bims-auttor Biomed News
on Autophagy and mTOR
Issue of 2020‒10‒11
forty-four papers selected by
Viktor Korolchuk
Newcastle University


  1. Mech Ageing Dev. 2020 Oct 03. pii: S0047-6374(20)30175-5. [Epub ahead of print]192 111379
    Haidurov A, Budanov AV.
      Sestrins are a family of stress-responsive antioxidant proteins responsible for regulation of cell viability and metabolism. The best known Sestrin targets are mTORC1 and mTORC2 kinases that control different cellular processes including growth, viability, autophagy, and mitochondrial metabolism. Inactivation of the single Sestrin gene in invertebrates has an adverse impact on their healthspan and longevity, whereas each of the three Sestrin genes in mammals and other vertebrate organisms has a different impact on maintenance of a particular tissue, affecting its stress tolerance, function and regenerative capability. As a result, Sestrins attenuate ageing and suppress development of many age-related diseases including myocardial infarction, muscle atrophy, diabetes, and immune dysfunction, but exacerbate development of chronic obstructive pulmonary disease. Moreover, Sestrins play opposite roles in carcinogenesis in different tissues. Stem cells support tissue remodelling that influences ageing, and Sestrins might suppress ageing and age-related pathologies through control of stem cell biology. In this review, we will discuss the potential link between Sestrins, stem cells, and ageing.
    Keywords:  Cell death; GATOR1/2; Mitochondria; Sestrin; mTORC1/2
    DOI:  https://doi.org/10.1016/j.mad.2020.111379
  2. Autophagy. 2020 Oct 04.
    Nowosad A, Besson A.
      The tumor suppressor CDKN1B/p27Kip1 binds to and inhibits cyclin-CDK complexes in the nucleus, inducing cell cycle arrest. However, when in the cytoplasm, CDKN1B may promote tumorigenesis. Notably, cytoplasmic CDKN1B was reported to promote macroautophagy/autophagy in response to nutrient shortage by a previously unknown mechanism. In our recent work, we found that during prolonged amino acid starvation, CDKN1B promotes autophagy via an MTORC1-dependent pathway. A fraction of CDKN1B translocates to lysosomes, where it interacts with the Ragulator subunit LAMTOR1, preventing Ragulator assembly, which is required for MTORC1 activation in response to amino acids. Therefore, CDKN1B represses MTORC1 activity, leading to nuclear translocation of the transcription factor TFEB and activation of lysosomal function, enhancing starvation-induced autophagy flux and apoptosis. In contrast, cells lacking CDKN1B survive starvation despite reduced autophagy, due to elevated MTORC1 activation. These findings reveal that, by directly repressing MTORC1 activity, CDKN1B couples the cell cycle and cell growth machineries during metabolic stress.
    DOI:  https://doi.org/10.1080/15548627.2020.1831217
  3. Autophagy. 2020 Oct 06. 1-21
    Garcia EJ, Liao PC, Tan G, Vevea JD, Sing CN, Tsang CA, McCaffery JM, Boldogh IR, Pon LA.
      Our previous studies reveal a mechanism for lipid droplet (LD)-mediated proteostasis in the endoplasmic reticulum (ER) whereby unfolded proteins that accumulate in the ER in response to lipid imbalance-induced ER stress are removed by LDs and degraded by microlipophagy (µLP), autophagosome-independent LD uptake into the vacuole (the yeast lysosome). Here, we show that dithiothreitol- or tunicamycin-induced ER stress also induces µLP and identify an unexpected role for vacuolar membrane dynamics in this process. All stressors studied induce vacuolar fragmentation prior to µLP. Moreover, during µLP, fragmented vacuoles fuse to form cup-shaped structures that encapsulate and ultimately take up LDs. Our studies also indicate that proteins of the endosome sorting complexes required for transport (ESCRT) are upregulated, required for µLP, and recruited to LDs, vacuolar membranes, and sites of vacuolar membrane scission during µLP. We identify possible target proteins for LD-mediated ER proteostasis. Our live-cell imaging studies reveal that one potential target (Nup159) localizes to punctate structures that colocalizes with LDs 1) during movement from ER membranes to the cytosol, 2) during microautophagic uptake into vacuoles, and 3) within the vacuolar lumen. Finally, we find that mutations that inhibit LD biogenesis, homotypic vacuolar membrane fusion or ESCRT function inhibit stress-induced autophagy of Nup159 and other ER proteins. Thus, we have obtained the first direct evidence that LDs and µLP can mediate ER stress-induced ER proteostasis, and identified direct roles for ESCRT and vacuolar membrane fusion in that process.
    Keywords:  ER stress; erad; escrt; lipid droplet proteome; microautophagy; microlipophagy; unfolded protein response; vacuolar membrane fusion; vacuole
    DOI:  https://doi.org/10.1080/15548627.2020.1826691
  4. Kaohsiung J Med Sci. 2020 Oct 05.
    Lin PW, Chu ML, Liu HS.
      Metabolism consists of diverse life-sustaining chemical reactions in living organisms. Autophagy is a highly conservative process that responds to various internal and external stresses. Both processes utilize surrounding resources to provide energy and nutrients for the cell. Autophagy progression may proceed to the degradative or secretory pathway determined by Rab family proteins. The former is a degradative and lysosome-dependent catabolic process that produces energy and provides nutrients for the synthesis of essential proteins. The degradative pathway also balances the energy source of the cell and regulates tissue homeostasis. The latter is a newly discovered pathway in which the autophagosome is fused with the plasma membrane. Secretory autophagy participates in diverse functions and diseases ranging from the spread of viral particles to cancer and neurodegenerative diseases. Aberrant metabolism in the body causes various metabolic syndromes. This review explores the relationships among autophagy, metabolism, and related diseases.
    Keywords:  autophagy; metabolic syndrome; secretory autophagy
    DOI:  https://doi.org/10.1002/kjm2.12299
  5. EMBO Rep. 2020 Oct 07. e50733
    Gao J, Kurre R, Rose J, Walter S, Fröhlich F, Piehler J, Reggiori F, Ungermann C.
      The mechanism and regulation of fusion between autophagosomes and lysosomes/vacuoles are still only partially understood in both yeast and mammals. In yeast, this fusion step requires SNARE proteins, the homotypic vacuole fusion and protein sorting (HOPS) tethering complex, the RAB7 GTPase Ypt7, and its guanine nucleotide exchange factor (GEF) Mon1-Ccz1. We and others recently identified Ykt6 as the autophagosomal SNARE protein. However, it has not been resolved when and how lipid-anchored Ykt6 is recruited onto autophagosomes. Here, we show that Ykt6 is recruited at an early stage of the formation of these carriers through a mechanism that depends on endoplasmic reticulum (ER)-resident Dsl1 complex and COPII-coated vesicles. Importantly, Ykt6 activity on autophagosomes is regulated by the Atg1 kinase complex, which inhibits Ykt6 through direct phosphorylation. Thus, our findings indicate that the Ykt6 pool on autophagosomal membranes is kept inactive by Atg1 phosphorylation, and once an autophagosome is ready to fuse with vacuole, Ykt6 dephosphorylation allows its engagement in the fusion event.
    Keywords:   SNARE ; COPII vesicles; Dsl1 complex; Ykt6; autophagy
    DOI:  https://doi.org/10.15252/embr.202050733
  6. Front Immunol. 2020 ;11 572960
    Fekete T, Ágics B, Bencze D, Bene K, Szántó A, Tarr T, Veréb Z, Bácsi A, Pázmándi K.
      To detect replicating viruses, dendritic cells (DCs) utilize cytoplasmic retinoic acid inducible gene-(RIG) I-like receptors (RLRs), which play an essential role in the subsequent activation of antiviral immune responses. In this study, we aimed to explore the role of the mammalian target of rapamycin (mTOR) in the regulation of RLR-triggered effector functions of human monocyte-derived DCs (moDCs) and plasmacytoid DCs (pDCs). Our results show that RLR stimulation increased the phosphorylation of the mTOR complex (mTORC) 1 and mTORC2 downstream targets p70S6 kinase and Akt, respectively, and this process was prevented by the mTORC1 inhibitor rapamycin as well as the dual mTORC1/C2 kinase inhibitor AZD8055 in both DC subtypes. Furthermore, inhibition of mTOR in moDCs impaired the RLR stimulation-triggered glycolytic switch, which was reflected by the inhibition of lactate production and downregulation of key glycolytic genes. Blockade of mTOR diminished the ability of RLR-stimulated moDCs and pDCs to secret type I interferons (IFNs) and pro-inflammatory cytokines, while it did not affect the phenotype of DCs. We also found that mTOR blockade decreased the phosphorylation of Tank-binding kinase 1 (TBK1), which mediates RLR-driven cytokine production. In addition, rapamycin abrogated the ability of both DC subtypes to promote the proliferation and differentiation of IFN-y and Granzyme B producing CD8 + T cells. Interestingly, AZD8055 was much weaker in its ability to decrease the T cell proliferation capacity of DCs and was unable to inhibit the DC-triggered production of IFN-y and Granyzme B by CD8 + T cells. Here we demonstrated for the first time that mTOR positively regulates the RLR-mediated antiviral activity of human DCs. Further, we show that only selective inhibition of mTORC1 but not dual mTORC1/C2 blockade suppresses effectively the T cell stimulatory capacity of DCs that should be considered in the development of new generation mTOR inhibitors and in the improvement of DC-based vaccines.
