bims-auttor Biomed News
on Autophagy and mTOR
Issue of 2022‒05‒22
fifty-one papers selected by
Viktor Korolchuk, Newcastle University
  1. Prolonged deprivation of arginine or leucine induces PI3K/Akt-dependent reactivation of mTORC1.
  2. Targeting Selective Autophagy as a Therapeutic Strategy for Viral Infectious Diseases.
  3. Crossing paths: Recent insights in the interplay between autophagy and intracellular trafficking in plants.
  4. Phase Separation in Regulation of Autophagy.
  5. Myotubularin-related phosphatase 5 is a critical determinant of autophagy in neurons.
  6. N7-methylguanosine (m7G) tRNA modification: a novel autophagy modulator in cancer.
  7. The Interplay Between Autophagy and RNA Homeostasis: Implications for Amyotrophic Lateral Sclerosis and Frontotemporal Dementia.
  8. Autophagy regulation by RNA alternative splicing and implications in human diseases.
  9. The non-essential TSC complex component TBC1D7 restricts tissue mTORC1 signaling and brain and neuron growth.
  10. Are mitophagy enhancers therapeutic targets for Alzheimer's disease?
  11. Piperazine-Based Mitochondria-Immobilized pH Fluorescent Probe for Imaging Endogenous ONOO- and Real-Time Tracking of Mitophagy.
  12. Close association of polarization and LC3, a marker of autophagy, in axon determination in mouse hippocampal neurons.
  13. A dual-response fluorescent probe for simultaneously monitoring polarity and ATP during autophagy.
  14. Distinct designer diamines promote mitophagy, and thereby enhance healthspan in C. elegans and protect human cells against oxidative damage.
  15. Can We Treat Neurodegenerative Proteinopathies by Enhancing Protein Degradation?
  16. Cytoskeleton grows p62 condensates for autophagy.
  17. Understanding the contributions of VPS35 and the retromer in neurodegenerative disease.
  18. Autophagy targets Hd1 for vacuolar degradation to regulate rice flowering.
  19. Targeting autophagy in colorectal cancer: an update on pharmacological small-molecule compounds.
  20. mTOR substrate phosphorylation in growth control.
  21. A CRISPR view on autophagy.
  22. Protective autophagy by thymidine causes resistance to rapamycin in colorectal cancer cells in vitro.
  23. Vacuole fragmentation depends on a novel Atg18-containing retromer-complex.
  24. Retinoic acid increases the cellular cholesterol predominantly in a mTOR-independent manner.
  25. SIK2 Improving Mitochondrial Autophagy Restriction Induced by Cerebral Ischemia-Reperfusion in Rats.
  26. Glia-Neurons Cross-Talk Regulated Through Autophagy.
  27. HSc70 interactome reveal major role of macroautophagy and minor role of chaperone mediated autophagy in K-Ras G12V cell proliferation and survival.
  28. The disruption of human trophoblast functions by autophagy activation through PI3K/AKT/mTOR pathway induced by exposure to titanium carbide (Ti3C2) MXene.
  29. cPKCγ Deficiency Exacerbates Autophagy Impairment and Hyperphosphorylated Tau Buildup through the AMPK/mTOR Pathway in Mice with Type 1 Diabetes Mellitus.
  30. Ambra1 deficiency impairs mitophagy in skeletal muscle.
  31. Directing LRRK2 to membranes of the endolysosomal pathway triggers RAB phosphorylation and JIP4 recruitment.
  32. A novel ATG5 interaction with Ku70 potentiates DNA repair upon genotoxic stress.
  33. Pyrimidine-Based Fluorescent Probe for Monitoring Mitophagy via Detection of Mitochondrial pH Variation.
  34. The RALF1-FERONIA Complex Interacts with and Activates TOR Signaling in Response to Low Nutrients.
  35. Type I collagen reduces lipid accumulation during adipogenesis of preadipocytes 3T3-L1 via the YAP-mTOR-autophagy axis.
  36. Monitoring of ATG4 Protease Activity During Autophagy in the Model Microalga Chlamydomonas reinhardtii.
  37. SQSTM1 and its MAP1LC3B-binding domain induce forced mitophagy to degrade mitochondrial carryover during mitochondrial replacement therapy.
  38. Nuclear ingression of cytoplasmic bodies accompanies a boost in autophagy.
  39. Tumor suppressor p53 restrains cancer cell dissemination by modulating mitochondrial dynamics.
  40. Aspartate metabolism in endothelial cells activates the mTORC1 pathway to initiate translation during angiogenesis.
  41. PINK1/Parkin-mediated mitophagy in cardiovascular disease: From pathogenesis to novel therapy.
  42. Mitochondrial-derived vesicles: Recent insights.
  43. Lipid droplet degradation by autophagy connects mitochondria metabolism to Prox1-driven expression of lymphatic genes and lymphangiogenesis.
  44. Cx43-mediated sorting of miRNAs into extracellular vesicles.
  45. mTORC2 mediates structural plasticity in distal nociceptive endings that contributes to pain hypersensitivity following inflammation.
  46. Neuron-astrocyte transmitophagy is altered in Alzheimer's disease.
  47. Ispinesib as an Effective Warhead for the Design of Autophagosome-Tethering Chimeras: Discovery of Potent Degraders of Nicotinamide Phosphoribosyltransferase (NAMPT).
  48. Identification of a modulator of the actin cytoskeleton, mitochondria, nutrient metabolism and lifespan in yeast.
  49. Chaperone Mediated Autophagy Protects Cardiomyocytes Against Hypoxic-Cell Death.
  50. Heat shock protein A4 ablation leads to skeletal muscle myopathy associated with dysregulated autophagy and induced apoptosis.
  51. Integrating the effects of sucrose intake on the brain and white adipose tissue: Could autophagy be a possible link?
  1. J Biol Chem. 2022 May 13. pii: S0021-9258(22)00470-7. [Epub ahead of print] 102030
    Prolonged deprivation of arginine or leucine induces PI3K/Akt-dependent reactivation of mTORC1.
    Gwen R Buel, Huy Dang, John M Asara, John Blenis, Anders P Mutvei.
      The mechanistic target of rapamycin complex 1 (mTORC1) is a serine/threonine kinase complex that promotes anabolic processes including protein, lipid, and nucleotide synthesis, while suppressing catabolic processes such as macroautophagy. mTORC1 activity is regulated by growth factors and amino acids which signal through distinct but integrated molecular pathways: growth factors largely signal through the PI3K/Akt-dependent pathway, whereas the availabilities of amino acids leucine and arginine are communicated to mTORC1 by the Rag-GTPase pathway. While it is relatively well described how acute changes in leucine and arginine levels affect mTORC1 signaling, the effects of prolonged amino acid deprivation remain less well understood. Here, we demonstrate that prolonged deprivation of arginine and/or leucine leads to reactivation of mTORC1 activity, which reaches activation levels similar to those observed in nutrient-rich conditions. Surprisingly, we find that this reactivation is independent of the regeneration of amino acids by canonical autophagy or proteasomal degradation, but is dependent on PI3K/Akt signaling. Together, our data identify a novel crosstalk between the amino acid and PI3K/Akt signaling pathways upstream of mTORC1. These observations extend our understanding of the role of mTORC1 in growth-related diseases and indicate that dietary intervention by removal of leucine and/or arginine may be an ineffective therapeutic approach.
    DOI:  https://doi.org/10.1016/j.jbc.2022.102030
  2. Front Microbiol. 2022 ;13 889835
    Targeting Selective Autophagy as a Therapeutic Strategy for Viral Infectious Diseases.
    Yishan Liu, Tao Zhou, Jiajia Hu, Shouheng Jin, Jianfeng Wu, Xiangdong Guan, Yaoxing Wu, Jun Cui.
      Autophagy is an evolutionarily conserved lysosomal degradation system which can recycle multiple cytoplasmic components under both physiological and stressful conditions. Autophagy could be highly selective to deliver different cargoes or substrates, including protein aggregates, pathogenic proteins or superfluous organelles to lysosome using a series of cargo receptor proteins. During viral invasion, cargo receptors selectively target pathogenic components to autolysosome to defense against infection. However, viruses not only evolve different strategies to counteract and escape selective autophagy, but also utilize selective autophagy to restrict antiviral responses to expedite viral replication. Furthermore, several viruses could activate certain forms of selective autophagy, including mitophagy, lipophagy, aggrephagy, and ferritinophagy, for more effective infection and replication. The complicated relationship between selective autophagy and viral infection indicates that selective autophagy may provide potential therapeutic targets for human infectious diseases. In this review, we will summarize the recent progress on the interplay between selective autophagy and host antiviral defense, aiming to arouse the importance of modulating selective autophagy as future therapies toward viral infectious diseases.
    Keywords:  antiviral responses; macroautophagy (autophagy); selective autophagy; viral infectious diseases; virophagy
    DOI:  https://doi.org/10.3389/fmicb.2022.889835
  3. FEBS Lett. 2022 May 20.
    Crossing paths: Recent insights in the interplay between autophagy and intracellular trafficking in plants.
    Paul Gouguet, Suayb Üstün.
      Autophagy fulfils a crucial role in plant cellular homeostasis by recycling diverse cellular components ranging from protein complexes to whole organelles. Autophagy cargos are shuttled to the vacuole for degradation, thereby completing the recycling process. Canonical autophagy requires the lipidation and insertion of ATG8 proteins into double-membrane structures, termed autophagosomes, which engulf the cargo to be degraded. As such, the autophagy pathway actively contributes to intracellular membrane trafficking. Yet, the autophagic process is not fully considered a bona fide component of the canonical membrane trafficking pathway. However, recent findings have started to pinpoint the interconnection between classical membrane trafficking pathways and autophagy. This review details the latest advances in our comprehension of the interplay between these two pathways. Understanding the overlap between autophagy and canonical membrane trafficking pathways is important to illuminate the inner workings of both pathways in plant cells.
    Keywords:  Autophagy; amphisomes; endomembranes; lipids; non-canonical autophagy; secretory pathway; vacuolar degradation; vesicle trafficking
    DOI:  https://doi.org/10.1002/1873-3468.14404
  4. Front Cell Dev Biol. 2022 ;10 910640
    Phase Separation in Regulation of Autophagy.
    Yi Lu, Chunmei Chang.
      
    Keywords:  autophagosome formation; autophagy; autophagy substrate; cargo receptor; phase separation; selective autophagy
    DOI:  https://doi.org/10.3389/fcell.2022.910640
  5. Curr Biol. 2022 May 10. pii: S0960-9822(22)00671-6. [Epub ahead of print]
    Myotubularin-related phosphatase 5 is a critical determinant of autophagy in neurons.
    Jason P Chua, Karan Bedi, Michelle T Paulsen, Mats Ljungman, Elizabeth M H Tank, Erin S Kim, Jonathon P McBride, Jennifer M Colón-Mercado, Michael E Ward, Lois S Weisman, Sami J Barmada.
