RESUMO
Polyamines are molecules with multiple amino groups that are essential for cellular function. The major polyamines are putrescine, spermidine, spermine, and cadaverine. Polyamines are important for posttranscriptional regulation, autophagy, programmed cell death, proliferation, redox homeostasis, and ion channel function. Their levels are tightly controlled. High levels of polyamines are associated with proliferative pathologies such as cancer, whereas low polyamine levels are observed in aging, and elevated polyamine turnover enhances oxidative stress. Polyamine metabolism is implicated in several pathophysiological processes in the nervous, immune, and cardiovascular systems. Currently, manipulating polyamine levels is under investigation as a potential preventive treatment for several pathologies, including aging, ischemia/reperfusion injury, pulmonary hypertension, and cancer. Although polyamines have been implicated in many intracellular mechanisms, our understanding of these processes remains incomplete and is a topic of ongoing investigation. Here, we discuss the regulation and cellular functions of polyamines, their role in physiology and pathology, and emphasize the current gaps in knowledge and potential future research directions.
Assuntos
Poliaminas , Humanos , Animais , Poliaminas/metabolismo , Estresse Oxidativo/fisiologia , Homeostase/fisiologia , Autofagia/fisiologiaRESUMO
In order to develop prediction models of one-year treatment response in lupus nephritis, an approach using machine learning to combine traditional clinical data and novel urine biomarkers was undertaken. Contemporary lupus nephritis biomarkers were identified through an unbiased PubMed search. Thirteen novel urine proteins contributed to the top 50% of ranked biomarkers and were selected for measurement at the time of lupus nephritis flare. These novel markers along with traditional clinical data were incorporated into a variety of machine learning algorithms to develop prediction models of one-year proteinuria and estimated glomerular filtration rate (eGFR). Models were trained on 246 individuals from four different sub-cohorts and validated on an independent cohort of 30 patients with lupus nephritis. Seven models were considered for each outcome. Three-quarters of these models demonstrated good predictive value with areas under the receiver operating characteristic curve over 0.7. Overall, prediction performance was the best for models of eGFR response to treatment. Furthermore, the best performing models contained both traditional clinical data and novel urine biomarkers, including cytokines, chemokines, and markers of kidney damage. Thus, our study provides further evidence that a machine learning approach can predict lupus nephritis outcomes at one year using a set of traditional and novel biomarkers. However, further validation of the utility of machine learning as a clinical decision aid to improve outcomes will be necessary before it can be routinely used in clinical practice to guide therapy.
Assuntos
Lúpus Eritematoso Sistêmico , Nefrite Lúpica , Biomarcadores , Humanos , Nefrite Lúpica/diagnóstico , Nefrite Lúpica/tratamento farmacológico , Proteinúria/etiologia , Curva ROC , Exacerbação dos SintomasRESUMO
OBJECTIVES: We previously identified a hypomorphic variant, p.Arg90His (p.R90H) of neutrophil cytosolic factor 1 (NCF1, a regulatory subunit of phagocyte nicotinamide adenine dinucleotide phosphate (NADPH) oxidase 2 complex), as an putative causal variant for systemic lupus erythematosus (SLE), and established a knock-in (KI) H90 variant in the C57BL/6 background to study how this variant promotes lupus development. METHODS: Wild type (WT) and KI littermates were assessed for immune profiles and lupus-like features. Disease activity and renal damage of patients with SLE were assessed by systemic lupus erythematosus disease activity index (SLEDAI) and renal items of systemic lupus international collaborating clinics (SLICC), respectively. RESULTS: Compared with WT littermates, 5-week-old homozygous KI mice had reduced oxidative burst, splenomegaly, elevated type I interferon (IFN-I) scores, increased ratios of splenic follicular T helper 2 (Tfh2) to either T follicular regulatory (Tfr) or Tfh1 cells, increased ANA+ follicular, germinal centre and plasma cells without spontaneous kidney disease up to 1 year of age. Pristane treatment exacerbated the immune dysregulation and induced IFN-I-dependent kidney disease in 36-week-old H90 KI female mice. Decreased efferocytosis of macrophages derived from KI mice and patients with homozygous H90 SLE promoted elevated ratios of Tfh2/Tfr and Tfh2/Tfh1 as well as dysregulated humoral responses due to reduced voltage-gated proton channel 1 (Hv1)-dependent acidification of phagosome pH to neutralise the decreased electrogenic effect of the H90 variant, resulting in impaired maturation and phagosome proteolysis, and increased autoantibody production and kidney damage in mice and patients with SLE of multiple ancestries. CONCLUSIONS: A lupus causal variant, NCF1-H90, reduces macrophage efferocytosis, enhances Tfh2 responses and promotes autoantibody production and kidney damage in both mice and patients with SLE.
