Searching for signals of evolutionary selection in 168 genes related to immune function

Pathogens have played a substantial role in human evolution, with past infections shaping genetic variation at loci influencing immune function. We selected 168 genes known to be involved in the immune response, genotyped common single nucleotide polymorphisms across each gene in three population samples (CEPH Europeans from Utah, Han Chinese from Guangxi, and Yoruba Nigerians from Southwest Nigeria) and searched for evidence of selection based on four tests for non-neutral evolution: minor allele frequency (MAF), derived allele frequency (DAF), Fst versus heterozygosity and extended haplotype homozygosity (EHH). Six of the 168 genes show some evidence for non-neutral evolution in this initial screen, with two showing similar signals in independent data from the International HapMap Project. These analyses identify two loci involved in immune function that are candidates for having been subject to evolutionary selection, and highlight a number of analytical challenges in searching for selection in genome-wide polymorphism data. Electronic Supplementary Material Supplementary material is available for this article at and is accessible for authorized users. Emily C. Walsh, Pardis Sabeti, Holli B. Hutcheson, and Ben Fry have contributed equally to this work and Stephen O’Brien and David Altshuler have jointly supervised this project     

Inferring malaria parasite population structure from serological networks

The malaria parasite Plasmodium falciparum is characterized by high levels of genetic diversity at antigenic loci involved in virulence and immune evasion. Knowledge of the population structure and dynamics of these genes is important for designing control programmes and understanding the acquisition of immunity to malaria; however, high rates of homologous and non-homologous recombination as well as complex patterns of expression within hosts have hindered attempts to elucidate these structures experimentally. Here, we analyse serological data from Kenya using a novel network technique to deconstruct the relationships between patients' immune responses to different parasite isolates. We show that particular population structures and expression patterns produce distinctive signatures within serological networks of parasite recognition, which can be used to discriminate between competing hypotheses regarding the organization of these genes. Our analysis suggests that different levels of immune selection occur within different groups of the same multigene family leading to mixed population structures.     

Signatures of adaptation and genetic structure among the mainland populations of Pinus radiata (D. Don) inferred from SNP loci

Insights into the relative contributions of locus specific and genome-wide effects on population genetic diversity can be gained through separation of their resulting genetic signals. Here we explore patterns of adaptive and neutral genetic diversity in the disjunct natural populations of Pinus radiata (D. Don) from mainland California. A first-generation common garden of 447 individuals revealed significant differentiation of wood phenotypes among populations (P _ST), possibly reflecting local adaptation in response to environment. We subsequently screened all trees for genetic diversity at 149 candidate gene single nucleotide polymorphism (SNP) loci for signatures of adaptation. Ten loci were identified as being possible targets of diversifying selection following F _ST outlier tests. Multivariate canonical correlation performed on a data set of 444 individuals identified significant covariance between environment, adaptive phenotypes and outlier SNP diversity, lending support to the case for local adaptation suggested from F _ST and P _ST tests. Covariation among discrete sets of outlier SNPs and adaptive phenotypes (inferred from multivariate loadings) with environment are supported by existing studies of candidate gene function and genotype–phenotype association. Canonical analyses failed to detect significant correlations between environment and 139 non-outlier SNP loci, which were applied to estimate neutral patterns of genetic differentiation among populations (F _ST 4.3 %). Using this data set, significant hierarchical structure was detected, indicating three populations on the mainland. The hierarchical relationships based on neutral SNP markers (and SSR) were in contrast with those inferred from putatively adaptive loci, potentially highlighting the independent action of selection and demography in shaping genetic structure in this species.     

Lack of genetic diversity across diverse immune genes in an endangered mammal,the Tasmanian devil (Sarcophilus harrisii)