    Keywords:  RLR signaling; T cell stimulation; antiviral response; dendritic cell; mTOR
    DOI:  https://doi.org/10.3389/fimmu.2020.572960
  7. Nat Commun. 2020 10 07. 11(1): 5052
    May AI, Prescott M, Ohsumi Y.
      The mechanism and function of autophagy as a highly-conserved bulk degradation pathway are well studied, but the physiological role of autophagy remains poorly understood. We show that autophagy is involved in the adaptation of Saccharomyces cerevisiae to respiratory growth through its recycling of serine. On respiratory media, growth onset, mitochondrial initiator tRNA modification and mitochondrial protein expression are delayed in autophagy defective cells, suggesting that mitochondrial one-carbon metabolism is perturbed in these cells. The supplementation of serine, which is a key one-carbon metabolite, is able to restore mitochondrial protein expression and alleviate delayed respiratory growth. These results indicate that autophagy-derived serine feeds into mitochondrial one-carbon metabolism, supporting the initiation of mitochondrial protein synthesis and allowing rapid adaptation to respiratory growth.
    DOI:  https://doi.org/10.1038/s41467-020-18805-x
  8. Cell Rep. 2020 Oct 06. pii: S2211-1247(20)31219-5. [Epub ahead of print]33(1): 108230
    Moustafa-Kamal M, Kucharski TJ, El-Assaad W, Abbas YM, Gandin V, Nagar B, Pelletier J, Topisirovic I, Teodoro JG.
      mTOR is a serine/threonine kinase and a master regulator of cell growth and proliferation. Raptor, a scaffolding protein that recruits substrates to mTOR complex 1 (mTORC1), is known to be phosphorylated during mitosis, but the significance of this phosphorylation remains largely unknown. Here we show that raptor expression and mTORC1 activity are dramatically reduced in cells arrested in mitosis. Expression of a non-phosphorylatable raptor mutant reactivates mTORC1 and significantly reduces cytotoxicity of the mitotic poison Taxol. This effect is mediated via degradation of PDCD4, a tumor suppressor protein that inhibits eIF4A activity and is negatively regulated by the mTORC1/S6K pathway. Moreover, pharmacological inhibition of eIF4A is able to enhance the effects of Taxol and restore sensitivity in Taxol-resistant cancer cells. These findings indicate that the mTORC1/S6K/PDCD4/eIF4A axis has a pivotal role in the death versus slippage decision during mitotic arrest and may be exploited clinically to treat tumors resistant to anti-mitotic agents.
    Keywords:  PDCD4; S6K; Taxol; cell cycle; eIF4A; hippuristanol; mTORC1; mitosis; raptor
    DOI:  https://doi.org/10.1016/j.celrep.2020.108230
  9. Proc Natl Acad Sci U S A. 2020 Oct 06. pii: 202009838. [Epub ahead of print]
    Mor DE, Sohrabi S, Kaletsky R, Keyes W, Tartici A, Kalia V, Miller GW, Murphy CT.
      Metabolic dysfunction occurs in many age-related neurodegenerative diseases, yet its role in disease etiology remains poorly understood. We recently discovered a potential causal link between the branched-chain amino acid transferase BCAT-1 and the neurodegenerative movement disorder Parkinson's disease (PD). RNAi-mediated knockdown of Caenorhabditis elegans bcat-1 is known to recapitulate PD-like features, including progressive motor deficits and neurodegeneration with age, yet the underlying mechanisms have remained unknown. Using transcriptomic, metabolomic, and imaging approaches, we show here that bcat-1 knockdown increases mitochondrial respiration and induces oxidative damage in neurons through mammalian target of rapamycin-independent mechanisms. Increased mitochondrial respiration, or "mitochondrial hyperactivity," is required for bcat-1(RNAi) neurotoxicity. Moreover, we show that post-disease-onset administration of the type 2 diabetes medication metformin reduces mitochondrial respiration to control levels and significantly improves both motor function and neuronal viability. Taken together, our findings suggest that mitochondrial hyperactivity may be an early event in the pathogenesis of PD, and that strategies aimed at reducing mitochondrial respiration may constitute a surprising new avenue for PD treatment.
    Keywords:  Caenorhabditis elegans; Parkinson’s disease; branched-chain amino acid metabolism; metformin; mitochondria
    DOI:  https://doi.org/10.1073/pnas.2009838117
  10. BMC Mol Cell Biol. 2020 Oct 07. 21(1): 70
    Morshed S, Tasnin MN, Ushimaru T.
      BACKGROUND: Microautophagy, which degrades cargos by direct lysosomal/vacuolar engulfment of cytoplasmic cargos, is promoted after nutrient starvation and the inactivation of target of rapamycin complex 1 (TORC1) protein kinase. In budding yeast, microautophagy has been commonly assessed using processing assays with green fluorescent protein (GFP)-tagged vacuolar membrane proteins, such as Vph1 and Pho8. The endosomal sorting complex required for transport (ESCRT) system is proposed to be required for microautophagy, because degradation of vacuolar membrane protein Vph1 was compromised in ESCRT-defective mutants. However, ESCRT is also critical for the vacuolar sorting of most vacuolar proteins, and hence reexamination of the involvement of ESCRT in microautophagic processes is required.RESULTS: Here, we show that the Vph1-GFP processing assay is unsuitable for estimating the involvement of ESCRT in microautophagy, because Vph1-GFP accumulated highly in the prevacuolar class E compartment in ESCRT mutants. In contrast, GFP-Pho8 and Sna4-GFP destined for vacuolar membranes via an alternative adaptor protein-3 (AP-3) pathway, were properly localized on vacuolar membranes in ESCRT-deficient cells. Nevertheless, microautophagic degradation of GFP-Pho8 and Sna4-GFP after TORC1 inactivation was hindered in ESCRT mutants, indicating that ESCRT is indeed required for microautophagy after nutrient starvation and TORC1 inactivation.
    CONCLUSIONS: These findings provide evidence for the direct role of ESCRT in microautophagy induction.
    Keywords:  AP-3 pathway; ESCRT; Microautophagy; Pho8; VPS pathway; Vph1
    DOI:  https://doi.org/10.1186/s12860-020-00314-w
  11. Front Cell Neurosci. 2020 ;14 264
    Lottes EN, Cox DN.
      Cellular protein homeostasis, or proteostasis, is indispensable to the survival and function of all cells. Distinct from other cell types, neurons are long-lived, exhibiting architecturally complex and diverse multipolar projection morphologies that can span great distances. These properties present unique demands on proteostatic machinery to dynamically regulate the neuronal proteome in both space and time. Proteostasis is regulated by a distributed network of cellular processes, the proteostasis network (PN), which ensures precise control of protein synthesis, native conformational folding and maintenance, and protein turnover and degradation, collectively safeguarding proteome integrity both under homeostatic conditions and in the contexts of cellular stress, aging, and disease. Dendrites are equipped with distributed cellular machinery for protein synthesis and turnover, including dendritically trafficked ribosomes, chaperones, and autophagosomes. The PN can be subdivided into an adaptive network of three major functional pathways that synergistically govern protein quality control through the action of (1) protein synthesis machinery; (2) maintenance mechanisms including molecular chaperones involved in protein folding; and (3) degradative pathways (e.g., Ubiquitin-Proteasome System (UPS), endolysosomal pathway, and autophagy. Perturbations in any of the three arms of proteostasis can have dramatic effects on neurons, especially on their dendrites, which require tightly controlled homeostasis for proper development and maintenance. Moreover, the critical importance of the PN as a cell surveillance system against protein dyshomeostasis has been highlighted by extensive work demonstrating that the aggregation and/or failure to clear aggregated proteins figures centrally in many neurological disorders. While these studies demonstrate the relevance of derangements in proteostasis to human neurological disease, here we mainly review recent literature on homeostatic developmental roles the PN machinery plays in the establishment, maintenance, and plasticity of stable and dynamic dendritic arbors. Beyond basic housekeeping functions, we consider roles of PN machinery in protein quality control mechanisms linked to dendritic plasticity (e.g., dendritic spine remodeling during LTP); cell-type specificity; dendritic morphogenesis; and dendritic pruning.
