bims-auttor Biomed News
on Autophagy and mTOR
Issue of 2020–09–06
28 papers selected by
Viktor Korolchuk, Newcastle University
  1. Reconstitution of autophagosome nucleation defines Atg9 vesicles as seeds for membrane formation.
  2. The Roles of Ubiquitin in Mediating Autophagy.
  3. Epr1, a UPR-upregulated soluble autophagy receptor for reticulophagy.
  4. PDPK1 regulates autophagosome biogenesis by binding to PIK3C3.
  5. Autophagy under glucose starvation enhances protein translation initiation in response to re-addition of glucose in C2C12 myotubes.
  6. Amino acid starvation inhibits autophagy in lipid droplet-deficient cells through mitochondrial dysfunction.
  7. Ubiquitin, Autophagy and Neurodegenerative Diseases.
  8. Structural mechanism for amino acid-dependent Rag GTPase nucleotide state switching by SLC38A9.
  9. Keratinocyte autophagy enables the activation of keratinocytes and fibroblasts and facilitates wound healing.
  10. Beclin 2 negatively regulates innate immune signaling and tumor development.
  11. Decanoic acid inhibits mTORC1 activity independent of glucose and insulin signaling.
  12. Assays for Autophagy III: Observing Dynamic Protein Trafficking.
  13. Lysosome as a Central Hub for Rewiring PH Homeostasis in Tumors.
  14. Inhibition of proteasome reveals basal mitochondrial ubiquitination.
  15. The effect of trehalose on autophagy-related proteins and cyst growth in a hypomorphic Pkd1 mouse model of autosomal dominant polycystic kidney disease.
  16. Insights into Disease-Associated Tau Impact on Mitochondria.
  17. SH003 activates autophagic cell death by activating ATF4 and inhibiting G9a under hypoxia in gastric cancer cells.
  18. mTOR signaling regulates the morphology and migration of outer radial glia in developing human cortex.
  19. The DNA methyltransferase DNMT3A contributes to autophagy long-term memory.
  20. The regulation of mitochondrial homeostasis by the Ubiquitin Proteasome System.
  21. Antioxidant Modulation of mTOR and Sirtuin Pathways in Age-Related Neurodegenerative Diseases.
  22. Atg8-Family Proteins-Structural Features and Molecular Interactions in Autophagy and Beyond.
  23. The primary cilium and lipophagy translate mechanical forces to direct metabolic adaptation of kidney epithelial cells.
  24. The impact of autophagy during the development and survival of glioblastoma.
  25. Mechanistic Target of Rapamycin (mTOR) Signaling Activation Antagonizes Autophagy to Facilitate Zika Virus Replication.
  26. Autophagy-Induced Drug Resistance in Liver Cancer.
  27. Targeting Oxidative Phosphorylation Reverses Drug Resistance in Cancer Cells by Blocking Autophagy Recycling.
  28. A new mechanism for autophagy regulation of anti-tumor immune responses.
  1. Science. 2020 Sep 04. pii: eaaz7714. [Epub ahead of print]369(6508):
    Reconstitution of autophagosome nucleation defines Atg9 vesicles as seeds for membrane formation.
    Justyna Sawa-Makarska, Verena Baumann, Nicolas Coudevylle, Sören von Bülow, Veronika Nogellova, Christine Abert, Martina Schuschnig, Martin Graef, Gerhard Hummer, Sascha Martens.
      Autophagosomes form de novo in a manner that is incompletely understood. Particularly enigmatic are autophagy-related protein 9 (Atg9)-containing vesicles that are required for autophagy machinery assembly but do not supply the bulk of the autophagosomal membrane. In this study, we reconstituted autophagosome nucleation using recombinant components from yeast. We found that Atg9 proteoliposomes first recruited the phosphatidylinositol 3-phosphate kinase complex, followed by Atg21, the Atg2-Atg18 lipid transfer complex, and the E3-like Atg12-Atg5-Atg16 complex, which promoted Atg8 lipidation. Furthermore, we found that Atg2 could transfer lipids for Atg8 lipidation. In selective autophagy, these reactions could potentially be coupled to the cargo via the Atg19-Atg11-Atg9 interactions. We thus propose that Atg9 vesicles form seeds that establish membrane contact sites to initiate lipid transfer from compartments such as the endoplasmic reticulum.
    DOI:  https://doi.org/10.1126/science.aaz7714
  2. Cells. 2020 Sep 02. pii: E2025. [Epub ahead of print]9(9):
    The Roles of Ubiquitin in Mediating Autophagy.
    Zhangyuan Yin, Hana Popelka, Yuchen Lei, Ying Yang, Daniel J Klionsky.
      Ubiquitination, the post-translational modification essential for various intracellular processes, is implicated in multiple aspects of autophagy, the major lysosome/vacuole-dependent degradation pathway. The autophagy machinery adopted the structural architecture of ubiquitin and employs two ubiquitin-like protein conjugation systems for autophagosome biogenesis. Ubiquitin chains that are attached as labels to protein aggregates or subcellular organelles confer selectivity, allowing autophagy receptors to simultaneously bind ubiquitinated cargos and autophagy-specific ubiquitin-like modifiers (Atg8-family proteins). Moreover, there is tremendous crosstalk between autophagy and the ubiquitin-proteasome system. Ubiquitination of autophagy-related proteins or regulatory components plays significant roles in the precise control of the autophagy pathway. In this review, we summarize and discuss the molecular mechanisms and functions of ubiquitin and ubiquitination, in the process and regulation of autophagy.
    Keywords:  autophagy; lysosome; selective autophagy; ubiquitin; ubiquitination
    DOI:  https://doi.org/10.3390/cells9092025
  3. Autophagy. 2020 Aug 31.
    Epr1, a UPR-upregulated soluble autophagy receptor for reticulophagy.
    Dan Zhao, Li-Lin Du.
