Regulating transcription of the key pulmonary neuroendocrine lineage driver ASCL1

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The bHLH transcription factor ASCL1 (HASH1/MASH1) is essential for specification of the neuroendocrine (NE) lineage in the lung. NE cells are rare in pulmonary epithelium but they are thought to play diverse roles during lung development and homeostasis. Increases in NE cell number have been documented in many lung diseases in children and adults. ASCL1 function is balanced with NOTCH signaling to control progenitor proliferation and NE differentiation versus other cell types in the lung. Consistent with the role of ASCL1 as a lineage-specifying factor and in expanding progenitor populations, ASCL1 has been found to be a lineage-dependent oncogene in pulmonary NE tumors as well. With these important functions attributed to ASCL1, and its requirement for controlled spatial and temporal expression during development and in conditions of lung damage, it is surprising how little is known about regulation of ASCL1 gene transcription. This gap in knowledge reflects past technical challenges in identifying and manipulating cis-regulatory elements (REs) found at large distances from the gene of interest, in addition to challenges of poor culture systems reflecting this NE population and the low fraction of NE cells in embryonic and adult lungs. REs functioning at long-distances to control key developmental genes are being discovered using advances in technologies that can interrogate and manipulate the spatial genome. Here we will exploit these technologies to gain much needed insights into transcriptional control of ASCL1 using newly reported protocols for generating pulmonary NE cells from human iPSCs and leveraging cell culture models of neuroendocrine cancer that express high levels of ASCL1. Each model has a particular strength that allows unique aspects of ASCL1 regulation to be uncovered. Aims include identifying and testing functions of long-range REs controlling ASCL1 during NE differentiation in the two models. Success in these aims will provide functional non-coding regulatory sequences controlling ASCL1 expression, and presumably, the NE lineage in lung development and homeostasis. This is important for future projects to identify molecular components of the signaling complexes working through these REs to reach the goal of providing an understanding of how a key lineage defining transcriptional regulator is controlled during development and disease.