Lung tuft cell transcriptomics

While the lung bears significant regenerative capacity, severe viral pneumonia can chronically impair lung function by triggering dysplastic remodeling. The connection between these enduring changes and chronic disease remains poorly understood.  We recently described the emergence of tuft cells within Krt5+ dysplastic regions after influenza injury. Using bulk and single cell transcriptomics, we characterized and delineated multiple distinct tuft cell populations that arise following influenza clearance. Distinct from intestinal tuft cells which rely on Type 2 immune signals for their expansion, neither IL-25 nor IL-4Ra signaling are required to drive tuft cell development in dysplastic/injured lungs. Furthermore, tuft cells were also observed upon bleomycin injury, suggesting that their development may be a general response to severe lung injury. While intestinal tuft cells promote growth and differentiation of surrounding epithelial cells, in the lungs of tuft cell deficient mice, Krt5+ dysplasia still occurs, goblet cell production is unchanged, and there remains no appreciable contribution of Krt5+ cells into more regionally appropriate alveolar Type 2 cells. Together, these findings highlight unexpected differences in signals necessary for lung tuft cell amplification and establish a framework for future elucidation of tuft cell functions in pulmonary health and disease. Bulk seq. 1000-3000 EpCam+ Trpm5-GFP+ cells from mice at day 43 post-influenza were sorted directly into lysis buffer from Takara SMART-Seq v4 and RNA / cDNA was amplified according to manufacturer’s instructions. All downstream library preparation and sequencing was performed by the Next-Generation Sequencing Core at the Perelman School of Medicine, University of Pennsylvania. In brief, libraries were sequenced on a NovaSeq sequencer at 100SR, raw FASTQ files were imported into R and reads mapped with Kallisto, and differential expression performed by Limma. Raw data is deposited at GEO.