, in addition to shared autoantigen reactivities, the patients display high inter-individual variation in their autoantigen profles, which collectively target many autoantigens [106]. Among the autoantigen proteins, we found reactivity to many cancer-testis antigens, implying a central role of the thymus in antitumor
immunity. These included members of MAGE-A, MAGE-B, and GAGE families, and sperm-specifc proteins, highlighting the role of AIRE in modulating immune responses to tumor antigens and implications for cancer immunotherapy [107]. The APECED autoantigens seem to have two major origins, which are the proteins expressed in thymic medullary epithelial cells and lymphoid cells [107]. The presence of autoantibodies varies between patients, and many are found in only a limited number of patients [106]. The development of a unique antibody repertoire may depend on an individual patient’s genetic background or may signal an indirect immune response to tissue destruction without a direct role in pathogenesis. Strikingly, many of these autoantigens are intracellular, and although some of them have a tissue-specifc expression like steroidogenic enzymes, others are expressed widely or ubiquitously in many tissues. In addition, the majority of identifed human autoantigens are not mTEC-specifc (and likely not AIRE-specifc), which questions the current paradigm of AIRE function in the thymus.
1.8 Autoantibodies to Cytokines Add Another Puzzle to the Tolerance Paradigm
An even more intriguing feature of APECED is the development of autoantibodies to cytokines, the modulators of immune responses. The most prominent are high-titer autoantibodies to type 1 interferon, mainly targeting IFN-α and IFN-ω with a prevalence of over 95% and much less IFN-β, which is found in around 20% of the patients. Anti-IFN-α and IFN-ω autoantibodies can be detected early in life, even before the clinical symptoms, and because of their high prevalence, they have become diagnostic or even prognostic markers for APECED [108–111]. They are highly neutralizing, specifc for native conformational epitopes, and inhibit IFNstimulated gene expression [97, 111, 112]. Interestingly, their neutralization capacity correlates inversely with the onset of clinical T1D in those APECED patients who have T1D-specifc
GAD65 autoantibodies suggesting that naturally arising autoantibodies may be protective against the development of T1D and that these autoantibodies may also have the potential for the therapeutic use [106].
IFN-α and IFN-ω are critical modulators in the protection of viral infections. Nevertheless, APECED patients do not develop severe viral infections that are seen in patients with inherited defciencies in interferon-receptor genes. The likely explanation is that viral infections are avoided by intact IFN-β function. It should be noted that some APECED patients have been reported to develop severe manifestations of skin varicella zoster (shingles) or mucosal herpes simplex infections [113]. Recent reports from the COVID-19 pandemic period showed that the presence of neutralizing autoantibodies to type 1 IFNs in APECED patients also confers a high risk of life-threatening COVID-19 pneumonia [114].
The susceptibility to CMC is associated with the immune response to Th17 cells and cytokines [115, 116]. APECED patients develop autoantibodies to Th17 cytokines, and they also emerge at an early stage of the disease. The autoantibodies to IL-22 are the most common, as they are present in over 90% of the patients. Anti-IL-17F and IL-17A autoantibodies are less frequent; anti-IL17F antibodies are found in approximately 75%, and anti-IL-17A in around 35–50% of patients with CMC [115]. Interestingly, APECED patients do not develop neutralizing autoantibodies to IL-17B and IL-17C. Accordingly, the production of IL-22 and IL-17F cytokines by stimulated blood and skin T cells is impaired in patients with CMC [115], and the neutralizing anti-IL-22 autoantibodies are present in patient saliva [117]. The contribution of Th17 cytokine autoantibodies to mucocutaneous protection needs further clarifcation, as in some patients, CMC occurs without detectable neutralizing autoantibodies, suggesting a broader dysfunction of Th17mediated immunity in APECED patients. Furthermore, the patients also have autoantibodies to type 3 IFN (IFN-λ), interleukins 5, 6, 18, and several other cytokines [107, 118].
P. Peterson
1.9 Do We Have a Full Understanding of How AIRE Regulates Immune Tolerance?
The reason for the development of anti-cytokine autoantibodies remains unknown, especially as the cytokines are not expressed tissue-specifcally and as autoimmune targets do not belong to the category of AIRE-induced organ-specifc autoantigens. Autoantibodies to type 1 IFNs are not entirely specifc to APECED. They are present in other thymus-related immune disorders such as in patients with thymoma [111], RAGhypomorphic patients who lack AIRE expression [119], IPEX patients [120] as well as in patients with systemic lupus erythematosus with increased expression of type 1 IFN [121]. Recent studies in COVID-19 patients have established that autoantibodies to type 1 IFN are associated with a severe course of COVID-19 and adverse reactions to the yellow fever vaccine and West Nile virus encephalitis [122, 123].
Yet, the main issue is that the development of high-titer neutralizing autoantibodies to multiple cytokines before the onset of the human disease does not ft into the current textbook paradigm of thymic tolerance due to the lack of AIREdependent control of cytokine expression. Their presence also hints at so far unknown thymic mechanism that keeps the balance of immune tolerance in humans. As they impact the type 1 IFN responses in the patients, they cannot be without an effect on disease, rather they have a key role in APECED pathology. The differences between human disease and mouse disease models have been raised and these concerns are justifed [124]. Although the regulation of ectopic gene expression in the mouse thymus is well-established, Aire-defcient mice develop relatively mild disease and the only animals that become sick are those backcrossed to NOD background, the autoimmune-prone mouse strain developing T1D. However, on NOD background, Aire knockout animals manifest not endocrine but exocrine pancreatitis, which is extremely rare in human APECED, and without destruction of the pancreatic islets although insulin expression in
the mouse thymus is controlled by Aire. Furthermore, the mouse models do not develop autoantibodies to type 1 IFN, which are present in almost 100% of the patients. In contrast to mice, Aire-defcient rats develop autoantibodies to type 1 IFNs and other cytokines, and seem to more accurately resemble APECED but also suggest differences in mouse and rat immune systems [125]. Further animal studies should focus on rat knockout to reveal the full mechanism of the breakdown of immune tolerance related to this monogenic autoimmune disease and lead to the implementation of therapeutic approaches for patients [126].
In conclusion, studies in patients and corresponding mouse models have provided valuable insights into understanding thymic immune tolerance and the factors that contribute to autoimmunity. Nevertheless, the autoantibody fndings in human disease suggest that important gaps remain in the prevailing framework, which needs to be solved and might need focused human studies or new animal models with higher relevance to human disease.
