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Systemic lupus erythematosus (SLE) is a heterogeneous autoimmune disease with established genetic susceptibility. Genetic studies have revealed that SLE is largely a complex genetic disease influenced by the cumulative effect of multiple genetic variants, most of which are nonprotein coding, dispersed across the genome. Research has successfully identified more than 120 risk loci, defined mainly by haplotypes (a set of variants that are coinherited), that are associated with increased SLE susceptibility. Current efforts use publicly available bioinformatics data, fine-mapping and/or dense imputation, and molecular studies to identify the functional variants that drive the haplotype association and pathologic effect. In this chapter, we emphasize the complex genetic heritability of SLE by highlighting several risk associations from the current literature that demonstrate the complexity and recent advances in our understanding of how non-HLA SLE-susceptibility loci contribute to disease pathology.

It is generally agreed that disease features of systemic lupus erythematosus (SLE) manifest in genetically predisposed individuals after encountering environmental triggers. Genome wide association studies provide an unbiased approach to screening hundreds of thousands of single nucleotide polymorphisms and have enhanced the discovery of genetic risk loci associated with SLE. Further advancements in the field (large-scale replication studies, meta-analysis, next generation sequencing, fine mapping of disease characteristics, and functional studies) have led a rapid increase in the discovery of SLE risk variants, approaching 150 loci identified. Although functions of a number of identified loci remain unknown, most risk gene products participate in various aspects of the immune system, including the clearance of immune complexes, type I interferon and NFkB pathways, B and T cell signaling, and transcriptional regulation, which are highlighted herein. Updated progress in lupus genetics, the identification of new causal variants, the discovery of novel pathways and regulatory mechanisms, and advances in utilizing SLE-risk loci to predict disease susceptibility and therapeutic strategies are reviewed. Here we emphasize the progress made in the last few years that has greatly enhanced our understanding of SLE pathogenesis, which will serve to guide the development of novel therapies to treat the disease.

Genetic variation within the major histocompatibility complex (MHC) contributes substantial risk for systemic lupus erythematosus, but high gene density, extreme polymorphism and extensive linkage disequilibrium (LD) have made fine mapping challenging. To address the problem, we compared two association techniques in two ancestrally diverse populations, African Americans (AAs) and Europeans (EURs). We observed a greater number of Human Leucocyte Antigen (HLA) alleles in AA consistent with the elevated level of recombination in this population. In EUR we observed 50 different A—C—B—DRB1—DQA—DQB multilocus haplotype sequences per hundred individuals; in the AA sample, these multilocus haplotypes were twice as common compared to Europeans. We also observed a strong narrow class II signal in AA as opposed to the long-range LD observed in EUR that includes class I alleles. We performed a Bayesian model choice of the classical HLA alleles and a frequentist analysis that combined both single nucleotide polymorphisms (SNPs) and classical HLA alleles. Both analyses converged on a similar subset of risk HLA alleles: in EUR HLA– B*08:01 + B*18:01 + (DRB1*15:01 frequentist only) + DQA*01:02 + DQB*02:01 + DRB3*02 and in AA HLA–C*17:01 + B*08:01 + DRB1*15:03 + (DQA*01:02 frequentist only) + DQA*02:01 + DQA*05:01+ DQA*05:05 + DQB*03:19 + DQB*02:02. We observed two additional independent SNP associations in both populations: EUR rs146903072 and rs501480; AA rs389883 and rs114118665. The DR2 serotype was best explained by DRB1*15:03 + DQA*01:02 in AA and by DRB1*15:01 + DQA*01:02 in EUR. The DR3 serotype was best explained by DQA*05:01 in AA and by DQB*02:01 in EUR. Despite some differences in underlying HLA allele risk models in EUR and AA, SNP signals across the extended MHC showed remarkable similarity and significant concordance in direction of effect for risk-associated variants.

Human leukocyte antigen (HLA) is a key genetic factor conferring risk of systemic lupus erythematosus (SLE), but precise independent localization of HLA effects is extremely challenging. As a result, the contribution of specific HLA alleles and amino-acid residues to the overall risk of SLE and to risk of specific autoantibodies are far from completely understood. Here, we dissected (a) overall SLE association signals across HLA, (b) HLA-peptide interaction, and (c) residue-autoantibody association. Classical alleles, SNPs, and amino-acid residues of eight HLA genes were imputed across 4,915 SLE cases and 13,513 controls from Eastern Asia. We performed association followed by conditional analysis across HLA, assessing both overall SLE risk and risk of autoantibody production. DR15 alleles HLA-DRB1*15:01 (P = 1.4x10-27, odds ratio (OR) = 1.57) and HLA-DQB1*06:02 (P = 7.4x10-23, OR = 1.55) formed the most significant haplotype (OR = 2.33). Conditioned protein-residue signals were stronger than allele signals and mapped predominantly to HLA-DRB1 residue 13 (P = 2.2x10-75) and its proxy position 11 (P = 1.1x10-67), followed by HLA-DRB1-37 (P = 4.5x10-24). After conditioning on HLA-DRB1, novel associations at HLA-A-70 (P = 1.4x10-8), HLA-DPB1-35 (P = 9.0x10-16), HLA-DQB1-37 (P = 2.7x10-14), and HLA-B-9 (P = 6.5x10-15) emerged. Together, these seven residues increased the proportion of explained heritability due to HLA to 2.6%. Risk residues for both overall disease and hallmark autoantibodies (i.e., nRNP: DRB1-11, P = 2.0x10-14; DRB1-13, P = 2.9x10-13; DRB1-30, P = 3.9x10-14) localized to the peptide-binding groove of HLA-DRB1. Enrichment for specific amino-acid characteristics in the peptide-binding groove correlated with overall SLE risk and with autoantibody presence. Risk residues were in primarily negatively charged side-chains, in contrast with rheumatoid arthritis. We identified novel SLE signals in HLA Class I loci (HLA-A, HLA-B), and localized primary Class II signals to five residues in HLA-DRB1, HLA-DPB1, and HLA-DQB1. These findings provide insights about the mechanisms by which the risk residues interact with each other to produce autoantibodies and are involved in SLE pathophysiology., Author summary The Human leukocyte antigen (HLA) region is a key genetic factor conferring risk of systemic lupus erythematosus (SLE). In spite of multiple SLE association signals identified in the HLA region, only amino-acid residues within HLA-DRB1 have been specifically described previously. In this study, we performed an imputation-based analysis on individuals with East Asian ancestry, and characterized SLE risk within the HLA region for all involved independent genes (HLA-DRB1, HLA-DPB1, HLA-DQB1, HLA-A, and HLA-B). Furthermore, we identified a characteristic SLE risk residue signature as well as a pattern of specific nRNP and Ro/La autoantibody residues located in the peptide-binding grooves, suggesting their key involvement in autoantibody production.

The interplay of genetic predispositions, epigenetic changes, and environmental exposures contributes to the complex etiology of systemic lupus erythematosus (SLE). During the last decade, genome-wide association studies (GWAS) followed by meta-analyses and large-scale replication/ fine-mapping studies have yielded a substantial number of genomic loci linked to SLE susceptibility, which helps to enhance our understanding of SLE pathogenesis to the molecular level. This chapter reviews recent progress toward identifying causative variants at the SLE-associated loci, highlights key immune regulatory genes with specific SLE phenotype features shared among major immune-mediated diseases, and discusses molecular pathways perturbed by these disease loci revealing new insights into the disease pathogenesis.

Objective Systemic lupus erythematosus (SLE) is a chronic autoimmune disease with a strong genetic component. We undertook the present work to perform the first genome-wide association study on individuals from the Americas who are enriched for Native American heritage. Methods We analyzed 3,710 individuals from the US and 4 countries of Latin America who were diagnosed as having SLE, and healthy controls. Samples were genotyped with HumanOmni1 BeadChip. Data on out-of-study controls genotyped with HumanOmni2.5 were also included. Statistical analyses were performed using SNPtest and SNPGWA. Data were adjusted for genomic control and false discovery rate. Imputation was performed using Impute2 and, for classic HLA alleles, HiBag. Odds ratios (ORs) and 95% confidence intervals (95% CIs) were calculated. Results The IRF5–TNPO3 region showed the strongest association and largest OR for SLE (rs10488631: genomic control–adjusted P [Pgcadj] = 2.61 × 10−29, OR 2.12 [95% CI 1.88–2.39]), followed by HLA class II on the DQA2–DQB1 loci (rs9275572: Pgcadj = 1.11 × 10−16, OR 1.62 [95% CI 1.46–1.80] and rs9271366: Pgcadj = 6.46 × 10−12, OR 2.06 [95% CI 1.71–2.50]). Other known SLE loci found to be associated in this population were ITGAM, STAT4, TNIP1, NCF2, and IRAK1. We identified a novel locus on 10q24.33 (rs4917385: Pgcadj = 1.39 × 10−8) with an expression quantitative trait locus (eQTL) effect (Peqtl = 8.0 × 10−37 at USMG5/miR1307), and several new suggestive loci. SLE risk loci previously identified in Europeans and Asians were corroborated. Local ancestry estimation showed that the HLA allele risk contribution is of European ancestral origin. Imputation of HLA alleles suggested that autochthonous Native American haplotypes provide protection against development of SLE. Conclusion Our results demonstrate that studying admixed populations provides new insights in the delineation of the genetic architecture that underlies autoimmune and complex diseases.

