Notch-Dependent Differentiation of Adult Airway Basal Stem Cells

Abstract

Summary The epithelium lining the airways of the adult human lung is composed of ciliated and secretory cells together with undifferentiated basal cells (BCs). The composition and organization of this epithelium is severely disrupted in many respiratory diseases. However, little is known about the mechanisms controlling airway homeostasis and repair after epithelial damage. Here, we exploit the mouse tracheobronchial epithelium, in which BCs function as resident stem cells, as a genetically tractable model of human small airways. Using a reporter allele we show that the low level of Notch signaling at steady state is greatly enhanced during repair and the generation of luminal progenitors. Loss-of-function experiments show that Notch signaling is required for the differentiation, but not self-renewal, of BCs. Moreover, sustained Notch activation in BCs promotes their luminal differentiation, primarily towards secretory lineages. We also provide evidence that this function of Notch signaling is conserved in BCs from human airways. Notch signaling is active in steady state airways and increased during repairNotch is required for differentiation, but not self-renewal, of airway basal cellsNotch promotes luminal differentiation of mouse basal stem cellsOur data suggest this mechanism is conserved in human basal cells

Cx26 regulates proliferation of repairing basal airway epithelial cells

“…One interpretation of these results is that the expression of Cx26 is triggered in daughter cells of an expanding, but not yet differentiating, population of BCs, in response to proliferative signals generated by the injured epithelium. Cx26 was not detected in CK8-positive cells, which represent early progenitors of SC and CC lineages (Rock et al, 2011), but is present in a population of CK14-positive HAECs. An early report in the rat tracheal epithelium and later mouse geneticlineage tracing studies showed that the proportion of CK14-expressing BCs transiently increased following epithelial damage, suggesting that CK14 is up-regulated when BCs are activated (Hong et al, 2004;Ghosh et al, 2010).…”

“…Typical markers of BCs are the transcription factor Trp63 (or p63), cytokeratin (CK)5 and CK14. On the other hand, CK8 is a marker of well-differentiated CCs and SCs that is also detected in early progenitors of these cell lineages during repair (Beers, Morrissey, 2011;Rock et al, 2011;Wansleeben et al, 2013). The mechanisms synchronizing proliferation and differentiation of activated BCs within the repairing human airway epithelium remain, however, poorly known.…”

GRHL2 coordinates regeneration of a polarized mucociliary epithelium from basal stem cells

“…Specifically, recent studies have implicated Notch3 in promoting the differentiation of adult basal stem cells into parabasal progenitors and Notch1 and 2 in fate selection between secretory and ciliated linages (Rock et al, 2011b;Danahay et al, 2015;Mori et al, 2015;Pardo-Saganta et al, 2015). ChIP-Seq has shown Grhl2 occupancy of binding sites near NOT CH1, NOT CH3, JAG1, and JAG2 (Gao et al, 2013).…”

“…Samples were embedded in paraffin, sectioned at 7 µm, and stained as described previously (Rock et al, 2011b). For whole-mount staining of ALI culture, after fixation and washing with PBS, the membrane was cut from the insert and stained by standard protocols.…”

“…At steady state, these basal cells (BCs) are relatively quiescent, but after a loss of luminal cells they proliferate and differentiate into ciliated and secretory cells (Borthwick et al, 2001;Hogan et al, 2014). Considerable progress is being made in understanding the transcriptional circuits and signaling pathways that regulate the self-renewal of BCs and the lineage choices of their descendants (Rock et al, 2011b;Tata et al, 2013;Paul et al, 2014;Tadokoro et al, 2014;Zhao et al, 2014;Danahay et al, 2015;Mori et al, 2015;Pardo-Saganta et al, 2015;Watson et al, 2015). However, less is known about the cellular mechanisms that drive morphological changes in the daughters of the BCs as they acquire apical-basal polarity and integrate seamlessly into the epithelium.…”