bims-auttor Biomed News
on Autophagy and mTOR
Issue of 2021–05–09
29 papers selected by
Viktor Korolchuk, Newcastle University
  1. Prohibitin depletion extends lifespan of a TORC2/SGK-1 mutant through autophagy and the mitochondrial UPR.
  2. Activation and targeting of ATG8 protein lipidation.
  3. Mendelian neurodegenerative disease genes involved in autophagy.
  4. Manifestations of Age on Autophagy, Mitophagy and Lysosomes in Skeletal Muscle.
  5. Atg2 and Atg9: Intermembrane and interleaflet lipid transporters driving autophagy.
  6. TOR Signaling in Caenorhabditis elegans Development, Metabolism, and Aging.
  7. Loss of Ambra1 promotes melanoma growth and invasion.
  8. Cyclosporine A blocks autophagic flux in tubular epithelial cells by impairing TFEB-mediated lysosomal function.
  9. Dendritic cell PIK3C3/VPS34 controls the pathogenicity of CNS autoimmunity independently of LC3-associated phagocytosis.
  10. The SARS-CoV-2 protein ORF3a inhibits fusion of autophagosomes with lysosomes.
  11. The Contribution of Lysosomes to DNA Replication.
  12. Mitophagy in plants.
  13. Upregulation of brain cholesterol levels inhibits mitophagy in Alzheimer disease.
  14. Rapamycin protects against aristolochic acid nephropathy in mice by potentiating mammalian target of rapamycin‑mediated autophagy.
  15. Suppressor of cytokine signaling 2 is induced in Huntington's disease and involved in autophagy.
  16. RAG-ulating mTORC1 with amino acids.
  17. MicroRNA-34a activation in tuberous sclerosis complex during early brain development may lead to impaired corticogenesis.
  18. Organelle-Specific Autophagy in Cellular Aging and Rejuvenation.
  19. Conformational stabilization of optineurin by the dynamic interaction of linear polyubiquitin.
  20. CHIR99021 Augmented the Function of Late Endothelial Progenitor Cells by Preventing Replicative Senescence.
  21. mTOR in Alzheimer disease and its earlier stages: Links to oxidative damage in the progression of this dementing disorder.
  22. Lysosomal nitric oxide determines transition from autophagy to ferroptosis after exposure to plasma-activated Ringer's lactate.
  23. The human vault RNA enhances tumorigenesis and chemoresistance through the lysosome in hepatocellular carcinoma.
  24. Metabolic flexibility via mitochondrial BCAA carrier SLC25A44 is required for optimal fever.
  25. Blue LED causes autophagic cell death in human osteosarcoma by increasing ROS generation and dephosphorylating EGFR.
  26. Lysosome biogenesis: Regulation and functions.
  27. Minimized combinatorial CRISPR screens identify genetic interactions in autophagy.
  28. Relationship Between Autophagy and Metabolic Syndrome Characteristics in the Pathogenesis of Atherosclerosis.
  29. Autophagy in Hepatic Steatosis: A Structured Review.
  1. Aging Cell. 2021 May 03. e13359
    Prohibitin depletion extends lifespan of a TORC2/SGK-1 mutant through autophagy and the mitochondrial UPR.
    Patricia de la Cruz-Ruiz, Blanca Hernando-Rodríguez, Mercedes M Pérez-Jiménez, María Jesús Rodríguez-Palero, Manuel D Martínez-Bueno, Antoni Pla, Roxani Gatsi, Marta Artal-Sanz.
      Mitochondrial prohibitins (PHB) are highly conserved proteins with a peculiar effect on lifespan. While PHB depletion shortens lifespan of wild-type animals, it enhances longevity of a plethora of metabolically compromised mutants, including target of rapamycin complex 2 (TORC2) mutants sgk-1 and rict-1. Here, we show that sgk-1 mutants have impaired mitochondrial homeostasis, lipogenesis and yolk formation, plausibly due to alterations in membrane lipid and sterol homeostasis. Remarkably, all these features are suppressed by PHB depletion. Our analysis shows the requirement of SRBP1/SBP-1 for the lifespan extension of sgk-1 mutants and the further extension conferred by PHB depletion. Moreover, although the mitochondrial unfolded protein response (UPRmt ) and autophagy are induced in sgk-1 mutants and upon PHB depletion, they are dispensable for lifespan. However, the enhanced longevity caused by PHB depletion in sgk-1 mutants requires both, the UPRmt and autophagy, but not mitophagy. We hypothesize that UPRmt induction upon PHB depletion extends lifespan of sgk-1 mutants through autophagy and probably modulation of lipid metabolism.
    Keywords:  SGK-1; UPRmt; autophagy; lipogenesis; mitochondria; prohibitin
    DOI:  https://doi.org/10.1111/acel.13359
  2. Cell Discov. 2020 May 05. 6(1): 23
    Activation and targeting of ATG8 protein lipidation.
    Sascha Martens, Dorotea Fracchiolla.
      ATG8 family proteins are evolutionary conserved ubiquitin-like modifiers, which become attached to the headgroup of the membrane lipid phosphatidylethanolamine in a process referred to as lipidation. This reaction is carried out analogous to the conjugation of ubiquitin to its target proteins, involving the E1-like ATG7, the E2-like ATG3 and the E3-like ATG12-ATG5-ATG16 complex, which determines the site of lipidation. ATG8 lipidation is a hallmark of autophagy where these proteins are involved in autophagosome formation, the fusion of autophagosomes with lysosomes and cargo selection. However, it has become evident that ATG8 lipidation also occurs in processes that are not directly related to autophagy. Here we discuss recent insights into the targeting of ATG8 lipidation in autophagy and other pathways with special emphasis on the recruitment and activation of the E3-like complex.
    DOI:  https://doi.org/10.1038/s41421-020-0155-1
  3. Cell Discov. 2020 May 05. 6(1): 24
    Mendelian neurodegenerative disease genes involved in autophagy.
    Eleanna Stamatakou, Lidia Wróbel, Sandra Malmgren Hill, Claudia Puri, Sung Min Son, Motoki Fujimaki, Ye Zhu, Farah Siddiqi, Marian Fernandez-Estevez, Marco M Manni, So Jung Park, Julien Villeneuve, David Chaim Rubinsztein.