Assuntos
Nefropatias/imunologia , Lúpus Eritematoso Sistêmico/imunologia , Macrófagos/imunologia , NADPH Oxidases/genética , Células T Auxiliares Foliculares/imunologia , Animais , Autoanticorpos/imunologia , Técnicas de Introdução de Genes , Humanos , Nefropatias/genética , Lúpus Eritematoso Sistêmico/genética , Lúpus Eritematoso Sistêmico/patologia , Macrófagos/patologia , Camundongos , Camundongos Endogâmicos C57BL , Polimorfismo de Nucleotídeo ÚnicoRESUMO
OBJECTIVES: Families that contain multiple siblings affected with childhood onset of systemic lupus erythematosus (SLE) likely have strong genetic predispositions. We performed whole exome sequencing (WES) to identify familial rare risk variants and to assess their effects in lupus. METHODS: Sanger sequencing validated the two ultra-rare, predicted pathogenic risk variants discovered by WES and identified additional variants in 562 additional patients with SLE. Effects of a splice site variant and a frameshift variant were assessed using a Minigene assay and CRISPR/Cas9-mediated knock-in (KI) mice, respectively. RESULTS: The two familial ultra-rare, predicted loss-of-function (LOF) SAT1 variants exhibited X-linked recessive Mendelian inheritance in two unrelated African-American families. Each LOF variant was transmitted from the heterozygous unaffected mother to her two sons with childhood-onset SLE. The p.Asp40Tyr variant affected a splice donor site causing deleterious transcripts. The young hemizygous male and homozygous female Sat1 p.Glu92Leufs*6 KI mice spontaneously developed splenomegaly, enlarged glomeruli with leucocyte infiltration, proteinuria and elevated expression of type I interferon-inducible genes. SAT1 is highly expressed in neutrophils and encodes spermidine/spermine-N1-acetyltransferase 1 (SSAT1), a rate-limiting enzyme in polyamine catabolism. Young male KI mice exhibited neutrophil defects and decreased proportions of Foxp3 +CD4+ T-cell subsets. Circulating neutrophil counts and proportions of Foxp3 +CD4+ T cells correlated with decreased plasma levels of spermine in treatment-naive, incipient SLE patients. CONCLUSIONS: We identified two novel SAT1 LOF variants, showed the ability of the frameshift variant to confer murine lupus, highlighted the pathogenic role of dysregulated polyamine catabolism and identified SAT1 LOF variants as new monogenic causes for SLE.
Assuntos
Doenças Genéticas Ligadas ao Cromossomo X , Lúpus Eritematoso Sistêmico , Animais , Criança , Feminino , Humanos , Masculino , Camundongos , Predisposição Genética para Doença , Homozigoto , Lúpus Eritematoso Sistêmico/genética , Espermina/sangue , Doenças Genéticas Ligadas ao Cromossomo X/genética , Acetiltransferases/genéticaRESUMO
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.
Assuntos
Sequência de Aminoácidos , Autoanticorpos/imunologia , Suscetibilidade a Doenças , Antígenos de Histocompatibilidade Classe II/química , Antígenos de Histocompatibilidade Classe II/imunologia , Antígenos de Histocompatibilidade Classe I/química , Antígenos de Histocompatibilidade Classe I/imunologia , Lúpus Eritematoso Sistêmico/etiologia , Alelos , Substituição de Aminoácidos , Povo Asiático , Feminino , Predisposição Genética para Doença , Variação Genética , Antígenos de Histocompatibilidade Classe I/genética , Antígenos de Histocompatibilidade Classe II/genética , Humanos , Masculino , Razão de Chances , Polimorfismo de Nucleotídeo ÚnicoRESUMO
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.
Assuntos
Predisposição Genética para Doença , Lúpus Eritematoso Sistêmico/genética , Complexo Principal de Histocompatibilidade/genética , Polimorfismo de Nucleotídeo Único , Negro ou Afro-Americano/genética , Feminino , Estudos de Associação Genética , Haplótipos , Humanos , Masculino , Modelos Genéticos , População Branca/genéticaRESUMO
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.