Background Genetic susceptibility of SLE in general is attributed to the overall risk of multiple common variants that each confers a small effect. However, in few cases, highly penetrant single gene variants have been reported as monogenic forms of SLE. To explore novel risk variants, we carried out whole-exome sequencing to identify underlying monogenic causes from two multiplex families that each family has two boys with childhood onset lupus nephritis. Methods We sequenced the whole exome of lupus patients and their parents using the Illumina’s instrument Hiseq2000. We conducted variant calling and annotation using the Genome Analysis Toolkit GATK and ANNOVAR, respectively. Our findings of exome-seq was confirmed using the Sanger sequencing. Results Using bioinformatics, we focused only on potential loss-of-function variants. In addition, by using the recessive inheritance model and allele frequency 2 00 000 individuals). In one family, the SAT1 frameshift mutation was transmitted from the mother to the two sons affected with SLE but not to the unaffected son. Conclusions We identified SAT1 as a novel gene associated with monogenic lupus. SAT1 encodes the spermidine/spermine-N1-acetyltransferase (SSAT), a rate-limiting enzyme that regulates the catabolism of polyamine. We hypothesize that loss-of-function SAT1 variants may cause dysregulated polyamine homeostasis which confers risk of SLE.

Author(s): Lanata, Cristina M; Nititham, Joanne; Taylor, Kimberly E; Chung, Sharon A; Torgerson, Dara G; Seldin, Michael F; Pons-Estel, Bernardo A; Tusie-Luna, Teresa; Tsao, Betty P; Morand, Eric F; Alarcon-Riquelme, Marta E; Criswell, Lindsey A | Abstract: ObjectiveAfrican Americans, East Asians, and Hispanics with systemic lupus erythematous (SLE) are more likely to develop lupus nephritis (LN) than are SLE patients of European descent. The etiology of this difference is not clear, and this study was undertaken to investigate how genetic variants might explain this effect.MethodsIn this cross-sectional study, 1244 SLE patients from multiethnic case collections were genotyped for 817,810 single-nucleotide polymorphisms (SNPs) across the genome. Continental genetic ancestry was estimated utilizing the program ADMIXTURE. Gene-based testing and pathway analysis was performed within each ethnic group and meta-analyzed across ethnicities. We also performed candidate SNP association tests with SNPs previously established as risk alleles for SLE, LN, and chronic kidney disease (CKD). Association testing and logistic regression models were performed with LN as the outcome, adjusted for continental ancestries, sex, disease duration, and age.ResultsWe studied 255 North European, 263 South European, 238 Hispanic, 224 African American and 264 East Asian SLE patients, of whom 606 had LN (48.7%). In genome-wide gene-based and candidate SNP analyses, we found distinct genes, pathways and established risk SNPs associated with LN for each ethnic group. Gene-based analyses showed significant associations between variation in ZNF546 (p = 1.0E-06), TRIM15 (p = 1.0E-06), and TRIMI0 (p = 1.0E-06) and LN among South Europeans, and TTC34 (p = 8.0E-06) was significantly associated with LN among Hispanics. The SNP rs8091180 in NFATC1 was associated with LN (OR 1.43, p = 3.3E-04) in the candidate SNP meta-analysis with the highest OR among African-Americans (OR 2.17, p = 0.0035).ConclusionDistinct genetic factors are associated with the risk of LN in SLE patients of different ethnicities. CKD risk alleles may play a role in the development of LN in addition to SLE-associated risk variants. These findings may further explain the clinical heterogeneity of LN risk and response to therapy observed between different ethnic groups.

Systemic lupus erythematosus (SLE or lupus) (OMIM: 152700) is a chronic autoimmune disease with debilitating inflammation that affects multiple organ systems. The STAT1-STAT4 locus is one of the first and most highly replicated genetic loci associated with lupus risk. We performed a fine-mapping study to identify plausible causal variants within the STAT1-STAT4 locus associated with increased lupus disease risk. Using complementary frequentist and Bayesian approaches in trans-ancestral Discovery and Replication cohorts, we found one variant whose association with lupus risk is supported across ancestries in both the Discovery and Replication cohorts: rs11889341. In B cell lines from patients with lupus and healthy controls, the lupus risk allele of rs11889341 was associated with increased STAT1 expression. We demonstrated that the transcription factor HMGA1, a member of the HMG transcription factor family with an AT-hook DNA-binding domain, has enriched binding to the risk allele compared with the non-risk allele of rs11889341.We identified a genotype-dependent repressive element in the DNA within the intron of STAT4 surrounding rs11889341. Consistent with expression quantitative trait locus (eQTL) analysis, the lupus risk allele of rs11889341 decreased the activity of this putative repressor. Altogether, we present a plausible molecular mechanism for increased lupus risk at the STAT1-STAT4 locus in which the risk allele of rs11889341, the most probable causal variant, leads to elevated STAT1 expression in B cells due to decreased repressor activity mediated by increased binding of HMGA1. © The Author(s) 2018. Published by Oxford University Press. All rights reserved.

Leptin is abnormally elevated in the plasma of patients with systemic lupus erythematosus (SLE), where it is thought to promote and/or sustain pro-inflammatory responses. Whether this association could reflect an increased genetic susceptibility to develop SLE is not known, and studies of genetic associations with leptin-related polymorphisms in SLE patients have been so far inconclusive. Here we genotyped DNA samples from 15,706 SLE patients and healthy matched controls from four different ancestral groups, to correlate polymorphisms of genes of the leptin pathway to risk for SLE. It was found that although several SNPs showed weak associations, those associations did not remain significant after correction for multiple testing. These data do not support associations between defined leptin-related polymorphisms and increased susceptibility to develop SLE. © 2015 Elsevier Inc.

Complement receptor 2 (CR2/CD21) is predominantly expressed on the surface of mature B cells where it forms part of a coreceptor complex that functions, in part, to modulate B-cell receptor signal strength. CR2/CD21 expression is tightly regulated throughout B-cell development such that CR2/CD21 cannot be detected on pre-B or terminally differentiated plasma cells. CR2/CD21 expression is upregulated at B-cell maturation and can be induced by IL-4 and CD40 signaling pathways. We have previously characterized elements in the proximal promoter and first intron of CR2/CD21 that are involved in regulating basal and tissue-specific expression. We now extend these analyses to the CR2/CD21 core promoter. We show that in mature B cells, CR2/CD21 transcription proceeds from a focused TSS regulated by a non-consensus TATA box, an initiator element and a downstream promoter element. Furthermore, occupancy of the general transcriptional machinery in pre-B versus mature B-cell lines correlate with CR2/CD21 expression level and indicate that promoter accessibility must switch from inactive to active during the transitional B-cell window.

Our understanding on genetic basis of SLE has been advanced through genome-wide association studies. We review recent progress in lupus genetics with a focus on SLE-associated loci that have been functionally characterized, and discuss the potential for clinical translation of genetics data. Over 100 loci have been confirmed to show robust association with SLE and many share with other immune-mediated diseases. Although causative variants captured at these established loci are limited, they guide biological studies of gene targets for functional characterization which highlight the importance of aberrant recognition of self-nucleic acid, type I interferon overproduction, and defective immune cell signaling underlying the pathogenesis of SLE. Increasing examples illustrate a predictive value of genetic findings in susceptibility/prognosis prediction, clinical classification, and pharmacological implication. Genetic findings provide a foundation for better understanding of disease pathogenic mechanisms and opportunities for target selection in lupus drug development.

Systemic lupus erythematosus (SLE) is a heterogeneous autoimmune disease with a strong genetic component characterized by autoantibody production and a type I interferon signature. Here we report a missense variant (g.74779296G>A; p.Arg90His) in NCF1, encoding the p47phox subunit of the phagocyte NADPH oxidase (NOX2), as the putative underlying causal variant that drives a strong SLE-associated signal detected by the Immunochip in the GTF2IRD1-GTF2I region at 7q11.23 with a complex genomic structure. We show that the p.Arg90His substitution, which is reported to cause reduced reactive oxygen species (ROS) production, predisposes to SLE (odds ratio (OR) = 3.47 in Asians (Pmeta = 3.1 × 10-104), OR = 2.61 in European Americans, OR = 2.02 in African Americans) and other autoimmune diseases, including primary Sjögren's syndrome (OR = 2.45 in Chinese, OR = 2.35 in European Americans) and rheumatoid arthritis (OR = 1.65 in Koreans). Additionally, decreased and increased copy numbers of NCF1 predispose to and protect against SLE, respectively. Our data highlight the pathogenic role of reduced NOX2-derived ROS levels in autoimmune diseases.

