Lupus Genetics

Abstract

Lupus erythematosus is an autoimmune disorder with an important genetic component. Studies in monozygotic twins have revealed a concordance rate of 50% indicating that environmental factors might play a significant role in the development of the disease. Genes that are implicated in the pathogenesis of lupus erythematosus include HLA, Interferon genes, complement genes, cytokine genes (TNF, IL-10, IL-1β, IL-17, IL-23), NF-κB genes, ITGAM gene, PPP2CA genes, SIAE genes, SLAMF molecules, PTPN22, BLK, BANK1, PD-1 and X-linked genes (AIRE gene and others). Epigenetic factors which alter only the expression but not the DNA structure may also interfere with the development of the disease.

1. Introduction

Autoimmunity occurs when a component of a certain tissue of the human body becomes immunogenic with consequent production of autoantibodies against it.

For induction of autoimmunity three conditions are required.

1. Self-antigen.

2. An inflammatory environment.

3. A genetic predisposition.

For the production of a self-antigen, two mechanisms are implicated. A part of an inflammatory agent, a virus, for instance, may have similarities with a component of human tissue and in this way it becomes immunogenic; this mechanism is called molecular mimicry – a mechanism quite commonly encountered in nature. Another mechanism is epitope spreading when fragments of damaged tissue during the inflammatory process become immunogenic. Stimuli that may induce an inflammation are multiple: microbes, viruses, chemicals, stress, etc. As all human beings are exposed to various agents very frequently, it is quite difficult to identify the initial moment autoimmunity occurs and the responsible factor [1].

2. Lupus erythematosus

Lupus is the prototype of autoimmune diseases with B-cell hyperactivity resulting in the production of anti-DNA autoantibodies and manifests with abnormalities of internal organs. Renal insufficiency, hemolytic anemia, arterial and venous thromboses are common in SLE. Skin eruptions may also be observed in patients with SLE. Females are more frequently affected with the disease female: male ratio 6-10:1. Prevalence of SLE is relatively low 1 in 2000, in general population. Despite this fact, SLE remains an important health care problem and is associated with a significant financial burden to the community because of the young age of the individuals suffering from the disease. A study of survival from 2000 to 2002 has shown that almost 4% of all patients hospitalized in New York AND Pennsylvania with lupus die [2, 3].

Lupus is a disease with a strong genetic component as it runs in families [4].

3. Genes associated with SLE

Genes associated with lupus include genes of the HLA of the MHC group of genes and non-HLA genes such as interferon genes, autophagy genes and the X-linked group genes and others. Among them, the HLA genes are those that play the most important role in the pathogenesis of lupus erythematosus. The different groups of these genes will be discussed in detail below [5].

4. HLA genes of the MHC and lupus

The human MHC genes are located in a segment of chromosome 6 (6p21.3) that consists of three areas (I, II and III) encoding three major classes of proteins: class I human leukocyte antigens, HLA divided into antigens (A, B and Cw), HLA class II (DP, DQ and DR) and class III whose components are complement, tumor necrosis factor (TNFa) and heat shock proteins. The HLA genes were the first to be associated with SLE and this since 1970. Studies in the past have identified HLA A1, B8 – a weak association in some studies with other studies showing no association at all -DR2, DR3, DQW1, DRW52, C4 null ancestral haplotypes as susceptibility genes for lupus erythematosus. HLA DR2 has been found in 75% of white patients with SLE (normal subjects 24%), 75% are positive for DR3 (normal subjects 25%), 75% are positive for DQW1 (normal subjects 55%) and 65% haveDRW52 (normal subjects 46%) [6]. Data suggest that the HLA-D region is exerting its effect on certain autoantibody responses in lupus erythematosus. Recently, the DRB1 gene polymorphisms have been associated with different sub-groups of systemic lupus erythematosus.

Different groups of SLE are defined by autoantibodies status, HLA-DRB1 polymorphisms, immunological and clinical manifestations.

Recent research has identified four different groups described below:

1. Subgroup 1 is dominated by anti-SSA/R060/Ro52/SSB autoantibodies and is strongly associated with HLA-DRB103. Discoid lesions are more common in this subgroup.

