Single cell transcriptomic atlases of the mouse and human lungs reveal new insights into pulmonary diseases

Since the advent of cell theory in the mid-19th century, scientists have used microscopy to document hundreds of cell types' positions, structures, and niches across our bodies and those of other multicellular organisms. Concurrently, molecular functions were identified for roughly half of the proteins encoded by our genes, which keep cells (and by extension us) alive and healthy. Each cell type uses different sets of genes to perform their essential functions, but mapping which cells use which genes has, historically, been done one cell type and/or one gene or pathway at a time. That changed a decade ago, when we captured the full gene expression of a single cell, and we can now profile hundreds of thousands to millions of cells simultaneously in a single experiment. Although single cell RNA sequencing (scRNAseq) studies have begun providing compendia of cell expression profiles, it has proven more difficult to systematically identify and localize all molecular types in individual organs to create a full molecular cell atlas and to elucidate their molecular functions. This is particularly important for the lung because diseases affecting it are the leading cause of both morbidity and mortality in the United States. In this dissertation, I use droplet- and plate-based scRNAseq applied to ~36,000 mouse cells and ~75,000 human cells across all lung tissue compartments and, combined with a multi-pronged cell annotation approach, define the gene expression profiles and anatomical locations of 40 and 58 cell populations in the mouse and human lungs, respectively, including 41 of 45 previously known human cell types or subtypes and 14 new ones. These comprehensive molecular atlases elucidate the biochemical functions of lung cell types and the cell-selective transcription factors and optimal markers for making and monitoring them; defines the cell targets of circulating hormones and predicts local signaling interactions; and identifies the cell types directly affected by lung disease genes and respiratory viruses. Comparison of the mouse and human atlases identifies 17 molecular types that appear to have been gained or lost during lung evolution and others whose expression profiles have been substantially altered, revealing extensive plasticity of cell types and gene expression. Stromal cells lie beneath epithelial and endothelial cells and help give them shape in development, but their developmental origin, lineage hierarchy, molecular functions, and disease relevance in the lung are understudied compared to their epithelial, endothelial, and immune cell counterparts. In this dissertation, I also leverage existing atlases of developing mouse and human lungs to identify distinct molecular trajectories that give rise to each adult stromal cell type, which include transcription factors and signaling receptors that could serve as master regulators in specifying their fate. I create a mouse line to mark and manipulate one stromal type, pericytes, and discover another line to mark and manipulate another type, myofibroblasts, and use lineage tracing and clonal analysis with both to identify a subset of pericytes that are proliferative but restricted in their fate and to discover that myofibroblasts, long thought proliferative in response to injury, are actually quiescent in adulthood. Finally, I end by implicating pericytes as the cell of origin in a familial form of pulmonary hypertension and by revealing that myofibroblasts have the capacity to respond to destruction of alveolar entrance rings in adult, part of emphysema's pathological course. Both discoveries shed new light on pulmonary diseases that are refractory to treatment and have poor prognoses and could lead to the rational discovery of new, more effective treatments

Keywords: Biochemistry.
Advisor: Krasnow Mark
Artandi Steven E.
Beachy Philip Arden
Harbury Pehr