Systemic lupus erythematosus (SLE) is an autoimmune disease with marked gender and ethnic disparities. We report a large transancestral association study of SLE using Immunochip genotype data from 27,574 individuals of European (EA), African (AA) and Hispanic Amerindian (HA) ancestry. We identify 58 distinct non-HLA regions in EA, 9 in AA and 16 in HA (∼50% of these regions have multiple independent associations); these include 24 novel SLE regions (P, Systemic lupus erythematosus (SLE) is an autoimmune disease with a strong ethnic and gender bias. In a transancestral genetic association study, Langefeld et al. identify 24 novel regions associated with risk to lupus and propose a cumulative hits hypothesis for loci conferring risk to SLE.

Exploiting genotyping, DNA sequencing, imputation and trans-ancestral mapping, we used Bayesian and frequentist approaches to model the IRF5-TNPO3 locus association, now implicated in two immunotherapies and seven autoimmune diseases. Specifically, in systemic lupus erythematosus (SLE), we resolved separate associations in the IRF5 promoter (all ancestries) and with an extended European haplotype. We captured 3230 IRF5-TNPO3 high-quality, common variants across 5 ethnicities in 8395 SLE cases and 7367 controls. The genetic effect from the IRF5 promoter can be explained by any one of four variants in 5.7 kb (P-valuemeta = 6 × 10-49; OR = 1.38-1.97). The second genetic effect spanned an 85.5-kb, 24-variant haplotype that included the genes IRF5 and TNPO3(P-valuesEU = 10-27-10-32, OR = 1.7-1.81). Many variants at the IRF5 locus with previously assigned biological function are not members of either final credibleset of potential causal variants identified herein. In addition to the known biologically functional variants, we demonstrated that the risk allele of rs4728142, a variant in the promoter among the lowest frequentist probability and highest Bayesian posterior probability, was correlated with IRF5 expression and differentially binds the transcription factor ZBTB3. Our analytical strategy provides a novel framework for future studies aimed at dissecting etiological genetic effects. Finally, both SLE elements of the statistical model appear to operate in Sjögren's syndrome and systemic sclerosis whereas only the IRF5-TNPO3 gene-spanning haplotype is associated with primary biliary cirrhosis, demonstrating the nuance of similarity and difference in autoimmune disease risk mechanisms at IRF5-TNPO3. © The Author 2014.

In a Genome Wide Association Study (GWAS) of individuals of European ancestry afflicted with Systemic Lupus Erythematosus (SLE) the extensive utilization of imputation, stepwise multiple regression, lasso regularization, and increasing study power by utilizing False Discovery Rate (FDR) instead of a Bonferroni multiple test correction enabled us to identify 13 novel non-human leukocyte antigen (HLA) genes and confirmed the association of 4 genes previously reported to be associated. Novel genes associated with SLE susceptibility included two transcription factors (EHF, and MED1), two components of the NFκB pathway (RASSF2 and RNF114), one gene involved in adhesion and endothelial migration (CNTN6), and two genes involved in antigen presentation (BIN1 and SEC61G). In addition, the strongly significant association of multiple single nucleotide polymorphisms (SNPs) in the HLA region was assigned to HLA alleles and serotypes and deconvoluted into four primary signals. The novel SLE-associated genes point to new directions for both the diagnosis and treatment of this debilitating autoimmune disease.

Objective.Little is known about the genetic etiology of systemic lupus erythematosus (SLE) in individuals of African ancestry, despite its higher prevalence and greater disease severity. Overproduction of nitric oxide (NO) and reactive oxygen species are implicated in the pathogenesis and severity of SLE, making NO synthases and other reactive intermediate-related genes biological candidates for disease susceptibility. We analyzed variation in reactive intermediate genes for association with SLE in 2 populations with African ancestry.Methods.A total of 244 single-nucleotide polymorphisms (SNP) from 53 regions were analyzed in non-Gullah African Americans (AA; 1432 cases and 1687 controls) and the genetically more homogeneous Gullah of the Sea Islands of South Carolina (133 cases and 112 controls). Single-marker, haplotype, and 2-locus interaction tests were computed for these populations.Results.The glutathione reductase geneGSR(rs2253409; p = 0.0014, OR 1.26, 95% CI 1.09–1.44) was the most significant single SNP association in AA. In the Gullah, the NADH dehydrogenaseNDUFS4(rs381575; p = 0.0065, OR 2.10, 95% CI 1.23–3.59) and NO synthase geneNOS1(rs561712; p = 0.0072, OR 0.62, 95% CI 0.44–0.88) were most strongly associated with SLE. When both populations were analyzed together,GSRremained the most significant effect (rs2253409; p = 0.00072, OR 1.26, 95% CI 1.10–1.44). Haplotype and 2-locus interaction analyses also uncovered different loci in each population.Conclusion.These results suggest distinct patterns of association with SLE in African-derived populations; specific loci may be more strongly associated within select population groups.

The human immune system defends the body against microbes and other threats. However, if this process goes wrong the immune system can attack the body’s own healthy cells, which can lead to serious autoimmune diseases. Systemic lupus erythematosus (SLE) is an autoimmune disease in which immune cells often attack internal organs – including the kidneys, nervous system and heart. Over the past decade, multiple genes have been linked with an increased risk of SLE. However, it is largely unknown how the sequences of these genes differ between individuals with SLE and healthy individuals, and the precise changes that lead to an increased risk of SLE are also not clear. Now, Raj, Rai et al. have determined the genetic sequences of over 700 people with SLE and over 500 healthy individuals and looked for differences that influence susceptibility to the disease. The vast majority of differences were discovered in stretches of DNA that regulate the expression of nearby genes, rather than in DNA that encodes the structures of proteins. Notably, extensive differences were found in a region of the human genome that regulates the production of proteins called Human Leukocyte Antigen class II molecules; which are known to play a critical role in activating the immune system. Raj, Rai et al. found that slight changes to the regulatory DNA sequences resulted in an overabundance of these proteins, which led to a hyperactive immune system that is strongly associated with SLE. Future studies could now ask if the changes to the regulatory DNA sequences highlighted by Raj, Rai et al. increase susceptibility to other autoimmune disorders as well. It may also be possible to use the increased understanding of how the immune system is regulated to develop new ways to minimize the rejection of organ transplants.

The complex etiology of systemic lupus erythematosus (SLE) could be attributed to a combination of genetic predispositions, epigenetic changes, and environmental exposures. The genome-wide association study (GWAS) that screens hundreds of thousands of single nucleotide polymorphisms across the genome based on a hypothesis-free approach has accelerated discovery of genetic variants predisposing individuals to SLE. After the initial GWAS discovery, subsequent large-scale replication studies and meta-analyses have identified and confirmed over 50 SLE-risk loci that most are shared in multiple ancestries. We review recent progress toward identifying causative variants at the SLE-risk loci, highlighting genes with functions in innate and adaptive immune responses that contribute to disease susceptibility and specific manifestations or end organ damage in SLE. These findings have enhanced our understanding of SLE pathogenesis to the molecular level and will guide the development of new therapeutic interventions in the near future.

ObjectivesFollowing up the systemic lupus erythematosus (SLE) genome-wide association studies (GWAS) identification of NMNAT2 at rs2022013, we fine-mapped its 150 kb flanking regions containing NMNAT2 and SMG7 in a 15 292 case–control multi-ancestry population and tested functions of identified variants.MethodsWe performed genotyping using custom array, imputation by IMPUTE 2.1.2 and allele specific functions using quantitative real-time PCR and luciferase reporter transfections. SLE peripheral blood mononuclear cells (PBMCs) were cultured with small interfering RNAs to measure antinuclear antibody (ANA) and cyto/chemokine levels in supernatants using ELISA.ResultsWe confirmed association at NMNAT2 in European American (EA) and Amerindian/Hispanic ancestries, and identified independent signal at SMG7 tagged by rs2702178 in EA only (p=2.4×10−8, OR=1.23 (95% CI 1.14 to 1.32)). In complete linkage disequilibrium with rs2702178, rs2275675 in the promoter region robustly associated with SMG7 mRNA levels in multiple expression quantitative trait locus (eQTL) datasets. Its risk allele was dose-dependently associated with decreased SMG7 mRNA levels in PBMCs of 86 patients with SLE and 119 controls (p=1.1×10−3 and 6.8×10−8, respectively) and conferred reduced transcription activity in transfected HEK-293 (human embryonic kidney cell line) and Raji cells (p=0.0035 and 0.0037, respectively). As a critical component in the nonsense-mediated mRNA decay pathway, SMG7 could regulate autoantigens including ribonucleoprotein (RNP) and Smith (Sm). We showed SMG7 mRNA levels in PBMCs correlated inversely with ANA titres of patients with SLE (r=−0.31, p=0.01), and SMG7 knockdown increased levels of ANA IgG and chemokine (C-C motif) ligand 19 in SLE PBMCs (p=2.0×10−5 and 2.0×10−4, respectively).ConclusionWe confirmed NMNAT2 and identified independent SMG7 association with SLE. The inverse relationship between levels of the risk allele-associated SMG7 mRNAs and ANA suggested the novel contribution of mRNA surveillance pathway to SLE pathogenesis.

