Sestrin mediates detection of and adaptation to low-leucine diets in Drosophila.

Mechanistic target of rapamycin complex 1 (mTORC1) regulates cell growth and metabolism in response to multiple nutrients, including the essential amino acid leucine1. Recent work in cultured mammalian cells established the Sestrins as leucine-binding proteins that inhibit mTORC1 signalling during leucine deprivation2,3, but their role in the organismal response to dietary leucine remains elusive. Here we find that Sestrin-null flies (Sesn-/-) fail to inhibit mTORC1 or activate autophagy after acute leucine starvation and have impaired development and a shortened lifespan on a low-leucine diet. Knock-in flies expressing a leucine-binding-deficient Sestrin mutant (SesnL431E) have reduced, leucine-insensitive mTORC1 activity. Notably, we find that flies can discriminate between food with or without leucine, and preferentially feed and lay progeny on leucine-containing food. This preference depends on Sestrin and its capacity to bind leucine. Leucine regulates mTORC1 activity in glial cells, and knockdown of Sesn in these cells reduces the ability of flies to detect leucine-free food. Thus, nutrient sensing by mTORC1 is necessary for flies not only to adapt to, but also to detect, a diet deficient in an essential nutrient.

SAR1B senses leucine levels to regulate mTORC1 signalling.

The mTOR complex 1 (mTORC1) controls cell growth in response to amino acid levels1. Here we report SAR1B as a leucine sensor that regulates mTORC1 signalling in response to intracellular levels of leucine. Under conditions of leucine deficiency, SAR1B inhibits mTORC1 by physically targeting its activator GATOR2. In conditions of leucine sufficiency, SAR1B binds to leucine, undergoes a conformational change and dissociates from GATOR2, which results in mTORC1 activation. SAR1B-GATOR2-mTORC1 signalling is conserved in nematodes and has a role in the regulation of lifespan. Bioinformatic analysis reveals that SAR1B deficiency correlates with the development of lung cancer. The silencing of SAR1B and its paralogue SAR1A promotes mTORC1-dependent growth of lung tumours in mice. Our results reveal that SAR1B is a conserved leucine sensor that has a potential role in the development of lung cancer.

Activation of GCN2 in macrophages promotes white adipose tissue browning and lipolysis under leucine deprivation.

We have previously shown that leucine deprivation stimulates browning and lipolysis in white adipose tissue (WAT), which helps to treat obesity. Adipose tissue macrophages (ATMs) significantly influence WAT browning and lipolysis. However, it is unclear whether ATMs are involved in leucine deprivation-induced browning and lipolysis in WAT; the associated signals remain to be elucidated. Here, we investigated the role of ATMs and the possible mechanisms involved in WAT browning and lipolysis under leucine-deprivation conditions. In this study, macrophages were depleted in mice by injecting clodronate-liposomes (CLOD) into subcutaneous white adipose tissues. Then, mice lacking general control nonderepressible 2 kinase (GCN2), which is a sensor of amino acid starvation, specifically in Lyz2-expressing cells, were generated to investigate the changes in leucine deprivation-induced WAT browning and lipolysis. We found leucine deprivation decreased the accumulation and changed the polarization of ATMs. Ablation of macrophages by CLOD impaired WAT browning and lipolysis under leucine-deprivation conditions. Mechanistically, leucine deprivation activated GCN2 signals in macrophages. Myeloid-specific abrogation of GCN2 in mice blocked leucine deprivation-induced browning and lipolysis in WAT. Further analyses revealed that GCN2 activation in macrophages reduced the expression of monoamine oxidase A (MAOA), resulting in increased norepinephrine (NE) secretion from macrophages to adipocytes, and this resulted in enhanced WAT browning and lipolysis. Moreover, the injection of CL316,243, a ß3-adrenergic receptor agonist, and inhibition of MAOA effectively increased the level of NE, leading to the enhancement of browning and lipolysis of WAT in myeloid GCN2 knockout mice under leucine deprivation. Collectively, our results demonstrate a novel function of GCN2 signals in macrophages, that is, regulating WAT browning and lipolysis under leucine deprivation. Our study provides important hints for possible treatment for obesity.

