PROJECT SUMMARY The enteric nervous system (ENS) plays a vital role in gastrointestinal biology by regulating gut motility, secretion, and absorption, and by interacting with the immune system. Dysregulation of the ENS thus contributes to many diseases, including the inflammatory bowel diseases (IBDs) ulcerative colitis (UC) and Crohn’s disease (CD). IBDs are associated with increased density of enteric neurons and activation of enteric glial cells, and existing evidence suggests that these effects directly contribute to the pathobiology of IBDs. Understanding the role of ENS components in UC and CD is thus critical for developing targeted therapies. Recent papers both from our group and from other laboratories have demonstrated that some ENS glial cells switch fates to become neurons in response to inflammation. The ENS glia thus contain a subset of neuronal progenitor cells, and inflammatory signals trigger this population to generate new neurons. However, it is not clear how glial cells alter their transcriptional program to generate neurons, and what subgroups of glia are involved in this process is unknown. The goal of this project is to test the hypothesis that specific transcription factors with known roles in prenatal neurogenesis induce postnatal neurogenesis in the ENS in response to inflammation. Phox2b is a transcription factor with critical roles in both ENS and central nervous system neurogenesis, and preliminary data shows that it induces cultured glia to adopt a neuronal phenotype. I hypothesize that colonic inflammation will trigger Phox2b-mediated neuronal fate acquisition in a subset of enteric glial cells. To test this hypothesis, I will use single cell RNA sequencing and a conditional Phox2b knockout mouse model. Completion of this project will significantly enhance our understanding of enteric glia in the pathophysiology of IBDs, and will reveal molecular pathways that may represent therapeutic targets. This work will also reveal glial subpopulations with potential to be harnessed therapeutically to regenerate missing neurons. In addition to the proposed research, this fellowship will provide training to plan, execute, and analyze data from experiments utilizing RNA sequencing techniques, to breed mice to generate tissue-specific conditional knockout models, to culture and manipulate primary enteric glial cells, to effectively communicate scientific data in writing and orally, and to manage and lead a scientific project. Upon completion of the fellowship, I will be prepared to launch my own career as an independent, NIH-funded scientist studying gastrointestinal biology.