bims-auttor Biomed News
on Autophagy and mTOR
Issue of 2023–09–10
fifty-one papers selected by
Viktor Korolchuk, Newcastle University
  1. PTEN activates chaperone-mediated autophagy to regulate metabolism.
  2. Lysosomes mediate the mitochondrial UPR via mTORC1-dependent ATF4 phosphorylation.
  3. Epigenetic regulation of autophagy-related genes: Implications for neurodevelopmental disorders.
  4. Neurodegenerative disease-associated inclusion bodies are cleared by selective autophagy in budding yeast.
  5. The separate axes of TECPR1 and ATG16L1 in CASM.
  6. CCDC50 promotes tumor growth through regulation of lysosome homeostasis.
  7. Relating aging and autophagy: a new perspective towards the welfare of human health.
  8. Caveolin-3 loss linked with the P104L LGMD-1C mutation modulates skeletal muscle mTORC1 signalling and cholesterol homeostasis.
  9. Autophagy-mediated surveillance of Rim4-mRNA interaction safeguards programmed meiotic translation.
  10. Iron-inhibited autophagy via transcription factor ZFP27 in Parkinson's disease.
  11. Mammalian autophagosomes form from finger-like phagophores.
  12. 4E-BP1 expression in embryonic postmitotic neurons mitigates mTORC1-induced cortical malformations and behavioral seizure severity but does not prevent epilepsy in mice.
  13. The biased M3 mAChR ligand PD 102807 mediates qualitatively distinct signaling to regulate airway smooth muscle phenotype.
  14. Altered Expression of Autophagy Biomarkers in Hippocampal Neurons in a Multiple Sclerosis Animal Model.
  15. TFEB regulates cellular labile iron and prevents ferroptosis in a TfR1-dependent manner.
  16. mTOR Signaling: New Insights into Cancer, Cardiovascular Diseases, Diabetes and Aging.
  17. Multifaceted membrane interactions of human Atg3 promote LC3-phosphatidylethanolamine conjugation during autophagy.
  18. A twin UGUA motif directs the balance between gene isoforms through CFIm and the mTORC1 signaling pathway.
  19. Podocyte Injury in Diabetic Kidney Disease in a Mouse Model Involves TRPC6-mediated Calpain Activation Impairing Autophagy.
  20. Astrovirus replication is dependent on induction of double-membrane vesicles through a PI3K-dependent, LC3-independent pathway.
  21. Lysosome-targeted fluorescent probes: Design mechanism and biological applications.
  22. A non-canonical role: The Hippo pathway regulator MAP4K2 controls autophagy and cell survival upon energy stress.
  23. Degradation of Cyclin-Dependent Kinase 9/Cyclin T1 by Optimized Microtubule-Associated Protein 1 Light Chain 3 Beta-Recruiting Coumarin Analogs.
  24. The nonautophagic functions of autophagy-related proteins.
  25. Pharmacotherapy developments in Autophagy inhibitors for bladder cancer.
  26. Rescuing the Golgi from heat damages by ATG8: restoration rather than clean-up.
  27. HOPS-dependent lysosomal fusion controls Rab19 availability for ciliogenesis in polarized epithelial cells.
  28. Fatty acid oxidation regulates cellular senescence by modulating the autophagy-SIRT1 axis.
  29. Coat protein of rice stripe virus enhances autophagy activity through interaction with cytosolic glyceraldehyde-3-phosphate dehydrogenases, a negative regulator of plant autophagy.
  30. Inhibition of mitochondrial fatty acid β-oxidation activates mTORC1 pathway and protein synthesis via Gcn5-dependent acetylation of raptor in zebrafish.
  31. CD44 connects autophagy decline and ageing in the vascular endothelium.
  32. RUNDC1 inhibits autolysosome formation and survival of zebrafish via clasping ATG14-STX17-SNAP29 complex.
  33. A selective ER-phagy exerts neuroprotective effects via modulation of α-synuclein clearance in parkinsonian models.
  34. Kangfuxin liquid reduces the ultraviolet B-induced photodamage of HaCaT cells by regulating autophagy.
  35. ROS-mediated mitophagy and necroptosis regulate osteocytes death caused by TCP particles in MLO-Y4 cells.
  36. Kuwanon H Inhibits Melanoma Growth through Cytotoxic Endoplasmic Reticulum Stress and Impaired Autophagy Flux.
  37. Mitochondrial dynamics in health and disease: mechanisms and potential targets.
  38. The Combination of Gold and Silver Food Nanoparticles with Gluten Peptides Alters the Autophagic Pathway in Intestinal Crypt-like Cells.
  39. The role and implication of autophagy in cholangiocarcinoma.
  40. ATG3 proteins possess a unique amphipathic α-helix essential for the Atg8/LC3 lipidation reaction.
  41. Fighting Against Drug-resistant Tumor by The Induction of Excessive Mitophagy with Transferrin Nanomedicine.
  42. Crosstalk between autophagy and metabolic regulation of (CAR) T cells: therapeutic implications.
  43. 3-Acetyldeoxynivalenol induces apoptosis, barrier dysfunction and endoplasmic reticulum stress by inhibiting mTORC1-dependent autophagy in porcine enterocytes.
  44. Glucolipotoxicity Suppressed Autophagy and Insulin Contents in Human Islets, and Attenuation of PERK Activity Enhanced Them in an ATG7-Dependent Manner.
  45. ERK1/2-Dependent Phosphorylation of GABAB1(S867/T872), Controlled by CaMKIIβ, Is Required for GABAB Receptor Degradation under Physiological and Pathological Conditions.
  46. The role of glial autophagy in Alzheimer's disease.
  47. MAGED2 Depletion Promotes Stress-Induced Autophagy by Impairing the cAMP/PKA Pathway.
  48. Damaged mitochondria recruit the effector NEMO to activate NF-κB signaling.
  49. The N-end rule pathway regulates ER stress-induced clusterin release to the cytosol where it directs misfolded proteins for degradation.
  50. Autophagy/ferroptosis in colorectal cancer: Carcinogenic view and nanoparticle-mediated cell death regulation.
  51. PPARɑ Ligand Caudatin Improves Cognitive Functions and Mitigates Alzheimer's Disease Defects By Inducing Autophagy in Mice Models.
  1. Autophagy. 2023 Sep 08. 1-2
    PTEN activates chaperone-mediated autophagy to regulate metabolism.
    S Joseph Endicott, Richard A Miller.
      PTEN is a negative modulator of the INS-PI3K-AKT pathway and is an essential regulator of metabolism and cell growth. PTEN is one of the most commonly mutated tumor suppressors in cancer. However, PTEN overexpression extends the lifespan of both sexes of mice. We recently showed that PTEN is necessary and sufficient to activate chaperone-mediated autophagy (CMA) in the mouse liver and cultured cells. Selective protein degradation via CMA is required to suppress glycolysis and fatty acid synthesis when PTEN is overexpressed. Thus, activation of CMA downstream of PTEN might modulate health and metabolism through selective degradation of key metabolic enzymes.
    Keywords:  Aging; PTEN; autophagy; chaperone-mediated autophagy; metabolism
    DOI:  https://doi.org/10.1080/15548627.2023.2255966
  2. Cell Discov. 2023 Sep 07. 9(1): 92
    Lysosomes mediate the mitochondrial UPR via mTORC1-dependent ATF4 phosphorylation.
    Terytty Yang Li, Qi Wang, Arwen W Gao, Xiaoxu Li, Yu Sun, Adrienne Mottis, Minho Shong, Johan Auwerx.
      Lysosomes are central platforms for not only the degradation of macromolecules but also the integration of multiple signaling pathways. However, whether and how lysosomes mediate the mitochondrial stress response (MSR) remain largely unknown. Here, we demonstrate that lysosomal acidification via the vacuolar H+-ATPase (v-ATPase) is essential for the transcriptional activation of the mitochondrial unfolded protein response (UPRmt). Mitochondrial stress stimulates v-ATPase-mediated lysosomal activation of the mechanistic target of rapamycin complex 1 (mTORC1), which then directly phosphorylates the MSR transcription factor, activating transcription factor 4 (ATF4). Disruption of mTORC1-dependent ATF4 phosphorylation blocks the UPRmt, but not other similar stress responses, such as the UPRER. Finally, ATF4 phosphorylation downstream of the v-ATPase/mTORC1 signaling is indispensable for sustaining mitochondrial redox homeostasis and protecting cells from ROS-associated cell death upon mitochondrial stress. Thus, v-ATPase/mTORC1-mediated ATF4 phosphorylation via lysosomes links mitochondrial stress to UPRmt activation and mitochondrial function resilience.
    DOI:  https://doi.org/10.1038/s41421-023-00589-1
  3. Autophagy. 2023 Sep 06. 1-14
    Epigenetic regulation of autophagy-related genes: Implications for neurodevelopmental disorders.
    Elly I Lewerissa, Nael Nadif Kasri, Katrin Linda.
      Macroautophagy/autophagy is an evolutionarily highly conserved catabolic process that is important for the clearance of cytosolic contents to maintain cellular homeostasis and survival. Recent findings point toward a critical role for autophagy in brain function, not only by preserving neuronal health, but especially by controlling different aspects of neuronal development and functioning. In line with this, mutations in autophagy-related genes are linked to various key characteristics and symptoms of neurodevelopmental disorders (NDDs), including autism, micro-/macrocephaly, and epilepsy. However, the group of NDDs caused by mutations in autophagy-related genes is relatively small. A significant proportion of NDDs are associated with mutations in genes encoding epigenetic regulatory proteins that modulate gene expression, so-called chromatinopathies. Intriguingly, several of the NDD-linked chromatinopathy genes have been shown to regulate autophagy-related genes, albeit in non-neuronal contexts. From these studies it becomes evident that tight transcriptional regulation of autophagy-related genes is crucial to control autophagic activity. This opens the exciting possibility that aberrant autophagic regulation might underly nervous system impairments in NDDs with disturbed epigenetic regulation. We here summarize NDD-related chromatinopathy genes that are known to regulate transcriptional regulation of autophagy-related genes. Thereby, we want to highlight autophagy as a candidate key hub mechanism in NDD-related chromatinopathies.Abbreviations: ADNP: activity dependent neuroprotector homeobox; ASD: autism spectrum disorder; ATG: AutTophaGy related; CpG: cytosine-guanine dinucleotide; DNMT: DNA methyltransferase; EHMT: euchromatic histone lysine methyltransferase; EP300: E1A binding protein p300; EZH2: enhancer of zeste 2 polycomb repressive complex 2 subunit; H3K4me3: histone 3 lysine 4 trimethylation; H3K9me1/2/3: histone 3 lysine 9 mono-, di-, or trimethylation; H3K27me2/3: histone 3 lysine 27 di-, or trimethylation; hiPSCs: human induced pluripotent stem cells; HSP: hereditary spastic paraplegia; ID: intellectual disability; KANSL1: KAT8 regulatory NSL complex subunit 1; KAT8: lysine acetyltransferase 8; KDM1A/LSD1: lysine demethylase 1A; MAP1LC3B: microtubule associated protein 1 light chain 3 beta; MTOR: mechanistic target of rapamycin kinase; MTORC1: mechanistic target of rapamycin complex 1; NDD: neurodevelopmental disorder; PHF8: PHD finger protein 8; PHF8-XLID: PHF8-X linked intellectual disability syndrome; PTM: post-translational modification; SESN2: sestrin 2; YY1: YY1 transcription factor; YY1AP1: YY1 associated protein 1.
