Nutrient-sensing Kinase Research Articles

Cytotoxic stress caused by azalamellarin D (AzaD) interferes with cellular protein translation by targeting the nutrient-sensing kinase mTOR.

Analogs of pyrrole alkaloid lamellarins exhibit anticancer activity by modulating multiple cellular events. Lethal doses of several lamellarins were found to enhance autophagy flux in HeLa cells, suggesting that lamellarins may modulate protein homeostasis through the interference of proteins or kinases controlling energy and nutrient metabolism. To further delineate molecular mechanisms and their targets, our results herein show that azalamellarin D (AzaD) cytotoxicity could cause translational attenuation, as indicated by a change in eIF2α phosphorylation. Intriguingly, acute AzaD treatment promoted the phosphorylation of GCN2, a kinase that transduces the integrated stress response (ISR), and prolonged exposure to AzaD could increase the levels of the phosphorylated forms of eIF2α and the other ISR kinase protein kinase R (PKR). However, the effects of AzaD on ISR signalling were marginally abrogated in cells with genetic deletion of GCN2 and PKR, and evaluation of protein target engagement by cellular thermal shift assay (CETSA) revealed no significant interaction between AzaD and ISR kinases. Further investigation revealed that acute AzaD treatment negatively affected mechanistic target of rapamycin (mTOR) phosphorylation and signalling. The analyses by CETSA and computational modelling indicated that mTOR may be a possible protein target for AzaD. These findings indicate the potential for developing lamellarins as novel agents for cancer treatment.

Ecl family genes: Factors linking starvation and lifespan extension in Schizosaccharomyces pombe.

In the fission yeast Schizosaccharomyces pombe, the duration of survival in the stationary phase, termed the chronological lifespan (CLS), is affected by various environmental factors and the corresponding gene activities. The ecl family genes were identified in the genomic region encoding non-coding RNA as positive regulators of CLS in S. pombe, and subsequently shown to encode relatively short proteins. Several studies revealed that ecl family genes respond to various nutritional starvation conditions via different mechanisms, and they are additionally involved in stress resistance, autophagy, sexual differentiation, and cell cycle control. Recent studies reported that Ecl family proteins strongly suppress target of rapamycin complex 1, which is a conserved eukaryotic nutrient-sensing kinase complex that also regulates longevity in a variety of organisms. In this review, we introduce the regulatory mechanisms of Ecl family proteins and discuss their emerging findings.

Nutrient-sensing mTORC1 and AMPK pathways in chronic kidney diseases.

Nutrients such as glucose, amino acids and lipids are fundamental sources for the maintenance of essential cellular processes and homeostasis in all organisms. The nutrient-sensing kinases mechanistic target of rapamycin (mTOR) and AMP-activated protein kinase (AMPK) are expressed in many cell types and have key roles in the control of cell growth, proliferation, differentiation, metabolism and survival, ultimately contributing to the physiological development and functions of various organs, including the kidney. Dysregulation of these kinases leads to many human health problems, including cancer, neurodegenerative diseases, metabolic disorders and kidney diseases. In the kidney, physiological levels of mTOR and AMPK activity are required to support kidney cell growth and differentiation and to maintain kidney cell integrity and normal nephron function, including transport of electrolytes, water and glucose. mTOR forms two functional multi-protein kinase complexes, mTOR complex 1 (mTORC1) and mTOR complex 2 (mTORC2). Hyperactivation of mTORC1 leads to podocyte and tubular cell dysfunction and vulnerability to injury, thereby contributing to the development of chronic kidney diseases, including diabetic kidney disease, obesity-related kidney disease and polycystic kidney disease. Emerging evidence suggests that targeting mTOR and/or AMPK could be an effective therapeutic approach to controlling or preventing these diseases.

A phosphorylation network controls the stability of alpha‐arrestins Aly1/Art6 and Aly2/Art3

