Thymic stromal lymphopoietin (TSLP), an IL7-like cytokine produced by bronchial epithelial cells is upregulated in asthma and induces dendritic cell maturation supporting a Th2 response. Environmental pollutants, including tobacco smoke and diesel exhaust particles upregulate TSLP suggesting that TSLP may be an interface between environmental pollution and immune responses in asthma. Since asthma is prevalent in urban communities, variants in the TSLP gene may be important in asthma susceptibility in these populations.

Primary biliary cirrhosis (PBC) is a classical autoimmune liver disease for which effective immunomodulatory therapy is lacking. Here we perform meta-analyses of discovery data sets from genome-wide association studies of European subjects (n=2,764 cases and 10,475 controls) followed by validation genotyping in an independent cohort (n=3,716 cases and 4,261 controls). We discover and validate six previously unknown risk loci for PBC (Pcombined<5 × 10(-8)) and used pathway analysis to identify JAK-STAT/IL12/IL27 signalling and cytokine-cytokine pathways, for which relevant therapies exist.

Diabetic kidney disease (DKD) is the most common etiology of chronic kidney disease (CKD) in the industrialized world and accounts for much of the excess mortality in patients with diabetes mellitus. Approximately 45% of U.S. patients with incident end-stage kidney disease (ESKD) have DKD. Independent of glycemic control, DKD aggregates in families and has higher incidence rates in African, Mexican, and American Indian ancestral groups relative to European populations. The Family Investigation of Nephropathy and Diabetes (FIND) performed a genome-wide association study (GWAS) contrasting 6,197 unrelated individuals with advanced DKD with healthy and diabetic individuals lacking nephropathy of European American, African American, Mexican American, or American Indian ancestry. A large-scale replication and trans-ethnic meta-analysis included 7,539 additional European American, African American and American Indian DKD cases and non-nephropathy controls. Within ethnic group meta-analysis of discovery GWAS and replication set results identified genome-wide significant evidence for association between DKD and rs12523822 on chromosome 6q25.2 in American Indians (P = 5.74x10-9). The strongest signal of association in the trans-ethnic meta-analysis was with a SNP in strong linkage disequilibrium with rs12523822 (rs955333; P = 1.31x10-8), with directionally consistent results across ethnic groups. These 6q25.2 SNPs are located between the SCAF8 and CNKSR3 genes, a region with DKD relevant changes in gene expression and an eQTL with IPCEF1, a gene co-translated with CNKSR3. Several other SNPs demonstrated suggestive evidence of association with DKD, within and across populations. These data identify a novel DKD susceptibility locus with consistent directions of effect across diverse ancestral groups and provide insight into the genetic architecture of DKD.

The minor allele of the R620W missense single-nucleotide polymorphism (SNP) (rs2476601) in the hematopoietic-specific protein tyrosine phosphatase gene, PTPN22, has been associated with multiple autoimmune diseases, including rheumatoid arthritis (RA). These genetic data, combined with biochemical evidence that this SNP affects PTPN22 function, suggest that this phosphatase is a key regulator of autoimmunity. To determine whether other genetic variants in PTPN22 contribute to the development of RA, we sequenced the coding regions of this gene in 48 white North American patients with RA and identified 15 previously unreported SNPs, including 2 coding SNPs in the catalytic domain. We then genotyped 37 SNPs in or near PTPN22 in 475 patients with RA and 475 individually matched controls (sample set 1) and selected a subset of markers for replication in an additional 661 patients with RA and 1,322 individually matched controls (sample set 2). Analyses of these results predict 10 common (frequency >1%) PTPN22 haplotypes in white North Americans. The sole haplotype found to carry the previously identified W620 risk allele was strongly associated with disease in both sample sets, whereas another haplotype, identical at all other SNPs but carrying the R620 allele, showed no association. R620W, however, does not fully explain the association between PTPN22 and RA, since significant differences between cases and controls persisted in both sample sets after the haplotype data were stratified by R620W. Additional analyses identified two SNPs on a single common haplotype that are associated with RA independent of R620W, suggesting that R620W and at least one additional variant in the PTPN22 gene region influence RA susceptibility.