ATF4-Mediated Upregulation of REDD1 and Sestrin2 Suppresses mTORC1 Activity during Prolonged Leucine Deprivation.

BACKGROUND: The protein kinase target of rapamycin (mTOR) in complex 1 (mTORC1) is activated by amino acids and in turn upregulates anabolic processes. Under nutrient-deficient conditions, e.g., amino acid insufficiency, mTORC1 activity is suppressed and autophagy is activated. Intralysosomal amino acids generated by autophagy reactivate mTORC1. However, sustained mTORC1 activation during periods of nutrient insufficiency would likely be detrimental to cellular homeostasis. Thus, mechanisms must exist to prevent amino acids released by autophagy from reactivating the kinase. OBJECTIVE: The objective of the present study was to test whether mTORC1 activity is inhibited during prolonged leucine deprivation through ATF4-dependent upregulation of the mTORC1 suppressors regulated in development and DNA damage response 1 (REDD1) and Sestrin2. METHODS: Mice (8 wk old; C57Bl/6 × 129SvEV) were food deprived (FD) overnight and one-half were refed the next morning. Mouse embryo fibroblasts (MEFs) deficient in ATF4, REDD1, and/or Sestrin2 were deprived of leucine for 0-16 h. mTORC1 activity and ATF4, REDD1, and Sestrin2 expression were assessed in liver and cell lysates. RESULTS: Refeeding FD mice resulted in activation of mTORC1 in association with suppressed expression of both REDD1 and Sestrin2 in the liver. In cells in culture, mTORC1 exhibited a triphasic response to leucine deprivation, with an initial suppression followed by a transient reactivation from 2 to 4 h and a subsequent resuppression after 8 h. Resuppression occurred concomitantly with upregulated expression of ATF4, REDD1, and Sestrin2. However, in cells lacking ATF4, neither REDD1 nor Sestrin2 expression was upregulated by leucine deprivation, and resuppression of mTORC1 was absent. Moreover, in cells lacking either REDD1 or Sestrin2, mTORC1 resuppression was attenuated, and in cells lacking both proteins resuppression was further blunted. CONCLUSIONS: The results suggest that leucine deprivation upregulates expression of both REDD1 and Sestrin2 in an ATF4-dependent manner, and that upregulated expression of both proteins is involved in resuppression of mTORC1 during prolonged leucine deprivation.

Nutritional deficiency dermatitis related to branched-chain amino acid restriction in a child with maple syrup urine disease.

We present a one-year-old girl with maple syrup urine disease with dermatitis secondary to the restriction of amino acids as part of the treatment. We present the clinical evolution and histopathological correlation.

A novel function of B-cell translocation gene 1 (BTG1) in the regulation of hepatic insulin sensitivity in mice via c-Jun.

Insulin resistance is one of the major factors contributing to metabolic diseases, but the underlying mechanisms are still poorly understood. As an important cofactor, B-cell translocation gene 1 (BTG1) is involved in many physiologic processes; however, the direct effect of BTG1 on insulin sensitivity has not been described. In our study, BTG1 overexpression or knockdown improved or impaired insulin signaling in vitro, respectively. In addition, adenovirus-mediated BTG1 overexpression improved insulin sensitivity in wild-type (WT) and insulin-resistant leptin-receptor mutated (db/db) mice. In addition, transgenic BTG1-overexpressing mice were resistant to high-carbohydrate diet-induced insulin resistance. Adenovirus-mediated BTG1 knockdown consistently impaired insulin sensitivity in WT and insulin-sensitive leucine-deprived mice. Moreover, hepatic BTG1 expression was increased by leucine deprivation via the mammalian target of rapamycin/ribosomal protein S6 kinase 1 pathway. Furthermore, c-Jun expression was up-regulated by BTG1, and adenovirus-mediated c-Jun knockdown blocked BTG1-improved insulin signaling and insulin sensitivity in vitro and in vivo. Finally, BTG1 promoted c-Jun expression via stimulating c-Jun and retinoic acid receptor activities. Taken together, these results identify a novel function for BTG1 in the regulation of hepatic insulin sensitivity and provide important insights into the nutritional regulation of BTG1 expression.- Xiao, F., Deng, J., Yu, J., Guo, Y., Chen, S., Guo, F. A novel function of B-cell translocation gene 1 (BTG1) in the regulation of hepatic insulin sensitivity in mice via c-Jun.

