Rationale: Coronavirus disease (COVID-19) is caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). ACE2 (angiotensin-converting enzyme 2), and TMPRSS2 (transmembrane protease serine 2) mediate viral infection of host cells. We reasoned that differences in ACE2 or TMPRSS2 gene expression in sputum cells among patients with asthma may identify subgroups at risk for COVID-19 morbidity.Objectives: To determine the relationship between demographic features and sputum ACE2 and TMPRSS2 gene expression in asthma.Methods: We analyzed gene expression for ACE2 and TMPRSS2, and for ICAM-1 (intercellular adhesion molecule 1) (rhinovirus receptor as a comparator) in sputum cells from 330 participants in SARP-3 (Severe Asthma Research Program-3) and 79 healthy control subjects.Measurements and Main Results: Gene expression of ACE2 was lower than TMPRSS2, and expression levels of both genes were similar in asthma and health. Among patients with asthma, male sex, African American race, and history of diabetes mellitus were associated with higher expression of ACE2 and TMPRSS2. Use of inhaled corticosteroids (ICS) was associated with lower expression of ACE2 and TMPRSS2, but treatment with triamcinolone acetonide did not decrease expression of either gene. These findings differed from those for ICAM-1, where gene expression was increased in asthma and less consistent differences were observed related to sex, race, and use of ICS.Conclusions: Higher expression of ACE2 and TMPRSS2 in males, African Americans, and patients with diabetes mellitus provides rationale for monitoring these asthma subgroups for poor COVID-19 outcomes. The lower expression of ACE2 and TMPRSS2 with ICS use warrants prospective study of ICS use as a predictor of decreased susceptibility to SARS-CoV-2 infection and decreased COVID-19 morbidity.

The type 2 cytokine-high asthma endotype (T2H) is characterized by IL-13-driven mucus obstruction of the airways. To further investigate this incompletely understood pathobiology, we characterize IL-13 effects on human airway epithelial cell cultures using single-cell RNA sequencing, finding that IL-13 generates a distinctive transcriptional state for each cell type. Specifically, we discover a mucus secretory program induced by IL-13 in all cell types which converts both mucus and defense secretory cells into a metaplastic state with emergent mucin production and secretion, while leading to ER stress and cell death in ciliated cells. The IL-13-remodeled epithelium secretes a pathologic, mucin-imbalanced, and innate immunity-depleted proteome that arrests mucociliary motion. Signatures of IL-13-induced cellular remodeling are mirrored by transcriptional signatures characteristic of the nasal airway epithelium within T2H versus T2-low asthmatic children. Our results reveal the epithelium-wide scope of T2H asthma and present candidate therapeutic targets for restoring normal epithelial function.

Background

Global gene expression levels are known to be highly dependent upon gross demographic features including age, yet identification of age-related genomic indicators has yet to be comprehensively undertaken in a disease and treatment-specific context.

Methods

We used gene expression data from CD4+ lymphocytes in the Asthma BioRepository for Integrative Genomic Exploration (Asthma BRIDGE), an open-access collection of subjects participating in genetic studies of asthma with available gene expression data. Replication population participants were Puerto Rico islanders recruited as part of the ongoing Genes environments & Admixture in Latino Americans (GALA II), who provided nasal brushings for transcript sequencing. The main outcome measure was chronic asthma control as derived by questionnaires. Genomic associations were performed using regression of chronic asthma control score on gene expression with age in years as a covariate, including a multiplicative interaction term for gene expression times age.

Results

The SMARCD1 gene (SWI/SNF-related matrix-associated actin-dependent regulator of chromatin subfamily D member 1) interacted with age to influence chronic asthma control on inhaled corticosteroids, with a doubling of expression leading to an increase of 1.3 units of chronic asthma control per year (95% CI [0.86, 1.74], p = 6 × 10^- 9), suggesting worsening asthma control with increasing age. This result replicated in GALA II (p = 3.8 × 10^- 8). Cellular assays confirmed the role of SMARCD1 in glucocorticoid response in airway epithelial cells.

Conclusion

Focusing on age-dependent factors may help identify novel indicators of asthma medication response. Age appears to modulate the effect of SMARCD1 on asthma control with inhaled corticosteroids.