      The endoplasmic reticulum (ER) is a major site of protein folding. Perturbations in the folding capacity of the ER result in ER stress. ER stress triggers autophagic degradation of the ER (reticulophagy). Molecular mechanisms underlying ER stress-induced reticulophagy remain largely unknown. Our recent study identified a soluble protein, Epr1, as an autophagy receptor for ER stress-induced reticulophagy in the fission yeast Schizosaccharomyces pombe. Epr1 can interact simultaneously with Atg8 and a VAP family integral ER membrane protein, and thereby act as a bridging molecule between them. VAP family proteins contribute to reticulophagy by not only connecting Atg8 to the ER membrane through Epr1, but also by supporting the ER-plasma membrane contact. The expression of Epr1 is upregulated during ER stress in a manner dependent on the unfolded protein response (UPR) regulator Ire1. Ire1 promotes reticulophagy by upregulating Epr1.
    Keywords:  ER stress; ER-phagy; ER-plasma membrane contact; Ire1; VAP; reticulophagy; selective autophagy; unfolded protein response (UPR)
    DOI:  https://doi.org/10.1080/15548627.2020.1816665
  4. Autophagy. 2020 Sep 02.
    PDPK1 regulates autophagosome biogenesis by binding to PIK3C3.
    Boli Hu, Yina Zhang, Tingjuan Deng, Jinyan Gu, Juan Liu, Hui Yang, Yuting Xu, Yan Yan, Fan Yang, Heng Zhang, Yulan Jin, Jiyong Zhou.
      PDPK1 (3-phosphoinositide dependent protein kinase 1) is a phosphorylation-regulated kinase that plays a central role in activating multiple signaling pathways and cellular processes. Here, this study shows that PDPK1 turns on macroautophagy/autophagy as a SUMOylation-regulated kinase. In vivo data demonstrate that the SUMO modification of PDPK1 is a physiological feature in the brain and that it can be induced by viral infections. The SUMOylated PDPK1 regulates its own phosphorylation and subsequent activation of the AKT1 (AKT serine/threonine kinase 1)-MTOR (mechanistic target of rapamycin kinase) pathway. However, SUMOylation of PDPK1 is inhibited by binding to PIK3C3 (phosphatidylinositol 3-kinase catalytic subunit type 3). The nonSUMOylated PDPK1 then tethers LC3 to the endoplasmic reticulum to initiate autophagy, and it acts as a key component in forming the autophagic vacuole. Collectively, this study reveals the intricate molecular regulation of PDPK1 by post-translational modification in controlling autophagosome biogenesis, and it highlights the role of PDPK1 as a sensor of cellular stress and regulator of autophagosome biogenesis.
    Keywords:  AKT1-MTOR; PDPK1; PIK3C3; SUMOylation; autophagy
    DOI:  https://doi.org/10.1080/15548627.2020.1817279
  5. FEBS Open Bio. 2020 Sep 03.
    Autophagy under glucose starvation enhances protein translation initiation in response to re-addition of glucose in C2C12 myotubes.
    Naoya Nakai, Saki Kitai, Noriko Iida, Sachika Inoue, Kazuhiko Higashida.
      Proteolysis is known to play a crucial role in maintaining skeletal muscle mass and function. Autophagy is a conserved intracellular process for the bulk degradation of proteins in lysosomes. Although nutrient starvation is known to induce autophagy, the effect of nutrient repletion following starvation on the mTOR pathway-mediated protein translation remains unclear. In the present study, we examined the effect of glucose starvation on the initiation of protein translation in response to glucose re-addition in C2C12 myotubes. Glucose starvation decreased the phosphorylation of p70 S6 kinase (p70S6K), a bonafide marker for protein translation initiation. Following re-addition of glucose, phosphorylation of p70S6K markedly increased only in glucose-starved cells. Inhibiting autophagy using pharmacological inhibitors diminished the effect of glucose re-addition on the phosphorylation of p70S6K, whereas inhibition of the ubiquitin-proteasome system did not exert any effect. In conclusion, autophagy under glucose starvation partially accounts for the activation of translation initiation by re-addition of glucose.
    Keywords:  autophagy; glucose starvation; mTORC1; p70 S6 kinase; protein synthesis
    DOI:  https://doi.org/10.1002/2211-5463.12970
  6. Biochem J. 2020 Sep 04. pii: BCJ20200551. [Epub ahead of print]
    Amino acid starvation inhibits autophagy in lipid droplet-deficient cells through mitochondrial dysfunction.
    Pierre Voisin, Marianne Bernard, Thierry Bergès, Matthieu Regnacq.
      Lipid droplets are ubiquitous organelles in eukaryotes that act as storage sites for neutral lipids. Under normal growth conditions they are not required in the yeast Saccharomyces cerevisiae. However, recent works have shown that lipid droplets are required for autophagy to proceed in response to nitrogen starvation and that they play an essential role in maintaining ER homeostasis. Autophagy is a major catabolic pathway that helps degradation and recycling of potentially harmful proteins and organelles. It can be pharmacologically induced by rapamycin even in the absence of lipid droplets. Here, we show that amino acid starvation is responsible for autophagy failure in lipid droplet-deficient yeast.  It not only fails to induce autophagy but also inhibits rapamycin-induced autophagy. The general amino acid control pathway is not involved in this paradoxical effect of amino acid shortage. We correlate the autophagy failure with mitochondria aggregation and we show that amino acid starvation-induced autophagy is restored in lipid droplet-deficient yeast by increasing mitochondrial biomass physiologically (respiration) or genetically (REG1 deletion). Our results establish a new functional link between lipid droplets, ER and mitochondria during nitrogen starvation-induced autophagy.
    Keywords:  Autophagy; Saccharomyces cerevisiae; catabolite repression; lipid droplets; mitochondrial dysfunction
    DOI:  https://doi.org/10.1042/BCJ20200551
  7. Cells. 2020 Sep 02. pii: E2022. [Epub ahead of print]9(9):
    Ubiquitin, Autophagy and Neurodegenerative Diseases.
    Yoshihisa Watanabe, Katsutoshi Taguchi, Masaki Tanaka.