Assuntos
Alelos , Lúpus Eritematoso Sistêmico/genética , Polimorfismo Genético , Locos de Características Quantitativas , Fator de Transcrição STAT1/genética , Fator de Transcrição STAT4/genética , Feminino , Humanos , Lúpus Eritematoso Sistêmico/epidemiologia , Masculino , Fatores de RiscoRESUMO
Genetic variants at chromosomal region 11q23.3, near the gene ETS1, have been associated with systemic lupus erythematosus (SLE), or lupus, in independent cohorts of Asian ancestry. Several recent studies have implicated ETS1 as a critical driver of immune cell function and differentiation, and mice deficient in ETS1 develop an SLE-like autoimmunity. We performed a fine-mapping study of 14,551 subjects from multi-ancestral cohorts by starting with genotyped variants and imputing to all common variants spanning ETS1. By constructing genetic models via frequentist and Bayesian association methods, we identified 16 variants that are statistically likely to be causal. We functionally assessed each of these variants on the basis of their likelihood of affecting transcription factor binding, miRNA binding, or chromatin state. Of the four variants that we experimentally examined, only rs6590330 differentially binds lysate from B cells. Using mass spectrometry, we found more binding of the transcription factor signal transducer and activator of transcription 1 (STAT1) to DNA near the risk allele of rs6590330 than near the non-risk allele. Immunoblot analysis and chromatin immunoprecipitation of pSTAT1 in B cells heterozygous for rs6590330 confirmed that the risk allele increased binding to the active form of STAT1. Analysis with expression quantitative trait loci indicated that the risk allele of rs6590330 is associated with decreased ETS1 expression in Han Chinese, but not other ancestral cohorts. We propose a model in which the risk allele of rs6590330 is associated with decreased ETS1 expression and increases SLE risk by enhancing the binding of pSTAT1.
Assuntos
Predisposição Genética para Doença , Lúpus Eritematoso Sistêmico/genética , Proteína Proto-Oncogênica c-ets-1/genética , Fator de Transcrição STAT1/genética , Alelos , Animais , Povo Asiático , Teorema de Bayes , Genótipo , Haplótipos , Humanos , Camundongos , Ligação Proteica , Proteína Proto-Oncogênica c-ets-1/metabolismo , Fator de Transcrição STAT1/metabolismoRESUMO
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.
Assuntos
Lúpus Eritematoso Sistêmico/genética , Regiões Promotoras Genéticas , Transcrição Gênica , Quinases da Família src/genética , Alelos , Cromossomos Humanos Par 8 , Ensaio de Desvio de Mobilidade Eletroforética , Feminino , Predisposição Genética para Doença , Haplótipos , Humanos , Masculino , Polimorfismo de Nucleotídeo ÚnicoRESUMO
Objective: The importance of hypomethylation in SLE is well recognized; however, the significance of hypermethylation has not been well characterized. We screened hypermethylated marks in SLE and investigated their possible implications. Methods: DNA methylation marks were screened in SLE whole-blood DNA by microarray, and two marks ( CD3Z and VHL hypermethylations) were confirmed by a methylation single-base extension method in two independent ethnic cohorts consisting of 207 SLE patients and 151 controls. The correlation with clinical manifestations and the genetic influence on those epigenetic marks were analysed. Results: Two epigenetic marks, CD3Z and VHL hypermethylation, were significantly correlated with SLE: CD3Z hypermethylation (odds ratio = 7.76; P = 1.71 × 10 -13 ) and VHL hypermethylation (odds ratio = 3.77; P = 3.20 × 10 -8 ), and the increased CD3Z methylation was correlated with downregulation of the CD3ζ-chain in SLE T cells. In addition, less genetic influence on CD3Z methylation relative to VHL methylation was found in analyses of longitudinal and twin samples. Furthermore, a higher CD3Z methylation level was significantly correlated with a higher SLE disease activity index and more severe clinical manifestations, such as proteinuria, haemolytic anaemia and thrombocytopenia, whereas VHL hypermethylation was not. Conclusion: CD3Z hypermethylation is an SLE risk factor that can be modified by environmental factors and is associated with more severe SLE clinical manifestations, which are related to deranged T cell function by downregulating the CD3ζ-chain.
Assuntos
Complexo CD3/genética , Metilação de DNA/genética , Lúpus Eritematoso Sistêmico/genética , Linfócitos T/metabolismo , Proteína Supressora de Tumor Von Hippel-Lindau/genética , Adulto , Complexo CD3/metabolismo , Estudos de Casos e Controles , Regulação para Baixo , Epigênese Genética , Feminino , Humanos , Lúpus Eritematoso Sistêmico/imunologia , Lúpus Eritematoso Sistêmico/fisiopatologia , Masculino , Pessoa de Meia-Idade , Razão de Chances , República da Coreia , Linfócitos T/imunologia , Estados Unidos , Adulto JovemRESUMO
PURPOSE OF REVIEW: 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. RECENT FINDINGS: 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.