    Keywords:  Autophagy related gene expression; chromatinopathies; epigenetics; neurodevelopmental disorders; neuronal autophagy
    DOI:  https://doi.org/10.1080/15548627.2023.2250217
  4. Autophagy Rep. 2023 ;pii: 2236407. [Epub ahead of print]2(1):
    Neurodegenerative disease-associated inclusion bodies are cleared by selective autophagy in budding yeast.
    Austin Folger, Chuan Chen, Marie-Helene Kabbaj, Karina Frey, Yanchang Wang.
      Protein misfolding, aggregation, and accumulation cause neurodegenerative disorders. One such disorder, Huntington's disease, is caused by an increased number of glutamine-encoding trinucleotide repeats CAG in the first exon of the huntingtin (HTT) gene. Mutant proteins of Htt exon 1 with polyglutamine expansion are prone to aggregation and form pathological inclusion bodies in neurons. Extensive studies have shown that misfolded proteins are cleared by the ubiquitin-proteasome system or autophagy to alleviate their cytotoxicity. Misfolded proteins can form small soluble aggregates or large insoluble inclusion bodies. Previous works have elucidated the role of autophagy in the clearance of misfolded protein aggregates, but autophagic clearance of inclusion bodies remains poorly characterized. Here we use mutant Htt exon 1 with 103 polyglutamine (Htt103QP) as a model substrate to study the autophagic clearance of inclusion bodies in budding yeast. We found that the core autophagy-related proteins were required for Htt103QP inclusion body autophagy. Moreover, our evidence indicates that the autophagy of Htt103QP inclusion bodies is selective. Interestingly, Cue5/Tollip, a known autophagy receptor for aggrephagy, is dispensable for this inclusion body autophagy. From the known selective autophagy receptors in budding yeast, we identified three that are essential for inclusion body autophagy. Amyloid beta peptide (Aβ42) is a major component of amyloid plaques found in Alzheimer's disease brains. Interestingly, a similar selective autophagy pathway contributes to the clearance of Aβ42 inclusion bodies in budding yeast. Therefore, our results reveal a novel autophagic pathway specific for inclusion bodies associated with neurodegenerative diseases, which we have termed IBophagy.
    Keywords:  Autophagy; Aβ42; IBophagy; inclusion body; mutant Huntingtin
    DOI:  https://doi.org/10.1080/27694127.2023.2236407
  5. Autophagy. 2023 Sep 07. 1-2
    The separate axes of TECPR1 and ATG16L1 in CASM.
    Namrita Kaur, Sven R Carlsson, Alf Håkon Lystad.
      Conjugation of ATG8 to single membranes (CASM) is a fundamental cellular process that entails the conjugation of mammalian Atg8 homologs, here referred to as ATG8, to phosphatidylethanolamine (PE) and phosphatidylserine (PS) on endolysosomal compartments. Our current research, together with recent reports from the Randow, Wu, and Wileman labs, has uncovered yet another layer to this process. We discovered that, in addition to ATG16L1-containing complexes, TECPR1 (tectonin beta-propeller repeat containing 1)-containing ATG12-ATG5 E3 complexes can facilitate CASM, thereby providing a broader understanding of this pathway.
    Keywords:  CASM; DysF; SopF; membrane damage; non-canonical autophagy; sphingomyelin
    DOI:  https://doi.org/10.1080/15548627.2023.2255462
  6. EMBO Rep. 2023 Sep 06. e56948
    CCDC50 promotes tumor growth through regulation of lysosome homeostasis.
    Penghui Jia, Tian Tian, Zibo Li, Yicheng Wang, Yuxin Lin, Weijie Zeng, Yu Ye, Miao He, Xiangrong Ni, Ji'an Pan, Xiaonan Dong, Jian Huang, Chun-Mei Li, Deyin Guo, Panpan Hou.
      The maintenance of lysosome homeostasis is crucial for cell growth. Lysosome-dependent degradation and metabolism sustain tumor cell survival. Here, we demonstrate that CCDC50 serves as a lysophagy receptor, promoting tumor progression and invasion by controlling lysosomal integrity and renewal. CCDC50 monitors lysosomal damage, recognizes galectin-3 and K63-linked polyubiquitination on damaged lysosomes, and specifically targets them for autophagy-dependent degradation. CCDC50 deficiency causes the accumulation of ruptured lysosomes, impaired autophagic flux, and superfluous reactive oxygen species, consequently leading to cell death and tumor suppression. CCDC50 expression is associated with malignancy, progression to metastasis, and poor overall survival in human melanoma. Targeting CCDC50 suppresses tumor growth and lung metastasis, and enhances the effect of BRAFV600E inhibition. Thus, we demonstrate critical roles of CCDC50-mediated clearance of damaged lysosomes in supporting tumor growth, hereby identifying a potential therapeutic target of melanoma.
    Keywords:  CCDC50; cell death; lysophagy; lysosome damage; melanoma
    DOI:  https://doi.org/10.15252/embr.202356948
  7. EXCLI J. 2023 ;22 732-748
    Relating aging and autophagy: a new perspective towards the welfare of human health.
    Sougata Ghosh Chowdhury, Rachayeeta Ray, Parimal Karmakar.
      The most common factor that contributes to aging is the loss of proteostasis, resulting in an excess amount of non-functional/damaged proteins. These proteins lead to various age-associated phenotypes such as cellular senescence and dysfunction in the nutrient-sensing pathways. Despite the various factors that can contribute to aging, it is still a process that can be changed. According to recent advances in the field of biology, the ability to alter the pathways that are involved in aging can improve the lifespan of a person. Autophagy is a process that helps in preserving survival during stressful situations, such as starvation. It is a common component of various anti-aging interventions, including those that target the insulin/IGF-1 and rapamycin signaling pathways. It has been shown that altered autophagy is a common feature of old age and its impaired regulation could have significant effects on the aging process. This review aims to look into the role of autophagy in aging and how it can be used to improve one's health.
    Keywords:  aging; autophagy; life span; misfolded protein
    DOI:  https://doi.org/10.17179/excli2023-6300
  8. J Cachexia Sarcopenia Muscle. 2023 Sep 06.
    Caveolin-3 loss linked with the P104L LGMD-1C mutation modulates skeletal muscle mTORC1 signalling and cholesterol homeostasis.
    Dinesh S Shah, Raid B Nisr, Gabriela Krasteva-Christ, Harinder S Hundal.
       BACKGROUND: Caveolins are the principal structural components of plasma membrane caveolae. Dominant pathogenic mutations in the muscle-specific caveolin-3 (Cav3) gene isoform, such as the limb girdle muscular dystrophy type 1C (LGMD-1C) P104L mutation, result in dramatic loss of the Cav3 protein and pathophysiological muscle weakness/wasting. We hypothesize that such muscle degeneration may be linked to disturbances in signalling events that impact protein turnover. Herein, we report studies assessing the effects of Cav3 deficiency on mammalian or mechanistic target of rapamycin complex 1 (mTORC1) signalling in skeletal muscle cells.
    METHODS: L6 myoblasts were stably transfected with Cav3P104L or expression of native Cav3 was abolished by CRISPR/Cas9 genome editing (Cav3 knockout [Cav3KO]) prior to performing subcellular fractionation and immunoblotting, analysis of real-time mitochondrial respiration or fixed cell immunocytochemistry. Skeletal muscle from wild-type and Cav3-/- mice was processed for immunoblot analysis of downstream mTORC1 substrate phosphorylation.
    RESULTS: Cav3 was detected in lysosomal-enriched membranes isolated from L6 myoblasts and observed by confocal microscopy to co-localize with lysosomal-specific markers. Cav3P104L expression, which results in significant (~95%) loss of native Cav3, or CRISPR/Cas9-mediated Cav3KO, reduced amino acid-dependent mTORC1 activation. The decline in mTORC1-directed signalling was detected by immunoblot analysis of L6 muscle cells and gastrocnemius Cav3-/- mouse muscle as judged by reduced phosphorylation of mTORC1 substrates that play key roles in the initiation of protein synthesis (4EBP1S65 and S6K1T389 ). S6K1T389 and 4EBP1S65 phosphorylation reduced by over 75% and 80% in Cav3KO muscle cells and by over 90% and 30% in Cav3-/- mouse skeletal muscle, respectively. The reduction in protein synthetic capacity in L6 muscle cells was confirmed by analysis of puromycylated peptides using the SUnSET assay. Cav3 loss was also associated with a 26% increase in lysosomal cholesterol, and pharmacological manipulation of lysosomal cholesterol was effective in replicating the reduction in mTORC1 activity observed in Cav3KO cells. Notably, re-expression of Cav3 in Cav3KO myoblasts normalized lysosomal cholesterol content, which coincided with a recovery in protein translation and an associated increase in mTORC1-directed phosphorylation of downstream targets.
    CONCLUSIONS: Our findings indicate that Cav3 can localize on lysosomal membranes and is a novel regulator of mTORC1 signalling in muscle. Cav3 deficiency associated with the Cav3P104L mutation impairs mTORC1 activation and protein synthetic capacity in skeletal muscle cells, which may be linked to disturbances in lysosomal cholesterol trafficking and contribute to the pathology of LGMD-1C.
    Keywords:  LGMD-1C; amino acid; caveolin-3; caveolinopathy; lysosome; mTORC1; skeletal muscle
    DOI:  https://doi.org/10.1002/jcsm.13317
  9. Cell Rep. 2023 Sep 01. pii: S2211-1247(23)01062-8. [Epub ahead of print]42(9): 113051
    Autophagy-mediated surveillance of Rim4-mRNA interaction safeguards programmed meiotic translation.
    Rudian Zhang, Wenzhi Feng, Suhong Qian, Fei Wang.
      In yeast meiosis, autophagy is active and essential. Here, we investigate the fate of Rim4, a meiosis-specific RNA-binding protein (RBP), and its associated transcripts during meiotic autophagy. We demonstrate that Rim4 employs a nuclear localization signal (NLS) to enter the nucleus, where it loads its mRNA substrates before nuclear export. Upon reaching the cytoplasm, active autophagy selectively spares the Rim4-mRNA complex. During meiotic divisions, autophagy preferentially degrades Rim4 in an Atg11-dependent manner, coinciding with the release of Rim4-bound mRNAs for translation. Intriguingly, these released mRNAs also become vulnerable to autophagy. In vitro, purified Rim4 and its RRM-motif-containing variants activate Atg1 kinase in meiotic cell lysates and in immunoprecipitated (IP) Atg1 complexes. This suggests that the conserved RNA recognition motifs (RRMs) of Rim4 are involved in stimulating Atg1 and thereby facilitating selective autophagy. Taken together, our findings indicate that autophagy surveils Rim4-mRNA interaction to ensure stage-specific translation during meiosis.
    Keywords:  Atg1; Atg11; CP: Cell biology; Pab1; RBP; Rim4; mRNAs; meiosis; nucleus; selective autophagy
    DOI:  https://doi.org/10.1016/j.celrep.2023.113051
  10. J Cell Mol Med. 2023 Sep 05.
    Iron-inhibited autophagy via transcription factor ZFP27 in Parkinson's disease.
    Yinying Wang, Qian Wen, Rongsha Chen, Zhichao Gan, Xinwei Huang, Pengfei Wang, Xia Cao, Ninghui Zhao, Zhongshan Yang, Jinyuan Yan.