The Tasmanian devil (Sarcophilus harrisii) is threatened with extinction due to the spread of devil facial tumour disease. Polymorphisms in immune genes can provide adaptive potential to resist diseases. Previous studies in diversity at immune loci in wild species have almost exclusively focused on genes of the major histocompatibility complex (MHC); however, these genes only account for a fraction of immune gene diversity. Devils lack diversity at functionally important immunity loci, including MHC and Toll‐like receptor genes. Whether there are polymorphisms at devil immune genes outside these two families is unknown. Here, we identify polymorphisms in a wide range of key immune genes, and develop assays to type single nucleotide polymorphisms (SNPs) within a subset of these genes. A total of 167 immune genes were examined, including cytokines, chemokines and natural killer cell receptors. Using genome‐level data from ten devils, SNPs within coding regions, introns and 10 kb flanking genes of interest were identified. We found low polymorphism across 167 immune genes examined bioinformatically using whole‐genome data. From this data, we developed long amplicon assays to target nine genes. These amplicons were sequenced in 29–220 devils and found to contain 78 SNPs, including eight SNPS within exons. Despite the extreme paucity of genetic diversity within these genes, signatures of balancing selection were exhibited by one chemokine gene, suggesting that remaining diversity may hold adaptive potential. The low functional diversity may leave devils highly vulnerable to infectious disease, and therefore, monitoring and preserving remaining diversity will be critical for the long‐term management of this species. Examining genetic variation in diverse immune genes should be a priority for threatened wildlife species. This study can act as a model for broad‐scale immunogenetic diversity analysis in threatened species.     

Evidence for selection at cytokine loci in a natural population of field voles (Microtus agrestis)

Individuals in natural populations are frequently exposed to a wide range of pathogens. Given the diverse profile of gene products involved in responses to different types of pathogen, this potentially results in complex pathogen-specific selection pressures acting on a broad spectrum of immune system genes in wild animals. Thus far, studies into the evolution of immune genes in natural populations have focused almost exclusively on the Major Histocompatibility Complex (MHC). However, the MHC represents only a fraction of the immune system and there is a need to broaden research in wild species to include other immune genes. Here, we examine the evidence for natural selection in a range of non-MHC genes in a natural population of field voles (Microtus agrestis). We concentrate primarily on genes encoding cytokines, signalling molecules critical in eliciting and mediating immune responses and identify signatures of natural selection acting on several of these genes. In particular, genetic diversity within Interleukin 1 beta and Interleukin 2 appears to have been maintained through balancing selection. Taken together with previous findings that polymorphism within these genes is associated with variation in resistance to multiple pathogens, this suggests that pathogen-mediated selection may be an important force driving genetic diversity at cytokine loci in voles and other natural populations. These results also suggest that, along with the MHC, preservation of genetic variation within cytokine genes should be a priority for the conservation genetics of threatened wildlife populations.     

Single nucleotide polymorphisms generated by genotyping by sequencing to characterize genome-wide diversity,linkage disequilibrium,and selective sweeps in cultivated watermelon

Background

A large single nucleotide polymorphism (SNP) dataset was used to analyze genome-wide diversity in a diverse collection of watermelon cultivars representing globally cultivated, watermelon genetic diversity. The marker density required for conducting successful association mapping depends on the extent of linkage disequilibrium (LD) within a population. Use of genotyping by sequencing reveals large numbers of SNPs that in turn generate opportunities in genome-wide association mapping and marker-assisted selection, even in crops such as watermelon for which few genomic resources are available. In this paper, we used genome-wide genetic diversity to study LD, selective sweeps, and pairwise F_ST distributions among worldwide cultivated watermelons to track signals of domestication.

Results

We examined 183 Citrullus lanatus var. lanatus accessions representing domesticated watermelon and generated a set of 11,485 SNP markers using genotyping by sequencing. With a diverse panel of worldwide cultivated watermelons, we identified a set of 5,254 SNPs with a minor allele frequency of ≥ 0.05, distributed across the genome. All ancestries were traced to Africa and an admixture of various ancestries constituted secondary gene pools across various continents. A sliding window analysis using pairwise F_ST values was used to resolve selective sweeps. We identified strong selection on chromosomes 3 and 9 that might have contributed to the domestication process. Pairwise analysis of adjacent SNPs within a chromosome as well as within a haplotype allowed us to estimate genome-wide LD decay. LD was also detected within individual genes on various chromosomes. Principal component and ancestry analyses were used to account for population structure in a genome-wide association study. We further mapped important genes for soluble solid content using a mixed linear model.

Conclusions

Information concerning the SNP resources, population structure, and LD developed in this study will help in identifying agronomically important candidate genes from the genomic regions underlying selection and for mapping quantitative trait loci using a genome-wide association study in sweet watermelon.

Electronic supplementary material

The online version of this article (doi:10.1186/1471-2164-15-767) contains supplementary material, which is available to authorized users.     