    Keywords:  autophagy; chaperone; dendrite; developmental homeostasis; neurological disease; proteostasis network; ribosome; ubiquitin-proteasome system (UPS)
    DOI:  https://doi.org/10.3389/fncel.2020.00264
  12. J Appl Toxicol. 2020 Jul 06.
    Deng Q, Jiang L, Mao L, Song XH, He CQ, Li XL, Zhang ZH, Zeng HC, Chen JX, Long DX.
      As an organophosphorus ester, tri-ortho-cresyl phosphate (TOCP) has been widely used in agriculture and industry. It is reported that TOCP can induce organophosphate-induced delayed neuropathy (OPIDN) in sensitive animal and human species. However, the exact molecular mechanisms underlying TOCP-induced neurotoxicity are still unknown. In this study, we found that TOCP could induce autophagy by activating protein kinase C alpha (PKCα) signaling in neuroblastoma SK-N-SH cells. PKCα activators could positively regulate TOCP-induced autophagy by increasing the expression levels of neighbor BRCA1 gene protein 1 (NBR1), LC3 and P62 autophagic receptor protein. Furthermore, PKCα activation impaired the ubiquitin-proteasome system (UPS), resulting in inhibition of proteasome activity and accumulation of ubiquitinated proteins. UPS dysfunction could stimulate autophagy to serve as a compensatory pathway, which contributed to the accumulation of the abnormally hyperphosphorylated tau proteins and degradation of impaired proteins of the MAP 2 and NF-H families in neurodegenerative disorders.
    Keywords:  OPIDN; autophagy; protein kinase C alpha; tri-ortho-cresyl phosphate; ubiquitin-proteasome system
    DOI:  https://doi.org/10.1002/jat.3999
  13. Vet Pathol. 2020 Oct 05. 300985820959243
    Syrjä P, Palviainen M, Jokinen T, Kyöstilä K, Lohi H, Roosje P, Anderegg L, Leeb T, Sukura A, Eskelinen EL.
      Lagotto Romagnolo breed dogs develop a progressive neurological disease with intracellular vacuolar storage when homozygous for a variant in the autophagy-related gene 4D (ATG4D). A lysosomal enzyme deficiency has not been proven in this disease, despite its overlapping morphology with lysosomal storage diseases. Instead, basal autophagy was altered in fibroblasts from affected dogs. The aim of this study was to clarify the origin of the limiting membrane of the accumulating vacuoles and determine whether altered basal autophagy affects the extracellular release of vesicles in cells from diseased dogs. When assessed by immunoelectron microscopy, the membrane of the cytoplasmic vacuoles in affected tissues contained ATG4D, markers for autolysosomes (microtubule-associated protein 1A/B light chain 3 and lysosome-associated membrane protein 2) and for recycling endosomes (transferrin receptor 2), indicating that the vacuoles are hybrid organelles between endocytic and autophagic pathways. Ultracentrifugation, nanoparticle tracking analysis, and mass spectrometry were used to analyze the vesicles released from cultured fibroblasts of affected and control dogs. The amount of extracellular vesicles (EVs) released from affected fibroblasts was significantly increased during basal conditions in comparison to controls. This difference disappeared during starvation. The basal EV proteome of affected cells was enriched with cytosolic, endoplasmic reticulum, and mitochondrial proteins. Heat shock proteins and chaperones, some of which are known substrates of basal autophagy, were identified among the proteins unique to EVs of affected cells. An increased release of extracellular vesicles may serve as a compensatory mechanism in disposal of intracellular proteins during dysfunctional basal autophagy in this spontaneous disease.
    Keywords:  ATG4D; NTA; basal autophagy; canine; disease model; extracellular vesicles; immunoelectron microscopy; mass spectrometry
    DOI:  https://doi.org/10.1177/0300985820959243
  14. Nat Commun. 2020 10 05. 11(1): 4979
    Omer A, Barrera MC, Moran JL, Lian XJ, Di Marco S, Beausejour C, Gallouzi IE.
      Cellular senescence is a known driver of carcinogenesis and age-related diseases, yet senescence is required for various physiological processes. However, the mechanisms and factors that control the negative effects of senescence while retaining its benefits are still elusive. Here, we show that the rasGAP SH3-binding protein 1 (G3BP1) is required for the activation of the senescent-associated secretory phenotype (SASP). During senescence, G3BP1 achieves this effect by promoting the association of the cyclic GMP-AMP synthase (cGAS) with cytosolic chromatin fragments. In turn, G3BP1, through cGAS, activates the NF-κB and STAT3 pathways, promoting SASP expression and secretion. G3BP1 depletion or pharmacological inhibition impairs the cGAS-pathway preventing the expression of SASP factors without affecting cell commitment to senescence. These SASPless senescent cells impair senescence-mediated growth of cancer cells in vitro and tumor growth in vivo. Our data reveal that G3BP1 is required for SASP expression and that SASP secretion is a primary mediator of senescence-associated tumor growth.
    DOI:  https://doi.org/10.1038/s41467-020-18734-9
  15. Autophagy. 2020 Oct 04.
    Banerjee A, Guardia CM, Christenson ET, Zhou W, Tan XF, Lian T, Faraldo-Gómez JD, Bonifacino JS, Jiang J.
      ATG9, the only transmembrane protein in the core macroautophagy/autophagy machinery, is a key player in the early stages of autophagosome formation. Yet, the lack of a high-resolution structure of ATG9 was a major impediment in understanding its three-dimensional organization and function. We recently solved a high-resolution cryoEM structure of the ubiquitously expressed human ATG9A isoform. The structure revealed that ATG9A is a domain-swapped homotrimer with a unique fold, and has an internal network of branched cavities. In cellulo analyses demonstrated the functional importance of the cavity-lining residues. These cavities could serve as conduits for transport of hydrophilic moieties, such as lipid headgroups, across the bilayer. Finally, structure-guided molecular dynamics predicted that ATG9A has membrane-bending properties, which is consistent with its localization to highly curved membranes.
    Keywords:  ATG9A; autophagosome; cryo-EM; membrane curvature; molecular dynamics; transmembrane protein
    DOI:  https://doi.org/10.1080/15548627.2020.1830522
  16. Sci Rep. 2020 Oct 09. 10(1): 16886
    Hollville E, Joers V, Nakamura A, Swahari V, Tansey MG, Moy SS, Deshmukh M.
      Mitochondrial quality control is essential for the long-term survival of postmitotic neurons. The E3 ubiquitin ligase Parkin promotes the degradation of damaged mitochondria via mitophagy and mutations in Parkin are a major cause of early-onset Parkinson's disease (PD). Surprisingly however, mice deleted for Parkin alone are rather asymptomatic for PD-related pathology, suggesting that other complementary or redundant mitochondrial quality control pathways may exist in neurons. Mitochondrial damage is often accompanied by the release of toxic proteins such as cytochrome c. We have reported that once in the cytosol, cytochrome c is targeted for degradation by the E3 ligase CUL9 in neurons. Here we examined whether CUL9 and Parkin cooperate to promote optimal neuronal survival in vivo. We generated mice deficient for both Cul9 and Parkin and examined them for PD-related phenotypes. Specifically, we conducted assays to examine behavioural deficits (locomotor, sensory, memory and learning) and loss of dopaminergic neurons in both males and females. Our results show that the loss of Cul9 and Parkin together did not enhance the effect of Parkin deficiency alone. These results indicate that while both Parkin and CUL9 participate in mitochondrial quality control, neurons likely have multiple redundant mechanisms to ensure their long-term survival.
    DOI:  https://doi.org/10.1038/s41598-020-73854-y
  17. Curr Med Chem. 2020 Oct 06.
    Yang M, Li C, Yang S, Xiao Y, Chen W, Gao P, Jiang N, Xiong S, Wei L, Zhang Q, Yang J, Zeng L, Sun L.
