Highly efficient genome editing in primary bronchial epithelial cells establishes FOXJ1 as essential for ciliation in human airways

The structure and composition of the bronchial epithelium is altered in respiratory diseases such as COPD and asthma, in which goblet cell hyperplasia and reduced numbers of ciliated cells impair mucociliary clearance. We describe a robust genome editing pipeline to interrogate modulators of primary human bronchial epithelial cell (HBEC) differentiation and function. By employing plasmid- and virus-free delivery of CRISPR/Cas9 to human airway basal cells we achieve highly efficient gene inactivation without the need for positive selection. Genome edited cells are differentiated at air liquid interface (ALI) into a pseudo-stratified epithelium. We focus on profiling ciliation using imaging cytometry coupled to confocal microscopy and immunohistochemistry. To our knowledge, this is the first study to describe highly efficient genome editing of ALI cultured primary HBECs. As proof of concept, we establish that inactivation of the gene encoding the transcription factor FOXJ1 in primary human airway basal cells precludes ciliation in ALI differentiated bronchial epithelia.