Selective protein trafficking controls the repertoire of membrane proteins at the cell surface. This contingent of surface proteins regulates nutrient/metabolite balance and response to extracellular cues. Environmental changes trigger cellular signaling that drives transitions in the plasma membrane proteome. These alterations are achieved in no small part through regulation of the α‐arrestins, an emergent and powerful class of protein trafficking adaptors. The α‐arrestins are largely cytosolic proteins that are transiently recruited to membranes via binding to membrane protein motifs. α‐Arrestins bring with them a ubiquitin ligase, which stimulates ubiquitination and subsequent endocytosis of membrane proteins. Phosphorylation of α‐arrestins is key to regulating their trafficking function; dephosphorylated α‐arrestins are generally ‘active’ endocytic adaptors. While some kinases and phosphatases for α‐arrestins are known, the degree of α‐arrestins phosphorylation is staggering, with >40 phosphorylation sites identified for a single α‐arrestin, and suggests we still have much to learn about α‐arrestins regulation. We sought to determine what kinases and phosphates regulate paralogous α‐arrestins Aly1 and Aly2 using a genetic screen in yeast. Using an array of all known non‐essential kinase and phosphatase deletions, we determined which of these influenced α‐arrestin‐mediated resistance to rapamycin, an inhibitor of the TORC1 nutrient‐sensing kinase. We identified a large cohort of kinases and phosphates that influenced Aly‐dependent phenotypes, causing electrophoretic mobility changes and, in many cases, diminishing the abundance of these α‐arrestins. We focused our studies on the Sit4 protein phosphatase, a key regulator downstream of TORC1 signaling able to influence other α‐arrestins, identified in our screen. Strikingly, Aly1 and Aly2 were hyperphosphorylated and destabilized in the absence of Sit4, suggesting that excessive phosphorylation may promote degradation of these α‐arrestins. For Aly2, but not Aly1, degradation in the sit4∆ cells could be reversed by loss of the vacuolar protease Pep4, indicating that vacuolar degradation predominates for Aly2 under these conditions. Loss of the Npr1 kinase in sit4∆ cells restored Aly protein abundances and reversed the hyperphosphorylation, demonstrating that this kinase is responsible for the excess Aly phosphorylation in sit4∆ cells. This is a remarkable finding as typically Npr1 is considered inactive in sit4∆, however, we suggest that Npr1 is selectively active in the absence of Sit4, able to modify some substrates but not others. We define new features of TORC1 signaling in regulating α‐arrestin phosphorylation and stability.

Phosphate Restriction Promotes Longevity via Activation of Autophagy and the Multivesicular Body Pathway.

Nutrient limitation results in an activation of autophagy in organisms ranging from yeast, nematodes and flies to mammals. Several evolutionary conserved nutrient-sensing kinases are critical for efficient adaptation of yeast cells to glucose, nitrogen or phosphate depletion, subsequent cell-cycle exit and the regulation of autophagy. Here, we demonstrate that phosphate restriction results in a prominent extension of yeast lifespan that requires the coordinated activity of autophagy and the multivesicular body pathway, enabling efficient turnover of cytoplasmic and plasma membrane cargo. While the multivesicular body pathway was essential during the early days of aging, autophagy contributed to long-term survival at later days. The cyclin-dependent kinase Pho85 was critical for phosphate restriction-induced autophagy and full lifespan extension. In contrast, when cell-cycle exit was triggered by exhaustion of glucose instead of phosphate, Pho85 and its cyclin, Pho80, functioned as negative regulators of autophagy and lifespan. The storage of phosphate in form of polyphosphate was completely dispensable to in sustaining viability under phosphate restriction. Collectively, our results identify the multifunctional, nutrient-sensing kinase Pho85 as critical modulator of longevity that differentially coordinates the autophagic response to distinct kinds of starvation.

Arginine Signaling and Cancer Metabolism.

Simple SummaryIn this review, we describe arginine’s role as a signaling metabolite, epigenetic regulator and mitochondrial modulator in cancer cells, and summarize recent progress in the application of arginine deprivation as a cancer therapy.Arginine is an amino acid critically involved in multiple cellular processes including the syntheses of nitric oxide and polyamines, and is a direct activator of mTOR, a nutrient-sensing kinase strongly implicated in carcinogenesis. Yet, it is also considered as a non- or semi-essential amino acid, due to normal cells’ intrinsic ability to synthesize arginine from citrulline and aspartate via ASS1 (argininosuccinate synthase 1) and ASL (argininosuccinate lyase). As such, arginine can be used as a dietary supplement and its depletion as a therapeutic strategy. Strikingly, in over 70% of tumors, ASS1 transcription is suppressed, rendering the cells addicted to external arginine, forming the basis of arginine-deprivation therapy. In this review, we will discuss arginine as a signaling metabolite, arginine’s role in cancer metabolism, arginine as an epigenetic regulator, arginine as an immunomodulator, and arginine as a therapeutic target. We will also provide a comprehensive summary of ADI (arginine deiminase)-based arginine-deprivation preclinical studies and an update of clinical trials for ADI and arginase. The different cell killing mechanisms associated with various cancer types will also be described.