      Diabetic nephropathy (DN) is a common microvascular complication of diabetes and one of the leading causes of end-stage renal disease. Tubular damage is an early change and characteristic of DN, and mitochondrial dysfunction plays an important role in the development of DN. Therefore, the timely removal of damaged mitochondria in tubular cells is an effective treatment strategy for DN. Mitophagy is a type of selective autophagy that ensures the timely elimination of damaged mitochondria to protect cells from oxidative stress. In this review, we summarize our understanding of mitochondrial dysfunction and dynamic disorders in tubular cells in DN and the molecular mechanism of mitophagy. Finally, the role of mitophagy in DN and its feasibility as a therapeutic target for DN are discussed.
    Keywords:  Mitophagy; autophagy.; diabetic nephropathy; mitochondria; oxidative stress; tubular cells
    DOI:  https://doi.org/10.2174/0929867327666201006152656
  18. Autophagy. 2020 Oct 09.
    Manganelli V, Matarrese P, Antonioli M, Gambardella L, Vescovo T, Gretzmeier C, Longo A, Capozzi A, Recalchi S, Riitano G, Misasi R, Dengjel J, Malorni W, Fimia GM, Sorice M, Garofalo T.
      Mitochondria-associated membranes (MAMs) are essential communication subdomains of the endoplasmic reticulum (ER) that interact with mitochondria. We previously demonstrated that, upon macroautophagy/autophagy induction, AMBRA1 is recruited to the BECN1 complex and relocalizes to MAMs, where it regulates autophagy by interacting with raft-like components. ERLIN1 is an endoplasmic reticulum lipid raft protein of the prohibitin family. However, little is known about its association with the MAM interface and its involvement in autophagic initiation. In this study, we investigated ERLIN1 association with MAM raft-like microdomains and its interaction with AMBRA1 in the regulation of the autophagic process. We show that ERLIN1 interacts with AMBRA1 at MAM raft-like microdomains, which represents an essential condition for autophagosome formation upon nutrient starvation, as demonstrated by knocking down ERLIN1 gene expression. Moreover, this interaction depends on the "integrity" of key molecules, such as ganglioside GD3 and MFN2. Indeed, knocking down ST8SIA1/GD3-synthase or MFN2 expression impairs AMBRA1-ERLIN1 interaction at the MAM level and hinders autophagy. In conclusion, AMBRA1-ERLIN1 interaction within MAM raft-like microdomains appears to be pivotal in promoting the formation of autophagosomes.
    Keywords:  AMBRA1; ERLIN1; autophagy; lipid rafts; mitochondria associated membranes
    DOI:  https://doi.org/10.1080/15548627.2020.1834207
  19. Biochem Biophys Res Commun. 2020 Sep 30. pii: S0006-291X(20)31867-2. [Epub ahead of print]
    Sakane H, Urabe J, Nakahira S, Hino K, Miyata N, Akasaki K.
      Lysosomal integral membrane protein-2 (LIMP-2) is a type III transmembrane protein that is highly glycosylated and mainly localized to the lysosomal membrane. The diverse functions of LIMP-2 are currently being uncovered; however, its participation in macroautophagy, usually described as autophagy, has not yet been well-investigated. To determine the possible involvement of LIMP-2 in autophagic activity, we examined the intracellular amount of microtubule-associated protein 1 light chain 3 (LC3)-II, which is well-correlated with autophagosome levels, in exogenous rat LIMP-2-expressing COS7 and HEK293 cells. Transient or stable expression of LIMP-2-myc significantly increased the levels of LC3-II. Conversely, knockdown of LIMP-2 decreased the LC3-II levels in NIH3T3 cells. Furthermore, approaches using lysosomal protease inhibitors and mCherry-GFP-LC3 fluorescence suggested that exogenous expression of LIMP-2 increased the biogenesis of autophagosomes rather than decreased the lysosomal turnover of LC3-II. Considering the results of the biochemical assay and the quantitative fluorescence assay together, it is suggested that LIMP-2 has a possible involvement in autophagic activity, especially autophagosome biogenesis.
    Keywords:  LC3-II; LIMP-2; Lysosomes; Macroautophagy
    DOI:  https://doi.org/10.1016/j.bbrc.2020.09.114
  20. Annu Rev Cell Dev Biol. 2020 Oct 06. 36 265-289
    Moehlman AT, Youle RJ.
      Maintaining mitochondrial health is essential for the survival and function of eukaryotic organisms. Misfunctioning mitochondria activate stress-responsive pathways to restore mitochondrial network homeostasis, remove damaged or toxic proteins, and eliminate damaged organelles via selective autophagy of mitochondria, a process termed mitophagy. Failure of these quality control pathways is implicated in the pathogenesis of Parkinson's disease and other neurodegenerative diseases. Impairment of mitochondrial quality control has been demonstrated to activate innate immune pathways, including inflammasome-mediated signaling and the antiviral cyclic GMP-AMP synthase (cGAS)/stimulator of interferon genes (STING)-regulated interferon response. Immune system malfunction is a common hallmark in many neurodegenerative diseases; however, whether inflammation suppresses or exacerbates disease pathology is still unclear. The goal of this review is to provide a historical overview of the field, describe mechanisms of mitochondrial quality control, and highlight recent advances on the emerging role of mitochondria in innate immunity and inflammation.
    Keywords:  immunity; inflammation; mitochondria; mitophagy; neurodegeneration
    DOI:  https://doi.org/10.1146/annurev-cellbio-021820-101354
  21. EMBO Rep. 2020 Oct 09. e50214
    Yang L, Liang J, Lam SM, Yavuz A, Shui G, Ding M, Huang X.
      Lipid droplets (LDs) are dynamic cytoplasmic organelles present in most eukaryotic cells. The appearance of LDs in neurons is not usually observed under physiological conditions, but is associated with neural diseases. It remains unclear how LD dynamics is regulated in neurons and how the appearance of LDs affects neuronal functions. We discovered that mutations of two key lipolysis genes atgl-1 and lid-1 lead to LD appearance in neurons of Caenorhabditis elegans. This neuronal lipid accumulation protects neurons from hyperactivation-triggered neurodegeneration, with a mild decrease in touch sensation. We also discovered that reduced biosynthesis of polyunsaturated fatty acids (PUFAs) causes similar effects and synergizes with decreased lipolysis. Furthermore, we demonstrated that these changes in lipolysis and PUFA biosynthesis increase PUFA partitioning toward triacylglycerol, and reduced incorporation of PUFAs into phospholipids increases neuronal protection. Together, these results suggest the crucial role of neuronal lipolysis in cell-autonomous regulation of neural functions and neurodegeneration.
    Keywords:   Caenorhabditis elegans ; lipid droplet; lipolysis; neurodegeneration; polyunsaturated fatty acid
    DOI:  https://doi.org/10.15252/embr.202050214
  22. Biochem Biophys Res Commun. 2020 Oct 02. pii: S0006-291X(20)31873-8. [Epub ahead of print]
    Kim K, Park JE, Yeom J, Park N, Trần TT, Kang MJ.
      The organisms have the capacity to sense and adapt to their surroundings for their life in a dynamic environment. In response to amino acid starvation, cells activate a rectifying physiological program, termed the integrated stress response (ISR), to restore cellular homeostasis. General controlled non-repressed (GCN2) kinase is a master regulator of the ISR and modulates protein synthesis in response to amino acid starvation. We previously established the GCN2/ATF4/4E-BP pathway in development and aging. Here, we investigated the tissue-specific roles of GCN2 upon dietary restriction of amino acid in a Drosophila model. The knockdown of GCN2 in the gut and fat body, an energy sensing organ in Drosophila, abolished the beneficial effect of GCN2 in lifespan extension upon dietary restriction of amino acids. Proteome analysis in an autosomal dominant retinitis pigmentosa (ADRP) model showed that dietary restriction of amino acids regulates the synthesis of proteins in several pathways, including mitochondrial translation, mitochondrial gene expression, and regulation of biological quality, and that gcn2-mutant flies have reduced levels of these mitochondria-associated proteins, which may contribute to retinal degeneration in ADRP. These results indicate that the tissue-specific regulation of GCN2 contributes to normal physiology and ADRP progression.
    Keywords:  Aging; Autosomal dominant retinitis pigmentosa; Drosophila; GCN2; Integrated stress response; Mitochondria; Proteomics
    DOI:  https://doi.org/10.1016/j.bbrc.2020.09.120
  23. mBio. 2020 Oct 06. pii: e00852-20. [Epub ahead of print]11(5):
    Bhushan J, Radke JB, Perng YC, Mcallaster M, Lenschow DJ, Virgin HW, Sibley LD.