      Autophagy is a conserved, multi-step process of capturing proteolytic cargo in autophagosomes for lysosome degradation. The capacity to remove toxic proteins that accumulate in neurodegenerative disorders attests to the disease-modifying potential of the autophagy pathway. However, neurons respond only marginally to conventional methods for inducing autophagy, limiting efforts to develop therapeutic autophagy modulators for neurodegenerative diseases. The determinants underlying poor autophagy induction in neurons and the degree to which neurons and other cell types are differentially sensitive to autophagy stimuli are incompletely defined. Accordingly, we sampled nascent transcript synthesis and stabilities in fibroblasts, induced pluripotent stem cells (iPSCs), and iPSC-derived neurons (iNeurons), thereby uncovering a neuron-specific stability of transcripts encoding myotubularin-related phosphatase 5 (MTMR5). MTMR5 is an autophagy suppressor that acts with its binding partner, MTMR2, to dephosphorylate phosphoinositides critical for autophagy initiation and autophagosome maturation. We found that MTMR5 is necessary and sufficient to suppress autophagy in iNeurons and undifferentiated iPSCs. Using optical pulse labeling to visualize the turnover of endogenously encoded proteins in live cells, we observed that knockdown of MTMR5 or MTMR2, but not the unrelated phosphatase MTMR9, significantly enhances neuronal degradation of TDP-43, an autophagy substrate implicated in several neurodegenerative diseases. Our findings thus establish a regulatory mechanism of autophagy intrinsic to neurons and targetable for clearing disease-related proteins in a cell-type-specific manner. In so doing, our results not only unravel novel aspects of neuronal biology and proteostasis but also elucidate a strategy for modulating neuronal autophagy that could be of high therapeutic potential for multiple neurodegenerative diseases.
    Keywords:  RNA stability; TDP-43; autophagosome; iPSCs; induced pluripotent stem cells; macroautophagy; myotubularin; neuronal autophagy; optical pulse labeling; phosphoinositide
    DOI:  https://doi.org/10.1016/j.cub.2022.04.053
  6. Autophagy. 2022 May 16.
    N7-methylguanosine (m7G) tRNA modification: a novel autophagy modulator in cancer.
    Hui Han, Siyi Zheng, Shuibin Lin.
      Macroautophagy/autophagy, a physiological process that is involved in tumorigenesis, is regulated at genetic and epigenetic levels. Emerging reports suggest that aberrant RNA modifications cause dysregulated autophagy and affect tumorigenesis, while the role of RNA modifications in the regulation of autophagy in cancers remains unclear. In a recent study, we describe a new role for the tRNA m7G methyltransferase complex components METTL1 and WDR4 as negative regulators of MTORC1-mediated autophagy in esophageal squamous cell carcinoma (ESCC). METTL1 and WDR4 show abnormally high expression in ESCC tissues, and are associated with poor ESCC prognosis. Targeting METTL1 or WDR4 leads to decreased expression of m7G-modified tRNAs and reduces the translation of a subset of oncogenic transcripts, including the genes related to the MTOR signaling pathway and negative regulators of autophagy in an m7G-related codon-dependent manner, thereby resulting in hyperactivated MTORC1-mediated autophagy via dephosphorylation of ULK1 and finally causes cell death in ESCC. Our findings provide a new layer of translation regulation mechanism mediated by tRNA m7G modification, link translational machinery with autophagic machinery, and suggest that METTL1 and its downstream signaling axis could be potential therapeutic targets for ESCC treatment.
    Keywords:  Autophagy; METTL1; esophageal squamous cell carcinoma; m7G; tRNA modification
    DOI:  https://doi.org/10.1080/15548627.2022.2077551
  7. Front Cell Dev Biol. 2022 ;10 838402
    The Interplay Between Autophagy and RNA Homeostasis: Implications for Amyotrophic Lateral Sclerosis and Frontotemporal Dementia.
    O H Houghton, S Mizielinska, P Gomez-Suaga.
      Amyotrophic lateral sclerosis and frontotemporal dementia are neurodegenerative disorders that lie on a disease spectrum, sharing genetic causes and pathology, and both without effective therapeutics. Two pathways that have been shown to play major roles in disease pathogenesis are autophagy and RNA homeostasis. Intriguingly, there is an increasing body of evidence suggesting a critical interplay between these pathways. Autophagy is a multi-stage process for bulk and selective clearance of malfunctional cellular components, with many layers of regulation. Although the majority of autophagy research focuses on protein degradation, it can also mediate RNA catabolism. ALS/FTD-associated proteins are involved in many stages of autophagy and autophagy-mediated RNA degradation, particularly converging on the clearance of persistent pathological stress granules. In this review, we will summarise the progress in understanding the autophagy-RNA homeostasis interplay and how that knowledge contributes to our understanding of the pathobiology of ALS/FTD.
    Keywords:  C9orf72; RNA; RNA-binding proteins; amyotrophic lateral sclerosis; autophagy; frontotemporal dementia; granulophagy; stress granules
    DOI:  https://doi.org/10.3389/fcell.2022.838402
  8. Nat Commun. 2022 May 18. 13(1): 2735
    Autophagy regulation by RNA alternative splicing and implications in human diseases.
    Patricia González-Rodríguez, Daniel J Klionsky, Bertrand Joseph.
      Autophagy and RNA alternative splicing are two evolutionarily conserved processes involved in overlapping physiological and pathological processes. However, the extent of functional connection is not well defined. Here, we consider the role for alternative splicing and generation of autophagy-related gene isoforms in the regulation of autophagy in recent work. The impact of changes to the RNA alternative splicing machinery and production of alternative spliced isoforms on autophagy are reviewed with particular focus on disease relevance. The use of drugs targeting both alternative splicing and autophagy as well as the selective regulation of single autophagy-related protein isoforms, are considered as therapeutic strategies.
    DOI:  https://doi.org/10.1038/s41467-022-30433-1
  9. Cell Rep. 2022 May 17. pii: S2211-1247(22)00595-2. [Epub ahead of print]39(7): 110824
    The non-essential TSC complex component TBC1D7 restricts tissue mTORC1 signaling and brain and neuron growth.
    Sandra Schrötter, Christopher J Yuskaitis, Michael R MacArthur, Sarah J Mitchell, Aaron M Hosios, Maria Osipovich, Margaret E Torrence, James R Mitchell, Gerta Hoxhaj, Mustafa Sahin, Brendan D Manning.
      The tuberous sclerosis complex (TSC) 1 and 2 proteins associate with TBC1D7 to form the TSC complex, which is an essential suppressor of mTOR complex 1 (mTORC1), a ubiquitous driver of cell and tissue growth. Loss-of-function mutations in TSC1 or TSC2, but not TBC1D7, give rise to TSC, a pleiotropic disorder with aberrant activation of mTORC1 in various tissues. Here, we characterize mice with genetic deletion of Tbc1d7, which are viable with normal growth and development. Consistent with partial loss of function of the TSC complex, Tbc1d7 knockout (KO) mice display variable increases in tissue mTORC1 signaling with increased muscle fiber size but with strength and motor defects. Their most pronounced phenotype is brain overgrowth due to thickening of the cerebral cortex, with enhanced neuron-intrinsic mTORC1 signaling and growth. Thus, TBC1D7 is required for full TSC complex function in tissues, and the brain is particularly sensitive to its growth-suppressing activities.
    Keywords:  CP: Developmental biology; CP: Neuroscience; TBC1D7; Tsc1; Tsc2; brain; gait; growth; hamartin; mTOR; megalencephaly; mouse model; neurons; rapamycin; tuberin; tuberous sclerosis complex
    DOI:  https://doi.org/10.1016/j.celrep.2022.110824
  10. Biomed Pharmacother. 2022 May;pii: S0753-3322(22)00307-9. [Epub ahead of print]149 112918
    Are mitophagy enhancers therapeutic targets for Alzheimer's disease?
    Jangampalli Adi Pradeepkiran, Ashly Hindle, Sudhir Kshirsagar, P Hemachandra Reddy.
      Healthy mitochondria are essential for functional bioenergetics, calcium signaling, and balanced redox homeostasis. Dysfunctional mitochondria are a central aspect of aging and neurodegenerative diseases such as Alzheimer's disease (AD). The formation and accumulation of amyloid beta (Aβ) and hyperphosphorylated tau (P-tau) play large roles in the cellular changes seen in AD, including mitochondrial dysfunction, synaptic damage, neuronal loss, and defective mitophagy. Mitophagy is the cellular process whereby damaged mitochondria are selectively removed, and it plays an important role in mitochondrial quality control. Dysfunctional mitochondria are associated with increased reactive oxygen species and increased levels of Aβ, P-tau and Drp1, which together trigger mitophagy and autophagy. Impaired mitophagy causes the progressive accumulation of defective organelles and damaged mitochondria, and it has been hypothesized that the restoration of mitophagy may offer therapeutic benefits to AD patients. This review highlights the challenges of pharmacologically inducing mitophagy through two different signaling cascades: 1) The PINK1/parkin-dependent pathway and 2) the PINK1/parkin-independent pathway, with an emphasis on abnormal mitochondrial interactions with Aβ and P-Tau, which alter mitophagy in an age-dependent manner. This article also summarizes recent studies on the effects of mitophagy enhancers, including urolithin A, NAD+, actinonin, and tomatidine, on mutant APP/Aβ and mutant Tau. Findings from our lab have revealed that mitophagy enhancers can suppress APP/Aβ-induced and mutant Tau-induced mitochondrial and synaptic dysfunctions in mouse and cell line models of AD. Finally, we discuss the mechanisms underlying the beneficial health effects of mitophagy enhancers like urolithin A, NAD+, resveratrol and spermidine in AD.
    Keywords:  Alzheimer’s disease; Aβ; Mitochondrial dysfunction; Mitophagy; Phosphorylated tau
    DOI:  https://doi.org/10.1016/j.biopha.2022.112918
  11. ACS Appl Bio Mater. 2022 May 15.
    Piperazine-Based Mitochondria-Immobilized pH Fluorescent Probe for Imaging Endogenous ONOO- and Real-Time Tracking of Mitophagy.
    Minglu Li, Yue Huang, Shengmei Song, Shaomin Shuang, Chuan Dong.
      ONOO- is mainly produced in mitochondria, and dysfunctional and damaged mitochondria are degraded in lysosomes through autophagy, so it is important to synthesize a single probe for dual detection of ONOO- and mitophagy. Unfortunately, mitochondria-immobilized fluorescent probes for dual detection of ONOO- and mitophagy have not yet been developed. Hence, we first reported a piperazine-based mitochondria-immobilized red-emitting fluorescent probe (PMR), which not only can detect ONOO- but also could be used to image cellular mitophagy by the pH variations because of the protonation of the piperazine moiety. PMR was designed and prepared by introducing a piperazine ring as the pH response group, a lipophilic cation as the targeting mitochondria moiety, and benzyl chloride for immobilizing mitochondrial proteins through thiol groups. PMR displayed an enhanced fluorescence response at 640 nm through mitochondrial acidification. Using these advantages of PMR, which was successfully used for visualizing the mitophagy process induced by rapamycin or starvation, and chloroquine can inhibit rapamycin-induced mitophagy and prevent the fusion of autophagosomes and lysosomes. PMR also showed good sensitivity with a detection limit of 23 nM to ONOO-, which was successfully applied in imaging exogenous/endogenous ONOO-. Combining the above design, PMR may be used to study the detailed function of the mitophagy and ONOO--associated physiological and pathological processes.