Acknowledgments The chapter is dedicated to the late Professors Kai Krohn and Nobuyoshi Shimizu. The author thanks all the collaborations throughout the studies and apologizes for not acknowledging or citing signifcant contributions by many authors who have contributed to the feld due to the space limits.
References
1. Spinner MW, Blizzard RM, Childs B. Clinical and genetic heterogeneity in idiopathic Addison’s disease and hypoparathyroidism. J Clin Endocrinol Metab. 1968;28(6):795–804.
2. Blizzard RM, Kyle M. Studies of the adrenal antigens and antibodies in Addison’s disease. J Clin Invest. 1963;42:1653–60.
3. Neufeld M, Maclaren NK, Blizzard RM. Two types of autoimmune Addison’s disease associated with different polyglandular autoimmune (PGA) syndromes. Medicine (Baltimore). 1981;60(5):355–62.
4. Heinonen E, Krohn K, Perheentupa J, Aro A, Pelkonen R. Association of precipitating antiadrenal anti-adrenal antibodies with moniliasispolyendocrinopathy syndrome. Ann Clin Res. 1976;8(4):262–5.
5. Uibo R, Aavik E, Peterson P, Perheentupa J, Aranko S, Pelkonen R, et al. Autoantibodies to cytochrome P450 enzymes P450scc, P450c17, and P450c21 in autoimmune polyglandular disease types I and II and in isolated Addison’s disease. J Clin Endocrinol Metab. 1994;78(2):323–8.
6. Krohn K, Uibo R, Aavik E, Peterson P, Savilahti K. Identifcation by molecular cloning of an autoantigen associated with Addison’s disease as steroid 17 alpha-hydroxylase. Lancet. 1992;339(8796):770–3.
7. Peterson P, Uibo R, Peranen J, Krohn K. Immunoprecipitation of steroidogenic enzyme autoantigens with autoimmune polyglandular syndrome type I (APS I) sera; further evidence for independent humoral immunity to P450c17 and P450c21. Clin Exp Immunol. 1997;107(2):335–40.
8. Aaltonen J, Bjorses P, Sandkuijl L, Perheentupa J, Peltonen L. An autosomal locus causing autoimmune disease: autoimmune polyglandular disease typeIassigned to chromosome 21. Nat Genet. 1994;8(1):83–7.
9. Scott HS, Kyriakou DS, Peterson P, Heino M, Tahtinen M, Krohn K, et al. Characterization of a novel gene, C21orf2, on human chromosome 21q22.3 and its exclusion as the APECED gene by mutation analysis. Genomics. 1998;47(1):64–70.
10. Kudoh J, Nagamine K, Asakawa S, Abe I, Kawasaki K, Maeda H, et al. Localization of 16 exons to a 450kb region involved in the autoimmune polyglandular disease type I (APECED) on human chromosome 21q22.3. DNA Res. 1997;4(1):45–52.
11. Heino M, Peterson P, Kudoh J, Nagamine K, Lagerstedt A, Ovod V, et al. Autoimmune regulator is expressed in the cells regulating immune tolerance in thymus medulla. Biochem Biophys Res Commun. 1999;257(3):821–5.
12. Hanahan D. Peripheral-antigen-expressing cells in thymic medulla: factors in self-tolerance and autoimmunity. Curr Opin Immunol. 1998;10(6):656–62.
13. Nagamine K, Peterson P, Scott HS, Kudoh J, Minoshima S, Heino M, et al. Positional cloning of the APECED gene. Nat Genet. 1997;17(4):393–8.
14. Consortium F-GA. An autoimmune disease, APECED, caused by mutations in a novel gene featuring two PHD-type zinc-fnger domains. Nat Genet. 1997;17(4):399–403.
15. Aapola U, Lyle R, Krohn K, Antonarakis SE, Peterson P. Isolation and initial characterization of the mouse Dnmt3l gene. Cytogenet Cell Genet. 2001;92(1–2):122–6.
16. Aapola U, Kawasaki K, Scott HS, Ollila J, Vihinen M, Heino M, et al. Isolation and initial characterization of a novel zinc fnger gene, DNMT3L, on 21q22.3, related to the cytosine-5-methyltransferase 3 gene family. Genomics. 2000;65(3):293–8.
17. Rosatelli M, Meloni A, Devoto M, Cao A, Scott H, Peterson P, et al. A common mutation in Sardinian autoimmune polyendocrinopathy-candidiasis-ectodermal dystrophy patients. Hum Genet. 1998;103(4):428–34.
18. Heino M, Scott HS, Chen Q, Peterson P, Maebpaa U, Papasavvas MP, et al. Mutation analyses of North American APS-1 patients. Hum Mutat. 1999;13(1):69–74.
19. Scott HS, Heino M, Peterson P, Mittaz L, Lalioti MD, Betterle C, et al. Common mutations in autoimmune polyendocrinopathy-candidiasis-ectodermal dystrophy patients of different origins. Mol Endocrinol. 1998;12(8):1112–9.
20. Klein L, Kyewski B. “Promiscuous” expression of tissue antigens in the thymus: a key to T-cell tolerance and autoimmunity? J Mol Med. 2000;78(9):483–94.
21. Klein L, Klugmann M, Nave KA, Tuohy VK, Kyewski B. Shaping of the autoreactive T-cell repertoire by a splice variant of self protein expressed in thymic epithelial cells. Nat Med. 2000;6(1):56–61.
22. Anderson MS, Venanzi ES, Klein L, Chen Z, Berzins SP, Turley SJ, et al. Projection of an immunological self shadow within the thymus by the aire protein. Science. 2002;298(5597):1395–401.
23. Saltis M, Criscitiello MF, Ohta Y, Keefe M, Trede NS, Goitsuka R, et al. Evolutionarily conserved and divergent regions of the autoimmune regulator (Aire) gene: a comparative analysis. Immunogenetics. 2008;60(2):105–14.
24. Miller RD. Those other mammals: the immunoglobulins and T cell receptors of marsupials and monotremes. Semin Immunol. 2010;22(1):3–9.
25. Akiyoshi H, Hatakeyama S, Pitkanen J, Mouri Y, Doucas V, Kudoh J, et al. Subcellular expression of autoimmune regulator is organized in a spatiotemporal manner. J Biol Chem. 2004;279(32):33984–91.