Objective American Indian-Europeans, Asians, and African Americans have an excess morbidity from systemic lupus erythematosus (SLE) and a higher prevalence of lupus nephritis than do Caucasians. The aim of this study was to analyze the relationship between genetic ancestry and sociodemographic characteristics and clinical features in a large cohort of American Indian-European SLE patients. Methods A total of 2,116 SLE patients of American Indian-European origin and 4,001 SLE patients of European descent for whom we had clinical data were included in the study. Genotyping of 253 continental ancestry-informative markers was performed on the Illumina platform. Structure and Admixture software were used to determine genetic ancestry proportions of each individual. Logistic regression was used to test the association between genetic ancestry and sociodemographic and clinical characteristics. Odds ratios (ORs) were calculated with 95% confidence intervals (95% CIs). Results The average American Indian genetic ancestry of 2,116 SLE patients was 40.7%. American Indian genetic ancestry conferred increased risks of renal involvement (P less than 0.0001, OR 3.50 [95% CI 2.63- 4.63]) and early age at onset (P less than 0.0001). American Indian ancestry protected against photosensitivity (P less than 0.0001, OR 0.58 [95% CI 0.44-0.76]), oral ulcers (P less than 0.0001, OR 0.55 [95% CI 0.42-0.72]), and serositis (P less than 0.0001, OR 0.56 [95% CI 0.41-0.75]) after adjustment for age, sex, and age at onset. However, age and sex had stronger effects than genetic ancestry on malar rash, discoid rash, arthritis, and neurologic involvement. Conclusion In general, American Indian genetic ancestry correlates with lower sociodemographic status and increases the risk of developing renal involvement and SLE at an earlier age. Copyright © 2012 by the American College of Rheumatology.

Objective Candidate gene and genome-wide association studies have identified several disease susceptibility loci in lupus patients. These studies have largely been performed in lupus patients who are Asian or of European ancestry. This study was undertaken to examine whether some of these same susceptibility loci increase lupus risk in African American individuals. Methods Single-nucleotide polymorphisms tagging 15 independent lupus susceptibility loci were genotyped in a set of 1,724 lupus patients and 2,024 healthy controls of African American descent. The loci examined included PTPN22, FCGR2A, TNFSF4, STAT4, CTLA4, PDCD1, PXK, BANK1, MSH5 (HLA region), CFB (HLA region), C8orf13-BLK region, MBL2, KIAA1542, ITGAM, and MECP2/IRAK1. Results We found the first evidence of genetic association between lupus in African American patients and 5 susceptibility loci (C8orf13-BLK, BANK1, TNFSF4, KIAA1542, and CTLA4; P = 8.0 × 10−6, P = 1.9 × 10−5, P = 5.7 × 10−5, P = 0.00099, and P = 0.0045, respectively). Further, we confirmed the genetic association between lupus and 5 additional lupus susceptibility loci (ITGAM, MSH5, CFB, STAT4, and FCGR2A; P = 7.5 × 10−11, P = 5.2 × 10−8, P = 8.7 × 10−7, P = 0.0058, and P = 0.0070, respectively), and provided evidence, for the first time, of genome-wide significance for the association between lupus in African American patients and ITGAM and MSH5 (HLA region). Conclusion These findings provide evidence of novel genetic susceptibility loci for lupus in African Americans and demonstrate that the majority of lupus susceptibility loci examined confer lupus risk across multiple ethnicities.

Systemic lupus erythematosus (SLE) is considered to be the prototypic autoimmune disease, with a complex genetic architecture influenced by environmental factors. We sought to replicate a putative association at 11p13 not yet exceeding genome-wide significance (p < 5 × 10-8) identified in a genome-wide association study (GWAS). Our GWA scan identified two intergenic SNPs located between PDHX and CD44 showing suggestive evidence of association with SLE in cases of European descent (rs2732552, p = 0.004, odds ratio [OR] = 0.78; rs387619, p = 0.003, OR = 0.78). The replication cohort consisted of >15,000 subjects, including 3562 SLE cases and 3491 controls of European ancestry, 1527 cases and 1811 controls of African American (AA) descent, and 1265 cases and 1260 controls of Asian origin. We observed robust association at both rs2732552 (p = 9.03 × 10-8, OR = 0.83) and rs387619 (p = 7.7 × 10-7, OR = 0.83) in the European samples with pmeta = 1.82 × 10-9 for rs2732552. The AA and Asian SLE cases also demonstrated association at rs2732552 (p = 5 × 10-3, OR = 0.81 and p = 4.3 × 10-4, OR = 0.80, respectively). A meta-analysis of rs2732552 for all racial and ethnic groups studied produced pmeta = 2.36 × 10-13. This locus contains multiple regulatory sites that could potentially affect expression and functions of CD44, a cell-surface glycoprotein influencing immunologic, inflammatory, and oncologic phenotypes, or PDHX, a subunit of the pyruvate dehydrogenase complex. © 2011 The American Society of Human Genetics.

Supported by grants from the NIH (5P01-AI-083194 to Drs. Lessard, Rasmussen, Gaffney, Alarcon-Riquelme, Criswell, Jacob, Kimberly, Vyse, Harley, Sivils, Langefeld, and Tsao; 5P01-AR-049084 to Drs. Kimberly and Langefeld; and R01-AR-043814 and R21-AR-065626 to Dr. Tsao), the Oklahoma Medical Research Foundation (to Drs. Lessard, Gaffney, and Sivils), the Alliance for Lupus Research (to Drs. Criswell and Tsao), and the Wake Forest School of Medicine Center for Public Health Genomics (to Dr. Langefeld). Dr. Criswell is recipient of a Kirkland Scholar Award. Sample collection and phenotyping of the subjects utilized in this study was funded by the Ministry for Health and Welfare, Republic of Korea (Korea Healthcare Technology R&D Project grant HI13C2124 to Dr. Bae, and HI13C1754 to Dr. Song). The out-of-study Korean control data were provided by the Korean Biobank Project, which is supported by the Korea Centers for Disease Control and Prevention at the Korea National Institute of Health.

A classic T-cell phenotype in systemic lupus erythematosus (SLE) is the downregulation and replacement of the CD3ζ chain that alters T-cell receptor signaling. However, genetic associations with SLE in the human CD247 locus that encodes CD3ζ are not well established and require replication in independent cohorts. Our aim was therefore to examine, localize and validate CD247-SLE association in a large multiethnic population. We typed 44 contiguous CD247 single-nucleotide polymorphisms (SNPs) in 8922 SLE patients and 8077 controls from four ethnically distinct populations. The strongest associations were found in the Asian population (11 SNPs in intron 1, 4.99 × 10(-4) < P < 4.15 × 10(-2)), where we further identified a five-marker haplotype (rs12141731-rs2949655-rs16859085-rs12144621-rs858554; G-G-A-G-A; P(hap) = 2.12 × 10(-5)) that exceeded the most associated single SNP rs858554 (minor allele frequency in controls = 13%; P = 4.99 × 10(-4), odds ratio = 1.32) in significance. Imputation and subsequent association analysis showed evidence of association (P < 0.05) at 27 additional SNPs within intron 1. Cross-ethnic meta-analysis, assuming an additive genetic model adjusted for population proportions, showed five SNPs with significant P-values (1.40 × 10(-3) < P< 3.97 × 10(-2)), with one (rs704848) remaining significant after Bonferroni correction (P(meta) = 2.66 × 10(-2)). Our study independently confirms and extends the association of SLE with CD247, which is shared by various autoimmune disorders and supports a common T-cell-mediated mechanism. National Institutes of Health grants: (UL1RR025741, K24AR002138, P602AR30692, P01AR49084, UL1TR000165, P01AI083194, RO1AR43814, P60AR053308, UL1TR000004, AR43727, R21AI070304, RO1AR057172, UL1RR025014, R01AR051545-03, UL1RR029882, P60AR062755, P30AR53483, U19AI082714, P30GM103510, U01AI101934, AI063274, AR056360, AI083194, R37AI024717, P01083194, P01AR049084, PR094002); Northwestern University Feinberg School of Medicine; University of Alabama Birmingham; National Institute of Arthritis and Musculoskeletal and Skin Diseases; University of California Los Angeles; University of California San Francisco; Hopkins University; University of Colorado School of Medicine; University of Southern California; Seattle Children's Research Institute Arthritis Foundation; Medical University of South Carolina; Oklahoma Medical Research Foundation; Cincinnati Children's Hospital Medical Center; US Departments of Defense grant: (PR094002); Veterans Affairs; Alliance for Lupus Research; Kirkland Scholar Award; Korea Healthcare technology R & D project: (A121983); Ministry for Health and Welfare; Republic of Korea; Swedish Research Council; Instituto de Salud Carlos III grant: (PS09/00129); European Union FEDER funds; Fundação para a Ciência e Tecnologia fellowships: (SFRH/BPD/29354/2006, SFRH/BPD/34648/2007).

