One of the most fundamental issues in cell biology is how cells integrate growth-stimulating and inhibitory signals to ultimately regulate a diversity of key cellular functions, including gene expression, autophagy, organelle biogenesis, and cell growth. mTOR is a serine/threonine kinase that regulates proliferation, cell cycle, and autophagy in response to energy levels, growth factors, and nutrients. mTOR responds to numerous stresses and its dysregulation leads to cancer, metabolic disease, and diabetes. In cells, mTOR exists as two structurally and functionally distinct complexes termed mTOR complex 1 (mTORC1) and mTOR complex 2 (mTORC2). mTORC1 couples energy and nutrient abundance to cell growth and proliferation by balancing anabolic (protein synthesis and nutrient storage) and catabolic processes (autophagy and the utilization of energy stores). Active mTORC1 localizes to late endosomes/lysosomes and this distribution is thought to be critical for the ability of mTORC1 to sense and respond to variations in the levels of amino acids. The transcription factor EB (TFEB) is a member of the basic helix-loop-helix leucine-zipper family of transcription factors that controls lysosomal biogenesis and autophagy by positively regulating genes belonging to the Coordinated Lysosomal Expression and Regulation (CLEAR) network. Importantly, we have found that mTORC1 controls the activity and cellular localization of TFEB. Under nutrient-rich conditions, mTORC1 phophorylates TFEB in S211, thus promoting binding of TFEB to the cytosolic chaperone 14-3-3 and retention of TFEB in the cytosol. Upon amino acids deprivation, dissociation of the TFEB/14-3-3 complex results in delivery of TFEB to the nucleus and up-regulation of genes that leads to induction of autophagy, biogenesis of lysosomes, and increased lysosomal degradation. We also found that TFEB is recruited to lysosomes through direct interaction with active Rag GTPases. This Rag-mediated redistribution of TFEB to the lysosomal surface facilitates the phosphorylation of TFEB by mTORC1 and constitutes an efficient way to link nutrient availability to TFEB inactivation. Inhibition of the interaction between TFEB and Rags results in accumulation of TFEB in the nucleus and constitutive activation of autophagy under nutrient rich conditions, thus indicating that recruitment of TFEB to lysosomes is critical for the proper control of this transcription factor. We have also identified the transcription factor E3 (TFE3) as novel regulator of lysosomal formation and function. Similar to TFEB, the recruitment of TFE3 to lysosomes is mediated by active Rag GTPases and this step is critical for mTORC1-mediated phosphorylation of TFE3 and retention in the cytosol. Over-expression of TFE3 results in increased autophagy and enhanced lysosomal biogenesis, as evidenced by an increase in the number of lysosomes and lysosomal activity. In contrast, depletion of endogenous TFE3 entirely abolishes the cellular response to starvation, thus confirming the crucial role of TFE3 in nutrient sensing and energy metabolism. More recently we have addresses the participation of TFEB and TFE3 in cellular adaptation to different types of stress. We found that TFEB and TFE3 play an important role in the cellular response to ER stress. Treatment with ER stressors causes translocation of TFEB and TFE3 to the nucleus in a process that is dependent on PERK and calcineurin but not on mTORC1. Activated TFEB and TFE3 enhance cellular response to stress by inducing direct transcriptional upregulation of ATF4 and other unfolded protein response (UPR) genes. Under conditions of prolonged ER stress, TFEB and TFE3 contribute to cell death, thus revealing an unexpected role for these proteins in controlling cell fate. Finally, we found that TFEB and TFE3 cooperate in the transcriptional regulation of the innate immune response. TFEB and TFE3 are rapidly recruited to the nucleus of activated macrophages where they promote lysosomal biogenesis, autophagy induction, as well as expression of a number of cytokines, chemokines, and other immune-related genes involved in the regulation and activation of the innate immune response. In summary, our work evidences a broader role of TFEB and TFE3 in the cellular response to stress than previously anticipated and reveals an integrated cooperation between different cellular stress pathways.
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