Wednesday, April 2, 2025
4/2/2025
Macrophage NUDT21-mediated alternative polyadenylation in lung injury - PROJECT SUMMARY ABSTRACT The main goal of this proposal is to define the role of macrophage Nudix Hydrolase 21 (NUDT21, also known as CFIm25) mediated alternative polyadenylation (APA) during acute respiratory distress syndrome (ARDS). ARDS is an inflammatory lung disease with high morbidity and mortality. While there is currently no specific therapy available, finding novel approaches to prevent or treat ARDS would be of the highest significance and impact. Our preliminary studies suggest that NUDT21-mediated APA is implicated in ARDS. APA is a post-transcriptional mechanism of RNA regulation that allows adding a poly(A) tail at different polyadenylation sites (PAS), generating transcripts with variable 3' UTR lengths. Previous studies from our group and others identified NUDT21 as a master regulator of APA whose suppression promotes 3' UTR shortening of target genes. Transcripts with shortened 3' UTR bypass microRNA-mediated suppression and thus may increase protein production. Our studies demonstrated that NUDT21 was downregulated in macrophages in both human and mouse ARDS. NUDT21 depletion in macrophages led to 3'UTR shortening and increased protein expression of genes involved in important proinflammatory pathways, and promoted cytokine releases. Functional studies with myeloid-specific Nudt21 knockout (Nudt21f/f LysmCre) mice confirmed increased lung inflammation and injury in these mice compared to LysmCre controls in murine ARDS models. At last, we found that hypoxia represses NUDT21 expression in macrophages. Further screening of potential miRNAs targeting NUDT21 indicated that miR-181a was induced under hypoxia and repressed NUDT21 expression in macrophages. Based on these findings, we hypothesize that hypoxia-mediated NUDT21 repression in macrophages exaggerates ARDS by promoting the expression of proinflammatory factors through APA. We designed three Aims to test this hypothesis: Aim 1: Demonstrate the functional role of myeloid NUDT21 in murine ARDS models. Aim 2: Determinethe mechanisms for NUDT21 repressionand assess their potential as therapeutic targets in ARDS. Aim 3: Molecular investigation of the hypoxia/NUDT21/APA axis in human ARDS lungs. Our study is significant because it will elucidate a novel mechanism for increased inflammation that could contribute to the pathogenesis of ARDS. Our innovations include demonstrating the hypoxia/miR-181a/NUDT21/APA axis as a novel proinflammatory pathway in ARDS, new resources (e.g., Nudt21 knockout and overexpression mice, and lung and macrophage samples from ARDS patients), as well as cutting-edge technologies including mass cytometry to quantitate cytokines and markers of inflammatory cells and PolyAMiner bioinformatics algorithm for identifying APA genes from single-cell RNA sequencing data. Ultimately, results from these studies will facilitate the development of novel therapeutic strategies against this deadly disease.
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