Assuntos
Loci Gênicos , Predisposição Genética para Doença , Lúpus Eritematoso Sistêmico/genética , Polimorfismo de Nucleotídeo Único , Estudo de Associação Genômica Ampla , Humanos , Transdução de Sinais/genéticaRESUMO
OBJECTIVES: Systemic lupus erythematosus (SLE; OMIM 152700) is characterised by the production of antibodies to nuclear antigens. We previously identified variants in complement receptor 2 (CR2/CD21) that were associated with decreased risk of SLE. This study aimed to identify the causal variant for this association. METHODS: Genotyped and imputed genetic variants spanning CR2 were assessed for association with SLE in 15 750 case-control subjects from four ancestral groups. Allele-specific functional effects of associated variants were determined using quantitative real-time PCR, quantitative flow cytometry, electrophoretic mobility shift assay (EMSA) and chromatin immunoprecipitation (ChIP)-PCR. RESULTS: The strongest association signal was detected at rs1876453 in intron 1 of CR2 (pmeta=4.2×10(-4), OR 0.85), specifically when subjects were stratified based on the presence of dsDNA autoantibodies (case-control pmeta=7.6×10(-7), OR 0.71; case-only pmeta=1.9×10(-4), OR 0.75). Although allele-specific effects on B cell CR2 mRNA or protein levels were not identified, levels of complement receptor 1 (CR1/CD35) mRNA and protein were significantly higher on B cells of subjects harbouring the minor allele (p=0.0248 and p=0.0006, respectively). The minor allele altered the formation of several DNA protein complexes by EMSA, including one containing CCCTC-binding factor (CTCF), an effect that was confirmed by ChIP-PCR. CONCLUSIONS: These data suggest that rs1876453 in CR2 has long-range effects on gene regulation that decrease susceptibility to lupus. Since the minor allele at rs1876453 is preferentially associated with reduced risk of the highly specific dsDNA autoantibodies that are present in preclinical, active and severe lupus, understanding its mechanisms will have important therapeutic implications.
Assuntos
Anticorpos Antinucleares/sangue , Lúpus Eritematoso Sistêmico/genética , Receptores de Complemento 3d/genética , Adolescente , Adulto , Subpopulações de Linfócitos B/imunologia , Estudos de Casos e Controles , DNA/imunologia , Predisposição Genética para Doença , Variação Genética , Genótipo , Haplótipos , Humanos , Lúpus Eritematoso Sistêmico/imunologia , Pessoa de Meia-Idade , Fenótipo , Polimorfismo de Nucleotídeo Único , Receptores de Complemento 3b/biossíntese , Medição de Risco/métodos , Fatores de Transcrição/metabolismo , Adulto JovemRESUMO
OBJECTIVES: Following up the systemic lupus erythematosus (SLE) genome-wide association studies (GWAS) identification of NMNAT2 at rs2022013, we fine-mapped its 150â kb flanking regions containing NMNAT2 and SMG7 in a 15â 292 case-control multi-ancestry population and tested functions of identified variants. METHODS: We 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. RESULTS: We 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). CONCLUSION: We 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.
Assuntos
Anticorpos Antinucleares/metabolismo , Proteínas de Transporte/genética , Leucócitos Mononucleares/imunologia , Lúpus Eritematoso Sistêmico/genética , Nicotinamida-Nucleotídeo Adenililtransferase/genética , Alelos , Indígena Americano ou Nativo do Alasca/genética , Feminino , Predisposição Genética para Doença , Estudo de Associação Genômica Ampla , Técnicas de Genotipagem , Células HEK293 , Hispânico ou Latino/genética , Humanos , Desequilíbrio de Ligação , Lúpus Eritematoso Sistêmico/imunologia , Masculino , Linhagem , RNA Mensageiro/metabolismo , Reação em Cadeia da Polimerase em Tempo Real , Fatores de Risco , População Branca/genéticaRESUMO
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 [P(meta) = 5.20×10(-14); odds ratio, 95% confidence interval = 0.82 (0.78-0.87)], and two missense variants, rs1990760 (Ala946Thr) [P(meta) = 3.08×10(-7); 0.88 (0.84-0.93)] and rs10930046 (Arg460His) [P(dom) = 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.