      Parkinson's disease (PD) is a challenge because of the ageing of the population and the disease's complicated pathogenesis. Accumulating evidence showed that iron and autophagy were involved in PD. Nevertheless, the molecular mechanism and role of iron and autophagy in PD are not yet elucidated. In the present study, it was shown that PD mice had significant motor dysfunction, increased iron content, less dopamine neurons and more α-synuclein accumulation in the substantia nigra. Meanwhile, PD mice treated with deferoxamine exhibited less iron content, relieved the dyskinesia and had a significant increase in dopamine neurons and a significant decrease in α-synuclein. Autophagy induced by LC3 was inhibited in PD models with iron treatment. Following verification showed that iron aggregation restrained insulin-like growth factor 2 (IGF2) and transcription factor zinc finger protein 27 (ZFP27) in PD models. In addition, LC3-induced autophagy flux was reduced with ZFP27 knockdown. Furthermore, ZFP27 affected autophagy by regulating LC3 promoter activity. These data suggest that iron deposition inhibits IGF2 and ZFP27 to reduce LC3-induced autophagy, and ultimately decrease dopamine neurons, accelerating PD progression. Our findings provide a novel insight that ZFP27-mediated iron-related autophagy and IGF2 may activate the downstream kinase gene to trigger autophagy in the PD model.
    Keywords:  IGF2; Parkinson's disease; ZFP27; autophagy; iron
    DOI:  https://doi.org/10.1111/jcmm.17946
  11. Dev Cell. 2023 Aug 30. pii: S1534-5807(23)00410-0. [Epub ahead of print]
    Mammalian autophagosomes form from finger-like phagophores.
    Claudia Puri, Matthew J Gratian, David C Rubinsztein.
      The sequence of morphological intermediates that leads to mammalian autophagosome formation and closure is a crucial yet poorly understood issue. Previous studies have shown that yeast autophagosomes evolve from cup-shaped phagophores with only one closure point, and mammalian studies have inferred that mammalian phagophores also have single openings. Our superresolution microscopy studies in different human cell lines in conditions of basal and nutrient-deprivation-induced autophagy identified autophagosome precursors with multifocal origins that evolved into unexpected finger-like phagophores with multiple openings before becoming more spherical structures. Compatible phagophore structures were observed with whole-mount and conventional electron microscopy. This sequence of events was visualized using advanced SIM2 superresolution live microscopy. The finger-shaped phagophore apertures remained open when ESCRT function was compromised. The efficient closure of autophagic structures is important for their release from the recycling endosome. This has important implications for understanding how autophagosomes form and capture various cargoes.
    Keywords:  CHMP2A; ESCRT complex; RAB11A; VPS4A; autophagosome; autophagy; mitophagy; phagophore; recycling endosome; superresolution microscopy
    DOI:  https://doi.org/10.1016/j.devcel.2023.08.016
  12. Front Neurosci. 2023 ;17 1257056
    4E-BP1 expression in embryonic postmitotic neurons mitigates mTORC1-induced cortical malformations and behavioral seizure severity but does not prevent epilepsy in mice.
    Lena H Nguyen, Manas Sharma, Angelique Bordey.
      Hyperactivation of the mechanistic target of rapamycin complex 1 (mTORC1) pathway during neurodevelopment leads to focal cortical malformations associated with intractable seizures. Recent evidence suggests that dysregulated cap-dependent translation downstream of mTORC1 contributes to cytoarchitectural abnormalities and seizure activity. Here, we examined whether reducing cap-dependent translation by expressing a constitutively active form of the translational repressor, 4E-BP1, downstream of mTORC1 would prevent the development of cortical malformations and seizures. 4E-BP1CA was expressed embryonically either in radial glia (neural progenitor cells) that generate cortical layer 2/3 pyramidal neurons or in migrating neurons destined to layer 2/3 using a conditional expression system. In both conditions, 4E-BP1CA expression reduced mTORC1-induced neuronal hypertrophy and alleviated cortical mislamination, but a subset of ectopic neurons persisted in the deep layers and the white matter. Despite the above improvements, 4E-BP1CA expression in radial glia had no effects on seizure frequency and further exacerbated behavioral seizure severity associated with mTORC1 hyperactivation. In contrast, conditional 4E-BP1CA expression in migratory neurons mitigated the severity of behavioral seizures but the seizure frequency remained unchanged. These findings advise against targeting 4E-BPs by 4E-BP1CA expression during embryonic development for seizure prevention and suggest the presence of a development-dependent role for 4E-BPs in mTORC1-induced epilepsy.
    Keywords:  4E-BP; cortical development; corticogenesis; epilepsy; mTOR; malformation of cortical development; seizures
    DOI:  https://doi.org/10.3389/fnins.2023.1257056
  13. J Biol Chem. 2023 Sep 01. pii: S0021-9258(23)02237-8. [Epub ahead of print] 105209
    The biased M3 mAChR ligand PD 102807 mediates qualitatively distinct signaling to regulate airway smooth muscle phenotype.
    Eric Tompkins, Bogdana Mimic, Raymond B Penn, Tonio Pera.
      Airway smooth muscle (ASM) cells attain a hypercontractile phenotype during obstructive airway diseases. We recently identified a biased M3 muscarinic acetylcholine receptor (mAChR) ligand, PD 102807, that induces GRK-/arrestin-dependent AMPK activation to inhibit TGF-β-induced hypercontractile ASM phenotype. Conversely, the balanced mAChR agonist, methacholine (MCh), activates AMPK yet does not regulate ASM phenotype. In the current study, we demonstrate that PD 102807- and MCh-induced AMPK activation both depend on Ca2+/calmodulin-dependent kinase kinases (CaMKKs). However, MCh-induced AMPK activation is calcium-dependent and mediated by CaMKK1 and CaMKK2 isoforms. In contrast, PD 102807-induced signaling is calcium-independent and mediated by the atypical subtype protein kinase C-iota and the CaMKK1 (but not CaMKK2) isoform. Both MCh- and PD 102807-induced AMPK activation involve the AMPK α1 isoform. PD 102807-induced AMPK α1 (but not AMPK α2) isoform activation mediates inhibition of the mammalian target of rapamycin complex 1 (mTORC1) in ASM cells, as demonstrated by increased Raptor phosphorylation as well as inhibition of phospho-S6 protein and SRE-luciferase activity. The mTORC1 inhibitor rapamycin and the AMPK activator metformin both mimic the ability of PD 102807 to attenuate TGF-β-induced α-smooth muscle actin expression (a marker of hypercontractile ASM). These data indicate that PD 102807 transduces a signaling pathway (AMPK-mediated mTORC1 inhibition) qualitatively distinct from canonical M3 mAChR signaling to prevent pathogenic remodeling of ASM, thus demonstrating PD 102807 is a biased M3 mAChR ligand with therapeutic potential for the management of obstructive airway disease.
    Keywords:  Asthma; G protein-coupled receptor; airway; airway smooth muscle; biased receptor pharmacology
    DOI:  https://doi.org/10.1016/j.jbc.2023.105209
  14. Int J Mol Sci. 2023 Aug 25. pii: 13225. [Epub ahead of print]24(17):
    Altered Expression of Autophagy Biomarkers in Hippocampal Neurons in a Multiple Sclerosis Animal Model.
    Sabrina Ceccariglia, Diego Sibilia, Ornella Parolini, Fabrizio Michetti, Gabriele Di Sante.
      Multiple Sclerosis (MS) is a chronic inflammatory disease that affects the brain and spinal cord. Inflammation, demyelination, synaptic alteration, and neuronal loss are hallmarks detectable in MS. Experimental autoimmune encephalomyelitis (EAE) is an animal model widely used to study pathogenic aspects of MS. Autophagy is a process that maintains cell homeostasis by removing abnormal organelles and damaged proteins and is involved both in protective and detrimental effects that have been seen in a variety of human diseases, such as cancer, neurodegenerative diseases, inflammation, and metabolic disorders. This study is aimed at investigating the autophagy signaling pathway through the analysis of the main autophagic proteins including Beclin-1, microtubule-associated protein light chain (LC3, autophagosome marker), and p62 also called sequestosome1 (SQSTM1, substrate of autophagy-mediated degradation) in the hippocampus of EAE-affected mice. The expression levels of Beclin-1, LC3, and p62 and the Akt/mTOR pathway were examined by Western blot experiments. In EAE mice, compared to control animals, significant reductions of expression levels were detectable for Beclin-1 and LC3 II (indicating the reduction of autophagosomes), and p62 (suggesting that autophagic flux increased). In parallel, molecular analysis detected the deregulation of the Akt/mTOR signaling. Immunofluorescence double-labeling images showed co-localization of NeuN (neuronal nuclear marker) and Beclin-1, LC3, and p62 throughout the CA1 and CA3 hippocampal subfields. Taken together, these data demonstrate that activation of autophagy occurs in the neurons of the hippocampus in this experimental model.
    Keywords:  autophagy; experimental autoimmune encephalomyelitis; hippocampus; mouse; neurons
    DOI:  https://doi.org/10.3390/ijms241713225
  15. Free Radic Biol Med. 2023 Sep 06. pii: S0891-5849(23)00622-6. [Epub ahead of print]
    TFEB regulates cellular labile iron and prevents ferroptosis in a TfR1-dependent manner.
    Leilei Chen, Yue Ma, Xizhen Ma, Lin Liu, Xianhui Jv, Ang Li, Qingqing Shen, Wenting Jia, Le Qu, Limin Shi, Junxia Xie.
      Autophagy is a major clearance pathway for misfolded α-synuclein which promotes ferroptosis through NCOA4-mediated ferritin degradation. The regulation of these two processes to achieve improved neuroprotection in Parkinson's disease (PD) must be elucidated. Transcription factor EB (TFEB) is a master regulator of both autophagy and lysosome biogenesis, and lysosomes are important cellular iron storage organelles; however, the role of TFEB in ferroptosis and iron metabolism remains unclear. In this study, TFEB overexpression promoted the clearance of misfolded α-synuclein and prevented ferroptosis and iron overload. TFEB overexpression up-regulated transferrin receptor 1 (TfR1) synthesis and increased the localization of TfR1 in the lysosome, facilitating lysosomal iron import and transient lysosomal iron storage. TFEB overexpression increased the levels of cellular iron-safe storage proteins (both ferritin light and heavy chains). These functions in iron metabolism maintain the cellular labile iron at a low level and electrical activity, even under iron overload conditions. Notably, lower levels of cellular labile iron and the upregulation of ferritin light and heavy chains were reversed after TfR1 knockdown in cells overexpressing TFEB, indicating that TFEB regulates cellular labile iron and suppresses ferroptosis in a TfR1 dependent manner. Taken together, this evidence of the regulation of iron metabolism enriches our understanding of the function of TFEB. In addition, TFEB overexpression protects against ferroptosis and iron overload and provides a new direction and perspective for autophagy regulation in PD.
    Keywords:  Ferroptosis; Iron metabolism; Lysosome; TFEB; TfR1
    DOI:  https://doi.org/10.1016/j.freeradbiomed.2023.09.004
  16. Int J Mol Sci. 2023 Sep 04. pii: 13628. [Epub ahead of print]24(17):
    mTOR Signaling: New Insights into Cancer, Cardiovascular Diseases, Diabetes and Aging.
    Anindita Das, Flávio Reis.
      The mechanistic/mammalian target of rapamycin (mTOR), a member of the phosphoinositide 3-kinase (PI3K) related kinase family, integrates intracellular and environmental cues that coordinate a diverse set of cellular/tissue functions, such as cell growth, proliferation, metabolism, autophagy, apoptosis, longevity, protein/lipid/nucleotide synthesis, and tissue regeneration and repair [...].
    DOI:  https://doi.org/10.3390/ijms241713628
  17. Nat Commun. 2023 Sep 07. 14(1): 5503
    Multifaceted membrane interactions of human Atg3 promote LC3-phosphatidylethanolamine conjugation during autophagy.
    Yansheng Ye, Erin R Tyndall, Van Bui, Maria C Bewley, Guifang Wang, Xupeng Hong, Yang Shen, John M Flanagan, Hong-Gang Wang, Fang Tian.
