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Rare variants in non-coding regulatory regions of the genome that affect gene expression in systemic lupus erythematosus.

ABSTRACT: Personalized medicine approaches are increasingly sought for diseases with a heritable component. Systemic lupus erythematosus (SLE) is the prototypic autoimmune disease resulting from loss of immunologic tolerance, but the genetic basis of SLE remains incompletely understood. Genome wide association studies (GWAS) identify regions associated with disease, based on common single nucleotide polymorphisms (SNPs) within them, but these SNPs may simply be markers in linkage disequilibrium with other, causative mutations. Here we use an hierarchical screening approach for prediction and testing of true functional variants within regions identified in GWAS; this involved bioinformatic identification of putative regulatory elements within close proximity to SLE SNPs, screening those regions for potentially causative mutations by high resolution melt analysis, and functional validation using reporter assays. Using this approach, we screened 15 SLE associated loci in 143 SLE patients, identifying 7 new variants including 5 SNPs and 2 insertions. Reporter assays revealed that the 5 SNPs were functional, altering enhancer activity. One novel variant was linked to the relatively well characterized rs9888739 SNP at the ITGAM locus, and may explain some of the SLE heritability at this site. Our study demonstrates that non-coding regulatory elements can contain private sequence variants affecting gene expression, which may explain part of the heritability of SLE.

SUBMITTER: Jones SA

PROVIDER: S-EPMC6817816 | biostudies-literature | 2019 Oct

REPOSITORIES: biostudies-literature

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Rare variants in non-coding regulatory regions of the genome that affect gene expression in systemic lupus erythematosus.

Jones Sarah A SA Cantsilieris Stuart S Fan Huapeng H Cheng Qiang Q Russ Brendan E BE Tucker Elena J EJ Harris James J Rudloff Ina I Nold Marcel M Northcott Melissa M Dankers Wendy W Toh Andrew E J AEJ White Stefan J SJ Morand Eric F EF

Scientific reports 20191028 1

Personalized medicine approaches are increasingly sought for diseases with a heritable component. Systemic lupus erythematosus (SLE) is the prototypic autoimmune disease resulting from loss of immunologic tolerance, but the genetic basis of SLE remains incompletely understood. Genome wide association studies (GWAS) identify regions associated with disease, based on common single nucleotide polymorphisms (SNPs) within them, but these SNPs may simply be markers in linkage disequilibrium with other ...[more]

PMID: 31659207

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Project description:INTRODUCTION:The majority of the genetic variance of systemic lupus erythematosus (SLE) remains unexplained by the common disease-common variant hypothesis. Rare variants, which are not detectable by genome-wide association studies because of their low frequencies, are predicted to explain part of this "missing heritability." However, recent studies identifying rare variants within known disease-susceptibility loci have failed to show genetic associations because of their extremely low frequencies, leading to the questioning of the contribution of rare variants to disease susceptibility. A common (minor allele frequency = 17.4% in cases) nonsynonymous coding variant rs1143679 (R77H) in ITGAM (CD11b), which forms half of the heterodimeric integrin receptor, complement receptor 3 (CR3), is robustly associated with SLE and has been shown to impair CR3-mediated phagocytosis. METHODS:We resequenced ITGAM in 73 SLE cases and identified two previously unidentified, case-specific nonsynonymous variants, F941V and G1145S. Both variants were genotyped in 2,107 and 949 additional SLE cases, respectively, to estimate their frequencies in a disease population. An in vitro model was used to assess the impact of F941V and G1145S, together with two nonsynonymous ITGAM polymorphisms, A858V (rs1143683) and M441T (rs11861251), on CR3-mediated phagocytosis. A paired two-tailed t test was used to compare the phagocytic capabilities of each variant with that of wild-type CR3. RESULTS:Both rare variants, F941V and G1145S, significantly impair CR3-mediated phagocytosis in an in vitro model (61% reduction, P = 0.006; 26% reduction, P = 0.0232). However, neither of the common variants, M441T and A858V, had an effect on phagocytosis. Neither rare variant was observed again in the genotyping of additional SLE cases, suggesting that their frequencies are extremely low. CONCLUSIONS:Our results add further evidence to the functional importance of ITGAM in SLE pathogenesis through impaired phagocytosis. Additionally, this study provides a new example of the identification of rare variants in common-allele-associated loci, which, because of their extremely low frequencies, are not statistically associated. However, the demonstration of their functional effects adds support to their contribution to disease risk, and questions the current notion of dismissing the contribution of very rare variants on purely statistical analyses.