Assuntos
Negro ou Afro-Americano/genética , RNA Helicases DEAD-box/genética , Lúpus Eritematoso Sistêmico/genética , Alelos , Antígenos Nucleares/genética , Antígenos Nucleares/imunologia , Apoptose/genética , Autoanticorpos/genética , Autoanticorpos/imunologia , Mapeamento Cromossômico , Proteínas de Ligação a DNA/genética , Proteínas de Ligação a DNA/imunologia , Predisposição Genética para Doença , Genoma Humano , Haplótipos , Humanos , Inflamação/genética , Helicase IFIH1 Induzida por Interferon , Autoantígeno Ku , Lúpus Eritematoso Sistêmico/imunologia , Polimorfismo de Nucleotídeo Único , Ligação Proteica , População Branca/genéticaRESUMO
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 [Pâ=â6.5×10(-10), odds ratio (OR) (95%CI)â=â1.27 (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 (Pmetaâ=â7.5×10(-11), ORâ=â1.24 [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 (R(2)â=â0.255, Pâ=â0.001). 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 (Pâ=â0.0003). 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 â=â2.0×10(-19), ORâ=â1.25 [1.20-1.32]), which confers allelic effect on transcript turnover via differential binding to the epigenetic factor miR-3148.
Assuntos
Predisposição Genética para Doença , Lúpus Eritematoso Sistêmico/genética , MicroRNAs/genética , Receptor 7 Toll-Like , Regiões 3' não Traduzidas , Negro ou Afro-Americano/genética , Alelos , Povo Asiático/genética , Regulação da Expressão Gênica/genética , Células HEK293 , Hispânico ou Latino/genética , Humanos , Lúpus Eritematoso Sistêmico/metabolismo , Polimorfismo de Nucleotídeo Único , Receptor 7 Toll-Like/genética , Receptor 7 Toll-Like/metabolismo , População BrancaRESUMO
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.
Assuntos
Predisposição Genética para Doença , Estudo de Associação Genômica Ampla , Lúpus Eritematoso Sistêmico/genética , Ligante OX40/genética , Negro ou Afro-Americano/genética , Alelos , Povo Asiático/genética , Mapeamento Cromossômico , Feminino , Genótipo , Haplótipos , Hispânico ou Latino/genética , Humanos , Desequilíbrio de Ligação , Lúpus Eritematoso Sistêmico/patologia , Linfócitos/patologia , Masculino , NF-kappa B/genética , Polimorfismo de Nucleotídeo Único , População Branca/genéticaRESUMO
Immunoregulatory cytokine interleukin-10 (IL-10) is elevated in sera from patients with systemic lupus erythematosus (SLE) correlating with disease activity. The established association of IL10 with SLE and other autoimmune diseases led us to fine map causal variant(s) and to explore underlying mechanisms. We assessed 19 tag SNPs, covering the IL10 gene cluster including IL19, IL20 and IL24, for association with SLE in 15,533 case and control subjects from four ancestries. The previously reported IL10 variant, rs3024505 located at 1 kb downstream of IL10, exhibited the strongest association signal and was confirmed for association with SLE in European American (EA) (Pâ=â2.7×10â»8, ORâ=â1.30), but not in non-EA ancestries. SNP imputation conducted in EA dataset identified three additional SLE-associated SNPs tagged by rs3024505 (rs3122605, rs3024493 and rs3024495 located at 9.2 kb upstream, intron 3 and 4 of IL10, respectively), and SLE-risk alleles of these SNPs were dose-dependently associated with elevated levels of IL10 mRNA in PBMCs and circulating IL-10 protein in SLE patients and controls. Using nuclear extracts of peripheral blood cells from SLE patients for electrophoretic mobility shift assays, we identified specific binding of transcription factor Elk-1 to oligodeoxynucleotides containing the risk (G) allele of rs3122605, suggesting rs3122605 as the most likely causal variant regulating IL10 expression. Elk-1 is known to be activated by phosphorylation and nuclear localization to induce transcription. Of interest, phosphorylated Elk-1 (p-Elk-1) detected only in nuclear extracts of SLE PBMCs appeared to increase with disease activity. Co-expression levels of p-Elk-1 and IL-10 were elevated in SLE T, B cells and monocytes, associated with increased disease activity in SLE B cells, and were best downregulated by ERK inhibitor. Taken together, our data suggest that preferential binding of activated Elk-1 to the IL10 rs3122605-G allele upregulates IL10 expression and confers increased risk for SLE in European Americans.