      Autophagosome formation, a crucial step in macroautophagy (autophagy), requires the covalent conjugation of LC3 proteins to the amino headgroup of phosphatidylethanolamine (PE) lipids. Atg3, an E2-like enzyme, catalyzes the transfer of LC3 from LC3-Atg3 to PEs in targeted membranes. Here we show that the catalytically important C-terminal regions of human Atg3 (hAtg3) are conformationally dynamic and directly interact with the membrane, in collaboration with its N-terminal membrane curvature-sensitive helix. The functional relevance of these interactions was confirmed by in vitro conjugation and in vivo cellular assays. Therefore, highly curved phagophoric rims not only serve as a geometric cue for hAtg3 recruitment, but also their interaction with hAtg3 promotes LC3-PE conjugation by targeting its catalytic center to the membrane surface and bringing substrates into proximity. Our studies advance the notion that autophagosome biogenesis is directly guided by the spatial interactions of Atg3 with highly curved phagophoric rims.
    DOI:  https://doi.org/10.1038/s41467-023-41243-4
  18. Elife. 2023 09 04. pii: e85036. [Epub ahead of print]12
    A twin UGUA motif directs the balance between gene isoforms through CFIm and the mTORC1 signaling pathway.
    R Samuel Herron, Alexander K Kunisky, Jessica R Madden, Vivian I Anyaeche, May Z Maung, Hun-Way Hwang.
      Alternative polyadenylation (APA) generates mRNA isoforms and diversifies gene expression. Here we report the discovery that the mTORC1 signaling pathway balances the expression of two Trim9/TRIM9 isoforms through APA regulation in human and mouse. We showed that CFIm components, CPSF6 and NUDT21, promote the short Trim9/TRIM9 isoform (Trim9-S/TRIM9-S) expression. In addition, we identified an evolutionarily conserved twin UGUA motif, UGUAYUGUA, in TRIM9-S polyadenylation site (PAS) that is critical for its regulation by CPSF6. We found additional CPSF6-regulated PASs with similar twin UGUA motifs in human and experimentally validated the twin UGUA motif functionality in BMPR1B, MOB4, and BRD4-L. Importantly, we showed that inserting a twin UGUA motif into a heterologous PAS was sufficient to confer regulation by CPSF6 and mTORC1. Our study reveals an evolutionarily conserved mechanism to regulate gene isoform expression by mTORC1 and implicates possible gene isoform imbalance in cancer and neurological disorders with mTORC1 pathway dysregulation.
    Keywords:  CLIP; Tsc1; alternative polyadenylation; chromosomes; gene expression; human; isoform; mTORC1; mouse; tuberous sclerosis complex
    DOI:  https://doi.org/10.7554/eLife.85036
  19. J Am Soc Nephrol. 2023 Sep 06.
    Podocyte Injury in Diabetic Kidney Disease in a Mouse Model Involves TRPC6-mediated Calpain Activation Impairing Autophagy.
    Yann Salemkour, Dilemin Yildiz, Léa Dionet, Daan C T Hart, Kim A T Verheijden, Ryuta Sato, Nassim Mahtal, Jean-Daniel Delbet, Emmanuel Letavernier, Marion Rabant, Alexandre Karras, Johan van der Vlag, Tom Nijenhuis, Pierre-Louis Tharaux, Olivia Lenoir.
       BACKGROUND: Diabetic kidney disease is associated with impaired podocyte autophagy and subsequent podocyte injury. The regulation of podocyte autophagy is unique, because it minimally utilizes the mTOR and AMPK pathways. Thus, the molecular mechanisms underlying the impaired autophagy in podocytes in diabetic kidney disease remain largely elusive.
    METHODS: This study investigated how the calcium channel TRPC6 and the cysteine protease calpains deleteriously affect podocyte autophagy in diabetic kidney disease in mice. We demonstrated that TRPC6 knockdown in podocytes increased the autophagic flux due to decreased cysteine protease calpains activity. Diabetic kidney disease was induced in vivo using streptozotocin with unilateral nephrectomy in the BTBRob/ob mouse model.
    RESULTS: Diabetes increased TRPC6 expression in podocytes in vivo with decreased podocyte autophagic flux. Transgenic overexpression of the endogenous calpain inhibitor calpastatin, as well as pharmacological inhibition of calpain activity, normalized podocyte autophagic flux, reduced nephrin loss, and prevented the development of albuminuria in diabetic mice. In kidney biopsies from patients with diabetes, we further confirmed that TRPC6 overexpression in podocytes correlates with decreased calpastatin expression, autophagy blockade, and podocyte injury.
    CONCLUSION: Overall, we discovered a new mechanism that connects TRPC6 and calpain activity to impaired podocyte autophagy, increased podocyte injury, and development of proteinuria in the context of diabetic kidney disease. Therefore, targeting TRPC6 and/or calpain to restore podocyte autophagy might be a promising therapeutic strategy for diabetic kidney disease.
    DOI:  https://doi.org/10.1681/ASN.0000000000000212
  20. J Virol. 2023 Sep 05. e0102523
    Astrovirus replication is dependent on induction of double-membrane vesicles through a PI3K-dependent, LC3-independent pathway.
    Theresa Bub, Virginia Hargest, Shaoyuan Tan, Maria Smith, Ana Vazquez-Pagan, Tim Flerlage, Pamela Brigleb, Victoria Meliopoulos, Brett Lindenbach, Harish N Ramanathan, Valerie Cortez, Jeremy Chase Crawford, Stacey Schultz-Cherry.
      Human astrovirus is a positive-sense, single-stranded RNA virus. Astrovirus infection causes gastrointestinal symptoms and can lead to encephalitis in immunocompromised patients. Positive-strand RNA viruses typically utilize host intracellular membranes to form replication organelles, which are potential antiviral targets. Many of these replication organelles are double-membrane vesicles (DMVs). Here, we show that astrovirus infection leads to an increase in DMV formation through a replication-dependent mechanism that requires some early components of the autophagy machinery. Results indicate that the upstream class III phosphatidylinositol 3-kinase (PI3K) complex, but not LC3 conjugation machinery, is utilized in DMV formation. Both chemical and genetic inhibition of the PI3K complex lead to significant reduction in DMVs, as well as viral replication. Elucidating the role of autophagy machinery in DMV formation during astrovirus infection reveals a potential target for therapeutic intervention for immunocompromised patients.IMPORTANCEThese studies provide critical new evidence that astrovirus replication requires formation of double-membrane vesicles, which utilize class III phosphatidylinositol 3-kinase (PI3K), but not LC3 conjugation autophagy machinery, for biogenesis. These results are consistent with replication mechanisms for other positive-sense RNA viruses suggesting that targeting PI3K could be a promising therapeutic option for not only astrovirus, but other positive-sense RNA virus infections.
    Keywords:  astrovirus; autophagy; class III PI3K; double-membrane vesicle; replication organelle
    DOI:  https://doi.org/10.1128/jvi.01025-23
  21. Bioorg Chem. 2023 Sep 03. pii: S0045-2068(23)00493-5. [Epub ahead of print]140 106832
    Lysosome-targeted fluorescent probes: Design mechanism and biological applications.
    Xiangning Duan, Qin Tong, Chengxiao Fu, Linxi Chen.
      As an integral organelle in the eukaryote, the lysosome is the degradation center and metabolic signal center in living cells, and partakes in significant physiological processes such as autophagy, cell death and cellular senescence. Fluorescent probe has become a favorite tool for studying organelles and their chemical microenvironments because of its high specificity and non-destructive merits. Over recent years, it has been reported that increasingly new lysosome-targeted probes play a major role in the diagnosis and monitor of diseases, in particular cancer and neurodegenerative diseases. In order to deepen the relevant research on lysosome, it is challenging and inevitability to design novel lysosomal targeting probes. This review first introduces the concepts of lysosome and its closely related biological activities, and then introduces the fluorescent probes for lysosome in detail according to different detection targets, including targeting mechanism, biological imaging, and application in diseases. Finally, we summarize the specific challenges and discuss the future development direction facing the current lysosome-targeted fluorescent probes. We hope that this review can help biologists grasp the application of fluorescent probes and broaden the research ideas of researchers targeting fluorescent probes so as to design more accurate and functional probes for application in diseases.
    Keywords:  Cell imaging; Fluorescent probes; Lysosome; Sensing microenvironment
    DOI:  https://doi.org/10.1016/j.bioorg.2023.106832
  22. Mol Cell. 2023 Sep 07. pii: S1097-2765(23)00657-3. [Epub ahead of print]83(17): 3043-3045
    A non-canonical role: The Hippo pathway regulator MAP4K2 controls autophagy and cell survival upon energy stress.
    Anne Schreiber.
      In this issue, Seo et al.1 report a non-canonical function of the Hippo kinase MAP4K2 in energy stress response by regulating autophagy and cell survival, with relevance and therapeutic potential for head and neck cancer treatment.
    DOI:  https://doi.org/10.1016/j.molcel.2023.08.020
  23. J Med Chem. 2023 Sep 06.
    Degradation of Cyclin-Dependent Kinase 9/Cyclin T1 by Optimized Microtubule-Associated Protein 1 Light Chain 3 Beta-Recruiting Coumarin Analogs.
    Yanping Zeng, Jian Xiao, Yuanxin Xu, Fan Wei, Lina Tian, Yinglei Gao, Yi Chen, Youhong Hu.
      Autophagy is an efficient and attractive protein degradation pathway in addition to the ubiquitin-proteasome system. Herein, systematic optimization of coumarin analogs linked with the CDK9 inhibitor SNS-032 is reported that may bind to cyclin-dependent kinase 9 (CDK9) and microtubule-associated protein 1 light chain 3 beta (LC3B) simultaneously, which leads to the selective autophagic degradation of targeted CDK9/cyclin T1 and is different from the PROTAC degrader THAL-SNS-032. Further mechanism studies revealed an autophagy-lysosome pathway, where the degraders possibly formed a ternary complex with CDK9 and LC3B. In addition, degrader 10 showed antitumor efficacy in vivo. Our work optimized a potent LC3B recruiter and demonstrated the feasibility of autophagy-tethering compounds (ATTECs), which could be applied for the degradation of diverse intracellular pathogenic proteins to treat related diseases.
    DOI:  https://doi.org/10.1021/acs.jmedchem.3c00828
  24. Autophagy. 2023 Sep 08. 1-15
    The nonautophagic functions of autophagy-related proteins.
    Jia-Ni Shang, Chen-Ge Yu, Rui Li, Yan Xi, Yue Jenny Jian, Nan Xu, Su Chen.
       ABBREVIATIONS: ATG: autophagy related; BECN1: beclin 1; cAMP: cyclic adenosine monophosphate; dsDNA: double-stranded DNA; EMT: epithelial-mesenchymal transition; IFN: interferon; ISCs: intestinal stem cells; MAP1LC3/LC3: microtubule associated protein 1 light chain 3; MAPK/JNK: mitogen-activated protein kinase/c-Jun N-terminal kinases; MTOR: mechanistic target of rapamycin kinase; STING1: stimulator of interferon response cGAMP interactor 1; UVRAG: UV radiation resistance associated; VPS: vacuolar protein sorting.
    Keywords:  autophagy-related proteins; cargo transport; cell growth and senescence; metabolic homeostasis; nonautophagic functions; signal transduction
    DOI:  https://doi.org/10.1080/15548627.2023.2254664
  25. Expert Opin Pharmacother. 2023 Sep 05.
    Pharmacotherapy developments in Autophagy inhibitors for bladder cancer.
    Celine Chaaya, Ghady Zgheib, Fadi El Karak.