Towards the modulation of oxidative damage, apoptosis and tight junction protein in response to dietary leucine deficiency: A likely cause of ROS-induced gill structural integrity impairment.

The current study explored the protective effect of leucine on antioxidant status, apoptosis and tight junction damage in the gill of grass carp (Ctenopharyngodon idella Val.). The trial was conducted by feeding grass carp with six graded level of leucine (7.1, 8.9, 11.0, 13.3, 15.2 and 17.1 g kg-1 diet) for 8 weeks. The fish were fed to apparent satiation 4 times per day. The results indicated that compared with the leucine deficiency group, 8.9-11.3 g leucine kg-1 diet supplementations decreased protein carbonyl (PC), malondialdehyde (MDA) and ROS contents, which may be partially attributed to the improvement of antioxidant status in the gill by increasing hydroxyl radical capacity and anti-superoxide radicals, glutathione contents and the activities and mRNA levels of Cu/Zn superoxide dismutase (SOD1), glutathione peroxidase (GPx), catalase (CAT), glutathione-S-transferase (GST) and glutathione reductase (GR), that referring to the up-regulation of nuclear factor erythroid 2-related factor 2 (Nrf2) mRNA expression. Moreover, leucine deficiency induced DNA fragmentation via the up-regulation of caspase-3, caspase-8 and caspase-9 expressions and down-regulation of target of rapamycin and ribosomal S6 protein kinase 1 expressions. Furthermore, leucine deficiency increased interleukin-1ß (IL-1ß), interleukin-8 (IL-8) and tumor necrosis factor-α (TNF-α) mRNA expression and decreased IL-10 and transforming growth factor ß (TGF-ß), which was partly related to nuclear factor κB (NF-κB) and its inhibitor (IκB). In contrast, the relative mRNA expression of IL-1, IL-8 and TNF-α was down-regulated with 8.9-11.3 g leucine kg-1 diet supplementations. Finally, the relative mRNA expression of tight junction protein, including occludin, zonula occludens-1, claudin b, claudin 3 and claudin 12 was up-regulated with leucine diet supplementations. Our results indicate that leucine protected the fish gill structural integrity partially because of the inhibition of apoptosis, the improvement of antioxidant status, the regulation of tight junction protein and related signalling molecules mRNA expressions in the fish gill.

Leptin signaling is required for leucine deprivation-enhanced energy expenditure.

Leptin signaling in the hypothalamus is crucial in energy homeostasis. We have previously shown that dietary deprivation of the essential amino acid leucine in mice stimulates fat loss by increasing energy expenditure. The involvement of leptin signaling in this regulation, however, has not been reported. Here, we show that leucine deprivation promotes leptin signaling in mice maintained on an otherwise normal diet and restores leptin responses in mice maintained on a high fat diet, a regimen known to induce leptin resistance. In addition, we found that leucine deprivation stimulated energy expenditure, and fat loss was largely blocked in db/db mice homozygous for a mutation in leptin receptor and a knock-in mouse line Y3F with abrogation of leptin receptor Tyr(1138)-mediated signal transducer and activator transcript 3 signaling. Overall, our studies describe a novel link between hypothalamic leptin signaling and stimulation of energy expenditure under leucine deprivation.

CREB/TRH pathway in the central nervous system regulates energy expenditure in response to deprivation of an essential amino acid.