    Keywords:  ONOO−; fluorescent probe; mitochondria-immobilized; mitophagy; pH
    DOI:  https://doi.org/10.1021/acsabm.2c00213
  12. Exp Neurol. 2022 May 11. pii: S0014-4886(22)00137-6. [Epub ahead of print] 114112
    Close association of polarization and LC3, a marker of autophagy, in axon determination in mouse hippocampal neurons.
    Naoki Segi, Tomoya Ozaki, Yuji Suzuki, Jun Ouchida, Shiro Imagama, Kenji Kadomatsu, Kazuma Sakamoto.
      The autophagy-lysosome pathway is a cellular clearance system for intracellular organelles, macromolecules and microorganisms. It is indispensable for cells not only to maintain their homeostasis but also to achieve more active cellular processes such as differentiation. Therefore, impairment or disruption of the autophagy-lysosome pathway leads to a wide spectrum of human diseases, ranging from several types of neurodegenerative diseases to malignancies. In elongating axons, autophagy preferentially occurs at growth cones, and disruption of autophagy is closely associated with incapacity for axonal regeneration after injury in the central nervous system. However, the roles of autophagy in developing neurons remain elusive. In particular, whether autophagy is involved in axon-dendrite determination is largely unclear. Using primary cultured mouse embryonic hippocampal neurons, we here showed the polarized distribution of autophagosomes among minor processes of neurons at stage 2. Time-lapse observation of neurons from GFP-LC3 transgenic mice demonstrated that an "LC3 surge"-i.e., a rapid accumulation of autophagic marker LC3 that continues for several hours in one minor process-proceeded the differentiation of neurons into axons. In addition, pharmacological activation and inhibition of autophagy by trehalose and bafilomycin, respectively, accelerated and delayed axonal determination. Taken together, our findings revealed the close association between LC3, a marker of autophagy, and axon determination in developing neurons.
    Keywords:  Autophagy; Axon determination; Trehalose
    DOI:  https://doi.org/10.1016/j.expneurol.2022.114112
  13. J Mater Chem B. 2022 May 18.
    A dual-response fluorescent probe for simultaneously monitoring polarity and ATP during autophagy.
    Wen-Li Jiang, Zhi-Qing Wang, Zhi-Ke Tan, Guo-Jiang Mao, Junjie Fei, Chun-Yan Li.
      Autophagy plays a vital role in maintaining intracellular homeostasis through a lysosome-dependent intracellular degradation pathway, which is closely related to the polarity and ATP. Herein, the first example of the dual-response fluorescent probe Lyso-NRB was reported for visualizing the fluctuation of polarity and ATP in lysosomes during autophagy. Probe Lyso-NRB is non-fluorescent. After the decrease of polarity, Lyso-NRB exhibits significant green emission due to the unique intramolecular charge transfer (ICT) effect. Upon the addition of ATP, the probe can react with ATP to rapidly open the spirocycle of rhodamine and a strong red emission can be observed. Moreover, Lyso-NRB exhibits a high sensitivity and selectivity toward polarity and ATP. Most importantly, the probe possesses a good lysosome-targeting ability and is used for the real-time monitoring of lysosome polarity and ATP fluctuations during H2O2 or starvation induced autophagy in living cells. Interestingly, it is found that that ATP deficiency can induce autophagy to increase lysosome polarity. Furthermore, the probe is applied for imaging the change of polarity and ATP under oxidative stress induced autophagy in zebrafish. Therefore, this work holds great potential for tracking the autophagy procedure by detecting the changes of lysosome polarity and ATP, which makes it a potentially powerful tool for understanding the roles of autophagy in diverse biological processes.
    DOI:  https://doi.org/10.1039/d2tb00575a
  14. Autophagy. 2022 May 17.
    Distinct designer diamines promote mitophagy, and thereby enhance healthspan in C. elegans and protect human cells against oxidative damage.
    Vijigisha Srivastava, Veronica Zelmanovich, Virendra Shukla, Rachel Abergel, Irit Cohen, Shmuel A Ben-Sasson, Einav Gross.
      Impaired mitophagy is a primary pathogenic event underlying diverse aging-associated diseases such as Alzheimer and Parkinson diseases and sarcopenia. Therefore, augmentation of mitophagy, the process by which defective mitochondria are removed, then replaced by new ones, is an emerging strategy for preventing the evolvement of multiple morbidities in the elderly population. Based on the scaffold of spermidine (Spd), a known mitophagy-promoting agent, we designed and tested a family of structurally related compounds. A prototypic member, 1,8-diaminooctane (VL-004), exceeds Spd in its ability to induce mitophagy and protect against oxidative stress. VL-004 activity is mediated by canonical aging genes and promotes lifespan and healthspan in C. elegans. Moreover, it enhances mitophagy and protects against oxidative injury in rodent and human cells. Initial structural characterization suggests simple rules for the design of compounds with improved bioactivity, opening the way for a new generation of agents with a potential to promote healthy aging.
    Keywords:  Aging; Caenorhabditis elegans; diamine; healthspan; lifespan; mitochondrial autophagy; mitophagy; oxidative stress; spermidine
    DOI:  https://doi.org/10.1080/15548627.2022.2078069
  15. Mov Disord. 2022 May 17.
    Can We Treat Neurodegenerative Proteinopathies by Enhancing Protein Degradation?
    Simone Engelender, Leonidas Stefanis, Salvatore Oddo, Arianna Bellucci.
      Neurodegenerative proteinopathies are defined as a class of neurodegenerative disorders, with either genetic or sporadic age-related onset, characterized by the pathological accumulation of aggregated protein deposits. These mainly include Alzheimer's disease (AD), Parkinson's disease (PD), amyotrophic lateral sclerosis (ALS), Huntington's disease (HD) as well as frontotemporal lobar degeneration (FTLD). The deposition of abnormal protein aggregates in the brain of patients affected by these disorders is thought to play a causative role in neuronal loss and disease progression. On that account, the idea of improving the clearance of pathological protein aggregates has taken hold as a potential therapeutic strategy. Among the possible approaches to pursue for reducing disease protein accumulation, there is the stimulation of the main protein degradation machineries of eukaryotic cells: the ubiquitin proteasomal system (UPS) and autophagy lysosomal pathway (ALP). Of note, several clinical trials testing the efficacy of either UPS- or ALP-active compounds are currently ongoing. Here, we discuss the main gaps and controversies emerging from experimental studies and clinical trials assessing the therapeutic efficacy of modulators of either the UPS or ALP in neurodegenerative proteinopathies, to gather whether they may constitute a real gateway from these disorders. © 2022 International Parkinson and Movement Disorder Society.
    Keywords:  autophagy lysosomal pathway; neurodegenerative proteinopathies; pathological protein aggregates; therapeutic strategies; ubiquitin proteasomal system
    DOI:  https://doi.org/10.1002/mds.29058
  16. Cell Res. 2022 May 19.
    Cytoskeleton grows p62 condensates for autophagy.
    Nobuo N Noda.
      
    DOI:  https://doi.org/10.1038/s41422-022-00671-5
  17. Neurobiol Dis. 2022 May 16. pii: S0969-9961(22)00160-7. [Epub ahead of print] 105768
    Understanding the contributions of VPS35 and the retromer in neurodegenerative disease.
    Erin T Williams, Xi Chen, P Anthony Otero, Darren J Moore.
      Perturbations of the endolysosomal pathway have been suggested to play an important role in the pathogenesis of several neurodegenerative diseases, including Parkinson's disease (PD) and Alzheimer's disease (AD). Specifically, VPS35 and the retromer complex play an important role in the endolysosomal system and are implicated in the pathophysiology of these diseases. A single missense mutation in VPS35, Asp620Asn (D620N), is known to cause late-onset, autosomal dominant familial PD. In this review, we focus on the emerging role of the PD-linked D620N mutation in causing retromer dysfunction and dissect its implications in neurodegeneration. Additionally, we will discuss how VPS35 and the retromer are linked to AD, amyotrophic lateral sclerosis, and primary tauopathies. Interestingly, reduced levels of VPS35 and other retromer components have been observed in post-mortem brain tissue, suggesting a role for the retromer in the pathophysiology of these diseases. This review will provide a comprehensive dive into the mechanisms of VPS35 dysfunction in neurodegenerative diseases. Furthermore, we will highlight outstanding questions in the field and the retromer as a therapeutic target for neurodegenerative disease at large.
    Keywords:  Alzheimer's disease; Amyotrophic lateral sclerosis; Autophagy; Cargo; Endosome; Golgi; LRRK2; Lysosome; Mitochondria; Parkinson's disease; Retromer complex; Tau; Tauopathies; VPS35; Vesicular sorting
    DOI:  https://doi.org/10.1016/j.nbd.2022.105768
  18. Mol Plant. 2022 May 18. pii: S1674-2052(22)00154-X. [Epub ahead of print]
    Autophagy targets Hd1 for vacuolar degradation to regulate rice flowering.
    Zhi Hu, Zhipan Yang, Yi Zhang, Aihong Zhang, Qingtao Lu, Ying Fang, Congming Lu.
      Flowering time (heading date) is a critical agronomic trait that determines the yield and regional adaptability of crops. Heading date 1 (Hd1) is a central regulator of photoperiodic flowering in rice (Oryza sativa). However, how the homeostasis of Hd1 protein is achieved is poorly understood. Here, we report that the nuclear autophagy pathway mediates Hd1 degradation in the dark to regulate flowering. Loss of autophagy function results in an accumulation of Hd1 and delays flowering in both short-day and long-day conditions. In the dark, nucleus-localized Hd1 is recognized as a substrate for autophagy and is subjected to vacuolar degradation via the autophagy protein OsATG8. The Hd1-OsATG8 interaction is required for autophagic degradation of Hd1 in the dark. Our study reveals a new mechanism by which Hd1 protein homeostasis is regulated by autophagy to control rice flowering. Our study also indicates that the regulation of flowering by autophagic degradation of Hd1 orthologs may have arisen over the course of mesangiosperm evolution, which would have increased their flexibility and adaptability to the environment by modulating flowering time.
    Keywords:  Autophagy; Hd1 homeostasis; heading date; rice
    DOI:  https://doi.org/10.1016/j.molp.2022.05.006
  19. Drug Discov Today. 2022 May 16. pii: S1359-6446(22)00200-8. [Epub ahead of print]
    Targeting autophagy in colorectal cancer: an update on pharmacological small-molecule compounds.
    Zixiang Li, Wen Si, Wenke Jin, Zhaoxin Yuan, Yi Chen, Leilei Fu.
      Autophagy, an evolutionarily highly conserved cellular degradation process, plays the Janus role (either cytoprotective or death-promoting) in colorectal cancer, so the targeting of several key autophagic pathways with small-molecule compounds may be a new therapeutic strategy. In this review, we discuss autophagy-associated cell death pathways and key cytoprotective autophagy pathways in colorectal cancer. Moreover, we summarize a series of small-molecule compounds that have the potential to modulate autophagy-associated cell death or cytoprotective autophagy for therapeutic purposes. Taken together, these findings demonstrate the Janus role of autophagy in colorectal cancer, and shed new light on the exploitation of a growing number of small-molecule compounds to target autophagy in future cancer drug discovery. Teaser: Autophagy plays both cytoprotective and death-promoting roles in colorectal cancer, so the targeting of some autophagic pathways with pharmacological small-molecule compounds is a potential therapeutic strategy.