       INTRODUCTION: Autophagy is an intracellular process that plays a key role in the cellular homeostasis. Recently, it has been described as a potential therapeutic target in oncology, whether by activating or inhibiting its different cascades. Autophagy inhibitors interact with different molecular processes of the hallmarks of cancer.
    AREAS COVERED: Multiple proteins of the autophagy cascade could be aimed by specific inhibitors in many tumors, notably bladder cancer. In fact, bladder cancer has been increasing in prevalence over the last decade, and resistance to conventional treatment has been extensively reported in the literature. Autophagy inhibitors in bladder cancer have been described in preclinical studies to increase the sensitivity of the tumor to chemotherapy and radiotherapy. This paper is a review of the literature, which selected randomized trials, cohort studies, case-control studies documenting the relationship between autophagy inhibitors and bladder cancer treatment.
    EXPERT OPINION: Autophagy is a promising pathway for cancer cell targeting that opens the horizons for a potential new therapeutic area in particular the multidisciplinary management of bladder cancer.
    Keywords:  Autophagy inhibitors; bladder cancer; hallmarks of cancer; novel therapy
    DOI:  https://doi.org/10.1080/14656566.2023.2254697
  26. Stress Biol. 2023 Jun 26. 3(1): 19
    Rescuing the Golgi from heat damages by ATG8: restoration rather than clean-up.
    Anni Luo, Jian-Xiang Liu.
      High temperature stress poses significant adverse effects on crop yield and quality. Yet the molecular mechanisms underlying heat stress tolerance in plants/crops, especially regarding the organellar remodeling and homeostasis, are largely unknown. In a recent study, Zhou et al. reported that autophagy-related 8 (ATG8), a famous regulator involved in autophagy, plays a new role in Golgi restoration upon heat stress. Golgi apparatus is vacuolated following short-term acute heat stress, and ATG8 is translocated to the dilated Golgi membrane and interacts with CLATHRIN LIGHT CHAIN 2 (CLC2) to facilitate Golgi restoration, which is dependent on the ATG conjugation system, but not of the upstream autophagic initiators. These exciting findings broaden the fundamental role of ATG8, and elucidate the organelle-level restoration mechanism of Golgi upon heat stress in plants.
    Keywords:  ATG8; Autophagy; CLC2; ER; Golgi restoration; Heat stress
    DOI:  https://doi.org/10.1007/s44154-023-00100-6
  27. J Cell Sci. 2024 03 01. pii: jcs261047. [Epub ahead of print]137(5):
    HOPS-dependent lysosomal fusion controls Rab19 availability for ciliogenesis in polarized epithelial cells.
    Huxley K Hoffman, Rytis Prekeris.
      Primary cilia are sensory cellular organelles crucial for organ development and homeostasis. Ciliogenesis in polarized epithelial cells requires Rab19-mediated clearing of apical cortical actin to allow the cilium to grow from the apically docked basal body into the extracellular space. Loss of the lysosomal membrane-tethering homotypic fusion and protein sorting (HOPS) complex disrupts this actin clearing and ciliogenesis, but it remains unclear how the ciliary function of HOPS relates to its canonical function in regulating late endosome-lysosome fusion. Here, we show that disruption of HOPS-dependent lysosomal fusion indirectly impairs actin clearing and ciliogenesis by disrupting the targeting of Rab19 to the basal body, and that this effect is specific to polarized epithelial cells. We also find that Rab19 functions in endolysosomal cargo trafficking in addition to having its previously identified role in ciliogenesis. In summary, we show that inhibition of lysosomal fusion leads to the abnormal accumulation of Rab19 on late endosomes, thus depleting Rab19 from the basal body and thereby disrupting Rab19-mediated actin clearing and ciliogenesis in polarized epithelial cells.
    Keywords:  Cilia; Epithelial polarity; HOPS complex; Lysosomal fusion; Rab19; Renal epithelia
    DOI:  https://doi.org/10.1242/jcs.261047
  28. BMB Rep. 2023 Sep 08. pii: 5938. [Epub ahead of print]
    Fatty acid oxidation regulates cellular senescence by modulating the autophagy-SIRT1 axis.
    Seungyeon Yang, Subin Moon, Soojung Claire Hur, Seung Min Jeong.
      Senescence, a cellular process through which damaged or dysfunctional cells suppress their cell cycle, contributes to aging or age-related functional declines. Cell metabolism has been closely correlated with aging processes and it is widely recognized that metabolic changes underlie cellular alterations with aging. Here, we report that fatty acid oxidation (FAO) serves as a critical regulator of cellular senescence and uncover the underlying mechanism by which FAO inhibition induces senescence. Pharmacological or genetic ablation of FAO results in a p53-dependent induction of cellular senescence in human fibroblasts, whereas enhancing FAO suppresses replicative senescence. We find that FAO inhibition promotes cellular senescence through acetyl-CoA, independent of energy depletion. Mechanistically, the increased formation of autophagosome following FAO inhibition leads to a reduction in SIRT1 protein levels, thereby contributing to senescence induction. Finally, we find that the inhibition of autophagy or the enforced expression of SIRT1 can rescue the induction of senescence as a result of FAO inhibition. Collectively, our study reveals a distinctive role for the FAO-autophagy-SIRT1 axis in the regulation of cellular senescence.
  29. Stress Biol. 2023 Mar 23. 3(1): 3
    Coat protein of rice stripe virus enhances autophagy activity through interaction with cytosolic glyceraldehyde-3-phosphate dehydrogenases, a negative regulator of plant autophagy.
    Wanying Zhao, Li Wang, Lipeng Li, Tong Zhou, Fei Yan, Heng Zhang, Ying Zhu, Ida Bagus Andika, Liying Sun.
      Viral infection commonly induces autophagy, leading to antiviral responses or conversely, promoting viral infection or replication. In this study, using the experimental plant Nicotiana benthamiana, we demonstrated that the rice stripe virus (RSV) coat protein (CP) enhanced autophagic activity through interaction with cytosolic glyceraldehyde-3-phosphate dehydrogenase 2 (GAPC2), a negative regulator of plant autophagy that binds to an autophagy key factor, autophagy-related protein 3 (ATG3). Competitive pull-down and co-immunoprecipitation (Co-IP)assays showed that RSV CP activated autophagy by disrupting the interaction between GAPC2 and ATG3. An RSV CP mutant that was unable to bind GAPC2 failed to disrupt the interaction between GAPC2 and ATG3 and therefore lost its ability to induce autophagy. RSV CP enhanced the autophagic degradation of a viral movement protein (MP) encoded by a heterologous virus, citrus leaf blotch virus (CLBV). However, the autophagic degradation of RSV-encoded MP and RNA-silencing suppressor (NS3) proteins was inhibited in the presence of CP, suggesting that RSV CP can protect MP and NS3 against autophagic degradation. Moreover, in the presence of MP, RSV CP could induce the autophagic degradation of a remorin protein (NbREM1), which negatively regulates RSV infection through the inhibition of viral cell-to-cell movement. Overall, our results suggest that RSV CP induces a selective autophagy to suppress the antiviral factors while protecting RSV-encoded viral proteins against autophagic degradation through an as-yet-unknown mechanism. This study showed that RSV CP plays dual roles in the autophagy-related interaction between plants and viruses.
    Keywords:  ATG3; Autophagy; Coat protein; GAPC; RSV
    DOI:  https://doi.org/10.1007/s44154-023-00084-3
  30. J Biol Chem. 2023 Sep 01. pii: S0021-9258(23)02248-2. [Epub ahead of print] 105220
    Inhibition of mitochondrial fatty acid β-oxidation activates mTORC1 pathway and protein synthesis via Gcn5-dependent acetylation of raptor in zebrafish.
    Wen-Hao Zhou, Yuan Luo, Rui-Xin Li, Pascal Degrace, Tony Jourdan, Fang Qiao, Li-Qiao Chen, Mei-Ling Zhang, Zhen-Yu Du.
      Pharmacological inhibition of mitochondrial fatty acid oxidation (FAO) has been clinically used to alleviate certain metabolic diseases by remodeling cellular metabolism. However, mitochondrial FAO inhibition also leads to mTORC1 activation-related protein synthesis and tissue hypertrophy, but the mechanism remains unclear. Here, by using a mitochondrial FAO inhibitor (Mildronate or Etomoxir) or knocking out carnitine palmitoyltransferase-1, we revealed that mitochondrial FAO inhibition activated the mTORC1 pathway through Gcn5-dependent Raptor acetylation. Mitochondrial FAO inhibition significantly promoted glucose catabolism and increased intracellular acetyl-CoA levels. In response to the increased intracellular acetyl-CoA, acetyltransferase Gcn5 activated mTORC1 by catalyzing Raptor acetylation through direct interaction. Further investigation also screened Raptor deacetylases HDAC class II and identified HDAC7 as a potential regulator of Raptor. These results provide a possible mechanistic explanation for the mTORC1 activation after mitochondrial FAO inhibition and also bring light to reveal the roles of nutrient metabolic remodeling in regulating protein acetylation by affecting acetyl-CoA production.
    Keywords:  Gcn5; Raptor; acetyl-CoA; mTORC1; mitochondrial FAO inhibition
    DOI:  https://doi.org/10.1016/j.jbc.2023.105220
  31. Nat Commun. 2023 Sep 08. 14(1): 5524
    CD44 connects autophagy decline and ageing in the vascular endothelium.
    Lu Zhang, Peichang Yang, Jingxuan Chen, Zhiqiang Chen, Zhihui Liu, Gaoqing Feng, Fangfang Sha, Zirui Li, Zaoyi Xu, Yating Huang, Xiaotong Shi, Xuebiao Li, Jiatian Cui, Chenyi Zhang, Pei Fan, Liuqing Cui, Yunpeng Shen, Guangzhou Zhou, Hongjuan Jing, Shiwei Ma.
      The decline of endothelial autophagy is closely related to vascular senescence and disease, although the molecular mechanisms connecting these outcomes in vascular endothelial cells (VECs) remain unclear. Here, we identify a crucial role for CD44, a multifunctional adhesion molecule, in controlling autophagy and ageing in VECs. The CD44 intercellular domain (CD44ICD) negatively regulates autophagy by reducing PIK3R4 and PIK3C3 levels and disrupting STAT3-dependent PtdIns3K complexes. CD44 and its homologue clec-31 are increased in ageing vascular endothelium and Caenorhabditis elegans, respectively, suggesting that an age-dependent increase in CD44 induces autophagy decline and ageing phenotypes. Accordingly, CD44 knockdown ameliorates age-associated phenotypes in VECs. The endothelium-specific CD44ICD knock-in mouse is shorter-lived, with VECs exhibiting obvious premature ageing characteristics associated with decreased basal autophagy. Autophagy activation suppresses the premature ageing of human and mouse VECs overexpressing CD44ICD, function conserved in the CD44 homologue clec-31 in C. elegans. Our work describes a mechanism coordinated by CD44 function bridging autophagy decline and ageing.
    DOI:  https://doi.org/10.1038/s41467-023-41346-y
  32. Cell Death Differ. 2023 Sep 08.
    RUNDC1 inhibits autolysosome formation and survival of zebrafish via clasping ATG14-STX17-SNAP29 complex.
    Rui Zhang, Yuyan Yang, Chao He, Xin Zhang, Vincenzo Torraca, Shen Wang, Nan Liu, Jiaren Yang, Shicheng Liu, Jinglei Yuan, Dongzhi Gou, Shi Li, Xueying Dong, Yufei Xie, Junling He, Hua Bai, Mengyu Hu, Zhiquan Liao, Yuan Huang, Hao Lyu, Shuai Xiao, Dong Guo, Declan William Ali, Marek Michalak, Cong Ma, Xing-Zhen Chen, Jingfeng Tang, Cefan Zhou.