To discover new rheumatoid arthritis (RA) risk loci, we systematically examined 370 SNPs from 179 independent loci with P < 0.001 in a published meta-analysis of RA genome-wide association studies (GWAS) of 3,393 cases and 12,462 controls. We used Gene Relationships Across Implicated Loci (GRAIL), a computational method that applies statistical text mining to PubMed abstracts, to score these 179 loci for functional relationships to genes in 16 established RA disease loci. We identified 22 loci with a significant degree of functional connectivity. We genotyped 22 representative SNPs in an independent set of 7,957 cases and 11,958 matched controls. Three were convincingly validated: CD2-CD58 (rs11586238, P = 1 x 10(-6) replication, P = 1 x 10(-9) overall), CD28 (rs1980422, P = 5 x 10(-6) replication, P = 1 x 10(-9) overall) and PRDM1 (rs548234, P = 1 x 10(-5) replication, P = 2 x 10(-8) overall). An additional four were replicated (P < 0.0023): TAGAP (rs394581, P = 0.0002 replication, P = 4 x 10(-7) overall), PTPRC (rs10919563, P = 0.0003 replication, P = 7 x 10(-7) overall), TRAF6-RAG1 (rs540386, P = 0.0008 replication, P = 4 x 10(-6) overall) and FCGR2A (rs12746613, P = 0.0022 replication, P = 2 x 10(-5) overall). Many of these loci are also associated to other immunologic diseases.

Accounting for population genetic substructure is important in reducing type 1 errors in genetic studies of complex disease. As efforts to understand complex genetic disease are expanded to different continental populations the understanding of genetic substructure within these continents will be useful in design and execution of association tests. In this study, population differentiation (Fst) and Principal Components Analyses (PCA) are examined using >200 K genotypes from multiple populations of East Asian ancestry. The population groups included those from the Human Genome Diversity Panel [Cambodian, Yi, Daur, Mongolian, Lahu, Dai, Hezhen, Miaozu, Naxi, Oroqen, She, Tu, Tujia, Naxi, Xibo, and Yakut], HapMap [ Han Chinese (CHB) and Japanese (JPT)], and East Asian or East Asian American subjects of Vietnamese, Korean, Filipino and Chinese ancestry. Paired Fst (Wei and Cockerham) showed close relationships between CHB and several large East Asian population groups (CHB/Korean, 0.0019; CHB/JPT, 00651; CHB/Vietnamese, 0.0065) with larger separation with Filipino (CHB/Filipino, 0.014). Low levels of differentiation were also observed between Dai and Vietnamese (0.0045) and between Vietnamese and Cambodian (0.0062). Similarly, small Fst's were observed among different presumed Han Chinese populations originating in different regions of mainland of China and Taiwan (Fst's <0.0025 with CHB). For PCA, the first two PC's showed a pattern of relationships that closely followed the geographic distribution of the different East Asian populations. PCA showed substructure both between different East Asian groups and within the Han Chinese population. These studies have also identified a subset of East Asian substructure ancestry informative markers (EASTASAIMS) that may be useful for future complex genetic disease association studies in reducing type 1 errors and in identifying homogeneous groups that may increase the power of such studies.

Case-control genetic studies of complex human diseases can be confounded by population stratification. This issue can be addressed using panels of ancestry informative markers (AIMs) that can provide substantial population substructure information. Previously, we described a panel of 128 SNP AIMs that were designed as a tool for ascertaining the origins of subjects from Europe, Sub-Saharan Africa, Americas, and East Asia.

