Regulation of Liver Metabolism by Autophagy

Intracellular components must be recycled for cells to maintain energy and ensure quality control of proteins and organelles. Autophagy is a highly conserved recycling process that involves degradation of cellular constituents in lysosomes. Although autophagy regulates a number of cell functions, it was first found to maintain energy balance in liver cells. As our understanding of autophagy has increased, we have found its connections to energy regulation in liver cells to be tight and complex. We review 3 mechanisms by which hepatic autophagy monitors and regulates cellular metabolism. Autophagy provides essential components (amino acids, lipids, and carbohydrates) required to meet the cell’s energy needs, and it also regulates energy supply by controlling the number, quality, and dynamics of the mitochondria. Finally, autophagy also modulates levels of enzymes in metabolic pathways. In light of the multiple ways in which autophagy participates to control liver metabolism, it is no surprise that dysregulation of autophagy has been associated with metabolic diseases such as obesity, diabetes, or metabolic syndrome, as well as liver-specific disorders such as fatty liver, nonalcoholic steatohepatitis, and hepatocellular carcinoma. We discuss some of these connections and how hepatic autophagy might serve as a therapeutic target in common metabolic disorders. Intracellular components must be recycled for cells to maintain energy and ensure quality control of proteins and organelles. Autophagy is a highly conserved recycling process that involves degradation of cellular constituents in lysosomes. Although autophagy regulates a number of cell functions, it was first found to maintain energy balance in liver cells. As our understanding of autophagy has increased, we have found its connections to energy regulation in liver cells to be tight and complex. We review 3 mechanisms by which hepatic autophagy monitors and regulates cellular metabolism. Autophagy provides essential components (amino acids, lipids, and carbohydrates) required to meet the cell’s energy needs, and it also regulates energy supply by controlling the number, quality, and dynamics of the mitochondria. Finally, autophagy also modulates levels of enzymes in metabolic pathways. In light of the multiple ways in which autophagy participates to control liver metabolism, it is no surprise that dysregulation of autophagy has been associated with metabolic diseases such as obesity, diabetes, or metabolic syndrome, as well as liver-specific disorders such as fatty liver, nonalcoholic steatohepatitis, and hepatocellular carcinoma. We discuss some of these connections and how hepatic autophagy might serve as a therapeutic target in common metabolic disorders. Recycling is the basis of cellular survival by contributing to reduce cellular waste, to preserve cellular energy, and to adapt to environmental challenges by regulating abundance of intracellular components.1Singh R. Cuervo A.M. Autophagy in the cellular energetic balance.Cell Metab. 2011; 13: 495-504Abstract Full Text Full Text PDF PubMed Scopus (577) Google Scholar, 2Mizushima N. The pleiotropic role of autophagy: from protein metabolism to bactericide.Cell Death Differ. 2005; 12: 1535-1541Crossref PubMed Scopus (404) Google Scholar Autophagy, the process that mediates degradation of intracellular constituents in lysosomes, is the ultimate example of efficient cellular recycling because it seamlessly couples cellular quality control and cellular energetics.2Mizushima N. The pleiotropic role of autophagy: from protein metabolism to bactericide.Cell Death Differ. 2005; 12: 1535-1541Crossref PubMed Scopus (404) Google Scholar Autophagy contributes to maintain a positive energetic balance through degradation and recycling of proteins,3Ezaki J. Matsumoto N. Takeda-Ezaki M. et al.Liver autophagy contributes to the maintenance of blood glucose and amino acid levels.Autophagy. 2011; 7: 727-736Crossref PubMed Scopus (207) Google Scholar, 4Kuma A. Hatano M. Matsui M. et al.The role of autophagy during the early neonatal starvation period.Nature. 