    Keywords:  autophagy; autophagy-associated cell death; colorectal cancer; cytoprotective autophagy; small-molecule compound
    DOI:  https://doi.org/10.1016/j.drudis.2022.05.011
  20. Cell. 2022 May 12. pii: S0092-8674(22)00460-3. [Epub ahead of print]
    mTOR substrate phosphorylation in growth control.
    Stefania Battaglioni, Don Benjamin, Matthias Wälchli, Timm Maier, Michael N Hall.
      The target of rapamycin (TOR), discovered 30 years ago, is a highly conserved serine/threonine protein kinase that plays a central role in regulating cell growth and metabolism. It is activated by nutrients, growth factors, and cellular energy. TOR forms two structurally and functionally distinct complexes, TORC1 and TORC2. TOR signaling activates cell growth, defined as an increase in biomass, by stimulating anabolic metabolism while inhibiting catabolic processes. With emphasis on mammalian TOR (mTOR), we comprehensively reviewed the literature and identified all reported direct substrates. In the context of recent structural information, we discuss how mTORC1 and mTORC2, despite having a common catalytic subunit, phosphorylate distinct substrates. We conclude that the two complexes recruit different substrates to phosphorylate a common, minimal motif.
    DOI:  https://doi.org/10.1016/j.cell.2022.04.013
  21. Trends Cell Biol. 2022 May 14. pii: S0962-8924(22)00107-6. [Epub ahead of print]
    A CRISPR view on autophagy.
    Jin Rui Liang, Jacob E Corn.
      Autophagy is a fundamental pathway for the degradation of cytoplasmic content in response to pleiotropic extracellular and intracellular stimuli. Recent advances in the autophagy field have demonstrated that different organelles can also be specifically targeted for autophagy with broad implications on cellular and organismal health. This opens new dimensions in the autophagy field and more unanswered questions on the rationale and underlying mechanisms to degrade different organelles. Functional genomics via clustered regularly interspaced short palindromic repeats (CRISPR)-Cas9-based screening has gained popularity in the autophagy field to understand the common and unique factors that are implicated in the signaling, recognition, and execution of different cargo-specific autophagies. We focus on recent applications of CRISPR-based screens in the autophagy field, their discoveries, and the future directions of autophagy screens.
    Keywords:  CRISPR; autophagy; functional genomics; genome-wide screens; organellophagy
    DOI:  https://doi.org/10.1016/j.tcb.2022.04.006
  22. Cancer Drug Resist. 2021 ;4(3): 719-727
    Protective autophagy by thymidine causes resistance to rapamycin in colorectal cancer cells in vitro.
    I V Bijnsdorp, Godefridus J Peters.
      Aim: Thynidine phosphorylase (TP) acts as a proangiogenic growth factor which may regulate mammalian Target of Rapamycin (mTOR). We investigated whether the TP substrate thymidine and overexpression of TP affected mTOR signaling by comparing Colo320 (TP deficient) cells and its TP-transfected variant (Colo320TP1). Methods: Drug resistance was assessed with the sulforhodamine B assay, protein expression with Western blotting, cell cycle distribution and cell death with Fluorescence-activated cell sorting analysis, and autophagy with immunofluorescence. Results: Colo320 and Colo320TP1 cells had comparable levels of sensitivity to the mTOR inhibitor rapamycin. Thymidine treatment led to 13- and 50-fold resistance to rapamycin in Colo320 and Colo320TP1 cells, respectively. In Colo320TP1 cells, the thymidine phosphorylase inhibitor (TPI) reversed the thymidine induced resistance to rapamycin, but not in Colo320 cells, indicating a role for TP in the protection. Thymidine increased p70/S6k-phosphorylation (downstream of mTOR) in Colo320TP1, but it was not affected in Colo320. As a mechanism behind resistance, we studied the levels of autophagy and found that, in Colo320TP1 cells, autophagy was highly induced by thymidine-rapamycin, which was decreased by TPI. In addition, the autophagy inhibitor 3-methyl-adenine completely inhibited autophagy and its protection. Conclusion: Rapamycin resistance in TP-expressing cancer cells may therefore be related to thymidine-mediated autophagy activation.
    Keywords:  Thymidine phosphorylase; autophagy; mTOR; rapamycin; thymidine; thymidine phosphorylase inhibitor
    DOI:  https://doi.org/10.20517/cdr.2021.21
  23. Autophagy. 2022 May 15. 1-18
    Vacuole fragmentation depends on a novel Atg18-containing retromer-complex.
    Lisa Marquardt, Matthew Taylor, Florian Kramer, Kerstin Schmitt, Gerhard H Braus, Oliver Valerius, Michael Thumm.
      The yeast PROPPIN Atg18 folds as a β-propeller with two binding sites for phosphatidylinositol-3-phosphate (PtdIns3P) and PtdIns(3,5)P2 at its circumference. Membrane insertion of an amphipathic loop of Atg18 leads to membrane tubulation and fission. Atg18 has known functions at the PAS during macroautophagy, but the functional relevance of its endosomal and vacuolar pool is not well understood. Here we show in a proximity-dependent labeling approach and by co-immunoprecipitations that Atg18 interacts with Vps35, a central component of the retromer complex. The binding of Atg18 to Vps35 is competitive with the sorting nexin dimer Vps5 and Vps17. This suggests that Atg18 within the retromer can substitute for both the phosphoinositide binding and the membrane bending capabilities of these sorting nexins. Indeed, we found that Atg18-retromer is required for PtdIns(3,5)P2-dependent vacuolar fragmentation during hyperosmotic stress. The Atg18-retromer is further involved in the normal sorting of the integral membrane protein Atg9. However, PtdIns3P-dependent macroautophagy and the selective cytoplasm-to-vacuole targeting (Cvt) pathway are only partially affected by the Atg18-retromer. We expect that this is due to the plasticity of the different sorting pathways within the endovacuolar system.Abbreviations:BAR: bin/amphiphysin/Rvs; FOA: 5-fluoroorotic acid; PAS: phagophore assembly site; PROPPIN: beta-propeller that binds phosphoinositides; PtdIns3P: phosphatidylinositol-3-phosphate; PX: phox homology.
    Keywords:  Atg18; Atg9; PROPPIN; Vps35; retrograde transport; retromer; vacuolar fragmentation
    DOI:  https://doi.org/10.1080/15548627.2022.2072656
  24. Immunol Res. 2022 May 18.
    Retinoic acid increases the cellular cholesterol predominantly in a mTOR-independent manner.
    Satya Prakash, Ambak Kumar Rai.
      Retinoic acid (RA) plays a role in the mounting immune response and controls several functions of the human body, including cholesterol homeostasis. The synthesis, uptake, and efflux of cellular cholesterol are significantly linked to the mammalian target of rapamycin complex-1 (mTORC1). Activation of mTORC1 promotes the synthesis and uptake of the cholesterol and suppresses its efflux, thus causing accumulation of cellular cholesterol. It is intriguing to know the effect of a high dose of RA on cholesterol accumulation in macrophages (mφ) and whether it is via mTOR activation. It is important to note that the long-term treatment of RA in humans is safe. Therefore, we chose a high dose of RA to observe its effect, which may be implicated in diseases like visceral leishmaniasis, where cholesterol deficiency is established. In the present study, we found the increased expression of RAPTOR, a regulatory component of the mTORC1 complex, in mφ upon treatment with RA. We observed the increased expression of SREBP2, LDLR, and PCSK9 in RA-treated mφ under sufficient cholesterol conditions, which further increased cellular cholesterol levels. Notably, their expressions were decreased when the mTOR pathway was inhibited by rapamycin. However, treatment with rapamycin did not result in the loss of cellular cholesterol in RA-treated mφ. Comparison with rapamycin-treated mφ suggests that RA induces cellular cholesterol levels in a mTORC1-independent manner.
    Keywords:  Cholesterol; Macrophage; Retinoic acid (RA); mTOR
    DOI:  https://doi.org/10.1007/s12026-022-09292-x
  25. Front Pharmacol. 2022 ;13 683898
    SIK2 Improving Mitochondrial Autophagy Restriction Induced by Cerebral Ischemia-Reperfusion in Rats.
    Ran Zhang, Yun Liu, Wenhua Zhong, Zebo Hu, Chao Wu, Mengyao Ma, Yi Zhang, Xiangyun He, Lin Wang, Shu Li, Yun Hong.
      Previous studies have shown that Salt-induced kinase-2(SIK2) is involved in the regulation of various energy-metabolism-related reactions, and it also can regulate angiogenesis after cerebral ischemia-reperfusion. However, it is unclear whether SIK2 can regulate energy metabolism in cerebral ischemia-reperfusion injury. As mitochondria plays an important role in energy metabolism, whether SIK2 regulates energy metabolism through affecting mitochondrial changes is also worth to be explored. In this study, rats were treated with adeno-associated virus-SIK2-Green fluorescent protein (AAV-SIK2-GFP) for the overexpression of SIK2 before middle cerebral artery occlusion (MCAO). We found that SIK2 overexpression could alleviate the neuronal damage, reduce the area of cerebral infarction, and increase the adenosine triphosphate (ATP) content, which could promote the expression of phosphorylated-mammalian target of rapamycin-1 (p-mTORC1), hypoxia-inducible factor-1α (HIF-1α), phosphatase and tensin homologue-induced putative kinase 1 (PINK1) and E3 ubiquitinligating enzyme (Parkin). Transmission electron microscopy revealed that SIK2 overexpression enhanced mitochondrial autophagy. It is concluded that SIK2 can ameliorate neuronal injury and promote the energy metabolism by regulating the mTOR pathway during cerebral ischemia-reperfusion, and this process is related to mitochondrial autophagy.
    Keywords:  MCAO; SIK2; cerebral ischemia-reperfusion; energy metabolism; mitochondrial autophagy
    DOI:  https://doi.org/10.3389/fphar.2022.683898
  26. Front Physiol. 2022 ;13 886273
    Glia-Neurons Cross-Talk Regulated Through Autophagy.
    Milena Damulewicz, Kornel Szypulski, Elzbieta Pyza.
      Autophagy is a self-degradative process which plays a role in removing misfolded or aggregated proteins, clearing damaged organelles, but also in changes of cell membrane size and shape. The aim of this phenomenon is to deliver cytoplasmic cargo to the lysosome through the intermediary of a double membrane-bound vesicle (autophagosome), that fuses with a lysosome to form autolysosome, where cargo is degraded by proteases. Products of degradation are transported back to the cytoplasm, where they can be re-used. In the present study we showed that autophagy is important for proper functioning of the glia and that it is involved in the regulation of circadian structural changes in processes of the pacemaker neurons. This effect is mainly observed in astrocyte-like glia, which play a role of peripheral circadian oscillators in the Drosophila brain.
    Keywords:  Drosophila; autophagy; circadian clock; neuronal plasticity; sleep
    DOI:  https://doi.org/10.3389/fphys.2022.886273
  27. J Proteomics. 2022 May 17. pii: S1874-3919(22)00138-5. [Epub ahead of print] 104614
    HSc70 interactome reveal major role of macroautophagy and minor role of chaperone mediated autophagy in K-Ras G12V cell proliferation and survival.
    Vipin Kumar Yadav, Poorwa Awasthi, Rachna Behl, Amit Kumar.