For admixture mapping studies in Mexican Americans (MAM), we define a genomewide single-nucleotide-polymorphism (SNP) panel that can distinguish between chromosomal segments of Amerindian (AMI) or European (EUR) ancestry. These studies used genotypes for >400,000 SNPs, defined in EUR and both Pima and Mayan AMI, to define a set of ancestry-informative markers (AIMs). The use of two AMI populations was necessary to remove a subset of SNPs that distinguished genotypes of only one AMI subgroup from EUR genotypes. The AIMs set contained 8,144 SNPs separated by a minimum of 50 kb with only three intermarker intervals >1 Mb and had EUR/AMI FST values >0.30 (mean FST = 0.48) and Mayan/Pima FST values <0.05 (mean FST < 0.01). Analysis of a subset of these SNP AIMs suggested that this panel may also distinguish ancestry between EUR and other disparate AMI groups, including Quechuan from South America. We show, using realistic simulation parameters that are based on our analyses of MAM genotyping results, that this panel of SNP AIMs provides good power for detecting disease-associated chromosomal segments for genes with modest ethnicity risk ratios. A reduced set of 5,287 SNP AIMs captured almost the same admixture mapping information, but smaller SNP sets showed substantial drop-off in admixture mapping information and power. The results will enable studies of type 2 diabetes, rheumatoid arthritis, and other diseases among which epidemiological studies suggest differences in the distribution of ancestry-associated susceptibility.

Primary biliary cholangitis (PBC) is an autoimmune liver disease with a strong hereditary component. Here, we report a genome-wide association study that included 1,122 PBC cases and 4,036 controls of Han Chinese descent, with subsequent replication in a separate cohort of 907 PBC cases and 2,127 controls. Our results show genome-wide association of 14 PBC risk loci including previously identified 6p21 (HLA-DRA and DPB1), 17q12 (ORMDL3), 3q13.33 (CD80), 2q32.3 (STAT1/STAT4), 3q25.33 (IL12A), 4q24 (NF-κB) and 22q13.1 (RPL3/SYNGR1). We also identified variants in IL21, IL21R, CD28/CTLA4/ICOS, CD58, ARID3A and IL16 as novel PBC risk loci. These new findings and histochemical studies showing enhanced expression of IL21 and IL21R in PBC livers (particularly in the hepatic portal tracks) support a disease mechanism in which the deregulation of the IL21 signalling pathway, in addition to CD4 T-cell activation and T-cell co-stimulation are critical components in the development of PBC.

Primary biliary cirrhosis (PBC) is an uncommon autoimmune disease with a homogeneous clinical phenotype that reflects incomplete disease concordance in monozygotic (MZ) twins. We have taken advantage of a unique collection consisting of genomic DNA and mRNA from peripheral blood cells of female MZ twins (n = 3 sets) and sisters of similar age (n = 8 pairs) discordant for disease. We performed a genome-wide study to investigate differences in (i) DNA methylation (using a custom tiled four-plex array containing tiled 50-mers 19,084 randomly chosen methylation sites), (ii) copy number variation (CNV) (with a chip including markers derived from the 1000 Genomes Project, all three HapMap phases, and recently published studies), and/or (iii) gene expression (by whole-genome expression arrays). Based on the results obtained from these three approaches we utilized quantitative PCR to compare the expression of candidate genes. Importantly, our data support consistent differences in discordant twins and siblings for the (i) methylation profiles of 60 gene regions, (ii) CNV of 10 genes, and (iii) the expression of 2 interferon-dependent genes. Quantitative PCR analysis showed that 17 of these genes are differentially expressed in discordant sibling pairs. In conclusion, we report that MZ twins and sisters discordant for PBC manifest particular epigenetic differences and highlight the value of the epigenetic study of twins.