2004; 432: 1032-1036Crossref PubMed Scopus (2386) Google Scholar glycogen,5Raben N. Hill V. Shea L. et al.Suppression of autophagy in skeletal muscle uncovers the accumulation of ubiquitinated proteins and their potential role in muscle damage in Pompe disease.Hum Mol Genet. 2008; 17: 3897-3908Crossref PubMed Scopus (255) Google Scholar or lipids.6Singh R. Kaushik S. Wang Y. et al.Autophagy regulates lipid metabolism.Nature. 2009; 458: 1131-1135Crossref PubMed Scopus (2635) Google Scholar Regulated turnover through autophagy also ensures renewal and proper functioning of proteome and intracellular organelles.7Mizushima N. Levine B. Cuervo A.M. et al.Autophagy fights disease through cellular self-digestion.Nature. 2008; 451: 1069-1075Crossref PubMed Scopus (5144) Google Scholar Cellular metabolism has been tightly associated with autophagy since the early days when de Duve et al8Deter R.L. Baudhuin P. De Duve C. Participation of lysosomes in cellular autophagy induced in rat liver by glucagon.J Cell Biol. 1967; 35: C11-C16Crossref PubMed Google Scholar showed that starvation and glucagon, a glycogenolytic agent, induced hepatic autophagy. Since then, many new functions have been linked to autophagic activity such as control of the cell cycle, immune response, development, differentiation, or cell death.9Zhang H. Baehrecke E.H. Eaten alive: novel insights into autophagy from multicellular model systems.Trends Cell Biol. 2015; 25: 376-387Abstract Full Text Full Text PDF PubMed Scopus (81) Google Scholar, 10Cecconi F. Levine B. The role of autophagy in mammalian development: cell makeover rather than cell death.Dev Cell. 2008; 15: 344-357Abstract Full Text Full Text PDF PubMed Scopus (443) Google Scholar However, the role of autophagy in cellular metabolism has persisted as one of its main functions because of the identification of new additional ways in which autophagy contributes to cellular energetics. We recently learned that, added to the previously shown replenishment of the free pool of amino acids through protein breakdown, autophagy also contributes to mobilization and hydrolysis of lipid stores and glycogen.5Raben N. Hill V. Shea L. et al.Suppression of autophagy in skeletal muscle uncovers the accumulation of ubiquitinated proteins and their potential role in muscle damage in Pompe disease.Hum Mol Genet. 2008; 17: 3897-3908Crossref PubMed Scopus (255) Google Scholar, 6Singh R. Kaushik S. Wang Y. et al.Autophagy regulates lipid metabolism.Nature. 2009; 458: 1131-1135Crossref PubMed Scopus (2635) Google Scholar Furthermore, a selective form of autophagy known as mitophagy controls the number and functionality of the mitochondrial network responsible for adenosine triphosphate (ATP) generation from different energy stores.11Lemasters J.J. Variants of mitochondrial autophagy: types 1 and 2 mitophagy and micromitophagy (type 3).Redox Biol. 2014; 2: 749-754Crossref PubMed Scopus (204) Google Scholar, 12Zhu J. Wang K.Z. Chu C.T. After the banquet: mitochondrial biogenesis, mitophagy, and cell survival.Autophagy. 2013; 9: 1663-1676Crossref PubMed Scopus (220) Google Scholar Finally, enzymes and other proteins that participate in metabolic pathways such as glycolysis, lipophagy, or lipolysis can be timely and selectively degraded by autophagy, contributing in this way to modulate the energetic flux through these metabolic pathways.13Schneider J.L. Suh Y. Cuervo A.M. Deficient chaperone-mediated autophagy in liver leads to metabolic dysregulation.Cell Metab. 2014; 20: 417-432Abstract Full Text Full Text PDF PubMed Scopus (198) Google Scholar, 14Shpilka T. Welter E. Borovsky N. et al.Fatty acid synthase is preferentially degraded by autophagy upon nitrogen starvation in yeast.Proc Natl Acad Sci U S A. 2015; 112: 1434-1439Crossref PubMed Scopus (46) Google Scholar In this review, we provide a brief description of the autophagic pathways in the mammalian liver and then summarize recent findings on the role of these autophagy variants in the metabolic process and metabolic disorders in the liver, the organ most tightly related with whole-body energetics. Autophagy stands out among