bims-auttor Biomed News
on Autophagy and mTOR
Issue of 2022‒03‒06
39 papers selected by
Viktor Korolchuk
Newcastle University

  1. Curr Opin Cell Biol. 2022 Feb 28. pii: S0955-0674(22)00010-2. [Epub ahead of print]75 102064
      The homeostasis of cells depends on the selective degradation of damaged or superfluous cellular components. Autophagy is the major pathway that recognizes such components, sequesters them in de novo formed autophagosomes and delivers them to lysosomes for degradation. The recognition of specific cargo and the biogenesis of autophagosomes involve a dedicated machinery of autophagy related (ATG) proteins. Intense research over the past decades has revealed insights into the function of autophagy proteins and mechanisms that govern cargo recognition. Other aspects including the molecular mechanisms involved in the onset of human diseases are less well understood. However, autophagic dysfunctions, caused by age related decline in autophagy or mutations in ATG proteins, are directly related to a large number of human pathologies including neurodegenerative disorders. Here, we review most recent discoveries and breakthroughs in selective autophagy and its relationship to neurodegeneration.
  2. Front Cell Dev Biol. 2022 ;10 793328
      Efficient proteostasis is crucial for somatic maintenance, and its decline during aging leads to cellular dysfunction and disease. Selective autophagy is a form of autophagy mediated by receptors that target specific cargoes for degradation and is an essential process to maintain proteostasis. The protein Sequestosome 1 (p62/SQSTM1) is a classical selective autophagy receptor, but it also has roles in the ubiquitin-proteasome system, cellular metabolism, signaling, and apoptosis. p62 is best known for its role in clearing protein aggregates via aggrephagy, but it has recently emerged as a receptor for other forms of selective autophagy such as mitophagy and lipophagy. Notably, p62 has context-dependent impacts on organismal aging and turnover of p62 usually reflects active proteostasis. In this review, we highlight recent advances in understanding the role of p62 in coordinating the ubiquitin-proteasome system and autophagy. We also discuss positive and negative effects of p62 on proteostatic status and their implications on aging and neurodegeneration. Finally, we relate the link between defective p62 and diseases of aging and examine the utility of targeting this multifaceted protein to achieve proteostatic benefits.
    Keywords:  aging; autophagy; neurodegenerative diseases; p62 (sequestosome 1(SQSTM1)); proteasome
  3. J Cell Biol. 2022 May 02. pii: e202107151. [Epub ahead of print]221(5):
      Autophagy is a conserved eukaryotic lysosomal degradation pathway that responds to environmental and cellular cues. Autophagy is essential for the meiotic exit and sporulation in budding yeast, but the underlying molecular mechanisms remain unknown. Here, we show that autophagy is maintained during meiosis and stimulated in anaphase I and II. Cells with higher levels of autophagy complete meiosis faster, and genetically enhanced autophagy increases meiotic kinetics and sporulation efficiency. Strikingly, our data reveal that the conserved phosphatase Cdc14 regulates meiosis-specific autophagy. Cdc14 is activated in anaphase I and II, accompanying its subcellular relocation from the nucleolus to the cytoplasm, where it dephosphorylates Atg13 to stimulate Atg1 kinase activity and thus autophagy. Together, our findings reveal a meiosis-tailored mechanism that spatiotemporally controls meiotic autophagy activity to ensure meiosis progression, exit, and sporulation.
  4. Autophagy. 2022 Feb 28. 1-14
      Chloroquine (CQ), a lysosomotropic agent, is commonly used to inhibit lysosomal degradation and macroautophagy/autophagy. Here we investigated the cell-extrinsic effects of CQ on secretion. We showed that lysosomal and autophagy inhibition by CQ altered the secretome, and induced the release of Atg8 orthologs and autophagy receptors. Atg8-family proteins, in particular, were secreted inside small extracellular vesicles (sEVs) in a lipidation-dependent manner. CQ treatment enhanced the release of Atg8-family proteins inside sEVs. Using full-length ATG16L1 and an ATG16L1 mutant that enables Atg8-family protein lipidation on double but not on single membranes, we demonstrated that LC3B is released in two distinct sEV populations: one enriched with SDCBP/Syntenin-1, CD63, and endosomal lipidated LC3B, and another that contains LC3B but is not enriched with SDCBP/Syntenin-1 or CD63, and which our data supports as originating from a double-membrane source. Our findings underscore the context-dependency of sEV heterogeneity and composition, and illustrate the integration of autophagy and sEV composition in response to lysosomal inhibition.Abbreviations: ACTB: actin beta; ANOVA: analysis of variance; ATG4B: autophagy related 4B cysteine peptidase; Atg8: autophagy related 8; ATG16L1: autophagy related 16 like 1; ATP5F1A/ATP5a: ATP synthase F1 subunit alpha; CALCOCO2: calcium binding and coiled-coil domain 2; CASP3: caspase 3; CASP7: caspase 7; CQ: chloroquine; CD9: CD9 molecule; CD63: CD63 molecule; DAPI: 4',6-diamidino-2-phenylindole; DQ-BSA: dye quenched-bovine serum albumin; ER: endoplasmic reticulum; ERN1/IRE1a: endoplasmic reticulum to nucleus signaling 1; EV: extracellular vesicles; FBS: fetal bovine serum; FDR: false discovery rate; GABARAP: GABA type A receptor-associated protein; GABARAPL2: GABA type A receptor associated protein like 2; GAPDH: glyceraldehyde-3-phosphate dehydrogenase; GFP: green fluorescent protein; GO: gene ontology; HCQ: hydroxychloroquine; HSP90AA1: heat shock protein 90 alpha family class A member 1; IP: immunoprecipitation; KO: knockout; LAMP2: lysosomal associated membrane protein 2; LIR: LC3-interacting region; LMNA: lamin A/C; MAP1LC3B/LC3B: microtubule associated protein 1 light chain 3 beta; MS: mass spectrometry; NBR1: NBR1 autophagy cargo receptor; NCOA4: nuclear receptor coactivator 4; NTA: nanoparticle tracking analysis; PE: phosphatidylethanolamine; PECA: probe-level expression change averaging; SDCBP/syntenin-1: syndecan binding protein; SD: standard deviation; SE: secreted; sEV: small extracellular vesicles; SQSTM1/p62: sequestosome 1; TAX1BP1: Tax1 binding protein 1; TEM: transmission electron microscopy; TMT: tandem-mass tag; TSG101: tumor susceptibility 101; ULK1: unc-51 like autophagy activating kinase 1; WC: whole cell.
    Keywords:  ATG16L1; Atg8; CD63; MAP1LC3B; SDCBP/syntenin-1; autophagy; chloroquine; endosome; extracellular vesicle; lysosome
  5. Biochem Soc Trans. 2022 Feb 28. 50(1): 621-632
      Mutations in the leucine-rich repeat kinase 2 (LRRK2) gene are associated with familial and sporadic forms of Parkinson's disease (PD), for which the LRRK2 locus itself represents a risk factor. Idiopathic and LRRK2-related PD share the main clinical and neuropathological features, thus animals harboring the most common LRRK2 mutations, i.e. G2019S and R1441C/G, have been generated to replicate the parkinsonian phenotype and investigate the underlying pathological mechanisms. Most LRRK2 rodent models, however, fail to show the main neuropathological hallmarks of the disease i.e. the degeneration of dopaminergic neurons in the substantia nigra pars compacta and presence of Lewy bodies or Lewy body-like aggregates of α-synuclein, lacking face validity. Rather, they manifest dysregulation in cellular pathways and functions that confer susceptibility to a variety of parkinsonian toxins/triggers and model the presymptomatic/premotor stages of the disease. Among such susceptibility factors, dysregulation of synaptic activity and proteostasis are evident in LRRK2 mutants. These abnormalities are also manifest in the PD brain and represent key events in the development and progression of the pathology. The present minireview covers recent articles (2018-2021) investigating the role of LRRK2 and LRRK2 mutants in the regulation of synaptic activity and autophagy-lysosomal pathway. These articles confirm a perturbation of synaptic vesicle endocytosis and glutamate release in LRRK2 mutants. Likewise, LRRK2 mutants show a marked impairment of selective forms of autophagy (i.e. mitophagy and chaperone-mediated autophagy) and lysosomal function, with minimal perturbations of nonselective autophagy. Thus, LRRK2 rodents might help understand the contribution of these pathways to PD.
