Investigating the role of NKX2-1 in human lung development
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Citation
Abstract
NK2 Homeobox 1 (NKX2-1) is a critically important transcription factor in lung development, with all lung epithelia being derived from an NKX2-1+ progenitor pool. Nkx2-1-/- mice have severely hypoplastic lungs and tracheoesophageal fistula. Loss of NKX2-1 in the context of lung adenocarcinoma results in fate loss and is associated with progression to metastasis. Humans with heterozygous NKX2-1 mutations develop respiratory insufficiency, hypothyroidism, and neurological problems, a disease known broadly as brain-lung-thyroid syndrome. Despite its known importance, the role of NKX2-1 in human development is not fully defined. Here, we use an induced pluripotent stem cell (iPSC) model of early lung development, patient-derived iPSC lines with NKX2-1 mutations, and clustered short interspersed short palindromic repeats (CRISPR) gene editing to study NKX2-1’s role in development. Using this platform, we identified broad transcriptomic differences between mutant and CRISPR-corrected cells that increasingly diverged over developmental time, with mutant alveolar cells failing to properly induce alveolar type 2 (ATII) cell fate, while also upregulating non-lung endodermal markers, of which many are NKX2-1 genomic target sites. Based on the finding that we saw more transcriptomic differences in alveolar versus airway differentiations, we hypothesized that two functioning copies of NKX2-1 are needed for proper alveolar, but not airway development. We performed in vivo competition assays between Nkx2-1 heterozygous and wild-type cells via blastocyst complementation, revealing Nkx2-1 heterozygous cells are at a competitive disadvantage in alveolar, but not airway development. Next, we identified aberrant transcription factor activity consequential to aberrant NKX2-1 activity, highlighting how crosstalk between transcription factors can influence alveolar development. Of note, we identified aberrant mothers against decapentaplegic homolog 3 (SMAD3) activity in mutant cells and were able to rescue gene expression of some NKX2-1 targets via treatment with a transforming growth factor beta (TGFβ) pathway inhibitor. In conclusion, these findings significantly advance knowledge of a critically important transcription factor in human lung development. Future directions will further tease apart NKX2-1’s crosstalk with other developmentally important pathways and seek to identify the role NKX2-1 plays in chromatin remodeling.
Description
2024