26. Pitkanen J, Vahamurto P, Krohn K, Peterson P. Subcellular localization of the autoimmune regulator protein. Characterization of nuclear targeting and transcriptional activation domain. J Biol Chem. 2001;276(22):19597–602.
27. Pitkanen J, Doucas V, Sternsdorf T, Nakajima T, Aratani S, Jensen K, et al. The autoimmune regulator protein has transcriptional transactivating properties and interacts with the common coactivator CREB-binding protein. J Biol Chem. 2000;275(22):16802–9.
28. Gibson TJ, Ramu C, Gemund C, Aasland R. The APECED polyglandular autoimmune syndrome protein, AIRE-1, contains the SAND domain and is probably a transcription factor. Trends Biochem Sci. 1998;23(7):242–4.
29. Saare M, Hämarik U, Venta R, Panarina M, Zucchelli C, Pihlap M, et al. SP140L, an evolutionarily recent member of the SP100 family, is an autoantigen in primary biliary cirrhosis. J Immunol Res. 2015;2015:526518.
30. Bloch DB, Nakajima A, Gulick T, Chiche JD, Orth D, de La Monte SM, et al. Sp110 localizes to the PML-Sp100 nuclear body and may function as a nuclear hormone receptor transcriptional coactivator. Mol Cell Biol. 2000;20(16):6138–46.
31. Roscioli T, Cliffe ST, Bloch DB, Bell CG, Mullan G, Taylor PJ, et al. Mutations in the gene encoding
P. Peterson
the PML nuclear body protein Sp110 are associated with immunodefciency and hepatic veno-occlusive disease. Nat Genet. 2006;38(6):620–2.
32. Bottomley MJ, Stier G, Pennacchini D, Legube G, Simon B, Akhtar A, et al. NMR structure of the frst PHD fnger of autoimmune regulator protein (AIRE1). Insights into autoimmune polyendocrinopathy-candidiasis-ectodermal dystrophy (APECED) disease. J Biol Chem. 2005;280(12):11505–12.
33. Chignola F, Gaetani M, Rebane A, Org T, Mollica L, Zucchelli C, et al. The solution structure of the frst PHD fnger of autoimmune regulator in complex with non-modifed histone H3 tail reveals the antagonistic role of H3R2 methylation. Nucleic Acids Res. 2009;37(9):2951–61.
34. Org T, Rebane A, Kisand K, Laan M, Haljasorg U, Andreson R, et al. AIRE activated tissue specifc genes have histone modifcations associated with inactive chromatin. Hum Mol Genet. 2009;18(24):4699–710.
35. Org T, Chignola F, Hetenyi C, Gaetani M, Rebane A, Liiv I, et al. The autoimmune regulator PHD fnger binds to non-methylated histone H3K4 to activate gene expression. EMBO Rep. 2008;9(4):370–6.
36. Koh AS, Kingston RE, Benoist C, Mathis D. Global relevance of Aire binding to hypomethylated lysine-4 of histone-3. Proc Natl Acad Sci U S A. 2010;107(29):13016–21.
37. Koh AS, Kuo AJ, Park SY, Cheung P, Abramson J, Bua D, et al. Aire employs a histone-binding module to mediate immunological tolerance, linking chromatin regulation with organ-specifc autoimmunity. Proc Natl Acad Sci U S A. 2008;105(41):15878–83.
38. Gaetani M, Matafora V, Saare M, Spiliotopoulos D, Mollica L, Quilici G, et al. AIRE-PHD fngers are structural hubs to maintain the integrity of chromatin-associated interactome. Nucleic Acids Res. 2012;40(22):11756–68.
39. Giraud M, Yoshida H, Abramson J, Rahl PB, Young RA, Mathis D, et al. Aire unleashes stalled RNA polymerase to induce ectopic gene expression in thymic epithelial cells. Proc Natl Acad Sci U S A. 2012;109(2):535–40.
40. Oven I, Brdicková N, Kohoutek J, Vaupotic T, Narat M, Peterlin BM. AIRE recruits P-TEFb for transcriptional elongation of target genes in medullary thymic epithelial cells. Mol Cell Biol. 2007;27(24):8815–23.
41. Yoshida H, Bansal K, Schaefer U, Chapman T, Rioja I, Proekt I, et al. Brd4 bridges the transcriptional regulators, Aire and P-TEFb, to promote elongation of peripheral-tissue antigen transcripts in thymic stromal cells. Proc Natl Acad Sci U S A. 2015;112(32):E4448–57.
42. Abramson J, Giraud M, Benoist C, Mathis D. Aire’s partners in the molecular control of immunological tolerance. Cell. 2010;140(1):123–35.
43. Liiv I, Rebane A, Org T, Saare M, Maslovskaja J, Kisand K, et al. DNA-PK contributes to the phosphorylation of AIRE: importance in transcriptional activity. Biochim Biophys Acta. 2008;1783(1):74–83.
44. Guha M, Saare M, Maslovskaja J, Kisand K, Liiv I, Haljasorg U, et al. DNA breaks and chromatin structural changes enhance the transcription of autoimmune regulator target genes. J Biol Chem. 2017;292(16):6542–54.
45. Zumer K, Low AK, Jiang H, Saksela K, Peterlin BM. Unmodifed histone H3K4 and DNA-dependent protein kinase recruit autoimmune regulator to target genes. Mol Cell Biol. 2012;32(8):1354–62.
46. Maslovskaja J, Rebane A, Peterson P. AIREregulated gene expression mainly occurs on transcriptional level and is promoter-independent. Scand J Immunol. 2010;71(6):488.
47. Meredith M, Zemmour D, Mathis D, Benoist C. Aire controls gene expression in the thymic epithelium with ordered stochasticity. Nat Immunol. 2015;16(9):942–9.
48. Bansal K, Yoshida H, Benoist C, Mathis D. The transcriptional regulator Aire binds to and activates super-enhancers. Nat Immunol. 2017;18(3):263–73.
49. Mathis D, Benoist C. Aire. Annu Rev Immunol. 2009;27:287–312.
50. Peterson P, Org T, Rebane A. Transcriptional regulation by AIRE: molecular mechanisms of central tolerance. Nat Rev Immunol. 2008;8(12):948–57.