    Keywords:  Parkinson's disease; autophagy; leucine rich repeat kinase; lysosomes; synaptic transmission
  6. Neurochem Int. 2022 Feb 23. pii: S0197-0186(22)00036-5. [Epub ahead of print]155 105311
      Mechanistic/mammalian target of rapamycin (mTOR) belongs to the phosphatidylinositol kinase-related kinase (PIKK) family. mTOR signaling is required for the commencement of essential cell functions including autophagy. mTOR primarily governs cell growth in response to favourable nutrients and other growth stimuli. However, it also influences aging and other aspects of nutrient-related physiology such as protein synthesis, ribosome biogenesis, and cell proliferation in adults with very limited growth. The major processes for survival such as synaptic plasticity, memory storage and neuronal recovery involve a significant mTOR activity. mTOR dysregulation is becoming a prevalent motif in a variety of human diseases, including cancer, neurological disorders, and other metabolic syndromes. The use of rapamycin to prolong life in different animal models may be attributable to the multiple roles played by mTOR signaling in various processes involved in ageing, protein translation, autophagy, stem cell pool turnover, inflammation, and cellular senescence. mTOR activity was found to be altered in AD brains and rodent models, supporting the notion that aberrant mTOR activity is one of the key events contributing to the onset and progression of AD hallmarks This review assesses the molecular association between the mTOR signaling pathway and pathogenesis of Alzheimer's disease. The research data supporting this theme are also reviewed.
    Keywords:  Alzheimer's disease; Apoptosis; Cognition; Macroautophagy; Rapamycin; mTOR
  7. Autophagy. 2022 Feb 27. 1-2
      Macroautophagy/autophagy plays crucial roles in aging and the pathogenesis of age-related diseases. Studies in various animal models demonstrate the conserved requirement for autophagy-related genes in multiple anti-aging interventions. A recent study from the Shirasu-Hiza lab showed that a newly designed intermittent time-restricted feeding (iTRF) dietary regimen can robustly extend fly healthspan and lifespan through circadian rhythm-dependent activation of autophagy. The night-specific induction of autophagy is both necessary and sufficient for iTRF-mediated health benefits. The study provides the intriguing possibility that novel behavioral or pharmaceutical interventions that promote night-specific autophagy can be used to promote healthy aging.
    Keywords:  Aging; autophagy; circadian rhythm; lifespan; time-restricted feeding
  8. Autophagy. 2022 Feb 27. 1-2
      The endoplasmic reticulum (ER) carries out essential cellular functions ranging from protein trafficking to metabolite signaling. ER function is maintained in part by quality control pathways including ER degradation by selective autophagy (reticulophagy) during conditions of cellular stress. Reticulophagy is known to be important for cellular responses to starvation and protein folding stress, but no natural role during development had been identified. While investigating ER remodeling during the conserved cell differentiation process of meiosis in budding yeast, we unexpectedly observed developmentally regulated reticulophagy that was driven by expression of the autophagy receptor Atg40. This reticulophagy was coordinated with massive morphological rearrangement of the ER, including movement of most cortical ER away from the cell periphery. As meiotic reticulophagy prevents specific ER subpopulations from being inherited by gametes, we propose that it serves a quality control role, preventing deleterious material from being passed on to subsequent generations.
    Keywords:  Atg40; ERphagy; endoplasmic reticulum; gametes; meiosis; quality control; reticulophagy
  9. Cell Mol Life Sci. 2022 Mar 01. 79(3): 167
      The cellular defense mechanisms against cumulative endo-lysosomal stress remain incompletely understood. Here, we identify Ubr1 as a protein quality control (QC) E3 ubiquitin-ligase that counteracts proteostasis stresses by facilitating endosomal cargo-selective autophagy for lysosomal degradation. Astrocyte regulatory cluster membrane protein MLC1 mutations cause endosomal compartment stress by fusion and enlargement. Partial lysosomal clearance of mutant endosomal MLC1 is accomplished by the endosomal QC ubiquitin ligases, CHIP and Ubr1 via ESCRT-dependent route. As a consequence of the endosomal stress, a supportive QC mechanism, dependent on both Ubr1 and SQSTM1/p62 activities, targets ubiquitinated and arginylated MLC1 mutants for selective endosomal autophagy (endophagy). This QC pathway is also activated for arginylated Ubr1-SQSTM1/p62 autophagy cargoes during cytosolic Ca2+-assault. Conversely, the loss of Ubr1 and/or arginylation elicited endosomal compartment stress. These findings underscore the critical housekeeping role of Ubr1 and arginylation-dependent endophagy/autophagy during endo-lysosomal proteostasis perturbations and suggest a link of Ubr1 to Ca2+ homeostasis and proteins implicated in various diseases including cancers and brain disorders.
    Keywords:  Lysosome; Protein homeostasis network; Protein stability; Regeneration; Reprogramming; Stress response
  10. Cell Rep. 2022 Mar 01. pii: S2211-1247(22)00171-1. [Epub ahead of print]38(9): 110444
      Accumulation of senescent cells affects organismal aging and the prevalence of age-associated disease. Emerging evidence suggests that activation of autophagy protects against age-associated diseases and promotes longevity, but the roles and regulatory mechanisms of autophagy in cellular senescence are not well understood. Here, we identify the transcription factor, MondoA, as a regulator of cellular senescence, autophagy, and mitochondrial homeostasis. MondoA protects against cellular senescence by activating autophagy partly through the suppression of an autophagy-negative regulator, Rubicon. In addition, we identify peroxiredoxin 3 (Prdx3) as another downstream regulator of MondoA essential for mitochondrial homeostasis and autophagy. Rubicon and Prdx3 work independently to regulate senescence. Furthermore, we find that MondoA knockout mice have exacerbated senescence during ischemic acute kidney injury (AKI), and a decrease of MondoA in the nucleus is correlated with human aging and ischemic AKI. Our results suggest that decline of MondoA worsens senescence and age-associated disease.
    Keywords:  C. elegans; MondoA; Rubicon; aging; autophagy; cellular senescence; kidney; mitochondrial homeostasis; mml-1; peroxiredoxin 3
  11. Acta Mater Med. 2022 ;1(1): 42-55
      Maintaining neuronal integrity and functions requires precise mechanisms controlling organelle and protein quality. Alzheimer's disease (AD) is characterized by functional defects in the clearance and recycling of intracellular components. As such, neuronal homeostasis involves autophagy, mitophagy, and apoptosis. Compromised activity in these cellular processes may cause pathological phenotypes of AD. Dysfunction of mitochondria is one of the hallmarks of AD. Mitophagy is a critical mitochondria quality control system, and the impaired mitophagy is observed in AD. Myeloid cell leukemia 1 (MCL1), a member of the pro-survival B-cell lymphoma protein 2 (BCL2) family, is a mitochondria-targeted protein that contributes to maintaining mitochondrial integrity. Mcl1 knockout mice display peri-implantation lethality. The studies on conditional Mcl1 knockout mice demonstrate that MCL1 plays a central role in neurogenesis and neuronal survival during brain development. Accumulating evidence reveals the critical role of MCL1 as a regulator of neuronal autophagy, mitophagy, and survival. In this review, we discuss the emerging neuroprotective function of MCL1 and how dysregulation of MCL1 signaling is involved in the pathogenesis of AD. As the pro-survival BCL2 family of proteins are promising targets of pharmacological intervention with BH3 mimetic drugs, we also discuss the promise of MCL1-targeting therapy in AD.