the different post-translational regulatory mechanisms in liver because of its broad range of physiological functions in this organ. The versatility of the 2 key steps of the autophagy process, breakdown and recycling, along with the co-existence of different autophagic pathways in the same cell, allow for this functional diversity. Three types of autophagy co-exist in hepatocytes, macroautophagy, microautophagy, and chaperone-mediated autophagy (CMA) (Figure 1), and connections with the hepatic energetic balance have been established for all of them. Here, we briefly summarize the main characteristics that differentiate each of these autophagic processes, but readers interested in a more comprehensive description are encouraged to consult recent reviews on this topic.9Zhang H. Baehrecke E.H. Eaten alive: novel insights into autophagy from multicellular model systems.Trends Cell Biol. 2015; 25: 376-387Abstract Full Text Full Text PDF PubMed Scopus (81) Google Scholar, 15Feng Y. He D. Yao Z. et al.The machinery of macroautophagy.Cell Res. 2014; 24: 24-41Crossref PubMed Scopus (1318) Google Scholar, 16Shen H.M. Mizushima N. At the end of the autophagic road: an emerging understanding of lysosomal functions in autophagy.Trends Biochem Sci. 2014; 39: 61-71Abstract Full Text Full Text PDF PubMed Scopus (264) Google Scholar, 17Kaushik S. Cuervo A.M. Chaperone-mediated autophagy: a unique way to enter the lysosome world.Trends Cell Biol. 2012; 22: 407-417Abstract Full Text Full Text PDF PubMed Scopus (594) Google Scholar Macroautophagy, the better-characterized autophagic pathway, starts with the formation of the limiting membrane of the autophagosome, a double-membrane vesicle that delivers the cytosolic material targeted for degradation (cargo) to lysosomes. This limiting membrane, or nascent phagophore, forms through the assembly of proteins and lipids from different cellular organelles such as the endoplasmic reticulum (ER), Golgi, mitochondria, endocytic system, or plasma membrane (Figure 1). Cargo sequestration by autophagy in response to starvation occurs, for the most part, in bulk. However, a better characterization of autophagy activated in response to other stimuli has led to the discovery of selective forms of macroautophagy based on the cargo recruited for degradation.18Stolz A. Ernst A. Dikic I. Cargo recognition and trafficking in selective autophagy.Nat Cell Biol. 2014; 16: 495-501Crossref PubMed Scopus (817) Google Scholar In these instances, formation of the pre-autophagosome structure occurs in close proximity to the cargo thanks to autophagy receptors, such as P62SQSTM1, NBR1, NDP52, and optineurin.18Stolz A. Ernst A. Dikic I. Cargo recognition and trafficking in selective autophagy.Nat Cell Biol. 2014; 16: 495-501Crossref PubMed Scopus (817) Google Scholar These receptors simultaneously bind cargo and key components of the autophagy machinery known as autophagy-related proteins (ATGs).18Stolz A. Ernst A. Dikic I. Cargo recognition and trafficking in selective autophagy.Nat Cell Biol. 2014; 16: 495-501Crossref PubMed Scopus (817) Google Scholar The elongation of this membrane around the cargo occurs through the coordinated action of a cascade of ATGs that conjugate among themselves and with lipids.19Mizushima N. Yoshimori T. Ohsumi Y. The role of Atg proteins in autophagosome formation.Ann Rev Cell Dev Biol. 2011; 27: 107-132Crossref PubMed Scopus (2153) Google Scholar Upon sealing of the autophagosome membrane, this cargo-containing vesicle moves along microtubules to fuse with the lysosome and deliver the sequestered cargo in this hydrolytic organelle16Shen H.M. Mizushima N. At the end of the autophagic road: an emerging understanding of lysosomal functions in autophagy.Trends Biochem Sci. 2014; 39: 61-71Abstract Full Text Full Text PDF PubMed Scopus (264) Google Scholar (Figure 1). After hydrolysis, the resultant molecules, amino acids, lipids, and carbohydrate moieties reach the cytosol through transporters and permeases for recycling.16Shen H.M. Mizushima N. At the end of the autophagic road: an emerging understanding of lysosomal functions in autophagy.Trends Biochem Sci. 2014; 39: 61-71Abstract Full Text Full Text PDF PubMed Scopus (264) Google Scholar How cells sense the need for a boost in autophagy activity has yet to be well defined and likely may occur through multiple mechanisms depending on the type of stimulus. Recent studies have shed some light on one of these mechanisms by showing that nutrient deprivation can be sensed by signaling cascades present on the primary cilia, such as the Sonic Hedgehog pathway.20Pampliega O. Orhon I. Patel B. et al.Functional interaction between autophagy and ciliogenesis.Nature. 