Genome-wide association studies (GWAS) have implicated the IL33 locus in asthma, but the underlying mechanisms remain unclear. Here, we identify a 5 kb region within the GWAS-defined segment that acts as an enhancer-blocking element in vivo and in vitro. Chromatin conformation capture showed that this 5 kb region loops to the IL33 promoter, potentially regulating its expression. We show that the asthma-associated single nucleotide polymorphism (SNP) rs1888909, located within the 5 kb region, is associated with IL33 gene expression in human airway epithelial cells and IL-33 protein expression in human plasma, potentially through differential binding of OCT-1 (POU2F1) to the asthma-risk allele. Our data demonstrate that asthma-associated variants at the IL33 locus mediate allele-specific regulatory activity and IL33 expression, providing a mechanism through which a regulatory SNP contributes to genetic risk of asthma.

Background: Genome-wide association studies of asthma have revealed robust associations with variation across the human leukocyte antigen (HLA) complex with independent associations in the HLA class I and class II regions for both childhood-onset asthma (COA) and adult-onset asthma (AOA). However, the specific variants and genes contributing to risk are unknown.Methods: We used Bayesian approaches to perform genetic fine-mapping for COA and AOA (n=9432 and 21,556, respectively;n=318,167 shared controls) in White British individuals from the UK Biobank and to perform expression quantitative trait locus (eQTL) fine-mapping in immune (lymphoblastoid cell lines,n=398; peripheral blood mononuclear cells,n=132) and airway (nasal epithelial cells,n=188) cells from ethnically diverse individuals. We also examined putatively causal protein coding variation from protein crystal structures and conducted replication studies in independent multi-ethnic cohorts from the UK Biobank (COAn=1686; AOAn=3666; controlsn=56,063).Results: Genetic fine-mapping revealed both shared and distinct causal variation between COA and AOA in the class I region but only distinct causal variation in the class II region. Both gene expression levels and amino acid variation contributed to risk. Our results from eQTL fine-mapping and amino acid visualization suggested that theHLA-DQA1*03:01 allele and variation associated with expression of the nonclassicalHLA-DQA2andHLA-DQB2genes accounted entirely for the most significant association with AOA in GWAS. Our studies also suggested a potentially prominent role for HLA-C protein coding variation in the class I region in COA. We replicated putatively causal variant associations in a multi-ethnic cohort.Conclusions: We highlight roles for both gene expression and protein coding variation in asthma risk and identified putatively causal variation and genes in the HLA region. A convergence of genomic, transcriptional, and protein coding evidence implicates theHLA-DQA2andHLA-DQB2genes andHLA-DQA1*03:01 allele in AOA.

Following corneal injury, coordinated cellular and protein interactions occur at the wound site to restore tissue homeostasis. Regulation of this response is required to prevent the development of chronic inflammation, abnormal neovascularization, and fibrosis. The chemokine CCL2 and its primary receptor CCR2 are key regulators of the inflammatory and neovascular responses to injury. In this study, we investigated the role of macrophage-associated matrix metalloproteinase 12 (MMP12) in the regulation of CCL2 and CCR2 after corneal wounding. Using two corneal injury models, we examined the temporal and spatial expression of CCL2 and CCR2 in Mmp12^−/− and wild-type (WT) mice. Our data showed that MMP12 downregulated CCL2 and CCR2 expression in a manner dependent on the timing and mechanism of injury. We also examined the effect of CCL2 on the injury response in Mmp12^−/− and WT corneas. We found that macrophage infiltration and neovascularization following CCL2 blockade was significantly reduced in Mmp12^−/− corneas as compared with WT corneas. These findings indicate that MMP12 inhibits corneal inflammation and neovascularization after injury through its regulation of CCL2.

Most genetic studies of asthma and allergy have focused on common variation in individuals primarily of European ancestry. Studying the role of rare variation in quantitative phenotypes and in asthma phenotypes in populations of diverse ancestries can provide additional, important insights into the development of these traits. We sought to examine the contribution of rare variants to different asthma- or allergy-associated quantitative traits in children with diverse ancestries and explore their role in asthma phenotypes. We examined whole-genome sequencing data from children participants in longitudinal studies of asthma (n = 1035; parent-identified as 67% Black and 25% Hispanic) to identify rare variants (minor allele frequency < 0.01). We assigned variants to genes and tested for associations using an omnibus variant-set test between each of 24,902 genes and 8 asthma-associated quantitative traits. On combining our results with external data on predicted gene expression in humans and mouse knockout studies, we identified 3 candidate genes. A burden of rare variants in each gene and in a combined 3-gene score was tested for its associations with clinical phenotypes of asthma. Finally, published single-cell gene expression data in lower airway mucosal cells after allergen challenge were used to assess transcriptional responses to allergen. Rare variants in USF1 were significantly associated with blood neutrophil count (P = 2.18 × 10^-7); rare variants in TNFRSF21 with total IgE (P = 6.47 × 10^-6) and PIK3R6 with eosinophil count (P = 4.10 × 10^-5) reached suggestive significance. These 3 findings were supported by independent data from human and mouse studies. A burden of rare variants in TNFRSF21 and in a 3-gene score was associated with allergy-related phenotypes in cohorts of children with mild and severe asthma. Furthermore, TNFRSF21 was significantly upregulated in bronchial basal epithelial cells from adults with allergic asthma but not in adults with allergies (but not asthma) after allergen challenge. We report novel associations between rare variants in genes and allergic and inflammatory phenotypes in children with diverse ancestries, highlighting TNFRSF21 as contributing to the development of allergic asthma.

Mononuclear phagocytes (MP) have central importance in innate immunity, inflammation, and fibrosis. Recruited MPs, such as macrophages, are plastic cells and can switch from an inflammatory to a restorative phenotype during the healing process. However, the role of the MPs in corneal wound healing is not completely understood. The purpose of this study is to characterize the kinetics of recruited MPs and evaluate the role of macrophage metalloelastase (MMP12) in the healing process, using an in vivo corneal chemical injury model. Unwounded and wounded corneas of wild-type (WT) and Mmp12^−/− mice were collected at 1, 3, and 6 days after chemical injury and processed for flow cytometry analysis. Corneal MP phenotype significantly changed over time with recruited Ly6C^high (proinflammatory) cells being most abundant at 1 day post-injury. Ly6C^int cells were highly expressed at 3 days post-injury and Ly6C^neg (patrolling) cells became the predominant cell type at 6 days post-injury. CD11c⁺ dendritic cells were abundant in corneas from Mmp12^−/− mice at 6 days post-injury. These findings show the temporal phenotypic plasticity of recruited MPs and provide valuable insight into the role of the MPs in the corneal repair response, which may help guide the future development of MP-targeted therapies.