      Constitutively active K-Ras oncogene mutation at G12V changes the proteome of cells and activates macroautophagy for cell advantage. Inhibition of macroautophagy impairs K-Ras mediated tumor progression to a limited extent with increase of spontaneous tumors due to poorly understood mechanisms. Here, we show that inhibition of macroautophagy in K-Ras G12V mouse embryonic fibroblasts (MEFs) hyper activates chaperon mediated autophagy (CMA). Quantitative identification of CMA substrates through co-immunoprecipitation of CMA component heat shock cognate 70 (Hsc70) demonstrates a shift of proteins from macroautophagy to CMA mediated degradation. However, macroautophagy impairment show significant inhibition on proliferation and CMA hyper activation provides a basal support to macroautophagy-inhibited MEFs for survival. On the other hand, K-Ras G12V MEFs impaired of CMA reduces number of Hsc70 clients but activated macroautophagy significantly compensated CMA loss. Nonetheless, co-inhibition of CMA and macroautophagy had a synergistic detrimental effect on both proliferation and survival of MEFs expressing K-Ras G12V mutant. Our results point to K-Ras G12V MEFs dependency on macroautophagy and CMA partly compensates its loss for survival but not hyper-proliferation; implicating that targeting both macroautophagy and CMA as a promising therapeutic target in G12V mutation associated K-Ras cancers. SIGNIFICANCE: The present study provides a framework of Hsc70 interacting proteins, which differentially interact with Hsc70 in response to autophagy alterations. The role of proteins accumulation and induced proteo-toxicity could be underlying factor in macroautophagy and CMA co-inhibited K-Ras G12V MEFs phenotype. Our study provides rational for adaptive mechanisms in K-Ras tumors inhibited with different autophagy pathways and also supports targeting both macroautophagy and CMA simultaneously as therapeutic target. At the same time current study will help in characterizing the underlying cellular processes that may play a role in escaping tutor suppressor role CMA and macroautophagy in cancers harboring K-Ras G12V mutation that may be further utilized to identify molecular targets for K-Ras-driven cancers.
    Keywords:  CMA; Cell prolifertion; Immunoprecipitation; K-ras; Macro-autophagy; Proliferation
    DOI:  https://doi.org/10.1016/j.jprot.2022.104614
  28. Food Chem Toxicol. 2022 May 12. pii: S0278-6915(22)00326-X. [Epub ahead of print] 113128
    The disruption of human trophoblast functions by autophagy activation through PI3K/AKT/mTOR pathway induced by exposure to titanium carbide (Ti3C2) MXene.
    Limei Yang, Le Hu, Hongyu Tang, Xuemei Chen, Xueqing Liu, Yue Zhang, Yixian Wen, Yongxiu Yang, Yanqing Geng.
      Ti3C2 MXene, as a novel nanomaterial, has attracted great attention due to its promising properties in biomedical applications. However, the potential effects of Ti3C2 MXene on trophoblast functions have not been investigated. Here, we found that Ti3C2 MXene exposure weakened the extension ability of villus explants in vitro. We employed human trophoblast HTR-8/SVneo cells to reveal the underlying molecular mechanisms by which Ti3C2 MXene exposure affected trophoblast functions. Results showed that Ti3C2 MXene entered cells and mostly deposited in the cytoplasm, inhibiting cell migration and invasion abilities. Furthermore, we found that Ti3C2 MXene exposure elevated autophagy through the inhibition of the PI3K/AKT/mTOR pathway. Meanwhile, the application of an autophagy inhibitor (3-MA) prevented autophagy and restored cell viability, resulting in the recovery of cell migration and invasion abilities. These indicated that the cellular dysfunction induced by Ti3C2 MXene may be mediated by autophagy activation. Our results indicated that autophagy is a key factor in eliciting HTR-8/SVneo dysfunction after Ti3C2 MXene exposure, which could therefore damage placental development. Autophagy inhibition is a potential therapeutic strategy for alleviating the placental toxicity of nanoparticles.
    Keywords:  Autophagy; HTR-8/SVneo; Ti(3)C(2) MXene; Trophoblast invasion; Trophoblast migration
    DOI:  https://doi.org/10.1016/j.fct.2022.113128
  29. Neurosci Bull. 2022 May 21.
    cPKCγ Deficiency Exacerbates Autophagy Impairment and Hyperphosphorylated Tau Buildup through the AMPK/mTOR Pathway in Mice with Type 1 Diabetes Mellitus.
    Jiayin Zheng, Yue Wang, Yue Liu, Song Han, Ying Zhang, Yanlin Luo, Yi Yan, Junfa Li, Li Zhao.
      Type 1 diabetes mellitus (T1DM)-induced cognitive dysfunction is common, but its underlying mechanisms are still poorly understood. In this study, we found that knockout of conventional protein kinase C (cPKC)γ significantly increased the phosphorylation of Tau at Ser214 and neurofibrillary tangles, but did not affect the activities of GSK-3β and PP2A in the hippocampal neurons of T1DM mice. cPKCγ deficiency significantly decreased the level of autophagy in the hippocampal neurons of T1DM mice. Activation of autophagy greatly alleviated the cognitive impairment induced by cPKCγ deficiency in T1DM mice. Moreover, cPKCγ deficiency reduced the AMPK phosphorylation levels and increased the phosphorylation levels of mTOR in vivo and in vitro. The high glucose-induced Tau phosphorylation at Ser214 was further increased by the autophagy inhibitor and was significantly decreased by an mTOR inhibitor. In conclusion, these results indicated that cPKCγ promotes autophagy through the AMPK/mTOR signaling pathway, thus reducing the level of phosphorylated Tau at Ser214 and neurofibrillary tangles.
    Keywords:  AMPK/mTOR signaling pathway; Autophagy; Conventional protein kinase C (cPKC)γ; Phosphorylated Tau; Tau
    DOI:  https://doi.org/10.1007/s12264-022-00863-4
  30. J Cachexia Sarcopenia Muscle. 2022 May 20.
    Ambra1 deficiency impairs mitophagy in skeletal muscle.
    Lisa Gambarotto, Samuele Metti, Martina Chrisam, Cristina Cerqua, Patrizia Sabatelli, Andrea Armani, Carlo Zanon, Marianna Spizzotin, Silvia Castagnaro, Flavie Strappazzon, Paolo Grumati, Matilde Cescon, Paola Braghetta, Eva Trevisson, Francesco Cecconi, Paolo Bonaldo.
      BACKGROUND: Maintaining healthy mitochondria is mandatory for muscle viability and function. An essential surveillance mechanism targeting defective and harmful mitochondria to degradation is the selective form of autophagy called mitophagy. Ambra1 is a multifaceted protein with well-known autophagic and mitophagic functions. However, the study of its role in adult tissues has been extremely limited due to the embryonic lethality caused by full-body Ambra1 deficiency.METHODS: To establish the role of Ambra1 as a positive regulator of mitophagy, we exploited in vivo overexpression of a mitochondria-targeted form of Ambra1 in skeletal muscle. To dissect the consequence of Ambra1 inactivation in skeletal muscle, we generated muscle-specific Ambra1 knockout (Ambra1fl/fl :Mlc1f-Cre) mice. Mitochondria-enriched fractions were obtained from muscles of fed and starved animals to investigate the dynamics of the mitophagic flux.
    RESULTS: Our data show that Ambra1 has a critical role in the mitophagic flux of adult murine skeletal muscle and that its genetic inactivation leads to mitochondria alterations and myofibre remodelling. Ambra1 overexpression in wild-type muscles is sufficient to enhance mitochondria clearance through the autophagy-lysosome system. Consistently with this, Ambra1-deficient muscles display an abnormal accumulation of the mitochondrial marker TOMM20 by +76% (n = 6-7; P < 0.05), a higher presence of myofibres with swollen mitochondria by +173% (n = 4; P < 0.05), and an alteration in the maintenance of the mitochondrial membrane potential and a 34% reduction in the mitochondrial respiratory complex I activity (n = 4; P < 0.05). Lack of Ambra1 in skeletal muscle leads to impaired mitophagic flux, without affecting the bulk autophagic process. This is due to a significantly decreased recruitment of DRP1 (n = 6-7 mice; P < 0.01) and Parkin (n = 6-7 mice; P < 0.05) to the mitochondrial compartment, when compared with controls. Ambra1-deficient muscles also show a marked dysregulation of the endolysosome compartment, as the incidence of myofibres with lysosomal accumulation is 20 times higher than wild-type muscles (n = 4; P < 0.05). Histologically, Ambra1-deficient muscles of both 3- and 6-month-old animals display a significant decrease of myofibre cross-sectional area and a 52% reduction in oxidative fibres (n = 6-7; P < 0.05), thus highlighting a role for Ambra1 in the proper structure and activity of skeletal muscle.
    CONCLUSIONS: Our study indicates that Ambra1 is critical for skeletal muscle mitophagy and for the proper maintenance of functional mitochondria.
    Keywords:  Ambra1; Mitochondria; Mitophagy; Skeletal muscle
    DOI:  https://doi.org/10.1002/jcsm.13010
  31. Neurobiol Dis. 2022 May 14. pii: S0969-9961(22)00161-9. [Epub ahead of print] 105769
    Directing LRRK2 to membranes of the endolysosomal pathway triggers RAB phosphorylation and JIP4 recruitment.
    Jillian H Kluss, Luis Bonet-Ponce, Patrick A Lewis, Mark R Cookson.
      Coding mutations in the Leucine-rich repeat kinase 2 (LRRK2) gene, which are associated with dominantly inherited Parkinson's disease (PD), lead to an increased activity of the encoded LRRK2 protein kinase. As such, kinase inhibitors are being considered as therapeutic agents for PD. It is therefore of interest to understand the mechanism(s) by which LRRK2 is activated during cellular signaling. Lysosomal membrane damage represents one way of activating LRRK2 and leads to phosphorylation of downstream RAB substrates and recruitment of the motor adaptor protein JIP4. However, it is unclear whether the activation of LRRK2 would be seen at other membranes of the endolysosomal system, where LRRK2 has also shown to be localized, or whether these signaling events can be induced without membrane damage. Here, we use a rapamycin-dependent oligomerization system to direct LRRK2 to various endomembranes including the Golgi apparatus, lysosomes, the plasma membrane, recycling, early, and late endosomes. Irrespective of membrane location, the recruitment of LRRK2 to membranes results in local accumulation of phosphorylated RAB10, RAB12, and JIP4. We also show that endogenous RAB29, previously nominated as an activator of LRRK2 based on overexpression, is not required for activation of LRRK2 at the Golgi nor lysosome. We therefore conclude that LRRK2 signaling to RAB10, RAB12, and JIP4 can be activated once LRRK2 is accumulated at any cellular organelle along the endolysosomal pathway.
    Keywords:  Endolysosomal membranes; LRRK2; Parkinson's disease
    DOI:  https://doi.org/10.1016/j.nbd.2022.105769
  32. Sci Rep. 2022 May 17. 12(1): 8134
    A novel ATG5 interaction with Ku70 potentiates DNA repair upon genotoxic stress.
    Sinem Demirbag-Sarikaya, Yunus Akkoc, Sıla Turgut, Secil Erbil-Bilir, Nur Mehpare Kocaturk, Joern Dengjel, Devrim Gozuacik.