      Autophagy serves as a pro-survival mechanism for a cell or a whole organism to cope with nutrient stress. Our understanding of the molecular regulation of this fusion event remains incomplete. Here, we identified RUNDC1 as a novel ATG14-interacting protein, which is highly conserved across vertebrates, including zebrafish and humans. By gain and loss of function studies, we demonstrate that RUNDC1 negatively modulates autophagy by blocking fusion between autophagosomes and lysosomes via inhibiting the assembly of the STX17-SNAP29-VAMP8 complex both in human cells and the zebrafish model. Moreover, RUNDC1 clasps the ATG14-STX17-SNAP29 complex via stimulating ATG14 homo-oligomerization to inhibit ATG14 dissociation. This also prevents VAMP8 from binding to STX17-SNAP29. We further identified that phosphorylation of RUNDC1 Ser379 is crucial to inhibit the assembly of the STX17-SNAP29-VAMP8 complex via promoting ATG14 homo-oligomerization. In line with our findings, RunDC1 is crucial for zebrafish in their response to nutrient-deficient conditions. Taken together, our findings demonstrate that RUNDC1 is a negative regulator of autophagy that restricts autophagosome fusion with lysosomes by clasping the ATG14-STX17-SNAP29 complex to hinder VAMP8 binding.
    DOI:  https://doi.org/10.1038/s41418-023-01215-z
  33. Proc Natl Acad Sci U S A. 2023 Sep 12. 120(37): e2221929120
    A selective ER-phagy exerts neuroprotective effects via modulation of α-synuclein clearance in parkinsonian models.
    Dong Yeol Kim, Jin Young Shin, Ji Eun Lee, Ha Na Kim, Seok Jong Chung, Han Soo Yoo, Sang Jin Kim, Hwa Jin Cho, Eun-Jae Lee, Soo Jeong Nam, Seong Heon Kim, Jaewon Jang, Seung Eun Lee, Phil Hyu Lee.
      The endoplasmic reticulum (ER) is selectively degraded by ER-phagy to maintain cell homeostasis. α-synuclein accumulates in the ER, causing ER stress that contributes to neurodegeneration in Parkinson's disease (PD), but the role of ER-phagy in α-synuclein modulation is largely unknown. Here, we investigated the mechanisms by which ER-phagy selectively recognizes α-synuclein for degradation in the ER. We found that ER-phagy played an important role in the degradation of α-synuclein and recovery of ER function through interaction with FAM134B, where calnexin is required for the selective FAM134B-mediated α-synuclein clearance via ER-phagy. Overexpression of α-synuclein in the ER of the substantia nigra (SN) resulted in marked loss of dopaminergic neurons and motor deficits, mimicking PD characteristics. However, enhancement of ER-phagy using FAM134B overexpression in the SN exerted neuroprotective effects on dopaminergic neurons and recovered motor performance. These data suggest that ER-phagy represents a specific ER clearance mechanism for the degradation of α-synuclein.
    Keywords:  ER-phagy; FAM134B; Parkinson’s disease; protection; α-synuclein
    DOI:  https://doi.org/10.1073/pnas.2221929120
  34. Biosci Biotechnol Biochem. 2023 Sep 08. pii: zbad130. [Epub ahead of print]
    Kangfuxin liquid reduces the ultraviolet B-induced photodamage of HaCaT cells by regulating autophagy.
    Xianghong Lin, Jimin Chu, Yang Xiang, Miao He, Qiong Ma, Jingxian Duan, Yan Wang, Sujiao Sun.
      Kangfuxin liquid (KFX), an extract of the American cockroach, has been clinically proven to be effective in various skin damage disorders, but there are no reports on its use in photodamage. We explored the effect of KFX on ultraviolet B (UVB) induced photodamage and whether its mechanism was related to autophagy. We found that KFX treatment reduced UVB induced reactive oxygen species (ROS) production and improved the vitality of cells inhibited by UVB irradiation. The expression of LC3 (A/B), which was inhibited after UVB irradiation, could be rescued by KFX treatment. Furthermore, KFX may upregulate the level of cellular autophagy by regulating the AMPK-mTOR signaling pathway. When the autophagy inhibitor wortmannin was used to inhibit autophagy, the protective effect of KFX on cells was diminished or even disappeared. Our study suggests that KFX may resist UVB-mediated oxidative stress damage of HaCaT through the induction of autophagy.
    Keywords:  HaCaT; autophagy; kangfuxin liquid; photodamage; ultraviolet B
    DOI:  https://doi.org/10.1093/bbb/zbad130
  35. Toxicology. 2023 Sep 05. pii: S0300-483X(23)00214-7. [Epub ahead of print] 153627
    ROS-mediated mitophagy and necroptosis regulate osteocytes death caused by TCP particles in MLO-Y4 cells.
    Pei Yang, Bingbing Xu, Ruirong Zhu, Tao Zhang, Zihan Wang, Qiao Lin, Ming Yan, Zhangsen Yu, Hongjiao Mao, Yun Zhang.
      Our previous data have revealed TCP particles caused cell death of osteocytes, comprising over 95% of all bone cells, which contribute to periprosthetic osteolysis, joint loosening and implant failure, but its mechanisms are not fully understood. Here, we reported that TCP particles inhibited cell viability of osteocytes MLO-Y4, and caused cell death. TCP particles caused mitochondrial impairment and increased expressions of LC-3 II, Parkin and PINK 1, accompanied by the elevation of autophagy flux and intracellular acidic components, the accumulation of LC-3II, PINK1 and Parkin in damaged mitochondria, and p62 reduction. The increased LC-3II expression and cell death extent were significantly enhanced by the autophagy inhibitor Baf A1, compared with Baf A1 (or TCP particles) alone, indicating that TCP particles increase autophagic flux and lead to cell even death of MLO-Y4 cells, closely associated with mitophagy. Furthermore, TCP particles induced propidium iodide (PI) uptake and the phosphorylation of RIP1, RIP3 and MLKL, thereby increasing necroptosis in MLO-Y4 cells. The pro-necroptotic effect was alleviated by the RIP1 inhibitor Nec-1 or the MLKL inhibitor NSA. Additionally, TCP particles promoted the production of intracellular reactive oxygen species (ROS) and mitochondrial ROS (mtROS), and increased TXNIP expression, but decreased protein levels of TRX1, Nrf2, HO-1 and NQO1, leading to oxidative stress. The ROS scavenger NAC remarkably reversed mitophagy and necroptosis caused by TCP particles, suggesting that ROS is responsible for mitophagy and necroptosis. Collectively, ROS-mediated mitophagy and necroptosis regulate osteocytes death caused by TCP particles in MLO-Y4 cells, which enhances osteoclastogenesis and periprosthetic osteolysis.
    Keywords:  TCP particles; mitophagy; necroptosis; osteocytes; reactive oxygen species
    DOI:  https://doi.org/10.1016/j.tox.2023.153627
  36. J Agric Food Chem. 2023 Sep 06.
    Kuwanon H Inhibits Melanoma Growth through Cytotoxic Endoplasmic Reticulum Stress and Impaired Autophagy Flux.
    Xin Hu, Guangzhao Pan, Jili Luo, Xinyue Gao, Yuhang Mu, Zhi Wang, Xiaosong Hu, Chongyang Li, Muhammad Nadeem Abbas, Kui Zhang, Ying Zheng, Hongjuan Cui.
      Although great progress has been made recently in targeted and immune-based therapies, additional treatments are needed for most melanoma patients due to acquired chemoresistance, recurrence, or metastasis. Elevated autophagy is required for the pathogenesis of melanoma to attenuate metabolic stress, protecting cancer cells from chemotherapeutics or radiation. Thus, intervention with autophagy is a promising strategy for melanoma treatment. Here, we examined a novel antimelanoma natural compound named kuwanon H (KuH), which significantly inhibited melanoma cell growth in vitro/vivo. Mechanistically, KuH induced cytotoxic endoplasmic reticulum (ER) stress, which inhibited cell viability and induced apoptosis. Meanwhile, KuH-induced ER stress mediated autophagysome formation through the ATF4-DDIT3-TRIB3-AKT-MTOR axis. Importantly, KuH impaired autophagy flux, which contributed to the anticancer effects of KuH. Finally, our results showed that KuH enhanced the sensitivity of melanoma cells to cisplatin, both in vitro and in vivo, by impairing autophagy degradation of reactive oxygen species and damaged mitochondria. Our findings indicate that KuH is a promising candidate anticancer natural product for melanoma therapy.
    Keywords:  ER stress; anticancer natural product; autophagy; cisplatin; kuwanon H; melanoma
    DOI:  https://doi.org/10.1021/acs.jafc.3c02257
  37. Signal Transduct Target Ther. 2023 Sep 06. 8(1): 333
    Mitochondrial dynamics in health and disease: mechanisms and potential targets.
    Wen Chen, Huakan Zhao, Yongsheng Li.
      Mitochondria are organelles that are able to adjust and respond to different stressors and metabolic needs within a cell, showcasing their plasticity and dynamic nature. These abilities allow them to effectively coordinate various cellular functions. Mitochondrial dynamics refers to the changing process of fission, fusion, mitophagy and transport, which is crucial for optimal function in signal transduction and metabolism. An imbalance in mitochondrial dynamics can disrupt mitochondrial function, leading to abnormal cellular fate, and a range of diseases, including neurodegenerative disorders, metabolic diseases, cardiovascular diseases and cancers. Herein, we review the mechanism of mitochondrial dynamics, and its impacts on cellular function. We also delve into the changes that occur in mitochondrial dynamics during health and disease, and offer novel perspectives on how to target the modulation of mitochondrial dynamics.
    DOI:  https://doi.org/10.1038/s41392-023-01547-9
  38. Int J Mol Sci. 2023 Aug 22. pii: 13040. [Epub ahead of print]24(17):
    The Combination of Gold and Silver Food Nanoparticles with Gluten Peptides Alters the Autophagic Pathway in Intestinal Crypt-like Cells.
    Clara Mancuso, Eric Tremblay, Elisa Gnodi, Steve Jean, Jean-François Beaulieu, Donatella Barisani.
      Metallic nanoparticles (mNPs) are widely used as food additives and can interact with gliadin triggering an immune response, but evaluation of the effects on crypts, hypertrophic in celiac subjects, is still lacking. This study evaluated the effects of gold and silver mNPs in combination with gliadin on crypt-like cells (HIEC-6). Transmission electron microscopy (TEM) was used to evaluate gliadin-mNP aggregates in cells. Western blot and immunofluorescence analysis assessed autophagy-related molecule levels (p62, LC3, beclin-1, EGFR). Lysosome functionality was tested with acridine orange (AO) and Magic Red assays. TEM identified an increase in autophagic vacuoles after exposure to gliadin + mNPs, as also detected by significant increments in LC3-II and p62 expression. Immunofluorescence confirmed the presence of mature autophagosomes, showing LC3 and p62 colocalization, indicating an altered autophagic flux, further assessed with EGFR degradation, AO and Magic Red assays. The results showed a significant reduction in lysosomal enzyme activity and a modest reduction in acidity. Thus, gliadin + mNPs can block the autophagic flux inducing a lysosomal defect. The alteration of this pathway, essential for cell function, can lead to cell damage and death. The potential effects of this copresence in food should be further characterized to avoid a negative impact on celiac disease subjects.
    Keywords:  autophagy; celiac disease; dietary metallic nanoparticles; food additives; gluten peptides; lysosome dysfunction
    DOI:  https://doi.org/10.3390/ijms241713040
  39. Cell Death Discov. 2023 Sep 04. 9(1): 332
    The role and implication of autophagy in cholangiocarcinoma.