Project description:Recently, common variants within or near ATG5, which is a key autophagy gene required for the formation of autophagosomes, have been identified as a candidate gene of systemic lupus erythematosus (SLE) by several genome-wide association studies. Moreover, elevated ATG5 expression was observed in SLE as well as other autoimmune diseases. However, no significant associations between variants within ATG5 and SLE were identified in several Chinese populations. The present study was conducted to further check the genetic role of ATG5 by associating both common and rare variants of ATG5 in Chinese patients with lupus nephritis (LN), a major phenotype with poor prognosis in SLE.To detect the association of common variants of ATG5 with LN, 7 tagging single nucleotide polymorphisms (SNPs) designed in immunochip and 4 SNPs reported to be associated with SLE were genotyped in 500 LN patients and 500 healthy controls. Furthermore, direct sequencing of exons and their flanking regions in 90 LN patients, 30 SLE patients, and 60 healthy controls were performed. Functional genomic annotation was performed by using public databases.None of the 11 tagging SNPs was observed to be associated with LN. By sequencing, 13 variants were identified, including 5 common SNPs, 7 not previously described, and 1 reported as rare variants (<1%) in the Single Nucleotide Polymorphism Database or the 1000 Genome project. None of the 5 common SNPs showed significant association between patients and controls, whereas increased frequencies of rare or novel variants were observed in patients compared with healthy controls, with 6/90 in LN patients, 2/30 in SLE patients, and 1/163 in healthy controls. Although these rare variants were observed to be located in the flanking regions of exons instead of missense mutations, patients carrying them tended to have severe clinical phenotype, and in silicon analysis suggested their regulatory effects.Increased frequencies of rare variants of ATG5 were identified in patients with LN and SLE compared with healthy controls, highlighting a likely important role of rare ATG5 variants in Chinese SLE patients.

Project description:ObjectiveThe multisystem involvement and high heterogeneity of systemic lupus erythematosus (SLE) lead to great challenges in its diagnosis and treatment. The purpose of this study was to find new lncRNAs in peripheral blood mononuclear cells of SLE patients by transcriptome sequencing and explore their potential as biomarkers and their correlation with clinical features.Materials and methodsTranscriptome sequencing was used to screen differentially expressed lncRNAs (DELs) and mRNAs (DEMs). The expression of these selected lncRNAs and mRNAs in SLE patients and healthy controls was verified by qPCR. DAVID and WebGestalt were used to perform enrichment analysis. Cytoscape was used to construct a protein-protein network, a coexpression network, and a competitive endogenous RNA network to reveal the regulatory mechanisms of lncRNAs at the transcriptome level.ResultsA total of 1737 DELs and 4078 DEMs were identified between SLE patients and healthy controls. Ten lncRNAs and eight genes were verified by qPCR in a larger sample set. The lncRNA NONHSAT101022.2 was significantly downregulated in SLE patients and was also significantly related to the activity and severity of disease. The upregulated genes were enriched in defense and the immune response, while the downregulated genes were mainly enriched in SLE-related pathways. Topology network analysis revealed that the lncRNAs were involved in regulation at the transcriptome level, including acting directly on mRNA or indirectly affecting gene expression by acting on miRNA.ConclusionIn this work, we identified many mRNAs and novel lncRNAs by transcriptome sequencing. The functions and regulatory mechanisms of these lncRNAs were analyzed by bioinformatic methods. The novel lncRNA NONHSAT101022.2 is significantly downregulated in SLE patients and is significantly related to the activity and severity of disease. Additionally, we propose that NONHSAT101022.2 may enhance the signal transduction of β2-AR by cis regulating LMBRD2, inducing NK cells to produce high levels of IFN-γ and thereby exacerbating SLE.

Dataset Information

Rare variants in non-coding regulatory regions of the genome that affect gene expression in systemic lupus erythematosus.

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Rare variants in non-coding regulatory regions of the genome that affect gene expression in systemic lupus erythematosus.

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