51. Akiyama T, Shimo Y, Yanai H, Qin J, Ohshima D, Maruyama Y, et al. The tumor necrosis factor family receptors RANK and CD40 cooperatively establish the thymic medullary microenvironment and selftolerance. Immunity. 2008;29(3):423–37.
52. Rossi SW, Kim MY, Leibbrandt A, Parnell SM, Jenkinson WE, Glanville SH, et al. RANK signals from CD4(+)3(−) inducer cells regulate development of Aire-expressing epithelial cells in the thymic medulla. J Exp Med. 2007;204(6):1267–72.
53. Irla M, Hugues S, Gill J, Nitta T, Hikosaka Y, Williams IR, et al. Autoantigen-specifc interactions with CD4+ thymocytes control mature medullary thymic epithelial cell cellularity. Immunity. 2008;29(3):451–63.
54. Haljasorg U, Bichele R, Saare M, Guha M, Maslovskaja J, Kõnd K, et al. A highly conserved NF-κB-responsive enhancer is critical for thymic expression of Aire in mice. Eur J Immunol. 2015;45(12):3246–56.
55. LaFlam TN, Seumois G, Miller CN, Lwin W, Fasano KJ, Waterfeld M, et al. Identifcation of a novel cisregulatory element essential for immune tolerance. J Exp Med. 2015;212(12):1993–2002.
56. Murumagi A, Vahamurto P, Peterson P. Characterization of regulatory elements and methylation pattern of the autoimmune regulator (AIRE) promoter. J Biol Chem. 2003;278(22):19784–90.
57. Herzig Y, Nevo S, Bornstein C, Brezis MR, Ben-Hur S, Shkedy A, et al. Transcriptional programs that control expression of the autoimmune regulator gene Aire. Nat Immunol. 2017;18(2):161–72.
58. Murumägi A, Silvennoinen O, Peterson P. Ets transcription factors regulate AIRE gene pro-
moter. Biochem Biophys Res Commun. 2006;348(2):768–74.
59. Gardner JM, Metzger TC, McMahon EJ, Au-Yeung BB, Krawisz AK, Lu W, et al. Extrathymic Aireexpressing cells are a distinct bone marrow-derived population that induce functional inactivation of CD4+ T cells. Immunity. 2013;39(3):560–72.
60. Poliani PL, Kisand K, Marrella V, Ravanini M, Notarangelo LD, Villa A, et al. Human peripheral lymphoid tissues contain autoimmune regulator-expressing dendritic cells. Am J Pathol. 2010;176(3):1104–12.
61. Gardner JM, Devoss JJ, Friedman RS, Wong DJ, Tan YX, Zhou X, et al. Deletional tolerance mediated by extrathymic Aire-expressing cells. Science. 2008;321(5890):843–7.
62. Anderson MS, Venanzi ES, Chen Z, Berzins SP, Benoist C, Mathis D. The cellular mechanism of Aire control of T cell tolerance. Immunity. 2005;23(2):227–39.
63. Liston A, Lesage S, Wilson J, Peltonen L, Goodnow CC. Aire regulates negative selection of organ-specifc T cells. Nat Immunol. 2003;4(4):350–4.
64. Yang S, Fujikado N, Kolodin D, Benoist C, Mathis D. Immune tolerance. Regulatory T cells generated early in life play a distinct role in maintaining selftolerance. Science. 2015;348(6234):589–94.
65. Malchow S, Leventhal DS, Lee V, Nishi S, Socci ND, Savage PA. Aire enforces Immune tolerance by directing autoreactive T cells into the regulatory T cell lineage. Immunity. 2016;44(5):1102–13.
66. Malchow S, Leventhal DS, Nishi S, Fischer BI, Shen L, Paner GP, et al. Aire-dependent thymic development of tumor-associated regulatory T cells. Science. 2013;339(6124):1219–24.
67. Laakso SM, Laurinolli TT, Rossi LH, Lehtoviita A, Sairanen H, Perheentupa J, et al. Regulatory T cell defect in APECED patients is associated with loss of naive FOXP3(+) precursors and impaired activated population. J Autoimmun. 2010;35(4):351–7.
68. Kekäläinen E, Tuovinen H, Joensuu J, Gylling M, Franssila R, Pöntynen N, et al. A defect of regulatory T cells in patients with autoimmune polyendocrinopathy-candidiasis-ectodermal dystrophy. J Immunol. 2007;178(2):1208–15.
69. Ryan KR, Lawson CA, Lorenzi AR, Arkwright PD, Isaacs JD, Lilic D. CD4+CD25+ T-regulatory cells are decreased in patients with autoimmune polyendocrinopathy candidiasis ectodermal dystrophy. J Allergy Clin Immunol. 2005;116(5):1158–9.
70. Wolff AS, Oftedal BE, Kisand K, Ersvaer E, Lima K, Husebye ES. Flow cytometry study of blood cell subtypes refects autoimmune and infammatory processes in autoimmune polyendocrine syndrome type I. Scand J Immunol. 2010;71(6):459–67.
71. Sng J, Ayoglu B, Chen JW, Schickel JN, Ferre EMN, Glauzy S, et al. AIRE expression controls the peripheral selection of autoreactive B cells. Sci Immunol. 2019;4(34):eaav6778.
72. Sansom SN, Shikama-Dorn N, Zhanybekova S, Nusspaumer G, Macaulay IC, Deadman ME, et al. Population and single-cell genomics reveal the Aire dependency, relief from polycomb silencing, and distribution of self-antigen expression in thymic epithelia. Genome Res. 2014;24(12):1918–31.
73. Brennecke P, Reyes A, Pinto S, Rattay K, Nguyen M, Küchler R, et al. Single-cell transcriptome analysis reveals coordinated ectopic gene-expression patterns in medullary thymic epithelial cells. Nat Immunol. 2015;16(9):933–41.
74. Michelson DA, Hase K, Kaisho T, Benoist C, Mathis D. Thymic epithelial cells co-opt lineage-defning transcription factors to eliminate autoreactive T cells. Cell. 2022;185(14):2542–58.e18.
75. Farr AG, Dooley JL, Erickson M. Organization of thymic medullary epithelial heterogeneity: implications for mechanisms of epithelial differentiation. Immunol Rev. 2002;189:20–7.
76. Derbinski J, Gabler J, Brors B, Tierling S, Jonnakuty S, Hergenhahn M, et al. Promiscuous gene expression in thymic epithelial cells is regulated at multiple levels. J Exp Med. 2005;202(1):33–45.