Anti-nuclear antibodies to speckled 100 kDa (sp100) and glycoprotein 210 (gp210) are specific serologic markers of primary biliary cholangitis (PBC) of uncertain/controversial clinical or prognostic significance. To study the genetic determinants associated with sp100 and gp210 autoantibody subphenotypes, we performed a genome-wide association analysis of 930 PBC cases based on their autoantibody status, followed by a replication study in 1,252 PBC cases. We confirmed single-nucleotide polymorphisms rs492899 (P = 3.27 × 10-22 ; odds ratio [OR], 2.90; 95% confidence interval [CI], 2.34-3.66) and rs1794280 (P = 5.78 × 10-28 ; OR, 3.89; 95% CI, 3.05-4.96) in the human major histocompatibility complex (MHC) region associated with the sp100 autoantibody. However, no genetic variant was identified as being associated with the gp210 autoantibody. To further define specific classical human leukocyte antigen (HLA) alleles or amino acids associated with the sp100 autoantibody, we imputed 922 PBC cases (211 anti-sp100-positive versus 711 negative cases) using a Han Chinese MHC reference database. Conditional analysis identified that HLA-DRβ1-Asn77/Arg74, DRβ1-Ser37, and DPβ1-Lys65 were major determinants for sp100 production. For the classical HLA alleles, the strongest association was with DRB1*03:01 (P = 1.51 × 10-9 ; OR, 2.97; 95% CI, 2.06-4.29). Regression analysis with classical HLA alleles identified DRB1*03:01, DRB1*15:01, DRB1*01, and DPB1*03:01 alleles can explain most of the HLA association with sp100 autoantibody. Conclusion: This study indicated significant genetic predisposition to the sp100 autoantibody, but not the gp210 autoantibody, subphenotype in PBC patients. Additional studies will be necessary to determine if these findings have clinical significance to PBC pathogenesis and/or therapeutics.

The CD28 locus is associated with susceptibility to a variety of autoimmune and immune-mediated inflammatory diseases including primary sclerosing cholangitis (PSC). Previously, we linked the CD28 pathway in PSC disease pathology and found that vitamin D could maintain CD28 expression. Here, we assessed whether the PSC-associated CD28 risk variant A (rs7426056) affects CD28 expression and T cell function in healthy individuals (n = 14 AA, n = 14 AG, n = 14 GG). Homozygotes for the PSC disease risk allele (AA) showed significantly lower CD28 mRNA expression ex-vivo than either GG or AG (p < 0.001) in total peripheral blood mononuclear cells. However, the CD28 risk variant alone was not sufficient to explain CD28 protein loss on CD4+ T cells. All genotypes responded equally to vitamin D as indicated by induction of a regulatory phenotype and an increased anti-inflammatory/pro-inflammatory cytokine ratio. A genotypic effect on response to TNFα stimuli was detected, which was inhibited by vitamin D. Together our results show: (a) an altered gene expression in carriers of the susceptible CD28 variant, (b) no differences in protein levels on CD4+ T cells, and

We have performed a meta-analysis of the major-histocompatibility-complex (MHC) region in systemic lupus erythematosus (SLE) to determine the association with both SNPs and classical human-leukocyte-antigen (HLA) alleles. More specifically, we combined results from six studies and well-known out-of-study control data sets, providing us with 3,701 independent SLE cases and 12,110 independent controls of European ancestry. This study used genotypes for 7,199 SNPs within the MHC region and for classical HLA alleles (typed and imputed). Our results from conditional analysis and model choice with the use of the Bayesian information criterion show that the best model for SLE association includes both classical loci (HLA-DRB1(∗)03:01, HLA-DRB1(∗)08:01, and HLA-DQA1(∗)01:02) and two SNPs, rs8192591 (in class III and upstream of NOTCH4) and rs2246618 (MICB in class I). Our approach was to perform a stepwise search from multiple baseline models deduced from a priori evidence on HLA-DRB1 lupus-associated alleles, a stepwise regression on SNPs alone, and a stepwise regression on HLA alleles. With this approach, we were able to identify a model that was an overwhelmingly better fit to the data than one identified by simple stepwise regression either on SNPs alone (Bayes factor [BF] > 50) or on classical HLA alleles alone (BF > 1,000).