Family-based methods are a potentially powerful tool to identify trait-defining genetic variants in extended families, particularly when used to complement conventional association analysis. We utilized two-point linkage analysis and single variant association analysis to evaluate whole exome sequencing (WES) data from 1205 Hispanic Americans (78 families) from the Insulin Resistance Atherosclerosis Family Study. WES identified 211,612 variants above the minor allele frequency threshold of ≥0.005. These variants were tested for linkage and/or association with 50 cardiometabolic traits after quality control checks. Two-point linkage analysis yielded 10,580,600 logarithm of the odds (LOD) scores with 1148 LOD scores ≥3, 183 LOD scores ≥4, and 29 LOD scores ≥5. The maximal novel LOD score was 5.50 for rs2289043:T>C, in UNC5C with subcutaneous adipose tissue volume. Association analysis identified 13 variants attaining genome-wide significance (P < 5 × 10^-08 ), with the strongest association between rs651821:C>T in APOA5 and triglyceride levels (P  =  3.67 × 10^-10 ). Overall, there was a 5.2-fold increase in the number of informative variants detected by WES compared to exome chip analysis in this population, nearly 30% of which were novel variants relative to the Database of Single Nucleotide Polymorphisms (dbSNP) build 138. Thus, integration of results from two-point linkage and single-variant association analysis from WES data enabled identification of novel signals potentially contributing to cardiometabolic traits.

Air pollution particulate matter <2.5 μm (PM_2.5) exposure is associated with poor respiratory outcomes. Mechanisms underlying PM_2.5-induced lung pathobiology are poorly understood but likely involve cellular and molecular changes to the airway epithelium. We extracted and chemically characterized the organic and water-soluble components of air pollution PM_2.5 samples, then determined the whole transcriptome response of human nasal mucociliary airway epithelial cultures to a dose series of PM_2.5 extracts. We found that PM_2.5 organic extract (OE), but not water-soluble extract, elicited a potent, dose-dependent transcriptomic response from the mucociliary epithelium. Exposure to a moderate OE dose modified the expression of 424 genes, including activation of aryl hydrocarbon receptor signaling and an IL-1 inflammatory program. We generated an OE-response gene network defined by eight functional enrichment groups, which exhibited high connectivity through CYP1A1, IL1A, and IL1B. This OE exposure also robustly activated a mucus secretory expression program (>100 genes), which included transcriptional drivers of mucus metaplasia (SPDEF and FOXA3). Exposure to a higher OE dose modified the expression of 1,240 genes and further exacerbated expression responses observed at the moderate dose, including the mucus secretory program. Moreover, the higher OE dose significantly increased the MUC5AC/MUC5B gel-forming mucin expression ratio and strongly downregulated ciliated cell expression programs, including key ciliating cell transcription factors (e.g., FOXJ1 and MCIDAS). Chronic OE stimulation induced mucus metaplasia-like remodeling characterized by increases in MUC5AC⁺ secretory cells and MUC5AC mucus secretions. This epithelial remodeling may underlie poor respiratory outcomes associated with high PM_2.5 exposure.

Linkage studies of complex genetic diseases have been largely replaced by genome-wide association studies, due in part to limited success in complex trait discovery. However, recent interest in rare and low-frequency variants motivates re-examination of family-based methods. In this study, we investigated the performance of two-point linkage analysis for over 1.6 million single-nucleotide polymorphisms (SNPs) combined with single variant association analysis to identify high impact variants, which are both strongly linked and associated with cardiometabolic traits in up to 1414 Hispanics from the Insulin Resistance Atherosclerosis Family Study (IRASFS). Evaluation of all 50 phenotypes yielded 83 557 000 LOD (logarithm of the odds) scores, with 9214 LOD scores ⩾3.0, 845 ⩾4.0 and 89 ⩾5.0, with a maximal LOD score of 6.49 (rs12956744 in the LAMA1 gene for tumor necrosis factor-α (TNFα) receptor 2). Twenty-seven variants were associated with P<0.005 as well as having an LOD score >4, including variants in the NFIB gene under a linkage peak with TNFα receptor 2 levels on chromosome 9. Linkage regions of interest included a broad peak (31 Mb) on chromosome 1q with acute insulin response (max LOD=5.37). This region was previously documented with type 2 diabetes in family-based studies, providing support for the validity of these results. Overall, we have demonstrated the utility of two-point linkage and association in comprehensive genome-wide array-based SNP genotypes.