      Ubiquitin signals play various roles in proteolytic and non-proteolytic functions. Ubiquitin signals are recognized as targets of the ubiquitin-proteasome system and the autophagy-lysosome pathway. In autophagy, ubiquitin signals are required for selective incorporation of cargoes, such as proteins, organelles, and microbial invaders, into autophagosomes. Autophagy receptors possessing an LC3-binding domain and a ubiquitin binding domain are involved in this process. Autophagy activity can decline as a result of genetic variation, aging, or lifestyle, resulting in the onset of various neurodegenerative diseases. This review summarizes the selective autophagy of neurodegenerative disease-associated protein aggregates via autophagy receptors and discusses its therapeutic application for neurodegenerative diseases.
    Keywords:  autophagy; autophagy–lysosome pathway; neurodegenerative diseases; ubiquitin; ubiquitin–proteasome system
    DOI:  https://doi.org/10.3390/cells9092022
  8. Nat Struct Mol Biol. 2020 Aug 31.
    Structural mechanism for amino acid-dependent Rag GTPase nucleotide state switching by SLC38A9.
    Simon A Fromm, Rosalie E Lawrence, James H Hurley.
      The Rag GTPases (Rags) recruit mTORC1 to the lysosomal membrane in response to nutrients, where it is then activated in response to energy and growth factor availability. The lysosomal folliculin (FLCN) complex (LFC) consists of the inactive Rag dimer, the pentameric scaffold Ragulator, and the FLCN:FNIP2 (FLCN-interacting protein 2) GTPase activating protein (GAP) complex, and prevents Rag dimer activation during amino acid starvation. How the LFC is disassembled upon amino acid refeeding is an outstanding question. Here we show that the cytoplasmic tail of the human lysosomal solute carrier family 38 member 9 (SLC38A9) destabilizes the LFC and thereby triggers GAP activity of FLCN:FNIP2 toward RagC. We present the cryo-EM structures of Rags in complex with their lysosomal anchor complex Ragulator and the cytoplasmic tail of SLC38A9 in the pre- and post-GTP hydrolysis state of RagC, which explain how SLC38A9 destabilizes the LFC and so promotes Rag dimer activation.
    DOI:  https://doi.org/10.1038/s41594-020-0490-9
  9. Autophagy. 2020 Aug 31.
    Keratinocyte autophagy enables the activation of keratinocytes and fibroblasts and facilitates wound healing.
    Lei Qiang, Seungwon Yang, Yan-Hong Cui, Yu-Ying He.
      Macroautophagy/autophagy is a cellular catabolic process that is implicated in several physiological and pathological processes. However, the role of epidermal autophagy in wound healing remains unknown. Here, using mice with genetic ablation of the essential Atg5 (autophagy related 5) or Atg7 in their epidermis to inhibit autophagy, we show that keratinocyte autophagy regulates wound healing in mice. Wounding induces the expression of autophagy genes in mouse skin. Epidermis-specific autophagy deficiency inhibits wound closure, re-epithelialization, keratinocyte proliferation and differentiation, dermal granulation tissue formation, and infiltration of immune cells including macrophages, neutrophils, and mast cells, while it does not affect angiogenesis. Using cytokine array screening, we found that autophagy deficiency inhibits the transcription and production of the cytokine CCL2/MCP-1 by TNF. At the molecular level, TNF induces autophagic flux and the expression of autophagy genes through NFKB in epidermal keratinocytes. TNF promotes CCL2 transcription through the autophagy-AMPK-BRAF-MAPK/ERK-activator protein 1 (AP-1) pathway. Indeed, treating mice with recombinant CCL2 can reverse the effect of autophagy deficiency in keratinocytes. At the cellular level, we found that CCL2 induction via autophagy in keratinocytes is required not only for keratinocyte migration and proliferation but also for dermal fibroblast activation. Our findings demonstrate a critical role of epidermal autophagy in wound healing in vivo and elucidate a critical molecular machinery coordinating keratinocyte-fibroblast interaction in skin repair.
    Keywords:  CCL2/MCP-1; TNF; autophagy; differentiation; fibroblast; inflammation; keratinocyte; migration; proliferation; wound healing
    DOI:  https://doi.org/10.1080/15548627.2020.1816342
  10. J Clin Invest. 2020 Aug 31. pii: 133283. [Epub ahead of print]
    Beclin 2 negatively regulates innate immune signaling and tumor development.
    Motao Zhu, Guangtong Deng, Peng Tan, Changsheng Xing, Cuiping Guan, Chongming Jiang, Yinlong Zhang, Bo Ning, Chaoran Li, Bingnan Yin, Kaifu Chen, Yuliang Zhao, Helen Y Wang, Beth Levine, Guangjun Nie, Rong-Fu Wang.
      Beclin 2 plays a critical role in metabolic regulation and obesity, but its functions in innate immune signaling and cancer development remain largely unknown. Here, we identified Beclin 2 as a critical negative regulator of inflammation and lymphoma development. Mice with homozygous ablation of BCL2-interacting protein 2 (Becn2) developed splenomegaly and lymphadenopathy and markedly increased ERK1/2 and NF-κB signaling for proinflammatory cytokine production. Beclin 2 targeted the key signaling kinases MEKK3 and TAK1 for degradation through an ATG9A-dependent, but ATG16L/Beclin 1/LC3-independent, autophagic pathway. Mechanistically, Beclin 2 recruited MEKK3 or TAK1 through ATG9A to form a complex (Beclin 2-ATG9A-MEKK3) on ATG9A+ vesicles upon ULK1 activation. Beclin 2 further interacted with STX5 and STX6 to promote the fusion of MEKK3- or TAK1-associated ATG9A+ vesicles to phagophores for subsequent degradation. Importantly, Becn2-deficient mice had a markedly increased incidence of lymphoma development, with persistent STAT3 activation. Myeloid-specific ablation of MEKK3 (Map3k3) completely rescued the phenotypes (splenomegaly, higher amounts of proinflammatory cytokines, and cancer incidence) of Becn2-deficient mice. Hence, our findings have identified an important role of Beclin 2 in the negative regulation of innate immune signaling and tumor development through an ATG9A-dependent, but ATG16L/Beclin 1/LC3-independent, autophagic pathway, thus providing a potential target for the treatment of inflammatory diseases and cancer.