We previously reported association of FCGR2B-Ile232Thr with systemic lupus erythematosus (SLE) in three Asian populations. Because polymorphism of CD72, another inhibitory receptor of B cells, was associated with murine SLE, we identified human CD72 polymorphisms, tested their association with SLE and examined genetic interaction with FCGR2B in the Japanese (160 SLE, 277 controls), Thais (87 SLE, 187 controls) and Caucasians (94 families containing SLE members). Four polymorphisms and six rare variations were detected. The former constituted two major haplotypes that contained one or two repeats of 13 nucleotides in intron 8 (designated as *1 and *2, respectively). Although association with susceptibility to SLE was not detected, the *1 allele was significantly associated with nephritis among the Japanese patients (P=0.024). RT-PCR identified a novel alternatively spliced (AS) transcript that was expressed at the protein level in COS-7 transfectants. The ratio of AS/common isoforms was strikingly increased in individuals with *2/*2 genotype when compared with *1/*1 (P=0.000038) or *1/*2 (P=0.0085) genotypes. Using the two Asian cohorts, significant association of FCGR2B-232Thr/Thr with SLE was observed only in the presence of CD72-*1/*1 genotype (OR 4.63, 95% CI 1.47-14.6, P=0.009 versus FCGR2B-232Ile/Ile plus CD72-*2/*2). Minigene assays demonstrated that the 13-nucleotide repeat and 4 bp deletion within the same haplotype of intron 8 could regulate alternative splicing. The AS isoform lacks exon 8, and is deduced to contain 49 amino acid changes in the membrane-distal portion of the extracellular domain, where considerable amino acid changes are known in CD72(c) allele associated with murine SLE. These results indicated that the presence of CD72-*2 allele decreases risk for human SLE conferred by FCGR2B-232Thr, possibly by increasing the AS isoform of CD72.

Genetic predisposition has been firmly established as a key element in susceptibility to systemic lupus erythematosus (SLE). During the past three decades, association studies have assessed many genes for potential roles in predisposing to SLE. These studies have identified a few risk factors including hereditary deficiency of complement components, major histocompatibility complex class II alleles, and allelic variants for the Fc portion of IgG (FCGR) genes. In recent years, a few groups have completed linkage analyses in data sets from families containing multiple members affected with SLE. Results from these initial genome scans are encouraging; approximately eight chromosomal regions have been identified exhibiting evidence for significant linkage to SLE and have been confirmed using independent cohorts (1q23, 1q25-31, 1q41-42, 2q35-37, 4p16-15.2, 6p11-21, 12q24, and 16q12), suggesting the high likelihood of the presence of one or multiple SLE susceptibility genes at each locus. Another approach of linkage analyses conditioned on pedigrees where one affected member manifesting a particular clinical condition has also identified many chromosomal regions linked to SLE. Within several established susceptibility loci, evidence for association of positional candidate genes is emerging. Within 2q35-37, an intronic single nucleotide polymorphism (SNP) of the positional candidate gene program cell death 1 gene has been associated with SLE susceptibility. The SLE-associated SNP affects a transcription factor, RUNX1, binding site. Recently, SNPs of novel positional candidate genes that influence RUNX1 binding motifs have also been associated with other autoimmune diseases, suggesting the possibility of a common theme shared among susceptibility genes for autoimmune diseases. In the coming years, susceptibility genes responsible for the observed linkage will be identified, and will lead to further delineating genetic pathways involved in susceptibility to SLE.

Susceptibility to systemic lupus erythematosus (SLE) has a genetic component. In recent years, nine complete genome scans using family collections that differ greatly in ethnic compositions and geographic locations have identified several strong, confirmed SLE susceptibility loci. Evidence implicating individual gene polymorphisms (or haplotypes) within some of the linked intervals has been reported. This review highlights recent findings that may lead to the identification of putative genes and new insights in the pathogenesis of SLE.Eight of the best-supported SLE susceptibility loci are 1q23, 1q25-31, 1q41-42, 2q35-37, 4p16-15.2, 6p11-21, 12p24, and 16q12. These are chromosomal regions exhibiting genome-wide significance for linkage in single studies and suggestive evidence for linkage in other samples. Linkage analyses conditioning on pedigrees in which one affected member manifesting a particular clinical condition have also yielded many chromosomal regions linked to SLE. The linked interval on chromosome 6p has been narrowed to 0.5 approximately 1.0 Mb (million basepairs) of 3 MHC class II containing risk haplotypes in white subjects. Cumulative results have shown that hereditary deficiencies of complement component C4A (a MHC class III gene) confer risk for SLE in almost all ethnic groups studied. The FcgammaR genes (located at 1q23) have been convincingly demonstrated to play an important role in susceptibility to SLE (and/or lupus nephritis). The evidence for the intronic single nucleotide polymorphism of program cell death gene 1 (PDCD1 at 2q37) to confer susceptibility is promising but not yet compelling. Within several established susceptibility loci, evidence for association of positional candidate genes is emerging.Further replications of linkage and association are the immediate task. The respective contribution of each susceptibility gene, relationships between genotypes and phenotypes, and potential interactions between susceptibility gene products need to be elucidated. This line of investigation is now well poised to provide novel insights into how genetic variants can affect functional pathways leading to the development of SLE.

Systemic lupus erythematosus (SLE) is a complex, multifactorial autoimmune disease. Genetic factors are thought to contribute to its pathogenesis. There have been numerous recent advances in the study of murine and human lupus genetics. In well-defined experimental transgenic or gene-knockout mouse models, the development of lupus-like disease has implicated specific genes and pathways in the disease pathogenesis. Linkage analyses have mapped multiple susceptibility loci and disease suppressive loci using inbred strains of mice that spontaneously develop lupus-like disease. Elegant genetic dissection and function studies have led to the recent identification of two murine candidate susceptibility genes, Ifi202 (encoding an interferon-inducible protein) and Cr2 (encoding complement receptors 1 and 2). In human lupus, case- control studies have established associations of SLE with certain major histocompatibility class II alleles, complement deficiencies, and polymorphisms of Fc gamma receptor genes, a complement-related gene, and cytokine genes. During the past several years, linkage analyses using SLE multiplex families have provided many chromosomal regions for further exploration of susceptibility genes. Six regions exhibiting significant linkage to SLE are promising. Studies are underway to fine map these linked regions and to identify the genes in the susceptibility regions. An understanding of the genes involved in the development of lupus should provide targets for more focused therapy in lupus.

Genome-wide association studies have identified more than 50 robust loci associated with systemic lupus erythematosus (SLE) susceptibility, and follow-up studies help reveal candidate causative genetic variants and their biological relevance contributing to the development of SLE. Epigenetic modulation is emerging as an important mechanism for understanding how the implicated genes interact with environmental factors. We review recent progress toward identifying causative variants of SLE-associated loci and epigenetic impact on lupus, especially genetic-epigenetic interactions that modulate expression levels of SLE susceptibility genes.A few SLE-risk loci have been refined to localize likely causative variants responsible for the observed genome-wide association study signals. Few of such variants disrupt coding sequences resulting in gain or loss of function for the encoded protein, whereas most fall in noncoding regions with potential to regulate gene expression through alterations in transcriptional activity, splicing, mRNA stability and epigenetic modifications. Multiple key pathways related to the SLE pathogenesis have been indicated by the identified genetic risk factors, including type I interferon signaling pathway that can also be regulated by epigenetic changes occurred in SLE.These findings provide novel insights into the disease pathogenesis and promise better diagnostic accuracy and new therapeutic targets for patient management.

Efforts to identify lupus-associated causal variants in the FAM167A/BLK locus on 8p21 are hampered by highly associated noncausal variants. In this report, we used a trans-population mapping and sequencing strategy to identify a common variant (rs922483) in the proximal BLK promoter and a tri-allelic variant (rs1382568) in the upstream alternative BLK promoter as putative causal variants for association with systemic lupus erythematosus. The risk allele (T) at rs922483 reduced proximal promoter activity and modulated alternative promoter usage. Allelic differences at rs1382568 resulted in altered promoter activity in B progenitor cell lines. Thus, our results demonstrated that both lupus-associated functional variants contribute to the autoimmune disease association by modulating transcription of BLK in B cells and thus potentially altering immune responses.

