Posttranscriptional modification in lung development and regeneration

SUMMARY Lung regeneration involves the reuse of signaling pathways that are important for organ development. Previous studies have shown that these pathways converge to regulate the transcription of downstream targets. In contrast, little is known about the modifications of the resulting transcripts. In this proposal we will fill the knowledge gap and provide initial insights into the mechanism by which posttranscriptional regulation controls lung development and regeneration. We will study how Mettl3/14 complex-mediated RNA modifications are involved in lung epithelial differentiation. Our central hypothesis is that Mettl3/14-mediated m6A deposition on transcripts including Sox9 is critical for lung development and regeneration. We will test the hypothesis with three specific aims. Aim 1 will address the mechanism by which Mettl3/14-mediated RNA modification regulates lung development. We will use a combination of mouse genetics, Nanopore, m6a-SAC-sequencing and proteomics to identify and functionally test mRNA substrates that are modified by the Mettl3/14 complex. We will also study whether Sox9 transcripts are modified by the methylation complex. Aim 2 will focus on the Mettl3/14 complex-mediated RNA modification in alveolar regeneration. We will also investigate the role of Sox9 in alveolar regeneration through Sox9 loss- and gain-of-function studies. Aim 3 will examine whether the function of posttranscriptional regulation is conserved during the differentiation of human pluripotent stem cells-derived lung progenitor cells. Combined with the findings obtained from Aim 1, Aim 3 is expected to offer candidate transcripts that are regulated by the Mettl3/14 complex across species. Together these studies will help us gain a better understanding of the normal mechanism regulating lung development, meanwhile enabling insights into alveolar regeneration following injury.