      The lysosomal degradation pathway of macroautophagy (herein referred to as autophagy) plays a crucial role in cellular physiology by regulating the removal of unwanted cargoes such as protein aggregates and damaged organelles. Over the last five decades, significant progress has been made in understanding the molecular mechanisms that regulate autophagy and its roles in human physiology and diseases. These advances, together with discoveries in human genetics linking autophagy-related gene mutations to specific diseases, provide a better understanding of the mechanisms by which autophagy-dependent pathways can be potentially targeted for treating human diseases. Here, we review mutations that have been identified in genes involved in autophagy and their associations with neurodegenerative diseases.
    DOI:  https://doi.org/10.1038/s41421-020-0158-y
  4. Cells. 2021 Apr 29. pii: 1054. [Epub ahead of print]10(5):
    Manifestations of Age on Autophagy, Mitophagy and Lysosomes in Skeletal Muscle.
    Matthew Triolo, David A Hood.
      Sarcopenia is the loss of both muscle mass and function with age. Although the molecular underpinnings of sarcopenia are not fully understood, numerous pathways are implicated, including autophagy, in which defective cargo is selectively identified and degraded at the lysosome. The specific tagging and degradation of mitochondria is termed mitophagy, a process important for the maintenance of an organelle pool that functions efficiently in energy production and with relatively low reactive oxygen species production. Emerging data, yet insufficient, have implicated various steps in this pathway as potential contributors to the aging muscle atrophy phenotype. Included in this is the lysosome, the end-stage organelle possessing a host of proteolytic and degradative enzymes, and a function devoted to the hydrolysis and breakdown of defective molecular complexes and organelles. This review provides a summary of our current understanding of how the autophagy-lysosome system is regulated in aging muscle, highlighting specific areas where knowledge gaps exist. Characterization of the autophagy pathway with a particular focus on the lysosome will undoubtedly pave the way for the development of novel therapeutic strategies to combat age-related muscle loss.
    Keywords:  aging; autophagy; lysosomes; mitophagy; sarcopenia; skeletal muscle
    DOI:  https://doi.org/10.3390/cells10051054
  5. Biochim Biophys Acta Mol Cell Biol Lipids. 2021 Apr 28. pii: S1388-1981(21)00084-6. [Epub ahead of print]1866(8): 158956
    Atg2 and Atg9: Intermembrane and interleaflet lipid transporters driving autophagy.
    Nobuo N Noda.
      Autophagy, an intracellular degradation mechanism, involves de novo generation of autophagosomes that sequester and deliver cytoplasmic components to the lysosome for degradation. The mechanism behind autophagosomal membrane expansion has been a longstanding enigma in this field. Recent structural and biochemical analyses have revealed that two mysterious autophagy-related (Atg) proteins, Atg2 and Atg9, are novel types of intermembrane and interleaflet lipid transporters, respectively. This review summarizes recent discoveries surrounding Atg2 and Atg9 as a lipid transporter and discusses the molecular mechanism of autophagosomal membrane expansion driven by collaboration between these two lipid transporters.
    Keywords:  Atg2; Atg9; Autophagosome; Autophagy; Lipid transporter; Scramblase
    DOI:  https://doi.org/10.1016/j.bbalip.2021.158956
  6. Genetics. 2019 Oct 01. 213(2): 329-360
    TOR Signaling in Caenorhabditis elegans Development, Metabolism, and Aging.
    T Keith Blackwell, Aileen K Sewell, Ziyun Wu, Min Han.
      The Target of Rapamycin (TOR or mTOR) is a serine/threonine kinase that regulates growth, development, and behaviors by modulating protein synthesis, autophagy, and multiple other cellular processes in response to changes in nutrients and other cues. Over recent years, TOR has been studied intensively in mammalian cell culture and genetic systems because of its importance in growth, metabolism, cancer, and aging. Through its advantages for unbiased, and high-throughput, genetic and in vivo studies, Caenorhabditis elegans has made major contributions to our understanding of TOR biology. Genetic analyses in the worm have revealed unexpected aspects of TOR functions and regulation, and have the potential to further expand our understanding of how growth and metabolic regulation influence development. In the aging field, C. elegans has played a leading role in revealing the promise of TOR inhibition as a strategy for extending life span, and identifying mechanisms that function upstream and downstream of TOR to influence aging. Here, we review the state of the TOR field in C. elegans, and focus on what we have learned about its functions in development, metabolism, and aging. We discuss knowledge gaps, including the potential pitfalls in translating findings back and forth across organisms, but also describe how TOR is important for C. elegans biology, and how C. elegans work has developed paradigms of great importance for the broader TOR field.
    Keywords:   Caenorhabditis elegans development; DAF-15; NPRL-2; NPRL-3; Nprl2; Nprl3; RAGA-1; RSKS-1; RagA; RagC; Raptor; Rheb; Rheb-1; Rictor; S6 kinase; TOR; TORC1; TORC2; WormBook; aging; growth regulation; metabolism; nutrient signaling; sphingolipid
    DOI:  https://doi.org/10.1534/genetics.119.302504
  7. Nat Commun. 2021 05 05. 12(1): 2550
    Loss of Ambra1 promotes melanoma growth and invasion.
    Luca Di Leo, Valérie Bodemeyer, Francesca M Bosisio, Giuseppina Claps, Marco Carretta, Salvatore Rizza, Fiorella Faienza, Alex Frias, Shawez Khan, Matteo Bordi, Maria P Pacheco, Julie Di Martino, Jose J Bravo-Cordero, Colin J Daniel, Rosalie C Sears, Marco Donia, Daniel H Madsen, Per Guldberg, Giuseppe Filomeni, Thomas Sauter, Caroline Robert, Daniela De Zio, Francesco Cecconi.