    Keywords:  Alzheimer’s disease; Apoptosis; Autophagy; BH3-mimetics; MCL1; Mitochondria; Mitophagy
  12. Autophagy. 2022 Feb 28. 1-12
      Macroautophagy/autophagy is a tightly regulated catabolic process, which contributes at baseline level to cellular homeostasis, and upon its stimulation to the adaptive cellular response to intra- and extracellular stress stimuli. Decrease of autophagy activity is occurring upon aging and thought to contribute to age-related-diseases. Recently, we uncovered, upon autophagy induction, the role of de novo DNMT3A (DNA methyltransferase 3 alpha)-mediated DNA methylation on expression of the MAP1LC3 (microtubule associated protein 1 light chain 3) proteins, core components of the autophagy pathway, which resulted in reduced baseline autophagy activity. Here, we report that serine/threonine kinase ULK3 (unc-51 like kinase 3)-dependent activation of GLI1 (GLI family zinc finger 1) contributes to the transcriptional upregulation of DNMT3A gene expression upon autophagy induction, thereby bringing additional understanding of the long-term effect of autophagy induction and a possible mechanism for its decline upon aging, pathological conditions, or in response to treatment interventions.Abbreviations: CBZ: carbamazepine; ChIP: chromatin immunoprecipitation; Clon: clonidine; DNMT3A: DNA methyltransferase 3 alpha; GLI1: GLI family zinc finger 1; GLI2: GLI family zinc finger 2; MAP1LC3: microtubule associated protein 1 light chain 3; MTOR: mechanistic target of rapamycin kinase; PLA: proximity ligation assay; RT-qPCR: quantitative reverse transcription PCR; shRNA: small hairpin RNA; siRNA: small interfering RNA; Treh: trehalose; ULK3: unc-51 like kinase 3.
    Keywords:  Autophagy; DNMT3A; GLI1; ULK3; transcription
  13. Autophagy. 2022 Mar 03. 1-10
      Macroautophagy/autophagy, a fundamental cell process for nutrient recycling and defense against pathogens (termed xenophagy), is crucial to human health. ATG16L2 (autophagy related 16 like 2) is an autophagic protein and a paralog of ATG16L1. Both proteins are implicated in similar diseases such as cancer and other chronic diseases; however, most autophagy studies to date have primarily focused on the function of ATG16L1, with ATG16L2 remaining uncharacterized and understudied. Overexpression of ATG16L2 has been reported in various cancers including colorectal, gastric, and prostate carcinomas, whereas altered methylation of ATG16L2 has been associated with lung cancer formation and poorer response to therapy in leukemia. In addition, ATG16L2 polymorphisms have been implicated in a range of other diseases including inflammatory bowel diseases and neurodegenerative disorders. Despite this likely role in human health, the function of this enigmatic protein in autophagy remains unknown. Here, we review current studies on ATG16L2 and collate evidence that suggests that this protein is a potential modulator of autophagy as well as the implications this has on pathogenesis.Abbreviations: ATG5: autophagy related 5; ATG12: autophagy related 12; ATG16L1: autophagy related 16 like 1; ATG16L2: autophagy related 16 like 2; CD: Crohn disease; IBD: inflammatory bowel diseases; IRGM: immunity related GTPase M; MAP1LC3/LC3: microtubule associated protein 1 light chain 3; PE: phosphatidylethanolamine; RB1CC1: RB1 inducible coiled-coil 1; SLE: systemic lupus erythematosus; WIPI2B: WD repeat domain, phosphoinositide interacting 2B.
    Keywords:  ATG16L1; ATG16L2; autophagy; cancer; macroautophagy; xenophagy
  14. Cell Mol Gastroenterol Hepatol. 2022 Feb 28. pii: S2352-345X(22)00042-X. [Epub ahead of print]
      BACKGROUND & AIMS: Dysregulation of liver lipid metabolism is associated with the development and progression of non-alcoholic fatty liver disease (NAFLD), a spectrum of liver diseases including non-alcoholic steatohepatitis (NASH). In the liver, insulin controls lipid homeostasis by increasing triglyceride (TAG) synthesis, suppressing fatty acid oxidation, and enhancing TAG export via very low-density lipoproteins (VLDL). Downstream of insulin signaling, the mechanistic target of rapamycin complex 1 (mTORC1), is a key regulator of lipid metabolism. Here, we define the role of mTORC1 activity in mouse models of NASH and investigate the mTORC1-dependent mechanisms responsible for protection against liver damage in NASH.METHODS: Utilizing two rodent NASH-promoting diets, we demonstrate that hepatic mTORC1 activity was reduced in mice with NASH, whereas under conditions of insulin resistance and benign fatty liver, mTORC1 activity was elevated. To test the beneficial effects of hepatic mTORC1 activation in mouse models of NASH, we employed an acute, liver-specific knockout model of TSC1 (L-TSC-KO), a negative regulator of mTORC1.
    RESULTS: L-TSC-KO mice are protected from and have improved markers of NASH including reduced steatosis, decreased circulating transaminases, and reduced expression of inflammation and fibrosis genes. Mechanistically, protection from hepatic inflammation and fibrosis by constitutive mTORC1 occurred via promotion of the phosphatidylcholine synthesizing enzyme, CCTα, and enhanced VLDL-TAG export. Additionally, activation of mTORC1 protected from hepatic steatosis via negative feedback of the mTORC2-AKT-FOXO-SREBP1c lipogenesis axis.
    CONCLUSIONS: Collectively, this study identifies a protective role for mTORC1 signaling in the initiation and progression of NASH in mice via dual control of lipid export and synthesis.
    Keywords:  CCTα; FOXO1; insulin; non-alcoholic fatty liver disease; phosphatidylcholine
  15. Autophagy. 2022 Feb 28. 1-2
      Macroautophagy/autophagy-related protein Atg8/LC3 is important for autophagosome biogenesis and required for selective degradation of various substrates. In our recent study, we performed a yeast two-hybrid screening to identify proteins that interact with Atg8a, the Drosophila homolog of Atg8/LC3. The screening identified several Atg8a-interacting proteins. These proteins include: i) proteins which have already been experimentally verified to bind Atg8a, such as Atg1, DOR, ref(2)P and key (Kenny); ii) proteins for which their mammalian homologs interact with Atg8-family members, like Ank2, Atg4, and Nedd4; and iii) several novel Atg8a-interacting proteins, such as trc/STK38 and Tak1. We showed that Tak1, as well as its co-activator, Tab2, both interact with Atg8a and are substrates for selective autophagic clearance. We also determined that SH3PX1 interacts with Tab2 and is necessary for the effective regulation of the immune-deficiency (IMD) pathway. Our findings suggest a mechanism for the regulatory interactions between Tak1-Tab2-SH3PX1 and Atg8a, which contribute to the fine-tuning of the IMD pathway.
