Defining the Regulation and Function of Hepatic mTORC1 Signaling

Abstract

The protein kinase complex mechanistic target of rapamycin complex 1 (mTORC1) is at the center of an anabolic signaling pathway that is dysregulated in metabolic disease, as well as in human cancers and tumor syndromes. Mediators and targets of mTORC1 signaling have been characterized in cell culture models, but the regulation and function of mTORC1 in response to specific physiological cues and in specific tissue contexts has not been thoroughly examined. Utilizing knowledge gained from in vitro studies, we have developed a novel genetic mouse model to investigate the specific role of insulin-PI3K-AKT signaling in the physiological regulation and function of hepatic mTORC1. By analyzing mTORC1 signaling in liver tissue under a variety of experimental paradigms, including insulin and glucose treatment and fasting and refeeding, we demonstrated that the dominant mechanism of hepatic mTORC1 induction by insulin is AKT-mediated phosphorylation of the TSC complex, a critical negative regulator of mTORC1. However, we found that hepatic mTORC1 can be induced by feeding in a manner independent of this insulin-mediated mechanism. Specifically, we found that dietary protein was critical for hepatic mTORC1 activation by feeding and dominant over insulin signaling, and we present evidence that amino acids are sufficient to induce mTORC1 in primary hepatocytes. The reliance on insulin signaling for the activation of hepatic mTORC1 was increased in response to feeding a high carbohydrate, low protein diet. These data suggest a model whereby dietary composition impacts the contribution of insulin in the activation of mTORC1 in the liver. Strikingly, insulin-induced hepatic mTORC1 played a minimal role in impaired glucose homeostasis in diet-induced obese mice. To provide insight into the physiological functions of hepatic mTORC1, we defined novel targets of mTOR kinase activity through liver quantitative phosphoproteomics. We identified growth factor receptor bound protein 7 (Grb7) as a novel target of insulin-induced mTOR kinase activity. Our data suggests Grb7 inhibits insulin signaling in primary mouse hepatocytes and human hepatoma cells and may mediate mTORC1-driven feedback inhibition of insulin signaling, with a potential role in hepatic insulin resistance. Collectively, this study defines a hierarchy of signals regulating physiological mTORC1 signaling for the first time and identifies a set of physiological mTOR substrates that can be exploited to modulate mTORC1 function in physiological or pathological settings.