bims-auttor Biomed News
on Autophagy and mTOR
Issue of 2022‒02‒06
57 papers selected by
Viktor Korolchuk, Newcastle University
  1. Leucine retention in lysosomes is regulated by starvation.
  2. ER-phagy requires the assembly of actin at sites of contact between the cortical ER and endocytic pits.
  3. Defects in the autophagy lysosomal pathway in a cell model of disease-associated tau.
  4. OTUD7B deubiquitinates SQSTM1/p62 and promotes IRF3 degradation to regulate antiviral immunity.
  5. FK506-binding protein-like and FK506-binding protein 8 regulate dual leucine zipper kinase degradation and neuronal responses to axon injury.
  6. Autophagy takes it all - autophagy inducers target immune aging.
  7. Perinatal versus adult loss of ULK1 and ULK2 distinctly influences cardiac autophagy and function.
  8. Architecture of the endoplasmic reticulum plays a role in proteostasis.
  9. TDP-43 pathology: from noxious assembly to therapeutic removal.
  10. E3 ligase RNF167 and deubiquitinase STAMBPL1 modulate mTOR and cancer progression.
  11. Homocysteine Suppresses Autophagy via AMPK-mTOR-TFEB Signaling in Human THP-1 Macrophages.
  12. Rapamycin ameliorates age-related hearing loss in C57BL/6J mice by enhancing autophagy in the SGNs.
  13. Deficiency of the lysosomal exonuclease PLD3 impacts the degradative route.
  14. Mapping Critical Residues in ATG11's Coiled-Coil 2 Domain that Block Multiple Interactions and Disrupt Selective Autophagy.
  15. Autophagy-related DjAtg1-1 plays critical role in planarian regeneration by regulating proliferation and cell death.
  16. Dendritic autophagy degrades postsynaptic proteins and is required for long-term synaptic depression in mice.
  17. FUNDC1: A Promising Mitophagy Regulator at the Mitochondria-Associated Membrane for Cardiovascular Diseases.
  18. ALFY localizes to early endosomes and cellular protrusions to facilitate directional cell migration.
  19. Stressed and disoriented: stromal autophagy regulates HSCs.
  20. Activation of autophagy attenuates motor deficits and extends lifespan in a C. elegans model of ALS.
  21. Living in α-syn: Tackling aggregates in Parkinson's disease.
  22. LAMTOR1 inhibition of TRPML1-dependent lysosomal calcium release regulates dendritic lysosome trafficking and hippocampal neuronal function.
  23. Vps501, a novel vacuolar SNX-BAR protein cooperates with the SEA complex to regulate TORC1 signaling.
  24. Synaptic stimulation induces tau clearance by enhancing autophagosomal/lysosomal degradation.
  25. The dual roles of autophagy and the GPCRs-mediating autophagy signaling pathway after cerebral ischemic stroke.
  26. LRRK2, GBA and their interaction in the regulation of autophagy: implications on therapeutics in Parkinson's disease.
  27. Suppression of PI3K signaling is linked to autophagy activation and the spatiotemporal induction of the lens organelle free zone.
  28. Environmental control of Pub1 (NEDD4 family E3 ligase) in Schizosaccharomyces pombe is regulated by TORC2 and Gsk3.
  29. Mammalian AKT, the Emerging Roles on Mitochondrial Function in Diseases.
  30. The importance of mitochondrial quality control for maintaining skeletal muscle function across healthspan.
  31. PI3K/Akt/mTOR pathway: a potential target for anti-SARS-CoV-2 therapy.
  32. VPS35 down regulation alters degradation pathways in neuronal cells.
  33. Apigenin Alleviates Intervertebral Disc Degeneration via Restoring Autophagy Flux in Nucleus Pulposus Cells.
  34. Beyond Protein Synthesis; The Multifaceted Roles of Tuberin in Cell Cycle Regulation.
  35. DNAJC5 affects the endo-lysosomal pathway, APP processing, and AD pathology in vitro and in vivo.
  36. Neuroprogenitor Cells From Patients With TBCK Encephalopathy Suggest Deregulation of Early Secretory Vesicle Transport.
  37. The role of mitophagy during oocyte aging in human, mouse, and Drosophila: implications for oocyte quality and mitochondrial disease.
  38. mTORC1 inhibitor effects on rapid ketamine-induced reductions in suicidal ideation in patients with treatment-resistant depression.
  39. Quantitative and Time-Resolved Monitoring of Organelle and Protein Delivery to the Lysosome with A Tandem Fluorescent Halo-GFP reporter.
  40. Modulation of histone deacetylase, the ubiquitin proteasome system, and autophagy underlies the neuroprotective effects of venlafaxine in a rotenone-induced Parkinson's disease model in rats.
  41. Targeting lipophagy as a potential therapeutic strategy for nonalcoholic fatty liver disease.
  42. Acetylshikonin induces autophagy-dependent apoptosis through the key LKB1-AMPK and PI3K/Akt-regulated mTOR signalling pathways in HL-60 cells.
  43. Significant impact of mTORC1 and ATF4 pathways in CHO cell recombinant protein production induced by CDK4/6 inhibitor.
  44. Mitochondria homeostasis: Biology and involvement in hepatic steatosis to NASH.
  45. Unraveling and targeting RAS-driven metabolic signaling for therapeutic gain.
  46. MEK/ERK-mediated oncogenic signals promote secretion of extracellular vesicles by controlling lysosome function.
  47. The interplay between oxidative stress and autophagy: focus on the development of neurological diseases.
  48. Protection of Quiescence and Longevity of IgG Memory B Cells by Mitochondrial Autophagy.
  49. Oridonin Alleviates LPS-Induced Depression by Inhibiting NLRP3 Inflammasome via Activation of Autophagy.
  50. Repeated social defeat stress inhibits development of hippocampus neurons through mitophagy and autophagy.
  51. Roles of humanin and derivatives on the pathology of neurodegenerative diseases and cognition.
  52. Involvement of autophagy in exacerbation of eosinophilic airway inflammation in a murine model of obese asthma.
  53. A farnesyltransferase inhibitor restores cognitive deficits in Tsc2+/- mice through inhibition of Rheb1.
  54. Differential gene expression analysis for the identification of potential biomarkers for Niemann-Pick type C disease.
  55. Dual Inhibition of mTORC1/2 Reduces Migration of Cholangiocarcinoma Cells by Regulation of Matrixmetalloproteinases.
  56. Lysosomal subcellular distribution in neurons: Are there synaptic lysosomes?
  57. The Effects of Combined Therapy With Metformin and Hydroxypropyl-β-Cyclodextrin in a Mouse Model of Niemann-Pick Disease Type C1.
  1. Proc Natl Acad Sci U S A. 2022 Feb 08. pii: e2114912119. [Epub ahead of print]119(6):
    Leucine retention in lysosomes is regulated by starvation.
    Urmi Bandyopadhyay, Pavlina Todorova, Natalya N Pavlova, Yuma Tada, Craig B Thompson, Lydia W S Finley, Michael Overholtzer.
      Cells acquire essential nutrients from the environment and utilize adaptive mechanisms to survive when nutrients are scarce. How nutrients are trafficked and compartmentalized within cells and whether they are stored in response to stress remain poorly understood. Here, we investigate amino acid trafficking and uncover evidence for the lysosomal transit of numerous essential amino acids. We find that starvation induces the lysosomal retention of leucine in a manner requiring RAG-GTPases and the lysosomal protein complex Ragulator, but that this process occurs independently of mechanistic target of rapamycin complex 1 activity. We further find that stored leucine is utilized in protein synthesis and that inhibition of protein synthesis releases lysosomal stores. These findings identify a regulated starvation response that involves the lysosomal storage of leucine.
    Keywords:  leucine; lysosome; mTOR
    DOI:  https://doi.org/10.1073/pnas.2114912119
  2. Proc Natl Acad Sci U S A. 2022 Feb 08. pii: e2117554119. [Epub ahead of print]119(6):
    ER-phagy requires the assembly of actin at sites of contact between the cortical ER and endocytic pits.
    Dongmei Liu, Muriel Mari, Xia Li, Fulvio Reggiori, Susan Ferro-Novick, Peter Novick.
      Fragments of the endoplasmic reticulum (ER) are selectively delivered to the lysosome (mammals) or vacuole (yeast) in response to starvation or the accumulation of misfolded proteins through an autophagic process known as ER-phagy. A screen of the Saccharomyces cerevisiae deletion library identified end3Δ as a candidate knockout strain that is defective in ER-phagy during starvation conditions, but not bulk autophagy. We find that loss of End3 and its stable binding partner Pan1, or inhibition of the Arp2/3 complex that is coupled by the End3-Pan1 complex to endocytic pits, blocks the association of the cortical ER autophagy receptor, Atg40, with the autophagosomal assembly scaffold protein Atg11. The membrane contact site module linking the rim of cortical ER sheets and endocytic pits, consisting of Scs2 or Scs22, Osh2 or Osh3, and Myo3 or Myo5, is also needed for ER-phagy. Both Atg40 and Scs2 are concentrated at the edges of ER sheets and can be cross-linked to each other. Our results are consistent with a model in which actin assembly at sites of contact between the cortical ER and endocytic pits contributes to ER sequestration into autophagosomes.
    Keywords:  actin; autophagy; endoplasmic reticulum
    DOI:  https://doi.org/10.1073/pnas.2117554119
  3. Alzheimers Dement. 2021 Dec;17 Suppl 3 e051303
    Defects in the autophagy lysosomal pathway in a cell model of disease-associated tau.
    Saskia J Pollack, Dina Dakkak, Tong Guo, Patricia Gómez-Suaga, Wendy Noble, Diane P Hanger.
      BACKGROUND: Intracellular accumulation of insoluble tau is an important hallmark of Alzheimer's disease (AD) and related tauopathies. We have previously identified in human tauopathy brain a truncated tau species (Tau35), comprising the C-terminal half with four microtubule-binding repeats. Minimal Tau35 expression in transgenic mice results in a progressive tauopathy phenotype including tau phosphorylation and aggregation, cognitive and behavioural abnormalities and impaired protein clearance. The autophagy-lysosomal pathway (ALP) plays a crucial role in the clearance of protein aggregates and defects in ALP are associated with the pathogenesis of AD. We sought to explore the effect of Tau35 expression on the ALP and whether autophagy is disrupted due to lysosomal dysfunction.METHODS: Chinese hamster ovary (CHO) cells stably expressing Tau35 (CHO-Tau35) or full-length human 2N4R tau (CHO-FL) were generated. Primary cortical neurons from Tau35 transgenic and wild-type mice were cultured for 14 days in vitro(DIV) and brain homogenates were prepared from mice aged 4 and 12 months. The effect of Tau35 on the ALP was examined using immunofluorescence and western blots.
    RESULT: Nuclear translocation of transcription factor EB (TFEB), a key mediator of lysosomal biogenesis, was significantly reduced in both CHO-FL and CHO-Tau35 cells. However, only CHO-Tau35 cells exhibited disrupted mammalian target of rapamycin complex 1 (mTORC1) activity and autophagic flux. Expression of ALP markers, including LC3-I/II, LAMP1, LAMP2 and cathepsin D, were also reduced in CHO-Tau35 cells and in post-symptomatic Tau35 transgenic mice.
    CONCLUSIONS: Our findings suggest that N-terminally cleaved tau damages both lysosomal clearance of cellular proteins and lysosomal biogenesis. The Tau35-expressing cultured neurons will provide a useful tool to explore molecular mechanisms underlying tau-induced lysosomal dysfunction, which may lead to the identification of novel therapeutic targets for dementia.
    DOI:  https://doi.org/10.1002/alz.051303
  4. Autophagy. 2022 Jan 31. 1-15
    OTUD7B deubiquitinates SQSTM1/p62 and promotes IRF3 degradation to regulate antiviral immunity.
    Weihong Xie, Shuo Tian, Jiahui Yang, Sihui Cai, Shouheng Jin, Tao Zhou, Yaoxing Wu, Zhiyun Chen, Yanqin Ji, Jun Cui.
      Deubiquitination plays an important role in the regulation of the crosstalk between macroautophagy/autophagy and innate immune signaling, yet its regulatory mechanisms are not fully understood. Here we identify the deubiquitinase OTUD7B as a negative regulator of antiviral immunity by targeting IRF3 (interferon regulatory factor 3) for selective autophagic degradation. Mechanistically, OTUD7B interacts with IRF3, and activates IRF3-associated cargo receptor SQSTM1/p62 (sequestosome 1) by removing its K63-linked poly-ubiquitin chains at lysine 7 (K7) to enhance SQSTM1 oligomerization. Moreover, viral infection increased the expression of OTUD7B, which forms a negative feedback loop by promoting IRF3 degradation to balance type I interferon (IFN) signaling. Taken together, our study reveals a specific role of OTUD7B in mediating the activation of cargo receptors in a substrate-dependent manner, which could be a potential target against excessive immune responses.Abbreviations: Baf A1: bafilomycin A1; CGAS: cyclic GMP-AMP synthase; DDX58/RIG-I: DExD/H-box helicase 58; DSS: dextran sodium sulfate; DUBs: deubiquitinating enzymes; GFP: green fluorescent protein; IFN: interferon; IKKi: IKBKB/IkappaB kinase inhibitor; IRF3: interferon regulatory factor 3; ISGs: interferon-stimulated genes; MAVS: mitochondrial antiviral signaling protein; MOI: multiplicity of infection; PAMPs: pathogen-associated molecular patterns; SeV: Sendai virus; siRNA: small interfering RNA; SQSTM1/p62: sequestosome 1; STING1: stimulator of interferon response cGAMP interactor 1; TBK1: TANK binding kinase 1; Ub: ubiquitin; WT: wild-type; VSV: vesicular stomatitis virus.