      The maintenance of cellular homeostasis in living organisms requires a balance between anabolic and catabolic reactions. Macroautophagy (autophagy herein) is determined as one of the major catabolic reactions. Autophagy is an evolutionarily conserved stress response pathway that is activated by various insults including DNA damage. All sorts of damage to DNA potentially cause loss of genetic information and trigger genomic instability. Most of these lesions are repaired by the activation of DNA damage response following DNA repair mechanisms. Here we describe, a novel protein complex containing the autophagy protein ATG5 and the non-homologous end-joining repair system proteins. We discovered for the first time that ATG5 interacted with both Ku80 (XRCC5) and Ku70 (XRCC6). This novel interaction is facilitated mainly via Ku70. Our results suggest that this interaction is dynamic and enhanced upon genotoxic stresses. Strikingly, we identified that ATG5-Ku70 interaction is necessary for DNA repair and effective recovery from genotoxic stress. Therefore, our results are demonstrating a novel, direct, dynamic, and functional interaction between ATG5 and Ku70 proteins that plays a crucial role in DNA repair under genotoxic stress conditions.
    DOI:  https://doi.org/10.1038/s41598-022-11704-9
  33. Chembiochem. 2022 May 16.
    Pyrimidine-Based Fluorescent Probe for Monitoring Mitophagy via Detection of Mitochondrial pH Variation.
    Jia Zhou, Wei Du, Tao Shao, Zheng Li, Duoteng Zhang, Lan Wang, Zhijie Fang, Jie Li, Qiong Wu, Chengwu Zhang, Lin Li.
      Impaired mitophagy hinders the clearance of damaged mitochondria, inducing pathological states. Knowledge of this phenomenon is key to diagnose certain diseases and understand their pathogenesis. Mitophagy involves an acidization process that could sever as an ideal detection target. In this work, we designed and synthesized a mitochondrial-targeting fluorescence probe, Z2, for evaluating pH variation. This probe exhibited remarkable "turn-on" fluorescence under acidic condition. In biological applications, Z2 showed a strong, specific pH detection capacity in Parkin-overexpressing HeLa cells during the mitophagy process. The "turn-on" fluorescence property of Z2 is also used to detect pH variation in Caenorhabditis elegans (C. elegans). This probe, as the novel pH assessment tool, may facilitate further research of mitophagy-associated pathological patterns.
    Keywords:  cell imaging; fluorescent probe; in vivo imaging; mitophagy; pH detection
    DOI:  https://doi.org/10.1002/cbic.202200217
  34. Mol Plant. 2022 May 17. pii: S1674-2052(22)00152-6. [Epub ahead of print]
    The RALF1-FERONIA Complex Interacts with and Activates TOR Signaling in Response to Low Nutrients.
    Limei Song, Guoyun Xu, Tingting Li, Huina Zhou, Qinlu Lin, Jia Chen, Long Wang, Dousheng Wu, Xiaoxu Li, Lifeng Wang, Sirui Zhu, Feng Yu.
      Target of rapamycin (TOR) kinase is an evolutionarily conserved major regulator of nutrient metabolism and organismal growth in eukaryotes. In plants, nutrients are remobilized and reallocated between shoots and roots under low nutrient conditions, and nitrogen and nitrogen-related nutrients (e.g., amino acids) are key upstream signals leading to TOR activation in shoots under low nutrient conditions. However, how these forms of nitrogen can be sensed to activate TOR in plants is still poorly understood. Here, we found that the plant receptor kinase FERONIA (FER) interacts with the TOR pathway to regulate nutrient (nitrogen and amino acid) signaling under low nutrient conditions, and exerts similar metabolic effects in response to nitrogen-deficiency in Arabidopsis. FER and its partner RIPK kinase interact with the TOR/RAPTOR complex to positively modulate TOR signaling activity. In this process, the receptor complex FER/RIPK can phosphorylate the TOR complex component RAPTOR1B. The RALF1 peptide, a ligand of the FER/RIPK receptor complex, increases TOR activation in the young leaf by enhancing FER-TOR interactions, leading to promotion of true leaf growth in Arabidopsis under low nutrient conditions. These data indicate plant prioritization of nutritional stress over RALF1-mediated inhibition of cell growth under low nutrient conditions. In addition, specific amino acids (e.g., Gln, Asp, and Gly) promote true leaf growth under nitrogen-deficient conditions via the FER-TOR axis. Our study reveals a mechanism by which the RALF1-FER pathway activates TOR in the plant adaptive response to low nutrients.
    Keywords:  FERONIA; TOR; amino acid response; energy metabolism; nitrogen response
    DOI:  https://doi.org/10.1016/j.molp.2022.05.004
  35. Biochim Biophys Acta Mol Cell Biol Lipids. 2022 May 17. pii: S1388-1981(22)00071-3. [Epub ahead of print] 159181
    Type I collagen reduces lipid accumulation during adipogenesis of preadipocytes 3T3-L1 via the YAP-mTOR-autophagy axis.
    Yanfang Gao, Ma Kai, Yu Kang, Weiwei Liu, Xiaoling Liu, Xinyu Long, Toshihiko Hayashi, Shunji Hattori, Kazunori Mizuno, Hitomi Fujisaki, Takashi Ikejima.
      The extracellular matrix (ECM) regulates cell behavior through signal transduction and provides a suitable place for cell survival. As one of the major components of the extracellular matrix, type I collagen is involved in regulating cell migration, proliferation and differentiation. We present a system in which 3T3-L1 preadipocyte cells are induced for adipogenic differentiation on type I collagen coated dishes. Our previous study has found that type I collagen inhibits adipogenic differentiation via YAP activation. Here we further reveal that type I collagen inactivates autophagy by up-regulating mTOR activity via the YAP pathway. Under collagen-coating conditions, co-localization of lysosomes with mTOR was increased and the level of downstream protein p-S6K was elevated, accompanied by a decrease in the level of autophagy. Autophagy is negatively correlated with adipogenesis under type I collagen coating. Through the YAP-autophagy axis, type I collagen improves glycolipid metabolism accompanied by increased mitochondrial content, enhanced glucose uptake, reduced release of free fatty acids (FFAs) and decreased intracellular lipid accumulation. Our findings provide insight into the strategy for dealing with obesity: Type I collagen or the drugs with inhibitory effects on autophagy or YAP, have a potential to accelerate the energy metabolism of adipose tissue, so as to better maintain the homeostasis of glucose and lipids in the body, which can be used for achieving weight loss.
    Keywords:  Autophagy; Glycolipid metabolism; Type I collagen; YAP; mTOR
    DOI:  https://doi.org/10.1016/j.bbalip.2022.159181
  36. Methods Mol Biol. 2022 ;2447 205-220
    Monitoring of ATG4 Protease Activity During Autophagy in the Model Microalga Chlamydomonas reinhardtii.
    José L Crespo, M Esther Pérez-Pérez.
      Deciphering the molecular mechanisms underlying the regulation of the ATG4 protease is essential to understand the regulation of ATG8 lipidation, a key step in the biogenesis of the autophagosome and hence in autophagy progression. Here, we describe two complementary approaches to monitor ATG4 proteolytic activity in the model green alga Chlamydomonas reinhardtii: an in vitro assay using recombinant ATG4 and recombinant ATG8 as substrate, and a cell-free assay using soluble total protein extract from Chlamydomonas and recombinant Chlamydomonas ATG8 as substrate. Both assays are followed by non-reducing SDS-PAGE and immuno-blot analysis. Given the high evolutionary conservation of the ATG8 maturation process, these assays have also been validated to monitor ATG4 activity in yeast using Chlamydomonas ATG8 as substrate.
    Keywords:  ATG4; Autophagy; Chlamydomonas; Microalgae; Non-reducing SDS-PAGE; Protease; Western-blot
    DOI:  https://doi.org/10.1007/978-1-0716-2079-3_17
  37. Autophagy. 2022 May 19. 1-2
    SQSTM1 and its MAP1LC3B-binding domain induce forced mitophagy to degrade mitochondrial carryover during mitochondrial replacement therapy.
    Xiao-Yan Fan, Shen Yin, Shi-Ming Luo.
      Mitophagy is a process that selectively degrades mitochondria in cells, and it involves a series of signaling events. Our recent paper shows that the ectopic expression of SQSTM1 and its MAP1LC3B-binding domain (Binding) at the mitochondrial outer membrane, can directly cause mitophagy. To distinguish this mitophagy from others, we called it forced mitophagy. Further results show that the forced mitophagy can degrade half of the mitochondria and their DNA in HeLa cells and mouse embryos. Meanwhile, there are no apparent effects on mitochondrial membrane potential (MMP), reactive oxygen species (ROS), mitosis and embryo development. Thus, the forced mitophagy was examined to selectively degrade mitochondrial carryover in the nuclear donor embryos' mitochondria by pre-labeling with Binding before mitochondrial replacement therapy (MRT). The results show that the forced mitophagy can reduce mitochondrial carryover from an average of 4% to 0.09% compared to the controls in mouse embryos and tissues. In addition, the offspring from MRT mice show negligible effects on growth, reproduction, exercise and behavior. Furthermore, results from human tri-pronuclear embryos show that the forced mitophagy results in undetectable mitochondrial carryover in 77% of embryos following MRT. Therefore, forced mitophagy is efficient and safe for degrading mitochondrial carryover in MRT.
    Keywords:  Forced mitophagy; NIX; SQSTM1; mitochondrial carryover; mitochondrial replacement therapy
    DOI:  https://doi.org/10.1080/15548627.2022.2077552
  38. Life Sci Alliance. 2022 Sep;pii: e202101160. [Epub ahead of print]5(9):
    Nuclear ingression of cytoplasmic bodies accompanies a boost in autophagy.
    Manon Garcia, Sylvain Kumanski, Alberto Elías-Villalobos, Chantal Cazevieille, Caroline Soulet, María Moriel-Carretero.
      Membrane contact sites are functional nodes at which organelles reorganize metabolic pathways and adapt to changing cues. In Saccharomyces cerevisiae, the nuclear envelope subdomain surrounding the nucleolus, very plastic and prone to expansion, can establish contacts with the vacuole and be remodeled in response to various metabolic stresses. While using genotoxins with unrelated purposes, we serendipitously discovered a fully new remodeling event at this nuclear subdomain: the nuclear envelope partitions into its regular contact with the vacuole and a dramatic internalization within the nucleus. This leads to the nuclear engulfment of a globular, cytoplasmic portion. In spite of how we discovered it, the phenomenon is likely DNA damage-independent. We define lipids supporting negative curvature, such as phosphatidic acid and sterols, as bona fide drivers of this event. Mechanistically, we suggest that the engulfment of the cytoplasm triggers a suction phenomenon that enhances the docking of proton pump-containing vesicles with the vacuolar membrane, which we show matches a boost in autophagy. Thus, our findings unveil an unprecedented remodeling of the nucleolus-surrounding membranes with impact on metabolic adaptation.
    DOI:  https://doi.org/10.26508/lsa.202101160
  39. Oncogenesis. 2022 May 19. 11(1): 26
    Tumor suppressor p53 restrains cancer cell dissemination by modulating mitochondrial dynamics.
    Trinh T T Phan, Yu-Chun Lin, Yu-Ting Chou, Chien-Wei Wu, Lih-Yuan Lin.