2013; 502: 194-200Crossref PubMed Scopus (302) Google Scholar Signaling by this pathway activates autophagy by recruiting ATGs to the base of the cilia using ciliary trafficking proteins. The unique position of this single-ciliary structure that protrudes from the cellular surface and the highly specialized nature of the membrane that surrounds the cilia, supports the idea that primary cilia also may contribute to modulate autophagy by transducing other signals such as those initiated by growth factors, hormone ligands, or even physical stimuli that lead to ciliary bending. This association of ATGs with the plasma membrane also may assist the activation of autophagy in response to changes in the neighboring cells. Thus, for example, under rich nutritional conditions, connexins, the main structural component of intercellular gap junctions, serve as endogenous repressors of autophagy by directly binding to ATG complexes involved in autophagosome formation. During nutrient scarcity, these connexins become platforms for recruitment of additional ATGs (ie, ATG14 and ATG9), driving the internalization of connexin ATGs from the plasma membrane to the recycling endocytic compartment for autophagosome formation.21Bejarano E. Yuste A. Patel B. et al.Connexins modulate autophagosome biogenesis.Nat Cell Biol. 2014; 16: 401-414Crossref PubMed Scopus (94) Google Scholar Once the autophagy-activating signal is transduced, ATGs control each of the steps of the autophagy pathway, from autophagosome formation to cargo degradation and recycling. Consequently, in addition to their reorganization from inactive to active complexes, changes in ATG expression levels in response to different transcriptional programs modulate the magnitude and duration of autophagy activation. One of the master transcriptional regulators of the autophagic program is the transcription factor EB, or TFEB,22Settembre C. Di Malta C. Polito V.A. et al.TFEB links autophagy to lysosomal biogenesis.Science. 2011; 332: 1429-1433Crossref PubMed Scopus (2009) Google Scholar that enhances autophagy flux at multiple levels. First, TFEB transcriptionally manages expression of ATG genes during sustained autophagy to prevent ATG protein depletion. Second, TFEB increases lysosomal biogenesis to avoid overwhelming this degradative organelle by the arriving autophagosomes. Finally, TFEB also controls genes required in other autophagy steps such as molecular motors for trafficking, soluble N-ethylmaleimide-sensitive factor attachment protein receptors (SNARES) for membrane fusion, and specific hydrolases to accommodate the lysosomal enzymatic load to the type of arriving cargo.23Settembre C. De Cegli R. Mansueto G. et al.TFEB controls cellular lipid metabolism through a starvation-induced autoregulatory loop.Nat Cell Biol. 2013; 15: 647-658Crossref PubMed Scopus (630) Google Scholar, 24O'Rourke E.J. Ruvkun G. MXL-3 and HLH-30 transcriptionally link lipolysis and autophagy to nutrient availability.Nat Cell Biol. 2013; 15: 668-676Crossref PubMed Scopus (208) Google Scholar However, TFEB and the recently described more potent members of this family such as TFE325Martina J.A. Diab H.I. Lishu L. et al.The nutrient-responsive transcription factor TFE3 promotes autophagy, lysosomal biogenesis, and clearance of cellular debris.Sci Signal. 2014; 7: ra9Crossref PubMed Scopus (387) Google Scholar are not the only transcriptional regulators of autophagy. The transcriptional autophagy network also is controlled by the sensing nuclear receptor farnesoid X receptor in the fed state, guaranteeing physiological proteostasis through basal recycling of proteins.26Seok S. Fu T. Choi S.E. et al.Transcriptional regulation of autophagy by an FXR-CREB axis.Nature. 