2. Subgroup 2 is dominated by anti-nucleosome/SmRNP/DNA/RNPA autoantibodies and is associated with HLA-DRB115. Nephritis is most common in this subgroup.

3. Subgroup 3 is characterized by anti-b2GPI-IgG/ANTI-cl-IgG/IgM autoantibodies and a higher frequency of HLA-DRB104 compared with other patients with SLE. Vascular events are more common in this subgroup.

4. Subgroup 4 was negative for all the above-investigated autoantibodies and was not associated with HLA-DRB1 [7].

5. HLA genes and age

Recent research has shown that HLA genes are implicated differently in the pathogenesis of lupus according to age. HLA genes are more implicated in the pathogenesis of lupus in older than in younger patients. This may be due to the fact that lupus in young patients is more related to infection while in older individuals is associated with the intake of drugs administered for various other ailments such as antihypertensive drugs, for instance, captopril [8, 9].

6. Lupus erythematosus genetics and other autoimmune diseases

An association of lupus with myasthenia gravis in a male patient has been HLA investigated and found positivity for HLA DRB1602-frequent in autoimmune disorders associated with the production of autoantibodies, DRB1401 frequent in late-onset myasthenia gravis, and A1B8 found in three of seven patients with both lupus erythematosus and myasthenia gravis [10, 11, 12].

7. Interferon genes are associated with lupus

INF—is a protein produced mainly by dendritic cells and lymphocytes following a viral infection and links innate and adaptive immunity. It confers resistance to viral infection and susceptibility to autoimmunity. Previous studies showed that interferon type I is strongly associated with the pathogenesis of lupus. The primary pathogenic factor in SLE escalates IFN-a signaling, which can activate STAT4, a transcription factor [13]. Genetic variations of this transcription factor have been associated with the risk of SLE and rheumatoid arthritis. Immature pDCs are activated through innate toll receptors TLRs, TLR7 and TLR9 by immune complexes to produce inflammatory cytokines including type I INF. High levels of serum IFN type I together with overexpression of IFN inducible genes have been found in individuals with SLE. The level of IFN correlated with the severity of the disease. The top 10 genes identified, associated with excessive production of IFN in viral infection, are I STAT1, IRF7, IRF5, IRF8 MX1, OASL, ISG15, IFIT3, IFIT1, OAS2 and GBP1 [14]. Among those, all associated with SLE only GBP1 was of recent association with the disease IRF5, IRF7 and IRF8 a family of transcription factors downstream of endosomal TLRs, are required for activating transcription of IFN-a and IFN-inducible genes [15, 16, 17]. Genetic variants on these three genes but especially variants of IRF5 and IRF7 have a functional impact on increased serum IFN and such impact depends on the presence of specific autoantibodies. The type I interferon pathway is central to disease pathogenesis. Hydroxychloroquine acts therapeutically on lupus by inhibiting the interferon pathway [18].

8. Complement genes and lupus

Evidence has revealed that complement deficiencies result in a reduced ability in the clearance of apoptotic cells that increase the production of autoantibodies and therefore SLE development in susceptible individuals. The C1q component – the first component of the classical complement pathway - that plays a significant role in the apoptotic cells is associated with SLE. C1q is encoded by three genes (a, b and c genes) all located in chromosome 1. The lupus autoantigens that are located in apoptotic debris may stimulate an inappropriate immune response. C1q may inhibit IFN-gamma production and, in this way, is involved in SLE development. When hereditary homozygous deficiency in any of the three C1q genes occurs, this leads to the development of SLE in all cases. C2, C4A and C4B genes which are part of the HLA class III genes – located at chromosome 6 – constitute components of the classical complement activation pathway. Observed in 0.01–0.02% of the general population C2 deficiency is the commonest but in lupus patients its prevalence is significantly higher depending on the region, 0.4% – 2.33% of people of European origin carry C2 deficiency – caused by a deletion on the DRW2 haplotype. These individuals will eventually develop lupus in their lifetime. C4 is important as a single gene defect. About 70% of the known cases of double homozygous C4 deficiency (C40, deficient at both C4A and C4B genes) result in a lupus phenotype. The C4A null allele is associated with almost every SLE population studied to date and may have an independent HLA effect [19].