      Tumor suppressor p53 plays a central role in preventing tumorigenesis. Here, we unravel how p53 modulates mitochondrial dynamics to restrain the metastatic properties of cancer cells. p53 inhibits the mammalian target of rapamycin complex 1 (mTORC1) signaling to attenuate the protein level of mitochondrial fission process 1 (MTFP1), which fosters the pro-fission dynamin-related protein 1 (Drp1) phosphorylation. This regulatory mechanism allows p53 to restrict cell migration and invasion governed by Drp1-mediated mitochondrial fission. Downregulating p53 expression or elevating the molecular signature of mitochondrial fission correlates with aggressive tumor phenotypes and poor prognosis in cancer patients. Upon p53 loss, exaggerated mitochondrial fragmentation stimulates the activation of the extracellular signal-regulated kinase 1/2 (ERK1/2) signaling resulting in epithelial-to-mesenchymal transition (EMT)-like changes in cell morphology, accompanied by accelerated matrix metalloproteinase 9 (MMP9) expression and invasive cell migration. Notably, blocking the activation of mTORC1/MTFP1/Drp1/ERK1/2 axis completely abolishes the p53 deficiency-driven cellular morphological switch, MMP9 expression, and cancer cell dissemination. Our findings unveil a hitherto unrecognized mitochondria-dependent molecular mechanism underlying the metastatic phenotypes of p53-compromised cancers.
    DOI:  https://doi.org/10.1038/s41389-022-00401-x
  40. Dev Cell. 2022 May 11. pii: S1534-5807(22)00286-6. [Epub ahead of print]
    Aspartate metabolism in endothelial cells activates the mTORC1 pathway to initiate translation during angiogenesis.
    Roxana E Oberkersch, Giovanna Pontarin, Matteo Astone, Marianna Spizzotin, Liaisan Arslanbaeva, Giovanni Tosi, Emiliano Panieri, Sara Ricciardi, Maria Francesca Allega, Alessia Brossa, Paolo Grumati, Benedetta Bussolati, Stefano Biffo, Saverio Tardito, Massimo M Santoro.
      Angiogenesis, the active formation of new blood vessels from pre-existing ones, is a complex and demanding biological process that plays an important role in physiological as well as pathological settings. Recent evidence supports cell metabolism as a critical regulator of angiogenesis. However, whether and how cell metabolism regulates endothelial growth factor receptor levels and nucleotide synthesis remains elusive. We here shown in both human cell lines and mouse models that during developmental and pathological angiogenesis, endothelial cells (ECs) use glutaminolysis-derived glutamate to produce aspartate (Asp) via aspartate aminotransferase (AST/GOT). Asp leads to mTORC1 activation which, in turn, regulates endothelial translation machinery for VEGFR2 and FGFR1 synthesis. Asp-dependent mTORC1 pathway activation also regulates de novo pyrimidine synthesis in angiogenic ECs. These findings identify glutaminolysis-derived Asp as a regulator of mTORC1-dependent endothelial translation and pyrimidine synthesis. Our studies may help overcome anti-VEGF therapy resistance by targeting endothelial growth factor receptor translation.
    Keywords:  angiogenesis; aspartate metabolism; endothelial metabolism; mTOR signalling; tumor angiogenesis
    DOI:  https://doi.org/10.1016/j.devcel.2022.04.018
  41. Int J Cardiol. 2022 May 17. pii: S0167-5273(22)00735-5. [Epub ahead of print]
    PINK1/Parkin-mediated mitophagy in cardiovascular disease: From pathogenesis to novel therapy.
    Yanze Wu, Ting Jiang, Jinghai Hua, Zhiping Xiong, Kai Dai, Hui Chen, Lei Li, Jingtian Peng, Xiaoping Peng, Zeqi Zheng, Wenjun Xiong.
      Cardiovascular disease(CVD)is one of the predominant causes of death and morbidity. Mitochondria play a key role in maintaining cardiac energy metabolism. However, mitochondrial dysfunction leads to excessive production of ROS, resulting in oxidative damage to cardiomyocytes and contributing to a variety of cardiovascular diseases. In such a case, the clearance of impaired mitochondria is necessary. Currently, most studies have indicated an essential role for mitophagy in maintaining cardiac homeostasis and regulating CVD-related metabolic transition. Recent studies have implicated that PTEN-induced putative kinase 1 (PINK1)/Parkin-mediated mitophagy has been implicated in maintaining cardiomyocyte homeostasis. Here, we discuss the physiological and pathological roles of PINK1/Parkin-mediated mitophagy in the cardiovascular system, as well as potential therapeutic strategies based on PINK1/Parkin-mediated mitophagy modulation, which are of great significance for the prevention and treatment of cardiovascular diseases.
    Keywords:  Cardiomyocyte; Cardiovascular disease; Mitochondria; Mitophagy; PINK1; Parkin
    DOI:  https://doi.org/10.1016/j.ijcard.2022.05.025
  42. J Cell Mol Med. 2022 May 18.
    Mitochondrial-derived vesicles: Recent insights.
    Lucia-Doina Popov.
      The generation of vesicles is a constitutive attribute of mitochondria inherited from bacterial ancestors. The physiological conditions and mild oxidative stress promote oxidation and dysfunction of certain proteins and lipids within the mitochondrial membranes; these constituents are subsequently packed as small mitochondrial-derived vesicles (MDVs) (70-150 nm in diameter) and are transported intracellularly to lysosomes and peroxisomes to be degraded. In this way, MDVs remove the damaged mitochondrial components, preserve mitochondrial structural and functional integrity and restore homeostasis. An outline of the current knowledge on MDVs seems to be necessary for understanding the potential impact of this research area in cellular (patho)physiology. The present synopsis is an attempt towards the accomplishment of this demand, highlighting also the still unclear issues related to MDVs. Here, we discuss (i) MDVs budding and generation (molecules and mechanisms), (ii) the distinct cargoes packed and transported by MDVs, (iii) the MDVs trafficking pathways and (iv) the biological role of MDVs, from quality controllers to the involvement in organellar crosstalk, mitochondrial antigen presentation and peroxisome de novo biogenesis. These complex roles uncover also mitochondria integration into the cellular environment. As the therapeutic exploitation of MDVs is currently limited, future insights into MDVs cell biology are expected to direct to novel diagnostic tools and treatments.
    Keywords:  PINK1; Parkin; Quality control; extracellular vesicles; lysosomes; peroxisome
    DOI:  https://doi.org/10.1111/jcmm.17391
  43. Nat Commun. 2022 May 19. 13(1): 2760
    Lipid droplet degradation by autophagy connects mitochondria metabolism to Prox1-driven expression of lymphatic genes and lymphangiogenesis.
    Odeta Meçe, Diede Houbaert, Maria-Livia Sassano, Tania Durré, Hannelore Maes, Marco Schaaf, Sanket More, Maarten Ganne, Melissa García-Caballero, Mila Borri, Jelle Verhoeven, Madhur Agrawal, Kathryn Jacobs, Gabriele Bergers, Silvia Blacher, Bart Ghesquière, Mieke Dewerchin, Johan V Swinnen, Stefan Vinckier, María S Soengas, Peter Carmeliet, Agnès Noël, Patrizia Agostinis.
      Autophagy has vasculoprotective roles, but whether and how it regulates lymphatic endothelial cells (LEC) homeostasis and lymphangiogenesis is unknown. Here, we show that genetic deficiency of autophagy in LEC impairs responses to VEGF-C and injury-driven corneal lymphangiogenesis. Autophagy loss in LEC compromises the expression of main effectors of LEC identity, like VEGFR3, affects mitochondrial dynamics and causes an accumulation of lipid droplets (LDs) in vitro and in vivo. When lipophagy is impaired, mitochondrial ATP production, fatty acid oxidation, acetyl-CoA/CoA ratio and expression of lymphangiogenic PROX1 target genes are dwindled. Enforcing mitochondria fusion by silencing dynamin-related-protein 1 (DRP1) in autophagy-deficient LEC fails to restore LDs turnover and lymphatic gene expression, whereas supplementing the fatty acid precursor acetate rescues VEGFR3 levels and signaling, and lymphangiogenesis in LEC-Atg5-/- mice. Our findings reveal that lipophagy in LEC by supporting FAO, preserves a mitochondrial-PROX1 gene expression circuit that safeguards LEC responsiveness to lymphangiogenic mediators and lymphangiogenesis.
    DOI:  https://doi.org/10.1038/s41467-022-30490-6
  44. EMBO Rep. 2022 May 20. e54312
    Cx43-mediated sorting of miRNAs into extracellular vesicles.
    Tania Martins-Marques, Marina C Costa, Steve Catarino, Isaura Simoes, Trond Aasen, Francisco J Enguita, Henrique Girao.
      Through the exchange of lipids, proteins, and nucleic acids, extracellular vesicles (EV) allow for cell-cell communication across distant cells and tissues to regulate a wide range of physiological and pathological processes. Although some molecular mediators have been discovered, the mechanisms underlying the selective sorting of miRNAs into EV remain elusive. Previous studies demonstrated that connexin43 (Cx43) forms functional channels at the EV surface, mediating the communication with recipient cells. Here, we show that Cx43 participates in the selective sorting of miRNAs into EV through a process that can also involve RNA-binding proteins. We provide evidence that Cx43 can directly bind to specific miRNAs, namely those containing stable secondary structure elements, including miR-133b. Furthermore, Cx43 facilitates the delivery of EV-miRNAs into recipient cells. Phenotypically, we show that Cx43-mediated EV-miRNAs sorting modulates autophagy. Overall, our study ascribes another biological role to Cx43, that is, the selective incorporation of miRNAs into EV, which potentially modulates multiple biological processes in target cells and may have implications for human health and disease.
    Keywords:  Connexin43; heterogeneous nuclear ribonucleoproteins; intercellular communication; miRNA; selective sorting
    DOI:  https://doi.org/10.15252/embr.202154312
  45. J Clin Invest. 2022 May 17. pii: e152635. [Epub ahead of print]
    mTORC2 mediates structural plasticity in distal nociceptive endings that contributes to pain hypersensitivity following inflammation.
    Calvin Wong, Omer Barkai, Feng Wang, Carolina Thörn Pérez, Shaya Lev, Weihua Cai, Shannon Tansley, Noosha Yousefpour, Mehdi Hooshmandi, Kevin C Lister, Mariam Latif, A Claudio Cuello, Masha Prager-Khoutorsky, Jeffrey S Mogil, Philippe Séguéla, Yves De Koninck, Alfredo Ribeiro-da-Silva, Alexander M Binshtok, Arkady Khoutorsky.