      The intracellular protozoan parasite Toxoplasma gondii is capable of infecting most nucleated cells, where it survives in a specially modified compartment called the parasitophorous vacuole (PV). Interferon gamma (IFN-γ) is the major cytokine involved in activating cell-autonomous immune responses to inhibit parasite growth within this intracellular niche. In HeLa cells, IFN-γ treatment leads to ubiquitination of susceptible parasite strains, recruitment of the adaptors p62 and NDP52, and engulfment in microtubule-associated protein 1 light chain 3 (LC3)-positive membranes that restrict parasite growth. IFN-γ-mediated growth restriction depends on core members of the autophagy (ATG) pathway but not the initiation or degradative steps in the process. To explore the connection between these different pathways, we used permissive biotin ligation to identify proteins that interact with ATG5 in an IFN-γ-dependent fashion. Network analysis of the ATG5 interactome identified interferon-stimulated gene 15 (ISG15), which is highly upregulated by IFN treatment, as a hub connecting the ATG complex with other IFN-γ-induced genes, suggesting that it forms a functional link between the pathways. Deletion of ISG15 resulted in impaired recruitment of p62, NDP52, and LC3 to the PV and loss of IFN-γ-restricted parasite growth. The function of ISG15 required conjugation, and a number of ISGylated targets overlapped with the IFN-γ-dependent ATG5 interactome, including the adapter p62. Collectively, our findings establish a role for ISG15 in connecting the ATG pathway with IFN-γ-dependent restriction of T. gondii in human cells.IMPORTANCE Interferon(s) provide the primary defense against intracellular pathogens, a property ascribed to their ability to upregulate interferon-stimulated genes. Due to the sequestered niche occupied by Toxoplasma gondii, the host has elaborated intricate ways to target the parasite within its vacuole. One such mechanism is the recognition by a noncanonical autophagy pathway that envelops the parasite-containing vacuole and stunts growth in human cells. Remarkably, autophagy-dependent growth restriction requires interferon-γ, yet none of the classical components of autophagy are induced by interferon. Our studies draw a connection between these pathways by demonstrating that the antiviral protein ISG15, which is normally upregulated by interferons, links the autophagy-mediated control to ubiquitination of the vacuole. These findings suggest a similar link between interferon-γ signaling and autophagy that may underlie defense against other intracellular pathogens.
    Keywords:  ATG5; BioID; ISGylation; LC3; autophagy; autophagy adaptors; intracellular parasites; parasite; parasitophorous vacuole; ubiquitin; ubiquitination
    DOI:  https://doi.org/10.1128/mBio.00852-20
  24. J Cell Sci. 2020 Oct 08. pii: jcs234930. [Epub ahead of print]133(19):
    Smith SF, Collins SE, Charest PG.
      The Ras oncogene is notoriously difficult to target with specific therapeutics. Consequently, there is interest to better understand the Ras signaling pathways to identify potential targetable effectors. Recently, the mechanistic target of rapamycin complex 2 (mTORC2) was identified as an evolutionarily conserved Ras effector. mTORC2 regulates essential cellular processes, including metabolism, survival, growth, proliferation and migration. Moreover, increasing evidence implicate mTORC2 in oncogenesis. Little is known about the regulation of mTORC2 activity, but proposed mechanisms include a role for phosphatidylinositol (3,4,5)-trisphosphate - which is produced by class I phosphatidylinositol 3-kinases (PI3Ks), well-characterized Ras effectors. Therefore, the relationship between Ras, PI3K and mTORC2, in both normal physiology and cancer is unclear; moreover, seemingly conflicting observations have been reported. Here, we review the evidence on potential links between Ras, PI3K and mTORC2. Interestingly, data suggest that Ras and PI3K are both direct regulators of mTORC2 but that they act on distinct pools of mTORC2: Ras activates mTORC2 at the plasma membrane, whereas PI3K activates mTORC2 at intracellular compartments. Consequently, we propose a model to explain how Ras and PI3K can differentially regulate mTORC2, and highlight the diversity in the mechanisms of mTORC2 regulation, which appear to be determined by the stimulus, cell type, and the molecularly and spatially distinct mTORC2 pools.
    Keywords:  Mechanistic target of rapamycin complex 2; Phosphatidylinositol 3-kinase; Ras GTPase
    DOI:  https://doi.org/10.1242/jcs.234930
  25. Ageing Res Rev. 2020 Oct 03. pii: S1568-1637(20)30326-3. [Epub ahead of print] 101191
    Pradeepkiran JA, Hemachandra Reddy P.
      Alzheimer's disease (AD) is a progressive, mental illness without cure. Several years of intense research on postmortem AD brains using cell and mouse models of AD have revealed that multiple cellular changes are involved in the disease process, including mitochondrial abnormalities, synaptic damage, and glial/astrocytic activation, in addition to age-dependent accumulation of amyloid beta (Aβ) and hyperphosphorylated tau (p-tau). Synaptic damage and mitochondrial dysfunction are early cellular changes in the disease process. Healthy and functionally active mitochondria are essential for cellular functioning. Dysfunctional mitochondria play a central role in aging and AD. Mitophagy is a cellular process whereby damaged mitochondria are selectively removed from cell and mitochondrial quality and biogenesis. Mitophagy impairments cause the progressive accumulation of defective organelle and damaged mitochondria in cells. In AD, increased levels of Aβ and p-tau can induce reactive oxygen species (ROS) production, causing excessive fragmentation of mitochondria and promoting defective mitophagy. The current article discusses the latest developments of mitochondrial research and also highlights multiple types of mitophagy, including Aβ and p-tau-induced mitophagy, stress-induced mitophagy, receptor-mediated mitophagy, ubiquitin mediated mitophagy and basal mitophagy. This article also discusses the physiological states of mitochondria, including fission-fusion balance, Ca2+ transport, and mitochondrial transport in normal and diseased conditions. Our article summarizes current therapeutic interventions, like chemical or natural mitophagy enhancers, that influence mitophagy in AD. Our article discusses whether a partial reduction of Drp1 can be a mitophagy enhancer and a therapeutic target for mitophagy in AD and other neurological diseases.
    Keywords:  Alzheimer’s disease; amyloid beta; mitochondrial dysfunction; mitophagy; phosphorylated tau
    DOI:  https://doi.org/10.1016/j.arr.2020.101191
  26. J Biol Chem. 2020 Oct 07. pii: jbc.RA120.013428. [Epub ahead of print]
    Guiney SJ, Adlard PA, Lei P, Mawal CH, Bush AI, Finkelstein DI, Ayton S.
      Neurodegeneration in Parkinson's disease (PD) can be recapitulated in animals by administration of α-synuclein pre-formed fibrils (PFFs) into the brain. However, the mechanism by which these PFFs induce toxicity is unknown. Iron is implicated in PD pathophysiology, so we investigated whether α-synuclein PFFs induce ferroptosis, an iron-dependent cell death pathway. A range of ferroptosis inhibitors were added to a striatal neuron-derived cell line (STHdhQ7/7 cells), a dopaminergic neuron-derived cell line (SN4741 cells) and WT primary cortical neurons, all of which had been intoxicated with α-synuclein PFFs. Viability was not recovered by these inhibitors except for liproxstatin-1, a best-in-class ferroptosis inhibitor, when used at high doses. High dose liproxstatin-1 visibly enlarged the area of a cell that contained acidic vesicles, and elevated the expression of several proteins associated with the autophagy-lysosomal pathway similarly to the known lysosomal inhibitors, chloroquine and bafilomycin A1. Consistent with high dose liproxstatin-1 protecting via a lysosomal mechanism, we further demonstrated that loss of viability induced by α-synuclein PFFs was attenuated by chloroquine and bafilomycin A1 as well as the lysosomal cysteine protease inhibitors, leupeptin, E-64D and Ca-074-Me, but not other autophagy or lysosomal enzyme inhibitors. We confirmed using immunofluorescence microscopy that heparin prevented uptake of α-synuclein PFFs into cells, but that chloroquine did not stop α-synuclein uptake into lysosomes despite impairing lysosomal function and inhibiting α-synuclein toxicity. Together, these data suggested that α-synuclein PFFs are toxic in functional lysosomes in vitro. Therapeutic strategies that prevent α-synuclein fibril uptake into lysosomes may be of benefit in PD.