9. Fc-gamma receptor gene polymorphisms and lupus erythematosus

The receptors for the Fc portion of IgG (FcγRs) play an important role in the clearance of immune complexes. They also present the modified antigen to the different populations of lymphocytes and are implicated in the modulation of inflammatory processes within the human immune system. Pre three families of FcγRs exist, the FcγRI, is the high-affinity receptor, while FcγRII and FcγRIII are low-affinity receptors. Failure of FcγR mediated clearance of immune complexes and control of inflammatory responses are thought to be predisposing factors for the development of SLE. The FCGR2/3 locus on chromosome 1q23.3 that encodes the low-affinity FcγRs is subject to both single nucleotide polymorphisms (SNP) and copy number variation (CNV). An SNP in the promoter region of FcγRIIb, also known as 2B.4, was found to be more frequently present in lupus [20].

10. Cytokines and SLE

IL-10 is a pivotal cytokine which inhibits T cells and antigen-presenting cells while enhancing B-cell survival and activity. IL-1β, IL-10 and TNF-a genes are important in the pathogenesis of SLE. It has been found that ILβ-511, IL-1β + 3953, IL-10-1082 and TNF-a-308 polymorphisms may be linked to the risk of lupus development and also to a specific phenotype. The high TNF alpha genotypes - 308AA - were associated with SLE independently of IL10 alleles, but the risk of developing CCLE and the prevalence of discoid lesion phenotype - in SLE - were higher in the high IL10/low TNF alpha producer group (−1082/−308GG). In addition, interaction between different cytokines modifies the appearance of autoantibodies. Patients who produce low levels of both TNF alpha and IL10 present anti-Sm-antibodies while patients who produce low IL10/high TNF alpha present more frequently antibodies to SSa and SSb. Furthermore, interleukin 23R gene polymorphisms especially the IL23Rrs10889677 confers SLE susceptibility to individuals of certain ethnicities, such as IL17 A haplotype polymorphisms. In addition, Interleukin-1 receptor antagonist gene polymorphism is a disease severity factor in SLE [21, 22, 23, 24].

11. Nuclear factor kappa-B (NF-κB) and lupus

NF-κB is an inflammation driving factor that mediates the release of IL-6, IL-12 and TNF. In healthy individuals, the A20 binding inhibitors of NF-κB (ABINs1–3) help keeping it in an inactive form in the cytoplasm. It is its activation followed by phosphorylation and degradation and subsequently induction of gene expression that is associated with the pathogenesis of SLE. The gene encoding the ABIN1 protein presents polymorphism that is associated with a predisposition for autoimmune disease. The TNIP1 (TNFAIP3-interacting protein 1) gene locus that encodes for the protein ABIN1 is associated with predisposition to SLE [25].

12. ITGAM gene and SLE

ITGAM produces macrophage antigen 1(MAC1), an adhesion molecule found on the surface of myeloid cells, natural killer cells and a subgroup of B cells. This is achieved when CD11b-integrin Am - encoded by ITGAM - is united to C818-integrin β2. Leucocyte adhesion and migration may be influenced by MAC1 as it binds to intercellular adhesion molecules - ICAM1 and ICAM2. In addition, MAC1 is a receptor for the cleaved complement factor IC3b. A polymorphism of this gene, particularly the R77H - a single nucleotide polymorphism -(SNP), is associated with SLE by modifying the structure of the gene products. This results in an increased production of proinflammatory cytokines and a reduced Ic3b phagocytosis and cellular adhesion. This makes this polymorphism a probable therapeutic target for lupus treatment. ITGAM gene polymorphisms result in defective clearance of immune complexes and apoptotic cells and lead to initiation and maintenance of autoimmune responses and chronic inflammation in SLE. ITGAM gene polymorphisms are associated with increased susceptibility to CCLE rather than to SLE [26].