Systemic lupus erythematosus (SLE) is a complex, multifactorial autoimmune disease. Genetic factors are believed to contribute to its pathogenesis. There have been numerous recent advances in the study of murine and human lupus genetics. In well-defined, experimental, transgenic or gene-knockout mouse models, the development of lupus-like disease has implicated specific genes and pathways in the disease pathogenesis. Linkage analyses have mapped multiple susceptibility loci and disease suppressive loci using inbred strains of mice that spontaneously develop lupus-like disease. Elegant genetic dissection has demonstrated that a component phenotype of SLE is displayed by each congenic strain carrying a single susceptibility locus on a resistant genetic background, whereas polycongenic strains exhibit fatal lupus nephritis. These studies suggest that genes in separate pathways can interact to augment or suppress the initiation and progression of systemic autoimmunity. In association studies of human lupus, the contributions of the MHC loci, Fcg receptors, various cytokines, components of the complement cascade, and proteins involved in apoptosis have been explored. Most recently, linkage analyses have been performed and provide many chromosomal regions for further exploration for susceptibility genes. Studies to identify the genes in the susceptibility regions are underway. An understanding of the genes involved in the development of lupus should provide targets for more focused therapy in lupus.

Objective Signaling molecules from the T cell receptor zeta/Fcepsilon receptor gamma (TCRzeta/FcRgamma) family play a critical role in the function of Fcgamma receptors and the TCR and are located on human chromosome 1, where lupus susceptibility genes are located. This study was undertaken to investigate the possibility of polymorphisms and/or mutations of TCRzeta in systemic lupus erythematosus (SLE). Methods We amplified the whole coding region of TCRzeta by reverse transcriptase-polymerase chain reaction (PCR) and directly sequenced the PCR products with a dye primer technique to facilitate heterozygote detection. Results An alternative splicing form of TCRzeta, with a CAG codon (glutamine) inserted at the splice junction of exons 4 and 5, was found both in SLE and in non-SLE subjects. Both splice isoforms of TCRzeta occurred in human mixed peripheral blood mononuclear cells, natural killer cells, and Jurkat T cells. In TCRzeta, 2 silent and 2 missense mutations were found, but neither coding change occurred in the immunoreceptor tyrosine-activation motif. No unique mutations were found in Caucasian, African American, Hispanic, Chinese, or Japanese SLE patients living in North America. Conclusion The uncommon and equal occurrence of novel single-nucleotide polymorphisms in both SLE patients and normal subjects makes it improbable that they play important roles in genetic susceptibility to SLE.

Although cumulative evidence suggests that a genetic predisposition plays a major role in development of systemic lupus erythematosus (SLE) and/or lupus nephritis (LN), the susceptibility genes are mostly unknown. The difficulty in identifying susceptibility genes is due in part to multiple genes with variable genetic effects and the diverse genetic backgrounds of human populations. In human SLE, genes of early components of complements as well as many polymorphic genes (including the MHC class II and class III, FcγR, mannose-binding protein, IL-6, Bcl-2, and IL-10 genes) have been associated with SLE or LN by population-based case-control or within-case studies. The contribution of some of these disease-associated genes to the presence or absence of clinical manifestations has been further tested in mice with targeted disruption of the specific candidate gene. In addition to SLE susceptibility genes, there may be a separate set of nephropathy susceptibility genes predisposing to LN as suggested by the familial clustering of endstage renal disease in African-Americans with LN. The availability of densely mapped genetic markers spanning the entire genome has enabled the identification of chromosomal regions linked to disease susceptibility genes without prior knowledge of the gene function. Our group has used known murine lupus susceptibility loci as a guide, and conducted linkage analysis of genetic markers located within a specific, possibly syntenic human chromosomal region. Evidence for linkage of a chromosome 1q41-42 region was observed in SLE-affected sib pairs from multiple ethnic groups. More recently, several groups have reported results of genome scans of SLE-affected sib pairs or pedigrees. These exciting recent developments in delineating the genetic basis of SLE or LN are summarized in this review.

Protein tyrosine phosphatase non-receptor type 22 (PTPN22) is a negative regulator of T-cell activation associated with several autoimmune diseases, including systemic lupus erythematosus (SLE). Missense rs2476601 is associated with SLE in individuals with European ancestry. Since the rs2476601 risk allele frequency differs dramatically across ethnicities, we assessed robustness of PTPN22 association with SLE and its clinical sub-phenotypes across four ethnically diverse populations. Ten SNPs were genotyped in 8220 SLE cases and 7369 controls from in European-Americans (EA), African-Americans (AA), Asians (AS), and Hispanics (HS). We performed imputation-based association followed by conditional analysis to identify independent associations. Significantly associated SNPs were tested for association with SLE clinical sub-phenotypes, including autoantibody profiles. Multiple testing was accounted for by using false discovery rate. We successfully imputed and tested allelic association for 107 SNPs within the PTPN22 region and detected evidence of ethnic-specific associations from EA and HS. In EA, the strongest association was at rs2476601 (P = 4.7 × 10(-9), OR = 1.40 (95% CI = 1.25-1.56)). Independent association with rs1217414 was also observed in EA, and both SNPs are correlated with increased European ancestry. For HS imputed intronic SNP, rs3765598, predicted to be a cis-eQTL, was associated (P = 0.007, OR = 0.79 and 95% CI = 0.67-0.94). No significant associations were observed in AA or AS. Case-only analysis using lupus-related clinical criteria revealed differences between EA SLE patients positive for moderate to high titers of IgG anti-cardiolipin (aCL IgG >20) versus negative aCL IgG at rs2476601 (P = 0.012, OR = 1.65). Association was reinforced when these cases were compared to controls (P = 2.7 × 10(-5), OR = 2.11). Our results validate that rs2476601 is the most significantly associated SNP in individuals with European ancestry. Additionally, rs1217414 and rs3765598 may be associated with SLE. Further studies are required to confirm the involvement of rs2476601 with aCL IgG.

Systemic lupus erythematosus (SLE) is an inflammatory autoimmune disease with a strong genetic component. African-Americans (AA) are at increased risk of SLE, but the genetic basis of this risk is largely unknown. To identify causal variants in SLE loci in AA, we performed admixture mapping followed by fine mapping in AA and European-Americans (EA). Through genome-wide admixture mapping in AA, we identified a strong SLE susceptibility locus at 2q22–24 (LOD = 6.28), and the admixture signal is associated with the European ancestry (ancestry risk ratio ∼1.5). Large-scale genotypic analysis on 19,726 individuals of African and European ancestry revealed three independently associated variants in the IFIH1 gene: an intronic variant, rs13023380 [Pmeta = 5.20×10−14; odds ratio, 95% confidence interval = 0.82 (0.78–0.87)], and two missense variants, rs1990760 (Ala946Thr) [Pmeta = 3.08×10−7; 0.88 (0.84–0.93)] and rs10930046 (Arg460His) [Pdom = 1.16×10−8; 0.70 (0.62–0.79)]. Both missense variants produced dramatic phenotypic changes in apoptosis and inflammation-related gene expression. We experimentally validated function of the intronic SNP by DNA electrophoresis, protein identification, and in vitro protein binding assays. DNA carrying the intronic risk allele rs13023380 showed reduced binding efficiency to a cellular protein complex including nucleolin and lupus autoantigen Ku70/80, and showed reduced transcriptional activity in vivo. Thus, in SLE patients, genetic susceptibility could create a biochemical imbalance that dysregulates nucleolin, Ku70/80, or other nucleic acid regulatory proteins. This could promote antibody hypermutation and auto-antibody generation, further destabilizing the cellular network. Together with molecular modeling, our results establish a distinct role for IFIH1 in apoptosis, inflammation, and autoantibody production, and explain the molecular basis of these three risk alleles for SLE pathogenesis., Author Summary African-Americans (AA) are at increased risk of systemic lupus erythematosus (SLE), but the genetic basis of this risk increase is largely unknown. We used admixture mapping to localize disease-causing genetic variants that differ in frequency across populations. This approach is advantageous for localizing susceptibility genes in recently admixed populations like AA. Our genome-wide admixture scan identified seven admixture signals, and we followed the best signal at 2q22–24 with fine-mapping, imputation-based association analysis and experimental validation. We identified two independent coding variants and a non-coding variant within the IFIH1 gene associated with SLE. Together with molecular modeling, our results establish a distinct role for IFIH1 in apoptosis, inflammation, and autoantibody production, and explain the molecular basis of these three risk alleles for SLE pathogenesis.

