The genome-wide association study (GWAS)-identified asthma susceptibility risk alleles on chromosome 17q21 increase the expression of ORMDL3 (ORMDL sphingolipid biosynthesis regulator 3) in lung tissue. Given the importance of epithelial integrity in asthma, we hypothesized that ORMDL3 directly impacted bronchial epithelial function. To determine whether and how ORMDL3 expression impacts the bronchial epithelium, in studies using both primary human bronchial epithelial cells and human bronchial epithelial cell line, 16HBE (16HBE14o-), we assessed the impact of ORMDL3 on autophagy. Studies included: autophagosome detection by electron microscopy, RFP-GFP-LC3B to assess autophagic activity, and Western blot analysis of autophagy-related proteins. Mechanistic assessments included immunoprecipitation assays, intracellular calcium mobilization assessments, and cell viability assays. Coexpression of ORMDL3 and autophagy-related genes was measured in primary human bronchial epithelial cells derived from 44 subjects. Overexpressing ORMDL3 demonstrated increased numbers of autophagosomes and increased levels of autophagy-related proteins LC3B, ATG3, ATG7, and ATG16L1. ORMDL3 overexpression promotes autophagy and subsequent cell death by impairing intracellular calcium mobilization through interacting with SERCA2. Strong correlation was observed between expression of ORMDL3 and autophagy-related genes in patient-derived bronchial epithelial cells. Increased ORMDL3 expression induces autophagy, possibly through interacting with SERCA2, thereby inhibiting intracellular calcium influx, and induces cell death, impairing bronchial epithelial function in asthma.
Keywords: asthma; autophagy; calcium mobilization; cell death; human bronchial epithelium.