    Hayat Khizar, Yufei Hu, Yanhua Wu, Jianfeng Yang.
      Cholangiocarcinoma (CCA) is a malignant tumor that originates from the biliary epithelial cells. It is characterized by a difficult diagnosis and limited treatment options. Autophagy is a cellular survival mechanism that maintains nutrient and energy homeostasis and eliminates intracellular pathogens. It is involved in various physiological and pathological processes, including the development of cancer. However, the role, mechanism, and potential therapeutic targets of autophagy in CCA have not been thoroughly studied. In this review, we introduce the classification, characteristics, process, and related regulatory genes of autophagy. We summarize the regulation of autophagy on the progression of CCA and collect the latest research progress on some autophagy modulators with clinical potential in CCA. In conclusion, combining autophagy modulators with immunotherapy, chemotherapy, and targeted therapy has great potential in the treatment of CCA. This combination may be a potential therapeutic target for CCA in the future.
    DOI:  https://doi.org/10.1038/s41420-023-01631-7
  40. Autophagy. 2023 Sep 07. 1-2
    ATG3 proteins possess a unique amphipathic α-helix essential for the Atg8/LC3 lipidation reaction.
    Taki Nishimura, Gianmarco Lazzeri, Sharon A Tooze, Roberto Covino.
      In our recent paper, we uncovered that ATG3 exhibits a large degree of structural dynamics on autophagic membranes to efficiently carry out LC3 lipidation. ATG3 proteins possess an amphipathic α-helix (AH) identified by a small number of bulky and hydrophobic residues. This biophysical fingerprint allows for transient membrane association of ATG3 and facilitates its enzymatic reaction. This study will pave the way for a structural and mechanistic understanding of how membrane association of ATG proteins is orchestrated during autophagosome formation.
    Keywords:  ATG3; All-atom MD simulation; LC3 lipidation; amphipathic α-helix; membrane protein dynamics
    DOI:  https://doi.org/10.1080/15548627.2023.2255458
  41. Macromol Biosci. 2023 Sep 07. e2300116
    Fighting Against Drug-resistant Tumor by The Induction of Excessive Mitophagy with Transferrin Nanomedicine.
    Yuanxiang Yu, Sijin Chen, Yupeng Wang, Dongfang Zhou, Dehua Wu.
      The effectiveness of chemotherapy is primarily hindered by drug resistance, and autophagy plays a crucial role in overcoming this resistance. In this project, we have developed a human transferrin nanomedicine known as HTf@DOX/Qu NPs, which contains quercetin (a drug to induces excessive autophagy) and doxorubicin. The purpose of this nanomedicine is to enhance mitophagy and combating drug-resistant cancer. Through in vitro studies, we have demonstrated that HTf@DOX/Qu NPs can effectively downregulate cyclooxygenase-2 (COX-2), leading to an excessive promotion of mitophagy and subsequent mitochondrial dysfunction via the PINK1/Parkin axis. Additionally, HTf@DOX/Qu NPs can upregulate proapoptotic proteins to induce cellular apoptosis, thereby effectively reversing drug resistance. Furthermore, in vivo results have shown that HTf@DOX/Qu NPs exhibit prolonged circulation in the bloodstream, enhanced drug accumulation in tumors, and superior therapeutic efficacy compared to individual chemotherapy in a drug-resistant tumor model. This study presents a promising strategy for combating multidrug-resistant cancers by exacerbating mitophagy through the use of transferrin nanoparticles. This article is protected by copyright. All rights reserved.
    Keywords:  cancer therapy; excessive mitophagy; multidrug resistance; quercetin; transferrin nanoparticles
    DOI:  https://doi.org/10.1002/mabi.202300116
  42. Front Immunol. 2023 ;14 1212695
    Crosstalk between autophagy and metabolic regulation of (CAR) T cells: therapeutic implications.
    Ahmad Reza Panahi Meymandi, Behnia Akbari, Tahereh Soltantoyeh, Jamshid Hadjati, Daniel J Klionsky, Behnam Badie, Hamid Reza Mirzaei.
      Despite chimeric antigen receptor (CAR) T cell therapy's extraordinary success in subsets of B-cell lymphoma and leukemia, various barriers restrict its application in solid tumors. This has prompted investigating new approaches for producing CAR T cells with superior therapeutic potential. Emerging insights into the barriers to CAR T cell clinical success indicate that autophagy shapes the immune response via reprogramming cellular metabolism and vice versa. Autophagy, a self-cannibalization process that includes destroying and recycling intracellular components in the lysosome, influences T cell biology, including development, survival, memory formation, and cellular metabolism. In this review, we will emphasize the critical role of autophagy in regulating and rewiring metabolic circuits in CAR T cells, as well as how the metabolic status of CAR T cells and the tumor microenvironment (TME) alter autophagy regulation in CAR T cells to restore functional competence in CAR Ts traversing solid TMEs.
    Keywords:  CAR T cell; adoptive cellular therapy (ACT); autophagy; metabolism; tumor microenvironment
    DOI:  https://doi.org/10.3389/fimmu.2023.1212695
  43. Chem Biol Interact. 2023 Aug 31. pii: S0009-2797(23)00362-9. [Epub ahead of print] 110695
    3-Acetyldeoxynivalenol induces apoptosis, barrier dysfunction and endoplasmic reticulum stress by inhibiting mTORC1-dependent autophagy in porcine enterocytes.
    Tongkun Zhang, Jun Bai, Guangye Chen, Zhaohui Chen, Shenming Zeng, Ying Yang, Zhenlong Wu.
      3-Acetyldeoxynivalenol (3-Ac-DON), the acetylated form of deoxynivalenol, which is widely present in mycotoxin-contaminated food, feed as well as in other natural sources. Ingestion of 3-Ac-DON may result in intestinal dysfunction, leading to gut diseases in humans and animals. Nevertheless, the molecular mechanism of 3-Ac-DON in intestinal epithelial cytotoxicity remains unclear. In this study, intestinal porcine epithelial cell line 1 (IPEC-1) cells were treated with different concentrations of 3-Ac-DON for 12 h and 24 h, respectively. The results showed that 3-Ac-DON caused cell viability decrease, cell cycle arrest in G1 phaseand depolarization of mitochondrial membrane potential. Also, Western blotting analysis showed that 3-Ac-DON significantly decreased the expression of tight junction proteins, inhibited autophagy and activated endoplasmic reticulum (ER) stress in IPEC cells (P < 0.05). Further investigation demonstrated that 3-Ac-DON caused apoptosis, ER stress and barrier dysfunction were reversed after pretreatment with the autophagy activator rapamycin (100 nM), indicating that autophagy plays a key role in the process of 3-Ac-DON-induced cell damage. In addition, we demonstrated that 3-Ac-DON inhibits the occurrence of autophagy mediated by mTORC1 protein. In conclusion, our research indicated that the mTORC1 protein and autophagy played a key role in the 3-Ac-DON-induced cytotoxic in IPEC cells, which will provide new therapeutic targets and ideas for 3-AC-DON-mediated intestinal injury.
    Keywords:  3-Acetyldeoxynivalenol; Apoptosis; Autophagy; Barrier function; Endoplasmic reticulum stress; Mechanistic target of rapamycin complex 1
    DOI:  https://doi.org/10.1016/j.cbi.2023.110695
  44. Diabetes Metab J. 2023 Sep 06.
    Glucolipotoxicity Suppressed Autophagy and Insulin Contents in Human Islets, and Attenuation of PERK Activity Enhanced Them in an ATG7-Dependent Manner.
    Seoil Moon, Ji Yoon Lim, Mirang Lee, Youngmin Han, Hongbeom Kim, Wooil Kwon, Jin-Young Jang, Mi Na Kim, Kyong Soo Park, Hye Seung Jung.
       Background: Administration of pancreatic endoplasmic reticulum kinase inhibitor (PERKi) improved insulin secretion and hyperglycemia in obese diabetic mice. In this study, autophagic balance was studied whether to mediate it.
    Methods: Human islets were isolated from living patients without diabetes. PERKi GSK2606414 effects were evaluated in the islets under glucolipotoxicity by palmitate. Islet insulin contents and secretion were measured. Autophagic flux was assessed by microtubule associated protein 1 light chain 3 (LC3) conversion, a red fluorescent protein (RFP)-green fluorescent protein (GFP)- LC3 tandem assay, and P62 levels. For mechanical analyses, autophagy was suppressed using 3-methyladenine in mouse islets. Small interfering RNA for an autophagy-related gene autophagy related 7 (Atg7) was transfected to interfere autophagy.
    Results: PERKi administration to mice decreased diabetes-induced P62 levels in the islets. Glucolipotoxicity significantly increased PERK phosphorylation by 70% and decreased insulin contents by 50% in human islets, and addition of PERKi (40 to 80 nM) recovered both. PERKi also enhanced glucose-stimulated insulin secretion (6-fold). PERKi up-regulated LC3 conversion suppressed by glucolipotoxicity, and down-regulated P62 contents without changes in P62 transcription, indicating enhanced autophagic flux. Increased autophagosome-lysosome fusion by PERKi was visualized in mouse islets, where PERKi enhanced ATG7 bound to LC3. Suppression of Atg7 eliminated PERKi-induced insulin contents and secretion. Conclusion: This study provided functional changes of human islets with regard to autophagy under glucolipotoxicity, and suggested modulation of autophagy as an anti-diabetic mechanism of PERKi.
    Keywords:  Autophagy; Diabetes mellitus; Humans; Insulin; Islets of Langerhans
    DOI:  https://doi.org/10.4093/dmj.2022.0366
  45. Int J Mol Sci. 2023 Aug 30. pii: 13436. [Epub ahead of print]24(17):
    ERK1/2-Dependent Phosphorylation of GABAB1(S867/T872), Controlled by CaMKIIβ, Is Required for GABAB Receptor Degradation under Physiological and Pathological Conditions.
    Musadiq A Bhat, Thomas Grampp, Dietmar Benke.
      GABAB receptor-mediated inhibition is indispensable for maintaining a healthy neuronal excitation/inhibition balance. Many neurological diseases are associated with a disturbed excitation/inhibition balance and downregulation of GABAB receptors due to enhanced sorting of the receptors to lysosomal degradation. A key event triggering the downregulation of the receptors is the phosphorylation of S867 in the GABAB1 subunit mediated by CaMKIIβ. Interestingly, close to S867 in GABAB1 exists another phosphorylation site, T872. Therefore, the question arose as to whether phosphorylation of T872 is involved in downregulating the receptors and whether phosphorylation of this site is also mediated by CaMKIIβ or by another protein kinase. Here, we show that mutational inactivation of T872 in GABAB1 prevented the degradation of the receptors in cultured neurons. We found that, in addition to CaMKIIβ, also ERK1/2 is involved in the degradation pathway of GABAB receptors under physiological and ischemic conditions. In contrast to our previous view, CaMKIIβ does not appear to directly phosphorylate S867. Instead, the data support a mechanism in which CaMKIIβ activates ERK1/2, which then phosphorylates S867 and T872 in GABAB1. Blocking ERK activity after subjecting neurons to ischemic stress completely restored downregulated GABAB receptor expression to normal levels. Thus, preventing ERK1/2-mediated phosphorylation of S867/T872 in GABAB1 is an opportunity to inhibit the pathological downregulation of the receptors after ischemic stress and is expected to restore a healthy neuronal excitation/inhibition balance.
    Keywords:  CaMKII; ERK1/2; GABAB receptors; cerebral ischemia; degradation; phosphorylation
    DOI:  https://doi.org/10.3390/ijms241713436
  46. Mol Psychiatry. 2023 Sep 07.
    The role of glial autophagy in Alzheimer's disease.