    Keywords:  Autophagy; IMD; LIR-motif; Sh3px1; Tab2; Tak1; inflammation
  16. Biomed Pharmacother. 2022 Feb 28. pii: S0753-3322(22)00150-0. [Epub ahead of print]148 112762
      Epigenetics refers to alterations in gene expressions that are reversible and stable, but do not involve changes in DNA sequences. In recent years, an increasing number of studies have shown that epigenetics plays a critical role in autophagy, which can be schematized as a biological process comprising of the following steps: autophagy signal activation, autophagic vesicle elongation, autophagosome maturation and autophagosome-lysosome fusion. As previously reported, autophagy can maintain intracellular homeostasis and autophagy dysfunction will lead to various diseases. For instance, the abnormal expression of genes involved in autophagy can result in the occurrence of many cancers and atherosclerosis. It is also well known that epigenetic modifications can affect autophagy related genes expressions and modulate other signaling molecular involved in autophagy. As an important epigenetic enzyme, LSD1 (lysine specific demethylase 1) plays an essential role in modulating autophagy. On one hand, LSD1 directly regulates autophagy-related genes expressions, including ATGs, Beclin-1, LC3 and SQSTM1/p62. On the other hand, inhibition of LSD1 can activate autophagy through regulating the activities of some other proteins such as p53, SESN2, mTORC1 and PTEN. Since autophagy activation is tightly related to the occurrence of various diseases and can be induced by LSD1 inhibition, development of LSD1 inhibitors will provide a new direction to treat such diseases. In this review, we described the mechanisms by which LSD1 regulates autophagy in different manners and how autophagic dysfunction leads to diseases occurrence. In addition, some LSD1 inhibitors used to treat diseases through modulating autophagy are also summarized in our review.
    Keywords:  Autophagy; Diseases; Inhibitors; LSD1
  17. Oncol Lett. 2022 Apr;23(4): 109
      LAPTM4B is upregulated in the majority of types of cancer and associated with cancer cell proliferation, survival and drug resistance, as well as poor patient prognosis. LAPTM4B knockdown inhibits autophagosome maturation in the context of metabolic stress. Autophagy is a homeostatic process that degrades and recycles intracellular components in response to metabolic stress. The function of autophagy is dual, as this process can either have a tumor suppressor or an oncogenic role. EGFR serves an important role in determining the tumor-suppressive or oncogenic roles of autophagy. EGFR family members regulate autophagy through various signaling pathways, including PI3K/AKT signaling. Notably, LAPTM4B also promotes cancer cell proliferation via the PI3K/AKT signaling pathway. In addition, LAPTM4B can enhance and prolong EGFR signal transduction by blocking active EGFR intraluminal sorting and lysosomal degradation. Thus, LAPTM4B may be associated with autophagy through EGFR signaling. The present review proposed that LAPTM4B participates in regulating autophagy through the EGFR pathway.
    Keywords:  EGFR; LAPTM4B; autophagy
  18. Front Cell Dev Biol. 2022 ;10 811701
      Autophagy is pivotal in the maintenance of organelle function and intracellular nutrient balance. Besides the role of autophagy in the homeostasis and physiology of the individual tissues and whole organism in vivo, dysregulated autophagy has been incriminated in the pathogenesis of a variety of diseases including metabolic diseases, neurodegenerative diseases, cardiovascular diseases, inflammatory or immunological disorders, cancer and aging. Search for autophagy modulators has been widely conducted to amend dysregulation of autophagy or pharmacologically modulate autophagy in those diseases. Current data support the view that autophagy modulation could be a new modality for treatment of metabolic syndrome associated with lipid overload, human-type diabetes characterized by deposition of islet amyloid or other diseases including neurodegenerative diseases, infection and cardiovascular diseases. While clinically available bona fide autophagy modulators have not been developed yet, it is expected that on-going investigation will lead to the development of authentic autophagy modulators that can be safely administered to patients in the near future and will open a new horizon for treatment of incurable or difficult diseases.
    Keywords:  autophagy; endoplasmic reticulum; lysosome; metabolic diseases; mitochondria; modulator
  19. Neuron. 2022 Mar 02. pii: S0896-6273(22)00107-6. [Epub ahead of print]110(5): 735-737
      In this issue of Neuron, Yang et al. show that autophagy machinery is tightly coupled to neuronal activity via endocytic cycling of the transmembrane protein ATG-9 at presynaptic terminals.
  20. Autophagy. 2022 Mar 01. 1-17
      TFEB (transcription factor EB) and TFE3 (transcription factor binding to IGHM enhancer 3) orchestrate the cellular response to a variety of stressors, including nutrient deprivation, oxidative stress and pathogens. Here we describe a novel interaction of TFEB and TFE3 with the FAcilitates Chromatin Transcription (FACT) complex, a heterodimeric histone chaperone consisting of SSRP1 and SUPT16H that mediates nucleosome disassembly and assembly, thus facilitating transcription. Extracellular stimuli, such as nutrient deprivation or oxidative stress, induce nuclear translocation and activation of TFEB and TFE3, which then associate with the FACT complex to regulate stress-induced gene transcription. Depletion of FACT does not affect TFEB activation, stability, or binding to the promoter of target genes. In contrast, reduction of FACT levels by siRNA or treatment with the FACT inhibitor curaxin, severely impairs induction of numerous antioxidant and lysosomal genes, revealing a crucial role of FACT as a regulator of cellular homeostasis. Furthermore, upregulation of antioxidant genes induced by TFEB over-expression is significantly reduced by curaxin, consistent with a role of FACT as a TFEB transcriptional activator. Together, our data show that chromatin remodeling at the promoter of stress-responsive genes by FACT is important for efficient expression of TFEB and TFE3 targets, thus providing a link between environmental changes, chromatin modifications and transcriptional regulation.Abbreviations: ADNP2, ADNP homeobox 2; ATP6V0D1, ATPase H+ transporting V0 subunit d1; ATP6V1A, ATPase H+ transporting V1 subunit A; ATP6V1C1, ATPase H+ transporting V1 subunit C1; CSNK2/CK2, casein kinase 2; CLCN7, chloride voltage-gated channel 7; CTSD, cathepsin D; CTSZ, cathepsin Z; EBSS, earle's balanced salt solution; FACT complex, facilitates chromatin transcription complex; FOXO3, forkhead box O3; HEXA, hexosaminidase subunit alpha; HIF1A, hypoxia inducible factor 1 subunit alpha; HMOX1, heme oxygenase 1; LAMP1, lysosomal associated membrane protein 1; MAFF, MAF bZIP transcription factor F; MAFG, MAF bZIP transcription factor G; MCOLN1, mucolipin TRP cation channel 1; MTORC1, mechanistic target of rapamycin kinase complex 1; NaAsO2, sodium arsenite; POLR2, RNA polymerase II; PPARGC1A, PPARG coactivator 1 alpha; PYROXD1, pyridine nucleotide-disulfide oxidoreductase domain 1; RRAGC, Ras related GTP binding C; SEC13, SEC13 homolog, nuclear pore and COPII coat complex component; SLC38A9, solute carrier family 38 member 9; SSRP1, structure specific recognition protein 1; SUPT16H, SPT16 homolog, facilitates chromatin remodeling subunit; TFEB, transcription factor EB; TFE3, transcription factor binding to IGHM enhancer 3; TXNRD1, thioredoxin reductase 1; UVRAG, UV radiation resistance associated; WDR59, WD repeat domain 59.
    Keywords:  Autophagy; FACT; TFE3; TFEB; chaperone; histone; lysosomes
  21. Proc Natl Acad Sci U S A. 2022 Mar 08. 119(10): e2107357119
      Significance The mechanistic target of rapamycin (mTOR) plays a central role in growth, metabolism, and aging. It is assembled into two multiprotein complexes, namely, mTORC1 and mTORC2. We previously demonstrated the efficacy of sirolimus in ARHL in mice by decreasing mTORC1. However, the aspect of mTORC2 regulation in the cochlea is poorly characterized. Herein, based on pharmacological and genetic interventions, we found that a high dose of sirolimus resulted in severe hearing loss by reducing the mTORC2/AKT signaling pathway in the cochlea. Furthermore, selective activation of mTORC2 could protect against hearing loss induced by acoustic trauma and cisplatin-induced ototoxicity. Hence, the therapeutic activation of mTORC2 in conjunction with decreasing mTORC1 might represent a promising and effective strategy in preventing hearing loss.