AMPK phosphorylates PPARδ to mediate its stabilization, inhibit glucose and glutamine uptake and colon tumor growth

Peroxisome proliferator–activated receptor δ (PPARδ) is a nuclear receptor transcription factor that plays an important role in the regulation of metabolism, inflammation, and cancer. In addition, the nutrient-sensing kinase 5′AMP-activated protein kinase (AMPK) is a critical regulator of cellular energy in coordination with PPARδ. However, the molecular mechanism of the AMPK/PPARδ pathway on cancer progression is still unclear. Here, we found that activated AMPK induced PPARδ-S50 phosphorylation in cancer cells, whereas the PPARδ/S50A (nonphosphorylation mimic) mutant reversed this event. Further analysis showed that the PPARδ/S50E (phosphorylation mimic) but not the PPARδ/S50A mutant increased PPARδ protein stability, which led to reduced p62/SQSTM1-mediated degradation of misfolded PPARδ. Furthermore, PPARδ-S50 phosphorylation decreased PPARδ transcription activity and alleviated PPARδ-mediated uptake of glucose and glutamine in cancer cells. Soft agar and xenograft tumor model analysis showed that the PPARδ/S50E mutant but not the PPARδ/S50A mutant inhibited colon cancer cell proliferation and tumor growth, which was associated with inhibition of Glut1 and SLC1A5 transporter protein expression. These findings reveal a new mechanism of AMPK-induced PPARδ-S50 phosphorylation, accumulation of misfolded PPARδ protein, and inhibition of PPARδ transcription activity contributing to the suppression of colon tumor formation.

Metabolic Stress and Immunity: Nutrient-Sensing Kinases and Tryptophan Metabolism.

The tryptophan catabolizing enzyme indoleamine 2,3-dioxygenase (IDO-1) has gained major attention due the immunoregulatory nature of this pathway. Both depletion of tryptophan concentrations as well as the accumulation of downstream metabolites are relevant for the mediation of the manifold consequences of increased tryptophan metabolism. Increased tryptophan catabolism is indicative for several chronic inflammatory disorders such as infections, autoimmune diseases or cancer. Low tryptophan availability is likely to be involved in the manifestation of a variety of comorbidities such as anemia, cachexia, depression and neurocognitive disturbances.Several nutrient sensing kinases are implicated in the downstream effects of dysregulated tryptophan metabolism. These include mechanisms that were conserved during evolution but have gained special features in multicellular eukaryotes, such as pathways regulated by eukaryotic translation initiation factor 2 (eIF-2)-alpha kinase (GCN2, also named general control nonderepressible 2 kinase), 5'-adenosine monophosphate (AMP)-activated protein kinase (AMPK) and target of rapamycin (TOR).The interplay between IDO-1 and above-mentioned pathway seems to be highly context dependent. A better understanding of the crosstalk is necessary to support the search for druggable targets for the treatment of inflammatory and autoimmune disorders.

Nutrient Signaling, Stress Response, and Inter-organelle Communication Are Non-canonical Determinants of Cell Fate

Isogenic cells manifest distinct cellular fates for a single stress; however, the nongenetic mechanisms driving such fates remain poorly understood. Here, we implement a robust multi-channel live-cell imaging approach to uncover noncanonical factors governing cell fate. We show that in response to acute glucose removal (AGR), budding yeast undergoes distinct fates, becoming either quiescent or senescent. Senescent cells fail to resume mitotic cycles following glucose replenishment but remain responsive to nutrient stimuli. Whereas quiescent cells manifest starvation-induced adaptation, senescent cells display perturbed endomembrane trafficking and defective nucleus-vacuole junction (NVJ) expansion. Surprisingly, senescence occurs even in the absence of lipid droplets. Importantly, we identify the nutrient-sensing kinase Rim15 as a key biomarker predicting cell fates before AGR stress. We propose that isogenic yeast challenged with acute nutrient shortage contains determinants influencing post-stress fate and demonstrate that specific nutrient signaling, stress response, trafficking, and inter-organelle biomarkers are early indicators for long-term fate outcomes.