    Keywords:  Parkinson disease; alpha-synuclein (a-synuclein); cell death; iron; lysosome
    DOI:  https://doi.org/10.1074/jbc.RA120.013428
  27. Trends Neurosci. 2020 Sep 30. pii: S0166-2236(20)30197-1. [Epub ahead of print]
    Keller CW, Münz C, Lünemann JD.
      The CNS accommodates a diverse myeloid immune cell compartment that maintains CNS homeostasis in the steady state while contributing to tissue injury during infectious, autoimmune, and neurodegenerative disease conditions. Autophagy and autophagy proteins play fundamental roles in myeloid cell-related immune functions. Many of these processes do not necessarily involve the canonical formation of a double-membrane structure known as the 'autophagosome' and reflect noncanonical functions of the autophagy machinery. Here, we illustrate recent insights, concepts, and outstanding questions regarding how autophagy pathways in myeloid cells contribute to brain health and disease.
    Keywords:  antigen presentation; autoimmunity; cytokine; dendritic cell; immune response; inflammasome; microglia; nervous system
    DOI:  https://doi.org/10.1016/j.tins.2020.09.003
  28. Toxicol Lett. 2020 Sep 30. pii: S0378-4274(20)30429-X. [Epub ahead of print]334 94-101
    Chen S, Tan S, Yang S, Chen G, Zhu L, Sun Z, Li H, Yao S.
      Silica dust mainly attacks alveolar macrophages (AMs). The apoptosis of AMs is correlated with the progress of silicosis. Our previous study showed that autophagic degradation was blocked in AMs from silicosis patients. However, the effects of nicotine on AM autophagy and apoptosis in silicosis are unknown. In this study, we collected AMs from twenty male workers exposed to silica and divided them into observer and silicosis patient groups, according to the tuberous pathological changes observed by X-ray. The AMs from both groups were exposed to nicotine. We found increased levels of LC3, p62, and cleaved caspase-3, decreased levels of LAMP2, and damaged lysosomes after nicotine stimulation of the AMs from both groups. We also found that the autophagy inhibitor 3-methyladenine (3MA) inhibited nicotine-induced apoptosis in the AMs. Furthermore, 3MA reversed both the nicotine-induced decrease in Bcl-2 and the increase in Bax in both groups. These results suggest that nicotine may induce apoptosis by blocking AM autophagic degradation in human silicosis.
    Keywords:  Alveolar macrophages; Apoptosis; Autophagic degradation; Autophagy; Nicotine; Silicosis
    DOI:  https://doi.org/10.1016/j.toxlet.2020.09.019
  29. Aging Dis. 2020 Oct;11(5): 1260-1275
    Li W, Kui L, Demetrios T, Gong X, Tang M.
      Mitochondria are classically known to be cellular energy producers. Given the high-energy demanding nature of neurons in the brain, it is essential that the mitochondrial pool remains healthy and provides a continuous and efficient supply of energy. However, mitochondrial dysfunction is inevitable in aging and neurodegenerative diseases. In Alzheimer's disease (AD), neurons experience unbalanced homeostasis like damaged mitochondrial biogenesis and defective mitophagy, with the latter promoting the disease-defining amyloid β (Aβ) and p-Tau pathologies impaired mitophagy contributes to inflammation and the aggregation of Aβ and p-Tau-containing neurotoxic proteins. Interventions that restore defective mitophagy may, therefore, alleviate AD symptoms, pointing out the possibility of a novel therapy. This review aims to illustrate mitochondrial biology with a focus on mitophagy and propose strategies to treat AD while maintaining mitochondrial homeostasis.
    Keywords:  Alzheimer’s disease; NAD+; mitochondria dysfunction; mitophagy
    DOI:  https://doi.org/10.14336/AD.2020.0105
  30. Annu Rev Cell Dev Biol. 2020 Oct 06. 36 165-189
    Mallucci GR, Klenerman D, Rubinsztein DC.
      As the world's population ages, neurodegenerative disorders are poised to become the commonest cause of death. Despite this, they remain essentially untreatable. Characterized pathologically both by the aggregation of disease-specific misfolded proteins and by changes in cellular stress responses, to date, therapeutic approaches have focused almost exclusively on reducing misfolded protein load-notably amyloid beta (Aβ) in Alzheimer's disease. The repeated failure of clinical trials has led to despondency over the possibility that these disorders will ever be treated. We argue that this is in fact a time for optimism: Targeting various generic stress responses is emerging as an increasingly promising means of modifying disease progression across these disorders. New treatments are approaching clinical trials, while novel means of targeting aggregates could eventually act preventively in early disease.
    Keywords:  autophagy; integrated stress response; neurodegenerative disease; neuroprotection; protein aggregation; therapy; unfolded protein response
    DOI:  https://doi.org/10.1146/annurev-cellbio-040320-120625
  31. Food Funct. 2020 Oct 08.
    Lu J, Meng Z, Chen Y, Yu L, Gao B, Zheng Y, Guan S.
      Apigenin, as a natural flavonoid, has been proved to have many biological effects. Our previous research has found the antiadipogenic effects of apigenin on HepG2 cells. Autophagy is intimately associated with the metabolism of lipid droplets (LDs) and is considered to be one of the lipid breakdown pathways. However, there is no study to elucidate the lipid-lowering mechanism of apigenin from the perspective of autophagy. Here, we investigated the possible role of apigenin in autophagy and lipid accumulation in palmitic acid (PA)-induced HepG2 cells. Our results showed that apigenin increased autophagosome formation and the LC3-II/I ratio, but decreased the p-mTOR/mTOR ratio and P62 protein expression. The effects of apigenin were blocked by chloroquine (CQ). Likewise, apigenin significantly stimulated autophagic flux in the cytoplasm. This effect also could be blocked by CQ. Moreover, apigenin decreased the lipid content and co-localization of LDs with LC3, and CQ could block these effects. Thus, we proposed that apigenin induced autophagy and stimulated autophagic lipid degradation in PA-treated HepG2 cells.
    DOI:  https://doi.org/10.1039/d0fo00949k
  32. J Biol Chem. 2020 10 05. pii: jbc.REV120.014391. [Epub ahead of print]
    Chen CG, Iozzo RV.
      The extracellular matrix encompasses a reservoir of bioactive macromolecules that modulates a cornucopia of biological functions. A prominent body of work posits matrix constituents as master regulators of autophagy and angiogenesis and provides molecular insight into how these two processes are coordinated. Here, we review current understanding of the molecular mechanisms underlying hyaluronan and its main synthesizer, hyaluronan synthase 2 (HAS2). We critically evaluate the regulation and roles of soluble proteoglycans in affecting autophagy and angiogenesis. Specifically, we assess the role of proteoglycan-evoked autophagy in regulating angiogenesis via HAS2-hyaluronan axis and ATG9A, a novel HAS2 binding partner. We discuss extracellular hyaluronan biology and the post-transcriptional and post-translational modifications that control HAS2. We highlight the emerging group of proteoglycans that utilize outside-in signaling to modulate autophagy and angiogenesis in cancer microenvironments and thoroughly review the most up-to-date understanding of endorepellin signaling in vascular endothelia, providing insight into the temporal complexities involved.
    Keywords:  angiogenesis; autophagy; endothelial cell; hyaluronan; hyaluronan synthase 2; proteoglycan; vascular biology
    DOI:  https://doi.org/10.1074/jbc.REV120.014391
  33. Insect Biochem Mol Biol. 2020 Oct 03. pii: S0965-1748(20)30173-9. [Epub ahead of print] 103484
    Santos-Araujo S, Bomfim L, Araripe LO, Bruno R, Ramos I, Gondim KC.
      Rhodnius prolixus is an obligatorily hematophagous insect known as an important vector of Chagas disease. Autophagy is a conserved cellular mechanism that acts in response to nutrient starvation, where components of the cytoplasm are sequestered by a double membrane organelle, named autophagosome, which is targeted to fuse with the lysosome for degradation. Lipophagy is the process of lipid degradation by selective autophagy, where autophagosomes sequester lipid droplets and degrade triacylglycerol (TAG) generating free fatty acids for β-oxidation. Here, two essential genes of the autophagic pathway, Atg6/Beclin1 (RpAtg6) and Atg8/LC3 (RpAtg8), were silenced and the storage of lipids during starvation in Rhodnius prolixus was monitored. We found that RNAi knockdown of both RpAtg6 and RpAtg8 resulted in higher levels of TAG in the fat body and the flight muscle, 24 days after the blood meal, as well as a larger average diameter of the lipid droplets in the fat body, as seen by Nile Red staining under the confocal fluorescence microscope. Silenced starved insects had lower survival rates when compared to control insects. Accordingly, when examined during the starvation period for monitored activity, silenced insects had lower spontaneous locomotor activity and lower forced flight rates. Furthermore, we found that some genes involved in lipid metabolism had their expression levels altered in silenced insects, such as the Brummer lipase (down regulated) and the adipokinetic hormone receptor (up regulated), suggesting that, as previously observed in mammalian models, the autophagy and neutral lipolysis machineries are interconnected at the transcriptional level. Altogether, our data indicate that autophagy in the fat body is important to allow insects to mobilize energy from lipid stores.