    Keywords:  Antiviral immunity; cargo receptor; deubiquitination; selective autophagy; type I interferon signaling
    DOI:  https://doi.org/10.1080/15548627.2022.2026098
  5. J Biol Chem. 2022 Jan 28. pii: S0021-9258(22)00087-4. [Epub ahead of print] 101647
    FK506-binding protein-like and FK506-binding protein 8 regulate dual leucine zipper kinase degradation and neuronal responses to axon injury.
    Bohm Lee, Yeonsoo Oh, Eunhye Cho, Aaron DiAntonio, Valeria Cavalli, Jung Eun Shin, Hae Woong Choi, Yongcheol Cho.
      The dual leucine zipper kinase (DLK) is a key regulator of axon regeneration and degeneration in response to neuronal injury; however, regulatory mechanisms of the DLK function via its interacting proteins are largely unknown. To better understand the molecular mechanism of DLK function, we performed yeast two-hybrid screening analysis and identified FK506-binding protein-like (FKBPL, also known as WAF-1/CIP1 stabilizing protein 39) as a DLK-binding protein. FKBPL binds to the kinase domain of DLK and inhibits its kinase activity. In addition, FKBPL induces DLK protein degradation through ubiquitin-dependent pathways. We further assessed other members in the FKBP protein family and found that FK506-binding protein 8 (FKBP8) also induced DLK degradation. We identified the lysine 271 residue in the kinase domain as a major site of DLK ubiquitination and SUMO3 conjugation, and was thus responsible for regulating FKBP8-mediated proteasomal degradation that was inhibited by the substitution of the lysine 271 to arginine. FKBP8-mediated degradation of DLK is mediated by autophagy pathway because knockdown of Atg5 inhibited DLK destabilization. We show that in vivo overexpression of FKBP8 delayed the progression of axon degeneration and suppressed neuronal death following axotomy in sciatic and optic nerves. Taken together, this study identified FKBPL and FKBP8 as novel DLK-interacting proteins that regulate DLK stability via the ubiquitin-proteasome and lysosomal protein degradation pathways.
    Keywords:  DLK; FKBP8; FKBPL; MAP3K12; SUMOylation; axon degeneration; ubiquitination
    DOI:  https://doi.org/10.1016/j.jbc.2022.101647
  6. Dis Model Mech. 2022 01 01. pii: dmm049345. [Epub ahead of print]15(1):
    Autophagy takes it all - autophagy inducers target immune aging.
    Heidi Zinecker, Anna Katharina Simon.
      Autophagy, as the key nutrient recycling pathway, enables eukaryotic cells to adapt to surging cellular stress during aging and, thereby, delays age-associated deterioration. Autophagic flux declines with age and, in turn, decreases in autophagy contribute to the aging process itself and promote senescence. Here, we outline how autophagy regulates immune aging and discuss autophagy-inducing interventions that target senescent immune cells, which are major drivers of systemic aging. We examine how cutting-edge technologies, such as single-cell omics methods hold the promise to capture the complexity of molecular and cellular phenotypes associated with aging, driving the development of suitable putative biomarkers and clinical bioassays. Finally, we debate the urgency to initiate large-scale human clinical trials. We give special preference to small molecule probes and to dietary interventions that can extend healthy lifespan and are affordable for most of the world's population.
    Keywords:  Aging; Autophagy; Autophagy inducer; Immune system
    DOI:  https://doi.org/10.1242/dmm.049345
  7. Autophagy. 2022 Feb 01. 1-17
    Perinatal versus adult loss of ULK1 and ULK2 distinctly influences cardiac autophagy and function.
    Matthew P Harris, Quan J Zhang, Cole T Cochran, Jessica Ponce, Sean Alexander, Ana Kronemberger, Jordan D Fuqua, Yuan Zhang, Ranan Fattal, Tyler Harper, Matthew L Murry, Chad E Grueter, E Dale Abel, Vitor A Lira.
      Impairments in macroautophagy/autophagy, which degrades dysfunctional organelles as well as long-lived and aggregate proteins, are associated with several cardiomyopathies; however, the regulation of cardiac autophagy remains insufficiently understood. In this regard, ULK1 and ULK2 are thought to play primarily redundant roles in autophagy initiation, but whether their function is developmentally determined, potentially having an impact on cardiac integrity and function remains unknown. Here, we demonstrate that perinatal loss of ULK1 or ULK2 in cardiomyocytes (cU1-KO and cU2-KO mice, respectively) enhances basal autophagy without altering autophagy machinery content while preserving cardiac function. This increased basal autophagy is dependent on the remaining ULK protein given that perinatal loss of both ULK1 and ULK2 in cU1/2-DKO mice impaired autophagy causing age-related cardiomyopathy and reduced survival. Conversely, adult loss of cardiac ULK1, but not of ULK2 (i.e., icU1-KO and icU2-KO mice, respectively), led to a rapidly developing cardiomyopathy, heart failure and early death. icU1-KO mice had impaired autophagy with robust deficits in mitochondrial respiration and ATP synthesis. Trehalose ameliorated autophagy impairments in icU1-KO hearts but did not delay cardiac dysfunction suggesting that ULK1 plays other critical, autophagy-independent, functions in the adult heart. Collectively, these results indicate that cardiac ULK1 and ULK2 are functionally redundant in the developing heart, while ULK1 assumes a more unique, prominent role in the adult heart.Abbreviations: ATG4: autophagy related 4, cysteine peptidase; ATG5: autophagy related 5; ATG7: autophagy related 7; ATG9: autophagy related 9; ATG13: autophagy related 13; CYCS: Cytochrome C; DNM1L, dynamin 1-like; MAP1LC3A: microtubule-associated protein 1 light chain 3 alpha; MAP1LC3B: microtubule-associated protein 1 light chain 3 beta; MFN1: mitofusin 1; MFN2: mitofusin 2; MT-CO1: mitochondrially encoded cytochrome c oxidase I; MYH: myosin, heavy polypeptide; NBR1: NBR1 autophagy cargo receptor; NDUFA9: NADH:ubiquinone oxidoreductase subunit A9; OPA1: OPA1, mitochondrial dynamin like GTPase; PPARGC1A, peroxisome proliferator activated receptor, gamma, coactivator 1 alpha; SDHA: succinate dehydrogenase complex, subunit A, flavoprotein (Fp); SQSTM1: sequestosome 1; ULK1: unc-51 like kinase 1; ULK2: unc-51 like kinase 2; UQCRC1: ubiquinol-cytochrome c reductase core protein 1.
    Keywords:  Age-related cardiomyopathy; MAP1LC3; NBR1; SQSTM1; dilated cardiomyopathy; heart failure; mitochondria; mitophagy
    DOI:  https://doi.org/10.1080/15548627.2021.2022289
  8. Autophagy. 2022 Jan 31. 1-2
    Architecture of the endoplasmic reticulum plays a role in proteostasis.
    Smriti Parashar, Susan Ferro-Novick.
      The endoplasmic reticulum (ER) forms a contiguous network of tubules and sheets. When errors in protein folding occur, misfolded proteins accumulate in the ER. Proteostasis can be restored by ER quality control pathways. Reticulophagy is an ER quality control pathway that uses resident autophagy receptors to link an ER domain to the autophagy machinery. We recently showed that the reticulophagy receptor RTN3L recruits the COPII cargo adaptor SEC24C to target disease-causing mutant proinsulin INS2Akita puncta to the lysosome for degradation. When reticulophagy is disrupted and delivery to the lysosome is blocked, large INS2Akita puncta accumulate in the ER. Photobleach analysis revealed that these puncta behave like liquid condensates and not aggregates, as previously suggested. Other reticulophagy substrates that are segregated into tubules behave like INS2Akita, whereas a substrate of the ER sheets receptor, RETREG1/FAM134B, appears to be less fluid. Large INS2Akita puncta also accumulate when ER sheets are proliferated by the loss of LNPK, or by overproduction of the sheets-producing protein, CKAP4/CLIMP63. Restoring the tubular network by overexpressing reticulons reverses this phenotype. Our findings revealed that fluid-like deleterious cargoes are segregated into tubules to prevent them from expanding and affecting cell health while they are waiting to undergo reticulophagy.
    Keywords:  ER structure; Lunapark; SEC24C; misfolded prohormones and neuropeptides; protein quality control; reticulophagy
    DOI:  https://doi.org/10.1080/15548627.2022.2030175
  9. Prog Neurobiol. 2022 Jan 28. pii: S0301-0082(22)00015-6. [Epub ahead of print] 102229
    TDP-43 pathology: from noxious assembly to therapeutic removal.
    Sean S Keating, Rebecca San Gil, Molly E V Swanson, Emma L Scotter, Adam K Walker.
      Our understanding of amyotrophic lateral sclerosis and frontotemporal dementia has advanced dramatically since the discovery of cytoplasmic TAR DNA-binding protein 43 (TDP-43) inclusions as the hallmark pathology of these neurodegenerative diseases. Recent studies have provided insights into the physiological function of TDP-43 as an essential DNA-/RNA-modulating protein, and the triggers and consequences of TDP-43 dysfunction and aggregation. The formation of TDP-43 pathology is a progressive process, involving the generation of multiple distinct protein species, each with varying biophysical properties and roles in neurodegeneration. Here, we explore how the pathogenic changes to TDP-43, including mislocalisation, misfolding, aberrant liquid-liquid phase separation, stress granule assembly, oligomerisation, and post-translational modification, drive disease-association aggregation in TDP-43 proteinopathies. We highlight how pathological TDP-43 species are formed and contribute to cellular dysfunction and toxicity, via both loss-of-function and gain-of-function mechanisms. We also review the role of protein homeostasis mechanisms, namely the ubiquitin proteasome system, autophagy-lysosome pathway, heat-shock response, and chaperone-mediated autophagy, in combating TDP-43 aggregation and discuss how their dysfunction likely promotes disease pathogenesis and progression. Finally, we evaluate pre-clinical studies aimed at enhancing TDP-43 protein clearance via these mechanisms and provide insight on promising strategies for future therapeutic advances. Harnessing the mechanisms that protect against or ameliorate TDP-43 pathology presents promising opportunities for developing disease-modifying treatments for these neurodegenerative diseases.
    Keywords:  ALS; FTD; LLPS; TDP-43 proteinopathy; motor neuron disease; neurodegeneration; protein degradation; proteostasis
    DOI:  https://doi.org/10.1016/j.pneurobio.2022.102229
  10. Mol Cell. 2022 Jan 25. pii: S1097-2765(22)00002-8. [Epub ahead of print]
    E3 ligase RNF167 and deubiquitinase STAMBPL1 modulate mTOR and cancer progression.
    Dong Wang, Chenchen Xu, Wenyu Yang, Jie Chen, Yuhui Ou, Yuanyuan Guan, Jialiang Guan, Ying Liu.
      The mTOR complex 1 (mTORC1) is an essential metabolic hub that coordinates cellular metabolism with the availability of nutrients, including amino acids. Sestrin2 has been identified as a cytosolic leucine sensor that transmits leucine status signals to mTORC1. In this study, we identify an E3 ubiquitin ligase RING finger protein 167 (RNF167) and a deubiquitinase STAMBPL1 that function in concert to control the polyubiquitination level of Sestrin2 in response to leucine availability. Ubiquitination of Sestrin2 promotes its interaction with GATOR2 and inhibits mTORC1 signaling. Bioinformatic analysis reveals decreased RNF167 expression and increased STAMBPL1 expression in gastric and colorectal tumors. Knockout of STAMBPL1 or correction of the heterozygous STAMBPL1 mutation in a human colon cancer cell line suppresses xenograft tumor growth. Lastly, a cell-permeable peptide that blocks the STAMBPL1-Sestrin2 interaction inhibits mTORC1 and provides a potential option for cancer therapy.
    Keywords:  Sestrin2; amino acid sensing; colorectal cancer; mTOR; tumorigenesis; ubiquitination
    DOI:  https://doi.org/10.1016/j.molcel.2022.01.002
  11. J Cardiovasc Pharmacol. 2022 Feb 03.
    Homocysteine Suppresses Autophagy via AMPK-mTOR-TFEB Signaling in Human THP-1 Macrophages.
    Yang Yu-Ping, Ren Yong-Gang, Cai Bi-Qing, Huang Dan-Dan.
      ABSTRACT: Hyperhomocysteinemia is an independent risk factor for atherosclerosis. It is known that macrophage autophagy plays a protective role in atherosclerosis, and that hyperhomocysteinemia is strongly linked to autophagy. Therefore, it is of great significance to study the molecular mechanisms underlying the effect of homocysteine (Hcy) on macrophage autophagy. This study aimed to investigate the effects of Hcy on autophagy in a human acute monocytic leukemia cell line (THP-1). The Hcy-treated THP-1 cells exhibited increased levels of the autophagy substrate SQSTM1(p62), and decreased levels of the autophagy markers LC3 II/I and Beclin-1, indicating a decrease in autophagy in vitro. Furthermore, western blotting showed that Hcy significantly increased the levels of p-mTOR and nuclear TFEB and decreased the levels of p-AMPK and cytoplasmic TFEB. These data suggest that Hcy inhibits autophagosome formation in human THP-1 macrophages through the AMPK-mTOR-TFEB signaling pathway. Our findings provide new insights into the mechanisms of atherosclerotic diseases caused by Hcy.
    DOI:  https://doi.org/10.1097/FJC.0000000000001232
  12. Neurosci Lett. 2022 Jan 31. pii: S0304-3940(22)00050-7. [Epub ahead of print] 136493
    Rapamycin ameliorates age-related hearing loss in C57BL/6J mice by enhancing autophagy in the SGNs.
    Hongyan Liu, Feitian Li, Xuanyi Li, Qianru Wu, Chunfu Dai.
