bims-auttor Biomed News
on Autophagy and mTOR
Issue of 2022‒03‒20
forty-four papers selected by
Viktor Korolchuk, Newcastle University
  1. Follicular lymphoma-associated mutations in the V-ATPase chaperone VMA21 activate autophagy creating a targetable dependency.
  2. Loss of RUBCN/rubicon in adipocytes mediates the upregulation of autophagy to promote the fasting response.
  3. ROOT HAIR DEFECTIVE3 Is a Receptor for Selective Autophagy of the Endoplasmic Reticulum in Arabidopsis.
  4. Rheb regulates nuclear mTORC1 activity independent of farnesylation.
  5. Single serine on TSC2 exerts biased control over mTORC1 activation mediated by ERK1/2 but not Akt.
  6. LC3B is an RNA-binding protein to trigger rapid mRNA degradation during autophagy.
  7. Cell Type-Specific Mechanistic Target of Rapamycin-Dependent Distortion of Autophagy Pathways in Lupus Nephritis.
  8. Regulation of basal autophagy by calmodulin availability.
  9. Asymmetric Pericentrosomal CD133 Endosomes Induce the Unequal Autophagic Activity During Cytokinesis in CD133-Positive Human Neuroblastoma Cells.
  10. Lysosomal Ca2+-mediated TFEB activation modulates mitophagy and functional adaptation of pancreatic β-cells to metabolic stress.
  11. Mitochondrial depolarization after acute ethanol treatment drives mitophagy in living mice.
  12. Phosphatidic acid suppresses autophagy through competitive inhibition by binding GAPC (glyceraldehyde-3-phosphate dehydrogenase) and PGK (phosphoglycerate kinase) proteins.
  13. The needs of a synapse-How local organelles serve synaptic proteostasis.
  14. GFP-LC3 High-Content Assay for Screening Autophagy Modulators.
  15. MAP kinase-dependent autophagy controls phorbol myristate acetate-induced macrophage differentiation of HL-60 leukemia cells.
  16. Material properties of phase-separated TFEB condensates regulate the autophagy-lysosome pathway.
  17. eIF3a regulation of mTOR signaling and translational control via HuR in cellular response to DNA damage.
  18. Rapamycin Improved Retinal Function and Morphology in a Mouse Model of Retinal Degeneration.
  19. The unfolded protein response transducer IRE1α promotes reticulophagy in podocytes.
  20. Identification of defined structural elements within TOR2 kinase required for TOR Complex 2 assembly and function in S. cerevisiae.
  21. A trio of ubiquitin ligases sequentially drives ubiquitylation and autophagic degradation of dysfunctional yeast proteasomes.
  22. Mitochondrial Dynamics, Mitophagy, and Mitochondria-Endoplasmic Reticulum Contact Sites Crosstalk Under Hypoxia.
  23. Acute Increase of Renal Perfusion Pressure Causes Rapid Activation of mTORC1 (Mechanistic Target Of Rapamycin Complex 1) and Leukocyte Infiltration.
  24. Glutamate delta 1 receptor regulates autophagy mechanisms and affects excitatory synapse maturation in the somatosensory cortex.
  25. Recombinant pro-CTSD (cathepsin D) enhances SNCA/α-Synuclein degradation in α-Synucleinopathy models.
  26. Maternal exposure to CeO2NPs derails placental development through trophoblast dysfunction mediated by excessive autophagy activation.
  27. Autistic-like behavior and cerebellar dysfunction in Bmal1 mutant mice ameliorated by mTORC1 inhibition.
  28. F-box protein FBXO41 plays vital role in arsenic trioxide-mediated autophagic death of cancer cells.
  29. The Role of Zinc in Axon Formation via the mTORC1 Pathway.
  30. Muscle Wasting in COPD: Not Enough Autophagy?
  31. Enhancing autophagy in Alzheimer's disease through drug repositioning.
  32. Autophagy flux inhibition mediated by lysosomal dysfunction participates in the cadmium exposure-induced cardiotoxicity in swine.
  33. Targeting lipophagy in macrophages improves repair in multiple sclerosis.
  34. Autophagy inhibitors enhance biomolecular delivery efficiency of extracellular vesicles.
  35. A Novel Copper(II) Indenoisoquinoline Complex Inhibits Topoisomerase I, Induces G2 Phase Arrest, and Autophagy in Three Adenocarcinomas.