77. Nishikawa Y, Nishijima H, Matsumoto M, Morimoto J, Hirota F, Takahashi S, et al. Temporal lineage tracing of Aire-expressing cells reveals a requirement for Aire in their maturation program. J Immunol. 2014;192(6):2585–92.
78. Yano M, Kuroda N, Han H, Meguro-Horike M, Nishikawa Y, Kiyonari H, et al. Aire controls the differentiation program of thymic epithelial cells in the medulla for the establishment of self-tolerance. J Exp Med. 2008;205(12):2827–38.
79. Matsumoto M. Contrasting models for the roles of Aire in the differentiation program of epithelial cells in the thymic medulla. Eur J Immunol. 2011;41(1):12–7.
80. Kisand K, Peterson P. Autoimmune polyendocrinopathy candidiasis ectodermal dystrophy. J Clin Immunol. 2015;35(5):463–78.
81. Husebye ES, Anderson MS, Kämpe O. Autoimmune polyendocrine syndromes. N Engl J Med. 2018;378(26):2543–4.
82. Ricotta EE, Ferré EMN, Schmitt MM, DiMaggio T, Lionakis MS. Prevalence of APECED-like clinical disease in an electronic health record database, USA. J Clin Immunol. 2022;42(4):904–6.
83. Ferre EM, Rose SR, Rosenzweig SD, Burbelo PD, Romito KR, Niemela JE, et al. Redefned clinical features and diagnostic criteria in autoimmune polyendocrinopathy-candidiasis-ectodermal dystrophy. JCI Insight. 2016;1(13):e88782.
84. Perheentupa J. Autoimmune polyendocrinopathycandidiasis-ectodermal dystrophy. J Clin Endocrinol Metab. 2006;91(8):2843–50.
85. Perheentupa J. APS-I/APECED: the clinical disease and therapy. Endocrinol Metab Clin N Am. 2002;31(2):295–320. vi
86. Bjorses P, Halonen M, Palvimo JJ, Kolmer M, Aaltonen J, Ellonen P, et al. Mutations in the AIRE gene: effects on subcellular location and transactivation function of the autoimmune polyendocrinopathy-candidiasis-ectodermal dystrophy protein. Am J Hum Genet. 2000;66(2):378–92.
87. Pearce SH, Cheetham T, Imrie H, Vaidya B, Barnes ND, Bilous RW, et al. A common and recurrent 13-bp deletion in the autoimmune regulator gene in British kindreds with autoimmune polyendocrinopathy type 1. Am J Hum Genet. 1998;63(6):1675–84.
88. Giordano C, Modica R, Allotta ML, Guarnotta V, Cervato S, Masiero S, et al. Autoimmune polyendocrinopathy-candidiasis-ectodermal-dystrophy (APECED) in Sicily: confrmation that R203X is the peculiar AIRE gene mutation. J Endocrinol Investig. 2012;35(4):384–8.
89. Cetani F, Barbesino G, Borsari S, Pardi E, Cianferotti L, Pinchera A, et al. A novel mutation of the autoimmune regulator gene in an Italian kindred with autoimmune polyendocrinopathy-candidiasisectodermal dystrophy, acting in a dominant fashion and strongly cosegregating with hypothyroid autoimmune thyroiditis. J Clin Endocrinol Metab. 2001;86(10):4747–52.
90. Su MA, Giang K, Zumer K, Jiang H, Oven I, Rinn JL, et al. Mechanisms of an autoimmunity syndrome in mice caused by a dominant mutation in Aire. J Clin Invest. 2008;118(5):1712–26.
91. Oftedal BE, Hellesen A, Erichsen MM, Bratland E, Vardi A, Perheentupa J, et al. Dominant mutations in the autoimmune regulator AIRE are associated with common organ-specifc autoimmune diseases. Immunity. 2015;42(6):1185–96.
92. Eriksson D, Bianchi M, Landegren N, Dalin F, Skov J, Hultin-Rosenberg L, et al. Common genetic variation in the autoimmune regulator (AIRE) locus is associated with autoimmune Addison’s disease in Sweden. Sci Rep. 2018;8(1):8395.
93. Laisk T, Lepamets M, Koel M, Abner E, Mägi R, Team EBR. Genome-wide association study identifes fve risk loci for pernicious anemia. Nat Commun. 2021;12(1):3761.
94. Oftedal BE, Assing K, Baris S, Safgren SL, Johansen IS, Jakobsen MA, et al. Dominant-negative heterozygous mutations in AIRE confer diverse autoimmune phenotypes. iScience. 2023;26(6):106818.
95. Bjorses P, Aaltonen J, Vikman A, Perheentupa J, Ben-Zion G, Chiumello G, et al. Genetic homogeneity of autoimmune polyglandular disease type I. Am J Hum Genet. 1996;59(4):879–86.
96. Halonen M, Eskelin P, Myhre AG, Perheentupa J, Husebye ES, Kämpe O, et al. AIRE mutations and human leukocyte antigen genotypes as determinants of the autoimmune polyendocrinopathy-candidiasisectodermal dystrophy phenotype. J Clin Endocrinol Metab. 2002;87(6):2568–74.
97. Kisand K, Lilic D, Casanova J-L, Peterson P, Meager A, Willcox N. Mucocutaneous candidiasis and
P. Peterson
autoimmunity against cytokines in APECED and thymoma patients: clinical and pathogenetic implications. Eur J Immunol. 2011;41(6):1517–27.
98. Ferré EMN, Lionakis MS. An AIREless breath: pneumonitis caused by impaired central immune tolerance. Front Immunol. 2020;11:609253.
99. Winqvist O, Gustafsson J, Rorsman F, Karlsson FA, Kampe O. Two different cytochrome P450 enzymes are the adrenal antigens in autoimmune polyendocrine syndrome type I and Addison’s disease. J Clin Invest. 1993;92(5):2377–85.
100. Winqvist O, Karlsson FA, Kampe O. 21-hydroxylase, a major autoantigen in idiopathic Addison’s disease. Lancet. 1992;339(8809):1559–62.
101. Bjork E, Velloso LA, Kampe O, Karlsson FA. GAD autoantibodies in IDDM, stiff-man syndrome, and autoimmune polyendocrine syndrome type I recognize different epitopes. Diabetes. 1994;43(1):161–5.