    Keywords:  Atg6/Beclin1; Atg8/LC3; Autophagy; Chagas disease vector; Lipophagy; Rhodnius prolixus
    DOI:  https://doi.org/10.1016/j.ibmb.2020.103484
  34. Autophagy. 2020 Oct 08.
    Lu H, Yang HL, Zhou WJ, Lai ZZ, Qiu XM, Fu Q, Zhao JY, Wang J, Li DJ, Li MQ.
      Deficiency in decidualization has been widely regarded as an important cause of spontaneous abortion. Generalized decidualization also includes massive infiltration and enrichment of NK cells. However, the underlying mechanism of decidual NK (dNK) cell residence remains largely unknown. Here, we observe that the increased macroautophagy/autophagy of decidual stromal cells (DSCs) during decidualization, facilitates the adhesion and retention of dNK cells during normal pregnancy. Mechanistically, this process is mediated through activation of the MITF-TNFRSF14/HVEM signaling, and further upregulation of multiple adhesion adhesions (e.g, Selectins and ICAMs) in a MMP9-dependent manner. Patients with unexplained spontaneous abortion display insufficient DSC autophagy and dNK cell residence. In addition, poor vascular remodeling of placenta, low implantation number and high ratio of embryo loss are observed in NK cell depletion mice. In therapeutic studies, low doses of rapamycin, a known autophagy inducer that significantly promotes endometrium autophagy and NK cell residence, and improves embryo absorption in spontaneous abortion mice models, which should be dependent on the activation of MITF-TNFRSF14/HVEM-MMP9-adhension molecules axis. This observation reveals novel molecular mechanisms underlying DSCs autophagy-driven dNK cell residence, and provides a potential therapeutic strategy to prevent spontaneous abortion.
    Keywords:  MITF; MMP9; NK cells; TNFRSF14/HVEM; abortion; autophagy; decidual stromal cells; early pregnancy; residence
    DOI:  https://doi.org/10.1080/15548627.2020.1833515
  35. Autophagy. 2020 Oct 08.
    Ruparelia AA, McKaige EA, Williams C, Schulze KE, Fuchs M, Oorschot V, Lacene E, Meregalli M, Lee C, Serrano RJ, Baxter EC, Monro K, Torrente Y, Ramm G, Stojkovic T, Lavoie JN, Bryson-Richardson RJ.
      Dominant de novo mutations in the co-chaperone BAG3 cause a severe form of myofibrillar myopathy, exhibiting progressive muscle weakness, muscle structural failure, and protein aggregation. To elucidate the mechanism of disease in, and identify therapies for, BAG3 myofibrillar myopathy, we generated two zebrafish models, one conditionally expressing BAG3P209L and one with a nonsense mutation in bag3. While transgenic BAG3P209L-expressing fish display protein aggregation, modeling the early phase of the disease, bag3 -/- fish exhibit exercise dependent fiber disintegration, and reduced swimming activity, consistent with later stages of the disease. Detailed characterization of the bag3 -/- fish, revealed an impairment in macroautophagic/autophagic activity, a defect we confirmed in BAG3 patient samples. Taken together, our data highlights that while BAG3P209L expression is sufficient to promote protein aggregation, it is the loss of BAG3 due to its sequestration within aggregates, which results in impaired autophagic activity, and subsequent muscle weakness. We therefore screened autophagy-promoting compounds for their effectiveness at removing protein aggregates, identifying nine including metformin. Further evaluation demonstrated metformin is not only able to bring about the removal of protein aggregates in zebrafish and human myoblasts but is also able to rescue the fiber disintegration and swimming deficit observed in the bag3 -/- fish. Therefore, repurposing metformin provides a promising therapy for BAG3 myopathy.
    Keywords:  BAG3; autophagy; metformin; muscle; myofibrillar myopathy; zebrafish
    DOI:  https://doi.org/10.1080/15548627.2020.1833500
  36. Cell Death Differ. 2020 Oct 09.
    Li GM, Li L, Li MQ, Chen X, Su Q, Deng ZJ, Liu HB, Li B, Zhang WH, Jia YX, Wang WJ, Ma JY, Zhang HL, Xie D, Zhu XF, He YL, Guan XY, Bi J.
      Dysregulation of the balance between cell proliferation and cell death is a central feature of malignances. Death-associated protein kinase 3 (DAPK3) regulates programmed cell death including apoptosis and autophagy. Our previous study showed that DAPK3 downregulation was detected in more than half of gastric cancers (GCs), which was related to tumor invasion, metastasis, and poor prognosis. However, the precise molecular mechanism underlying DAPK3-mediated tumor suppression remains unclear. Here, we showed that the tumor suppressive function of DAPK3 was dependent on autophagy process. Mass spectrometry, in vitro kinase assay, and immunoprecipitation revealed that DAPK3 increased ULK1 activity by direct ULK1 phosphorylation at Ser556. ULK1 phosphorylation by DAPK3 facilitates the ULK1 complex formation, the VPS34 complex activation, and autophagy induction upon starvation. The kinase activity of DAPK3 and ULK1 Ser556 phosphorylation were required for DAPK3-modulated tumor suppression. The coordinate expression of DAPK3 with ULK1 Ser556 phosphorylation was confirmed in clinical GC samples, and this co-expression was correlated with favorable survival outcomes in patients. Collectively, these findings indicate that the tumor-suppressor roles of DAPK3 in GC are associated with autophagy and that DAPK3 is a novel autophagy regulator, which can directly phosphorylate ULK1 and activate ULK1. Thus, DAPK3 might be a promising prognostic autophagy-associated marker.
    DOI:  https://doi.org/10.1038/s41418-020-00627-5
  37. Physiol Rep. 2020 Oct;8(19): e14599
    Miyazaki M, Moriya N, Takemasa T.
      The regulation of cellular protein synthesis is a critical determinant of skeletal muscle growth and hypertrophy in response to an increased workload such as resistance exercise. The mechanistic target of rapamycin complex 1 (mTORC1) and its upstream protein kinase Akt1 have been implicated as a central signaling pathway that regulates protein synthesis in the skeletal muscle; however, the precise molecular regulation of mTORC1 activity is largely unknown. This study employed germline Akt1 knockout (KO) mice to examine whether upstream Akt1 regulation is necessary for the acute activation of mTORC1 signaling in the plantaris muscle following mechanical overload. The phosphorylation states of S6 kinase 1, ribosomal protein S6, and eukaryotic translation initiation factor 4E-binding protein 1 which show the functional activity of mTORC1 signaling, were significantly increased in the skeletal muscle of both wildtype and Akt1 KO mice following an acute bout (3 and 12 hr) of mechanical overload. Akt1 deficiency did not affect load-induced alteration of insulin-like growth factor-1 (IGF-1)/IGF receptor mRNA expression. Also, no effect of Akt1 deficiency was observed on the overload-induced increase in the gene expressions of pax7 and myogenic regulatory factor of myogenin. These observations show that the upstream IGF-1/Akt1 regulation is dispensable for the acute activation of mTORC1 signaling and regulation of satellite cells in response to mechanical overload.
    Keywords:  Akt1; mechanistic target of rapamycin; protein synthesis; satellite cells
    DOI:  https://doi.org/10.14814/phy2.14599
  38. Trends Biochem Sci. 2020 Oct 01. pii: S0968-0004(20)30229-2. [Epub ahead of print]
    Saftig P, Puertollano R.
      Lysosomes are in the center of the cellular control of catabolic and anabolic processes. These membrane-surrounded acidic organelles contain around 70 hydrolases, 200 membrane proteins, and numerous accessory proteins associated with the cytosolic surface of lysosomes. Accessory and transmembrane proteins assemble in signaling complexes that sense and integrate multiple signals and transmit the information to the nucleus. This communication allows cells to respond to changes in multiple environmental conditions, including nutrient levels, pathogens, energy availability, and lysosomal damage, with the goal of restoring cellular homeostasis. This review summarizes our current understanding of the major molecular players and known pathways that are involved in control of metabolic and stress responses that either originate from lysosomes or regulate lysosomal functions.
