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Inhibition of nonalcoholic fatty liver disease in mice by selective inhibition of mTORC1

Nonalcoholic fatty liver disease (NAFLD) and nonalcoholic steatohepatitis (NASH) remain without effective therapies. The mechanistic target of rapamycin complex 1 (mTORC1) pathway is a potential therapeutic target, but conflicting interpretations have been proposed for how mTORC1 controls lipid homeostasis. We show that selective inhibition of mTORC1 signaling in mice, through deletion of the RagC/D guanosine triphosphatase–activating protein folliculin (FLCN), promotes activation of transcription factor E3 (TFE3) in the liver without affecting other mTORC1 targets and protects against NAFLD and NASH. Disease protection is mediated by TFE3, which both induces lipid consumption and suppresses anabolic lipogenesis. TFE3 inhibits lipogenesis by suppressing proteolytic processing and activation of sterol regulatory element–binding protein-1c (SREBP-1c) and by interacting with SREBP-1c on chromatin. Our data reconcile previously conflicting studies and identify selective inhibition of mTORC1 as a potential approach to treat NASH and NAFLD. Description mTORC1 as fatty liver disease target Inappropriate accumulation of fat in the liver causes serious life-threatening disease in humans. In mice, Gosis et al. explored the potential beneficial effects of controlling lipid metabolism by preventing signaling by the mTORC1 (mechanistic target of rapamycin complex 1) protein kinase complex (see the Perspective by Ginsberg and Mani). The authors inhibited some, but not all, signaling by mTORC1 by depleting the folliculin protein in the liver. Such mice had increased lipid consumption and decreased lipogenesis and were protected when fed a diet that normally induces nonalcoholic fatty liver disease. These effects appear to result in part from activation of the TFE3 transcription factor. Similar strategies might thus be useful in treating liver disease. —LBR Inhibiting a particular signal from the mTORC1 protein kinase complex protects mice from fatty liver disease. INTRODUCTION As many as 100 million people in the US have nonalcoholic fatty liver disease (NAFLD), characterized by increased liver lipid accumulation, which often leads to hepatocyte injury and fibrosis, characteristics of nonalcoholic steatohepatitis (NASH). NASH in turn can progress to cirrhosis and hepatocellular carcinoma. There are currently no US Food and Drug Administration–approved therapies for NAFLD or NASH. NAFLD occurs when there is disequilibrium between the processes of hepatic lipid synthesis and consumption. The nutrient sensor mechanistic target of rapamycin complex 1 (mTORC1) regulates several of these pathways. mTORC1 is thus an attractive target to modulate lipid homeostasis in the liver. However, mTORC1 also regulates numerous other cellular pathways, and blunting of mTORC1 modulation can lead to unexpected feedback loops and unwanted effects. RATIONALE We hypothesized that selective modulation of hepatic mTORC1 signaling could benefit liver lipid metabolism and prevent NAFLD. In non-liver cell types, the protein folliculin (FLCN) has been shown to confer substrate specificity to mTORC1. Deletion of FLCN inhibits mTORC1-mediated phosphorylation of the transcription factor E3/B (TFE3/B) family of transcription factors, without affecting mTORC1-driven phosphorylation of its canonical substrates ribosomal protein S6 kinase beta-1 (S6K1) and eukaryotic translation initiation factor 4E–binding protein 1 (4E-BP1). Unphosphorylated TFE3 translocates to the nucleus and activates genes that promote lysosomal biogenesis, mitochondrial biogenesis, and oxidative metabolism. We reasoned that suppression of FLCN in the liver might promote fatty acid oxidation and lipid clearance without untoward effects of generalized mTORC1 inhibition. RESULTS Hepatocyte-specific genetic deletion of Flcn in adult mice selectively inhibited mTORC1-mediated cytoplasmic sequestration of TFE3, with little effect on other mTORC1 targets, including S6K, 4E-BP1, and Lipin1. Hepatocyte loss of Flcn protected mice from both NAFLD and NASH and partially reversed these processes when already established. The protection against NAFLD and NASH required TFE3, which activated lipid clearance. Unleashed TFE3 additionally suppressed de novo lipogenesis. The latter was mediated in part by TFE3-mediated induction of insulin-induced gene 2 (Insig2) to inhibit proteolytic activation of sterol regulatory element–binding protein-1c (SREBP-1c), a critical lipogenic transcription factor. CONCLUSION Our data establish FLCN as a critical regulator of lipid homeostasis in the liver. Flcn deletion affords selective inhibition of mTORC1, leading to nuclear translocation and activation of the transcription factor TFE3, which coordinates hepatic lipid metabolic pathways to protect against NAFLD and NASH in mice. Thus, our data reveal FLCN as a promising target for the treatment of NAFLD and NASH. The data also illuminate previously published and seemingly conflicting data, which likely reflected different effects on each arm of mTORC1 signaling. There have been numerous attempts by many to develop disease-specific treatments for NAFLD and NASH, thus far without success. A recurrent problem has been the many compensatory responses by the liver to targeting any one pathway; for example, inhibitors of acetyl–coenzyme A carboxylase led to compensatory activation of SREBP-1c and consequent hyperlipidemia. Targeting FLCN is thus particularly attractive, in that loss of FLCN simultaneously and favorably affects multiple aspects of hepatic lipid homeostasis, including promoting fatty acid oxidation and lysosomal biogenesis and inhibiting de novo lipogenesis. Deletion of Flcn in the liver protects mice from NAFLD and NASH through selective suppression of mTORC1. Diets high in fat, carbohydrates, and cholesterol lead to NAFLD and NASH. When Flcn is simultaneously deleted, mTORC1 is selectively inhibited, preserving phosphorylation of canonical substrates S6K and 4E-BP1 while blocking phosphorylation of the transcription factor TFE3. Unphosphorylated TFE3 is released to the nucleus, where it activates lipid catabolism genes while suppressing de novo lipogenesis genes. [Image created using Biorender]