13. PPP2CA gene and SLE

The protein phosphatase PP2A consists of three subunits but evidence reveals only the involvement of the catalytic subunit PP2Ac in the pathogenesis of SLE. PP2A controls various cellular pathways including DNA replication, gene translation and cell differentiation. The enzymatic activity of thePP2Ac is augmented in SLE. PP2Ac is dysregulated in SLE, resulting in altered transcription factor activation in T cells that affects their function by decreasing their capacity to produce IL-2 and reducing their expression of the T cell receptor (TCR) associated signaling molecule CD3ζ [27].

14. SIAE gene and SLE

Cd22 - also known as SIGLEC2 - is a lectin that belongs to the sialic acid binding immunoglobulin-like lectin family. It is expressed by B cells and functions as a negative regulator of B cell activation. Accordingly, deficiencies of SIAE and CD22 produce phenotypes characterized by hyperactive B cells and spontaneous autoimmunity as in SLE in animal models. In fact, exon sequencing of the SIAE gene in a small group of patients with high titers of antinuclear antibodies (13 of whom had defined autoimmune diseases) found that 2 out of 19 had rare loss of function variants of SIAE. Loss of function alleles was found to be more common in patients with autoimmune diseases [27].

15. Signaling lymphocytic activation molecule SLAMF molecules

They comprise nine type I trans-membrane glycoprotein receptors that provide potent co-stimulatory signals for the TCR-CD3, complex and mediate regulatory signals between immune cells. Defective SLAMF signals result in B and T cell abnormalities and impaired antibody production. SLAMF 3 and SLAMF 6 molecules are expressed at higher levels on T cells in patients with lupus contributing to SLE pathogenesis [27].

16. PTPN22 and SLE

Protein tyrosine phosphatase non-receptor type 22 (PTPN22) is a negative regulator of T cell activation associated with several autoimmune diseases and SLE. In individuals of European ancestry, The rs2476601 is the most significantly with SLE associated polymorphism expressed at higher levels on T cells from patients with lupus contributing to SLE pathophysiology [27, 28].

17. BLK and BANK1 genes and lupus

Variants of those two genes which are present alone or in combination in a substantial proportion of lupus patients impair suppression of IRF5 and type-I IFN in human B cell lines and likely contribute to genetic risk [29].

18. PD-1 gene and lupus

PD-1 (Programmed Cell Death Protein 1) discovered in 1992 and described as the rheostat of the immune reaction, because expressed in cells of the lymphoid tissue it inhibits their effector action allowing tumour development, while its blockade induces a robust anti-tumour activity and at the same time the appearance of autoimmune phenomena. Antibodies against PD-1 are currently widely used in cancer chemotherapy but are associated with induction of autoimmunity and subsequently SLE [30].

19. Genetics of chronic cutaneous lupus erythematosus (CCLE)

CCLE in the past referred to as discoid lupus represents the papulosquamous scarring form of cutaneous lupus erythematosus. Lupus panniculitis and lupus timidus also belong to this group. Lesions similar to these of CCLE have been described in carriers of the X-linked granulomatous disease [31]. Initial research found an association of HLA-A1, B8, HLA-B15, HLA-DRB10303 and HLA-DQA1 haplotype with CCLE. HLA-B8 was more frequently found in white CCLE patients while the HLA-DRW6 was found in an increased proportion of patients with DLE of both white and black races [32]. An association of HLA-DRB116 with CCLE in Mexican mestizo patients, the HLA-DRB116 gene is associated with most immune disorders mediated by the production of autoantibodies [33]. In a homozygous C2 deficiency patient with CCLE lupus, the C2 deficiency gene was associated with HLA-A10, B18, DR2, C4A4B2, Bfs, on one chromosome and with HLA-A2, B7 DR2, C4A4B2, Bfs on the other [34]. Susceptibility to CCLE lupus was recently noticed in a high interleukin10/low TNF alpha producer genotype (−1082 gg-308GG) [35]. A variant of the gene encoding the TRAF31P2, a member of the TNF receptor pathway involved in autoimmune diseases - missense substitution Thr438Asn - has been associated with discoid lupus and folliculitis decalvans [36]. Recently studies have identified 10 hub genes associated with CCLE which are CXCL10, CCR7, FPR3, PPARGC1A, MMP9, IRF7, IL2RG, SOCS1, ISG15 andGSTM3. Although general signs may be absent from skin in CCLE, it is a site of important disease activity. Genetic mapping recently showed overlapping of regions in seven (7) chromosomes - mainly genes of interferons and tumor necrosis factor-alpha between SLE and CCLE. In conclusion, these findings indicate that a genetic risk is shared by both SLE and CCLE [37, 38]. Micro RNA associated with epigenetic processes has also been found decreased in the serum of patients with CCLE [39].