    Keywords:  hair cells; hearing; mTORC2
  22. Autophagy. 2022 Feb 27. 1-15
      The mechanisms by which the ATG16L1T300A polymorphism affects cell function and causes an increased risk for the development of Crohn disease remain incompletely understood. Here we report that healthy individuals and mice bearing this polymorphism, even as heterozygotes, manifest enhanced TLR, and NLR cytokine and chemokine responses due to increased activation of NFKB. We elucidated the mechanism of the NFKB abnormality and found that in the ATG16L1T300A cell, there is enhanced polyubiquitination of TRAF6 or RIPK2 resulting from the accumulation of SQSTM1/p62. Indeed, knockout of Sqstm1 in autophagy-deficient cells almost completely normalized TRAF6 or RIPK2 polyubiquitination and NFKB activation in these cells. Thus, by identifying that autophagy is a pathway-intrinsic homeostatic mechanism that restricts excessive TLR- or NLR-mediated inflammatory signaling, our findings shed new light on how the ATG16L1T300A polymorphism sets the stage for the occurrence of Crohn disease.Abbreviations: 3-MA: 3-methyladenine; ATG16L1: autophagy related 16 like 1; ATG7: autophagy related 7; BMDM: bone marrow-derived macrophage; CD: Crohn disease; CXCL: C-X-C motif chemokine ligand; IBD: inflammatory bowel disease; iBMDM: immortalized mouse BMDM; IL1B/IL-1β: interleukin 1 beta; IL6: interleukin 6; KI: knockin; KO: knockout; MAP1LC3/LC3: microtubule associated protein 1 light chain 3; LPS: lipopolysaccharide; MDP: muramyl dipeptide; MEF: mouse embryonic fibroblast; NFKB/NF-κB: nuclear factor kappa B; NFKBIA/IKBA: NFKB inhibitor alpha; NLR: NOD-like receptor; NOD: nucleotide-binding oligomerization domain containing; RIPK2: receptor interacting serine/threonine kinase 2; SNP: single nucleotide polymorphism; SQSTM1/p62: sequestosome 1; TLR: toll like receptor; TNF/TNF-α: tumor necrosis factor; TRAF6: TNF receptor associated factor 6; Ub: ubiquitin; WT: wild type.
    Keywords:  ATG16L1T300A; NFKB; NLR; TLR4; autophagy
  23. Nat Commun. 2022 Mar 04. 13(1): 1172
      Hypoxia is a physiological stress that frequently occurs in solid tissues. Autophagy, a ubiquitous degradation/recycling system in eukaryotic cells, renders cells tolerant to multiple stressors. However, the mechanisms underlying autophagy initiation upon hypoxia remains unclear. Here we show that protein arginine methyltransferase 5 (PRMT5) catalyzes symmetrical dimethylation of the autophagy initiation protein ULK1 at arginine 170 (R170me2s), a modification removed by lysine demethylase 5C (KDM5C). Despite unchanged PRMT5-mediated methylation, low oxygen levels decrease KDM5C activity and cause accumulation of ULK1 R170me2s. Dimethylation of ULK1 promotes autophosphorylation at T180, a prerequisite for ULK1 activation, subsequently causing phosphorylation of Atg13 and Beclin 1, autophagosome formation, mitochondrial clearance and reduced oxygen consumption. Further, expression of a ULK1 R170K mutant impaired cell proliferation under hypoxia. This study identifies an oxygen-sensitive methylation of ULK1 with an important role in hypoxic stress adaptation by promoting autophagy induction.
  24. Curr Opin Pharmacol. 2022 Mar 01. pii: S1471-4892(22)00019-4. [Epub ahead of print]63 102193
      Despite evidence for prominent metabolic dysfunction within multiple sclerosis (MS) lesions, the mechanisms controlling metabolic shifts in oligodendroglia are poorly understood. The cuprizone model of demyelination and remyelination is a valuable tool for assessing metabolic insult during oligodendrocyte death and myelin degradation, closely resembling the distal oligodendrogliopathy seen in Pattern III MS lesions. In this review we discuss how metabolic processes in oligodendrocytes are disrupted in both MS and the cuprizone model, as well as the evidence for mechanistic target of rapamycin (mTOR) signaling as a key regulator of oligodendroglial metabolic function and efficient remyelination.
  25. Autophagy. 2022 Feb 27. 1-22
      Massive infiltrated and enriched decidual macrophages (dMφ) have been widely regarded as important regulators of maternal-fetal immune tolerance and trophoblast invasion, contributing to normal pregnancy. However, the characteristics of metabolic profile and the underlying mechanism of dMφ residence remain largely unknown. Here, we observe that dMφ display an active glycerophospholipid metabolism. The activation of ENPP2-lysophosphatidic acid (LPA) facilitates the adhesion and retention, and M2 differentiation of dMφ during normal pregnancy. Mechanistically, this process is mediated through activation of the LPA receptors (LPAR1 and PPARG/PPARγ)-DDIT4-macroautophagy/autophagy axis, and further upregulation of multiple adhesion factors (e.g., cadherins and selectins) in a CLDN7 (claudin 7)-dependent manner. Additionally, poor trophoblast invasion and placenta development, and a high ratio of embryo loss are observed in Enpp2±, lpar1-/- or PPARG-blocked pregnant mice. Patients with unexplained spontaneous abortion display insufficient autophagy and cell residence of dMφ. In therapeutic studies, supplementation with LPA or the autophagy inducer rapamycin significantly promotes dMφ autophagy and cell residence, and improves embryo resorption in Enpp2± and spontaneous abortion mouse models, which should be dependent on the activation of DDIT4-autophagy-CLDN7-adhesion molecules axis. This observation reveals that inactivation of ENPP2-LPA metabolism and insufficient autophagy of dMφ result in resident obstacle of dMφ and further increase the risk of spontaneous abortion, and provides potential therapeutic strategies to prevent spontaneous abortion.Abbreviations: ACTB: actin beta; ADGRE1/F4/80: adhesion G protein-coupled receptor E1; Atg5: autophagy related 5; ATG13: autophagy related 13; BECN1: beclin 1; CDH1/E-cadherin: cadherin 1; CDH5/VE-cadherin: cadherin 5; CFSE: carboxyfluorescein succinimidyl ester; CLDN7: claudin 7; CSF1/M-CSF: colony stimulating factor 1; CSF2/GM-CSF: colony stimulating factor 2; Ctrl: control; CXCL10/IP-10: chemokine (C-X-C) ligand 10; DDIT4: DNA damage inducible transcript 4; dMφ: decidual macrophage; DSC: decidual stromal cells; ENPP2/ATX: ectonucleotide pyrophosphatase/phosphodiesterase 2; Enpp2±: Enpp2 heterozygous knockout mouse; ENPP2i/PF-8380: ENPP2 inhibitor; EPCAM: epithelial cell adhesion molecule; ESC: endometrial stromal cells; FGF2/b-FGF: fibroblast growth factor 2; GAPDH: glyceraldehyde-3-phosphate dehydrogenase; GPCPD1: glycerophosphocholine phosphodiesterase 1; HE: heterozygote; HIF1A: hypoxia inducible factor 1 subunit alpha; HNF4A: hepatocyte nuclear factor 4 alpha; HO: homozygote; ICAM2: intercellular adhesion molecule 2; IL: interleukin; ITGAV/CD51: integrin subunit alpha V; ITGAM/CD11b: integrin subunit alpha M; ITGAX/CD11b: integrin subunit alpha X; ITGB3/CD61: integrin subunit beta 3; KLRB1/NK1.1: killer cell lectin like receptor B1; KRT7/cytokeratin 7: keratin 7; LPA: lysophosphatidic acid; LPAR: lysophosphatidic acid receptor; lpar1-/-: lpar1 homozygous knockout mouse; LPAR1i/AM966: LPAR1 inhibitor; LY6C: lymphocyte antigen 6 complex, locus C1; LYPLA1: lysophospholipase 1; LYPLA2: lysophospholipase 2; Lyz2: lysozyme 2; MAP1LC3B: microtubule associated protein 1 light chain 3 beta; MARVELD2: MARVEL domain containing 2; 3-MA: 3-methyladenine; MBOAT2: membrane bound O-acyltransferase domain containing 2; MGLL: monoglyceride lipase; MRC1/CD206: mannose receptor C-type 1; MTOR: mechanistic target of rapamycin kinase; NP: normal pregnancy; PDGF: platelet derived growth factor; PLA1A: phospholipase A1 member A; PLA2G4A: phospholipase A2 group IVA; PLPP1: phospholipid phosphatase 1; pMo: peripheral blood monocytes; p-MTOR: phosphorylated MTOR; PPAR: peroxisome proliferator activated receptor; PPARG/PPARγ: peroxisome proliferator activated receptor gamma; PPARGi/GW9662: PPARG inhibitor; PTPRC/CD45: protein tyrosine phosphatase receptor type, C; Rapa: rapamycin; RHEB: Ras homolog, mTORC1 binding; SA: spontaneous abortion; SELE: selectin E; SELL: selectin L; siCLDN7: CLDN7-silenced; STAT: signal transducer and activator of transcription; SQSTM1: sequestosome 1; TJP1: tight junction protein 1; VCAM1: vascular cell adhesion molecule 1; WT: wild type.
