A revised airway epithelial hierarchy includes CFTR-expressing ionocytes

We combine single-cell RNA-seq and in vivo lineage tracing to study the cellular composition and hierarchy of the murine tracheal epithelium. We identify a new rare cell type, the FoxI1-positive pulmonary ionocyte; functional variations in club cells based on their proximodistal location; a distinct cell type that resides in high turnover squamous epithelial structures that we named “hillocks”; and disease-relevant subsets of tuft and goblet cells. With a new method, Pulse-Seq, we show that tuft, neuroendocrine, and ionocyte cells are continually and directly replenished by basal progenitor cells. Remarkably, the cystic fibrosis gene, CFTR, is predominantly expressed in the pulmonary ionocytes of both mouse and human. Foxi1 loss in murine ionocytes causes a loss of Cftr expression and disrupts airway fluid and mucus physiology, which are also altered in cystic fibrosis. By associating cell type-specific expression programs with key disease genes, we establish a new cellular narrative for airways disease.