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Clara cells (CCs) are a morphologically and operationally heterogeneous population of Secretoglobin Scgb1a1-expressing secretory cells that are crucial for airway homeostasis and post-injury repair. Analysis of the extent and origin of CC diversity are limited by knowledge of genes expressed in these cells and their precursors. To identify novel putative markers of CCs and explore the origins of CC diversity, we characterized global changes in gene expression in embryonic lungs in which CCs do not form due to conditional disruption of Notch signaling (Rbpjk CNULL ). Microarray profiling, Real Time PCR (qRT-PCR), and RNA  in situ  hybridization (ISH) identified eleven genes downregulated in the E18.5 airways of Rbpjk cnull  compared to controls, nearly half not previously known to mark CCs. ISH revealed that several genes had overlapping but distinct domains of expression of in the normal developing lung (E18.5). Notably,  Reg3g ,  Chad ,  Gabrp  and  Lrrc26  were enriched in proximal airways,  Hp  in the distal airways and  Upk3a  in clusters of cells surrounding Neuroepithelial Bodies (NEBs). Seven of the eleven genes, including  Reg3g ,  Hp , and  Upk3a , were expressed in the adult lung in CCs in a pattern similar to that observed in the developing airways. qRT-PCR-based analysis of gene expression of CCs isolated from different airway regions of B1-EGFP reporter mice corroborated the spatial enrichment in gene expression observed by ISH. Our study identifies candidate markers for CC-precursors and CCs and supports the idea that the diversification of the CC phenotype occurs already during embryonic development.

Analysis of Notch Signaling-Dependent Gene Expression in Developing Airways Reveals Diversity of Clara Cells