    Keywords:  Abortion; CLDN7; DDIT4; ENPP2; LPAR1; decidual macrophage; lysophosphatidic acid; trophoblast invasion
  26. Cell Rep. 2022 Mar 01. pii: S2211-1247(22)00173-5. [Epub ahead of print]38(9): 110446
      The factors that promote T cell expansion are not fully known. Creatine is an abundant circulating metabolite that has recently been implicated in T cell function; however, its cell-autonomous role in immune-cell function is unknown. Here, we show that creatine supports cell-intrinsic CD8+ T cell homeostasis. We further identify creatine kinase B (CKB) as the creatine kinase isoenzyme that supports these T cell properties. Loss of the creatine transporter (Slc6a8) or Ckb results in compromised CD8+ T cell expansion in response to infection without influencing adenylate energy charge. Rather, loss of Slc6a8 or Ckb disrupts naive T cell homeostasis and weakens TCR-mediated activation of mechanistic target of rapamycin complex 1 (mTORC1) signaling required for CD8+ T cell expansion. These data demonstrate a cell-intrinsic role for creatine transport and creatine transphosphorylation, independent of their effects on global cellular energy charge, in supporting CD8+ T cell homeostasis and effector function.
    Keywords:  CD8+ T cells; adoptive transfer; creatine kinase; creatine metabolism; infection
  27. Front Physiol. 2021 ;12 791691
      Throughout mammal erythroid differentiation, erythroblasts undergo enucleation and organelle clearance becoming mature red blood cell. Organelles are cleared by autophagic pathways non-specifically targeting organelles and cytosolic content or by specific mitophagy targeting mitochondria. Mitochondrial functions are essential to coordinate metabolism reprogramming, cell death, and differentiation balance, and also synthesis of heme, the prosthetic group needed in hemoglobin assembly. In mammals, mitochondria subcellular localization and mitochondria interaction with other structures as endoplasmic reticulum and nucleus might be of importance for the removal of the nucleus, that is, the enucleation. Here, we aim to characterize by electron microscopy the changes in ultrastructure of cells over successive stages of human erythroblast differentiation. We focus on mitochondria to gain insights into intracellular localization, ultrastructure, and contact with other organelles. We found that mitochondria are progressively cleared with a significant switch between PolyE and OrthoE stages, acquiring a rounded shape and losing contact sites with both ER (MAM) and nucleus (NAM). We studied intracellular vesicle trafficking and found that endosomes and MVBs, known to be involved in iron traffic and heme synthesis, are increased during BasoE to PolyE transition; autophagic structures such as autophagosomes increase from ProE to OrthoE stages. Finally, consistent with metabolic switch, glycogen accumulation was observed in OrthoE stage.
    Keywords:  autophagy; electron microscopy; erythropoiesis; mitochondria; vesicles
  28. Autophagy. 2022 Feb 27. 1-14
      Defective mitophagy contributes to normal aging and various neurodegenerative and cardiovascular diseases. The newly developed methodologies to visualize and quantify mitophagy allow for additional progress in defining the pathophysiological significance of mitophagy in various model organisms. However, current knowledge regarding mitophagy relevant to human physiology is still limited. Model organisms such as mice might not be optimal models to recapitulate all the key aspects of human disease phenotypes. The development of the human-induced pluripotent stem cells (hiPSCs) may provide an exquisite approach to bridge the gap between animal mitophagy models and human physiology. To explore this premise, we take advantage of the pH-dependent fluorescent mitophagy reporter, mt-Keima, to assess mitophagy in hiPSCs and hiPSC-derived cardiomyocytes (hiPSC-CMs). We demonstrate that mt-Keima expression does not affect mitochondrial function or cardiomyocytes contractility. Comparison of hiPSCs and hiPSC-CMs during different stages of differentiation revealed significant variations in basal mitophagy. In addition, we have employed the mt-Keima hiPSC-CMs to analyze how mitophagy is altered under certain pathological conditions including treating the hiPSC-CMs with doxorubicin, a chemotherapeutic drug well known to cause life-threatening cardiotoxicity, and hypoxia that stimulates ischemia injury. We have further developed a chemical screening to identify compounds that modulate mitophagy in hiPSC-CMs. The ability to assess mitophagy in hiPSC-CMs suggests that the mt-Keima hiPSCs should be a valuable resource in determining the role mitophagy plays in human physiology and hiPSC-based disease models. The mt-Keima hiPSCs could prove a tremendous asset in the search for pharmacological interventions that promote mitophagy as a therapeutic target.
    Keywords:  Cardiomyocytes; cardiomyopathy; induced pluripotent stem cells; mitochondrial; mitophagy; mt-Keima
  29. STAR Protoc. 2022 Mar 18. 3(1): 101018
      Following lysosomal damage, activation and nuclear translocation of transcription factor EB (TFEB) is the key event to maintain lysosomal homeostasis. Here, we describe steps to induce lysosomal damage in HeLa cells. This can be followed by monitoring the changes in TFEB localization using widefield fluorescence microscopy. As a complementary approach, we describe the use of immunoblotting to follow the activation and localization of TFEB in cell lysates. These protocols enable quantitative analysis of TFEB. For complete details on the use and execution of this protocol, please refer to Nakamura et al. (2020).