      Melanoma is the deadliest skin cancer. Despite improvements in the understanding of the molecular mechanisms underlying melanoma biology and in defining new curative strategies, the therapeutic needs for this disease have not yet been fulfilled. Herein, we provide evidence that the Activating Molecule in Beclin-1-Regulated Autophagy (Ambra1) contributes to melanoma development. Indeed, we show that Ambra1 deficiency confers accelerated tumor growth and decreased overall survival in Braf/Pten-mutated mouse models of melanoma. Also, we demonstrate that Ambra1 deletion promotes melanoma aggressiveness and metastasis by increasing cell motility/invasion and activating an EMT-like process. Moreover, we show that Ambra1 deficiency in melanoma impacts extracellular matrix remodeling and induces hyperactivation of the focal adhesion kinase 1 (FAK1) signaling, whose inhibition is able to reduce cell invasion and melanoma growth. Overall, our findings identify a function for AMBRA1 as tumor suppressor in melanoma, proposing FAK1 inhibition as a therapeutic strategy for AMBRA1 low-expressing melanoma.
    DOI:  https://doi.org/10.1038/s41467-021-22772-2
  8. J Cell Mol Med. 2021 May 04.
    Cyclosporine A blocks autophagic flux in tubular epithelial cells by impairing TFEB-mediated lysosomal function.
    Zhi-Hang Li, Ning An, Xi-Jie Huang, Chen Yang, Hong-Luan Wu, Xiao-Cui Chen, Qing-Jun Pan, Hua-Feng Liu.
      Cyclosporine A (CsA) is an immunosuppressor widely used for the prevention of acute rejection during solid organ transplantation. However, severe nephrotoxicity has substantially limited its long-term usage. Recently, an impaired autophagy pathway was suggested to be involved in the pathogenesis of chronic CsA nephrotoxicity. However, the underlying mechanisms of CsA-induced autophagy blockade in tubular cells remain unclear. In the present study, we observed that CsA suppressed the activation and expression of transcription factor EB (TFEB) by increasing the activation of mTOR, in turn promoting lysosomal dysfunction and autophagy flux blockade in tubular epithelial cells (TECs) in vivo and in vitro. Restoration of TFEB activation by Torin1-mediated mTOR inhibition significantly improved lysosomal function and rescued autophagy pathway activity, suppressing TEC injury. In summary, targeting TFEB-mediated autophagy flux represents a potential therapeutic strategy for CsA-induced nephrotoxicity.
    Keywords:  TFEB; autophagy; cyclosporine A; lysosome; mTOR; tubular epithelial cells
    DOI:  https://doi.org/10.1111/jcmm.16593
  9. Autophagy. 2021 May 07. 1-10
    Dendritic cell PIK3C3/VPS34 controls the pathogenicity of CNS autoimmunity independently of LC3-associated phagocytosis.
    Guan Yang, J Luke Postoak, Wenqiang Song, Jennifer Martinez, Jianhua Zhang, Lan Wu, Luc Van Kaer.
      PIK3C3/VPS34 is a key player in macroautophagy/autophagy and MAP1LC3/LC3-associated phagocytosis (LAP), which play critical roles in dendritic cell (DC) function. In this study, we assessed the contribution of PIK3C3 to DC function during experimental autoimmune encephalomyelitis (EAE), an animal model of multiple sclerosis (MS). We found that Pik3c3-deficient DCs exhibit attenuated capacity to reactivate encephalitogenic T cells in the central nervous system, leading to reduced incidence and severity of EAE in DC-specific Pik3c3-deficient mice. Additionally, animals with a DC-specific deficiency in Rb1cc1/Fip200 but not Rubcn were protected against EAE, suggesting that the EAE phenotype of DC-specific Pik3c3-deficient mice is due to defective canonical autophagy rather than LAP. Collectively, our studies have revealed a critical role of PIK3C3 in DC function and the pathogenicity of these cells during EAE, with important implications for the development of immunotherapies for autoimmune diseases such as MS.
    Keywords:  Autophagy; LC3-associated phagocytosis; PIK3C3/VPS34; dendritic cell; experimental autoimmune encephalomyelitis
    DOI:  https://doi.org/10.1080/15548627.2021.1922051
  10. Cell Discov. 2021 May 04. 7(1): 31
    The SARS-CoV-2 protein ORF3a inhibits fusion of autophagosomes with lysosomes.
    Yabin Zhang, Hao Sun, Rongjuan Pei, Binli Mao, Zhenyu Zhao, Huihui Li, Yong Lin, Kefeng Lu.
      Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has caused the ongoing coronavirus disease 2019 pandemic. How SARS-CoV-2 regulates cellular responses to escape clearance by host cells is unknown. Autophagy is an intracellular lysosomal degradation pathway for the clearance of various cargoes, including viruses. Here, we systematically screened 28 viral proteins of SARS-CoV-2 and identified that ORF3a strongly inhibited autophagic flux by blocking the fusion of autophagosomes with lysosomes. ORF3a colocalized with lysosomes and interacted with VPS39, a component of the homotypic fusion and protein sorting (HOPS) complex. The ORF3a-VPS39 interaction prohibited the binding of HOPS with RAB7, which prevented the assembly of fusion machinery, leading to the accumulation of unfused autophagosomes. These results indicated the potential mechanism by which SARS-CoV-2 escapes degradation; that is, the virus interferes with autophagosome-lysosome fusion. Furthermore, our findings will facilitate strategies targeting autophagy for conferring potential protection against the spread of SARS-CoV-2.
    DOI:  https://doi.org/10.1038/s41421-021-00268-z
  11. Cells. 2021 Apr 30. pii: 1068. [Epub ahead of print]10(5):
    The Contribution of Lysosomes to DNA Replication.
    Joanna Maria Merchut-Maya, Apolinar Maya-Mendoza.
      Lysosomes, acidic, membrane-bound organelles, are not only the core of the cellular recycling machinery, but they also serve as signaling hubs regulating various metabolic pathways. Lysosomes maintain energy homeostasis and provide pivotal substrates for anabolic processes, such as DNA replication. Every time the cell divides, its genome needs to be correctly duplicated; therefore, DNA replication requires rigorous regulation. Challenges that negatively affect DNA synthesis, such as nucleotide imbalance, result in replication stress with severe consequences for genome integrity. The lysosomal complex mTORC1 is directly involved in the synthesis of purines and pyrimidines to support DNA replication. Numerous drugs have been shown to target lysosomal function, opening an attractive avenue for new treatment strategies against various pathologies, including cancer. In this review, we focus on the interplay between lysosomal function and DNA replication through nucleic acid degradation and nucleotide biosynthesis and how these could be exploited for therapeutic purposes.