Maintaining functional major histocompatibility complex diversity under inbreeding: the case of a selfing vertebrate

Major histocompatibility complex (MHC) genes encode proteins that present pathogen-derived antigens to T-cells, initiating the adaptive immune response in vertebrates. Although populations with low MHC diversity tend to be more susceptible to pathogens, some bottlenecked populations persist and even increase in numbers despite low MHC diversity. Thus, the relative importance of MHC diversity versus genome-wide variability for the long-term viability of populations after bottlenecks and/or under high inbreeding is controversial. We tested the hypothesis that genome-wide inbreeding (estimated using microsatellites) should be more critical than MHC diversity alone in determining pathogen resistance in the self-fertilizing fish Kryptolebias marmoratus by analysing MHC diversity and parasite loads in natural and laboratory populations with different degrees of inbreeding. Both MHC and neutral diversities were lost after several generations of selfing, but we also found evidence of parasite selection acting on MHC diversity and of non-random loss of alleles, suggesting a possible selective advantage of those individuals with functionally divergent MHC, in accordance with the hypothesis of divergent allele advantage. Moreover, we found that parasite loads were better explained by including MHC diversity in the model than by genome-wide (microsatellites) heterozygosity alone. Our results suggest that immune-related overdominance could be the key in maintaining variables rates of selfing and outcrossing in K. marmoratus and other mixed-mating species.     

High level of functional polymorphism indicates a unique role of natural selection at human immune system loci

Several studies have shown that immune system proteins have on average a higher rate of amino acid evolution between different species of mammals than do most other proteins. To test whether immune-system-expressed loci show a correspondingly elevated rate of within-species nonsynonymous (amino acid altering) polymorphism, we examined gene diversity (heterozygosity) at 4,911 single nucleotide polymorphism (SNP) sites at 481 protein-coding loci. At loci with nonimmune functions, gene diversity at nonsynonymous SNP sites was typically lower than that at silent SNP sites (those not altering the amino acid sequence) in the same gene, a pattern that is an evidence of purifying selection acting to eliminate slightly deleterious variants. However, this pattern was not seen at nonsynonymous SNPs causing conservative amino acid replacements in immune system proteins, indicating that the latter are subject to a reduced level of functional constraint. Similarly, immune system genes showed higher gene diversities in their 5′ noncoding regions than did other proteins. These results identified certain immune system loci that are likely to be subject to balancing selection that acts to maintain polymorphism in either coding or regulatory regions. Electronic Supplementary Material Supplementary material is available for this article at .     

Divergence with gene flow in a population of thermophilic bacteria: a potential role for spatially varying selection

A fundamental goal of evolutionary biology is to understand how ecological diversity arises and is maintained in natural populations. We have investigated the contributions of gene flow and divergent selection to the distribution of genetic variation in an ecologically differentiated population of a thermophilic cyanobacterium (Mastigocladus laminosus) found along the temperature gradient of a nitrogen‐limited stream in Yellowstone National Park. For most loci sampled, gene flow appears to be sufficient to prevent substantial genetic divergence. However, one locus (rfbC) exhibited a comparatively low migration rate as well as other signatures expected for a gene experiencing spatially varying selection, including an excess of common variants, an elevated level of polymorphism and extreme genetic differentiation along the gradient. rfbC is part of an expression island involved in the production of the polysaccharide component of the protective envelope of the heterocyst, the specialized nitrogen‐fixing cell of these bacteria. SNP genotyping in the vicinity of rfbC revealed a ~5‐kbp region including a gene content polymorphism that is tightly associated with environmental temperature and therefore likely contains the target of selection. Two genes have been deleted both in the predominant haplotype found in the downstream region of White Creek and in strains from other Yellowstone populations of M. laminosus, which may result in the production of heterocysts with different envelope properties. This study implicates spatially varying selection in the maintenance of variation related to thermal performance at White Creek despite on‐going or recent gene flow.     