    Anna Litwiniuk, Grzegorz Roman Juszczak, Adrian Mateusz Stankiewicz, Kaja Urbańska.
      Although Alzheimer's disease is the most pervasive neurodegenerative disorder, the mechanism underlying its development is still not precisely understood. Available data indicate that pathophysiology of this disease may involve impaired autophagy in glial cells. The dysfunction is manifested as reduced ability of astrocytes and microglia to clear abnormal protein aggregates. Consequently, excessive accumulation of amyloid beta plaques and neurofibrillary tangles activates microglia and astrocytes leading to decreased number of mature myelinated oligodendrocytes and death of neurons. These pathologic effects of autophagy dysfunction can be rescued by pharmacological activation of autophagy. Therefore, a deeper understanding of the molecular mechanisms involved in autophagy dysfunction in glial cells in Alzheimer's disease may lead to the development of new therapeutic strategies. However, such strategies need to take into consideration differences in regulation of autophagy in different types of neuroglia.
    DOI:  https://doi.org/10.1038/s41380-023-02242-5
  47. Int J Mol Sci. 2023 Aug 30. pii: 13433. [Epub ahead of print]24(17):
    MAGED2 Depletion Promotes Stress-Induced Autophagy by Impairing the cAMP/PKA Pathway.
    Sadiq Nasrah, Aline Radi, Johanna K Daberkow, Helmut Hummler, Stefanie Weber, Elie Seaayfan, Martin Kömhoff.
      Melanoma-associated antigen D2 (MAGED2) plays an essential role in activating the cAMP/PKA pathway under hypoxic conditions, which is crucial for stimulating renal salt reabsorption and thus explaining the transient variant of Bartter's syndrome. The cAMP/PKA pathway is also known to regulate autophagy, a lysosomal degradation process induced by cellular stress. Previous studies showed that two members of the melanoma-associated antigens MAGE-family inhibit autophagy. To explore the potential role of MAGED2 in stress-induced autophagy, specific MAGED2-siRNA were used in HEK293 cells under physical hypoxia and oxidative stress (cobalt chloride, hypoxia mimetic). Depletion of MAGED2 resulted in reduced p62 levels and upregulation of both the autophagy-related genes (ATG5 and ATG12) as well as the autophagosome marker LC3II compared to control siRNA. The increase in the autophagy markers in MAGED2-depleted cells was further confirmed by leupeptin-based assay which concurred with the highest LC3II accumulation. Likewise, under hypoxia, immunofluorescence in HEK293, HeLa and U2OS cell lines demonstrated a pronounced accumulation of LC3B puncta upon MAGED2 depletion. Moreover, LC3B puncta were absent in human fetal control kidneys but markedly expressed in a fetal kidney from a MAGED2-deficient subject. Induction of autophagy with both physical hypoxia and oxidative stress suggests a potentially general role of MAGED2 under stress conditions. Various other cellular stressors (brefeldin A, tunicamycin, 2-deoxy-D-glucose, and camptothecin) were analyzed, which all induced autophagy in the absence of MAGED2. Forskolin (FSK) inhibited, whereas GNAS Knockdown induced autophagy under hypoxia. In contrast to other MAGE proteins, MAGED2 has an inhibitory role on autophagy only under stress conditions. Hence, a prominent role of MAGED2 in the regulation of autophagy under stress conditions is evident, which may also contribute to impaired fetal renal salt reabsorption by promoting autophagy of salt-transporters in patients with MAGED2 mutation.
    Keywords:  Bartter’s syndrome; G-alpha-S; MAGED2; autophagy; cAMP; cellular stress; hypoxia nephrogenesis
    DOI:  https://doi.org/10.3390/ijms241713433
  48. Mol Cell. 2023 Sep 07. pii: S1097-2765(23)00641-X. [Epub ahead of print]83(17): 3188-3204.e7
    Damaged mitochondria recruit the effector NEMO to activate NF-κB signaling.
    Olivia Harding, Elisabeth Holzer, Julia F Riley, Sascha Martens, Erika L F Holzbaur.
      Failure to clear damaged mitochondria via mitophagy disrupts physiological function and may initiate damage signaling via inflammatory cascades, although how these pathways intersect remains unclear. We discovered that nuclear factor kappa B (NF-κB) essential regulator NF-κB effector molecule (NEMO) is recruited to damaged mitochondria in a Parkin-dependent manner in a time course similar to recruitment of the structurally related mitophagy adaptor, optineurin (OPTN). Upon recruitment, NEMO partitions into phase-separated condensates distinct from OPTN but colocalizing with p62/SQSTM1. NEMO recruitment, in turn, recruits the active catalytic inhibitor of kappa B kinase (IKK) component phospho-IKKβ, initiating NF-κB signaling and the upregulation of inflammatory cytokines. Consistent with a potential neuroinflammatory role, NEMO is recruited to mitochondria in primary astrocytes upon oxidative stress. These findings suggest that damaged, ubiquitinated mitochondria serve as an intracellular platform to initiate innate immune signaling, promoting the formation of activated IKK complexes sufficient to activate NF-κB signaling. We propose that mitophagy and NF-κB signaling are initiated as parallel pathways in response to mitochondrial stress.
    Keywords:  ALS; NEMO; NF-κB; NF-κB effector molecule; Parkin; Parkinson’s disease; SQSTM1/p62; amyotrophic lateral sclerosis; cell stress; innate immunity; mitophagy; neurodegeneration; neuroinflammation; optineurin nuclear factor kappa B; phase separation; ubiquitin
    DOI:  https://doi.org/10.1016/j.molcel.2023.08.005
  49. Cell Rep. 2023 Sep 01. pii: S2211-1247(23)01070-7. [Epub ahead of print]42(9): 113059
    The N-end rule pathway regulates ER stress-induced clusterin release to the cytosol where it directs misfolded proteins for degradation.
    Sandeep Satapathy, Holly Walker, James Brown, Yann Gambin, Mark R Wilson.
      Previous work suggests that cell stress induces release of the normally secreted chaperone clusterin (CLU) into the cytosol. We analyzed the localization of CLU in healthy and stressed cells, the mechanism of its cytosolic release, and its interactions with cytosolic misfolded proteins. Key results of this study are the following: (1) full-length CLU is released to the cytosol during stress, (2) the CLU N-terminal D1 residue is recognized by the N-end rule pathway and together with the enzyme ATE1 is essential for cytosolic release, (3) CLU can form stable complexes with cytosolic misfolded proteins and direct them to the proteasome and autophagosomes, and (4) cytosolic CLU protects cells from hypoxic stress and the cytosolic overexpression of an aggregation-prone protein. Collectively, the results suggest that enhanced cytosolic release of CLU is a stress response that can inhibit the toxicity of misfolded proteins and facilitate their targeted degradation via both autophagy and the proteasome.
    Keywords:  CP: Cell biology; N-End Rule Pathway; autophagy; clusterin; cytoprotection; cytosolic release; intracellular proteostasis; proteasome
    DOI:  https://doi.org/10.1016/j.celrep.2023.113059
  50. Environ Res. 2023 Sep 03. pii: S0013-9351(23)01810-8. [Epub ahead of print] 117006
    Autophagy/ferroptosis in colorectal cancer: Carcinogenic view and nanoparticle-mediated cell death regulation.
    Zhibin Zhang, Yintao Zhao, Yuman Wang, Yutang Zhao, Jianen Guo.
      The cell death mechanisms have a long history of being evaluated in diseases and pathological events. The ability of triggering cell death is considered to be a promising strategy in cancer therapy, but some mechanisms have dual functions in cancer, requiring more elucidation of underlying factors. Colorectal cancer (CRC) is a disease and malignant condition of colon and rectal that causes high mortality and morbidity. The autophagy targeting in CRC is therapeutic importance and this cell death mechanism can interact with apoptosis in inhibiting or increasing apoptosis. Autophagy has interaction with ferroptosis as another cell death pathway in CRC and can accelerate ferroptosis in suppressing growth and invasion. The dysregulation of autophagy affects the drug resistance in CRC and pro-survival autophagy can induce drug resistance. Therefore, inhibition of protective autophagy enhances chemosensitivity in CRC cells. Moreover, autophagy displays interaction with metastasis and EMT as a potent regulator of invasion in CRC cells. The same is true for ferroptosis, but the difference is that function of ferroptosis is determined and it can reduce viability. The lack of ferroptosis can cause development of chemoresistance in CRC cells and this cell death mechanism is regulated by various pathways and mechanisms that autophagy is among them. Therefore, current review paper provides a state-of-art analysis of autophagy, ferroptosis and their crosstalk in CRC. The nanoparticle-mediated regulation of cell death mechanisms in CRC causes changes in progression. The stimulation of ferroptosis and control of autophagy (induction or inhibition) by nanoparticles can impair CRC progression. The engineering part of nanoparticle synthesis to control autophagy and ferroptosis in CRC still requires more attention.
    Keywords:  Cancer therapy; Colorectal cancer; Ferroptosis; Nanoparticles; autophagy
    DOI:  https://doi.org/10.1016/j.envres.2023.117006
  51. J Neuroimmune Pharmacol. 2023 Sep 08.
    PPARɑ Ligand Caudatin Improves Cognitive Functions and Mitigates Alzheimer's Disease Defects By Inducing Autophagy in Mice Models.
    Senthilkumar Krishnamoorthi, Ashok Iyaswamy, Sravan Gopalkrishnashetty Sreenivasmurthy, Abhimanyu Thakur, Karthick Vasudevan, Gaurav Kumar, Xin-Jie Guan, Kejia Lu, Isha Gaurav, Cheng-Fu Su, Zhou Zhu, Jia Liu, Yuxuan Kan, Selvaraj Jayaraman, Zhiqiang Deng, Ka Kit Chua, King-Ho Cheung, Zhijun Yang, Ju-Xian Song, Min Li.
      The autophagy-lysosomal pathway (ALP) is a major cellular machinery involved in the clearance of aggregated proteins in Alzheimer disease (AD). However, ALP is dramatically impaired during AD pathogenesis via accumulation of toxic amyloid beta (Aβ) and phosphorylated-Tau (phospho-Tau) proteins in the brain. Therefore, activation of ALP may prevent the increased production of Aβ and phospho-Tau in AD. Peroxisome proliferator-activated receptor alpha (PPARα), a transcription factor that can activate autophagy, and transcriptionally regulate transcription factor EB (TFEB) which is a key regulator of ALP. This suggests that targeting PPARα, to reduce ALP impairment, could be a viable strategy for AD therapy. In this study, we investigated the anti-AD activity of Caudatin, an active constituent of Cynanchum otophyllum (a traditional Chinese medicinal herb, Qing Yang Shen; QYS). We found that Caudatin can bind to PPARα as a ligand and augment the expression of ALP in microglial cells and in the brain of 3XTg-AD mice model. Moreover, Caudatin could activate PPARα and transcriptionally regulates TFEB-augmented lysosomal degradation of Aβ and phosphor-Tau aggregates in AD cell models. Oral administration of Caudatin decreased AD pathogenesis and ameliorated the cognitive dysfunction in 3XTg-AD mouse model. Conclusively, Caudatin can be a potential AD therapeutic agent via activation of PPARα-dependent ALP.
    Keywords:  Alzheimer’s disease (AD); Amyloid beta (Aβ); Autophagy-lysosomal pathway (ALP); Caudatin; Peroxisome proliferator-activated receptor alpha (PPARα); Phospho-Tau
    DOI:  https://doi.org/10.1007/s11481-023-10083-w
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