    Keywords:  Autophagy; Cell Biology; Inflammation; Innate immunity
    DOI:  https://doi.org/10.1172/JCI133283
  11. Proc Natl Acad Sci U S A. 2020 Sep 02. pii: 202008980. [Epub ahead of print]
    Decanoic acid inhibits mTORC1 activity independent of glucose and insulin signaling.
    Eleanor C Warren, Stephanie Dooves, Eleonora Lugarà, Joseph Damstra-Oddy, Judith Schaf, Vivi M Heine, Mathew C Walker, Robin S B Williams.
      Low-glucose and -insulin conditions, associated with ketogenic diets, can reduce the activity of the mechanistic target of rapamycin complex 1 (mTORC1) signaling pathway, potentially leading to a range of positive medical and health-related effects. Here, we determined whether mTORC1 signaling is also a target for decanoic acid, a key component of the medium-chain triglyceride (MCT) ketogenic diet. Using a tractable model system, Dictyostelium, we show that decanoic acid can decrease mTORC1 activity, under conditions of constant glucose and in the absence of insulin, measured by phosphorylation of eukaryotic translation initiation factor 4E-binding protein 1 (4E-BP1). We determine that this effect of decanoic acid is dependent on a ubiquitin regulatory X domain-containing protein, mediating inhibition of a conserved Dictyostelium AAA ATPase, p97, a homolog of the human transitional endoplasmic reticulum ATPase (VCP/p97) protein. We then demonstrate that decanoic acid decreases mTORC1 activity in the absence of insulin and under high-glucose conditions in ex vivo rat hippocampus and in tuberous sclerosis complex (TSC) patient-derived astrocytes. Our data therefore indicate that dietary decanoic acid may provide a new therapeutic approach to down-regulate mTORC1 signaling.
    Keywords:  Dictyostelium discoideum; decanoic acid; epilepsy; mTOR; tuberous sclerosis complex
    DOI:  https://doi.org/10.1073/pnas.2008980117
  12. Methods Mol Biol. 2021 ;2196 211-222
    Assays for Autophagy III: Observing Dynamic Protein Trafficking.
    Jing Zhu, Zhiping Xie.
      Macroautophagy, by its very nature, is a protein trafficking process. Cargos are transported and processed. Atg proteins come and go. In this chapter, we present three assays to monitor these dynamic events: a non-radioactive pulse-chase labeling assay to monitor the transport of prApe1 and two fluorescent microscopy-based assays to assess the trafficking of Atg8 and Atg9.
    Keywords:  Ape1; Atg8; Atg9; Autophagy; Cvt pathway; Fluorescent microscopy; Intracellular trafficking; Pulse-chase; Yeast
    DOI:  https://doi.org/10.1007/978-1-0716-0868-5_16
  13. Cancers (Basel). 2020 Aug 27. pii: E2437. [Epub ahead of print]12(9):
    Lysosome as a Central Hub for Rewiring PH Homeostasis in Tumors.
    Ran Chen, Marja Jäättelä, Bin Liu.
      Cancer cells generate large quantities of cytoplasmic protons as byproducts of aberrantly activated aerobic glycolysis and lactate fermentation. To avoid potentially detrimental acidification of the intracellular milieu, cancer cells activate multiple acid-removal pathways that promote cytosolic alkalization and extracellular acidification. Accumulating evidence suggests that in addition to the well-characterized ion pumps and exchangers in the plasma membrane, cancer cell lysosomes are also reprogrammed for this purpose. On the one hand, the increased expression and activity of the vacuolar-type H+-ATPase (V-ATPase) on the lysosomal limiting membrane combined with the larger volume of the lysosomal compartment increases the lysosomal proton storage capacity substantially. On the other hand, enhanced lysosome exocytosis enables the efficient release of lysosomal protons to the extracellular space. Together, these two steps dynamically drive proton flow from the cytosol to extracellular space. In this perspective, we provide mechanistic insight into how lysosomes contribute to the rewiring of pH homeostasis in cancer cells.
    Keywords:  V-ATPase; lysosomal exocytosis; lysosome; pH regulation
    DOI:  https://doi.org/10.3390/cancers12092437
  14. J Proteomics. 2020 Aug 31. pii: S1874-3919(20)30317-1. [Epub ahead of print] 103949
    Inhibition of proteasome reveals basal mitochondrial ubiquitination.
    Prasad Sulkshane, Inbar Duek, Jonathan Ram, Anita Thakur, Noa Reis, Tamar Ziv, Michael H Glickman.