      The encoding of noxious stimuli into action potential firing is largely mediated by nociceptive free nerve endings. Tissue inflammation, by changing the intrinsic properties of the nociceptive endings, leads to nociceptive hyperexcitability, and thus to the development of inflammatory pain. Here, we showed that tissue inflammation-induced activation of the mammalian target of rapamycin complex 2 (mTORC2) triggers changes in the architecture of nociceptive terminals and leads to inflammatory pain. Pharmacological activation of mTORC2 induced elongation and branching of nociceptor peripheral endings and caused long-lasting pain hypersensitivity. Conversely, nociceptor-specific deletion of the mTORC2 regulatory protein, Rictor, prevented inflammation-induced elongation and branching of cutaneous nociceptive fibers and attenuated inflammatory pain hypersensitivity. Computational modelling demonstrated that mTORC2-mediated structural changes in the nociceptive terminal tree are sufficient to increase the excitability of nociceptors. Targeting mTORC2 using a single injection of antisense oligonucleotide against Rictor provided long-lasting alleviation of inflammatory pain hypersensitivity. Collectively, we showed that tissue inflammation-induced activation of mTORC2 causes structural plasticity of nociceptive free nerve endings in the epidermis and inflammatory hyperalgesia, representing a therapeutic target for inflammatory pain.
    Keywords:  Mouse models; Neuroscience; Pain; Signal transduction
    DOI:  https://doi.org/10.1172/JCI152635
  46. Neurobiol Dis. 2022 May 13. pii: S0969-9961(22)00145-0. [Epub ahead of print]170 105753
    Neuron-astrocyte transmitophagy is altered in Alzheimer's disease.
    Riikka Lampinen, Irina Belaya, Liudmila Saveleva, Jeffrey R Liddell, Dzhessi Rait, Mikko T Huuskonen, Raisa Giniatullina, Annika Sorvari, Liisi Soppela, Nikita Mikhailov, Isabella Boccuni, Rashid Giniatullin, Marcela Cruz-Haces, Julia Konovalova, Marja Koskuvi, Andrii Domanskyi, Riikka H Hämäläinen, Gundars Goldsteins, Jari Koistinaho, Tarja Malm, Sweelin Chew, Kirsi Rilla, Anthony R White, Nicholas Marsh-Armstrong, Katja M Kanninen.
      Under physiological conditions in vivo astrocytes internalize and degrade neuronal mitochondria in a process called transmitophagy. Mitophagy is widely reported to be impaired in neurodegeneration but it is unknown whether and how transmitophagy is altered in Alzheimer's disease (AD). Here we report that the internalization of neuronal mitochondria is significantly increased in astrocytes isolated from AD mouse brains. We also demonstrate that the degradation of neuronal mitochondria by astrocytes is increased in AD mice at the age of 6 months onwards. Furthermore, we demonstrate for the first time a similar phenomenon between human neurons and AD astrocytes, and in murine hippocampi in vivo. The results suggest the involvement of S100a4 in impaired mitochondrial transfer between neurons and AD astrocytes together with significant increases in the mitophagy regulator and reactive oxygen species in aged AD astrocytes. These findings demonstrate altered neuron-supporting functions of AD astrocytes and provide a starting point for studying the molecular mechanisms of transmitophagy in AD.
    Keywords:  Alzheimer's disease; Astrocytes; Mitochondria; Mitophagy; Transmitophagy
    DOI:  https://doi.org/10.1016/j.nbd.2022.105753
  47. J Med Chem. 2022 May 19.
    Ispinesib as an Effective Warhead for the Design of Autophagosome-Tethering Chimeras: Discovery of Potent Degraders of Nicotinamide Phosphoribosyltransferase (NAMPT).
    Guoqiang Dong, Ying Wu, Junfei Cheng, Long Chen, Rui Liu, Yu Ding, Shanchao Wu, Junhui Ma, Chunquan Sheng.
      Autophagosome-tethering compounds (ATTECs) are an emerging new technology in targeted protein degradation. However, effective tools and successful examples for autophagosome-tethering chimeras are still rather limited. Herein, ATTEC ispinesib was identified for the first time to be an effective warhead to design autophagosome-tethering chimeras. As a conceptual validation study, the first generation of autophagic degraders of nicotinamide phosphoribosyltransferase (NAMPT) were developed by connecting the NAMPT inhibitor and LC3-binding ispinesib through a flexible linker. In particular, compound A3 significantly induced the degradation of NAMPT through the autophagy-lysosomal pathway, leading to excellent cellular antitumor potency. Ispinesib may have broad applications in the design of potent autophagosome-tethering chimeras.
    DOI:  https://doi.org/10.1021/acs.jmedchem.1c02001
  48. Nat Commun. 2022 May 16. 13(1): 2706
    Identification of a modulator of the actin cytoskeleton, mitochondria, nutrient metabolism and lifespan in yeast.
    Cierra N Sing, Enrique J Garcia, Thomas G Lipkin, Thomas M Huckaba, Catherine A Tsang, Arielle C Coughlin, Emily J Yang, Istvan R Boldogh, Ryo Higuchi-Sanabria, Liza A Pon.
      In yeast, actin cables are F-actin bundles that are essential for cell division through their function as tracks for cargo movement from mother to daughter cell. Actin cables also affect yeast lifespan by promoting transport and inheritance of higher-functioning mitochondria to daughter cells. Here, we report that actin cable stability declines with age. Our genome-wide screen for genes that affect actin cable stability identified the open reading frame YKL075C. Deletion of YKL075C results in increases in actin cable stability and abundance, mitochondrial fitness, and replicative lifespan. Transcriptome analysis revealed a role for YKL075C in regulating branched-chain amino acid (BCAA) metabolism. Consistent with this, modulation of BCAA metabolism or decreasing leucine levels promotes actin cable stability and function in mitochondrial quality control. Our studies support a role for actin stability in yeast lifespan, and demonstrate that this process is controlled by BCAA and a previously uncharacterized ORF YKL075C, which we refer to as actin, aging and nutrient modulator protein 1 (AAN1).
    DOI:  https://doi.org/10.1038/s41467-022-30045-9
  49. Am J Physiol Cell Physiol. 2022 May 18.
    Chaperone Mediated Autophagy Protects Cardiomyocytes Against Hypoxic-Cell Death.
    Rajeshwary Ghosh, Jennifer Jason Gillaspie, Kenneth S Campbell, J David Symons, Sihem Boudina, James Scott Pattison.
      Chaperone-mediated autophagy (CMA) is a chaperone-dependent process of selective cytosolic protein turnover that targets specific proteins to lysosomes for degradation. Enhancing protein degradation mechanisms have been shown beneficial in multiple models of cardiac disease, including myocardial infarction (MI) and ischemia-reperfusion (I/R) injury. However, the causal role of CMA in cardiomyocyte injury and death is largely unknown. Hypoxia is an important contributor to both MI and I/R damage, which are major precedent causes of heart failure. Upregulating CMA was hypothesized to protect against hypoxia-induced cardiomyocyte death. Lysosome-associated membrane protein 2a (Lamp2a) overexpression and knockdown were used to causally study CMA's role in hypoxically-stressed cardiomyocytes. LAMP2a protein levels were employed as both a primary indicator and driver of CMA function. Hypoxic stress was stimulated by CoCl2 treatment, which increased LAMP2a protein levels (+1.4-fold) and induced cardiomyocyte apoptosis (+3.2 - 4.0-fold). Lamp2a siRNA knockdown (-3.2-fold) of control cardiomyocytes increased apoptosis (+1.8-fold) suggesting that loss of CMA is detrimental for cardiomyocyte survival. However, there was neither an additive nor a synergistic effect on cell death when Lamp2a-silenced cells were treated with CoCl2. Conversely, Lamp2a overexpression (+3.0-fold) successfully reduced hypoxia-induced apoptosis by ~50%. LAMP2a was also significantly increased (+1.7-fold) in ischemic heart failure patient samples, similar to hypoxically-stressed cardiomyocytes. The failing ischemic hearts may have had insufficient CMA activation. This study for the first time establishes a protective role for CMA (via Lamp2a overexpression) against hypoxia-induced cardiomyocyte loss and reveals the intriguing possibility that CMA activation may offer a cardioprotective treatment for ischemic heart disease.
    Keywords:  Apoptosis; Cardiomyocytes; Chaperone-mediated autophagy; Hypoxia; Protein Degradation
    DOI:  https://doi.org/10.1152/ajpcell.00369.2021
  50. J Transl Med. 2022 05 14. 20(1): 229
    Heat shock protein A4 ablation leads to skeletal muscle myopathy associated with dysregulated autophagy and induced apoptosis.
    Manar Elkenani, Amal Z Barakat, Torsten Held, Daniel Marques Rodrigues, Sherok Mobarak, Surabhi Swarnka, Ibrahim M Adham, Belal A Mohamed.
      BACKGROUND: Molecular chaperones assist protein folding, facilitate degradation of misfolded polypeptides, and thereby maintain protein homeostasis. Impaired chaperone activity leads to defective protein quality control that is implicated in multiple skeletal muscle diseases. The heat shock protein A4 (HSPA4) acts as a co-chaperone for HSP70. Previously, we showed that Hspa4 deletion causes impaired protein homeostasis in the heart. However, its functional role in skeletal muscle has not been explored.METHODS: We performed a comparative phenotypic and biochemical analyses of Hspa4 knockout (KO) mice with wild-type (WT) littermates.
    RESULTS: HSPA4 is markedly upregulated in regenerating WT muscle in vivo, and in differentiated myoblasts in vitro. Hspa4-KO mice are marked by growth retardation and increased variability in body weight, accompanied by 35% mortality rates during the peri-weaning period. The surviving Hspa4-KO mice experienced progressive skeletal muscle myopathy, characterized by increased number of muscle fibers with centralized nuclei, heterogeneous myofiber size distribution, inflammatory cell infiltrates and upregulation of embryonic and perinatal myosin heavy chain transcripts. Hspa4-KO muscles demonstrated an accumulation of autophagosome-associated proteins including microtubule associated protein1 light chain 3-II (LC3-II) and p62/sequestosome accompanied by increased number of TUNEL-positive nuclei.
    CONCLUSIONS: Our findings underscore the indispensable role of HSPA4 in maintenance of muscle integrity through contribution in skeletal muscle autophagy and apoptosis, which might provide a novel therapeutic strategy for skeletal muscle morbidities.
    Keywords:  Autophagy; HSPs; Myopathy
    DOI:  https://doi.org/10.1186/s12967-022-03418-3
  51. Obesity (Silver Spring). 2022 May 17.
    Integrating the effects of sucrose intake on the brain and white adipose tissue: Could autophagy be a possible link?
    Pamela Mattar, Lilian Toledo-Valenzuela, María Paz Hernández-Cáceres, Daniel Peña-Oyarzún, Eugenia Morselli, Claudio Perez-Leighton.
      Excess dietary sucrose is associated with obesity and metabolic diseases. This relationship is driven by the malfunction of several cell types and tissues critical for the regulation of energy balance, including hypothalamic neurons and white adipose tissue (WAT). However, the mechanisms behind these effects of dietary sucrose are still unclear and might be independent of increased adiposity. Accumulating evidence has indicated that dysregulation of autophagy, a fundamental process for maintenance of cellular homeostasis, alters energy metabolism in hypothalamic neurons and WAT, but whether autophagy could mediate the detrimental effects of dietary sucrose on hypothalamic neurons and WAT that contribute to weight gain is a matter of debate. In this review, we examine the hypothesis that dysregulated autophagy in hypothalamic neurons and WAT is an adiposity-independent effect of sucrose that contributes to increased body weight gain. We propose that excess dietary sucrose leads to autophagy unbalance in hypothalamic neurons and WAT, which increases caloric intake and body weight, favoring the emergence of obesity and metabolic diseases.
    DOI:  https://doi.org/10.1002/oby.23411
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