102. Perniola R, Falorni A, Clemente MG, Forini F, Accogli E, Lobreglio G. Organ-specifc and nonorgan-specifc autoantibodies in children and young adults with autoimmune polyendocrinopathy-candidiasis-ectodermal dystrophy (APECED). Eur J Endocrinol. 2000;143(4):497–503.
103. Alimohammadi M, Björklund P, Hallgren A, Pöntynen N, Szinnai G, Shikama N, et al. Autoimmune polyendocrine syndrome type 1 and NALP5, a parathyroid autoantigen. N Engl J Med. 2008;358(10):1018–28.
104. Ekwall O, Sjoberg K, Mirakian R, Rorsman F, Kampe O. Tryptophan hydroxylase autoantibodies and intestinal disease in autoimmune polyendocrine syndrome type 1. Lancet. 1999;354(9178):568.
105. Husebye ES, Gebre-Medhin G, Tuomi T, Perheentupa J, Landin-Olsson M, Gustafsson J, et al. Autoantibodies against aromatic L-amino acid decarboxylase in autoimmune polyendocrine syndrome type I. J Clin Endocrinol Metab. 1997;82(1):147–50.
106. Meyer S, Woodward M, Hertel C, Vlaicu P, Haque Y, Kärner J, et al. AIRE-defcient patients harbor unique high-affnity disease-ameliorating autoantibodies. Cell. 2016;166:582.
107. Fishman D, Kisand K, Hertel C, Rothe M, Remm A, Pihlap M, et al. Autoantibody repertoire in APECED patients targets two distinct subgroups of proteins. Front Immunol. 2017;8:976.
108. Wolff ASB, Sarkadi AK, Marodi L, Kaerner J, Orlova E, Oftedal BEV, et al. Anti-cytokine autoantibodies preceding onset of autoimmune polyendocrine syndrome type I features in early childhood. J Clin Immunol. 2013;33(8):1341–8.
109. Meager A, Visvalingam K, Peterson P, Moll K, Murumagi A, Krohn K, et al. Anti-interferon autoantibodies in autoimmune polyendocrinopathy syndrome type 1. PLoS Med. 2006;3(7):e289.
110. Wolff A, Kisand K, Meager A, Willcox N, Peterson P, Husebye E. Cytokine antibodies in autoimmune polyendocrine syndrome type I correlates to chronic candidiasis. Scand J Immunol. 2011;73(4):379.
111. Meager A, Peterson P, Willcox N. Hypothetical review: thymic aberrations and type-I interferons; attempts to deduce autoimmunizing mechanisms from unexpected clues in monogenic and paraneoplastic syndromes. Clin Exp Immunol. 2008;154(1):141–51.
112. Kisand K, Link M, Wolff AS, Meager A, Tserel L, Org T, et al. Interferon autoantibodies associated with AIRE defciency decrease the expression of IFN-stimulated genes. Blood. 2008;112(7):2657–66.
113. Hetemäki I, Laakso S, Välimaa H, Kleino I, Kekäläinen E, Mäkitie O, et al. Patients with autoimmune polyendocrine syndrome type 1 have an increased susceptibility to severe herpesvirus infections. Clin Immunol. 2021;231:108851.
114. Bastard P, Orlova E, Sozaeva L, Lévy R, James A, Schmitt MM, et al. Preexisting autoantibodies to type I IFNs underlie critical COVID-19 pneumonia in patients with APS-1. J Exp Med. 2021;218(7):e20210554.
115. Kisand K, Bøe Wolff AS, Podkrajsek KT, Tserel L, Link M, Kisand KV, et al. Chronic mucocutaneous candidiasis in APECED or thymoma patients correlates with autoimmunity to Th17-associated cytokines. J Exp Med. 2010;207(2):299–308.
116. Puel A, Doffnger R, Natividad A, Chrabieh M, Barcenas-Morales G, Picard C, et al. Autoantibodies against IL-17A, IL-17F, and IL-22 in patients with chronic mucocutaneous candidiasis and autoimmune polyendocrine syndrome type I. J Exp Med. 2010;207(2):291–7.
117. Kaleviste E, Rühlemann M, Kärner J, Haljasmägi L, Tserel L, Org E, et al. IL-22 paucity in APECED is associated with mucosal and microbial alterations in oral cavity. Front Immunol. 2020;11:838.
118. Karner J, Pihlap M, Ranki A, Krohn K, Podkrajsek KT, Bratanic N, et al. IL-6-specifc autoantibodies among APECED and thymoma patients. Immun Infamm Dis. 2016;4(2):235–43.
119. Walter JE, Rosen LB, Csomos K, Rosenberg JM, Mathew D, Keszei M, et al. Broad-spectrum antibodies against self-antigens and cytokines in RAG defciency. J Clin Invest. 2015;125(11):4135–48.
120. Rosenberg JM, Maccari ME, Barzaghi F, Allenspach EJ, Pignata C, Weber G, et al. Neutralizing anticytokine autoantibodies against interferon-α in immunodysregulation polyendocrinopathy enteropathy X-linked. Front Immunol. 2018;9:544.
121. Bradford HF, Haljasmägi L, Menon M, McDonnell TCR, Särekannu K, Vanker M, et al. Inactive disease in patients with lupus is linked to autoantibodies to type I interferons that normalize blood IFNα and B cell subsets. Cell Rep Med. 2023;4(1):100894.
122. Manry J, Bastard P, Gervais A, Le Voyer T, Rosain J, Philippot Q, et al. The risk of COVID-19 death is much greater and age dependent with type I IFN autoantibodies. Proc Natl Acad Sci U S A. 2022;119(21):e2200413119.
123. Bastard P, Rosen LB, Zhang Q, Michailidis E, Hoffmann HH, Zhang Y, et al. Autoantibodies against type I IFNs in patients with life-threatening COVID-19. Science. 2020;370(6515):eabd4585.
124. Kekäläinen E, Miettinen A, Arstila TP. Does the defciency of Aire in mice really resemble human APECED? Nat Rev Immunol. 2007;7(10):1.
125. Ossart J, Moreau A, Autrusseau E, Ménoret S, Martin JC, Besnard M, et al. Breakdown of immune tolerance in AIRE-defcient rats induces a severe autoim-
P. Peterson
mune polyendocrinopathy-candidiasis-ectodermal dystrophy-like autoimmune disease. J Immunol. 2018;201(3):874–87.