We previously established an 80 kb haplotype upstream of TNFSF4 as a susceptibility locus in the autoimmune disease SLE. SLE-associated alleles at this locus are associated with inflammatory disorders, including atherosclerosis and ischaemic stroke. In Europeans, the TNFSF4 causal variants have remained elusive due to strong linkage disequilibrium exhibited by alleles spanning the region. Using a trans-ancestral approach to fine-map the locus, utilising 17,900 SLE and control subjects including Amerindian/Hispanics (1348 cases, 717 controls), African-Americans (AA) (1529, 2048) and better powered cohorts of Europeans and East Asians, we find strong association of risk alleles in all ethnicities; the AA association replicates in African-American Gullah (152,122). The best evidence of association comes from two adjacent markers: rs2205960-T (P = 1.71×10-34, OR = 1.43[1.26-1.60]) and rs1234317-T (P = 1.16×10-28, OR = 1.38[1.24-1.54]). Inference of fine-scale recombination rates for all populations tested finds the 80 kb risk and non-risk haplotypes in all except African-Americans. In this population the decay of recombination equates to an 11 kb risk haplotype, anchored in the 5′ region proximal to TNFSF4 and tagged by rs2205960-T after 1000 Genomes phase 1 (v3) imputation. Conditional regression analyses delineate the 5′ risk signal to rs2205960-T and the independent non-risk signal to rs1234314-C. Our case-only and SLE-control cohorts demonstrate robust association of rs2205960-T with autoantibody production. The rs2205960-T is predicted to form part of a decameric motif which binds NF-κBp65 with increased affinity compared to rs2205960-G. ChIP-seq data also indicate NF-κB interaction with the DNA sequence at this position in LCL cells. Our research suggests association of rs2205960-T with SLE across multiple groups and an independent non-risk signal at rs1234314-C. rs2205960-T is associated with autoantibody production and lymphopenia. Our data confirm a global signal at TNFSF4 and a role for the expressed product at multiple stages of lymphocyte dysregulation during SLE pathogenesis. We confirm the validity of trans-ancestral mapping in a complex trait. © 2013 Manku et al.

Many identified genetic risk factors for systemic lupus erythematosus (SLE) contribute to the function of the immune system, which has expanded our understanding of disease pathogenesis. We outline the genetic variants in recently identified SLE-associated loci, the immunological pathways affected by these gene products and the disease manifestations linked to these loci. Pathways potentially influenced by SLE risk variants include: apoptosis, DNA degradation and clearance of cellular debris; antigen presentation; type I interferon, Toll-like receptor and nuclear factor kappa κB activation; defective clearance of immune complexes containing nuclear antigens; B and T-cell function and signalling; and monocyte and neutrophil function and signalling. These identified SLE susceptibility loci are predominantly common variants that have been confirmed among multiple ancestries, suggesting shared mechanisms in disease aetiology. Ongoing genetic studies continue the investigation of specific functional variants, and their potential consequences on immune dysregulation, enhancing our understanding of links between genotypes and specific disease manifestations. The next generation of sequencing explores the identification of causal rare variants that may contribute robust genetic effects to developing SLE. Novel insights coming from genetic studies of SLE provide the opportunity to elucidate pathogenic mechanisms as well as contribute to the development of innovative therapeutic targets for this complex disease.

We previously reported that the G allele of rs3853839 at 3′untranslated region (UTR) of Toll-like receptor 7 (TLR7) was associated with elevated transcript expression and increased risk for systemic lupus erythematosus (SLE) in 9,274 Eastern Asians [P = 6.5×10−10, odds ratio (OR) (95%CI) = 1.27 (1.17–1.36)]. Here, we conducted trans-ancestral fine-mapping in 13,339 subjects including European Americans, African Americans, and Amerindian/Hispanics and confirmed rs3853839 as the only variant within the TLR7-TLR8 region exhibiting consistent and independent association with SLE (P meta = 7.5×10−11, OR = 1.24 [1.18–1.34]). The risk G allele was associated with significantly increased levels of TLR7 mRNA and protein in peripheral blood mononuclear cells (PBMCs) and elevated luciferase activity of reporter gene in transfected cells. TLR7 3′UTR sequence bearing the non-risk C allele of rs3853839 matches a predicted binding site of microRNA-3148 (miR-3148), suggesting that this microRNA may regulate TLR7 expression. Indeed, miR-3148 levels were inversely correlated with TLR7 transcript levels in PBMCs from SLE patients and controls (R2 = 0.255, P = 0.001). Overexpression of miR-3148 in HEK-293 cells led to significant dose-dependent decrease in luciferase activity for construct driven by TLR7 3′UTR segment bearing the C allele (P = 0.0003). Compared with the G-allele construct, the C-allele construct showed greater than two-fold reduction of luciferase activity in the presence of miR-3148. Reduced modulation by miR-3148 conferred slower degradation of the risk G-allele containing TLR7 transcripts, resulting in elevated levels of gene products. These data establish rs3853839 of TLR7 as a shared risk variant of SLE in 22,613 subjects of Asian, EA, AA, and Amerindian/Hispanic ancestries (Pmeta = 2.0×10−19, OR = 1.25 [1.20–1.32]), which confers allelic effect on transcript turnover via differential binding to the epigenetic factor miR-3148., Author Summary Systemic lupus erythematosus (SLE) is a debilitating autoimmune disease contributed to by excessive innate immune activation involving toll-like receptors (TLRs, particularly TLR7/8/9) and type I interferon (IFN) signaling pathways. TLR7 responds against RNA–containing nuclear antigens and activates IFN-α pathway, playing a pivotal role in the development of SLE. While a genomic duplication of Tlr7 promotes lupus-like disease in the Y-linked autoimmune accelerator (Yaa) murine model, the lack of common copy number variations at TLR7 in humans led us to identify a functional single nucleotide polymorphism (SNP), rs3853839 at 3′ UTR of the TLR7 gene, associated with SLE susceptibility in Eastern Asians. In this study, we fine-mapped the TLR7-TLR8 region and confirmed rs3853839 exhibiting the strongest association with SLE in European Americans, African Americans, and Amerindian/Hispanics. Individuals carrying the risk G allele of rs3853839 exhibited increased TLR7 expression at the both mRNA and protein level and decreased transcript degradation. MicroRNA-3148 (miR-3148) downregulated the expression of non-risk allele (C) containing transcripts preferentially, suggesting a likely mechanism for increased TLR7 levels in risk-allele carriers. This trans-ancestral mapping provides evidence for the global association with SLE risk at rs3853839, which resides in a microRNA–gene regulatory site affecting TLR7 expression.

Objectives The Xq28 region containing IRAK1 and MECP2 has been identified as a risk locus for systemic lupus erythematosus (SLE) in previous genetic association studies. However, due to the strong linkage disequilibrium between IRAK1 and MECP2 , it remains unclear which gene is affected by the underlying causal variant(s) conferring risk of SLE. Methods We fine-mapped ≥136 SNPs in a ∼227 kb region on Xq28, containing IRAK1, MECP2 and seven adjacent genes ( L1CAM, AVPR2, ARHGAP4, NAA10, RENBP, HCFC1 and TMEM187 ), for association with SLE in 15 783 case-control subjects derived from four different ancestral groups. Results Multiple SNPs showed strong association with SLE in European Americans, Asians and Hispanics at p −8 with consistent association in subjects with African ancestry. Of these, six SNPs located in the TMEM187-IRAK1-MECP2 region captured the underlying causal variant(s) residing in a common risk haplotype shared by all four ancestral groups. Among them, rs1059702 best explained the Xq28 association signals in conditional testings and exhibited the strongest p value in transancestral meta-analysis (p meta = 1.3×10 −27 , OR=1.43), and thus was considered to be the most likely causal variant. The risk allele of rs1059702 results in the amino acid substitution S196F in IRAK1 and had previously been shown to increase NF-κB activity in vitro. We also found that the homozygous risk genotype of rs1059702 was associated with lower mRNA levels of MECP2 , but not IRAK1 , in SLE patients (p=0.0012) and healthy controls (p=0.0064). Conclusions These data suggest contributions of both IRAK1 and MECP2 to SLE susceptibility.

Systemic lupus erythematosus (SLE) is an autoimmune disease with diverse clinical manifestations characterized by the development of pathogenic autoantibodies manifesting in inflammation of target organs such as the kidneys, skin and joints. Genome-wide association studies have identified genetic variants in the UBE2L3 region that are associated with SLE in subjects of European and Asian ancestry. UBE2L3 encodes an ubiquitin-conjugating enzyme, UBCH7, involved in cell proliferation and immune function. In this study, we sought to further characterize the genetic association in the region of UBE2L3 and use molecular methods to determine the functional effect of the risk haplotype. We identified significant associations between variants in the region of UBE2L3 and SLE in individuals of European and Asian ancestry that exceeded a Bonferroni-corrected threshold (P

ObjectiveSystemic lupus erythematosus (SLE; OMIM 152700) is a chronic autoimmune disease for which the aetiology includes genetic and environmental factors. ITGAM, integrin αM (complement component 3 receptor 3 subunit) encoding a ligand for intracellular adhesion molecule (ICAM) proteins, is an established SLE susceptibility locus. This study aimed to evaluate the independent and joint effects of genetic variations in the genes that encode ITGAM and ICAM.MethodsThe authors examined several markers in the ICAM1–ICAM4–ICAM5 locus on chromosome 19p13 and the single ITGAM polymorphism (rs1143679) using a large-scale case–control study of 17 481 unrelated participants from four ancestry populations. The single-marker association and gene–gene interaction were analysed for each ancestry, and a meta-analysis across the four ancestries was performed.ResultsThe A-allele of ICAM1–ICAM4–ICAM5 rs3093030, associated with elevated plasma levels of soluble ICAM1, and the A-allele of ITGAM rs1143679 showed the strongest association with increased SLE susceptibility in each of the ancestry populations and the trans-ancestry meta-analysis (ORmeta=1.16, 95% CI 1.11 to 1.22; p=4.88×10−10 and ORmeta=1.67, 95% CI 1.55 to 1.79; p=3.32×10−46, respectively). The effect of the ICAM single-nucleotide polymorphisms (SNPs) was independent of the effect of the ITGAM SNP rs1143679, and carriers of both ICAM rs3093030-AA and ITGAM rs1143679-AA had an OR of 4.08 compared with those with no risk allele in either SNP (95% CI 2.09 to 7.98; p=3.91×10−5).ConclusionThese findings are the first to suggest that an ICAM–integrin-mediated pathway contributes to susceptibility to SLE.