      Autophagy plays a pathogenic role in neurodegenerative disease. However, the involvement of autophagy in the pathogenesis of age-related hearing loss (ARHL) remains obscure. Naturally aged C57BL/6J mice were used to identify the role of autophagy in ARHL, and rapamycin, a mammalian target of rapamycin (mTOR) inhibitor, was administered for 34 weeks to explore the potential therapeutic effect of rapamycin in ARHL. We found that the number of autophagosomes and the expression of microtubule-associated protein 1 light chain 3B (LC3B) decreased as the mice aged. The expression of autophagy-related (Atg) proteins, including Beclin1 and Atg5, and the ratio of LC3-II/I was reduced in aged mice, while mTOR activity in aged mice gradually increased. Rapamycin improved the auditory brainstem response (ABR) threshold (at 8, 12, and 24 kHz). Further exploration demonstrated that spiral ganglion neuron (SGN) density was enhanced in response to administration of rapamycin. The rate of apoptosis in the basal turn SGNs was decreased, whereas autophagy activity was increased in the experimental group. Meanwhile, mTOR activity in the experimental group was decreased. Our findings indicate that age-related deficiency in autophagy may lead to increased apoptosis of aged SGNs. Rapamycin enhances autophagy of SGNs by inhibiting mTOR activation, resulting in amelioration of ARHL. Therapeutic strategy targeting autophagy may provide a potential approach for treating ARHL.
    DOI:  https://doi.org/10.1016/j.neulet.2022.136493
  13. Alzheimers Dement. 2021 Dec;17 Suppl 3 e050868
    Deficiency of the lysosomal exonuclease PLD3 impacts the degradative route.
    Zoë P Van Acker, Marine Bretou, Ragna Sannerud, Markus Damme, Wim Annaert.
      BACKGROUND: Phospholipase D3 (PLD3) is a 5'-3' exonuclease that resides in lysosomes where it is implicated in the regulation of inflammatory responses by degrading ssDNA. Polymorphisms in the PLD3 gene have been linked to late-onset Alzheimer's disease (LOAD; Cruchaga et al., PMID: 24336208), but contradictory data exist on the impact of PLD3 deficiency on APP processing. Given PLD3 is primarily expressed in neurons within the brain, we were interested in exploring its role in neuronal endolysosomal homeostasis.METHOD: We generated PLD3-/- neuroblastoma SH-SY5Y clonal cell lines, using Crispr/cas9 gene editing, that were subsequently stably rescued with lentiviral vector technology. For this, the wild-type PLD3 sequence was compared to variants harbouring LOAD-associated risk single-nucleotide polymorphisms (SNPs), including M6R, K228R, V232M, N236S, N284S and T426A. PLD3 exonuclease kinetics towards different substrates were measured using EFQO assays (Cappel et al., PMID: 33288674). Autophagy and mitophagy dysregulations, as possible important catalysts for DNA delivery to lysosomes, were further scrutinized with biochemical assays and confocal imaging. In parallel, we investigated the endolysosomal morphology by electron microscopy.
    RESULT: Firstly, most SNP variants, except the M6R and T426A variants, display a significantly reduced exonuclease activity; indicating a loss-of-function for these variants. Overall, knock-out of PLD3 leads to elevations in lysosomal and autophagosomal components, including a defect in mitophagy, lysosomal cholesterol accumulation and an accretion of the uncleaved/inactive CatD. These aberrant features can be rescued by re-introducing wild-type PLD3, but not PLD3 SNP-variants. This underscores a critical role for PLD3 in lysosomal homeostasis, likely mediated through its exonuclease activity. Given different variants elicit different exonuclease kinetics towards different ssDNA substrates, this suggests that PLD3 contributes to upholding lysosomal DNA levels within a narrow range, which is required for a normal lysosomal proteostasis.
    CONCLUSION: PLD3 depletion and exonuclease dysfunction-causing SNPs promote lysosomal impairment and, as such, a congestion of the degradative route. We hypothesize that this could relate to a dysfunctional exonuclease activity of PLD3 in lysosomes. Given the observation of engulfed mitochondria within autophagosomes, the substrates for PLD3 could potentially come from mtDNA.
    DOI:  https://doi.org/10.1002/alz.050868
  14. Front Cell Dev Biol. 2021 ;9 775364
    Mapping Critical Residues in ATG11's Coiled-Coil 2 Domain that Block Multiple Interactions and Disrupt Selective Autophagy.
    Mitchell D Meyer, Jasmine Winzeler, Sophia M Taylor, Alex Kilgore, Kimberly Edicha, Chase Chitwood, Zachary Spearin, S K Nadia Rahman Silvia, Ronith Chakraborty, Jesse E Smith, Bridget Kennedy, Carson Zois, Hayley Cawthon, Mukiri Gilruth, Steven K Backues.
      Selective autophagy is a conserved subcellular process that maintains the health of eukaryotic cells by targeting damaged or toxic cytoplasmic components to the vacuole/lysosome for degradation. A key player in the initiation of selective autophagy in S. Cerevisiae (baker's yeast) is a large adapter protein called Atg11. Atg11 has multiple predicted coiled-coil domains and intrinsically disordered regions, is known to dimerize, and binds and organizes other essential components of the autophagosome formation machinery, including Atg1 and Atg9. We performed systematic directed mutagenesis on the coiled-coil 2 domain of Atg11 in order to map which residues were required for its structure and function. Using yeast-2-hybrid and coimmunoprecipitation, we found only three residues to be critical: I562, Y565, and I569. Mutation of any of these, but especially Y565, could interfere with Atg11 dimerization and block its interaction with Atg1 and Atg9, thereby inactivating selective autophagy.
    Keywords:  Atg1; Atg11; Atg19; Atg9; coimmunoprecipitation; directed mutagenesis; selective autophagy; yeast-2-hybrid
    DOI:  https://doi.org/10.3389/fcell.2021.775364
  15. Cell Tissue Res. 2022 Feb 02.
    Autophagy-related DjAtg1-1 plays critical role in planarian regeneration by regulating proliferation and cell death.
    Baijie Jin, Jing Ren, Jinzi Chen, Zimei Dong, Guangwen Chen, Dezeng Liu.
      Autophagy is an intracellular degradation process and plays key roles in energy recycle and homeostasis maintenance during planarian regeneration. Although planarians provide an ideal model organism for studying autophagy in vivo, the molecular mechanism of planarian autophagy is still unknown. Here, we identify three autophagy-related (Atg) gene 1 homologs from Dugesia japonica and study their roles in planarian regeneration. Both DjATG1-1 and DjATG1-2 proteins show homology to vertebrate unc-51 like autophagy activating kinase 1 (ULK1) and ULK2, DjATG1-3 shows homology to vertebrate ULK3. In contrast to the ubiquitously expressed DjAtg1-1 and DjAtg1-3, DjAtg1-2 is mainly expressed in the intestine branches and epidermis. All the three DjAtg1s can respond to planarian regeneration and starvation. Both DjAtg1-1 and DjAtg1-2 are expressed in the reproductive organs of the starved sexual worms. DjAtg1-1 or DjAtg1-3 RNAi leads to head lysis and death of starved planarians, accompanied by exhaustion of neoblasts. DjAtg1-1 RNAi causes autophagy and regeneration defects and decreases proliferation and cell death; both DjAtg1-2 and DjAtg1-3 RNAi cause no autophagy or regeneration defect but increase cell death during regeneration. Our findings uncover the roles of DjAtg1s in autophagy and regeneration of planarian and highlight the links between proliferation, cell death, and autophagy during regeneration.
    Keywords:  Autophagy; DjAtg1s; Dugesia japonica; Planarian; Regeneration
    DOI:  https://doi.org/10.1007/s00441-022-03591-3
  16. Nat Commun. 2022 Feb 03. 13(1): 680
    Dendritic autophagy degrades postsynaptic proteins and is required for long-term synaptic depression in mice.
    Emmanouela Kallergi, Akrivi-Dimitra Daskalaki, Angeliki Kolaxi, Come Camus, Evangelia Ioannou, Valentina Mercaldo, Per Haberkant, Frank Stein, Kyriaki Sidiropoulou, Yannis Dalezios, Mikhail M Savitski, Claudia Bagni, Daniel Choquet, Eric Hosy, Vassiliki Nikoletopoulou.
      The pruning of dendritic spines during development requires autophagy. This process is facilitated by long-term depression (LTD)-like mechanisms, which has led to speculation that LTD, a fundamental form of synaptic plasticity, also requires autophagy. Here, we show that the induction of LTD via activation of NMDA receptors or metabotropic glutamate receptors initiates autophagy in the postsynaptic dendrites in mice. Dendritic autophagic vesicles (AVs) act in parallel with the endocytic machinery to remove AMPA receptor subunits from the membrane for degradation. During NMDAR-LTD, key postsynaptic proteins are sequestered for autophagic degradation, as revealed by quantitative proteomic profiling of purified AVs. Pharmacological inhibition of AV biogenesis, or conditional ablation of atg5 in pyramidal neurons abolishes LTD and triggers sustained potentiation in the hippocampus. These deficits in synaptic plasticity are recapitulated by knockdown of atg5 specifically in postsynaptic pyramidal neurons in the CA1 area. Conducive to the role of synaptic plasticity in behavioral flexibility, mice with autophagy deficiency in excitatory neurons exhibit altered response in reversal learning. Therefore, local assembly of the autophagic machinery in dendrites ensures the degradation of postsynaptic components and facilitates LTD expression.
    DOI:  https://doi.org/10.1038/s41467-022-28301-z
  17. Front Cell Dev Biol. 2021 ;9 788634
    FUNDC1: A Promising Mitophagy Regulator at the Mitochondria-Associated Membrane for Cardiovascular Diseases.
    Guoyong Li, Junli Li, Ruochen Shao, Jiahao Zhao, Mao Chen.
      Mitochondrial autophagy (or mitophagy) regulates the mitochondrial network and function to contribute to multiple cellular processes. The protective effect of homeostatic mitophagy in cardiovascular diseases (CVDs) has attracted increasing attention. FUN14 domain containing 1 (FUNDC1), an identified mitophagy receptor, plays an essential role in CVDs. Different expression levels of FUNDC1 and its phosphorylated state at different sites alleviate or exacerbate hypoxia and ischemia/reperfusion injury, cardiac hypertrophy, or metabolic damage through promotion or inhibition of mitophagy. In addition, FUNDC1 can be enriched at contact sites between mitochondria and the endoplasmic reticulum (ER), determining the formation of mitochondria-associated membranes (MAMs) that regulate cellular calcium (Ca2+) homeostasis and mitochondrial dynamics to prevent heart dysfunction. Moreover, FUNDC1 has also been involved in inflammatory cardiac diseases such as septic cardiomyopathy. In this review, we collect and summarize the evidence on the roles of FUNDC1 exclusively in various CVDs, describing its interactions with different cellular organelles, its involvement in multiple cellular processes, and its associated signaling pathways. FUNDC1 may become a promising therapeutic target for the prevention and management of various CVDs.
    Keywords:  FUNDC1; LC3; MAM; cardiovascular diseases; mitophagy
    DOI:  https://doi.org/10.3389/fcell.2021.788634
  18. J Cell Sci. 2022 Jan 31. pii: jcs.259138. [Epub ahead of print]
    ALFY localizes to early endosomes and cellular protrusions to facilitate directional cell migration.
    Kristiane Søreng, Serhiy Pankiv, Camilla Bergsmark, Ellen M Haugsten, Anette K Dahl, Laura R de la Ballina, Ai Yamamoto, Alf H Lystad, Anne Simonsen.
      Cell migration is a complex process underlying physiological and pathological processes such as brain development and cancer metastasis. The autophagy-linked FYVE protein (ALFY), an autophagy adaptor protein known to promote clearance of protein aggregates, has been implicated in brain development and neural migration during cerebral cortical neurogenesis in mice. However, a specific role of ALFY in cell motility and extracellular matrix adhesion during migration has not been investigated. Here we reveal a novel role for ALFY in the endocytic pathway and in cell migration. We show that ALFY localizes to RAB5 and EEA1 positive early endosomes in a PtdIns(3)P dependent manner and is highly enriched in cellular protrusions at the leading and lagging edge of migrating cells. We find that cells lacking ALFY have reduced attachment and altered protein levels and glycosylation of integrins, resulting in the inability to form a proper leading edge and loss of directional cell motility.
    Keywords:  ALFY; Endosome; Focal adhesion; Integrin; Migration; WDFY3
    DOI:  https://doi.org/10.1242/jcs.259138
  19. Blood. 2022 Feb 03. 139(5): 640-642
    Stressed and disoriented: stromal autophagy regulates HSCs.
    Stephanie N Hurwitz, Peter Kurre.
      
    DOI:  https://doi.org/10.1182/blood.2021014337
  20. Free Radic Biol Med. 2022 Jan 31. pii: S0891-5849(22)00041-7. [Epub ahead of print]
    Activation of autophagy attenuates motor deficits and extends lifespan in a C. elegans model of ALS.
    Hui Xu, Congcong Jia, Cheng Cheng, Haifeng Wu, Huaibin Cai, Weidong Le.
      Mutations in Cu/Zn-superoxide dismutase 1 (SOD1) are linked to amyotrophic lateral sclerosis (ALS). Using a line of ALS-related mutant human SOD1 (hSOD1) transgenic Caenorhabditis elegans, we tested the effects of metformin on the progression of ALS-like pathological abnormalities. We found that metformin significantly extended the lifespan, improved motor performance, and enhanced antioxidant activity of mutant worms. We further showed that metformin enhanced expression of lgg-1, daf-16, skn-1 and other genes known to regulate autophagy, longevity and oxidative stress in hSOD1 transgenic worms. Accordingly, overexpression of lgg-1 or daf-16 attenuated the aging and pathological abnormalities of mutant human SOD1 worms, while genetic deletion of lgg-1 or daf-16 abolished the beneficial effects of metformin. Collectively, we demonstrate that metformin protects against mutant SOD1-induced cytotoxicity in part through enhancement of autophagy and extends lifespan through daf-16 pathway. Our findings suggest that metformin could be further explored as a potential therapeutic agent in treating ALS.
    Keywords:  ALS; Autophagy; Lifespan; Metformin; Neuroprotection
    DOI:  https://doi.org/10.1016/j.freeradbiomed.2022.01.030
  21. Neuron. 2022 Feb 02. pii: S0896-6273(22)00055-1. [Epub ahead of print]110(3): 351-352
    Living in α-syn: Tackling aggregates in Parkinson's disease.