126. Besnard M, Sérazin C, Ossart J, Moreau A, Vimond N, Flippe L, et al. Anti-CD45RC antibody immunotherapy prevents and treats experimental autoimmune polyendocrinopathy-candidiasis-ectodermal dystrophy syndrome. J Clin Invest. 2022;132(7):e156507.
Learning the Autoimmune Pathogenesis Through the Study of Aire
Mitsuru Matsumoto and Minoru Matsumoto
Abstract
One of the diffculties in studying the pathogenesis of autoimmune diseases is that the disease is multifactorial involving sex, age, MHC, environment, and some genetic factors. Because defciency of Aire, a transcriptional regulator, is an autoimmune disease caused by a single gene abnormality, Aire is an ideal research target for approaching the enigma of autoimmunity, e.g., the mechanisms underlying Aire defciency can be studied using genetically modifed animals. Nevertheless, the exact mechanisms of the breakdown of self-tolerance due to Aire’s dysfunction have not yet been fully clarifed. This is due, at least in part, to the lack of information on the exact target genes controlled by Aire. State-of-theart research infrastructures such as single-cell analysis are now in place to elucidate the essential function of Aire. The knowledge gained through the study of Aire-mediated tolerance should help our understanding of the pathogenesis of autoimmune disease in general.
M. Matsumoto (*) Tokushima University, Tokushima, Japan e-mail: mitsuru@tokushima-u.ac.jp
M. Matsumoto
Department of Molecular Pathology, Tokushima University Graduate School of Biomedical Sciences, Tokushima, Japan
Keywords
Autoimmune disease · Aire · mTEC ·
Self-tolerance · Animal model · Single-cell analysis
2.1 Introduction
Recent biologics, mainly anti-cytokine monoclonal antibodies and inhibitors of the immune receptors, have had an extraordinary impact on the treatment of autoimmune diseases. It has been demonstrated clinically that the excessive production of TNF [1] and/or IL-6 [2] due to the breakdown of central and/or peripheral tolerance plays an important role in the pathogenesis of various autoimmune diseases. However, it is still diffcult to say that the exact mechanism of the breakdown of tolerance has been fully elucidated. This fact, along with the fact that anti-cytokine therapy is not effective for all autoimmune diseases, constitutes a major hurdle in developing a more fundamental and innovative therapy to improve or replace the current biologics. A possible breakthrough in solving this diffcult situation was brought about by the identifcation of AIRE, a gene responsible for autoimmune diseases that exhibits Mendelian inheritance (autosomal recessive) [3]. Positional cloning identifed the transcriptional regulator AIRE (autoimmune regulator) as a causative gene for autoimmune
© The Author(s), under exclusive license to Springer Nature Singapore Pte Ltd. 2024 M. Matsumoto (ed.), Basic Immunology and Its Clinical Application, Advances in Experimental Medicine and Biology 1444, https://doi.org/10.1007/978-981-99-9781-7_2
19
polyendocrinopathy-candidiasis-ectodermal dystrophy (APECED) [4, 5]. In APECED, mutations in just one gene invariably cause autoimmune diseases that target various glandular tissues [6]. Importantly, Aire expression is strongest in medullary thymic epithelial cells (mTECs), which serve to eliminate autoreactive T cells by presenting self-antigens (self-Ags) [7, 8]. Therefore, elucidation of disease pathology caused by Aire dysfunction should directly reveal the actual state of immunological tolerance. This may lead to the elucidation of the disease mechanisms of not only APECED but also other autoimmune diseases. From an experimental viewpoint, it is possible to create and use disease animal models produced by exactly the same pathological mechanisms as that of humans by modifying the Aire gene in rodents. Indeed, Airedefcient rodents developed autoimmune pathologies and have become an extremely useful disease model for studying the pathogenesis of APECED [9, 10]. In this way, Aire has become an ideal research target for investigating the pathogenesis of autoimmune diseases. Thus, the discovery of Aire has made true experimental medicine possible even for human autoimmune diseases.
2.2 Animal Models of Aire Defciency
Several groups have generated Aire-defcient mice independently and reported their phenotypes in detail. Although there are some minor phenotypic differences among the reports, all the Aire-defcient mice developed signs of autoimmunity including sialadenitis, hepatitis, and epididymitis in males [11–14]. Rather unexpectedly, Aire-defcient mice showed no major symptoms of APECED in humans, i.e., hypoparathyroidism, hypoadrenocorticism, and ectodermal dystrophy. Furthermore, they did not produce autoantibodies against the steroid genic enzymes (e.g., steroidogenic P450 cytochromes) [15] and cytokines (e.g., type I IFNs and IL-17) [16, 17], which are often seen in the sera from human patients. However, the core of the Aire-defcient
M. Matsumoto and M. Matsumoto
thymus-mediated autoimmunity was confrmed by the thymus graft experiments in mice [12, 13]. Interestingly, the phenotypes varied depending on the genetic background. Although Airedefcient mice on C57BL/6 showed mild autoimmune pathologies predominating in the sialadenitis, Aire-defcient mice on Balb/c specifcally showed autoimmune gastritis associated with autoantibodies against the gastric mucosa [13]. On NOD background, Aire-defcient mice developed lethal pneumonitis and severe destruction of pancreatic acinar cells [18, 19]. Irrespective of the genetic background, the most remarkable fnding was the reduced expression of many selfAgs (tissue-restricted Ags: TRAs) from mTECs that may account for the defective deletion of autoreactive T cells and/or the production of regulatory T cells (Tregs) [20]. However, although Aire-defcient mTECs on NOD showed dramatically reduced expression of insulin, they never developed insulitis and diabetes [18]. This discrepancy (i.e., reduced expression of insulin in mTECs without the development of insulitis) raised the question of the essential function of Aire in the control of self-Ags in mTECs (see below).
Aire-defcient rats were also generated using a zinc fnger nuclease technology, and they showed signs of ectodermal dystrophy (e.g., nail dystrophy), which was not observed in mice [21]. Interestingly, a monoclonal antibody against CD45RC, a molecule expressed at high levels by conventional T cells (CD45RChigh), their precursors, and terminally differentiated T cells but not by Tregs (CD45RClow/ ), was effective for both prevention and treatment of autoimmune manifestations in Aire-defcient rats [22].
