Abstract
The leukocyte-associated immunoglobulin-like receptor 1 (LAIR-1) is a collagen-binding inhibitory receptor important for the regulation of immune responses, expressed on the majority of peripheral blood mononuclear cells (PBMC). LAIR-2 is a soluble homolog that antagonizes LAIR-1 inhibitory function by binding the same ligands. We sought to investigate whether LAIR1 and LAIR2 single nucleotide polymorphisms (SNP) are associated with differential mRNA expression levels. We analyzed 14 SNPs of LAIR1 (6) and LAIR2 (8) by mass spectrometry-based genotyping and extracted mRNA from PBMC of 177 healthy subjects, followed by quantitative assays. Four SNPs of LAIR1 and two SNPs of LAIR2 mark differential mRNA levels in healthy individuals. To verify the biological relevance of these findings, we analyzed additional 515 individuals (282 patients and 233 controls) to check if LAIR1 and LAIR2 differential mRNA expression could be related to susceptibility to pemphigus foliaceus (PF), an autoimmune blistering skin disease endemic in Brazil. Two LAIR1 variants (rs56802430 G, OR = 1.52, p = 0.0329; rs11084332 C, OR = 0.57, p = 0.0022) and one LAIR2 (rs2287828 T+, OR = 1.9, p = 0.0097) contribute to differential susceptibility to PF. Furthermore, we demonstrate interactions among four LAIR2 SNPs (rs2042287, rs2287828, rs2277974 and rs114834145). A haplotype harboring these SNPs is strongly associated with higher LAIR2 mRNA levels (4.5-fold, p = 0.0069) and with higher susceptibility to PF (OR = 4.02, p = 0.0008). We suggest that LAIR1 and LAIR2 genetic variants are associated with regulation of gene expression and variable PF susceptibility, and show indirect association of LAIR2 differential mRNA expression with PF pathogenesis. Our data demonstrate how this relatively unknown disease can add invaluable knowledge regarding the role of LAIR1 and LAIR2 in immune responses.
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Acknowledgments
We thank the patients and healthy controls for volunteering for this study, and the staff of the Hospital Adventista do Pênfigo and of the Banco de Sangue do Hospital de Clínicas from the Federal University of Paraná for their support. We also thank Gabriel Adelman Cipolla and Liana Alves de Oliveira for the technical and intellectual input for this work. This study was funded by the Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq), Programa de Apoio a Núcleos de Excelência (PRONEX), Fundação Araucária, and Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES). We also thank CNPq and CAPES for the research fellowships granted for all authors, in especial the Science without Borders Program for the Young Talent Fellowship awarded for the second author.
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439_2015_1626_MOESM1_ESM.tif
Supplementary material 1. Fig. S1: Linkage disequilibrium among SNP pairs distributed over the genes LAIR1 and LAIR2. LD plots for LAIR1 (left) and LAIR2 (right) SNPs for the admixed total sample. The colors are indicative of D’/logarithm of odds (LOD), and values correspond to r2. Bright red color represents LOD score for LD ≥ 2 and D’ = 1, shades of pink/red represents LOD ≥ 2 and D’ < 1, blue color represents D’ = 1 but LOD < 2, and white squares represent LOD < 2 and D’ < 1. In this study, we defined high LD as D’= 1 and r2 ≥ 0.80. LD plots were constructed using genotype data from the admixed total sample, using Haploview 4.2 (Barrett et al. 2005) (TIFF 64 kb)
439_2015_1626_MOESM2_ESM.tif
Supplementary material 2. Fig. S2: Localization of the qPCR TaqMan assays (red brackets) and of SNPs (rs numbers) analyzed in this study in the LAIR1 (a) and LAIR2 (b) mRNA variants validated by NCBI RefSeq. At the left of each mRNA are the RefSeq accession numbers (NM_). The graphical representation of the mRNAs variants is made to scale. Light green boxes represent the 5′ (left) and 3′ (right) untranslated regions; dark green boxes represent the coding segments of the exons. Figures modified from NCBI Gene Viewer (TIFF 380 kb)
439_2015_1626_MOESM3_ESM.xlsx
Supplementary material 3. Table S1: Predicted regulatory effects of LAIR1 and LAIR2 SNPs that are associated with differential gene expression and/or with pemphigus foliaceus. In this table we show the SNPs investigated in the study and SNPs in LD (r2 > 0.80) in EUR (European), CEU (Northern European from Utah, USA), CLM (Colombian) and MXL (Mexican) populations of the 1000 Genomes project, their genomic positions and their predicted effects on DNA regulatory elements. Data was obtained using the HaploReg3 database (Ward and Kellis 2012), RegulomeDB (Boyle et al. 2012), F-SNP (Lee and Shatkay 2008), SIFT and PolyPhen predictions using Ensembl Variant Effect Predictor (McLaren et al. 2010), Genevar (Yang et al. 2010), miRNASNP (Gong et al. 2015), mirSNP (Liu et al. 2012), PolymiRTS (Bhattacharya et al. 2014) and microSNIPER (Barenboim et al. 2010). For the eQTL search with Genevar, the level of statistical significance was set at p < 0.001. miRNA binding predictions tools were only consulted for SNPs in the 3′UTR region (XLSX 19 kb)
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Camargo, C.M., Augusto, D.G. & Petzl-Erler, M.L. Differential gene expression levels might explain association of LAIR2 polymorphisms with pemphigus. Hum Genet 135, 233–244 (2016). https://doi.org/10.1007/s00439-015-1626-6
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DOI: https://doi.org/10.1007/s00439-015-1626-6