We conducted a genome-wide association study of rheumatoid arthritis in 2,418 cases and 4,504 controls from North America and identified an association at the REL locus, encoding c-Rel, on chromosome 2p13 (rs13031237, P = 6.01 x 10(-10)). Replication in independent case-control datasets comprising 2,604 cases and 2,882 controls confirmed this association, yielding an allelic OR = 1.25 (P = 3.08 x 10(-14)) for marker rs13031237 and an allelic OR = 1.21 (P = 2.60 x 10(-11)) for marker rs13017599 in the combined dataset. The combined dataset also provides definitive support for associations at both CTLA4 (rs231735; OR = 0.85; P = 6.25 x 10(-9)) and BLK (rs2736340; OR = 1.19; P = 5.69 x 10(-9)). c-Rel is an NF-kappaB family member with distinct functional properties in hematopoietic cells, and its association with rheumatoid arthritis suggests disease pathways that involve other recently identified rheumatoid arthritis susceptibility genes including CD40, TRAF1, TNFAIP3 and PRKCQ.

A substantial genetic contribution to systemic lupus erythematosus (SLE) risk is conferred by major histocompatibility complex (MHC) gene(s) on chromosome 6p21. Previous studies in SLE have lacked statistical power and genetic resolution to fully define MHC influences. We characterized 1,610 Caucasian SLE cases and 1,470 parents for 1,974 MHC SNPs, the highly polymorphic HLA-DRB1 locus, and a panel of ancestry informative markers. Single-marker analyses revealed strong signals for SNPs within several MHC regions, as well as with HLA-DRB1 (global p = 9.99 x 10(-16)). The most strongly associated DRB1 alleles were: *0301 (odds ratio, OR = 2.21, p = 2.53 x 10(-12)), *1401 (OR = 0.50, p = 0.0002), and *1501 (OR = 1.39, p = 0.0032). The MHC region SNP demonstrating the strongest evidence of association with SLE was rs3117103, with OR = 2.44 and p = 2.80 x 10(-13). Conditional haplotype and stepwise logistic regression analyses identified strong evidence for association between SLE and the extended class I, class I, class III, class II, and the extended class II MHC regions. Sequential removal of SLE-associated DRB1 haplotypes revealed independent effects due to variation within OR2H2 (extended class I, rs362521, p = 0.006), CREBL1 (class III, rs8283, p = 0.01), and DQB2 (class II, rs7769979, p = 0.003, and rs10947345, p = 0.0004). Further, conditional haplotype analyses demonstrated that variation within MICB (class I, rs3828903, p = 0.006) also contributes to SLE risk independent of HLA-DRB1*0301. Our results for the first time delineate with high resolution several MHC regions with independent contributions to SLE risk. We provide a list of candidate variants based on biologic and functional considerations that may be causally related to SLE risk and warrant further investigation.

Using a genome-wide single nucleotide polymorphism (SNP) panel, we observed population structure in a diverse group of Europeans and European Americans. Under a variety of conditions and tests, there is a consistent and reproducible distinction between "northern" and "southern" European population groups: most individual participants with southern European ancestry (Italian, Spanish, Portuguese, and Greek) have >85% membership in the "southern" population; and most northern, western, eastern, and central Europeans have >90% in the "northern" population group. Ashkenazi Jewish as well as Sephardic Jewish origin also showed >85% membership in the "southern" population, consistent with a later Mediterranean origin of these ethnic groups. Based on this work, we have developed a core set of informative SNP markers that can control for this partition in European population structure in a variety of clinical and genetic studies.

Admixture mapping requires a genomewide panel of relatively evenly spaced markers that can distinguish the ancestral origins of chromosomal segments in admixed individuals. Through use of the results of the International HapMap Project and specific selection criteria, the current study has examined the ability of selected single-nucleotide polymorphisms (SNPs) to extract continental ancestry information in African American subjects and to explore parameters for admixture mapping. Genotyping of two linguistically diverse West African populations (Bini and Kanuri Nigerians, who are Niger-Congo [Bantu] and Nilo-Saharan speakers, respectively), European Americans, and African Americans validated a genomewide set of >4,000 SNP ancestry-informative markers with mean and median F(ST) values >0.59 and mean and median Fisher's information content >2.5. This set of SNPs extracted a larger amount of ancestry information in African Americans than previously reported SNP panels and provides nearly uniform coverage of the genome. Moreover, in the current study, simulations show that this more informative panel improves power for admixture mapping in African Americans when ethnicity risk ratios are modest. This is particularly important in the application of admixture mapping in complex genetic diseases for which only modest ethnicity risk ratios of relevant susceptibility genes are expected.