BACKGROUND: In the central nervous system (CNS), thyrotropin-releasing hormone (TRH) has an important role in regulating energy balance. We previously showed that dietary deprivation of leucine in mice increases energy expenditure through CNS-dependent regulation. However, the involvement of central TRH in this regulation has not been reported. METHODS: Male C57J/B6 mice were maintained on a control or leucine-deficient diet for 7 days. Leucine-deprived mice were either third intracerebroventricular (i.c.v.) injected with a TRH antibody followed by intraperitoneal (i.p.) injection of triiodothyronine (T3) or i.c.v. administrated with an adenovirus of shCREB (cAMP-response element binding protein) followed by i.c.v. injection of TRH. Food intake and body weight were monitored daily. Oxygen consumption, physical activity and rectal temperature were assessed after the treatment. After being killed, the hypothalamus and the brown adipose tissue were collected and the expression of related genes and proteins related was analyzed. In other experiments, control or leucine-deficient medium incubated primary cultured neurons were either infected with adenovirus-mediated short hairpin RNA targeting extracellular signal-regulated kinases 1 and 2 (Ad-shERK1/2) or transfected with plasmid-overexpressing protein phosphatase 1 regulatory subunit 3C (PPP1R3C). RESULTS: I.c.v. administration of anti-TRH antibodies significantly reduced leucine deprivation-stimulated energy expenditure. Furthermore, the effects of i.c.v. TRH antibodies were reversed by i.p. injection of T3 during leucine deprivation. Moreover, i.c.v. injection of Ad-shCREB (adenovirus-mediated short hairpin RNA targeting CREB) significantly suppressed leucine deprivation-stimulated energy expenditure via modulation of TRH expression. Lastly, TRH expression was regulated by CREB, which was phosphorylated by ERK1/2 and dephosphorylated by PPP1R3C-containing protein Ser/Thr phosphatase type 1 (PP1) under leucine deprivation in vitro. CONCLUSIONS: Our data indicate a novel role for TRH in regulating energy expenditure via T3 during leucine deprivation. Furthermore, our findings reveal that TRH expression is activated by CREB, which is phosphorylated by ERK1/2 and dephosphorylated by PPP1R3C-containing PP1. Collectively, our studies provide novel insights into the regulation of energy homeostasis by the CNS in response to an essential amino-acid deprivation.

Leucine and methionine deficiency impairs immunity to gastrointestinal parasites during lactation.

Lactating rats reinfected with Nippostrongylus brasiliensis fed low-crude protein (CP) foods show reduced lactational performance and less resistance to parasites compared with their high-CP counterparts. Here, we hypothesised that feeding high-CP foods deficient in specific essential amino acids (AA) would result in similar penalties. Second-parity lactating rats, immunised with 1600 N. brasiliensis infective larvae before mating, were fed foods with either 250 (high protein; HP) or 150 (low protein; LP) g CP/kg, or were HP deficient in either leucine (HP-Leu) or methionine (HP-Met). On day 1 of lactation, litter size was standardised at twelve pups. On day 2, dams were either reinfected with 1600 N. brasiliensis larvae or sham-infected with PBS. Dams and litters were weighed daily until either day 8 or 11, when worm burdens, and inflammatory cells and systemic levels of N. brasiliensis-specific Ig isotypes were assessed. Data from five out of sixteen HP-Met rats were omitted due to very high levels of food refusals from parturition onwards. Relative to feeding HP foods, feeding LP, HP-Met and HP-Leu foods reduced dam weight gain and, to a lesser extent, litter weight gain, and increased the number of worm eggs in the colon, indicative of a reduction in resistance to parasites. However, only feeding LP and HP-Leu foods resulted in increased worm numbers, while none of the feeding treatments affected systemic Ig, mast and goblet cells, and eosinophil numbers. The present results support the view that resistance to parasites during lactation may be sensitive to specific essential AA scarcity.

Leucine limitation regulates myf5 and myoD expression and inhibits myoblast differentiation.