Assuntos
Predisposição Genética para Doença , Interleucina-10/genética , Lúpus Eritematoso Sistêmico/genética , Proteínas Elk-1 do Domínio ets/genética , Alelos , Povo Asiático , Regulação da Expressão Gênica , Estudo de Associação Genômica Ampla , Genótipo , Haplótipos , Hispânico ou Latino , Humanos , Interleucina-10/biossíntese , Íntrons , Lúpus Eritematoso Sistêmico/patologia , Polimorfismo de Nucleotídeo Único , Ligação Proteica , Regulação para Cima , População Branca/genética , Proteínas Elk-1 do Domínio ets/biossínteseRESUMO
Protein phosphatase 2A (PP2A) is a highly conserved and ubiquitous serine/threonine phosphatase. We have shown previously that PP2A expression is increased in T cells of systemic lupus erythematosus patients and that this increased expression and activity of PP2A plays a central role in the molecular pathogenesis of systemic lupus erythematosus. Although the control of PP2A expression has been the focus of many studies, many aspects of its regulation still remain poorly understood. In this study, we describe a novel mechanism of PP2A regulation. We propose that the transcription factor Ikaros binds to a variant site in the first intron of PP2A and modulates its expression. Exogenous expression of Ikaros leads to reduced levels of PP2Ac message as well as protein. Conversely, siRNA-enabled silencing of Ikaros enhances the expression of PP2A, suggesting that Ikaros acts as a suppressor of PP2A expression. A ChIP analysis further proved that Ikaros is recruited to this site in T cells. We also attempted to delineate the mechanism of Ikaros-mediated PP2Ac gene suppression. We show that Ikaros-mediated suppression of PP2A expression is at least partially dependent on the recruitment of the histone deacetylase HDAC1 to this intronic site. We conclude that the transcription factor Ikaros can regulate the expression of PP2A by binding to a site in the first intron and modulating chromatin modifications at this site via recruitment of HDAC1.
Assuntos
Regulação Enzimológica da Expressão Gênica/genética , Fator de Transcrição Ikaros/metabolismo , Íntrons , Proteína Fosfatase 2/genética , Proteínas Repressoras/metabolismo , Sequência de Bases , Sítios de Ligação , Células HEK293 , Histona Desacetilase 1/metabolismo , Humanos , Regiões Promotoras Genéticas/genética , Transporte ProteicoRESUMO
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.
Assuntos
Predisposição Genética para Doença/genética , Leptina/genética , Lúpus Eritematoso Sistêmico/genética , Polimorfismo de Nucleotídeo Único/genética , Estudos de Casos e Controles , Genótipo , HumanosRESUMO
Systemic lupus erythematosus (SLE) is a chronic heterogeneous autoimmune disorder characterized by the loss of tolerance to self-antigens and dysregulated interferon responses. The etiology of SLE is complex, involving both heritable and environmental factors. Candidate-gene studies and genome-wide association (GWA) scans have been successful in identifying new loci that contribute to disease susceptibility; however, much of the heritable risk has yet to be identified. In this study, we sought to replicate 1,580 variants showing suggestive association with SLE in a previously published GWA scan of European Americans; we tested a multiethnic population consisting of 7,998 SLE cases and 7,492 controls of European, African American, Asian, Hispanic, Gullah, and Amerindian ancestry to find association with the disease. Several genes relevant to immunological pathways showed association with SLE. Three loci exceeded the genome-wide significance threshold: interferon regulatory factor 8 (IRF8; rs11644034; p(meta-Euro) = 2.08 × 10(-10)), transmembrane protein 39A (TMEM39A; rs1132200; p(meta-all) = 8.62 × 10(-9)), and 17q21 (rs1453560; p(meta-all) = 3.48 × 10(-10)) between IKAROS family of zinc finger 3 (AIOLOS; IKZF3) and zona pellucida binding protein 2 (ZPBP2). Fine mapping, resequencing, imputation, and haplotype analysis of IRF8 indicated that three independent effects tagged by rs8046526, rs450443, and rs4843869, respectively, were required for risk in individuals of European ancestry. Eleven additional replicated effects (5 × 10(-8) < p(meta-Euro) < 9.99 × 10(-5)) were observed with CFHR1, CADM2, LOC730109/IL12A, LPP, LOC63920, SLU7, ADAMTSL1, C10orf64, OR8D4, FAM19A2, and STXBP6. The results of this study increase the number of confirmed SLE risk loci and identify others warranting further investigation.