A genome-wide perspective on the diversity and selection signatures in indigenous goats using 53 K single nucleotide polymorphism array

Tibetan goats, Taihang goats, Jining grey goats, and Meigu goats are the representative indigenous goats in China, found in Qinghai-Tibet Plateau, Western pastoral area, Northern and Southern agricultural regions. Very few studies have conducted a comprehensive analysis of the genomic diversity and selection of these breeds. We genotyped 96 unrelated individuals, using goat 53 K Illumina BeadChip array, of the following goat breeds: Tibetan (TG), Taihang (THG), Jining grey (JGG), and Meigu (MGG). A total of 45 951 single nucleotide polymorphisms were filtered to estimate the genetic diversity and selection signatures. All breeds had a high proportion (over 95%) of polymorphic loci. The observed and excepted heterozygosity ranged from 0.338 (MGG) to 0.402 (JGG) and 0.339 (MGG) to 0.395 (JGG), respectively. Clustering analysis displayed a genetically distinct lineage for each breed, and their Fst were greater than 0.25, indicating that they had a higher genetic differentiation between groups. Furthermore, effective population size reduced in all four populations, indicating a loss of genetic diversity. In addition, runs of homozygosity were mainly distributed in 5–10 Mb. Lastly, we identified signature genes, which were closely related to high-altitude adaptation (ADIRF) and prolificity (CNTROB, SMC3, and PTEN). This study provides a valuable resource for future studies on genome-wide perspectives on the diversity and selection signatures of Chinese indigenous goats.     

Multiple sexual ornaments signal heterozygosity in male blue tits

Higher individual genetic quality has been hypothesized to be associated with the expression of conspicuous ornaments. However, the relationship between multicomponent sexual signals and heterozygosity is poorly understood. In this study, we examined whether different ornaments, including song (repertoire size and bout length) and plumage coloration (yellow breast and blue crown), reflect individual genetic diversity in male blue tits (Aves: Cyanistes caeruleus). We estimated genetic diversity using 26 microsatellite markers that were classified as putatively functional (12 loci) and neutral (14 loci). We found that yellow breast carotenoid chroma, blue crown brightness, bout length and body condition were positively associated with heterozygosity at functional loci, but not with genetic diversity estimated at all typed loci or the subset of neutral markers. The lack of strong single‐locus effects and the presence of identity disequilibrium in our population suggest that the observed heterozygosity‐phenotype associations are driven by loci widely distributed across the genome. The predominant role of putatively functional loci evidences that the expression of secondary sexual characters is more tightly reflected by heterozygosity at genomic regions containing coding genes that are being actively expressed, a fact that may make ornamental traits more reliable indicators of the genetic quality of individuals. Overall, this study shows that multiple secondary sexual characters reflect male genetic diversity and lends support to the good‐genes‐as‐heterozygosity hypothesis. © 2015 The Linnean Society of London, Biological Journal of the Linnean Society, 2015, 115 , 362–375.     

Worldwide genetic structure in 37 genes important in telomere biology

Telomeres form the ends of eukaryotic chromosomes and are vital in maintaining genetic integrity. Telomere dysfunction is associated with cancer and several chronic diseases. Patterns of genetic variation across individuals can provide keys to further understanding the evolutionary history of genes. We investigated patterns of differentiation and population structure of 37 telomere maintenance genes among 53 worldwide populations. Data from 898 unrelated individuals were obtained from the genome-wide scan of the Human Genome Diversity Panel (HGDP) and from 270 unrelated individuals from the International HapMap Project at 716 single-nucleotide polymorphism (SNP) loci. We additionally compared this gene set to HGDP data at 1396 SNPs in 174 innate immunity genes. The majority of the telomere biology genes had low to moderate haplotype diversity (45-85%), high ancestral allele frequencies (>60%) and low differentiation (FST<0.10). Heterozygosity and differentiation were significantly lower in telomere biology genes compared with the innate immunity genes. There was evidence of evolutionary selection in ACD, TERF2IP, NOLA2, POT1 and TNKS in this data set, which was consistent in HapMap 3. TERT had higher than expected levels of haplotype diversity, likely attributable to a lack of linkage disequilibrium, and a potential cancer-associated SNP in this gene, rs2736100, varied substantially in genotype frequency across major continental regions. It is possible that the genes under selection could influence telomere biology diseases. As a group, there appears to be less diversity and differentiation in telomere biology genes than in genes with different functions, possibly due to their critical role in telomere maintenance and chromosomal stability.     