2014; 516: 108-111Crossref PubMed Scopus (297) Google Scholar During fasting, transcriptional regulation of the autophagy program is monitored by cyclic adenosine monophosphate response element-binding protein and peroxisome proliferator-activated receptor-alpha (PPARα).26Seok S. Fu T. Choi S.E. et al.Transcriptional regulation of autophagy by an FXR-CREB axis.Nature. 2014; 516: 108-111Crossref PubMed Scopus (297) Google Scholar, 27Lee J.M. Wagner M. Xiao R. et al.Nutrient-sensing nuclear receptors coordinate autophagy.Nature. 2014; 516: 112-115Crossref PubMed Scopus (352) Google Scholar CMA degrades a specific subset of proteins that cross the lysosomal membrane through the CMA receptor, the lysosome-associated membrane protein type 2A (LAMP-2A).17Kaushik S. Cuervo A.M. Chaperone-mediated autophagy: a unique way to enter the lysosome world.Trends Cell Biol. 2012; 22: 407-417Abstract Full Text Full Text PDF PubMed Scopus (594) Google Scholar All CMA substrate proteins contain in their amino acid sequence a pentapeptide motif (KFERQ)28Dice J.F. Peptide sequences that target cytosolic proteins for lysosomal proteolysis.Trends Biochem Sci. 1990; 15: 305-309Abstract Full Text PDF PubMed Scopus (529) Google Scholar that is selectively recognized by the cytosolic heat shock cognate protein of 70 kilodaltons (HSC70) (Figure 1). The substrate/chaperone complex docks at the lysosomal membrane through binding to monomeric LAMP-2A proteins that then organize into a multimeric complex required for translocation.29Bandyopadhyay U. Kaushik S. Varticovski L. et al.The chaperone-mediated autophagy receptor organizes in dynamic protein complexes at the lysosomal membrane.Mol Cell Biol. 2008; 28: 5747-5763Crossref PubMed Scopus (364) Google Scholar After unfolding and internalization, the substrate protein is degraded rapidly in the lysosomal lumen17Kaushik S. Cuervo A.M. Chaperone-mediated autophagy: a unique way to enter the lysosome world.Trends Cell Biol. 2012; 22: 407-417Abstract Full Text Full Text PDF PubMed Scopus (594) Google Scholar (Figure 1). LAMP-2A levels, which limit CMA activity, are controlled both through transcriptional activation and, more frequently, through direct changes in the stability of LAMP-2A at the lysosomal membrane.30Cuervo A.M. Dice J.F. Regulation of lamp2a levels in the lysosomal membrane.Traffic. 2000; 1: 570-583Crossref PubMed Scopus (225) Google Scholar CMA originally was described in liver as part of the hepatocyte response to nutritional changes.31Cuervo A.M. Knecht E. Terlecky S.R. et al.Activation of a selective pathway of lysosomal proteolysis in rat liver by prolonged starvation.Am J Physiol. 1995; 269: C1200-C1208Crossref PubMed Google Scholar Later studies have shown that CMA also functions as a defense mechanism against cellular insults in an effort to remove damaged proteins and thereby ensure appropriate hepatic proteostasis (reviewed by Schneider and Cuervo32Schneider J.L. Cuervo A.M. Liver autophagy: much more than just taking out the trash.Nat Rev Gastroenterol Hepatol. 2014; 11: 187-200Crossref PubMed Scopus (138) Google Scholar). The signaling mechanisms that regulate CMA activity are, for the most part, still unknown. Signaling through the nuclear retinoic acid receptor α negatively regulates CMA activity,33Anguiano J. Garner T.P. Mahalingam M. et al.Chemical modulation of chaperone-mediated autophagy by retinoic acid derivatives.Nat Chem Biol. 2013; 9: 374-382Crossref PubMed Scopus (145) Google Scholar whereas recent studies in T cells have identified the nuclear factor of activated T-cells (NFAT)-calcineurin axis behind CMA induction during T-cell activation.34Valdor R. Mocholi E. Botbol Y. et al.Chaperone-mediated autophagy regulates T cell responses through targeted degradation of negative regulators of T cell activation.Nat Immunol. 