    Christopher J Griffey, Ai Yamamoto.
      How do protein aggregates contribute to neurodegenerative disorders, and can they be therapeutically targeted? In this issue of Neuron, Stojkovska et al. (2022) show that aggregated α-synuclein disrupts ER and lysosomal function in Parkinson's disease patient-derived neurons and that combined enhancement of multiple arms of the proteostasis network improves these defects.
    DOI:  https://doi.org/10.1016/j.neuron.2022.01.016
  22. EMBO J. 2022 Jan 31. e108119
    LAMTOR1 inhibition of TRPML1-dependent lysosomal calcium release regulates dendritic lysosome trafficking and hippocampal neuronal function.
    Jiandong Sun, Yan Liu, Xiaoning Hao, Weiju Lin, Wenyue Su, Emerald Chiang, Michel Baudry, Xiaoning Bi.
      Lysosomes function not only as degradatory compartments but also as dynamic intracellular calcium ion stores. The transient receptor potential mucolipin 1 (TRPML1) channel mediates lysosomal Ca2+ release, thereby participating in multiple cellular functions. The pentameric Ragulator complex, which plays a critical role in the activation of mTORC1, is also involved in lysosomal trafficking and is anchored to lysosomes through its LAMTOR1 subunit. Here, we report that the Ragulator restricts lysosomal trafficking in dendrites of hippocampal neurons via LAMTOR1-mediated tonic inhibition of TRPML1 activity, independently of mTORC1. LAMTOR1 directly interacts with TRPML1 through its N-terminal domain. Eliminating this inhibition in hippocampal neurons by LAMTOR1 deletion or by disrupting LAMTOR1-TRPML1 binding increases TRPML1-mediated Ca2+ release and facilitates dendritic lysosomal trafficking powered by dynein. LAMTOR1 deletion in the hippocampal CA1 region of adult mice results in alterations in synaptic plasticity, and in impaired object-recognition memory and contextual fear conditioning, due to TRPML1 activation. Mechanistically, changes in synaptic plasticity are associated with increased GluA1 dephosphorylation by calcineurin and lysosomal degradation. Thus, LAMTOR1-mediated inhibition of TRPML1 is critical for regulating dendritic lysosomal motility, synaptic plasticity, and learning.
    Keywords:  LTD; LTP; calcineurin; calcium; dynein
    DOI:  https://doi.org/10.15252/embj.2021108119
  23. Traffic. 2022 Jan 30.
    Vps501, a novel vacuolar SNX-BAR protein cooperates with the SEA complex to regulate TORC1 signaling.
    Shreya Goyal, Verónica A Segarra, Nitika, Aaron M Stetcher, Andrew W Truman, Adam M Reitzel, Richard J Chi.
      The sorting nexins (SNX), constitute a diverse family of molecules that play varied roles in membrane trafficking, cell signaling, membrane remodeling, organelle motility and autophagy. In particular, the SNX-BAR proteins, a SNX subfamily characterized by a C-terminal dimeric Bin/Amphiphysin/Rvs (BAR) lipid curvature domain and a conserved Phox-homology domain, are of great interest. In budding yeast, many SNX-BARs proteins have well-characterized endo-vacuolar trafficking roles. Phylogenetic analyses allowed us to identify an additional SNX-BAR protein, Vps501, with a novel endo-vacuolar role. We report that Vps501 uniquely localizes to the vacuolar membrane and has physical and genetic interactions with the SEA complex to regulate TORC1 inactivation. We found cells displayed a severe deficiency in starvation-induced/nonselective autophagy only when SEA complex subunits are ablated in combination with Vps501, indicating a cooperative role with the SEA complex during TORC1 signaling during autophagy induction. Additionally, we found the SEACIT complex becomes destabilized in vps501Δsea1Δ cells, which resulted in aberrant endosomal TORC1 activity and subsequent Atg13 hyperphosphorylation. We have also discovered that the vacuolar localization of Vps501 is dependent upon a direct interaction with Sea1 and a unique lipid binding specificity that is also required for its function. This article is protected by copyright. All rights reserved.
    Keywords:  Atg27; SEA complex; SNX-BAR; TORC1; autophagy; retromer
    DOI:  https://doi.org/10.1111/tra.12833
  24. Alzheimers Dement. 2021 Dec;17 Suppl 3 e051678
    Synaptic stimulation induces tau clearance by enhancing autophagosomal/lysosomal degradation.
    Yvette Akwa, Chiara Di Malta, Elise Gondard, Anne Boiret, Andres M Lozano, Thomas Vaccari, Carmine Settembre, Etienne E Baulieu, Davide Tampellini.
      BACKGROUND: Alterations of synaptic excitability and reduced brain metabolism are some of the earliest changes associated Alzheimer's disease (AD) pathogenesis (Reiman et al., 2004; Sperling et al., 2009). Among different approaches for therapeutics, the stimulation of synaptic activity exerts protective effects in models of AD, and deep brain stimulation (DBS) has been shown to ameliorate AD symptoms in certain patients (Sankar et al., 2015; Swaab and Bao, 2010; Tampellini, 2015). Such positive effects might be the result of changes occurring at cellular level upon activity induction, which promote cellular mechanisms correcting neuronal and synaptic dysfunctions. We have demonstrated that synaptic stimulation, via DBS or other methods, is protective in mouse models of AD and frontotemporal dementia (FTD) by enhancing autophagy and lysosomal degradation of pathologic tau, and by protecting synapses (Akwa et al., 2018; Mann et al., 2018). Our more recent investigations are revealing the involvement of Transcription factor EB (TFEB), a pivotal player in lysosomal biogenesis and autophagy (Sardiello et al., 2009), and its downstream genes in the enhancement of lysosomal activity upon stimulation.METHOD: 3xTg mice received deep brain stimulation by electrode implantation in the entorhinal cortex (Mann et al., 2018). Cultured neurons were prepared from E15 PS19 mouse embryos (Akwa et al., 2018) and stimulated at 14 days in vitro (Ehlers, 2003). Confocal immunofluorescence, Western blot and statistical analyses were performed as described (Akwa et al., 2018). RT-qPCR was performed as described (Napolitano et al., 2018).
    RESULT: DBS was able to reduce levels of hyperphosphorylated and oligomeric (but not total) tau restoring levels of synaptic proteins back to wild-type in 3xTg mice. Pathological tau clearance occurred via autophagic pathway and lysosomal activity, the latest being enhanced by synaptic stimulation. TFEB was required for tau clearance as its inhibition prevented activity-induced reduction of pathological tau. In addition, RT-qPCR data analyses revealed increase expressions of TFEB downstream genes, including ATP6-V1H, ATP6-V0D1, and mucolipin-1 in neurons during synaptic stimulation.
    CONCLUSION: Protective effects of DBS/synaptic stimulation against pathological tau include the involvement TFEB and the enhancement of autophagy and lysosomal degradation.
    DOI:  https://doi.org/10.1002/alz.051678
  25. Mol Brain. 2022 02 02. 15(1): 14
    The dual roles of autophagy and the GPCRs-mediating autophagy signaling pathway after cerebral ischemic stroke.
    Weichen Hou, Yulei Hao, Li Sun, Yang Zhao, Xiangyu Zheng, Lei Song.
      Ischemic stroke, caused by a lack of blood supply in brain tissues, is the third leading cause of human death and disability worldwide, and usually results in sensory and motor dysfunction, cognitive impairment, and in severe cases, even death. Autophagy is a highly conserved lysosome-dependent process in which eukaryotic cells removal misfolded proteins and damaged organelles in cytoplasm, which is critical for energy metabolism, organelle renewal, and maintenance of intracellular homeostasis. Increasing evidence suggests that autophagy plays important roles in pathophysiological mechanisms under ischemic conditions. However, there are still controversies about whether autophagy plays a neuroprotective or damaging role after ischemia. G-protein-coupled receptors (GPCRs), one of the largest protein receptor superfamilies in mammals, play crucial roles in various physiological and pathological processes. Statistics show that GPCRs are the targets of about one-fifth of drugs known in the world, predicting potential values as targets for drug research. Studies have demonstrated that nutritional deprivation can directly or indirectly activate GPCRs, mediating a series of downstream biological processes, including autophagy. It can be concluded that there are interactions between autophagy and GPCRs signaling pathway, which provides research evidence for regulating GPCRs-mediated autophagy. This review aims to systematically discuss the underlying mechanism and dual roles of autophagy in cerebral ischemia, and describe the GPCRs-mediated autophagy, hoping to probe promising therapeutic targets for ischemic stroke through in-depth exploration of the GPCRs-mediated autophagy signaling pathway.
    Keywords:  Autophagy; G-protein-coupled receptors; Ischemic stroke; Macroautophagy; Neuropharmacology
    DOI:  https://doi.org/10.1186/s13041-022-00899-7
  26. Transl Neurodegener. 2022 01 31. 11(1): 5
    LRRK2, GBA and their interaction in the regulation of autophagy: implications on therapeutics in Parkinson's disease.
    Shirley Yin-Yu Pang, Rachel Cheuk Nam Lo, Philip Wing-Lok Ho, Hui-Fang Liu, Eunice Eun Seo Chang, Chi-Ting Leung, Yasine Malki, Zoe Yuen-Kiu Choi, Wing Yan Wong, Michelle Hiu-Wai Kung, David Boyer Ramsden, Shu-Leong Ho.
      Mutations in leucine-rich repeat kinase 2 (LRRK2) and glucocerebrosidase (GBA) represent two most common genetic causes of Parkinson's disease (PD). Both genes are important in the autophagic-lysosomal pathway (ALP), defects of which are associated with α-synuclein (α-syn) accumulation. LRRK2 regulates macroautophagy via activation of the mitogen activated protein kinase/extracellular signal regulated protein kinase (MAPK/ERK) kinase (MEK) and the calcium-dependent adenosine monophosphate (AMP)-activated protein kinase (AMPK) pathways. Phosphorylation of Rab GTPases by LRRK2 regulates lysosomal homeostasis and endosomal trafficking. Mutant LRRK2 impairs chaperone-mediated autophagy, resulting in α-syn binding and oligomerization on lysosomal membranes. Mutations in GBA reduce glucocerebrosidase (GCase) activity, leading to glucosylceramide accumulation, α-syn aggregation and broad autophagic abnormalities. LRRK2 and GBA influence each other: GCase activity is reduced in LRRK2 mutant cells, and LRRK2 kinase inhibition can alter GCase activity in GBA mutant cells. Clinically, LRRK2 G2019S mutation seems to modify the effects of GBA mutation, resulting in milder symptoms than those resulting from GBA mutation alone. However, dual mutation carriers have an increased risk of PD and earlier age of onset compared with single mutation carriers, suggesting an additive deleterious effect on the initiation of PD pathogenic processes. Crosstalk between LRRK2 and GBA in PD exists, but its exact mechanism is unclear. Drugs that inhibit LRRK2 kinase or activate GCase are showing efficacy in pre-clinical models. Since LRRK2 kinase and GCase activities are also altered in idiopathic PD (iPD), it remains to be seen if these drugs will be useful in disease modification of iPD.
    Keywords:  Autophagy; GBA; GCase; Interaction; LRRK2; Mutation; Parkinson’s disease; α-Synuclein
    DOI:  https://doi.org/10.1186/s40035-022-00281-6
  27. Exp Cell Res. 2022 Jan 28. pii: S0014-4827(22)00036-2. [Epub ahead of print] 113043
    Suppression of PI3K signaling is linked to autophagy activation and the spatiotemporal induction of the lens organelle free zone.
    Rifah Gheyas, Ramon Ortega-Alvarez, Daniel Chauss, Marc Kantorow, A Sue Menko.
      The terminal steps of lens cell differentiation require elimination of all organelles to create a central Organelle Free Zone (OFZ) that is required for lens function of focusing images on the retina. Previous studies show that the spatiotemporal elimination of these organelles during development is autophagy-dependent. We now show that the inhibition of PI3K signaling in lens organ culture results in the premature induction of autophagy within 24 h, including a significant increase in LAMP1+ lysosomes, and the removal of lens organelles from the center of the lens. Specific inhibition of just the PI3K/Akt signaling axis was directly linked to the elimination of mitochondria and ER, while pan-PI3K inhibitors that block all PI3K downstream signaling removed all organelles, including nuclei. Therefore, blocking the PI3K/Akt pathway was alone insufficient to remove nuclei. RNAseq analysis revealed increased mRNA levels of the endogenous inhibitor of PI3K activation, PIK3IP1, in differentiating lens fiber cells preceding the induction of OFZ formation. Co-immunoprecipitation confirmed that PIK3IP1 associates with multiple PI3K p110 isoforms just prior to formation of the OFZ, providing a likely endogenous mechanism for blocking all PI3K signaling and activating the autophagy pathway required to form the OFZ during lens development.
    Keywords:  Akt; Autophagy; Development; Lens; Organelle free zone (OFZ); PI3K
    DOI:  https://doi.org/10.1016/j.yexcr.2022.113043
  28. Life Sci Alliance. 2022 May;pii: e202101082. [Epub ahead of print]5(5):
    Environmental control of Pub1 (NEDD4 family E3 ligase) in Schizosaccharomyces pombe is regulated by TORC2 and Gsk3.
    Tingting Wang, Philip Woodman, Sean J Humphrey, Janni Petersen.