    Keywords:  TFEB; autophagy; lysosomes; mTOR; nutrient sensing; transcription factors
    DOI:  https://doi.org/10.1016/j.tibs.2020.09.004
  39. Cell Prolif. 2020 Oct 08. e12891
    Ouyang Y, Wu Q, Li J, Sun S, Sun S.
      Autophagy is a mechanism that enables cells to maintain cellular homeostasis by removing damaged materials and mobilizing energy reserves in conditions of starvation. Although nutrient availability strongly impacts the process of autophagy, the specific metabolites that regulate autophagic responses have not yet been determined. Recent results indicate that S-adenosylmethionine (SAM) represents a critical inhibitor of methionine starvation-induced autophagy. SAM is primarily involved in four key metabolic pathways: transmethylation, transsulphuration, polyamine synthesis and 5'-deoxyadenosyl 5'-radical-mediated biochemical transformations. SAM is the sole methyl group donor involved in the methylation of DNA, RNA and histones, modulating the autophagic process by mediating epigenetic effects. Moreover, the metabolites of SAM, such as homocysteine, glutathione, decarboxylated SAM and spermidine, also exert important influences on the regulation of autophagy. From our perspective, nuclear-cytosolic SAM is a conserved metabolic inhibitor that connects cellular metabolic status and the regulation of autophagy. In the future, SAM might be a new target of autophagy regulators and be widely used in the treatment of various diseases.
    DOI:  https://doi.org/10.1111/cpr.12891
  40. Annu Rev Cell Dev Biol. 2020 Oct 06. 36 115-139
    Roberts MA, Olzmann JA.
      Lipid droplets (LDs) are endoplasmic reticulum-derived organelles that consist of a core of neutral lipids encircled by a phospholipid monolayer decorated with proteins. As hubs of cellular lipid and energy metabolism, LDs are inherently involved in the etiology of prevalent metabolic diseases such as obesity and nonalcoholic fatty liver disease. The functions of LDs are regulated by a unique set of associated proteins, the LD proteome, which includes integral membrane and peripheral proteins. These proteins control key activities of LDs such as triacylglycerol synthesis and breakdown, nutrient sensing and signal integration, and interactions with other organelles. Here we review the mechanisms that regulate the composition of the LD proteome, such as pathways that mediate selective and bulk LD protein degradation and potential connections between LDs and cellular protein quality control.
    Keywords:  chaperone-mediated autophagy; endoplasmic reticulum; lipid droplet; lipophagy; metabolism; proteasome; protein targeting; triacylglycerol; ubiquitin
    DOI:  https://doi.org/10.1146/annurev-cellbio-031320-101827
  41. Autophagy. 2020 Oct 05. 1-18
    Kim YH, Kwak MS, Lee B, Shin JM, Aum S, Park IH, Lee MG, Shin JS.
      Nuclear protein HMGB1 is secreted in response to various stimuli and functions as a danger-associated molecular pattern. Extracellular HMGB1 induces inflammation, cytokine production, and immune cell recruitment via activation of various receptors. As HMGB1 does not contain an endoplasmic reticulum-targeting signal peptide, HMGB1 is secreted via the endoplasmic reticulum-Golgi independently via an unconventional secretion pathway. However, the mechanism underlying HMGB1 secretion remains largely unknown. Here, we investigated the role of secretory autophagy machinery and vesicular trafficking in HMGB1 secretion. We observed that HSP90AA1 (heat shock protein 90 alpha family class A member 1), a stress-inducible protein, regulates the translocation of HMGB1 from the nucleus to the cytoplasm and its secretion through direct interaction. Additionally, geldanamycin, an HSP90AA1 inhibitor, reduced HMGB1 secretion. GORASP2/GRASP55 (golgi reassembly stacking protein 2), ARF1Q71L (ADP ribosylation factor 1), and SAR1AT39N (secretion associated Ras related GTPase 1A), which promoted unconventional protein secretion, increased HMGB1 secretion. HMGB1 secretion was inhibited by an early autophagy inhibitor and diminished in ATG5-deficient cells even when GORASP2 was overexpressed. In contrast, a late autophagy inhibitor increased HMGB1 secretion under the same conditions. The multivesicular body formation inhibitor GW4869 dramatically decreased HMGB1 secretion under HMGB1 secretion-inducing conditions. Thus, we demonstrated that secretory autophagy and multivesicular body formation mediate HMGB1 secretion.
    Keywords:  GORASP2; HMGB1; HSP90AA1; MVB formation; autophagy; unconventional protein secretion
    DOI:  https://doi.org/10.1080/15548627.2020.1826690
  42. Cells. 2020 Oct 01. pii: E2221. [Epub ahead of print]9(10):
    Mebratu YA, Negasi ZH, Dutta S, Rojas-Quintero J, Tesfaigzi Y.
      Protein degradation is important for proper cellular physiology as it removes malfunctioning proteins or can provide a source for energy. Proteasomes and lysosomes, through the regulatory particles or adaptor proteins, respectively, recognize proteins destined for degradation. These systems have developed mechanisms to allow adaptation to the everchanging environment of the cell. While the complex recognition of proteins to be degraded is somewhat understood, the mechanisms that help switch the proteasomal regulatory particles or lysosomal adaptor proteins to adjust to the changing landscape of degrons, during infections or inflammation, still need extensive exploration. Therefore, this review is focused on describing the protein degradation systems and the possible sensors that may trigger the rapid adaptation of the protein degradation machinery.
    Keywords:  aggresome; autophagy; core particle; endosome; protein degradation; regulatory particle
    DOI:  https://doi.org/10.3390/cells9102221
  43. J Cell Physiol. 2020 Oct 06.
    Bloemberg D, Quadrilatero J.
      Due to the ever-expanding functions attributed to autophagy, there is widespread interest in understanding its contribution to human physiology; however, its specific cellular role as a stress-response mechanism is still poorly defined. To investigate autophagy's role in this regard, we repeatedly subjected cultured mouse myoblasts to two stresses with diverse impacts on autophagic flux: amino acid and serum withdrawal (Hank's balanced salt solution [HBSS]), which robustly induces autophagy, or low-level toxic stress (staurosporine, STS). We found that intermittent STS (int-STS) administration caused cell cycle arrest, development of enlarged and misshapen cells/nuclei, increased senescence-associated heterochromatic foci and senescence-associated β-galactosidase activity, and prevented myogenic differentiation. These features were not observed in cells intermittently incubated in HBSS (int-HB). While int-STS cells displayed less DNA damage (phosphorylated H2A histone family, member X content) and caspase activity when administered cisplatin, int-HB cells were protected from STS-induced cell death. Interestingly, STS-induced senescence was attenuated in autophagy related 7-deficient cells. Therefore, while repeated nutrient withdrawal did not cause senescence, autophagy was required for senescence caused by toxic stress. These results illustrate the context-dependent effects of different stressors, potentially highlighting autophagy as a distinguishing factor.
    Keywords:  autophagy; caspase; cell death; remodeling; senescence
    DOI:  https://doi.org/10.1002/jcp.30079
  44. Life Sci. 2020 Oct 01. pii: S0024-3205(20)31266-2. [Epub ahead of print]262 118513
    Fattahi S, Amjadi-Moheb F, Tabaripour R, Ashrafi GH, Akhavan-Niaki H.
      PI3K/AKT/mTOR pathway is one of the most important signaling pathways involved in normal cellular processes. Its aberrant activation modulates autophagy, epithelial-mesenchymal transition, apoptosis, chemoresistance, and metastasis in many human cancers. Emerging evidence demonstrates that some infections as well as epigenetic regulatory mechanisms can control PI3K/AKT/mTOR signaling pathway. In this review, we focused on the role of this pathway in gastric cancer development, prognosis, and metastasis, with an emphasis on epigenetic alterations including DNA methylation, histone modifications, and post-transcriptional modulations through non-coding RNAs fluctuations as well as H. pylori and Epstein-Barr virus infections. Finally, we reviewed different molecular targets and therapeutic agents in clinical trials as a potential strategy for gastric cancer treatment through the PI3K/AKT/mTOR pathway.
    Keywords:  Chemoresistance; Epigenetic modulation; Gastric cancer; Non-coding RNAs; PI3K/AKT/mTOR pathway; Targeted therapy
    DOI:  https://doi.org/10.1016/j.lfs.2020.118513