    Keywords:  DNA synthesis; autophagy; cancer; lysosomes; mTORC1
    DOI:  https://doi.org/10.3390/cells10051068
  12. Biochim Biophys Acta Gen Subj. 2021 Apr 28. pii: S0304-4165(21)00074-X. [Epub ahead of print] 129916
    Mitophagy in plants.
    Sakuya Nakamura, Shinya Hagihara, Masanori Izumi.
      Mitochondria play a central role in primary metabolism in plants as well as in heterotrophic eukaryotes. Plants must control the quality and number of mitochondria in response to a changing environment, across cell types and developmental stages. Mitophagy is defined as the degradation of mitochondria by autophagy, an evolutionarily conserved system for the removal and recycling of intracellular components. Recent studies have highlighted the importance of mitophagy in plant stress responses. This review article summarizes our current knowledge of plant mitophagy and discusses the underlying mechanisms. In plants, chloroplasts cooperate with mitochondria for energy production, and autophagy also targets chloroplasts through a process known as chlorophagy. Advances in plant autophagy studies now allow a comparative analysis of the autophagic turnover of mitochondria and chloroplasts, via the selective degradation of their soluble proteins, fragments, or entire organelles.
    Keywords:  Chlorophagy; Chloroplasts; Energy conversion; Mitochondria; Mitophagy; Plants
    DOI:  https://doi.org/10.1016/j.bbagen.2021.129916
  13. Autophagy. 2021 May 04. 1-3
    Upregulation of brain cholesterol levels inhibits mitophagy in Alzheimer disease.
    Vicente Roca-Agujetas, Cristina de Dios, Xenia Abadin, Anna Colell.
      Mitochondrial dysfunction is behind several neurodegenerative diseases, including Alzheimer disease (AD). Accumulation of damaged mitochondria is already observed at the early stages of AD and has been linked to impaired mitophagy, but the mechanisms underlying this alteration are still not fully known. In our recent study, we show that intracellular cholesterol enrichment can downregulate amyloid beta (Aβ)-induced mitophagy. Mitochondrial glutathione depletion resulting from high cholesterol levels promotes PINK1 (PTEN induced kinase 1)-mediated mitophagosome formation; however, mitophagy flux is ultimately disrupted, most likely due to fusion deficiency of endosomes-lysosomes caused by cholesterol. Meanwhile, in APP-PSEN1-SREBF2 mice, an AD mouse model that overexpresses the cholesterol-related transcription factor SREBF2, cholesterol accumulation prompts an oxidative- and age-dependent cytosolic aggregation of the mitophagy adaptor OPTN (optineurin), which prevents mitophagosome formation despite enhanced PINK1-PRKN/parkin signaling. Hippocampal neurons from postmortem brain of AD individuals reproduce the progressive accumulation of OPTN in aggresome-like structures accompanied by high levels of mitochondrial cholesterol in advanced stages of the disease. Overall, these data provide new insights into the impairment of the PINK1-PRKN mitophagy pathway in AD and suggest the combination of mitophagy inducers with strategies focused on restoring the cholesterol homeostasis and mitochondrial redox balance as a potential disease-modifying therapy for AD.
    Keywords:  Alzheimer disease; Mitophagy; PINK1; aggresomes; autophagy; cholesterol; optineurin; parkin
    DOI:  https://doi.org/10.1080/15548627.2021.1920814
  14. Mol Med Rep. 2021 Jul;pii: 495. [Epub ahead of print]24(1):
    Rapamycin protects against aristolochic acid nephropathy in mice by potentiating mammalian target of rapamycin‑mediated autophagy.
    Fan Lin, Yunqi Liu, Lili Tang, Xiaohui Xu, Xueli Zhang, Yifan Song, Bicheng Chen, Yeping Ren, Xiangdong Yang.
      Autophagy serves a crucial role in the etiology of kidney diseases, including drug‑induced renal impairment, inherited kidney disease, diabetic nephropathy and aristolochic acid nephropathy (AAN) and is, therefore, a potential target for treatment. We previously demonstrated that rapamycin could attenuate AAN in mice; however, the underlying mechanism remains to be elucidated. Therefore, whether the renal protective effect of rapamycin (an autophagy activator) is related to autophagy in aristolochic acid (AA)‑treated mice was of particular interest. The pathophysiological roles of rapamycin were investigated in AA‑induced nephrotoxicity in mice and the mechanisms of rapamycin action were explored by evaluating the modulation of autophagy in rapamycin‑treated mice and cultured renal tubular epithelial cells. Supplementation with rapamycin reversed AA‑induced kidney injury in mice and improved AA‑induced autophagy damage in vivo and in vitro. Mechanistically, rapamycin inhibited the renal expression of phosphorylated (p‑)mammalian target of rapamycin (mTOR) and p‑ribosomal S6 protein kinase 1, which in turn activated renal autophagy and decreased apoptosis, probably by removing AA‑elicited damaged mitochondria and misfolded proteins. The findings of the present study demonstrated that rapamycin protects against AA‑induced nephropathy by activating the mTOR‑autophagy axis and suggested that rapamycin may be a promising pharmacological target for the treatment of AAN.
    Keywords:  apoptosis; aristolochic acid nephropathy; autophagy; mTOR; rapamycin
    DOI:  https://doi.org/10.3892/mmr.2021.12134
  15. Biochem Biophys Res Commun. 2021 Apr 29. pii: S0006-291X(21)00707-5. [Epub ahead of print]559 21-27
    Suppressor of cytokine signaling 2 is induced in Huntington's disease and involved in autophagy.
    KyoungJoo Cho, Sejeong Kim, Seung Ho Choi.