      Cells respond to changing nutrient environments by adjusting the abundance of surface nutrient transporters and receptors. This can be achieved by modulating ubiquitin-dependent endocytosis, which in part is regulated by the NEDD4 family of E3 ligases. Here we report novel regulation of Pub1, a fission yeast Schizosaccharomyces pombe member of the NEDD4-family of E3 ligases. We show that nitrogen stress inhibits Pub1 function, thereby increasing the abundance of the amino acid transporter Aat1 at the plasma membrane and enhancing sensitivity to the toxic arginine analogue canavanine. We show that TOR complex 2 (TORC2) signalling negatively regulates Pub1, thus TORC2 mutants under nutrient stress have decreased Aat1 at the plasma membrane and are resistant to canavanine. Inhibition of TORC2 signalling increases Pub1 phosphorylation, and this is dependent on Gsk3 activity. Addition of the Tor inhibitor Torin1 increases phosphorylation of Pub1 at serine 199 (S199) by 2.5-fold, and Pub1 protein levels in S199A phospho-ablated mutants are reduced. S199 is conserved in NEDD4 and is located immediately upstream of a WW domain required for protein interaction. Together, we describe how the major TORC2 nutrient-sensing signalling network regulates environmental control of Pub1 to modulate the abundance of nutrient transporters.
    DOI:  https://doi.org/10.26508/lsa.202101082
  29. Aging Dis. 2022 Feb;13(1): 157-174
    Mammalian AKT, the Emerging Roles on Mitochondrial Function in Diseases.
    Xiaoxian Xie, Ruonan Shu, Chunan Yu, Zhengwei Fu, Zezhi Li.
      Mitochondrial dysfunction may play a crucial role in various diseases due to its roles in the regulation of energy production and cellular metabolism. Serine/threonine kinase (AKT) is a highly recognized antioxidant, immunomodulatory, anti-proliferation, and endocrine modulatory molecule. Interestingly, increasing studies have revealed that AKT can modulate mitochondria-mediated apoptosis, redox states, dynamic balance, autophagy, and metabolism. AKT thus plays multifaceted roles in mitochondrial function and is involved in the modulation of mitochondria-related diseases. This paper reviews the protective effects of AKT and its potential mechanisms of action in relation to mitochondrial function in various diseases.
    Keywords:  AKT; disease; energy dynamics/metabolism; mitochondrial autophagy; pathway signaling
    DOI:  https://doi.org/10.14336/AD.2021.0729
  30. Am J Physiol Cell Physiol. 2022 Feb 02.
    The importance of mitochondrial quality control for maintaining skeletal muscle function across healthspan.
    James Sligar, Danielle A Debruin, Nicholas J Saner, Ashleigh M Philp, Andrew Philp.
      As the principal energy-producing organelles of the cell, mitochondria support numerous biological processes related to metabolism, growth and regeneration in skeletal muscle. Deterioration in skeletal muscle functional capacity with age is thought to be driven in part by a reduction in skeletal muscle oxidative capacity and reduced fatigue resistance. Underlying this maladaptive response is the development of mitochondrial dysfunction caused by alterations in mitochondrial quality control (MQC), a term encompassing processes of mitochondrial synthesis (biogenesis), remodelling (dynamics) and degradation (mitophagy). Knowledge regarding the role and regulation of MQC in skeletal muscle and the influence of ageing in this process have rapidly advanced in the last decade. Given the emerging link between ageing and MQC, therapeutic approaches to manipulate MQC to prevent mitochondrial dysfuntion during ageing hold tremendous therapeutic potential.
    Keywords:  biogenesis; metabolism; mitochondria; mitophagy; skeletal muscle
    DOI:  https://doi.org/10.1152/ajpcell.00388.2021
  31. Immunol Res. 2022 Feb 02.
    PI3K/Akt/mTOR pathway: a potential target for anti-SARS-CoV-2 therapy.
    Soheila Fattahi, Zahra Khalifehzadeh-Esfahani, Mina Mohammad-Rezaei, Sahar Mafi, Morteza Jafarinia.
      Coronavirus disease 2019 (COVID-19) is a viral infection caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). A single-stranded RNA virus from a β-Coronaviridae family causes acute clinical manifestations. Its high death rate and severe clinical symptoms have turned it into the most significant challenge worldwide. Up until now, several effective COVID-19 vaccines have been designed and marketed, but our data on specialized therapeutic drugs for the treatment of COVID-19 is still limited. In order to synthesis virus particles, SARS-CoV-2 uses host metabolic pathways such as phosphoinositide3-kinase (PI3K)/protein kinase B (PKB, also known as AKT)/mammalian target of rapamycin (mTOR). mTOR is involved in multiple biological processes. Over-activation of the mTOR pathway improves viral replication, which makes it a possible target in COVID-19 therapy. Clinical data shows the hyperactivation of the mTOR pathway in lung tissues during respiratory viral infections. However, the exact impact of mTOR pathway inhibitors on the COVID-19 severity and death rate is yet to be thoroughly investigated. There are several mTOR pathway inhibitors. Rapamycin is the most famous inhibitor of mTORC1 among all. Studies on other respiratory viruses suggest that the therapeutic inhibitors of the mTOR pathway, especially rapamycin, can be a potential approach to anti-SARS-CoV-2 therapy. Using therapeutic methods that inhibit harmful immune responses can open a new chapter in treating severe COVID-19 disease. We highlighted the potential contribution of PI3K/Akt/mTOR inhibitors in the treatment of COVID-19.
    Keywords:  COVID-19; PI3K/Akt/ mTOR pathway; Rapamycin; SARS-CoV-2; mTOR inhibitors
    DOI:  https://doi.org/10.1007/s12026-022-09268-x
  32. Alzheimers Dement. 2021 Dec;17 Suppl 3 e057798
    VPS35 down regulation alters degradation pathways in neuronal cells.
    Alessia Filippone, Domenico Pratico, Emanuela Esposito.
      BACKGROUND: The vacuolar protein sorting 35 (VPS35) is the main component of the retromer recognition core complex system which regulates intracellular cargo protein sorting and trafficking. Downregulation of VPS35 has been linked to the pathogenesis of neurodegenerative disorders such Alzheimer's and Parkinson's disease via endosome dysregulation. Here we showed that the genetic manipulation of VPS35 has an effect on intracellular degradation pathways.METHOD: A neuronal cell line expressing the human APP Swedish variant (N2A APPSwe cell line) was used and VPS35 genetic manipulation was performed stimulating cells with VPS35 siRNA or Ctr siRNA. After 72 h, cells and lysates were used for Western blot and immunofluorescence analysis.
    RESULT: We found that down-regulation of VPS35 alters the autophagy flux by affecting both microtubule-associated proteins 1A/1B light chain 3B and sequestrosome-1 expression levels. This effect was associated with an intracellular accumulation of acidic and ubiquitinated aggregates suggesting that dysfunction of the retromer recognition core leads to a significant alteration in autophagy flux as well as in the ubiquitin-proteasome pathway.
    CONCLUSION: Our data demonstrate that besides cargo sorting and trafficking, VPS35 by supporting the integral function of the retromer complex system plays an important role also as a critical regulator of intracellular degradation pathways.
    DOI:  https://doi.org/10.1002/alz.057798
  33. Front Cell Dev Biol. 2021 ;9 787278
    Apigenin Alleviates Intervertebral Disc Degeneration via Restoring Autophagy Flux in Nucleus Pulposus Cells.
    Chenglong Xie, Yifeng Shi, Zuoxi Chen, Xin Zhou, Peng Luo, Chenxuan Hong, Naifeng Tian, Yaosen Wu, Yifei Zhou, Yan Lin, Haicheng Dou, Aimin Wu, Qishan Huang, Xiaolei Zhang, Xiangyang Wang.
      Oxidative stress-induced apoptosis and senescence of nucleus pulposus (NP) cells play a crucial role in the progression of intervertebral disc degeneration (IVDD). Accumulation of studies has shown that activated autophagy and enhanced autophagic flux can alleviate IVDD. In this study, we explored the effects of apigenin on IVDD in vitro and in vivo. Apigenin was found to inhibit tert-butyl hydroperoxide (TBHP)-induced apoptosis, senescence, and ECM degradation in NP cells. In addition, apigenin treatment can restore the autophagic flux blockage caused by TBHP. Mechanistically, we found that TBHP may induce autophagosome and lysosome fusion interruption and lysosomal dysfunction, while apigenin alleviates these phenomena by promoting the nuclear translocation of TFEB via the AMPK/mTOR signaling pathway. Furthermore, apigenin also exerts a protective effect against the progression of IVDD in the puncture-induced rat model. Taken together, these findings indicate that apigenin protects NP cells against TBHP-induced apoptosis, senescence, and ECM degradation via restoration of autophagic flux in vitro, and it also ameliorates IVDD progression in rats in vivo, demonstrating its potential for serving as an effective therapeutic agent for IVDD.
    Keywords:  TFEB; apoptosis; autophagic flux; autophagy; intervertebral disc degeneration; senescence
    DOI:  https://doi.org/10.3389/fcell.2021.787278
  34. Front Cell Dev Biol. 2021 ;9 806521
    Beyond Protein Synthesis; The Multifaceted Roles of Tuberin in Cell Cycle Regulation.
    E Fidalgo da Silva, J Fong, A Roye-Azar, A Nadi, C Drouillard, A Pillon, L A Porter.
      The ability of cells to sense diverse environmental signals, including nutrient availability and conditions of stress, is critical for both prokaryotes and eukaryotes to mount an appropriate physiological response. While there is a great deal known about the different biochemical pathways that can detect and relay information from the environment, how these signals are integrated to control progression through the cell cycle is still an expanding area of research. Over the past three decades the proteins Tuberin, Hamartin and TBC1D7 have emerged as a large protein complex called the Tuberous Sclerosis Complex. This complex can integrate a wide variety of environmental signals to control a host of cell biology events including protein synthesis, cell cycle, protein transport, cell adhesion, autophagy, and cell growth. Worldwide efforts have revealed many molecular pathways which alter Tuberin post-translationally to convey messages to these important pathways, with most of the focus being on the regulation over protein synthesis. Herein we review the literature supporting that the Tuberous Sclerosis Complex plays a critical role in integrating environmental signals with the core cell cycle machinery.
    Keywords:  TSC; cell cycle; cell growth; mTOR; tuberin; tuberous sclerosis complex
    DOI:  https://doi.org/10.3389/fcell.2021.806521
  35. Alzheimers Dement. 2021 Dec;17 Suppl 3 e054177
    DNAJC5 affects the endo-lysosomal pathway, APP processing, and AD pathology in vitro and in vivo.
    Niko Nykanen, Zhaohua Wang, Todd A Davis, Marie Nunez, Kevin O'Dell, John R Cirrito, Mark Sands, Bruno A Benitez.
      BACKGROUND: The DnaJ heat shock protein family member C5 (DNAJC5) gene encodes Cysteine String Protein-alpha (CSPα). CSPα is a key endo-lysosomal element of the misfolding-associated protein secretion (MAPS) machinery. MAPS eliminates misfolded cytosolic proteins, including alpha-synuclein, tau, TDP-43, huntingtin. Mutations in the DNAJC5 gene cause rare early-onset dementia called adult-onset Neuronal ceroid lipofuscinosis (ANCL). Data from CSPα-deficient mice and flies suggest that CSPα is critical for preventing age-dependent neurodegeneration. The endo-lysosome plays an essential role in normal and abnormal Amyloid-beta precursor protein (APP) processing and subsequent β-amyloidogenesis in Alzheimer's disease (AD). However, the role of CSPα in APP processing, trafficking, and amyloidogenesis is not well understood.METHODS: We used histological staining and whole transcriptome data from ANCL, AD patients, and age-matched pathology-free controls. Differential expression (DE) analysis was performed using DESeq2 software. We performed histological analysis of 5XFAD mouse models crossed with DNAJC5 mice. We used mouse neuroblastoma (N2A) cells stably expressing wild-type human APP695 (N2A695) and human wild-type (WT) and mutant DNAJC5. We used ELISA to quantify Aβ40 and Aβ42 in cell culture media and human brain lysates.
    RESULTS: CSPα co-localizes with endo-lysosomal and synaptic markers in N2A695 cells. CSPα overexpression affects lysosomal function and SNAP29-mediated exocytosis. Overexpression and knockdown of hCSPα-WT in N2A695 cells significantly affect extracellular Aβ40, Aβ42, full-length APP, and APP C-terminal fragments (CTF). N2A-APP cells expressing a gain-of-function DNAJC5 mutant displayed a significant increase in lysosomal and autophagy (LC3-II and p62) proteins, lysosomal exocytosis, and secreted levels of Aβ40 and Aβ42. ANCL brains showed considerable neuronal Aβ accumulation. ANCL brains exhibit a significant reduction of soluble and insoluble Aβ4 and Aβ42. Transcriptome analysis from ANCL brains shows changes in the mTOR pathway. DNAJC5 transcript levels are significantly reduced in AD cases compared to controls. A mouse AD model exhibits an inverse correlation between DNAJC5 transcript levels and Aβ plaques. 5XFAD mice haploinsufficient for DNAJC5 gene significantly increased the Aβ plaque burden and decreased Aβ plaque latency.
    CONCLUSIONS: Our results provide evidence of the novel and unexpected role of CSPα in endo-lysosomal function, lysosomal exocytosis, β-amyloidogenesis both in vitro and in vivo.
    DOI:  https://doi.org/10.1002/alz.054177
  36. Front Cell Neurosci. 2021 ;15 803302
    Neuroprogenitor Cells From Patients With TBCK Encephalopathy Suggest Deregulation of Early Secretory Vesicle Transport.
    Danielle de Paula Moreira, Angela May Suzuki, André Luiz Teles E Silva, Elisa Varella-Branco, Maria Cecília Zorél Meneghetti, Gerson Shigeru Kobayashi, Mariana Fogo, Merari de Fátima Ramires Ferrari, Rafaela Regina Cardoso, Naila Cristina Vilaça Lourenço, Karina Griesi-Oliveira, Elaine Cristina Zachi, Débora Romeo Bertola, Karina de Souza Weinmann, Marcelo Andrade de Lima, Helena Bonciani Nader, Andrea Laurato Sertié, Maria Rita Passos-Bueno.