Satellite cells are the major pool of muscle stem cells after birth; they represent an important component required to maintain muscle mass and functionality during life. The molecular mechanisms involved in myogenic differentiation are relatively well-known. However, the role of extracellular stimulus in the control of differentiation remains largely unresolved. Notably little is known about the impact of nutrients on this process. Here we have studied the role of leucine, an essential amino acid, in the control of myogenic differentiation. Leucine is a well-known regulator of muscle protein synthesis. It acts not only as a substrate for translation but also as a regulator of gene expression and signaling pathways such as those involving mTOR and GCN2. In this study we demonstrated that the lack of leucine abolishes the differentiation of both C2C12 myoblasts and primary satellite cells. This effect is associated with a modification of the pattern of expression of the myogenic regulatory factors (MRF) myf5 and myoD. We report an up-regulation of myf5 mRNA and a decrease of myoD protein level during leucine starvation. This study demonstrates the importance of a nutrient, leucine, in the control of the myogenic differentiation program.

The metabolic sensor GCN2 branches out.

GCN2 is a sensor of amino acid deprivation that triggers a repression of global protein synthesis while simultaneously inducing translation of specific proteins. In this issue of Cell Metabolism, Guo and Cavener (2007) present a much broader role for GCN2 in controlling lipid homeostasis in response to amino acid deprivation.

The GCN2 eIF2alpha kinase regulates fatty-acid homeostasis in the liver during deprivation of an essential amino acid.

Metabolic adaptation is required to cope with episodes of protein deprivation and malnutrition. GCN2 eIF2alpha kinase, a sensor of amino acid deficiency, plays a key role in yeast and mammals in modulating amino acid metabolism as part of adaptation to nutrient deprivation. The role of GCN2 in adaptation to long-term amino acid deprivation in mammals, however, is poorly understood. We found that expression of lipogenic genes and the activity of fatty acid synthase (FAS) in the liver are repressed and lipid stores in adipose tissue are mobilized in wild-type mice upon leucine deprivation. In contrast, GCN2-deficient mice developed liver steatosis and exhibited reduced lipid mobilization. Liver steatosis in Gcn2(-/-) mice was found to be caused by unrepressed expression of lipogenic genes, including Srebp-1c and Fas. Thus, our study identifies a novel function of GCN2 in regulating lipid metabolism during leucine deprivation in addition to regulating amino acid metabolism.

Isoleucine and leucine independently regulate mTOR signaling and protein synthesis in MAC-T cells and bovine mammary tissue slices.

Understanding the regulatory effects of individual amino acids (AA) on milk protein synthesis rates is important for improving protein and AA requirement models for lactation. The objective of this study was to examine the effects of individual essential AA (EAA) on cellular signaling and fractional protein synthesis rates (FSR) in bovine mammary cells. Omission of L-arginine, L-isoleucine, L-leucine, or all EAA reduced (P < 0.05) mammalian target of rapamycin (mTOR; Ser2448) and ribosomal protein S6 (rpS6; Ser235/236) phosphorylation in MAC-T cells. Phosphorylation of mTOR and rpS6 kinase 1 (S6K1; Thr389) decreased (P < 0.05) in the absence of L-isoleucine, L-leucine, or all EAA in lactogenic mammary tissue slices. Omission of L-tryptophan also reduced S6K1 phosphorylation (P = 0.01). Supplementation of L-leucine to media depleted of EAA increased mTOR and rpS6 and decreased eukaryotic elongation factor 2 (Thr56) phosphorylation (P < 0.05) in MAC-T cells. Supplementation of L-isoleucine increased mTOR, S6K1, and rpS6 phosphorylation (P < 0.05). No single EAA considerably affected eukaryotic initiation factor 2-α (eIF2α; Ser51) phosphorylation, but phosphorylation was reduced in response to provision of all EAA (P < 0.04). FSR declined when L-isoleucine (P = 0.01), L-leucine (P = 0.01), L-methionine (P = 0.02), or L-threonine (P = 0.07) was depleted in media and was positively correlated (R = 0.64, P < 0.01) with phosphorylation of mTOR and negatively correlated (R = -0.42, P = 0.01) with phosphorylation of eIF2α. Such regulation of protein synthesis will result in variable efficiency of transfer of absorbed EAA to milk protein and is incompatible with the assumption that a single nutrient limits protein synthesis that is encoded in current diet formulation strategies.