2014; 15: 1046-1054Crossref PubMed Scopus (125) Google Scholar Metabolites such as ketone bodies activate CMA,35Finn P.F. Dice J.F. Ketone bodies stimulate chaperone-mediated autophagy.J Biol Chem. 2005; 280: 25864-25870Abstract Full Text Full Text PDF PubMed Scopus (106) Google Scholar but whether it is because of their pro-oxidizing effects on proteins or by acting as second messengers in signaling pathways requires further investigation. Free fatty acids (FFAs) also activate CMA, however, their effect is biphasic and, as their levels increase, they inhibit CMA through destabilization of LAMP-2A at the lysosomal membrane.36Rodriguez-Navarro J.A. Kaushik S. Koga H. et al.Inhibitory effect of dietary lipids on chaperone-mediated autophagy.Proc Natl Acad Sci U S A. 2012; 109: E705-E714Crossref PubMed Scopus (156) Google Scholar Changes in the lipid composition of the lysosomal membrane with age also are behind the lower LAMP-2A levels and reduced CMA activity in aging.36Rodriguez-Navarro J.A. Kaushik S. Koga H. et al.Inhibitory effect of dietary lipids on chaperone-mediated autophagy.Proc Natl Acad Sci U S A. 2012; 109: E705-E714Crossref PubMed Scopus (156) Google Scholar Whether LAMP-2A instability is secondary to age-related changes in lipid metabolism, or a primary defect behind some of the metabolic changes of aging, remains unclear. In any case, this reciprocal interplay between lipid metabolism and CMA may constitute a vicious cycle that contributes to perpetuate the metabolic syndrome associated with aging. Interestingly, in support of the potential anti-aging effect of breaking this vicious circle by enhancing CMA activity, restoration of LAMP-2A levels in livers from old mice improves cellular homeostasis, enhances liver response to stress, and preserves normal hepatic function until late in life.37Zhang C. Cuervo A.M. Restoration of chaperone-mediated autophagy in aging liver improves cellular maintenance and hepatic function.Nat Med. 2008; 14: 959-965Crossref PubMed Scopus (383) Google Scholar Microautophagy occurs when small single-walled vesicles form at the lysosomal membrane and invaginate toward the lysosomal lumen while engulfing cytosolic components.38Mortimore G.E. Hutson N.J. Surmacz C.A. Quantitative correlation between proteolysis and macro- and microautophagy in mouse hepatocytes during starvation and refeeding.Proc Natl Acad Sci U S A. 1983; 80: 2179-2183Crossref PubMed Scopus (145) Google Scholar Although the original morphologic description of this process was performed in liver, the molecular mechanism and physiological relevance of hepatic microautophagy remains, for the most part, unknown. In fact, most of what is known about lysosomal microautophagy originates from studies in yeast where its selectivity for degradation of organelles, lipids, and even nuclear portions has been described.39Suzuki K. Selective autophagy in budding yeast.Cell Death Differ. 2013; 20: 43-48Crossref PubMed Scopus (89) Google Scholar, 40Uttenweiler A. Mayer A. Microautophagy in the yeast Saccharomyces cerevisiae.Methods Mol Biol. 2008; 445: 245-259Crossref PubMed Scopus (41) Google Scholar, 41van Zutphen T. 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Clement C.C. et al.Microautophagy of cytosolic proteins by late endosomes.Dev Cell. 2011; 20: 131-139Abstract Full Text Full Text PDF PubMed Scopus (584) Google Scholar The liver is essential for the maintenance of metabolic homeostasis and thereby the organismal energetic balance. Dysregulation of hepatic autophagy has been described in severe metabolic disorders such as obesity, fatty liver, and diabetes (reviewed by Schneider and Cuervo32Schneider J.L. Cuervo A.M. Liver autophagy: much more than just taking out the trash.Nat Rev Gastroenterol Hepatol. 