Rationale: Puerto Ricans have the highest childhood asthma prevalence in the United States (23.6%); however, the etiology is uncertain.Objectives: In this study, we sought to uncover the genetic architecture of lung function in Puerto Rican youth with and without asthma who were recruited from the island (n = 836).Methods: We used admixture-mapping and whole-genome sequencing data to discover genomic regions associated with lung function. Functional roles of the prioritized candidate SNPs were examined with chromatin immunoprecipitation sequencing, RNA sequencing, and expression quantitative trait loci data.Measurements and Main Results: We discovered a genomic region at 1q32 that was significantly associated with a 0.12-L decrease in the lung volume of exhaled air (95% confidence interval, -0.17 to -0.07; P = 6.62 × 10^-8) with each allele of African ancestry. Within this region, two SNPs were expression quantitative trait loci of TMEM9 in nasal airway epithelial cells and MROH3P in esophagus mucosa. The minor alleles of these SNPs were associated with significantly decreased lung function and decreased TMEM9 gene expression. Another admixture-mapping peak was observed on chromosome 5q35.1, indicating that each Native American ancestry allele was associated with a 0.15-L increase in lung function (95% confidence interval, 0.08-0.21; P = 5.03 × 10^-6). The region-based association tests identified four suggestive windows that harbored candidate rare variants associated with lung function.Conclusions: We identified common and rare genetic variants that may play a critical role in lung function among Puerto Rican youth. We independently validated an inflammatory pathway that could potentially be used to develop more targeted treatments and interventions for patients with asthma.

To identify genetic determinants of airway dysfunction, we performed a transcriptome-wide association study for asthma by combining RNA-seq data from the nasal airway epithelium of 681 children, with UK Biobank genetic association data. Our airway analysis identified 95 asthma genes, 58 of which were not identified by transcriptome-wide association analyses using other asthma-relevant tissues. Among these genes were MUC5AC, an airway mucin, and FOXA3, a transcriptional driver of mucus metaplasia. Muco-ciliary epithelial cultures from genotyped donors revealed that the MUC5AC risk variant increases MUC5AC protein secretion and mucus secretory cell frequency. Airway transcriptome-wide association analyses for mucus production and chronic cough also identified MUC5AC. These cis-expression variants were associated with trans effects on expression; the MUC5AC variant was associated with upregulation of non-inflammatory mucus secretory network genes, while the FOXA3 variant was associated with upregulation of type-2 inflammation-induced mucus-metaplasia pathway genes. Our results reveal genetic mechanisms of airway mucus pathobiology.

Coronavirus disease 2019 (COVID-19) outcomes vary from asymptomatic infection to death. This disparity may reflect different airway levels of the SARS-CoV-2 receptor, ACE2, and the spike protein activator, TMPRSS2. Here we explore the role of genetics and co-expression networks in regulating these genes in the airway, through the analysis of nasal airway transcriptome data from 695 children. We identify expression quantitative trait loci (eQTL) for both ACE2 and TMPRSS2, that vary in frequency across world populations. Importantly, we find TMPRSS2 is part of a mucus secretory network, highly upregulated by T2 inflammation through the action of interleukin-13, and that interferon response to respiratory viruses highly upregulates ACE2 expression. Finally, we define airway responses to coronavirus infections in children, finding that these infections upregulate IL6 while also stimulating a more pronounced cytotoxic immune response relative to other respiratory viruses. Our results reveal mechanisms likely influencing SARS-CoV-2 infectivity and COVID-19 clinical outcomes.

Baseline lung function, quantified as forced expiratory volume in the first second of exhalation (FEV₁), is a standard diagnostic criterion used by clinicians to identify and classify lung diseases. Using whole-genome sequencing data from the National Heart, Lung, and Blood Institute Trans-Omics for Precision Medicine project, we identified a novel genetic association with FEV₁ on chromosome 12 in 867 African American children with asthma (P = 1.26 × 10^-8, β = 0.302). Conditional analysis within 1 Mb of the tag signal (rs73429450) yielded one major and two other weaker independent signals within this peak. We explored statistical and functional evidence for all variants in linkage disequilibrium with the three independent signals and yielded nine variants as the most likely candidates responsible for the association with FEV₁ Hi-C data and expression QTL analysis demonstrated that these variants physically interacted with KITLG (KIT ligand, also known as SCF), and their minor alleles were associated with increased expression of the KITLG gene in nasal epithelial cells. Gene-by-air-pollution interaction analysis found that the candidate variant rs58475486 interacted with past-year ambient sulfur dioxide exposure (P = 0.003, β = 0.32). This study identified a novel protective genetic association with FEV₁, possibly mediated through KITLG, in African American children with asthma. This is the first study that has identified a genetic association between lung function and KITLG, which has established a role in orchestrating allergic inflammation in asthma.