    Keywords:  Antibody; Cell Biology; Cell culture; Cell separation/fractionation; Cell-based Assays; Microscopy
  30. Biomed Pharmacother. 2022 Feb 24. pii: S0753-3322(22)00115-9. [Epub ahead of print]148 112727
      Autophagy is an essential catabolic process in mammalian cells to maintain cellular integrity and viability by degrading the old and damaged cell organelles and other contents with the help of lysosomes. Deregulation in autophagy can be one of the major contributors leading to the continuous cell proliferation and development of tumors. Tetrandrine, a bisbenzylisoquinoline alkaloid known to have potent bioactivities such as anticancer, antimicrobial, anti-inflammatory, antidiabetic, antioxidant, immunosuppressive, cardiovascular, and calcium channel blocking effects. The present review evaluated the effectiveness of tetrandrine in targeting key proteins in the autophagy pathway to induce anticancer effect based on the available literature. An attempt is also made to understand the influence of tetrandrine in regulating autophagy by mTOR dependant and mTOR-independent pathways. In addition, the review also highlights the limitations involved and future perspectives in developing tetrandrine as a chemotherapeutic drug to treat cancer.
    Keywords:  Autophagy; Beclin-1; Cancer; MTOR inhibitor; Tetrandrine
  31. Autophagy. 2022 Mar 03. 1-17
      SARS-CoV-2 infections have resulted in a very large number of severe cases of COVID-19 and deaths worldwide. However, knowledge of SARS-CoV-2 infection, pathogenesis and therapy remains limited, emphasizing the urgent need for fundamental studies and drug development. Studies have shown that induction of macroautophagy/autophagy and hijacking of the autophagic machinery are essential for the infection and replication of SARS-CoV-2; however, the mechanism of this manipulation and the function of autophagy during SARS-CoV-2 infection remain unclear. In the present study, we identified ORF3a as an inducer of autophagy (in particular reticulophagy) and revealed that ORF3a localizes to the ER and induces RETREG1/FAM134B-related reticulophagy through the HMGB1-BECN1 (beclin 1) pathway. As a consequence, ORF3a induces ER stress and inflammatory responses through reticulophagy and then sensitizes cells to the acquisition of an ER stress-related early apoptotic phenotype and facilitates SARS-CoV-2 infection, suggesting that SARS-CoV-2 ORF3a hijacks reticulophagy and then disrupts ER homeostasis to induce ER stress and inflammatory responses during SARS-CoV-2 infection. These findings reveal the sequential induction of reticulophagy, ER stress and acute inflammatory responses during SARS-CoV-2 infection and imply the therapeutic potential of reticulophagy and ER stress-related drugs for COVID-19.
    Keywords:  ER stress; ORF3a; SARS-CoV-2; inflammatory response; reticulophagy
  32. J Exp Bot. 2022 Mar 05. pii: erac002. [Epub ahead of print]
      Physiological effects mediated by melatonin are attributable to its potent antioxidant activity as well as its role as a signaling molecule in inducing a vast array of melatonin-mediated genes. Here, we propose melatonin as a signaling molecule essential for protein quality control (PQC) in plants. PQC occurs by the coordinated activities of three systems: the chaperone network, autophagy, and the ubiquitin-proteasome system. With regard to the melatonin-mediated chaperone pathway, melatonin increases thermotolerance by induction of heat shock proteins and confers endoplasmic reticulum stress tolerance by increasing endoplasmic reticulum chaperone proteins. In chloroplasts, melatonin-induced chaperones, including Clps and CpHSP70s, play key roles in the PQC of chloroplast-localized proteins, such as Lhcb1, Lhcb4, and RBCL, during growth. Melatonin regulates PQC by autophagy processes, in which melatonin induces many autophagy (ATG) genes and autophagosome formation under stress conditions. Finally, melatonin-mediated plant stress tolerance is associated with up-regulation of stress-induced transcription factors, which are regulated by the ubiquitin-proteasome system. In this review, we propose that melatonin plays a pivotal role in PQC and consequently functions as a pleiotropic molecule under non-stress and adverse conditions in plants.
    Keywords:  AAA+ family of ATPase caseinolytic proteases; autophagosome; endoplasmic reticulum stress; heat shock proteins; melatonin; mitogen-activated protein kinase; ubiquitin ligases; unfolded protein response
  33. Autophagy. 2022 Feb 27. 1-12
      Mutations in the mitochondrial genome (mtDNA) are ubiquitous in humans and can lead to a broad spectrum of disorders. However, due to the presence of multiple mtDNA molecules in the cell, co-existence of mutant and wild-type mtDNAs (termed heteroplasmy) can mask disease phenotype unless a threshold of mutant molecules is reached. Importantly, the mutant mtDNA level can change across lifespan as mtDNA segregates in an allele- and cell-specific fashion, potentially leading to disease. Segregation of mtDNA is mainly evident in hepatic cells, resulting in an age-dependent increase of mtDNA variants, including non-synonymous potentially deleterious mutations. Here we modeled mtDNA segregation using a well-established heteroplasmic mouse line with mtDNA of NZB/BINJ and C57BL/6N origin on a C57BL/6N nuclear background. This mouse line showed a pronounced age-dependent NZB mtDNA accumulation in the liver, thus leading to enhanced respiration capacity per mtDNA molecule. Remarkably, liver-specific atg7 (autophagy related 7) knockout abolished NZB mtDNA accumulat ion, resulting in close-to-neutral mtDNA segregation through development into adulthood. prkn (parkin RBR E3 ubiquitin protein ligase) knockout also partially prevented NZB mtDNA accumulation in the liver, but to a lesser extent. Hence, we propose that age-related liver mtDNA segregation is a consequence of macroautophagic clearance of the less-fit mtDNA. Considering that NZB/BINJ and C57BL/6N mtDNAs have a level of divergence comparable to that between human Eurasian and African mtDNAs, these findings have potential implications for humans, including the safe use of mitochondrial replacement therapy.
    Keywords:  Atg7; NZB; heteroplasmy; mitochondria; mitophagy; parkin
  34. Biomed Pharmacother. 2022 Mar 01. pii: S0753-3322(22)00161-5. [Epub ahead of print]148 112773
      Growing evidence suggests that neuronal dysfunction in the endo-lysosomal and autophagic processes contributes to the onset and progression of neurodegenerative diseases such as Alzheimer's disease (AD). Since they are the primary cellular systems involved in the production and clearance of aggregated amyloid plaques, endo-lysosomal or autophagic equilibrium must be maintained throughout life. As a result, variations in the autophagic and endo-lysosomal torrent, as a measure of degenerative function in these sections or pathways, may have a direct impact on disease-related processes, such as Aß clearance from the brain and interneuronal deposition of Aß and tau aggregates, thus disrupting synaptic plasticity. The discovery of several chromosomal factors for Alzheimer's disease that are clinically linked to regulation of the endocytic pathway, including protein aggregation and removal, supports the theory that the endo-lysosomal/autophagic torrent is more susceptible to impairment, especially as people age, thus catalysing the onset of disease. Although the role of endo-lysosomal/autophagic dysfunction in neurodegeneration has progressed in recent years, the field remains underdeveloped. Because of its possible therapeutic implications in Alzheimer's disease, further study is needed to explain the possibilities for effective autophagy regulation.
    Keywords:  Alzheimer’s Disease; Endo-lysosomal; Neurology; Rab5 Protein
  35. Acta Mater Med. 2022 ;1(1): 24-41
      Neurodegenerative diseases (NDs) are characteristic with progression of neuron degeneration, resulting in dysfunction of cognition and mobility. Many neurodegenerative diseases are because of proteinopathies that results from unusual protein accumulations and aggregations. The aggregation of misfolded proteins like β-amyloid, α-synuclein, tau, and polyglutamates are hallmarked in Alzheimer's disease (AD), which are undruggable targets, and usually do not respond to conventional small-molecule agents. Therefore, developing novel technology and strategy for reducing the levels of protein aggregates would be critical for treatment of AD. Recently, the emerging proteolysis targeting chimeras (PRPTACs) technology has been significantly considered for artificial and selective degradation of aberrant target proteins. These engineered bifunctional molecules engage target proteins to be degraded by either the cellular degradation machinery in the ubiquitin-proteasome system (UPS) or via the autophagy-lysosome degradation pathway. Although the application of PROTACs technology is preferable than oligonucleotide and antibodies for treatment of NDs, many limitations such as their pharmacokinetic properties, tissue distribution and cell permeabilities, still need to be corrected. Herein, we review the recent advances in PROTACs technology with their limitation for pharmaceutical targeting of aberrant proteins involved in Alzheimer's diseases. We also review therapeutic potential of dysregulated signaling such as PI3K/AKT/mTOR axis for the management of AD.
