PROJECT SUMMARY/ABSTRACT From Greek philosopher Plato to recent animal and human studies, fasting has been recommended as a strategy to slow down aging and reduce the risk of many aging related diseases, such as insulin resistance, and type 2 diabetes. Understanding how fasting signals elicit cellular programs that establish “anti-aging” benefits will provide new therapeutic targets and strategies to prevent aging associated pathologies and expand healthspan. My findings show that during fasting, hepatocytes selectively remodel the translatome to activate a specific translation network of many longevity-related genes, which is overlooked by conventional transcriptomics analysis. This new mechanism involves a phosphorylation event of the major cap-binding protein eIF4E, which was perceived as a general translation factor. Strikingly, upon fasting, phosphorylation of eIF4E is activated and exquisitely induces target-specific translation, including mRNAs involved in lipid and glucose metabolism, despite that global translation is downregulated. Genetically inhibiting eIF4E phosphorylation leads to a reduction of ketogenesis and insulin sensitivity upon fasting or a fasting mimic diet. Furthermore, my preliminary data revealed that fatty acids (FAs), which are elevated in the blood upon fasting, are novel signaling molecules that induce the phosphorylation of eIF4E. These findings reveal a new paradigm for control of gene expression at the level of the proteome downstream of FA signaling that establishes metabolomic reprogramming and health benefits during fasting. Intriguingly, aging blunts this activation of eIF4E phosphorylation, and lack of eIF4E phosphorylation in mice exhibits early markers of insulin resistance at middle age. Thereby, I hypothesize that FAs activate a signaling pathway, that modulates the eIF4E-dependent translation network of mRNAs involved in ketogenesis and insulin pathway upon fasting. I further posit that dysregulation of this pathway during aging may be responsible for aging related insulin resistance. The specific aims of this project is to first identify the molecular mechanism underlying FA induced translation control, which includes identifying the functional cis regulatory motifs in mRNAs that render their specificity during fasting (Aim 1.1) by mutagenesis and functional assays, and revealing the upstream signals that are induced by FA (Aim 1.2) through unbiased labeling or a pull-down system coupled with mass spectrometry (MS). Secondly, I will determine the functional role of phospho-eIF4E dependent translation in aging associated insulin resistance (Aim 2.1). I will set up an aging mouse model employing eIF4E phosphorylation mutant mice coupled to metabolic assays and translatome analysis to detect how phospho-eIF4E dependent translation control affects insulin sensitivity upon aging. I will also set up an aging cell model to determine the mechanisms underlying the aging induced loss of activation of eIF4E phos...