Loss of protein kinase D activity demonstrates redundancy in cardiac glucose metabolism and preserves cardiac function in obesity

ObjectiveProtein kinase D (PKD) signaling has been implicated in stress-induced cardiac remodeling and function as well as metabolic processes including contraction-mediated cardiac glucose uptake. PKD has recently emerged as a nutrient-sensing kinase that is activated in high-lipid environments, such as in obesity. However, the role of PKD signaling in cardiac glucose metabolism and cardiac function in both normal and obese conditions remains unknown. MethodsA cardiac-specific and inducible dominant negative (DN) PKD mouse model was developed. Echocardiography was used to assess cardiac function, while metabolic phenotyping was performed, including stable isotope metabolomics on cardiac tissue in mice fed either regular chow or a high-fat diet (43% calories from fat). ResultsCardiac PKD activity declined by ∼90% following DN PKD induction in adult mice. The mice had diminished basal cardiac glucose clearance, suggesting impaired contraction-mediated glucose uptake, but normal cardiac function. In obesity studies, systolic function indices were reduced in control mice, but not in cardiac DN PKD mice. Using targeted stable isotope metabolomic analyses, no differences in glucose flux through glycolysis or the TCA cycle were observed between groups. ConclusionsThe data show that PKD contributes to cardiac dysfunction in obesity and highlight the redundancy in cardiac glucose metabolism that maintains cardiac glucose flux in vivo. The data suggest that impairments in contraction-mediated glucose uptake are unlikely to drive cardiac dysfunction in both normal and metabolic disease states.

Exposure of Pancreatic β-Cells to Excess Glucose Results in Bimodal Activation of mTORC1 and mTOR-Dependent Metabolic Acceleration.

SummaryChronic exposure of pancreatic β-cells to excess glucose can lead to metabolic acceleration and loss of stimulus-secretion coupling. Here, we examined how exposure to excess glucose (defined here as concentrations above 5 mM) affects mTORC1 signaling and the metabolism of β-cells. Acute exposure to excess glucose stimulated glycolysis-dependent mTORC1 signaling, without changes in the PI3K or AMPK pathways. Prolonged exposure to excess glucose led to hyperactivation of mTORC1 and metabolic acceleration, characterized by higher basal respiration and maximal respiratory capacity, increased energy demand, and enhanced flux through mitochondrial pyruvate metabolism. Inhibition of pyruvate transport to the mitochondria decelerated the metabolism of β-cells chronically exposed to excess glucose and re-established glucose-dependent mTORC1 signaling, disrupting a positive feedback loop for mTORC1 hyperactivation. mTOR inhibition had positive and negative impacts on various metabolic pathways and insulin secretion, demonstrating a role for mTOR signaling in the long-term metabolic adaptation of β-cells to excess glucose.

Mechanistic target of rapamycin signaling in mouse models of accelerated aging.

The mechanistic target of rapamycin (mTOR) is an essential nutrient-sensing kinase that integrates and regulates a number of fundamental cellular processes required for cell growth, cell motility, translation, metabolism, and autophagy. mTOR signaling has been implicated in the progression of many human diseases, and its dysregulation has been reported in several pathological processes, especially in age-related human diseases and mouse models of accelerated aging. In addition, many studies have demonstrated that the regulation of mTOR activity has a beneficial effect on longevity in several mouse models of aging. However, not all mouse models of accelerated aging show positive effects on aging-associated phenotypes in response to targeting mTOR signaling. Here, we review the effects of interventions that modulate mTOR signaling on aging-related phenotypes in different mouse models of accelerated aging and discuss their implications with respect to aging and aging-related disorders.

Tuberous sclerosis complex is required for tumor maintenance in MYC‐driven Burkitt's lymphoma

The tuberous sclerosis complex (TSC) 1/2 is a negative regulator of the nutrient‐sensing kinase mechanistic target of rapamycin complex (mTORC1), and its function is generally associated with tumor suppression. Nevertheless, biallelic loss of function of TSC1 or TSC2 is rarely found in malignant tumors. Here, we show that TSC1/2 is highly expressed in Burkitt's lymphoma cell lines and patient samples of human Burkitt's lymphoma, a prototypical MYC‐driven cancer. Mechanistically, we show that MYC induces TSC1 expression by transcriptional activation of the TSC1 promoter and repression of miR‐15a. TSC1 knockdown results in elevated mTORC1‐dependent mitochondrial respiration enhanced ROS production and apoptosis. Moreover, TSC1 deficiency attenuates tumor growth in a xenograft mouse model. Our study reveals a novel role for TSC1 in securing homeostasis between MYC and mTORC1 that is required for cell survival and tumor maintenance in Burkitt's lymphoma. The study identifies TSC1/2 inhibition and/or mTORC1 hyperactivation as a novel therapeutic strategy for MYC‐driven cancers.

