SMAD signaling in early foregut development and the basal cell program: mechanisms driving lung competence and airway maintenance
Embargo Date
2027-11-24
OA Version
Citation
Abstract
During foregut development, several domains are rapidly specified into organs that are diverse in terms of cellular composition and function, including the trachea and lungs. These respiratory airways and alveolar regions are lined with an epithelium that conducts the critical functions of gas exchange, host defense, filtration and hydration. While the respiratory epithelium is essential to human health, airway and alveolar disfunction contribute to a multitude of chronic diseases that are major contributors to death and disability worldwide. Though animal models continue to progress our understanding of respiratory epithelial development, mature cell function and disease, in-vitro culture of human cell and tissue types can broaden our investigative abilities, enabling precise chemical and physical environmental modulation. In these studies, I utilized cutting edge in-vitro technology to investigate both the early stages of human respiratory epithelial development as well as mature airway stem cell function. First, I leveraged the induced pluripotent stem cell (iPSC) directed differentiation platform to enhance our understanding of the molecular mechanisms that control early embryonic development of the gut tube endoderm, with a particular emphasis on the signals that define the presumptive lung domain. Using multifactorial design of experiment (DOE), I observed a sensitive concentration dependent role of bone morphogenic protein (BMP) on priming the foregut to a respiratory fate. RNA sequencing of iPSC derived foregut patterned with decreasing levels of BMP signaling revealed dose dependent patterning of the foregut in an anterior to posterior manner from pharynx to lung to liver. Using in-vivo and ex-vivo animal models of early development, I validated the specific anterior-posterior priming of liver, lung and pharyngeal pouch fates within iPSCs and confirmed the central role of BMP in driving this process. Second, I sought to gain insight into the molecular program of the airway basal cell (BC), the primary stem cell of the airway epithelium, with a focus on BC heterogeneity. I employed lentiviral barcoding technology on primary human BCs to understand how the transcriptional profile of BC subpopulations informs stem cell function. Barcoding uncovered significant heterogeneity in both proliferation and differentiation ability of uniquely labeled BC clones that did not correlate with their starting gene expression profile. Analysis revealed a subset of BC clones generated a large pool of daughter cells, including basal and differentiated cells. This multipotent group of clones was used to identify the most stem BCs which were enriched for key regulators of TGF-β, Notch, and Wnt signaling pathways, including Follistatin (FST), delta-like noncanonical notch ligand 2 (DLK2) and Dickkopf Wnt Signaling Pathway Inhibitor 3 (DKK3), as well as downstream targets of the Salvador-Warts-Hippo (SWH) signaling pathway, connective tissue growth factor (CTGF) and cysteine-rich angiogenic inducer 61 (CYR61). Modulation of Hippo pathway kinases large tumor suppressor kinase (LATS1/2) resulted in augmented stem cell ability ex vivo and increased expression of BC stemness markers. In summary, these studies reveal important insights into respiratory development and stem cell function that can help generate more effective iPSC derived respiratory cell types and lead to a better understanding of pathological airway remodeling in the context of disease.
Description
2025