Signatures of balancing selection are maintained at disease resistance loci following mating system evolution and a population bottleneck in the genus Capsella

ABSTRACT: BACKGROUND: Population bottlenecks can lead to a loss of variation at disease resistance loci, which could have important consequences for the ability of populations to adapt to pathogen pressure. Alternatively, current or past balancing selection could maintain high diversity, creating a strong heterogeneity in the retention of polymorphism across the genome of bottlenecked populations. We sequenced part of the LRR region of 9 NBS-LRR disease resistance genes in the outcrossing Capsella grandiflora and the recently derived, bottlenecked selfing species Capsella rubella, and compared levels and patterns of nucleotide diversity and divergence with genome-wide reference loci. RESULTS: In strong contrast with reference loci, average diversity at resistance loci was comparable between C. rubella and C. grandiflora, primarily due to two loci with highly elevated diversity indicative of past or present balancing selection. Average between-species differentiation was also reduced at the set of R-genes compared with reference loci, which is consistent with the maintenance of ancestral polymorphism. CONCLUSIONS: Historical or ongoing balancing selection on plant disease resistance genes is a likely contributor to the retention of ancestral polymorphism in some regions of the bottlenecked Capella rubella genome.     

Genetic structure in insular and mainland populations of house sparrows (Passer domesticus) and their hemosporidian parasites

Small and isolated populations usually exhibit low levels of genetic variability, and thus, they are expected to have a lower capacity to adapt to changes in environmental conditions, such as exposure to pathogens and parasites. Comparing the genetic variability of selectively neutral versus functional loci allows one to assess the evolutionary history of populations and their future evolutionary potential. The genes of the major histocompatibility complex (MHC) control immune recognition of parasites, and their unusually high diversity is genes which is likely driven by parasite‐mediated balancing selection. Here, we examined diversity and differentiation of neutral microsatellite loci and functional MHC class I genes in house sparrows (Passer domesticus), living in six insular and six mainland populations, and we aimed to determine whether their diversity or differentiation correlates with the diversity and the prevalence of infection of hemosporidian parasites. We found that island bird populations tended to have lower neutral genetic variability, whereas MHC variability gene was similar between island and mainland populations. Similarly, island populations tended to show greater genetic differentiation than mainland populations, especially at microsatellite markers. The maintenance of MHC genetic diversity and its less marked structure in the island populations could be attributed to balancing‐selection. The greater MHC differentiation among populations was negatively correlated with similarity in blood parasites (prevalence and diversity of parasite strains) between populations. Even at low prevalence and small geographical scale, haemosporidian parasites might contribute to structure the variability of immune genes among populations of hosts.     

Use of locally weighted scatterplot smoothing (LOWESS) regression to study selection signatures in Piedmontese and Italian Brown cattle breeds

Selection is the major force affecting local levels of genetic variation in species. The availability of dense marker maps offers new opportunities for a detailed understanding of genetic diversity distribution across the animal genome. Over the last 50 years, cattle breeds have been subjected to intense artificial selection. Consequently, regions controlling traits of economic importance are expected to exhibit selection signatures. The fixation index (F_st) is an estimate of population differentiation, based on genetic polymorphism data, and it is calculated using the relationship between inbreeding and heterozygosity. In the present study, locally weighted scatterplot smoothing (LOWESS) regression and a control chart approach were used to investigate selection signatures in two cattle breeds with different production aptitudes (dairy and beef). F_st was calculated for 42 514 SNP marker loci distributed across the genome in 749 Italian Brown and 364 Piedmontese bulls. The statistical significance of F_st values was assessed using a control chart. The LOWESS technique was efficient in removing noise from the raw data and was able to highlight selection signatures in chromosomes known to harbour genes affecting dairy and beef traits. Examples include the peaks detected for BTA2 in the region where the myostatin gene is located and for BTA6 in the region harbouring the ABCG2 locus. Moreover, several loci not previously reported in cattle studies were detected.     

Evidence of parasite-mediated disruptive selection on genetic diversity in a wild fish population