Nutrient-sensing kinase signaling in bovine immune cells is altered during the postpartum nutrient deficit: A possible role in transition cow inflammatory response

Transition dairy cows experience a nutrient deficit, particularly in the immediate postpartum period. At the same time, the inflammatory balance is altered and cows exhibit an immune response primed for inflammatory response rather than tolerance. The mechanistic link that might be underlying the immunological effects due to the lack in nutrients is not fully understood. Studies in other species demonstrate an orchestrating role of nutrient-sensing kinases in the determination of immune phenotypes and immune cell proliferation and differentiation. Our primary objective was to investigate changes in energy storage and signaling through the protein kinase B (AKT)/mechanistic target of rapamycin complex 1 (mTOR) pathway in bovine immune cells in the transition period, as well as the association with cytokine expression profiles. A secondary objective was to test if supplementation with branched-chain amino acids alone or in combination with oral propylene glycol had any effect on the measured parameters. To assess cellular energy storage, glycogen concentration was measured by an enzymatic-fluorometric method in peripheral blood mononuclear cells (PBMC) of multiparous Holstein cows (n = 72) at 3 time points in the transition period (21 d before, 7 and 28 d after calving). At the same time points, phosphorylation of proteins in the AKT/mTOR pathway was assessed by immunoblotting in PBMC from 60 animals. Whole-blood leukocyte cytokine gene expression of IL12B, IL6, IL1B, TNF, and IL10 was measured in samples from 50 animals by reverse-transcription quantitative PCR with and without stimulation of samples with 10 ng/mL of lipopolysaccharide. Compared with glycogen concentration of prepartum PBMC, glycogen concentration decreased by 37% on d 7 postpartum. The activation of AKT/mTOR in bovine PBMC postpartum was reduced compared with prepartum values. Results of reverse-transcription quantitative PCR showed an increase in cytokine gene expression postpartum compared with prepartum values. Supplementation with branched-chain amino acids alone or in combination with oral propylene glycol did not alter glycogen storage, AKT/mTOR activity, or inflammatory balance as assessed by the measured parameters in this study. We conclude that the nutrient deficit of the immediate postpartum period is sensed by bovine immune cells, and that it affects their energy storage as well as cellular signaling pathways postpartum. Temporal associations with changes in cytokine gene expression are intriguing and warrant further investigation of the role of this pathway as a possible link between metabolism and immune phenotype postpartum.

Differential control of ageing and lifespan by isoforms and splice variants across the mTOR network.

Ageing can be defined as the gradual deterioration of physiological functions, increasing the incidence of age-related disorders and the probability of death. Therefore, the term ageing not only reflects the lifespan of an organism but also refers to progressive functional impairment and disease. The nutrient-sensing kinase mTOR (mammalian target of rapamycin) is a major determinant of ageing. mTOR promotes cell growth and controls central metabolic pathways including protein biosynthesis, autophagy and glucose and lipid homoeostasis. The concept that mTOR has a crucial role in ageing is supported by numerous reports on the lifespan-prolonging effects of the mTOR inhibitor rapamycin in invertebrate and vertebrate model organisms. Dietary restriction increases lifespan and delays ageing phenotypes as well and mTOR has been assigned a major role in this process. This may suggest a causal relationship between the lifespan of an organism and its metabolic phenotype. More than 25 years after mTOR's discovery, a wealth of metabolic and ageing-related effects have been reported. In this review, we cover the current view on the contribution of the different elements of the mTOR signalling network to lifespan and age-related metabolic impairment. We specifically focus on distinct roles of isoforms and splice variants across the mTOR network. The comprehensive analysis of mouse knockout studies targeting these variants does not support a tight correlation between lifespan prolongation and improved metabolic phenotypes and questions the strict causal relationship between them.

Role of GCN2-Independent Signaling Through a Noncanonical PERK/NRF2 Pathway in the Physiological Responses to Dietary Methionine Restriction.

Restricting availability of essential amino acids (EAAs) limits aminoacylation of tRNAs by their cognate EAAs and activates the nutrient-sensing kinase, general control nonderepressible 2 (GCN2). Activated GCN2 phosphorylates eukaryotic initiation factor 2 (eIF2), altering gene-specific translation and initiating a transcriptional program collectively described as the integrated stress response (ISR). Central GCN2 activation by EAA deprivation is also linked to an acute aversive feeding response. Dietary methionine restriction (MR) produces a well-documented series of physiological responses (increased energy intake and expenditure, decreased adiposity, and increased insulin sensitivity), but the role of GCN2 in mediating them is unknown. Using Gcn2−/− mice, we found that the absence of GCN2 had no effect on the ability of MR to reduce body weight or adiposity, increase energy intake and expenditure, increase hepatic transcription and release of fibroblast growth factor 21, or improve insulin sensitivity. Interestingly, hepatic eIF2 phosphorylation by MR was uncompromised in Gcn2−/− mice. Instead, protein kinase R–like endoplasmic reticulum (ER) kinase (PERK) was activated in both intact and Gcn2−/− mice. PERK activation corresponded with induction of the ISR and the nuclear respiratory factor 2 antioxidant program but not ER stress. These data uncover a novel glutathione-sensing mechanism that functions independently of GCN2 to link dietary MR to its metabolic phenotype.