      Suppressor of cytokine signaling (SOCS) proteins are primarily feedback inhibitors of cytokine signaling. The two conserved domains of SOCS proteins have distinct functions. Src homology 2 (SH2) domain inhibits cytokine receptor, while SOCS box acts as an E3 ubiquitin ligase. SOCS2, a cytokine signaling suppressor, has been primarily implicated in regulating inflammatory conditions in neuronal diseases. However, SOCS proteins have been suggested to play diverse roles in healthy and diseased nervous system including neurodegenerative disorders. In this study, SOCS2 was found to be upregulated in Huntington's disease and was substantially induced in extended polyglutamine (polyQ)-expressing striatal cells. The induced level was augmented under aging conditions. In extended polyQ-expressing cells, downregulated SOCS2 improved autophagic dysfunction rather than altered inflammatory conditions. Overall, we suggest that SOCS2 involves in regulating autophagy by functioning as an E3 ligase in extended polyQ conditions, and consequently regulates cell damage and cell death type.
    Keywords:  Autophagy; E3 ligase; Huntington disease; Inflammation; Suppressor of cytokine signaling-2
    DOI:  https://doi.org/10.1016/j.bbrc.2021.04.089
  16. Nat Rev Mol Cell Biol. 2021 May 04.
    RAG-ulating mTORC1 with amino acids.
    Jenna L Jewell.
      
    DOI:  https://doi.org/10.1038/s41580-021-00378-2
  17. Neuropathol Appl Neurobiol. 2021 May 03.
    MicroRNA-34a activation in tuberous sclerosis complex during early brain development may lead to impaired corticogenesis.
    A Korotkov, N S Sim, M J Luinenburg, J J Anink, J van Scheppingen, T S Zimmer, A Bongaarts, D W M Broekaart, C Mijnsbergen, F E Jansen, W Van Hecke, W G M Spliet, P C van Rijen, M Feucht, J A Hainfellner, P Krsek, J Zamecnik, P B Crino, K Kotulska, L Lagae, A C Jansen, D J Kwiatkowski, S Jozwiak, P Curatolo, A Mühlebner, J H Lee, J D Mills, E A van Vliet, E Aronica.
       AIMS: Tuberous sclerosis complex (TSC) is a genetic disorder associated with dysregulation of the mechanistic target of rapamycin complex 1 (mTORC1) signalling pathway. Neurodevelopmental disorders, frequently present in TSC, are linked to cortical tubers in the brain. We previously reported microRNA-34a (miR-34a) among the most up-regulated miRs in tubers. Here, we characterized miR-34a expression in tubers with the focus on the early brain development and assessed the regulation of mTORC1 pathway and corticogenesis by miR-34a.
    METHODS: We analysed the expression of miR-34a in resected cortical tubers (n = 37) compared to autopsy-derived control tissue (n = 27). The effect of miR-34a overexpression on corticogenesis was assessed in mice at E18. The regulation of the mTORC1 pathway and the expression of the bioinformatically predicted target genes were assessed in primary astrocyte cultures from 3 patients with TSC and in SH-SY5Y cells following miR-34a transfection.
    RESULTS: The peak of miR-34a overexpression in tubers was observed during infancy, concomitant with the presence of pathological markers, particularly in giant cells and dysmorphic neurons. MiR-34a was also strongly expressed in fetal TSC cortex. Overexpression of miR-34a in mouse embryos decreased the percentage of cells migrated to the cortical plate. The transfection of miR-34a mimic in TSC astrocytes negatively regulated mTORC1 and decreased the expression of the target genes RAS related (RRAS) and NOTCH1.
    CONCLUSIONS: MiR-34a is most highly overexpressed in tubers during fetal and early postnatal brain development. MiR-34a can negatively regulate mTORC1, however, it may also contribute to abnormal corticogenesis in TSC.
    Keywords:  TSC; mechanistic target of rapamycin; miRNA; migration; neurodevelopmental disorder
    DOI:  https://doi.org/10.1111/nan.12717
  18. Adv Geriatr Med Res. 2021 ;pii: e210010. [Epub ahead of print]3(2):
    Organelle-Specific Autophagy in Cellular Aging and Rejuvenation.
    Tyler J Butsch, Bhaswati Ghosh, K Adam Bohnert.
      The health of a cell requires proper functioning, regulation, and quality control of its organelles, the membrane-enclosed compartments inside the cell that carry out its essential biochemical tasks. Aging commonly perturbs organelle homeostasis, causing problems to cellular health that can spur the initiation and progression of degenerative diseases and related pathologies. Here, we discuss emerging evidence indicating that age-related defects in organelle homeostasis stem in part from dysfunction of the autophagy-lysosome system, a pivotal player in cellular quality control and damage clearance. We also highlight natural examples from biology where enhanced activity of the autophagy-lysosome system might be harnessed to erase age-related organelle damage, raising potential implications for cellular rejuvenation.
    Keywords:  aging; autophagy; cell biology; organelles; rejuvenation
    DOI:  https://doi.org/10.20900/agmr20210010
  19. Biochem Biophys Res Commun. 2021 May 02. pii: S0006-291X(21)00723-3. [Epub ahead of print]559 203-209
    Conformational stabilization of optineurin by the dynamic interaction of linear polyubiquitin.
    Akira Kitamura, Rika Numazawa, Masataka Kinjo.
      Optineurin produces intracellular multi-functions involving autophagy, vesicular trafficking, and negative regulation of inflammation signaling through interaction with various proteins such as ATG8/LC3, Rab8, and polyubiquitin. Optineurin is a component of cytoplasmic inclusion bodies (IBs) in motor neurons from amyotrophic lateral sclerosis (ALS), and its mutation E478G, has been identified in patients with ALS. However, the mechanism by which polyubiquitin binding modulates the interaction partners of OPTN and ALS-associated IB formation is still unclear. To address this issue, we analyzed the interaction of Optineurin with Rab8 and LC3 in the absence and presence of linear polyubiquitin chains using fluorescence cross-correlation spectroscopy and IB formation efficiency of the E478G mutant of Optineurin during Rab8 depletion using fluorescence microscopy. Here, we hypothesize that linear polyubiquitin binding to Optineurin dynamically induces LC3 association and Rab8 dissociation, likely through a conformational change of Optineurin, and the dynamic conformational change may prevent the aggregate formation of mutant Optineurin.