      Biallelic pathogenic variants in TBCK cause encephaloneuropathy, infantile hypotonia with psychomotor retardation, and characteristic facies 3 (IHPRF3). The molecular mechanisms underlying its neuronal phenotype are largely unexplored. In this study, we reported two sisters, who harbored biallelic variants in TBCK and met diagnostic criteria for IHPRF3. We provided evidence that TBCK may play an important role in the early secretory pathway in neuroprogenitor cells (iNPC) differentiated from induced pluripotent stem cells (iPSC). Lack of functional TBCK protein in iNPC is associated with impaired endoplasmic reticulum-to-Golgi vesicle transport and autophagosome biogenesis, as well as altered cell cycle progression and severe impairment in the capacity of migration. Alteration in these processes, which are crucial for neurogenesis, neuronal migration, and cytoarchitecture organization, may represent an important causative mechanism of both neurodevelopmental and neurodegenerative phenotypes observed in IHPRF3. Whether reduced mechanistic target of rapamycin (mTOR) signaling is secondary to impaired TBCK function over other secretory transport regulators still needs further investigation.
    Keywords:  GM130; STAM; autophagy; clathrin; early secretory pathway; iPSC-neurodevelopmental disease modeling; mTOR; vesicle trafficking
    DOI:  https://doi.org/10.3389/fncel.2021.803302
  37. Reprod Fertil. 2021 Dec;2(4): R113-R129
    The role of mitophagy during oocyte aging in human, mouse, and Drosophila: implications for oocyte quality and mitochondrial disease.
    Rachel T Cox, Joanna Poulton, Suzannah A Williams.
      There is a worldwide trend for women to have their first pregnancy later in life. However, as oocyte quality declines with maternal aging, this trend leads to an increase in subfertility. The cellular mechanisms underlying this decline in oocyte competence are poorly understood. Oocyte mitochondria are the subcellular organelles that supply the energy that drives early embryogenesis, and thus their quality is critical for successful conception. Mitochondria contain their own DNA (mtDNA) and mutations in mtDNA cause mitochondrial diseases with severe symptoms, such as neurodegeneration and heart disease. Since mitochondrial function declines in tissues as humans age accompanied by an accumulation of mtDNA mutations, mtDNA is implicated as a cause of declining oocyte quality in older mothers. While this mutation load could be caused by declining accuracy of the mitochondrial replisome, age-related decline in mitochondrial quality control likely contributes, however knowledge is lacking. Mitophagy, a cellular process which specifically targets and recycles damaged mitochondria may be involved, but studies are scarce. And although assisted reproductive technologies can help older mothers, how these techniques affect the mechanisms that regulate mitochondrial and oocyte quality have not been studied. With the long-term goal of understanding the molecular mechanisms that control mitochondrial quality in the oocyte, model systems including Drosophila and mouse as well as human oocytes have been used. In this review, we explore the contribution of mitophagy to oocyte quality and the need for further systematic investigation in oocytes during maternal aging using different systems.Lay summary: Mitochondria are small parts of cells called organelles that generate the chemical energy needed for life. Hundreds of thousands of mitochondria in the developing eggs of the mother support the initial growth and development of the fertilized egg. However, due to increasingly diminished function over time, mitochondria generate less energy as we age, posing real problems for older women considering pregnancy. It is possible that this declining energy could be responsible for declining fertility as women age. Energy may decline because mitochondria fail and the cell's way of keeping them healthy become less efficient as we age. This review summarizes what is known about mitochondrial quality control in developing eggs as they age. In the future, understanding how the best mitochondria are selected and maintained in the egg, and hence the future baby, may enable older women with or without mitochondrial problems, to have healthy children.
    Keywords:  ARTs; Drosophila; human; mitochondria; mitophagy; mouse; mtDNA; oocyte; ovary
    DOI:  https://doi.org/10.1530/RAF-21-0060
  38. J Affect Disord. 2022 Jan 28. pii: S0165-0327(22)00115-X. [Epub ahead of print]
    mTORC1 inhibitor effects on rapid ketamine-induced reductions in suicidal ideation in patients with treatment-resistant depression.
    Lynnette A Averill, Christopher L Averill, Ralitza Gueorguieva, Samar Fouda, Mohamed Sherif, Kyung-Heup Ahn, Mohini Ranganathan, Deepak Cyril D'Souza, Steven M Southwick, Gerard Sanacora, Ronald S Duman, John H Krystal, Chadi G Abdallah.
      Suicide is a public health crisis with limited treatment options. Ketamine has demonstrated rapid and robust improvements in suicidal ideation (SI). The parent study for the secondary pilot analyses presented here was a double-blind, cross-over trial that found pretreatment with the mechanistic target of rapamycin complex 1 (mTORC1) prolonged the antidepressant effects of ketamine. Here we examined the effect of mTORC1 inhibition on ketamine's antisuicidal effects. Twenty patients in a major depressive episode were randomized to pretreatment with oral rapamycin (6 mg) or placebo prior to IV ketamine (0.5 mg/kg). We found ketamine administration resulted in significant improvements across all measures with the largest effect at 24 hrs with only the Beck Scale for Suicide remaining significant at the two-week follow-up. There were no significant main effects of pretreatment. While these analyses are pilot in nature and overall severity of SI was relatively low, the antisuicidal findings (no effect of rapamycin) being in contrast to the antidepressant effects (prolonged effect with rapamycin), suggest the rapid-acting antisuicidal and antidepressant effects of ketamine may be mechanistically distinct and the trajectories of response, recovery, and relapse may be independent. These findings provide additional evidence of ketamine's antisuicidal effects and highlight the importance of future studies that continue to examine potential differences in mechanisms and trajectory of outcomes.
    Keywords:  antidepressants; ketamine; mTOR; major depressive disorders; suicidal ideation; suicidality
    DOI:  https://doi.org/10.1016/j.jad.2022.01.104
  39. Mol Biol Cell. 2022 Feb 02. mbcE21100526
    Quantitative and Time-Resolved Monitoring of Organelle and Protein Delivery to the Lysosome with A Tandem Fluorescent Halo-GFP reporter.
    M Rudinskiy, T J Bergmann, M Molinari.
      Lysosomal degradative compartments hydrolyze macromolecules to generate basic building blocks that fuel metabolic pathways in our cells. They also remove misfolded proteins and control size, function and number of cytoplasmic organelles via constitutive and regulated autophagy. These catabolic processes attract interest because their defective functioning is linked to human disease and their molecular components are promising pharmacologic targets. The capacity to quantitatively assess them is highly sought for. Here, we present a tandem-fluorescent reporter consisting of a HaloTag-GFP chimera appended at the C- or at the N-terminus of select polypeptides to monitor protein and organelle delivery to the lysosomal compartment. The Halo-GFP changes color upon fluorescent pulse with cell-permeable HaloTag ligands and, again, upon delivery to acidic, degradative lysosomal compartments, where the fluorescent ligand-associated HaloTag is relatively stable, whereas the GFP portion is not, as testified by loss of the green fluorescence and generation of a protease-resistant, fluorescent HaloTag fragment. The Halo-GFP tandem fluorescent reporter presented in our study allows quantitative and, crucially, time-resolved analyses of protein and organelle transport to the lysosomal compartment by high resolution confocal laser scanning microscopy, antibody-free electrophoretic techniques and flow cytometry.
    DOI:  https://doi.org/10.1091/mbc.E21-10-0526
  40. Chem Biol Interact. 2022 Jan 31. pii: S0009-2797(22)00046-1. [Epub ahead of print]354 109841
    Modulation of histone deacetylase, the ubiquitin proteasome system, and autophagy underlies the neuroprotective effects of venlafaxine in a rotenone-induced Parkinson's disease model in rats.
    Khalid A El-Saiy, Rabab H Sayed, Ayman E El-Sahar, Esraa A Kandil.
      Parkinson's disease (PD) is a neurodegenerative disease characterized by motor and non-motor symptoms. Impairment of the ubiquitin proteasome system (UPS) and autophagy has been suggested to contribute to α-synuclein accumulation, which is identified as the pathological hallmark of PD. Recently, alteration in histone-3 acetylation has also been found to be correlated to PD. Interestingly, the histone deacetylase 6 (HDAC6) enzyme, which regulates the acetylation of histone-3, was shown to be involved in autophagy. Venlafaxine is an antidepressant that was proposed to inhibit HDAC expression in depressive rats' hippocampi. In this study, we aimed to examine the ability of venlafaxine to inhibit striatal HDAC6 and to enhance α-synuclein clearance through the activation of the UPS and autophagy, in addition to treating depression, which is the most debilitating non-motor symptom, in a rotenone model of PD. Venlafaxine administration was noted to decrease α-synuclein accumulation and preserve dopaminergic neurons along with restoration of striatal dopamine levels and motor recovery. Its administration augmented the UPS and autophagic markers (beclin-1, p62, and LC3) with consequent modulation of apoptotic indicators (Bax/Bcl-2 ratio, cytochrome c, and caspase-3). Additionally, venlafaxine inhibited HDAC6 with further enhancement of autophagy and restoration of histone-3 acetylation with subsequent increases in survival gene expressions (Bcl-2 and brain-derived neurotrophic factor). Chloroquine (autophagy inhibitor) was used to indicate the proposed pathway. Moreover, venlafaxine hampered depressive symptoms and improved hippocampal noradrenaline and serotonin levels. Collectively, venlafaxine is suggested to display neuroprotective effects with improvement of motor and non-motor PD symptoms.
    Keywords:  Autophagy; Depression; Histone deacetylase 6 (HDAC6); Parkinson's disease (PD); Ubiquitin proteasome system (UPS); Venlafaxine
    DOI:  https://doi.org/10.1016/j.cbi.2022.109841
  41. Biochem Pharmacol. 2022 Jan 31. pii: S0006-2952(22)00027-2. [Epub ahead of print]197 114933
    Targeting lipophagy as a potential therapeutic strategy for nonalcoholic fatty liver disease.
    Hong-Ying Li, Zong-Gen Peng.
      Nonalcoholic fatty liver disease (NAFLD) is becoming an increasingly serious disease worldwide. Unfortunately, no specific drug has been approved to treat NAFLD. Accumulating evidence suggests that lipotoxicity, which is induced by an excess of intracellular triacylglycerols (TAGs), is a potential mechanism underlying the ill-defined progression of NAFLD. Under physiological conditions, a balance is maintained between TAGs and free fatty acids (FFAs) in the liver. TAGs are catabolized to FFAs through neutral lipolysis and/or lipophagy, while FFAs can be anabolized to TAGs through an esterification reaction. However, in the livers of patients with NAFLD, lipophagy appears to fail. Reversing this abnormal state through several lipophagic molecules (mTORC1, AMPK, PLIN, etc.) facilitates NAFLD amelioration; therefore, restoring failed lipophagy may be a highly efficient therapeutic strategy for NAFLD. Here, we outline the lipophagy phases with the relevant important proteins and discuss the roles of lipophagy in the progression of NAFLD. Additionally, the potential candidate drugs with therapeutic value targeting these proteins are discussed to show novel strategies for future treatment of NAFLD.
    Keywords:  Lipophagy; Lipotoxicity; Nonalcoholic fatty liver disease; Signalling molecular; Therapeutic target
    DOI:  https://doi.org/10.1016/j.bcp.2022.114933
  42. J Cell Mol Med. 2022 Feb 02.
    Acetylshikonin induces autophagy-dependent apoptosis through the key LKB1-AMPK and PI3K/Akt-regulated mTOR signalling pathways in HL-60 cells.
    Meng-Di Wu, Yuan-Ying Zhang, Shu-Ying Yi, Bei-Bei Sun, Jing Lan, Han-Ming Jiang, Gang-Ping Hao.
      Acetylshikonin (ASK) is a natural naphthoquinone derivative of traditional Chinese medicine Lithospermum erythrorhyzon. It has been reported that ASK has bactericidal, anti-inflammatory and antitumour effects. However, whether ASK induces apoptosis and autophagy in acute myeloid leukaemia (AML) cells and the underlying mechanism are still unclear. Here, we explored the roles of apoptosis and autophagy in ASK-induced cell death and the potential molecular mechanisms in human AML HL-60 cells. The results demonstrated that ASK remarkably inhibited the cell proliferation, viability and induced apoptosis in HL-60 cells through the mitochondrial pathway, and ASK promoted cell cycle arrest in the S-phase. In addition, the increased formation of autophagosomes, the turnover from light chain 3B (LC3B) I to LC3B II and decrease of P62 suggested the induction of autophagy by ASK. Furthermore, ASK significantly decreased PI3K, phospho-Akt and p-p70S6K expression, while enhanced phospho-AMP-activated protein kinase (AMPK) and phospho-liver kinase B1(LKB1) expression. The suppression of ASK-induced the conversion from LC3B I to LC3B II caused by the application of inhibitors of AMPK (compound C) demonstrated that ASK-induced autophagy depends on the LKB1/AMPK pathway. These data suggested that the autophagy induced by ASK were dependent on the activation of LKB1/AMPK signalling and suppression of PI3K/Akt/mTOR pathways. The cleavage of the apoptosis-related markers caspase-3 and caspase-9 and the activity of caspase-3 induced by ASK were markedly reduced by inhibitor of AMPK (compound C), an autophagy inhibitor 3-methyladenine (3-MA) and another autophagy inhibitor chloroquine (CQ). Taken together, our data reveal that ASK-induced HL-60 cell apoptosis is dependent on the activation of autophagy via the LKB1/AMPK and PI3K/Akt-regulated mTOR signalling pathways.
    Keywords:  HL-60 cells; LKB1/AMPK; PI3K/Akt/mTOR; acetylshikonin (ASK); apoptosis; autophagy
    DOI:  https://doi.org/10.1111/jcmm.17202
  43. Biotechnol Bioeng. 2022 Feb 03.
    Significant impact of mTORC1 and ATF4 pathways in CHO cell recombinant protein production induced by CDK4/6 inhibitor.
    Meiping Chang, Steven Huhn, Luke Nelson, Michael Betenbaugh, Zhimei Du.