20. Genetics of subacute cutaneous lupus erythematosus (SCLE)

Subacute cutaneous lupus erythematosus represents a widespread, photosensitive, nonscarring, nonindurated form of lupus erythematosus., associated with a distinctive immunogenetic background including the production of Ro/SS-antibodies. Patients with subacute cutaneous lupus erythematosus present in most cases the HLA-A1, B8 and DR3 haplotype that is also observed in approximately 25% of the North American Caucasians; this haplotype is now referred as the 8.1 ancestral haplotype. Partial or complete deficiency in C2 and C4, whose genes are located on chromosome 6, has been reported in some patients with SCLEA single nucleotide polymorphism (SNP) in the TNF-alpha gene promoter (−308A) encoding excessive TNF-alpha expression has been associated with skin lesions in patients suffering from SCLE (the TNF, a gene is also located within the HLA region). Also, a robust association of photosensitive systemic lupus erythematosus and a complete congenital deficiency of C1q have been recently revealed. In addition, SCLE subphenotype was significantly associated with a single nucleotide polymorphism (SNP) in the second exon of the gene encoding the A chain of C1q, which is the molecule that initiates the classical pathway of complement. SCLE patients homozygous for this SNP had lower serum levels of Ciq antigen compared to SCLE patients not having this SNP. To date this C1q an SNP is the only genetic association of SCLE that lies outside the HLA region. Two recent studies are in accordance with finding that in both SCLE and CCLE IFN-pathways are increased, but CCLE does indeed express more IFN-gamma than SCLE [40, 41].

21. Genetics of neonatal lupus

Neonatal lupus erythematosus is noticed in the newborns of women suffering from SLE, and it is characterized by an erythematosus rash around the eyes similar to spectacles, a heart block and the presence of anti-Ro antibodies in the serum. Neonatal lupus has been associated with the HLA-DRB102, HLA-DRB103 and the-308A allele linked to higher TNF- alpha production; these are present in the majority of children with this rash [42].

22. Chilblain lupus erythematosus and the TREX1 gene

Chilblain lupus erythematosus is a rare subtype of chronic cutaneous lupus more frequently encountered in middle-aged women and can evolve into SLE in some cases. (18–20%) The clinical picture is characteristic of an acral distribution of bluish-red inflammatory skin lesions on upper and lower extremities following exposure to cold or damp weather. Rarely familial cases have been reported and are associated with autosomal dominant mutations in the TREX gene encoding the 3–5 DNA exonuclease. The TREX1 gene is located on chromosome 3p21. 0.5–3% of all patients with SLE carried mutations in TREX1 [43, 44].

23. Lupus and gender—X-linked genes associated with lupus

Lupus erythematosus is a disease that affects nine (9) times more frequently females than males. Females have a stronger immune system than males; this can be explained by evolutionary biology. Women are destined to be pregnant and during pregnancy their immune system is suppressed to tolerate the existence and development of the fetus. Therefore, their immune system has to be more robust than that of males to compensate for this relative deficiency during gestation - this is the compensation theory. According to the Lyon theory, women have two chromosomes, but one copy is turned off in every cell very early in embryonic development; this process is known as X inactivation. Recent research, however, revealed that 15 per cent of the genes of the supposedly inactivated X chromosome are still turned on that the number of certain proteins produced in women is an increase compared with that in men. Particularly in women with lupus, some genes are active on both X copies and this higher activity correlates with disease severity [45]. Anguera M has also recently discovered that in mice mature immune cells undergo significant dynamic changes that could make it easier for X-linked genes to get turned on when they should be off.