      Strict quality control for mitochondrial proteins is necessary to ensure cell homeostasis. Two cellular pathways-Ubiquitin Proteasome System (UPS) and autophagy-contribute to mitochondrial homeostasis under stressful conditions. Here, we investigate changes to the mitochondria proteome and to the ubiquitin landscape at mitochondria in response to proteasome inhibition. Treatment of HeLa cells devoid of Parkin, the primary E3 ligase responsible for mitophagy, with proteasome inhibitor MG132 for a few hours caused mitochondrial oxidative stress and fragmentation, reduced energy output, and increased mitochondrial ubiquitination without inducing mitophagy. Overexpression of Parkin did not show any induction of mitophagy in response to MG132 treatment. Analysis of ubiquitin chains on isolated mitochondria revealed predominance of K48, K29 and K63-linked polyubiquitin. Interestingly, of all ubiquitinated mitochondrial proteins detected in response to MG132 treatment, a majority (≥90%) were intramitochondrial irrespective of Parkin expression. However, overall levels of these ubiquitinated mitochondrial proteins did not change significantly upon proteasome inhibition when evaluated by quantitative proteomics (LFQ and SILAC), suggesting that only a small portion are ubiquitinated under basal conditions. Another aspect of proteasome inhibition is significant enrichment of UPS, lysosomal and phagosomal components, and other heat shock proteins associated with isolated mitochondria. Taken together, our study highlights a critical role of UPS for ubiquitinating and removing imported proteins as part of a basal mitochondrial quality control system independent of Parkin. SIGNIFICANCE: As centers of cellular bioenergetics, numerous metabolic pathways and signaling cascades, the health of mitochondria is of utmost importance for ensuring cell survival. Due to their unique physiology, mitochondria are constantly subjected to damaging oxidative radicals (ROS) and protein import-related stress due to buildup of unfolded aggregate-prone proteins. Thus, for quality control purposes, mitochondria are constantly under surveillance by Autophagy and the Ubiquitin Proteasome System (UPS), both of which share ubiquitin as a common signal. The ubiquitin landscape of mitochondria has been studied in detail under stressful conditions, however, little is known about basal mitochondrial ubiquitination. Our study reveals that the extent of ubiquitination at mitochondria greatly increases upon proteasome inhibition, pointing to a large number of potential substrates for proteasomal degradation. Interestingly, most of the ubiquitination occurs on intramitochondrial proteins, components of the electron transport chain (ETC) and matrix-resident metabolic enzymes in particular. Moreover, numerous cytosolic UPS components, chaperones and autophagy-lysosomal proteins were recruited to mitochondria upon proteasome inhibition. Taken together, this suggests that the levels and functions of mitochondrial proteins are constantly regulated through ubiquitin-dependent proteasomal degradation even under basal conditions. Unclogging mitochondrial import channels may provide a mechanism to alleviate stress associated with mitochondrial protein import or to adapt cells according to their metabolic needs. Therefore, targeting the mitochondrial ubiquitination/deubiquitination machinery, such as improving the therapeutic potency of proteasome inhibitors, may provide an additional therapeutic arsenal against tumors.
    Keywords:  Mitochondria; Mitostasis; Proteasome; Quantitative proteomics; Ubiquitin
    DOI:  https://doi.org/10.1016/j.jprot.2020.103949
  15. Cell Signal. 2020 Aug 28. pii: S0898-6568(20)30237-0. [Epub ahead of print] 109760
    The effect of trehalose on autophagy-related proteins and cyst growth in a hypomorphic Pkd1 mouse model of autosomal dominant polycystic kidney disease.
    Daniel J Atwood, Carolyn N Brown, Sara J Holditch, Deepak Pokhrel, Andrew Thorburn, Katharina Hopp, Charles L Edelstein.
      Autosomal dominant polycystic kidney disease (ADPKD) is a common inherited disorder characterized by kidney cyst growth often resulting in end-stage renal disease. There is growing attention on understanding the role of impaired autophagy in ADPKD. Trehalose (TRE) has been shown to increase both protein stability and aggregate clearance and induce autophagy in neurodegenerative diseases. TRE treatment in wild type mice compared to vehicle resulted in increased expression in the kidney of Atg12-5 complex and increased Rab9a, autophagy-related proteins that play a role in the formation of autophagosomes. Thus, the aim of the study was to determine the effect of TRE on cyst growth and autophagy-related proteins, in the hypomorphic Pkd1RC/RC mouse model of ADPKD. Pkd1RC/RC mice were treated 2% TRE in water from days 50 to 120 of age. TRE did not slow cyst growth or improve kidney function or affect proliferation and apoptosis in Pkd1RC/RC kidneys. In Pkd1RC/RC vs. wild type kidneys, expression of the Atg12-5 complex was inhibited by TRE resulting in increased free Atg12 and TRE was unable to rescue the deficiency of the Atg12-5 complex. Rab9a was decreased in Pkd1RC/RC vs. wild type kidneys and unaffected by TRE. The TRE-induced increase in p62, a marker of autophagic cargo, that was seen in normal kidneys was blocked in Pkd1RC/RC kidneys. In summary, the autophagy phenotype in Pkd1RC/RC kidneys was characterized by decreases in crucial autophagy-related proteins (Atg12-5 complex, Atg5, Atg16L1), decreased Rab9a and increased mTORC1 (pS6, p-mTORS2448) proteins. TRE increased Atg12-5 complex, Rab9a and p62 in normal kidneys, but was unable to rescue the deficiency in autophagy proteins or suppress mTORC1 in Pkd1RC/RC kidneys and did not protect against cyst growth.
    Keywords:  ADPKD; Apoptosis; Atg12; Atg5; PKD; Polycystic; Proliferation; Rab9a; TRE
    DOI:  https://doi.org/10.1016/j.cellsig.2020.109760
  16. Int J Mol Sci. 2020 Sep 01. pii: E6344. [Epub ahead of print]21(17):
    Insights into Disease-Associated Tau Impact on Mitochondria.
    Leonora Szabo, Anne Eckert, Amandine Grimm.
      Abnormal tau protein aggregation in the brain is a hallmark of tauopathies, such as frontotemporal lobar degeneration and Alzheimer's disease. Substantial evidence has been linking tau to neurodegeneration, but the underlying mechanisms have yet to be clearly identified. Mitochondria are paramount organelles in neurons, as they provide the main source of energy (adenosine triphosphate) to these highly energetic cells. Mitochondrial dysfunction was identified as an early event of neurodegenerative diseases occurring even before the cognitive deficits. Tau protein was shown to interact with mitochondrial proteins and to impair mitochondrial bioenergetics and dynamics, leading to neurotoxicity. In this review, we discuss in detail the different impacts of disease-associated tau protein on mitochondrial functions, including mitochondrial transport, network dynamics, mitophagy and bioenergetics. We also give new insights about the effects of abnormal tau protein on mitochondrial neurosteroidogenesis, as well as on the endoplasmic reticulum-mitochondria coupling. A better understanding of the pathomechanisms of abnormal tau-induced mitochondrial failure may help to identify new targets for therapeutic interventions.
    Keywords:  mitochondria; tau protein; tauopathies
    DOI:  https://doi.org/10.3390/ijms21176344
  17. Cell Death Dis. 2020 Sep 02. 11(8): 717
    SH003 activates autophagic cell death by activating ATF4 and inhibiting G9a under hypoxia in gastric cancer cells.
    Tae Woo Kim, Chunhoo Cheon, Seong-Gyu Ko.