      The CDK4/6 inhibitor has been shown to increase recombinant protein productivity in Chinese hamster ovary cells (CHO). Therefore, we investigated the mechanism that couples cell cycle inhibitor (CCI) treatment with protein productivity utilizing proteomics and phosphoproteomics. We identified mTORC1 as a critical early signaling event that preceded boosted productivity. Following CCI treatment, mTOR exhibited a transient increase in phosphorylation at a novel site that is also conserved in human and mouse. Upstream of mTORC1, increased phosphorylation of AKT1S1 and decreased phosphorylation of RB1 may provide molecular links between CDK4/6 inhibition and mTORC1. Downstream, increased EIF4EBP phosphorylation was observed, which can mediate cap-dependent translation. In addition, the collective effect of increased phosphorylation of RPS6, increased phosphorylation of regulators of RNA polymerase I, and increased protein expression in tRNA-aminoacylation pathway may contribute to enhancing the translational apparatus for increased productivity. In concert, an elevated stress response via GCN2/EIF2AK4-ATF4 axis persisted over the treatment course, which may link mTOR to downstream responses including the unfolded protein response (UPR) and autophagy to enhance proper protein folding and secretion. Together, this comprehensive proteomics and phosphoproteomics characterization of CCI treated CHO cells offers insights into understanding multiple aspects of signaling events resulting from CDK4/CDK6 inhibition. This article is protected by copyright. All rights reserved.
    Keywords:  Cell Signaling; Chinese hamster ovary; Mammalian biotechnology; Phosphoproteomics; Proteomics
    DOI:  https://doi.org/10.1002/bit.28050
  44. Acta Pharmacol Sin. 2022 Feb 01.
    Mitochondria homeostasis: Biology and involvement in hepatic steatosis to NASH.
    Yu-Feng Li, Zhi-Fu Xie, Qian Song, Jing-Ya Li.
      Mitochondrial biology and behavior are central to the physiology of liver. Multiple mitochondrial quality control mechanisms remodel mitochondrial homeostasis under physiological and pathological conditions. Mitochondrial dysfunction and damage induced by overnutrition lead to oxidative stress, inflammation, liver cell death, and collagen production, which advance hepatic steatosis to nonalcoholic steatohepatitis (NASH). Accumulating evidence suggests that specific interventions that target mitochondrial homeostasis, including energy metabolism, antioxidant effects, and mitochondrial quality control, have emerged as promising strategies for NASH treatment. However, clinical translation of these findings is challenging due to the complex and unclear mechanisms of mitochondrial homeostasis in the pathophysiology of NASH.
    Keywords:  NASH; liver; metabolism; mitochondria; mitochondrial homeostasis
    DOI:  https://doi.org/10.1038/s41401-022-00864-z
  45. Adv Cancer Res. 2022 ;pii: S0065-230X(21)00081-6. [Epub ahead of print]153 267-304
    Unraveling and targeting RAS-driven metabolic signaling for therapeutic gain.
    Jonathan M DeLiberty, Ryan Robb, Claire E Gates, Kirsten L Bryant.
      RAS mutations are among the most frequent oncogenic drivers observed in human cancers. With a lack of available treatment options, RAS-mutant cancers account for many of the deadliest cancers in the United States. Recent studies established that altered metabolic requirements are a hallmark of cancer, and many of these alterations are driven by aberrant RAS signaling. Specifically, RAS-driven cancers are characterized by upregulated glycolysis, the differential channeling of glycolytic intermediates, upregulated nutrient scavenging pathways such as autophagy and macropinocytosis, and altered glutamine utilization and mitochondrial function. This unique metabolic landscape promotes tumorigenesis, proliferation, survival in nutrient deficient environments and confers resistance to conventional cytotoxic and targeted therapies. Emerging work demonstrates how these dependencies can be therapeutically exploited in vitro and in vivo with many metabolic inhibitors currently in clinical trials. This review aims to outline the unique metabolic requirements induced by aberrant RAS signaling and how these altered dependencies present opportunities for therapeutic intervention.
    Keywords:  Autophagy; Glycolysis; Macropinocytosis; Metabolism; Mitochondria; RAS; Scavenging
    DOI:  https://doi.org/10.1016/bs.acr.2021.07.010
  46. Cancer Sci. 2022 Feb 02.
    MEK/ERK-mediated oncogenic signals promote secretion of extracellular vesicles by controlling lysosome function.
    Tomoya Hikita, Ryo Uehara, Reina E Itoh, Fumie Mitani, Mamiko Miyata, Takuya Yoshida, Rui Yamaguchi, Chitose Oneyama.
      Cancer cells secrete large amounts of extracellular vesicles (EVs) originating from multivesicular bodies (MVBs). Mature MVBs fuse either with the plasma membrane for release as EVs often referred as to exosomes or with lysosomes for degradation. However, the mechanisms regulating MVB fate remain unknown. Here, we investigated the regulators of MVB fate by analyzing the effects of signaling inhibitors on EV secretion from cancer cells engineered to secrete luciferase-labeled EVs. Inhibition of the oncogenic MEK/ERK pathway suppressed EV release and activated lysosome formation. MEK/ERK-mediated lysosomal inactivation impaired MVB degradation, resulting in increased EV secretion from cancer cells. Moreover, MEK/ERK inhibition prevented c-MYC expression and induced the nuclear translocation of MiT/TFE transcription factors, thereby promoting the activation of lysosome-related genes, including the gene encoding a subunit of vacuolar-type H+ -ATPase, which is responsible for lysosomal acidification and function. Furthermore, c-MYC upregulation was associated with lysosomal genes downregulation in MEK/ERK-activated renal cancer cells/tissues. These findings suggest that the MEK/ERK/c-MYC pathway controls MVB fate and promotes EV production in human cancers by inactivating lysosomal function.
    Keywords:  MEK/ERK; c-MYC; extracellular vesicles; lysosome; renal cancer
    DOI:  https://doi.org/10.1111/cas.15288
  47. Behav Brain Funct. 2022 Jan 29. 18(1): 3
    The interplay between oxidative stress and autophagy: focus on the development of neurological diseases.
    Marjan Talebi, Seyyed Ali Mohammadi Vadoud, Alireza Haratian, Mohsen Talebi, Tahereh Farkhondeh, Ali Mohammad Pourbagher-Shahri, Saeed Samarghandian.
      Regarding the epidemiological studies, neurological dysfunctions caused by cerebral ischemia or neurodegenerative diseases (NDDs) have been considered a pointed matter. Mount-up shreds of evidence support that both autophagy and reactive oxygen species (ROS) are involved in the commencement and progression of neurological diseases. Remarkably, oxidative stress prompted by an increase of ROS threatens cerebral integrity and improves the severity of other pathogenic agents such as mitochondrial damage in neuronal disturbances. Autophagy is anticipated as a cellular defending mode to combat cytotoxic substances and damage. The recent document proposes that the interrelation of autophagy and ROS creates a crucial function in controlling neuronal homeostasis. This review aims to overview the cross-talk among autophagy and oxidative stress and its molecular mechanisms in various neurological diseases to prepare new perceptions into a new treatment for neurological disorders. Furthermore, natural/synthetic agents entailed in modulation/regulation of this ambitious cross-talk are described.
    Keywords:  Autophagy; Neurodegenerative diseases; Neurological diseases; Oxidative stress; Signaling pathways, Alzheimer’s disease, Parkinson’s disease, Reactive Oxygen Species (ROS)
    DOI:  https://doi.org/10.1186/s12993-022-00187-3
  48. J Immunol. 2022 Jan 31. pii: ji2100969. [Epub ahead of print]
    Protection of Quiescence and Longevity of IgG Memory B Cells by Mitochondrial Autophagy.
    Srikanth Kodali, Min Li, Marietta M Budai, Min Chen, Jin Wang.
      The development of long-lived immune memory cells against pathogens is critical for the success of vaccines to establish protection against future infections. However, the mechanisms governing the long-term survival of immune memory cells remain to be elucidated. In this article, we show that the maintenance mitochondrial homeostasis by autophagy is critical for restricting metabolic functions to protect IgG memory B cell survival. Knockout of mitochondrial autophagy genes, Nix and Bnip3, leads to mitochondrial accumulation and increases in oxidative phosphorylation and fatty acid synthesis, resulting in the loss of IgG+ memory B cells in mice. Inhibiting fatty acid synthesis or silencing necroptosis gene Ripk3 rescued Nix-/-Bnip3-/- IgG memory B cells, indicating that mitochondrial autophagy is important for limiting metabolic functions to prevent cell death. Our results suggest a critical role for mitochondrial autophagy in the maintenance of immunological memory by protecting the metabolic quiescence and longevity of memory B cells.
    DOI:  https://doi.org/10.4049/jimmunol.2100969
  49. Front Med (Lausanne). 2021 ;8 813047
    Oridonin Alleviates LPS-Induced Depression by Inhibiting NLRP3 Inflammasome via Activation of Autophagy.
    Chunyan Li, Yuehua Zhu, Yuanyuan Wu, Meiyuan Fu, Yiling Wu, Yuehong Wu, Yinger Qiu, Hui Zhang, Mingxing Ding.
      Objective: Oridonin (Ori) is a diterpene compound that has multiple biological properties. Here, our study was conducted to observe the therapeutic effect of Ori on depression as well as to uncover the mechanism. Methods: Lipopolysaccharide (LPS)-induced depression models were established both in C57BL/6 mice and primary astrocytes, which were treated with Ori, autophagy agonist Rapamycin (Rap) and autophagy inhibitor 3-Methyladenine (3-MA). The depressive-like behaviors were assessed with behavioral tests. Autophagy was evaluated in the hippocampus and astrocytes by investigating autophagosomes under transmission electron microscope (TEM) and detecting LC3II/I, Beclin1 and P62 through western blotting. Astrocyte marker glial fibrillary acidic protein (GFAP) was investigated by immunofluorescence. NLRP3 inflammasome activation was evaluated by detecting IL-1β, NLRP3, ASC and Caspase-1 expression and reactive oxygen species (ROS) accumulation was quantified via DCFH-DA probe. Autolysosomes, autophagosomes and mitophagy were separately observed through mTag-Wasabi-LC3 plasmid, MitoTracker Deep Red staining, and TEM. Results: Our results showed that Ori administration alleviated LPS-induced depressive-like behaviors and increased GFAP expression in the hippocampus. Furthermore, Ori treatment promoted autophagy activation and cell viability as well as weakened NLRP3 inflammasome activation and ROS accumulation both in LPS-induced mice and astrocytes. Ori promoted the autophagic flux unblocked through enhancing fusion of autophagosomes with lysosomes as well as enhanced mitophagy in LPS-treated astrocytes. The therapeutic effect of Ori was enhanced by Rap and weakened by 3-MA. Conclusion: Collectively, our findings provided a promising antidepressant drug and uncovered that Ori alleviated LPS-induced depression by inhibiting NLRP3 inflammasome through activation of autophagy.
    Keywords:  LPS; NLRP3 inflammasome; Oridonin; autophagy; depression
    DOI:  https://doi.org/10.3389/fmed.2021.813047
  50. Brain Res Bull. 2022 Jan 31. pii: S0361-9230(22)00012-0. [Epub ahead of print]
    Repeated social defeat stress inhibits development of hippocampus neurons through mitophagy and autophagy.
    Lanmin Guo, Zhi-Mei Jiang, Rui-Xue Sun, Wei Pang, Xue Zhou, Mei-Ling Du, Meng-Xiang Chen, Xinyue Lv, Jing-Tao Wang.
      The social defeat stress model is commonly used to study depression and anxiety disorder, which can significantly affect the structure and function of neurons in the hippocampus; however, the relevant mechanism in neuronal loss has not been clearly defined. In the present study, a social defeat stress model was established in mice to evaluate the impact of social defeat stress on the structure of neurons in the hippocampus using Western blotting, immunofluorescence, Nissl staining, Golgi staining and transmission electron microscopy. The results demonstrated that social defeat stress leads to disruption of homeostasis in the hippocampus and the integrity of mitochondria in hippocampal neurons was markedly affected by enhanced mitophagy and autophagy resulting in inhibition of development and growth. These findings provide new insights into the mechanisms of neuronal development and growth due to social defeat stress, which should help in the development of new strategies to combat the effects of depression and anxiety disorder.
    Keywords:  Hippocampus neurons, Mitophagy, Autophagy; Social defeat stress
    DOI:  https://doi.org/10.1016/j.brainresbull.2022.01.009
  51. Biochim Biophys Acta Gen Subj. 2022 Jan 31. pii: S0304-4165(22)00015-0. [Epub ahead of print]1866(4): 130097
    Roles of humanin and derivatives on the pathology of neurodegenerative diseases and cognition.
    Kitti Thiankhaw, Kenneth Chattipakorn, Siriporn C Chattipakorn, Nipon Chattipakorn.
      BACKGROUND: Alzheimer's disease (AD), Parkinson's disease (PD), and age-related macular degeneration (AMD) are common among neurodegenerative diseases, but investigations into novel therapeutic approaches are currently limited. Humanin (HN) is a mitochondrial-derived peptide found in brain tissues of patients with familial AD and has been increasingly investigated in AD and other neurodegenerative diseases.SCOPE OF REVIEW: In this review, we summarize and discuss the effects of HN on the pathology of neurodegenerative diseases and cognition based on several studies from preclinical to clinical models. The association between cardiac ischemia-reperfusion (I/R) injury and brain are also included. Findings from in vitro studies and those involving mice provide the most fundamental information on the impact of HN and its potential association with clinical studies.
    MAJOR CONCLUSIONS: HN plays a considerable role in countering the progression and neuropathology of AD. Inhibition and reduction of oxidative stress and neuroinflammation of the original amyloid hypothesis is the mainstay mechanism. Multiple intracellular mechanisms will be elucidated, including those involved in the anti-apoptotic signaling cascades, the insulin signaling pathway, and mitochondrial function, and especially autophagic activity. These beneficial roles are also found following cardiac I/R injury. Cognitive improvement was found to be related to maintenance of synaptic integrity and neurotransmitter modulation. Small humanin-like peptide 2 demonstrates the neuroprotective effects in PD and AMD via prevention of mitochondrial loss.
