Siddiqui S*1,2, Johansson K*1,2,3, Joo A1,2, Bonser LR4,5, Koh KD4,5, Le Tonqueze O4,5, Bolourchi S1,2, Bautista RA1,2, Zlock L6, Roth TL3,7,8,9, Marson A3,9,10,11,12,13, Bhakta NR1, Ansel KM2,3, Finkbeiner WE6, Erle DJ4,5, Woodruff PG1,2,5
1 Department of Medicine, Division of Pulmonary and Critical Care Medicine, 2Sandler Asthma Basic Research Center, 3Department of Microbiology and Immunology, 4Lung Biology Center, 5Cardiovascular Research Institute, 6Department of Pathology, 7Biomedical Sciences Graduate Program, 8Diabetes Center, 9Innovative Genomics Institute, University of California, Berkeley, 10J. David Gladstone Institutes, 11Department of Medicine, Division of Infectious Diseases, 12Parker Institute for Cancer Immunotherapy and 13Chan Zuckerberg Biohub, San Francisco, CA, USA
* Equal contribution
Rational: Interleukin (IL)-13-induced goblet cell metaplasia contributes to airway remodeling and pathological mucus hypersecretion in asthma. MicroRNAs (miRNAs) are a distinct class of noncoding RNAs, about 20-22 nucleotides long, that mediate sequence-specific repression of target mRNAs. Cellular responses are modulated by miRNAs but their role in mucus regulation is largely unexplored. We hypothesized that airway epithelial miRNAs play a role in IL-13-induced mucus regulation.
Methods: We performed CRISPR/Cas9-editing of primary human bronchial epithelial cells (HBECs) to target a specific miRNA candidate, miR-141, by delivery of MIR141-targeting guide RNAs (gRNA) via electroporation. HBECs that received MIR141 gRNAs or non-targeting gRNA control were differentiated at air-liquid-interface (ALI) and epithelial mucus was induced by IL-13 stimulation.
Results: miR-141, a member of the miR-141/200 family, was identified as one of the most highly expressed miRNAs in human airway epithelium by miRNA sequencing. Analysis of bronchial brushings from asthmatic subjects revealed that miR-141 is reduced at baseline in asthma but is induced shortly after airway allergen challenge. Knock down of miR-141 resulted in decreased goblet cells frequency, intracellular MUC5AC and total secreted mucus. These effects correlated with a reduction in a goblet cell gene expression signature and enrichment of a basal cell gene expression signature defined by single cell RNA sequencing. Furthermore, intranasal administration of a sequence-specific miR-141 inhibitor in mice decreased Aspergillus-induced secreted mucus and mucus-expressing cells in the lung, and reduced airway hyper-responsiveness without affecting cellular inflammation.
Conclusions: We have identified a miRNA that regulates pathological airway mucus production in human cells and in mice and is amenable to therapeutic manipulation through an inhaled route.