      In gastric cancer (GC), hypoxia is one of the greatest obstacles to cancer therapy. In this present study, we report that SH003, an herbal formulation, induces ER stress via PERK-ATF4-CHOP signaling in GC. SH003-mediated ER stress inhibits G9a, a histone methyltransferase, by reducing STAT3 phosphorylation and activates autophagy, indicating to the dissociation of Beclin-1 and autophagy initiation from Bcl-2/Beclin-1 complex. However, the inhibition of PERK and CHOP inhibited SH003-induced cell death and autophagy activation. Moreover, targeting autophagy using specific siRNAs of LC3B or p62 or the autophagy inhibitor 3-MA also inhibited SH003-induced cell death in GC. Interestingly, SH003 induces BNIP3-mediated autophagic cell death under hypoxia than normoxia in GC. These findings reveal that SH003-induced ER stress regulates BNIP3-induced autophagic cell death via inhibition of STAT3-G9a axis under hypoxia in GC. Therefore, SH003 may an important tumor therapeutic strategy under hypoxia-mediated chemo-resistance.
    DOI:  https://doi.org/10.1038/s41419-020-02924-w
  18. Elife. 2020 Sep 02. pii: e58737. [Epub ahead of print]9
    mTOR signaling regulates the morphology and migration of outer radial glia in developing human cortex.
    Madeline G Andrews, Lakshmi Subramanian, Arnold R Kriegstein.
      Outer radial glial (oRG) cells are a population of neural stem cells prevalent in the developing human cortex that contribute to its cellular diversity and evolutionary expansion. The mammalian Target of Rapamycin (mTOR) signaling pathway is active in human oRG cells. Mutations in mTOR pathway genes are linked to a variety of neurodevelopmental disorders and malformations of cortical development. We find that dysregulation of mTOR signaling specifically affects oRG cells, but not other progenitor types, by changing the actin cytoskeleton through the activity of the Rho-GTPase, CDC42. These effects change oRG cellular morphology, migration, and mitotic behavior, but do not affect proliferation or cell fate. Thus, mTOR signaling can regulate the architecture of the developing human cortex by maintaining the cytoskeletal organization of oRG cells and the radial glia scaffold. Our study provides insight into how mTOR dysregulation may contribute to neurodevelopmental disease.
    Keywords:  human; human cortex; neuroscience; organoids; outer radial glia; regenerative medicine; stem cells
    DOI:  https://doi.org/10.7554/eLife.58737
  19. Autophagy. 2020 Sep 02.
    The DNA methyltransferase DNMT3A contributes to autophagy long-term memory.
    Patricia González-Rodríguez, Mathilde Cheray, Jens Füllgrabe, Maria Salli, Pinelopi Engskog-Vlachos, Lily Keane, Virginia Cunha, Agata Lupa, Wenbo Li, Qi Ma, Kristian Dreij, Michael G Rosenfeld, Bertrand Joseph.
      Macroautophagy/autophagy is a conserved catabolic pathway that targets cytoplasmic components for their degradation and recycling in an autophagosome-dependent lysosomal manner. Under physiological conditions, this process maintains cellular homeostasis. However, autophagy can be stimulated upon different forms of cellular stress, ranging from nutrient starvation to exposure to drugs. Thus, this pathway can be seen as a central component of the integrated and adaptive stress response. Here, we report that even brief induction of autophagy is coupled in vitro to a persistent downregulation of the expression of MAP1LC3 isoforms, which are key components of the autophagy core machinery. In fact, DNA-methylation mediated by de novo DNA methyltransferase DNMT3A of MAP1LC3 loci upon autophagy stimulation leads to the observed long-term decrease of MAP1LC3 isoforms at transcriptional level. Finally, we report that the downregulation of MAP1LC3 expression can be observed in vivo in zebrafish larvae and mice exposed to a transient autophagy stimulus. This epigenetic memory of autophagy provides some understanding of the long-term effect of autophagy induction and offers a possible mechanism for its decline upon aging, pathological conditions, or in response to treatment interventions.
    Keywords:  Autophagy; DNA methylation; MAP1LC3; epigenetics; transcription
    DOI:  https://doi.org/10.1080/15548627.2020.1816664
  20. Biochim Biophys Acta Bioenerg. 2020 Aug 27. pii: S0005-2728(20)30152-3. [Epub ahead of print] 148302
    The regulation of mitochondrial homeostasis by the Ubiquitin Proteasome System.
    Cynthia Alsayyah, Oznur Ozturk, Laetitia Cavellini, Naïma Belgareh-Touzé, Mickael M Cohen.
      From mitochondrial quality control pathways to the regulation of specific functions, the Ubiquitin Proteasome System (UPS) could be compared to a Swiss knife without which mitochondria could not maintain its integrity in the cell. Here, we review the mechanisms that the UPS employs to regulate mitochondrial function and efficiency. For this purpose, we depict how Ubiquitin and the Proteasome participate in diverse quality control pathways that safeguard entry into the mitochondrial compartment. A focus is then achieved on the UPS-mediated control of the yeast mitofusin Fzo1 which provides insights into the complex regulation of this particular protein in mitochondrial fusion. We ultimately dissect the mechanisms by which the UPS controls the degradation of mitochondria by autophagy in both mammalian and yeast systems. This organization should offer a useful overview of this abundant but fascinating literature on the crosstalks between mitochondria and the UPS.
    Keywords:  Mitochondria; Mitochondrial Quality Control; Mitochondrial fusion; Mitophagy; Proteasome; Ubiquitin
    DOI:  https://doi.org/10.1016/j.bbabio.2020.148302
  21. Mol Neurobiol. 2020 Aug 31.
    Antioxidant Modulation of mTOR and Sirtuin Pathways in Age-Related Neurodegenerative Diseases.
    Asmaa Abdullah, Nuraqila Mohd Murshid, Suzana Makpol.