Objective Systemic lupus erythematosus (SLE) is an autoimmune disease characterized by autoantibody production and altered type I interferon expression. Genetic surveys and genome-wide association studies have identified >30 SLE susceptibility genes. One of these genes, TNIP1, encodes the ABIN1 protein. ABIN1 functions in the immune system by restricting NF-?B signaling. The present study was undertaken to investigate the genetic factors that influence association with SLE in genes that regulate the NF-?B pathway. Methods We analyzed a dense set of genetic markers spanning TNIP1 and TAX1BP1, as well as the TNIP1 homolog TNIP2, in case-control populations of diverse ethnic origins. TNIP1, TNIP2, and TAX1BP1 were fine-mapped in a total of 8,372 SLE cases and 7,492 healthy controls from European-ancestry, African American, Hispanic, East Asian, and African American Gullah populations. Levels of TNIP1 messenger RNA (mRNA) and ABIN1 protein in Epstein-Barr virus-transformed human B cell lines were analyzed by quantitative reverse transcription-polymerase chain reaction and Western blotting, respectively. Results We found significant associations between SLE and genetic variants within TNIP1, but not in TNIP2 or TAX1BP1. After resequencing and imputation, we identified 2 independent risk haplotypes within TNIP1 in individuals of European ancestry that were also present in African American and Hispanic populations. Levels of TNIP1 mRNA and ABIN1 protein were reduced among subjects with these haplotypes, suggesting that they harbor hypomorphic functional variants that influence susceptibility to SLE by restricting ABIN1 expression. Conclusion Our results confirm the association signals between SLE and TNIP1 variants in multiple populations and provide new insight into the mechanism by which TNIP1 variants may contribute to SLE pathogenesis. Copyright © 2012 by the American College of Rheumatology.

Systemic lupus erythematosus (SLE) is a complex autoimmune disease characterized by autoantibody production and organ damage. Lupus nephritis (LN) is one of the most severe manifestations of SLE. Multiple studies reported associations between renal diseases and variants in the non-muscle myosin heavy chain 9 (MYH9) and the neighboring apolipoprotein L 1 (APOL1) genes. We evaluated 167 variants spanning MYH9 for association with LN in a multiethnic sample. The two previously identified risk variants in APOL1 were also tested for association with LN in European-Americans (EAs) (N=579) and African-Americans (AAs) (N=407). Multiple peaks of association exceeding a Bonferroni corrected P-value of P

In spite of the well-known clustering of multiple autoimmune disorders in families, analyses of specific shared genes and polymorphisms between systemic lupus erythematosus (SLE) and other autoimmune diseases (ADs) have been limited. Therefore, we comprehensively tested autoimmune variants for association with SLE, aiming to identify pleiotropic genetic associations between these diseases. We compiled a list of 446 non–Major Histocompatibility Complex (MHC) variants identified in genome-wide association studies (GWAS) of populations of European ancestry across 17 ADs. We then tested these variants in our combined Caucasian SLE cohorts of 1,500 cases and 5,706 controls. We tested a subset of these polymorphisms in an independent Caucasian replication cohort of 2,085 SLE cases and 2,854 controls, allowing the computation of a meta-analysis between all cohorts. We have uncovered novel shared SLE loci that passed multiple comparisons adjustment, including the VTCN1 (rs12046117, P = 2.02×10−06) region. We observed that the loci shared among the most ADs include IL23R, OLIG3/TNFAIP3, and IL2RA. Given the lack of a universal autoimmune risk locus outside of the MHC and variable specificities for different diseases, our data suggests partial pleiotropy among ADs. Hierarchical clustering of ADs suggested that the most genetically related ADs appear to be type 1 diabetes with rheumatoid arthritis and Crohn's disease with ulcerative colitis. These findings support a relatively distinct genetic susceptibility for SLE. For many of the shared GWAS autoimmune loci, we found no evidence for association with SLE, including IL23R. Also, several established SLE loci are apparently not associated with other ADs, including the ITGAM-ITGAX and TNFSF4 regions. This study represents the most comprehensive evaluation of shared autoimmune loci to date, supports a relatively distinct non–MHC genetic susceptibility for SLE, provides further evidence for previously and newly identified shared genes in SLE, and highlights the value of studies of potentially pleiotropic genes in autoimmune diseases., Author Summary It is well known that multiple autoimmune disorders cluster in families. However, all of the genetic variants that explain this clustering have not been discovered, and the specific genetic variants shared between systemic lupus erythematosus (SLE) and other autoimmune diseases (ADs) are not known. In order to better understand the genetic factors that explain this predisposition to autoimmunity, we performed a comprehensive evaluation of shared autoimmune genetic variants. First we considered results from 17 ADs and compiled a list with 446 significant genetic variants from these studies. We identified some genetic variants extensively shared between ADs, as well as the ADs that share the most variants. The genetic overlap between SLE and other ADs was modest. Next we tested how important all the 446 genetic variants were in our collection with a minimum of 1,500 SLE patients. Among the most significant variants in SLE, the majority had already been identified in previous studies, but we also discovered variants in two important immune genes. In summary, our data identified diseases with common genetic risk factors and novel SLE effects, and this supports a relatively distinct genetic susceptibility for SLE. This study helps delineate the genetic architecture of ADs.

Objective Several confirmed genetic susceptibility loci for lupus have been described. To date, no clear evidence for genetic epistasis in lupus has been established. The aim of this study was to test for gene–gene interactions in a number of known lupus susceptibility loci. Methods Eighteen single-nucleotide polymorphisms tagging independent and confirmed lupus susceptibility loci were genotyped in a set of 4,248 patients with lupus and 3,818 normal healthy control subjects of European descent. Epistasis was tested by a 2-step approach using both parametric and nonparametric methods. The false discovery rate (FDR) method was used to correct for multiple testing. Results We detected and confirmed gene–gene interactions between the HLA region and CTLA4, IRF5, and ITGAM and between PDCD1 and IL21 in patients with lupus. The most significant interaction detected by parametric analysis was between rs3131379 in the HLA region and rs231775 in CTLA4 (interaction odds ratio 1.19, Z = 3.95, P = 7.8 × 10−5 [FDR ≤0.05], P for multifactor dimensionality reduction = 5.9 × 10−45). Importantly, our data suggest that in patients with lupus, the presence of the HLA lupus risk alleles in rs1270942 and rs3131379 increases the odds of also carrying the lupus risk allele in IRF5 (rs2070197) by 17% and 16%, respectively (P = 0.0028 and P = 0.0047, respectively). Conclusion We provide evidence for gene–gene epistasis in systemic lupus erythematosus. These findings support a role for genetic interaction contributing to the complexity of lupus heritability.

Objective The overexpression of interferon (IFN)–inducible genes is a prominent feature of systemic lupus erythematosus (SLE); it serves as a marker for active and more severe disease, and is also observed in other autoimmune and inflammatory conditions. This study was undertaken to investigate the genetic variations responsible for sustained activation of IFN-responsive genes in SLE. Methods We systematically evaluated association of SLE with a total of 1,754 IFN pathway–related genes, including IFN-inducible genes known to be differentially expressed in SLE patients and their direct regulators. We used a 3-stage study design in which 2 cohorts (total of 939 SLE cases and 3,398 controls) were analyzed independently and jointly for association with SLE, and the results were adjusted for the number of comparisons. Results A total of 15,166 single-nucleotide polymorphisms (SNPs) passed all quality control filters; 305 of these SNPs demonstrated replicated association with SLE in both cohorts. Nine variants were further genotyped for confirmation in an average of 1,316 independent SLE cases and 3,215 independent controls. Association with SLE was confirmed for several genes, including those for the transmembrane receptor CD44 (CD44 [rs507230]; P = 3.98 × 10−12), the cytokine pleiotrophin (PTN [rs919581]; P = 5.38 × 10−4), the heat-shock protein DnaJ (DNAJA1 [rs10971259]; P = 6.31 × 10−3), and the nuclear import protein karyopherin α1 (KPNA [rs6810306]; P = 4.91 × 10−2). Conclusion This study expands the number of candidate genes that have been shown to be associated with SLE and highlights potential of pathway-based approaches for gene discovery. Identification of the causal alleles will help elucidate the molecular mechanisms responsible for activation of the IFN system in SLE.