    GENERAL SIGNIFICANCE: Comprehensive knowledge of HN effects on cognition and neurodegenerative diseases emphasizes its potential to treat a viable disease, as it ameliorates the pathogenesis of the disease.
    Keywords:  Autophagy; Brain; Dementia; Humanin; Mitochondria
    DOI:  https://doi.org/10.1016/j.bbagen.2022.130097
  52. Autophagy. 2022 Jan 31. 1-13
    Involvement of autophagy in exacerbation of eosinophilic airway inflammation in a murine model of obese asthma.
    Yuzo Suzuki, Yuya Aono, Norimichi Akiyama, Yasuoki Horiike, Hyogo Naoi, Ryo Horiguchi, Kiyoshi Shibata, Hironao Hozumi, Masato Karayama, Kazuki Furuhashi, Noriyuki Enomoto, Tomoyuki Fujisawa, Yutaro Nakamura, Naoki Inui, Takafumi Suda.
      Obesity is a common comorbidity in patients with asthma, and obese asthma patients present the most refractory phenotype among patients with severe asthma. Similar to the observations in non-obese asthma patients, clinical studies have revealed heterogeneity in obese asthma patients, including the occurrences of T helper (Th)2-high and Th2-low phenotypes. However, the mechanisms underlying obesity-related asthma are not completely understood. Though macroautophagy/autophagy is involved in asthma and obesity, its role in obesity-associated asthma is unknown. We hypothesized that autophagy is involved in the pathogenesis of obese asthma. For our investigations, we used high-fat diet-induced Atg5 (autophagy related 5)-deficient mice and epithelial cell-specific atg5-/- (Scgb1a1/CCSP-atg5-/-) obesity-induced mice. House dust mite (HDM)-sensitized atg5-/- obese mice exhibited marked eosinophilic inflammation and airway hyper-reactivity (AHR), compared to wild-type (WT) obese mice. Analyses of atg5-/- obese mice showed increased levels of Th2 cells but not ILC2s together with elevated expression of Th2 cytokines in the lung. In response to the HDM challenge, activated epithelial autophagy was observed in lean but not obese WT mice. Epithelium-specific deletion of Atg5 induced eosinophilic inflammation in Scgb1a1/CCSP-atg5-/- obese mice, and genetic analyses of epithelial cells from HDM-immunized atg5-/- obesity-induced mice showed an elevated expression of thymic stromal lymphopoietin (TSLP) and IL33. Notably, HDM-sensitized atg5-/- mice developed TSLP- and IL33-dependent eosinophilic inflammation and AHR. Our results suggest that autophagy contributes to the exacerbation of eosinophilic inflammation in obese asthma. Modulations of autophagy may be a therapeutic target in obesity-associated asthma.
    Keywords:  Asthma; IL33; autophagy; corticosteroid resistance; eosinophil inflammation; obesity; thymic stromal lymphopoietin
    DOI:  https://doi.org/10.1080/15548627.2022.2025571
  53. J Neurosci. 2022 Feb 04. pii: JN-RM-0449-21. [Epub ahead of print]
    A farnesyltransferase inhibitor restores cognitive deficits in Tsc2+/- mice through inhibition of Rheb1.
    Hiroko Sugiura, Tadayuki Shimada, Keiko Moriya-Ito, Jun-Ichi Goto, Hiroki Fujiwara, Rie Ishii, Hiroshi Shitara, Choji Taya, Satoshi Fujii, Toshiyuki Kobayashi, Okio Hino, Paul F Worley, Kanato Yamagata.
      Tuberous sclerosis complex (TSC) is caused by mutations in Tsc1 or Tsc2, whose gene products inhibit the small G-protein Rheb1. Rheb1 activates mTORC1, which may cause refractory epilepsy, intellectual disability and autism. The mTORC1 inhibitors have been used for TSC patients with intractable epilepsy. However, its effectiveness for cognitive symptoms remains unclear. We found a new signaling pathway for synapse formation through Rheb1 activation, but not mTORC1. Here, we show that treatment with the farnesyltransferase inhibitor lonafarnib increased unfarnesylated (inactive) Rheb1 levels and restored synaptic abnormalities in cultured Tsc2+/- neurons, whereas rapamycin did not enhance spine synapse formation. Lonafarnib treatment also restored the plasticity-related Arc expression in cultured Tsc2+/- neurons. Lonafarnib action was partly dependent on the Rheb1 reduction with syntenin. Oral administration of lonafarnib increased unfarnesylated protein levels without affecting mTORC1and MAP kinase signalings, and restored dendritic spine morphology in the hippocampi of male Tsc2+/- mice. In addition, lonafarnib treatment ameliorated contextual memory impairments and restored memory-related Arc expression in male Tsc2+/- mice in vivo Heterozygous Rheb1 knockout in male Tsc2+/- mice reproduced the results observed with pharmacological treatment. These results suggest that the Rheb1 activation may be responsible for synaptic abnormalities and memory impairments in Tsc2+/- mice, and its inhibition by lonafarnib could provide insight into potential treatment options for TSC-associated neuropsychiatric disorders (TANDs).SIGNIFICANCE STATEMENTTuberous sclerosis complex (TSC) is an autosomal dominant disease that causes neuropsychiatric symptoms, including intractable epilepsy, intellectual disability (ID) and autism. No pharmacological treatment for ID is reported so far. To develop a pharmacological treatment for ID, we investigated the mechanism of TSC and found Rheb1 activation is responsible for synaptic abnormalities in TSC neurons. To inhibit Rheb1 function, we used farnesyltransferase inhibitor lonafarnib, because farnesylation of Rheb1 is required for its activation. Lonafarnib treatment increased inactive Rheb1, and recovered proper synapse formation and plasticity-related Arc expression in TSC neurons. Furthermore, in vivo lonafarnib treatment restored contextual memory and Arc induction in TSC mice. Taken together, Rheb1 inhibition by lonafarnib could provide insight into potential treatments for TSC-associated ID.
    DOI:  https://doi.org/10.1523/JNEUROSCI.0449-21.2022
  54. Alzheimers Dement. 2021 Dec;17 Suppl 3 e056185
    Differential gene expression analysis for the identification of potential biomarkers for Niemann-Pick type C disease.
    Tamara Patricia Pradel-Bernal, Jared Rivera-Osorio, Maria Del Carmen Cardenas-Aguayo.
      BACKGROUND: Niemann-Pick disease (NP) is a rare neurovisceral lysosomal lipid storage disorder. NPC (NP type C) is characterized by the accumulation of unesterified cholesterol and other lipids within the cell, the function of lysosomes usually is impaired as well as the autophagy flux. To better understand the molecular pathways affected in this disease, we perform a bioinformatic study.METHOD: We used in silico methods to identify genes affected in this disease. The first approach was an analysis of microarray data from the public archive GEO2, accession number GSE124283. We studied gene expression of skin fibroblast from 55 individuals, 22 healthy controls (13 males and 9 females) and other 23 with NPC (12 males and 11 females), 16 patients suffering from infantile NPC, 5 juvenile NPC and 2 adults. The differential expression analysis was performed using limma package, genes with significative adjusted p value and fold change > 0.5 and < -0.5 were selected, we used topGO package to obtain the Gene Ontology.
    RESULT: The analysis revealed that cellular components enriched were vesicle membrane, vacuolar lumen lysosome, cytoplasm vesicle, cytoskeletal fiber, nucleus and nucleoplasm. For molecular function were affected catalytic activity, cadherin binding and enzyme binding, and the biological process enriched were the phagosome maturation, lysosome organization, vesicular transport, apoptotic process, gene expression, proteosome-mediated ubiquitin, cell cycle, cellular macromolecule biosynthetic process and regulation of cellular response to stress.
    CONCLUSION: Our results were consistent with changes in cell homeostasis reported from NPC patients. Abnormal protein degradation and metabolism might contribute to the higher level of mortality in this disease. We propose that some of the DEGs that are more related to NPC disease according to our study, could have value as potential biomarkers of this disease helping to unravel the molecular mechanisms involve in this disease.
    DOI:  https://doi.org/10.1002/alz.056185
  55. Front Cell Dev Biol. 2021 ;9 785979
    Dual Inhibition of mTORC1/2 Reduces Migration of Cholangiocarcinoma Cells by Regulation of Matrixmetalloproteinases.
    Katharina Joechle, Huda Jumaa, Kerstin Thriene, Claus Hellerbrand, Birte Kulemann, Stefan Fichtner-Feigl, Sven A Lang, Jessica Guenzle.
      Cholangiocarcinoma (CCA) is a rare but highly aggressive tumor entity for which systemic therapies only showed limited efficacy so far. As OSI-027-a dual kinase inhibitor targeting both mTOR complexes, mTORC1 and mTORC2 - showed improved anti-cancer effects, we sought to evaluate its impact on the migratory and metastatic capacity of CCA cells in vitro. We found that treatment with OSI-027 leads to reduced cell mobility and migration as well as a reduced surviving fraction in colony-forming ability. While neither cell viability nor proliferation rate was affected, OSI-027 decreased the expression of MMP2 and MMP9. Moreover, survival as well as anti-apoptotic signaling was impaired upon the use of OSI-027 as determined by AKT and MAPK blotting. Dual targeting of mTORC1/2 might therefore be a viable option for anti-neoplastic therapy in CCA.
    Keywords:  MTORC1/2; cholangiocarcinoma; invasion; matrix metalloproteinases; migration
    DOI:  https://doi.org/10.3389/fcell.2021.785979
  56. Alzheimers Dement. 2021 Dec;17 Suppl 3 e053844
    Lysosomal subcellular distribution in neurons: Are there synaptic lysosomes?
    César Augusto Vogado Cunha, Tatiana Burrinha, Claudia G Almeida.
      BACKGROUND: Despite Alzheimer's disease's first description 114 years ago1 , the mechanisms driving the most common late-onset forms' molecular etiology and neuropathophysiology remain elusive. While considering its elaborate display of pathological hallmarks, recent advances have described synapse loss as the most accurate correlate of disease progression2 . A wide array of data, including insights from human genetics, preclinical AD models, clinical research, and trials, has continuously linked a dysfunctional lysosomal network to pathological aging and disease onset. However, evidence correlating lysosomal dysfunction with early synapse loss is still lacking. Here, we aim to characterize the subcellular distribution of neuronal lysosomes, hypothesizing that these complex organelles localize to both presynaptic and postsynaptic terminals.METHOD: We have analyzed the subcellular distribution of synaptic and non-synaptic associated LAMP-1+ organelles in our established mature primary hippocampal and cortical mouse neuronal in vitro model and adult mouse brain synaptosomes. We applied quantitative single-cell analysis using ICY and ImageJ software for analytical purposes.
    RESULT: Our results show that the subcellular distribution of (endo)lysosomes is highly compartmentalized and dynamic. Based on the % of (endo)lysosomes in the neuronal soma, we documented that roughly 42% of (endo)lysosomes distribute along dendrites and 37% along the axon initial segment (AIS). Remarkably, we report that close to 8% of the total (endo)lysosomal pool was associated with synapsin, a presynaptic compartment marker. We also reveal that about 26% of (endo)lysosomes colocalized with PSD-95, a postsynaptic compartment marker, constituting 60% of the dendritic (endo)lysosomes. Moreover, we found LAMP-1 enriched in mouse brain synaptosomes against nuclear, non-nuclear, non-synaptosomal and membrane fractions. We are currently pushing to characterize these synaptic (endo)lysosomes' function.
    CONCLUSION: These results suggest that (endo)lysosomes are present at synapses. In the future we would like to establish whether its dysfunctional activity and trafficking act as a catalyst in the very early stages of synaptic dysfunction in the aging brain, potentially contributing to LOAD etiology and development. References: (1) Alzheimer, A., 1907. Uber einen eigenartigc Erkrankung der Hirnrinde. Allgemeine Zeitschrift fur Psyciatrie und Psychisch-Gerichtliche Medizin 64, 3. (2) Tampellini, D., Gouras, G., 2010. Synapses, synaptic activity and intraneuronal Aβ in Alzheimer's disease. Front Aging Neurosci. 2, 13. DOI: 10.3389/fnagi.2010.00013.
    DOI:  https://doi.org/10.1002/alz.053844
  57. Front Pharmacol. 2021 ;12 825425
    The Effects of Combined Therapy With Metformin and Hydroxypropyl-β-Cyclodextrin in a Mouse Model of Niemann-Pick Disease Type C1.
    Jiang Du, Xinlei Liu, Yan Zhang, Xiaojing Han, Chunya Ma, Yanli Liu, Lihong Guan, Liang Qiao, Juntang Lin.
      Niemann-Pick disease type C1 (NPC1) is a neurodegenerative disorder characterized by lysosomal storage of free cholesterol. 2-Hydroxypropyl-β-cyclodextrin (HPβCD) is a cyclic oligosaccharide derivative that is being developed to treat NPC1. Recently, metformin was reported to be beneficial in various neurodegenerative diseases, such as Alzheimer's and Huntington's diseases. In this study, we examined the effects of combined treatment with HPβCD and metformin on Npc1 -/- mice. Unfortunately, body weight and survival rates showed that cotreatment with metformin did not extend survival time and increase the body weight of HPβCD-treated Npc1 -/- mice. However, cotreatment with metformin reduced inflammatory response and inhibited the proinflammatory cytokine release in the brain, liver and spleen of HPβCD-treated Npc1 -/- mice. Furthermore, metformin did not reduce the free cholesterol levels in Npc1 -/- brain tissue or fibroblasts. In conclusion, our results demonstrate that metformin does not show beneficial effects on body weight or survival time but reduced the inflammatory response in a mouse model of NPC1 when combined with HPβCD.
    Keywords:  HPβCD; NPC1 disease; cholesterol accumulation; combined therapy; metformin
    DOI:  https://doi.org/10.3389/fphar.2021.825425
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