Intermittent Leucine Deprivation Produces Long-lasting Improvement in Insulin Sensitivity by Increasing Hepatic Gcn2 Expression.

Leucine deprivation improves insulin sensitivity; however, whether and how this effect can be extended are unknown. We hypothesized that intermittent leucine deprivation (ILD) might produce a long-term effect on improved insulin sensitivity via the formation of metabolic memory. Consistently, seven ILD cycles of treatment (1-day leucine-deficient diet, 3-day control diet) in mice produced a long-lasting (after a control diet was resumed for 49 days) effect on improved whole-body and hepatic insulin sensitivity in mice, indicating the potential formation of metabolic memory. Furthermore, the effects of ILD depended on hepatic general control nondepressible 2 (GCN2) expression, as verified by gain- and loss-of-function experiments. Moreover, ILD increased Gcn2 expression by reducing its DNA methylation at two CpG promoter sites controlled by demethylase growth arrest and DNA damage inducible b. Finally, ILD also improved insulin sensitivity in insulin-resistant mice. Thus, ILD induces long-lasting improvements in insulin sensitivity by increasing hepatic Gcn2 expression via a reduction in its DNA methylation. These results provide novel insights into understanding of the link between leucine deprivation and insulin sensitivity, as well as potential nutritional intervention strategies for treating insulin resistance and related diseases. We also provide evidence for liver-specific metabolic memory after ILD and novel epigenetic mechanisms for Gcn2 regulation.

Amino acid deprivation regulates the stress-inducible gene p8 via the GCN2/ATF4 pathway.

In mammals, the GCN2/ATF4 pathway has been described as the main pathway involved in the regulation of gene expression upon amino acid limitation. This regulation is notably conferred by the presence of a cis-element called Amino Acid Response Element (AARE) in the promoter of specific genes. In vivo, the notion of amino acid limitation is not limited to nutritional context, indeed several pathological situations are associated with alteration of endogenous amino acid availability. This is notably true in the context of tumour in which the alteration of the microenvironment can lead to a perturbation in nutrient availability. P8 is a small weakly folded multifunctional protein that is overexpressed in several kinds of cancers and whose expression is induced by different stresses. In this study we have demonstrated that amino acid starvation was also able to induce p8 expression. Moreover, we brought the evidence, in vitro and in vivo, that the GCN2/ATF4 pathway is involved in this regulation through the presence of an AARE in p8 promoter.

Identification of GCN2 as new redox regulator for oxidative stress prevention in vivo.

Constitution of oxidative defense systems and, correspondingly, oxidative stress prevention are highly dependent on amino acid supply. In vitro, experiments have demonstrated that amino acid availability participates to the homeostasis of reactive oxygen species. However the molecular mechanisms involved in the maintenance of redox homeostasis responsive to circulating amino acid levels remain unclear. As GCN2 is a protein kinase considered to be an important sensor for amino acids availability and a potential regulator of redox homeostasis, we hypothesized that this kinase can modulate redox homeostasis in vivo, in response to an amino acid-imbalanced diet. We investigated the response of GCN2+/+ and GCN2-/- mice to a long-term (24 weeks) leucine-imbalanced diet (EDΔLeu). In order to evaluate the oxidation level in each group of mice, we determined the degree of protein oxidation in the liver. Interestingly, GCN2-/- mice exhibited an increase in protein carbonylation, a marker of oxidative stress, in response to the EDΔLeu diet. These data correlate with a decrease in hepatic GPX1 expression, a major antioxidant enzyme, and a decrease in total GPX activity in the liver. Our results suggest that GCN2 and its downstream signaling pathway have an important role in the protection against oxidative injuries induced by an amino acid-imbalanced diet, and that it can play a critical role in the prevention of oxidative damage.

Gene expression and integrated stress response in HepG2/C3A cells cultured in amino acid deficient medium.