    Keywords:  ALS; FCCS; LC3; Optineurin; Polyubiquitin; Protein aggregation; Rab8
    DOI:  https://doi.org/10.1016/j.bbrc.2021.04.103
  20. Int J Mol Sci. 2021 Apr 30. pii: 4796. [Epub ahead of print]22(9):
    CHIR99021 Augmented the Function of Late Endothelial Progenitor Cells by Preventing Replicative Senescence.
    Vinoth Kumar Rethineswaran, Da Yeon Kim, Yeon-Ju Kim, WoongBi Jang, Seung Taek Ji, Le Thi Hong Van, Ly Thanh Truong Giang, Jong Seong Ha, Jisoo Yun, Jinsup Jung, Sang-Mo Kwon.
      Endothelial progenitor cells (EPCs) are specialized cells in circulating blood, well known for their ability to form new vascular structures. Aging and various ailments such as diabetes, atherosclerosis and cardiovascular disease make EPCs vulnerable to decreasing in number, which affects their migration, proliferation and angiogenesis. Myocardial ischemia is also linked to a reduced number of EPCs and their endothelial functional role, which hinders proper blood circulation to the myocardium. The current study shows that an aminopyrimidine derivative compound (CHIR99021) induces the inhibition of GSK-3β in cultured late EPCs. GSK-3β inhibition subsequently inhibits mTOR by blocking the phosphorylation of TSC2 and lysosomal localization of mTOR. Furthermore, suppression of GSK-3β activity considerably increased lysosomal activation and autophagy. The activation of lysosomes and autophagy by GSK-3β inhibition not only prevented replicative senescence of the late EPCs but also directed their migration, proliferation and angiogenesis. To conclude, our results demonstrate that lysosome activation and autophagy play a crucial role in blocking the replicative senescence of EPCs and in increasing their endothelial function. Thus, the findings provide an insight towards the treatment of ischemia-associated cardiovascular diseases based on the role of late EPCs.
    Keywords:  CHIR99021; EPC; GSK-3β; autophagy; lysosome; mTOR; senescence
    DOI:  https://doi.org/10.3390/ijms22094796
  21. Free Radic Biol Med. 2021 Apr 30. pii: S0891-5849(21)00260-4. [Epub ahead of print]169 382-396
    mTOR in Alzheimer disease and its earlier stages: Links to oxidative damage in the progression of this dementing disorder.
    M Perluigi, F Di Domenico, E Barone, D A Butterfield.
      Alzheimer's disease (AD) is the most prevalent form of dementia in the elderly population and has worldwide impact. The etiology of the disease is complex and results from the confluence of multiple mechanisms ultimately leading to neuronal loss and cognitive decline. Among risk factors, aging is the most relevant and accounts for several pathogenic events that contribute to disease-specific toxic mechanisms. Accumulating evidence linked the alterations of the mammalian target of rapamycin (mTOR), a serine/threonine protein kinase playing a key role in the regulation of protein synthesis and degradation, to age-dependent cognitive decline and pathogenesis of AD. To date, growing studies demonstrated that aberrant mTOR signaling in the brain affects several pathways involved in energy metabolism, cell growth, mitochondrial function and proteostasis. Recent advances associated alterations of the mTOR pathway with the increased oxidative stress. Disruption of all these events strongly contribute to age-related cognitive decline including AD. The current review discusses the main regulatory roles of mTOR signaling network in the brain, focusing on its role in autophagy, oxidative stress and energy metabolism. Collectively, experimental data suggest that targeting mTOR in the CNS can be a valuable strategy to prevent/slow the progression of AD.
    Keywords:  Alzheimer's disease; Oxidative stress; Protein aggregation; Proteostasis; mTOR
    DOI:  https://doi.org/10.1016/j.freeradbiomed.2021.04.025
  22. Redox Biol. 2021 Apr 23. pii: S2213-2317(21)00137-3. [Epub ahead of print]43 101989
    Lysosomal nitric oxide determines transition from autophagy to ferroptosis after exposure to plasma-activated Ringer's lactate.
    Li Jiang, Hao Zheng, Qinying Lyu, Shotaro Hayashi, Kotaro Sato, Yoshitaka Sekido, Kae Nakamura, Hiromasa Tanaka, Kenji Ishikawa, Hiroaki Kajiyama, Masaaki Mizuno, Masaru Hori, Shinya Toyokuni.
      Non-thermal plasma (NTP), an engineered technology to generate reactive species, induces ferroptosis and/or apoptosis specifically in various-type cancer cells. NTP-activated Ringer's lactate (PAL) is another modality for cancer therapy at preclinical stage. Here we found that PAL induces selective ferroptosis of malignant mesothelioma (MM) cells, where non-targeted metabolome screening identified upregulated citrulline-nitric oxide (.NO) cycle as a PAL target. .NO probe detected biphasic peaks transiently at PAL exposure with time-dependent increase, which was responsible for inducible . NO synthase (iNOS) overexpression through NF-κB activation. .NO and lipid peroxidation occupied lysosomes as a major compartment with increased TFEB expression. Not only ferrostatin-1 but inhibitors for . NO and/or iNOS could suppress this ferroptosis. PAL-induced ferroptosis accompanied autophagic process in the early phase, as demonstrated by an increase in essential amino acids, LC3B-II, p62 and LAMP1, transforming into the later phase with boosted lipid peroxidation. Therefore, .NO-mediated lysosomal impairment is central in PAL-induced ferroptosis.
    Keywords:  Autophagy; Ferroptosis; Malignant mesothelioma; Nitric oxide; Non-thermal plasma-activated Ringer's lactate
    DOI:  https://doi.org/10.1016/j.redox.2021.101989
  23. Autophagy. 2021 May 07. 1-13
    The human vault RNA enhances tumorigenesis and chemoresistance through the lysosome in hepatocellular carcinoma.