Despite the success of genome-wide association studies (GWAS) in detecting a large number of loci for complex phenotypes such as rheumatoid arthritis (RA) susceptibility, the lack of information on the causal genes leaves important challenges to interpret GWAS results in the context of the disease biology. Here, we genetically fine-map the RA risk locus at 19p13 to define causal variants, and explore the pleiotropic effects of these same variants in other complex traits. First, we combined Immunochip dense genotyping (n = 23,092 case/control samples), Exomechip genotyping (n = 18,409 case/control samples) and targeted exon-sequencing (n = 2,236 case/controls samples) to demonstrate that three protein-coding variants in TYK2 (tyrosine kinase 2) independently protect against RA: P1104A (rs34536443, OR = 0.66, P = 2.3 x 10(-21)), A928V (rs35018800, OR = 0.53, P = 1.2 x 10(-9)), and I684S (rs12720356, OR = 0.86, P = 4.6 x 10(-7)). Second, we show that the same three TYK2 variants protect against systemic lupus erythematosus (SLE, Pomnibus = 6 x 10(-18)), and provide suggestive evidence that two of the TYK2 variants (P1104A and A928V) may also protect against inflammatory bowel disease (IBD; P(omnibus) = 0.005). Finally, in a phenome-wide association study (PheWAS) assessing >500 phenotypes using electronic medical records (EMR) in >29,000 subjects, we found no convincing evidence for association of P1104A and A928V with complex phenotypes other than autoimmune diseases such as RA, SLE and IBD. Together, our results demonstrate the role of TYK2 in the pathogenesis of RA, SLE and IBD, and provide supporting evidence for TYK2 as a promising drug target for the treatment of autoimmune diseases.

Selective immunoglobulin A deficiency (IgAD) is the most common primary immunodeficiency in Europeans. Our genome-wide association study (GWAS) meta-analysis of 1,635 patients with IgAD and 4,852 controls identified four new significant (P < 5 × 10-8) loci and association with a rare IFIH1 variant (p.Ile923Val). Peak new variants (PVT1, P = 4.3 × 10-11; ATG13-AMBRA1, P = 6.7 × 10-10; AHI1, P = 8.4 × 10-10; CLEC16A, P = 1.4 × 10-9) overlapped with autoimmune markers (3/4) and correlated with 21 putative regulatory variants, including expression quantitative trait loci (eQTLs) for AHI1 and DEXI and DNase hypersensitivity sites in FOXP3+ regulatory T cells. Pathway analysis of the meta-analysis results showed striking association with the KEGG pathway for IgA production (pathway P < 0.0001), with 22 of the 30 annotated pathway genes containing at least one variant with P ≤ 0.05 in the IgAD meta-analysis. These data suggest that a complex network of genetic effects, including genes known to influence the biology of IgA production, contributes to IgAD.

Progress in defining the genetics of autoimmune disease has been dramatically enhanced by large scale genetic studies. Genome-wide approaches, examining hundreds or for some diseases thousands of cases and controls, have been implemented using high throughput genotyping and appropriate algorithms to provide a wealth of data over the last decade. These studies have identified hundreds of non-HLA loci as well as further defining HLA variations that predispose to different autoimmune diseases. These studies to identify genetic risk loci are also complemented by progress in gene expression studies including definition of expression quantitative trait loci (eQTL), various alterations in chromatin structure including histone marks, DNase I sensitivity, repressed chromatin regions as well as transcript factor binding sites. Integration of this information can partially explain why particular variations can alter proclivity to autoimmune phenotypes. Despite our incomplete knowledge base with only partial definition of hereditary factors and possible functional connections, this progress has and will continue to facilitate a better understanding of critical pathways and critical changes in immunoregulation. Advances in defining and understanding functional variants potentially can lead to both novel therapeutics and personalized medicine in which therapeutic approaches are chosen based on particular molecular phenotypes and genomic alterations.