    Keywords:  Alzheimer’s disease; Autophagy; PROTAC; protein degradation; ubiquitin-proteasome system
  36. ACS Chem Neurosci. 2022 Mar 01.
      α-Synuclein accumulation is implicated in the pathogenesis of neurodegenerative diseases, including Parkinson's disease (PD). Previously, we reported that Fas-associated factor 1 (FAF1), which plays a role in PD pathogenesis, potentiates α-synuclein accumulation through autophagy impairment in dopaminergic neurons. In this study, we show that KM-819, a FAF1-targeting compound, which has completed phase I clinical trials, interferes with α-synuclein accumulation in the mouse brain, as well as in human neuronal cells (SH-SY5Ys). KM-819 suppressed the accumulation of monomeric, oligomeric, and aggregated forms of α-synuclein in neuronal cells. Furthermore, KM-819 restored the turnover rate of α-synuclein in FAF1-overexpressing SH-SY5Y cells, implicating KM-819-mediated reconstitution of the α-synuclein degradative pathway. In addition, KM-819 reconstituted autophagic flux in FAF1-transfected SH-SY5Y cells, also suppressing α-synuclein-induced mitochondrial dysfunction. Moreover, oral administration of KM-819 also interfered with α-synuclein accumulation in the midbrain of mice overexpressing FAF1 via an adeno-associated virus system. Consistently, KM-819 reduced α-synuclein accumulation in both the hippocampus and the midbrain of human A53T α-synuclein transgenic mice. Collectively, these data imply that KM-819 may have therapeutic potential for patients with PD.
    Keywords:  A53T α-synuclein transgenic mice.; FAF1; KM-819; Parkinson’s disease; autophagy; α-synuclein
  37. Toxicol Res (Camb). 2022 Feb;11(1): 52-59
      NaAsO2-induced liver damage leads to autophagy, which plays an important role in cell quality control. Mitophagy plays an important role in hepatocyte damage, and PINK1 and Parkin constitute an important pathway in mitophagy. PINK1 selectively degrades abnormal mitochondria, and Parkin can recognize damaged mitochondria. However, the mechanism underlying the involvement of PINK1/Parkin in NaAsO2-induced mitophagy is unclear. Transfection plasmids containing dsRNA were used to interfere with the expression of Parkin in the following groups: the empty plasmid group was established by add the empty plasmid only, the PINK1-knockdown (KD) group was established by adding 5 μg of PINK1 dsRNA and then by adding 10 mM NaAsO2, and the Parkin-KD group was established by adding 5 μg of Parkin dsRNA and then by adding 10 mM NaAsO2. The expression of PINK1 and Parkin in autophagy was detected by western blotting and immunofluorescence staining. The ultrastructures of autophagosomes and mitochondria were observed by transmission electron microscopy. The successful KD of PINK1 and Parkin aggravated the NaAsO2-induced damage to mitophagy. The degeneration of mitochondrial vacuoles and the appearance of autophagosomes were detected in the NaAsO2, NaAsO2 + PINK1-KD and NaAsO2 + Parkin-KD groups. NaAsO2 can induce mitophagy in rat hepatocytes, and the silencing of PINK1 and Parkin can aggravate mitochondrial damage during this process. This study explored the mechanism of NaAsO2-induced mitophagy in BRL-3A cells after PINK1 and Parkin gene silencing.
    Keywords:  PINK1; Parkin; mitophagy; sodium arsenite
  38. Mol Biol Rep. 2022 Mar 02.
      PURPOSE: Cellular responses following cerebral ischemia/reperfusion injury are critical to recovery and survival after ischemic stroke. Understanding of these cellular responses can help the design of therapies to protect brain tissue and promote recovery after stroke. One of these cellular responses may be mediated by the AKT (protein kinase B) signal transduction pathway. This study was aimed to investigate the cerebral ischemia-induced alterations of AKT signaling and the upstream molecular pathways.METHODS: We modeled cerebral ischemia by middle cerebral artery occlusion in 2-3-month-old male C57BL/6J mice and then analyze the brain samples by using quantitative Western blots and phosphorylation/activation-dependent kinase antibodies. Cerebral ischemia was confirmed by staining of brain slices with 1% 2,3,5-triphenyltetrazolium chloride (TTC) and Nissl, as well as neurological assessments of the mice 24 h after ischemia-reperfusion surgery.
    RESULTS: We found marked downregulation of AKT within 12 h of cerebral ischemia/reperfusion, which leads to overactivation of glycogen synthase kinase-3β (GSK-3β). Furthermore, we found that the downregulation of AKT was mediated by downregulation of mTORC2 (the complex 2 of the mechanistic target of rapamycin) instead of its common upstream kinases, phosphatidylinositol 3-kinase and phosphoinositide-dependent kinase-1.
    CONCLUSION: Our findings provide new insight into the cellular responses to ischemia/reperfusion brain injury and will help develop new treatments targeting the AKT signaling pathway for the treatment of ischemic stroke.
    Keywords:  Cerebral ischemia/reperfusion; GSK-3β; Middle cerebral artery occlusion; Protein kinase B (AKT); Signal transduction; mTOR
  39. Brain. 2022 Mar 04. pii: awab303. [Epub ahead of print]
      Mitochondria are essential organelles found in every eukaryotic cell, required to convert food into usable energy. Therefore, it is not surprising that mutations in either mtDNA or nuclear DNA-encoded genes of mitochondrial proteins cause diseases affecting the oxidative phosphorylation system, which are heterogeneous from a clinical, genetic, biochemical and molecular perspective and can affect patients at any age. Despite all this, it is surprising that our understanding of the mechanisms governing mitochondrial gene expression and its associated pathologies remain superficial and therapeutic interventions largely unexplored. We recently showed that loss of the mitochondrial matrix protease caseinolytic protease proteolytic subunit (CLPP) ameliorates phenotypes in cells characterized by defects in oxidative phosphorylation maintenance. Here, we build upon this finding by showing that CLPP depletion is indeed beneficial in vivo for various types of neuronal populations, including Purkinje cells in the cerebellum and cortical and hippocampal neurons in the forebrain, as it strongly improves distinct phenotypes of mitochondria encephalopathy, driven by the deficiency of the mitochondrial aspartyl tRNA synthase DARS2. In the absence of CLPP, neurodegeneration of DARS2-deficient neurons is delayed as they present milder oxidative phosphorylation dysfunction. This in turn leads to a decreased neuroinflammatory response and significantly improved motor functions in both double-deficient models (Purkinje cell-specific or forebrain neuron-specific Dars2/Clpp double knockout mice). We propose that diminished turnover of respiratory complex I caused by the loss of CLPP is behind the improved phenotype in Dars2/Clpp double knockout animals, even though this intervention might not restore respiratory complex I activity but rather improve mitochondrial cristae morphology or help maintain the NAD+/NADH ratio inside mitochondria. These results also open the possibility of targeting CLPP activity in many other mitochondrial encephalopathies characterized by respiratory complex I instability.
    Keywords:  CLPP protease; DARS2 deficiency; LBSL; mitochondrial diseases