    Iolanda Ferro, Jacopo Gavini, Stefano Gallo, Lisamaria Bracher, Marc Landolfo, Daniel Candinas, Deborah M Stroka, Norbert Polacek.
      The small non-coding VTRNA1-1 (vault RNA 1-1) is known to confer resistance to apoptosis in several malignant cell lines and to also modulate the macroautophagic/autophagic flux in hepatocytes, thus highlighting its pro-survival role. Here we describe a new function of VTRNA1-1 in regulating in vitro and in vivo tumor cell proliferation, tumorigenesis and chemoresistance. Knockout (KO) of VTRNA1-1 in human hepatocellular carcinoma cells reduced nuclear localization of TFEB (transcription factor EB), leading to a downregulation of the coordinated lysosomal expression and regulation (CLEAR) network genes and lysosomal compartment dysfunction. We demonstrate further that impaired lysosome function due to loss of VTRNA1-1 potentiates the anticancer effect of conventional chemotherapeutic drugs. Finally, loss of VTRNA1-1 reduced drug lysosomotropism allowing higher intracellular compound availability and thereby significantly reducing tumor cell proliferation in vitro and in vivo. These findings reveal a so far unknown role of VTRNA1-1 in the intracellular catabolic compartment and describe its contribution to lysosome-mediated chemotherapy resistance.
    Keywords:  Chemoresistance; lysosome; non-coding RNA; tumorigenesis; vault RNA; vtRNA1-1
    DOI:  https://doi.org/10.1080/15548627.2021.1922983
  24. Elife. 2021 May 04. pii: e66865. [Epub ahead of print]10
    Metabolic flexibility via mitochondrial BCAA carrier SLC25A44 is required for optimal fever.
    Takeshi Yoneshiro, Naoya Kataoka, Jacquelyn M Walejko, Kenji Ikeda, Zachary Brown, Momoko Yoneshiro, Scott B Crown, Tsuyoshi Osawa, Juro Sakai, Robert W McGarrah, Phillip J White, Kazuhiro Nakamura, Shingo Kajimura.
      Importing necessary metabolites into the mitochondrial matrix is a crucial step of fuel choice during stress adaptation. Branched chain-amino acids (BCAA) are essential amino acids needed for anabolic processes, but they are also imported into the mitochondria for catabolic reactions. What controls the distinct subcellular BCAA utilization during stress adaptation is insufficiently understood. The present study reports the role of SLC25A44, a recently identified mitochondrial BCAA carrier (MBC), in the regulation of mitochondrial BCAA catabolism and adaptive response to fever in rodents. We found that mitochondrial BCAA oxidation in brown adipose tissue (BAT) is significantly enhanced during fever in response to the pyrogenic mediator prostaglandin E2 (PGE2) and psychological stress in mice and rats. Genetic deletion of MBC in a BAT-specific manner blunts mitochondrial BCAA oxidation and non-shivering thermogenesis following intracerebroventricular PGE2 administration. At a cellular level, MBC is required for mitochondrial BCAA deamination as well as the synthesis of mitochondrial amino acids and TCA intermediates. Together, these results illuminate the role of MBC as a determinant of metabolic flexibility to mitochondrial BCAA catabolism and optimal febrile responses. This study also offers an opportunity to control fever by rewiring the subcellular BCAA fate.
    Keywords:  cell biology; mouse
    DOI:  https://doi.org/10.7554/eLife.66865
  25. J Cell Mol Med. 2021 May 07.
    Blue LED causes autophagic cell death in human osteosarcoma by increasing ROS generation and dephosphorylating EGFR.
    Mingyu He, Gege Yan, Yang Wang, Rui Gong, Hong Lei, Shuting Yu, Xiaoqi He, Guanghui Li, Weijie Du, Tianshuai Ma, Manqi Gao, Meixi Yu, Shenzhen Liu, Zihang Xu, Elina Idiiatullina, Naufal Zagidullin, Valentin Pavlov, Benzhi Cai, Ye Yuan, Lei Yang.
      Osteosarcoma (OS) is the most common primary malignant bone tumour in adolescence. Lately, light-emitting diodes (LED)-based therapy has emerged as a new promising approach for several diseases. However, it remains unknown in human OS. Here, we found that the blue LED irradiation significantly suppressed the proliferation, migration and invasion of human OS cells, while we observed blue LED irradiation increased ROS production through increased NADPH oxidase enzymes NOX2 and NOX4, as well as decreased Catalase (CAT) expression levels. Furthermore, we revealed blue LED irradiation-induced autophagy characterized by alterations in autophagy protein markers including Beclin-1, LC3-II/LC3-I and P62. Moreover, we demonstrated an enhanced autophagic flux. The blockage of autophagy displayed a remarkable attenuation of anti-tumour activities of blue LED irradiation. Next, ROS scavenger N-acetyl-L-cysteine (NAC) and NOX inhibitor diphenyleneiodonium (DPI) blocked suppression of OS cell growth, indicating that ROS accumulation might play an essential role in blue LED-induced autophagic OS cell death. Additionally, we observed blue LED irradiation decreased EGFR activation (phosphorylation), which in turn led to Beclin-1 release and subsequent autophagy activation in OS cells. Analysis of EGFR colocalization with Beclin-1 and EGFR-immunoprecipitation (IP) assay further revealed the decreased interaction of EGFR and Beclin-1 upon blue LED irradiation in OS cells. In addition, Beclin-1 down-regulation abolished the effects of blue LED irradiation on OS cells. Collectively, we concluded that blue LED irradiation exhibited anti-tumour effects on OS by triggering ROS and EGFR/Beclin-1-mediated autophagy signalling pathway, representing a potential approach for human OS treatment.
    Keywords:  autophagy; beclin-1; blue light-emitting diodes (LED); cell death; epidermal growth factor receptor (EGFR); mitochondrial reactive oxygen species (ROS); osteosarcoma (OS)
    DOI:  https://doi.org/10.1111/jcmm.16412
  26. J Cell Biol. 2021 Jun 07. pii: e202102001. [Epub ahead of print]220(6):
    Lysosome biogenesis: Regulation and functions.
    Chonglin Yang, Xiaochen Wang.
