Investigating folliculin as a regulator of VLDL-TG secretion in non-alcoholic fatty liver disease and steatohepatitis - Abstract Non-alcoholic fatty liver disease (NAFLD) is a rapidly emerging public health risk. Approximately 35% of Americans have NAFLD, and if left untreated, NAFLD can progress to non-alcoholic steatohepatitis (NASH), cirrhosis, and hepatocellular carcinoma. Although obesity and Type II diabetes have emerged as risk factors, disease pathogenesis is poorly understood. Accordingly, there are no FDA-approved pharmacotherapies currently available for patients with NAFLD/NASH. An ideal therapeutic would target multiple hepatic lipid homeostatic processes, including de novo lipogenesis (DNL), fatty acid oxidation (FAO), and very-low-density lipoprotein (VLDL)-triglyceride (TG) secretion, as attempts to modulate a single lipid homeostatic process have often led to undesirable compensation from the remaining pathways. Our lab has identified the protein folliculin (FLCN) as a potential therapeutic target. Mice with hepatic deletion of FLCN are robustly protected against steatosis when challenged with NAFLD-inducing diets via an FLCN-mTORC1-TFE3 signaling axis. TFE3 suppresses DNL and induces genes involved in FAO to decrease hepatic TG content in FLCN-null livers. Given that VLDL-TG secretion is heavily regulated by TG availability, we expected to see a compensatory decrease in VLDL-TG secretion rate in FLCN-null mice. Strikingly, however, we saw marked upregulation of VLDL-TG secretion in FLCN deficient mice, which makes FLCN even more appealing as a therapeutic target. I hypothesize that upregulated VLDL-TG secretion is critical to the robust protection against steatosis afforded by liver deletion of FLCN, and that activated TFE3 is necessary and sufficient for VLDL-TG secretion. To test this, I will generate FLCN knockout (KO), TFE3 KO, and FLCN/TFE3 double knockout (DKO) mice and assess VLDL-TG secretion rates. I will overexpress constitutively active TFE3 in wild-type mice to test if TFE3 is sufficient for VLDL-TG secretion. My preliminary data further lead me to hypothesize that the upregulated VLDL-TG secretion in FLCN- null livers is mediated by TFE3-dependent transcriptional activation of CTP:phosphocholine cytidylyltransferase alpha (CCTa). CCTa is the rate-limiting enzyme in the synthesis of phosphatidylcholine, a critical necessary component for VLDL-TG secretion. My preliminary data show increased CCTa mRNA and protein levels in FLCN knockout mice. I will generate FLCN KO, CCTa KO, FLCN/CCTa DKO mice. I will assess their susceptibility to NAFLD/NASH and perform VLDL-TG secretion assays. Completion of this work would provide insight into the regulation of hepatic lipid homeostatic processes affected in NAFLD/NASH, as well as demonstrate FLCN to be a promising therapeutic target for treatment.