2.3 Molecular Biology of Aire
Because Aire is a transcriptional regulator, one important approach to knowing the exact function of Aire in immune tolerance would be to identify the target genes controlled by Aire. However, identifcation of Aire’s targets by the conventional biochemical approaches is not so easy due to the small amount of Aire protein in
2.1
of
primary mTECs and its association with the nuclear matrix [23, 24]. Accordingly, many functional analyses of each Aire’s domains such as two PHD domains together with a homogeneously staining region (HSR) and SAND domain were performed using in vitro transfection experiments (Fig. 2.1) [25]. Similarly, many studies identifying Aire’s targets and/or Aire’s partner proteins have been undertaken using the Aire-transfected cell lines [26]. Among them, Aire has been demonstrated to have a unique capacity to activate genes that are marked with chromatin of H3K4me0 in their promoters through the binding of the Aire-PHD1 domain [27, 28]. Genes marked with H3K4me0 chromatin are usually silent or expressed only at low levels. Indeed, it was demonstrated that Aire preferentially activates genes that are expressed at low levels and the genes that lacked H3K4me3 modifcations in their promoter regions [27]. Thus, Aire may be a histone-tail reader module that can sense the methylation status of histone H3 and augment the target gene expression in cooperation with many other transcription factors. There is another report suggesting that Aire is recruited to the repressive sites formed by the ATF7ip-MBD1 complex [29]. According to this study, Aire is considered to recognize the specifc methylated CpG dinucleotides provided by MBD1 to target the loci of TRAs, although how Aire induces the subsequent induction of the targets has not been addressed.
From the cytological characteristics, Aire forms the nuclear bodies (NBs) in the mTECs [23]. However, it remains to be determined whether
Aire NBs themselves are the major site of transcriptional regulation by Aire. Furthermore, it is not yet clear which form of Aire, its nucleoplasmic form or NB form, is mainly responsible for the physiological function of Aire. It is equally possible that Aire in NBs is functionally distinct from that existing homogenously in the nucleoplasm. Ideally, these questions need to be answered using the primary mTECs but not Aire-transfected cells, as discussed above. Technologies that overcome these biochemically tough conditions yet await to be developed.
2.4 Aire Controls the Diferentiation Program of mTECs
Besides the control of TRA expression from mTECs, we have argued that Aire plays an important role in the differentiation and maturation of mTECs [30]. Using a GFP-knockin mouse strain in which the GFP gene was introduced into the Aire locus in a manner allowing concomitant disruption of the Aire gene, we have demonstrated that Aireless GFP+ mTECs (homozygous for the Aire/GFP knockin allele) showed more globular cell shapes compared with Aire-suffcient mTECs [31]. Furthermore, Aire-defcient thymi showed much reduced numbers of Hassall’s corpusclelike structures in the medulla. Particularly, the Aire-dependent alteration of the shape of Aire+ mTECs suggested the cell-intrinsic role of Aire in the differentiation program of mTECs. We also found that Aire-expressing mTECs trans-regulate
Fig.
Structural motif
human AIRE
the production of other mTEC subsets, such as Ly6+ mTECs [32]. In the absence of Aire+ mTECs, Ly6+ mTECs were not produced. Thus, Aire controls the differentiation program of mTECs both cell-intrinsically and cellextrinsically, which together may contribute to the transcriptome of mTECs.
In elucidating the function of Aire, the activity of Aire in controlling the development of mTECs makes it complicated to identify the bona fde Aire’s targets. Namely, there are two modes of Aire’s action in controlling the transcriptome of mTECs. Aire controls the transcription of the targets directly within a cell (i.e., Aire’s direct effect on TRA expression). Aire also controls the transcriptome by controlling the development and/or composition of mTEC subpopulations (i.e., Aire’s indirect effect on TRAs). Because both gene control can contribute to the alteration of the overall transcriptome of mTECs, and ultimately contribute to the development of Aire-mediated autoimmunity, we frst need to distinguish the two groups of Aire’s targets (Aire’s direct targets vs. Aire’s indirect targets) to illustrate the clear picture of the Aire function.
2.5 The Tolerogenic Function of Aire
One of the landmarking discoveries using Airedefcient mice was that Aire-defcient mTECs showed the reduced expression of many TRAs. However, one immediate question that needs to be answered is whether it is relevant to the production of autoreactive T cells and to the development of organ-specifc autoimmunity caused by Aire defciency. This question was further highlighted by the report demonstrating that Aire defciency caused almost complete failure to delete the organ-specifc T cells in the thymus: this was demonstrated by crossing transgenic mice expressing hen egg lysozyme (HEL) under the control of insulin promotor with the
M. Matsumoto and M. Matsumoto
HEL-specifc CD4+ T cell transgenic line [33]. Remarkably, Aire defciency resulted in the defect in the negative selection of clonotypic T cells. However, whether this was due to the reduced HEL expression at the transcriptional level from Aire-defcient mTECs was not so clear [34]. Although endogenous insulin expression from Aire-defcient mTECs was almost gone [12, 13], expression levels of HEL driven by the insulin promoter from mTECs in the HEL-transgenic/Aire-defcient mice were rather modest [34].
We independently approached this issue using the RIP-OVA Tg model in which OVA was driven by the rat insulin promoter. Although RIP-OVA/Aire-defcient double-Tg showed the defect in the negative selection of clonotypic T cells of OT-II Tg, as reported in HELtransgenic/Aire-defcient mice, OVA expression levels from mTECs were not changed by the lack of Aire [35]. Thus, the defect in the clonotypic T cells in Aire-defcient mice may not be simply due to the reduced expression of model self-Ag of OVA. Because clonal deletion of OVA-specifc T cells requires bone marrow (BM)-derived Ag-presenting cells (APCs), we suspected that Ag transfer from mTECs to BM-derived APCs might be impaired by the lack of Aire. How the defect of Aire in mTECs results in the defective Ag transfer, however, remains unanswered. In this regard, we have recently demonstrated that self-Ag can be transferred not only from mature mTECs but also from immature mTECs [36]. In contrast, selfAg expressed in cortical TECs (cTECs) were not transferred to BM-derived APCs effciently. Elucidation of the exact mechanisms underlying Aire-dependent negative selection that does not depend on the alteration of the quantity of the self-Ag may reveal the precise function of Aire in immune tolerance.
Unexpectedly, we found that defective negative selection in the TCR-transgenic system was observed not only when Aire expression was abrogated by the generation of Aire-defcient
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