      In the human body, cell division and metabolism are expected to transpire uneventfully for approximately 25 years. Then, secondary metabolism and cell damage products accumulate, and ageing phenotypes are acquired, causing the progression of disease. Among these age-related diseases, neurodegenerative diseases have attracted considerable attention because of their irreversibility, the absence of effective treatment and their relationship with social and economic pressures. Mechanistic (formerly mammalian) target of rapamycin (mTOR), sirtuin (SIRT) and insulin/insulin growth factor 1 (IGF1) signalling pathways are among the most important pathways in ageing-associated conditions, such as neurodegeneration. These longevity-related pathways are associated with a diversity of various processes, including metabolism, cognition, stress reaction and brain plasticity. In this review, we discuss the roles of sirtuin and mTOR in ageing and neurodegeneration, with an emphasis on their regulation of autophagy, apoptosis and mitochondrial energy metabolism. The intervention of neurodegeneration using potential antioxidants, including vitamins, phytochemicals, resveratrol, herbals, curcumin, coenzyme Q10 and minerals, specifically aimed at retaining mitochondrial function in the treatment of Alzheimer's disease, Parkinson's disease and Huntington's disease is highlighted.
    Keywords:  Ageing; Antioxidants; Neurodegenerative diseases; Sirtuins; mTOR
    DOI:  https://doi.org/10.1007/s12035-020-02083-1
  22. Cells. 2020 Sep 01. pii: E2008. [Epub ahead of print]9(9):
    Atg8-Family Proteins-Structural Features and Molecular Interactions in Autophagy and Beyond.
    Nicole Wesch, Vladimir Kirkin, Vladimir V Rogov.
      Autophagy is a common name for a number of catabolic processes, which keep the cellular homeostasis by removing damaged and dysfunctional intracellular components. Impairment or misbalance of autophagy can lead to various diseases, such as neurodegeneration, infection diseases, and cancer. A central axis of autophagy is formed along the interactions of autophagy modifiers (Atg8-family proteins) with a variety of their cellular counter partners. Besides autophagy, Atg8-proteins participate in many other pathways, among which membrane trafficking and neuronal signaling are the most known. Despite the fact that autophagy modifiers are well-studied, as the small globular proteins show similarity to ubiquitin on a structural level, the mechanism of their interactions are still not completely understood. A thorough analysis and classification of all known mechanisms of Atg8-protein interactions could shed light on their functioning and connect the pathways involving Atg8-proteins. In this review, we present our views of the key features of the Atg8-proteins and describe the basic principles of their recognition and binding by interaction partners. We discuss affinity and selectivity of their interactions as well as provide perspectives for discovery of new Atg8-interacting proteins and therapeutic approaches to tackle major human diseases.
    Keywords:  Atg8; GABARAP; LC3; LIR motif; SAR; UBL; autophagy
    DOI:  https://doi.org/10.3390/cells9092008
  23. Nat Cell Biol. 2020 Sep;22(9): 1091-1102
    The primary cilium and lipophagy translate mechanical forces to direct metabolic adaptation of kidney epithelial cells.
    Caterina Miceli, Federica Roccio, Lucille Penalva-Mousset, Martine Burtin, Christine Leroy, Ivan Nemazanyy, Nicolas Kuperwasser, Marco Pontoglio, Gérard Friedlander, Etienne Morel, Fabiola Terzi, Patrice Codogno, Nicolas Dupont.
      Organs and cells must adapt to shear stress induced by biological fluids, but how fluid flow contributes to the execution of specific cell programs is poorly understood. Here we show that shear stress favours mitochondrial biogenesis and metabolic reprogramming to ensure energy production and cellular adaptation in kidney epithelial cells. Shear stress stimulates lipophagy, contributing to the production of fatty acids that provide mitochondrial substrates to generate ATP through β-oxidation. This flow-induced process is dependent on the primary cilia located on the apical side of epithelial cells. The interplay between fluid flow and lipid metabolism was confirmed in vivo using a unilateral ureteral obstruction mouse model. Finally, primary cilium-dependent lipophagy and mitochondrial biogenesis are required to support energy-consuming cellular processes such as glucose reabsorption, gluconeogenesis and cytoskeletal remodelling. Our findings demonstrate how primary cilia and autophagy are involved in the translation of mechanical forces into metabolic adaptation.
    DOI:  https://doi.org/10.1038/s41556-020-0566-0
  24. Open Biol. 2020 Sep;10(9): 200184
    The impact of autophagy during the development and survival of glioblastoma.
    Joanne E Simpson, Noor Gammoh.
      Glioblastoma is the most common and aggressive adult brain tumour, with poor median survival and limited treatment options. Following surgical resection and chemotherapy, recurrence of the disease is inevitable. Genomic studies have identified key drivers of glioblastoma development, including amplifications of receptor tyrosine kinases, which drive tumour growth. To improve treatment, it is crucial to understand survival response processes in glioblastoma that fuel cell proliferation and promote resistance to treatment. One such process is autophagy, a catabolic pathway that delivers cellular components sequestered into vesicles for lysosomal degradation. Autophagy plays an important role in maintaining cellular homeostasis and is upregulated during stress conditions, such as limited nutrient and oxygen availability, and in response to anti-cancer therapy. Autophagy can also regulate pro-growth signalling and metabolic rewiring of cancer cells in order to support tumour growth. In this review, we will discuss our current understanding of how autophagy is implicated in glioblastoma development and survival. When appropriate, we will refer to findings derived from the role of autophagy in other cancer models and predict the outcome of manipulating autophagy during glioblastoma treatment.
    Keywords:  autophagy; brain tumour; cancer; glioblastoma; receptor tyrosine kinase (RTK); therapy
    DOI:  https://doi.org/10.1098/rsob.200184
  25. J Virol. 2020 Sep 02. pii: JVI.01575-20. [Epub ahead of print]
    Mechanistic Target of Rapamycin (mTOR) Signaling Activation Antagonizes Autophagy to Facilitate Zika Virus Replication.
    Bikash R Sahoo, Aryamav Pattnaik, Arun S Annamalai, Rodrigo Franco, Asit K Pattnaik.