      Lysosomes are degradation centers and signaling hubs in cells and play important roles in cellular homeostasis, development, and aging. Changes in lysosome function are essential to support cellular adaptation to multiple signals and stimuli. Therefore, lysosome biogenesis and activity are regulated by a wide variety of intra- and extracellular cues. Here, we summarize current knowledge of the regulatory mechanisms of lysosome biogenesis, including synthesis of lysosomal proteins and their delivery via the endosome-lysosome pathway, reformation of lysosomes from degradative vesicles, and transcriptional regulation of lysosomal genes. We survey the regulation of lysosome biogenesis in response to nutrient and nonnutrient signals, the cell cycle, stem cell quiescence, and cell fate determination. Finally, we discuss lysosome biogenesis and functions in the context of organismal development and aging.
    DOI:  https://doi.org/10.1083/jcb.202102001
  27. Nucleic Acids Res. 2021 May 06. pii: gkab309. [Epub ahead of print]
    Minimized combinatorial CRISPR screens identify genetic interactions in autophagy.
    Valentina Diehl, Martin Wegner, Paolo Grumati, Koraljka Husnjak, Simone Schaubeck, Andrea Gubas, Varun Jayeshkumar Shah, Ibrahim H Polat, Felix Langschied, Cristian Prieto-Garcia, Konstantin Müller, Alkmini Kalousi, Ingo Ebersberger, Christian H Brandts, Ivan Dikic, Manuel Kaulich.
      Combinatorial CRISPR-Cas screens have advanced the mapping of genetic interactions, but their experimental scale limits the number of targetable gene combinations. Here, we describe 3Cs multiplexing, a rapid and scalable method to generate highly diverse and uniformly distributed combinatorial CRISPR libraries. We demonstrate that the library distribution skew is the critical determinant of its required screening coverage. By circumventing iterative cloning of PCR-amplified oligonucleotides, 3Cs multiplexing facilitates the generation of combinatorial CRISPR libraries with low distribution skews. We show that combinatorial 3Cs libraries can be screened with minimal coverages, reducing associated efforts and costs at least 10-fold. We apply a 3Cs multiplexing library targeting 12,736 autophagy gene combinations with 247,032 paired gRNAs in viability and reporter-based enrichment screens. In the viability screen, we identify, among others, the synthetic lethal WDR45B-PIK3R4 and the proliferation-enhancing ATG7-KEAP1 genetic interactions. In the reporter-based screen, we identify over 1,570 essential genetic interactions for autophagy flux, including interactions among paralogous genes, namely ATG2A-ATG2B, GABARAP-MAP1LC3B and GABARAP-GABARAPL2. However, we only observe few genetic interactions within paralogous gene families of more than two members, indicating functional compensation between them. This work establishes 3Cs multiplexing as a platform for genetic interaction screens at scale.
    DOI:  https://doi.org/10.1093/nar/gkab309
  28. Front Cell Dev Biol. 2021 ;9 641852
    Relationship Between Autophagy and Metabolic Syndrome Characteristics in the Pathogenesis of Atherosclerosis.
    Jing Xu, Munehiro Kitada, Yoshio Ogura, Daisuke Koya.
      Atherosclerosis is the main cause of mortality in metabolic-related diseases, including cardiovascular disease and type 2 diabetes (T2DM). Atherosclerosis is characterized by lipid accumulation and increased inflammatory cytokines in the vascular wall, endothelial cell and vascular smooth muscle cell dysfunction and foam cell formation initiated by monocytes/macrophages. The characteristics of metabolic syndrome (MetS), including obesity, glucose intolerance, dyslipidemia and hypertension, may activate multiple mechanisms, such as insulin resistance, oxidative stress and inflammatory pathways, thereby contributing to increased risks of developing atherosclerosis and T2DM. Autophagy is a lysosomal degradation process that plays an important role in maintaining cellular metabolic homeostasis. Increasing evidence indicates that impaired autophagy induced by MetS is related to oxidative stress, inflammation, and foam cell formation, further promoting atherosclerosis. Basal and mild adaptive autophagy protect against the progression of atherosclerotic plaques, while excessive autophagy activation leads to cell death, plaque instability or even plaque rupture. Therefore, autophagic homeostasis is essential for the development and outcome of atherosclerosis. Here, we discuss the potential role of autophagy and metabolic syndrome in the pathophysiologic mechanisms of atherosclerosis and potential therapeutic drugs that target these molecular mechanisms.
    Keywords:  atherosclerosis; autophagy; inflammation; metabolic syndrome; oxidative stress; type 2 diabetes
    DOI:  https://doi.org/10.3389/fcell.2021.641852
  29. Front Cell Dev Biol. 2021 ;9 657389
    Autophagy in Hepatic Steatosis: A Structured Review.
    Vitor de Miranda Ramos, Alicia J Kowaltowski, Pamela A Kakimoto.
      Steatosis is the accumulation of neutral lipids in the cytoplasm. In the liver, it is associated with overeating and a sedentary lifestyle, but may also be a result of xenobiotic toxicity and genetics. Non-alcoholic fatty liver disease (NAFLD) defines an array of liver conditions varying from simple steatosis to inflammation and fibrosis. Over the last years, autophagic processes have been shown to be directly associated with the development and progression of these conditions. However, the precise role of autophagy in steatosis development is still unclear. Specifically, autophagy is necessary for the regulation of basic metabolism in hepatocytes, such as glycogenolysis and gluconeogenesis, response to insulin and glucagon signaling, and cellular responses to free amino acid contents. Also, genetic knockout models for autophagy-related proteins suggest a critical relationship between autophagy and hepatic lipid metabolism, but some results are still ambiguous. While autophagy may seem necessary to support lipid oxidation in some contexts, other evidence suggests that autophagic activity can lead to lipid accumulation instead. This structured literature review aims to critically discuss, compare, and organize results over the last 10 years regarding rodent steatosis models that measured several autophagy markers, with genetic and pharmacological interventions that may help elucidate the molecular mechanisms involved.
    Keywords:  autophagy; liver; review; rodent models; steatosis
    DOI:  https://doi.org/10.3389/fcell.2021.657389
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