Identifying the processes maintaining genetic variability in wild populations is a major concern in conservation and evolutionary biology. Parasite-mediated selection may strongly affect genetic variability in wild populations. The inbreeding depression theory predicts that directional selection imposed by parasites should act against the most inbred hosts, thus favouring genetic diversity in wild populations. We have tested this prediction by evaluating the strength and shape of the relationship between the load of a harmful fin-feeder ectoparasite ( Tracheliastes polycolpus ) and the genome-wide genetic diversity (i.e. heterozygosity measured at a set of 15 microsatellites) of its fish host, the rostrum dace ( Leuciscus leuciscus ). Contrary to expectation, we found a nonlinear relationship between host genetic diversity and ectoparasite load, with hosts that were either homozygous or heterozygous harbouring significantly fewer parasites than hosts with an intermediate level of heterozygosity. This relationship suggests that parasites could increase the variance of global heterozygosity in this host population through disruptive selection on genetic diversity. Moreover, when genetic diversity was measured at each locus separately, we found two very strong positive associations between host genetic diversity and the ectoparasite load. This latter result has three main implications: (i) genome-wide effect cannot alone explain the nonlinear relationship between global heterozygosity and ectoparasite load, (ii) negative non-additive allelic interactions (i.e. underdominance) may be a mechanism for resisting ectoparasite infection, and (iii) ectoparasites may favour homozygosity at some loci in this host population.     

Gene Diversity and Genetic Differentiation in Natural Populations of Picea koraiensis

Vertical slab polyacrylamide gel electrophoresis was used to examine inheritance of four isoenzymes in megagametophytes of Picea koraiensis. Gene diversity and genetic differentiation in four natural populations of P. koraiensis in Northeastern China were analyzed at six loci coding for four enzyme systems. The proportion of loci polymorphism in population was 0.5. The average expected heterozygosity of all samples was 0. 314, and the average observed heterozygosity of all samples was 0. 316. The average number of alleles detected per locus was 2.50. The effective number of alleles per locus was 1. 71. Measurement of gene diversity for the six loci showed a 0. 0059 significance of interpopulation differentiation. More than ninety-nine percent of the total gene diversity resided within population. The mean genetic distance over all pairs of P. koraiensis was 0. 0110.     

Identifying footprints of directional and balancing selection in marine and freshwater three-spined stickleback (Gasterosteus aculeatus) populations

Natural selection is expected to leave an imprint on the neutral polymorphisms at the adjacent genomic regions of a selected gene. While directional selection tends to reduce within-population genetic diversity and increase among-population differentiation, the reverse is expected under balancing selection. To identify targets of natural selection in the three-spined stickleback ( Gasterosteus aculeatus ) genome, 103 microsatellite and two indel markers including expressed sequence tags (EST) and quantitative trait loci (QTL)-associated loci, were genotyped in four freshwater and three marine populations. The results indicated that a high proportion of loci (14.7%) might be affected by balancing selection and a lower proportion (2.8%) by directional selection. The strongest signatures of directional selection were detected in a microsatellite locus and two indel markers located in the intronic regions of the Eda-gene coding for the number of lateral plates. Yet, other microsatellite loci previously found to be informative in QTL-mapping studies revealed no signatures of selection. Two novel microsatellite loci ( Stn12 and Stn90 ) located in chromosomes I and VIII, respectively, showed signals of directional selection and might be linked to genomic regions containing gene(s) important for adaptive divergence. Although the coverage of the total genomic content was relatively low, the predominance of balancing selection signals is in agreement with the contention that balancing, rather than directional selection is the predominant mode of selection in the wild.     

Transcriptome Profiling of Human Ulcerative Colitis Mucosa Reveals Altered Expression of Pathways Enriched in Genetic Susceptibility Loci

Human colonic mucosa altered by inflammation due to ulcerative colitis (UC) displays a drastically altered pattern of gene expression compared with healthy tissue. We aimed to understand the underlying molecular pathways influencing these differences by analyzing three publically-available, independently-generated microarray datasets of gene expression from endoscopic biopsies of the colon. Gene set enrichment analysis (GSEA) revealed that all three datasets share 87 gene sets upregulated in UC lesions and 8 gene sets downregulated (false discovery rate <0.05). The upregulated pathways were dominated by gene sets involved in immune function and signaling, as well as the control of mitosis. We applied pathway analysis to genotype data derived from genome-wide association studies (GWAS) of UC, consisting of 5,584 cases and 11,587 controls assembled from eight European-ancestry cohorts. The upregulated pathways derived from the gene expression data showed a highly significant overlap with pathways derived from the genotype data (33 of 56 gene sets, hypergeometric P = 1.49×10^–19). This study supports the hypothesis that heritable variation in gene expression as measured by GWAS signals can influence key pathways in the development of disease, and that comparison of genetic susceptibility loci with gene expression signatures can differentiate key drivers of inflammation from secondary effects on gene expression of the inflammatory process.