The integrated stress response (ISR), a defense mechanism cells employ when under stress (e.g., amino acid deprivation), causes suppression of global protein synthesis along with the paradoxical increased expression of a host of proteins that are useful in combating various stresses. Genes that were similarly differentially expressed under conditions of either leucine- or cysteine-depletion were identified. Many of the genes known to contain an amino acid response element and to be induced in response to eIF2α phosphorylation and ATF4 heterodimer binding (ATF3, C/EBPß, SLC7A1, SLC7A11, and TRIB3), as well as others shown to be induced downstream of eIF2α phosphorylation (C/EBPγ, CARS, SARS, CLCN3, CBX4, and PPP1R15A) were among the upregulated genes. Evidence for the induction of the ISR in these cells also included the increased phosphorylation of eIF2α and increased protein abundance of ATF4, ATF3, and ASNS in cysteine- and leucine-depleted cells. Based on genes highly differentially expressed in both leucine- and cysteine-deficient cells, a list of 67 downregulated and 53 upregulated genes is suggested as likely targets of essential amino acid deprivation in mammalian cells.

Influence of genotype and nutrition on survival and metabolism of starving yeast.

Starvation of yeast cultures limited by auxotrophic requirements results in glucose wasting and failure to achieve prompt cell-cycle arrest when compared with starvation for basic natural nutrients like phosphate or sulfate. We measured the survival of yeast auxotrophs upon starvation for different nutrients and found substantial differences: When deprived of leucine or uracil, viability declined exponentially with a half-life of <2 days, whereas when the same strains were deprived of phosphate or sulfate, the half-life was approximately 10 days. The survival rates of nongrowing auxotrophs deprived of uracil or leucine depended on the carbon source available during starvation, but were independent of the carbon source during prior growth. We performed an enrichment procedure for mutants that suppress lethality during auxotrophy starvation. We repeatedly found loss-of-function mutations in a number of functionally related genes. Mutations in PPM1, which methylates protein phosphatase 2A, and target of rapamycin (TOR1) were characterized further. Deletion of PPM1 almost completely suppressed the rapid lethality and substantially suppressed glucose wasting during starvation for leucine or uracil. Suppression by a deletion of TOR1 was less complete. We suggest that, similar to the Warburg effect observed in tumor cells, starving yeast auxotrophs wastes glucose as a consequence of the failure of conserved growth control pathways. Furthermore, we suggest that our results on condition-dependent chronological lifespan have important implications for the interpretation and design of studies on chronological aging.

Deficiency in the Essential Amino Acids l-Isoleucine, l-Leucine and l-Histidine and Clinical Measures as Predictors of Moderate Depression in Elderly Women: A Discriminant Analysis Study.

Increases in depression are common in some elderly women. Elderly women often show moderate depressive symptoms, while others display minimal depressive symptoms. These discrepancies have produced contradictory and inconclusive outcomes, which have not been explained entirely by deficits in neurotransmitter precursors. Deficiency in some amino acids have been implicated in major depression, but its role in non-clinical elderly women is not well known. An analysis of essential amino acids, depression and the use of discriminant analysis can help to clarify the variation in depressive symptoms exhibited by some elderly women. The aim was to investigate the relationship of essential amino acids with affective, cognitive and comorbidity measures in elderly women without major depression nor severe mood disorders or psychosis, specifically thirty-six with moderate depressive symptoms and seventy-one with minimal depressive symptoms. The plasma concentrations of nineteen amino acids, Beck Depression Inventory (BDI) scores, Geriatric Depression Scale (GDS) scores, global cognitive scores and comorbidities were submitted to stepwise discriminant analysis to identify predictor variables. Seven predictors arose as important for belong to the group based on amino acid concentrations, with the moderate depressive symptoms group characterized by higher BDI, GDS and cognitive scores; fewer comorbidities; and lower levels of l-histidine, l-isoleucine and l-leucine. These findings suggest that elderly women classified as having moderate depressive symptoms displayed a deficiency in essential amino acids involved in metabolism, protein synthesis, inflammation and neurotransmission.