      Zika virus (ZIKV), a mosquito-transmitted flavivirus, is linked to microcephaly and other neurological defects in neonates and Guillain-Barré syndrome in adults. The molecular mechanisms regulating ZIKV infection and pathogenic outcomes are incompletely understood. Signaling by the mechanistic (mammalian) target of rapamycin (mTOR) kinase is important for cell survival and proliferation, and viruses are known to hijack this pathway for their replication. Here, we show that in human neuronal precursors and glial cells in culture, ZIKV infection activates both mTOR complex 1 (mTORC1) and 2 (mTORC2). Inhibition of mTOR kinase by torin1 or rapamycin results in reduction in ZIKV protein expression and progeny production. Depletion of Raptor, the defining subunit of mTORC1, by small interfering RNA (siRNA), negatively affects ZIKV protein expression and viral replication. Although depletion of Rictor, the unique subunit of mTORC2 or the mTOR kinase itself also inhibits the viral processes, the extent of inhibition is less pronounced. Autophagy is transiently induced early by ZIKV infection and impairment of autophagosome elongation by the Class III PI3K inhibitor 3-methyladenine (3-MA) enhances viral protein accumulation and progeny production. mTOR phosphorylates and inactivates ULK1 (S757) at later stages of ZIKV infection, suggesting a link between autophagy inhibition and mTOR activation by ZIKV. Accordingly, inhibition of ULK1 (by MRT68921) or autophagy (by 3-MA) reversed the effects of mTOR-inhibition, leading to increased levels of ZIKV protein expression and progeny production. Our results demonstrate that ZIKV replication requires the activation of both mTORC1 and mTORC2, which negatively regulates autophagy to facilitate ZIKV replication.IMPORTANCE The re-emergence of Zika virus (ZIKV) and its association with neurological complications necessitates studies on the molecular mechanisms that regulate ZIKV pathogenesis. The mTOR signaling cascade is tightly regulated and central to normal neuronal development and survival. Disruption of mTOR signaling can result in neurological abnormalities. In the studies reported here, we demonstrate for the first time that ZIKV infection results in activation of both mTORC1 and mTORC2 to promote the virus replication. Although autophagy is activated early in infection to counter virus replication, it is subsequently suppressed by the mTOR. These results reveal critical roles of mTOR signaling and autophagy in ZIKV infection and point to a possible mechanism underlying ZIKV-induced pathogenesis. Elucidating the role of mTOR signaling in ZIKV infection will provide insights into our understanding of the mechanisms of ZIKV-induced neurological complications and potential targets for therapeutic approaches.
    DOI:  https://doi.org/10.1128/JVI.01575-20
  26. Crit Rev Oncog. 2020 ;25(1): 21-30
    Autophagy-Induced Drug Resistance in Liver Cancer.
    Rahul Kumar Vempati, Rama Rao Malla.
      Autophagy is a self-destructive process that occurs in the cells during abnormal conditions like protein aggregation due to misfolding, nutrient deprivation, damage to vital cell organelles, pathogenic infections, and during cancer. Typically, autophagy plays a key role in the renovation of new cells by balancing the equilibrium between cell death and cell renewal. Dysregulation of autophagy has a profound effect on protein turnover, mitochondrial homeostasis, clearance of damaged organelles, and cellular metabolism, which lead to neurodegenerative, metabolic, and proliferative diseases. Despite its antitumorigenic role, autophagy can promote cell proliferation by enhancing chemotherapeutic resistance in liver cancer. In the present review, we provide a comprehensive overview and discussion on the role of autophagy in the drug-resistant mechanisms of liver cancer.
    DOI:  https://doi.org/10.1615/CritRevOncog.2020034969
  27. Cells. 2020 Sep 01. pii: E2013. [Epub ahead of print]9(9):
    Targeting Oxidative Phosphorylation Reverses Drug Resistance in Cancer Cells by Blocking Autophagy Recycling.
    Jae-Seon Lee, Ho Lee, Hyonchol Jang, Sang Myung Woo, Jong Bae Park, Seon-Hyeong Lee, Joon Hee Kang, Hee Yeon Kim, Jaewhan Song, Soo-Youl Kim.
      The greatest challenge in cancer therapy is posed by drug-resistant recurrence following treatment. Anticancer chemotherapy is largely focused on targeting the rapid proliferation and biosynthesis of cancer cells. This strategy has the potential to trigger autophagy, enabling cancer cell survival through the recycling of molecules and energy essential for biosynthesis, leading to drug resistance. Autophagy recycling contributes amino acids and ATP to restore mTOR complex 1 (mTORC1) activity, which leads to cell survival. However, autophagy with mTORC1 activation can be stalled by reducing the ATP level. We have previously shown that cytosolic NADH production supported by aldehyde dehydrogenase (ALDH) is critical for supplying ATP through oxidative phosphorylation (OxPhos) in cancer cell mitochondria. Inhibitors of the mitochondrial complex I of the OxPhos electron transfer chain and ALDH significantly reduce the ATP level selectively in cancer cells, terminating autophagy triggered by anticancer drug treatment. With the aim of overcoming drug resistance, we investigated combining the inhibition of mitochondrial complex I, using phenformin, and ALDH, using gossypol, with anticancer drug treatment. Here, we show that OxPhos targeting combined with anticancer drugs acts synergistically to enhance the anticancer effect in mouse xenograft models of various cancers, which suggests a potential therapeutic approach for drug-resistant cancer.
    Keywords:  ATP production; aldehyde dehydrogenase; cancer metabolism; energy metabolism; oxidative phosphorylation (OxPhos)
    DOI:  https://doi.org/10.3390/cells9092013
  28. Autophagy. 2020 Aug 31.
    A new mechanism for autophagy regulation of anti-tumor immune responses.
    Andrew Thorburn.
      In this commentary I discuss a recent paper that describes a new mechanism for how macroautophagy/autophagy regulates the immune response to cancer, and relate it to other recent studies in this area. These recent developments may allow more effective strategies to manipulate autophagy to improve cancer therapy.
    Keywords:  abscopal response; anti-tumor immunity; autophagy; cancer; radiotherapy
    DOI:  https://doi.org/10.1080/15548627.2020.1817286
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