2014; 11: 187-200Crossref PubMed Scopus (138) Google Scholar). Consequently, current efforts are directed to dissect the molecular pathways governing this recycling system and the mechanisms by which hepatic autophagy contributes to liver metabolism. Overall, these mechanisms can be grouped under 3 main functions as follows. Starvation is the best-characterized trigger for both macroautophagy and CMA activation. The first 4–6 hours of nutrient shortage inactivation of mechanistic target of rapamycin (mTOR), one of the best-characterized endogenous inhibitors of autophagy, leads to macroautophagy activation.43Kanazawa T. Taneike I. Akaishi R. et al.Amino acids and insulin control autophagic proteolysis through different signaling pathways in relation to mTOR in isolated rat hepatocytes.J Biol Chem. 2004; 279: 8452-8459Abstract Full Text Full Text PDF PubMed Scopus (179) Google Scholar Amino acids resulting from lysosomal degradation of cytosolic and organelle proteins sustain protein synthesis or directly feed the Krebs cycle to produce ATP and/or glucose3Ezaki J. Matsumoto N. Takeda-Ezaki M. et al.Liver autophagy contributes to the maintenance of blood glucose and amino acid levels.Autophagy. 2011; 7: 727-736Crossref PubMed Scopus (207) Google Scholar (Figure 2). Interestingly, some of the released amino acids inhibit autophagy, creating an auto-inhibitory feedback. The events connecting mTOR inactivation and autophagy induction have been elucidated. mTOR represses autophagy through direct phosphorylation and sequestration of Unc-51 like autophagy activating kinase 1 (ULK1), a kinase essential for autophagosome formation. Upon amino acid depletion, inactivation of mTOR is followed by cytosolic release of ULK1 and its relocation to sites of autophagosome biogenesis.44Egan D. Kim J. Shaw R.J. et al.The autophagy initiating kinase ULK1 is regulated via opposing phosphorylation by AMPK and mTOR.Autophagy. 2011; 7: 643-644Crossref PubMed Scopus (13) Google Scholar Once amino acids become available again, mTOR reactivation suppresses autophagy. Interestingly, the mechanism for autophagy inhibition seems to be amino acid–specific. For example, changes in leucine levels sensed by the leucyl–transfer RNA synthetase promote the relocation of the cytosolic mTOR complex to the lysosomal and endosomal membranes through binding to Rag–guanosine triphosphatase and RHEB.45Han J.M. Jeong S.J. Park M.C. et al.Leucyl-tRNA synthetase is an intracellular leucine sensor for the mTORC1-signaling pathway.Cell. 2012; 149: 410-424Abstract Full Text Full Text PDF PubMed Scopus (584) Google Scholar mTOR activation in this location results in inhibitory phosphorylation of TFEB, thus preventing its translocation into the nucleus, and ending the transcriptional autophagic program.46Settembre C. Zoncu R. Medina D.L. et al.A lysosome-to-nucleus signalling mechanism senses and regulates the lysosome via mTOR and TFEB.EMBO J. 2012; 31: 1095-1108Crossref PubMed Scopus (1250) Google Scholar The modulatory effect of other amino acids is less well characterized and apparently more complex. For example, both intracellular uptake of glutamine through SLC1A5 as well as its efflux activate mTOR, this is owing to coupling of the exit of this amino acid through the bidirectional transporter SLC7A5/SLC3A2 to intracellular influx of leucine and essential amino acids. Blockage of these transporters results in glutamine depletion and autophagy activation.47Nicklin P. Bergman P. Zhang B. et al.Bidirectional transport of amino acids regulates mTOR and autophagy.Cell. 2009; 136: 521-534Abstract Full Text Full Text PDF PubMed Scopus (1267) Google Scholar The physiologic relevance of autophagy activation as a source of amino acids is exemplified by the death of autophagy-deficient neonate mice during the period between the interruption at birth of the transplacental-nutrient supply and lactation. Administration of amino acids is sufficient to overcome the negative energetic balance of these autophagy-deficient neonates and guarantee their survival.4Kuma A. Hatano M. Matsui M. et al.The role of autophagy during the early neonatal starvation period.Nature. 2004; 432: 1032-1036Crossref PubMed Scopus (2386) Google Scholar In many cell types, if starvation persists beyond 8 hours, the contribution of macroautophagy to protein breakdown gradually decreases and it is replaced by activation of CMA, which peaks at 24 hours and can persist for up to 3 days into starvation.31Cuervo A.M. Knecht E. Terlecky S.R. et al.Activation of a selective pathway of lysosomal proteolysis in rat liver by prolonged starvation.Am J Physiol. 1995; 269: C1200-C1208Crossref PubMed G