Alternative polyadenylation as a genetic regulatory mechanism to bridge genome to phenome in the nervous system

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PROJECT SUMMARY Anorexia nervosa is the deadliest psychological disorder with an estimated 10% lifetime disease mortality rate, while over 1/3 of all cancer patients will die from disease-based anorexia, not the cancer itself. On the opposite end of the spectrum, over 2/3 of US adults are overweight or obese, and this number, as well as the rates of associated comorbidities such as heart disease, diabetes, and cancer, is only expected to increase in the coming years. Despite the opposite directionality of these eating disorders, dysfunctional eating in obesity and anorexia is mediated by common appetite circuitry in the central nervous system (CNS). Numerous studies have documented a coordinated and complex pattern of changes in multiple gene products in these appetite centers following periods of excessive or inadequate eating. These observations strongly suggest that the behavioral decision to eat excessively or inadequately is likely driven by a multitargeted, maladaptive genetic reprogramming process in CNS appetite centers. Thus, a core question is what global process could coordinate such changes in multiple gene products? My published studies have demonstrated that appetite changes align with changes in alternative polyadenylation (APA) in the hypothalamus. APA is a rapid, activity-dependent RNA processing mechanism that regulates mRNA transcript stability, maturation, and localization. I identified a significant APA pattern change on tissue inhibitor of metalloproteinases 2 (Timp2), a gene previously implicated in the development of an obese phenotype. Thus, I am exploring the hypothesis that Timp2 APA in the arcuate nucleus (ARC) of the hypothalamus meters the development of obesity. My proposed experiments in the F99 phase will show that 1) Timp2 mRNA is necessary for appetite control in the ARC, and that 2) APA regulation of ARC Timp2 is necessary to counteract hyperphagia and obesity. These studies will be the first to functionally link APA regulation to feeding behavior and will serve as the basis of further genome to behavioral phenome studies in my independent career. My Sponsor, Dr. Gary Wayman, and Co-Sponsors, Drs. Suzanne Appleyard and Emily Qualls-Creekmore, are established neuroscientists at Washington State University with expertise in molecular neuroscience (Wayman) and ingestive behavior (Appleyard and Qualls-Creekmore). My proposed Research and Training plan will strengthen my theoretical and technical understanding of neurogenetics. In the pre-doctoral F99 phase, I will learn shRNA and CRISPR/SaCas9 vector design and validation strategies, cell culture techniques, stereotaxic surgeries, and advanced metabolic analyses. In the postdoctoral K00 phase, I will build upon these skills and learn to use genetic mouse models, multi-omics, advanced bioinformatics, and AI computational models to map genome to phenome regulation. Overall, the proposed training will optimally position me to start an independent research career at a leading neuroscience research institute and advance our understanding of RNA regulation as a functional link between the genome and the behavioral phenome.