24. The AIRE gene

The AIRE gene was discovered in 1997 and plays an important role in autoimmune regulation. As it is expressed by cells in the thymus, it helps T cells to recognize if proteins presented to them are components of the self or non-self substances. It influences the expression of a wide variety of self-antigens in the thymus and is essential to the negative selection of self-reactive cells and the establishment of self-tolerance. The activity of AIRE is partially controlled by sex hormones, with estrogens and progesterone turning down the expression of AIRE while testosterone increasing its production. Under the influence of sex hormones, women at puberty make less AIRE than men, resulting in more self-reactive T cells escaping from the thymus and causing autoimmune disease. It has been discovered that the AIRE Ser196Ser synonymous variant is a risk factor for SLE.

Another X-linked gene is the gene for toll-like receptor7 or TLR-7 that encodes a protein that recognizes pathogens and increases the production of interferons - molecules directly implicated in the pathogenesis of lupus. The identification of TASL as the component that links endolysosomal TLRs to the IRF5 transcription factor via SLC15A4 provides a mechanistic explanation for the involvement of these proteins in systemic lupus erythematosus. Also, X-linked gene CXorf21 may contribute to sexual dimorphism in systemic lupus erythematosus [46, 47].

25. Epigenetic changes and SLE

Epigenetic processes are molecular events that affect gene expression by reorganizing the structure of chromatin without altering the DNA sequence [48]. Such mechanisms include:

1. CpG-DNA methylation.

2. Post-translational modification of histone tails.

3. micro RNAs (miRNAs).

Epigenetic patterns can be modified by environmental factors or internal ones such as medicines administered to the patients. SLE is one of the disorders where epigenetics plays a major role in the development of the disease. Monozygotic twins that are equipped with the same genome for SLE susceptibility present less than 50% concordance rate. This implies that other mechanisms, mostly epigenetics, are implicated in the pathogenesis of SLE.

26. CpG-DNA methylation

B and T cells from patients with SLE exhibit a global decrease in CpG-DNA methylation that correlates with disease activity. Increased methylation of the IL2 gene results in the failure of T cells in patients with SLE to express IL2, while other cytokine genes are overexpressed in T cells in patients with SLE as a result of CpG-DNA hypomethylation. Such genes include IL4, IL6, IL10, IL13, IL17 and genes encoding various surface molecules, namely CD6, CD11a, CD40L and CD70. All these events produce increased numbers of effector memory CD4+ and contribute to the proinflammatory phenotype of SLE.

27. Post-translational histone modifications

The most common histone modifications include acetylation, phosphorylation, methylation, ubiquitylation and citrullination of histone tails; however, histone modifications are even more complex than CpG-DNA methylation patterns and remain poorly understood. Histone modifications care altered in T cells from patients with SLE. These modifications result in increased TNF expression and subsequent monocyte maturation and cytokine expression. IL2 levels increase while IL17 levels augment as a result of the different expression of the relevant genes.

28. MicroRNAs

MicroRNAs are short non-coding(21–23 nucleotide) RNA molecules that act as post-transcriptional regulators of gene expression and function by forming duplexes with target mRNAs, resulting in mRNA degradation or translational arrest. MicroRNAs may be deregulated in SLE inducing the activation of type I IFN and NκB pathways and also by promoting the release of chemokines leading to exacerbation of inflammatory responses, also by reducing DNA methylation.

Molecular genetics of SLE will in the future provide the clinician with useful therapeutic tools and a cure will be achieved without even awareness of the event that initiated the inflammatory process.

Abbreviations

SLE	Systemic Lupus Erythematosus.
CCLE	Chronic Cutaneous Lupus Erythematosus.
SCLE	Subacute Cutaneous Lupus Erythematosus.