Nutrient-regulated Phosphorylation of ATG13 Inhibits Starvation-induced Autophagy

Autophagy is a conserved catabolic process that utilizes a defined series of membrane trafficking events to generate a de novo double-membrane vesicle termed the autophagosome, which matures by fusing to the lysosome. Subsequently, the lysosome facilitates the degradation and recycling of the cytoplasmic cargo. In yeast, the upstream signals that regulate the induction of starvation-induced autophagy are clearly defined. The nutrient-sensing kinase Tor inhibits the activation of autophagy by regulating the formation of the Atg1-Atg13-Atg17 complex, through hyperphosphorylation of Atg13. However, in mammals, the ortholog complex ULK1-ATG13-FIP200 is constitutively formed. As such, the molecular mechanism by which mTOR regulates mammalian autophagy is unknown. Here we report the identification and characterization of novel nutrient-regulated phosphorylation sites on ATG13: Ser-224 and Ser-258. mTOR directly phosphorylates ATG13 on Ser-258 while Ser-224 is modulated by the AMPK pathway. In ATG13 knock-out cells reconstituted with an unphosphorylatable mutant of ATG13, ULK1 kinase activity is more potent, and amino acid starvation induced more rapid ATG13 and ULK1 translocation. These events culminated in a more rapid starvation-induced autophagy response. Therefore, ATG13 phosphorylation plays a crucial role in autophagy regulation.

Cross-Talk between Carbon Metabolism and the DNA Damage Response in S. cerevisiae.

Yeast cells with DNA damage avoid respiration, presumably because products of oxidative metabolism can be harmful to DNA. We show that DNA damage inhibits the activity of the Snf1 (AMP-activated) protein kinase (AMPK), which activates expression of genes required for respiration. Glucose and DNA damage upregulate SUMOylation of Snf1, catalyzed by the SUMO E3 ligase Mms21, which inhibits SNF1 activity. The DNA damage checkpoint kinases Mec1/ATR and Tel1/ATM, as well as the nutrient-sensing protein kinase A (PKA), regulate Mms21 activity toward Snf1. Mec1 and Tel1 are required for two SNF1-regulated processes-glucose sensing and ADH2 gene expression-even without exogenous genotoxic stress. Our results imply that inhibition of Snf1 by SUMOylation is a mechanism by which cells lower their respiration in response to DNA damage. This raises the possibility that activation of DNA damage checkpoint mechanisms could contribute to aerobic fermentation (Warburg effect), a hallmark of cancer cells.

MTOR activates the VPS34-UVRAG complex to regulate autolysosomal tubulation and cell survival.

Lysosomes are essential organelles that function to degrade and recycle unwanted, damaged and toxic biological components. Lysosomes also act as signalling platforms in activating the nutrient-sensing kinase mTOR. mTOR regulates cellular growth, but it also helps to maintain lysosome identity by initiating lysosomal tubulation through a process termed autophagosome-lysosome reformation (ALR). Here we identify a lysosomal pool of phosphatidylinositol 3-phosphate that, when depleted by specific inhibition of the class III phosphoinositide 3-kinase VPS34, results in prolonged lysosomal tubulation. This tubulation requires mTOR activity, and we identified two direct mTOR phosphorylation sites on UVRAG (S550 and S571) that activate VPS34. Loss of these phosphorylation sites reduced VPS34 lipid kinase activity and resulted in an increase in number and length of lysosomal tubules. In cells in which phosphorylation at these UVRAG sites is disrupted, the result of impaired lysosomal tubulation alongside ALR activation is massive cell death. Our data imply that ALR is critical for cell survival under nutrient stress and that VPS34 is an essential regulatory element in this process.

Epidermal growth factor receptor signaling and the progression of diabetic nephropathy.

Epidermal growth factor receptor signaling and the progression of diabetic nephropathy.