Obesity affects a significant number of Americans and greatly increases the risk of developing cardiovascular disease (CVD). Understanding how obesity is regulated opens up new avenues of pharmacological interventions to treat obesity, concurrently reducing the risk of CVD and other co-morbidities. Nocturnin (NOC) is broadly expressed enzyme with exoribonuclease activity in in vitro assays and a repressive function against reporter RNAs in vivo. These observations are consistent with a function in RNA decay. Mice lacking NOC are resistant to diet-induced obesity and other studies have observed a role for NOC in regulating intestinal trafficking of dietary fats and promoting adipogenesis. These observations suggest that NOC may have a novel role in the regulation of lipid metabolism by targeting mRNAs for degradation. Though the broader physiological effects of NOC gene deletion have been described, discovering the molecular function of NOC remains an important goal.We have determined the first crystal structure of human NOC to aid in characterizing its RNA substrate specificity and catalytic mechanism. We developed biochemical and cell-based assays that will be used to study NOC function and analyze the effects of active site mutants. To further determine interactions in the active site that are important for binding and catalysis, NOC will be co-crystallized with RNA, further guiding functional assays in characterizing the enzyme in vitro and in vivo.To better describe the role of NOC in regulation of fat metabolism and adipogenesis, the identity of NOC targets will be identified. Toward this goal, I will initially focus on a list of putative NOC targets that were recently reported. Intriguingly, our gene ontology analysis indicates that this set of genes is enriched in functions linked to fat metabolism. The regulation of these putative NOC targets will be measured using loss of function approaches followed by quantitative measurement of gene expression. In summary, the experiments outlined in this proposal will yield a thorough understanding of the RNA substrate specificity of NOC and its functions in regulating fat metabolism. I expect to gain a better understanding of the NOC function and substrate specificity. Importantly, this research will provide a foundation for characterizing NOC as a potential drug target for obesity that will directly benefit treatment of CVD and other co-morbidities.
|Program type||Predoctoral Fellowship|
|Effective start/end date||01/01/2016 → 12/31/2017|