  36. Parallel kinase pathways stimulate actin polymerization at depolarized mitochondria.
  37. Autophagy-associated circRNA circATG7 facilitates autophagy and promotes pancreatic cancer progression.
  38. A molecular link between autophagy and circadian rhythm in plants.
  39. Ferritinophagy is involved in Bisphenol A-induced ferroptosis of renal tubular epithelial cells through the activation of the AMPK-mTOR-ULK1 pathway.
  40. Role of autophagy machinery dysregulation in bacterial chondronecrosis with osteomyelitis.
  41. Airway secretory cell fate conversion via YAP-mTORC1-dependent essential amino acid metabolism.
  42. Autophagy in Atherosclerosis: Not All Foam Cells Are Created Equal.
  43. A self-assembled leucine polymer sensitizes leukemic stem cells to chemotherapy by inhibiting autophagy in acute myeloid leukemia.
  44. Autophagy impairment in liver CD11c+ cells promotes non-alcoholic fatty liver disease through production of IL-23.
  1. Autophagy. 2022 Mar 15.
    Follicular lymphoma-associated mutations in the V-ATPase chaperone VMA21 activate autophagy creating a targetable dependency.
    Fangyang Wang, Ying Yang, Gabriel Boudagh, Eeva-Liisa Eskelinen, Daniel J Klionsky, Sami N Malek.
      The discovery of recurrent mutations in subunits and regulators of the vacuolar-type H+-translocating ATPase (V-ATPase) in follicular lymphoma (FL) highlights a role for macroautophagy/autophagy, amino-acid, and nutrient-sensing pathways in the pathogenesis of this disease. Here, we report on novel mutations in the ER-resident chaperone VMA21, which is involved in V-ATPase assembly in 12% of FL. Mutations in a novel VMA21 hotspot (p.93X) result in the removal of a C-terminal non-canonical ER retrieval signal thus causing VMA21 mislocalization to lysosomes. The resulting impairment in V-ATPase function prevents full lysosomal acidification and function, including impaired pH-dependent protein degradation as shown via lysosomal metabolomics and ultimately causes a degree of amino acid depletion in the cytoplasm. These deficiencies result in compensatory autophagy activation, as measured using multiple complementary assays in human and yeast cells. Of translational significance, the compensatory activation of autophagy creates a dependency for survival for VMA21-mutated primary human FL as shown using inhibitors to ULK1, the proximal autophagy-regulating kinase. Using high-throughput microscopy-based screening assays for autophagy-inhibiting compounds, we identify multiple clinical grade cyclin-dependent kinase inhibitors as promising drugs and thus provide new rationale for innovative clinical trials in FL harboring aberrant V-ATPase.
    Keywords:  Follicular lymphoma; VMA21 mutations; autophagy; lysosomal dysfunction; survival
    DOI:  https://doi.org/10.1080/15548627.2022.2050663
  2. Autophagy. 2022 Mar 14. 1-11
    Loss of RUBCN/rubicon in adipocytes mediates the upregulation of autophagy to promote the fasting response.
    Tadashi Yamamuro, Shuhei Nakamura, Kyosuke Yanagawa, Ayaka Tokumura, Tsuyoshi Kawabata, Atsunori Fukuhara, Hirofumi Teranishi, Maho Hamasaki, Iichiro Shimomura, Tamotsu Yoshimori.
      Upon fasting, adipocytes release their lipids that accumulate in the liver, thus promoting hepatic steatosis and ketone body production. However, the mechanisms underlying this process are not fully understood. In this study, we found that fasting caused a substantial decrease in the adipose levels of RUBCN/rubicon, a negative regulator of macroautophagy/autophagy, along with an increase in autophagy. Adipose-specific rubcn-knockout mice exhibited systemic fat loss that was not accelerated by fasting. Genetic inhibition of autophagy in adipocytes in fasted mice led to a reduction in fat loss, hepatic steatosis, and ketonemia. In terms of mechanism, autophagy decreased the levels of its substrates NCOA1/SRC-1 and NCOA2/TIF2, which are also coactivators of PPARG/PPARγ, leading to a fasting-induced reduction in the mRNA levels of adipogenic genes in adipocytes. Furthermore, RUBCN in adipocytes was degraded through the autophagy pathway, suggesting that autophagic degradation of RUBCN serves as a feedforward system for autophagy induction during fasting. Collectively, we propose that loss of adipose RUBCN promotes a metabolic response to fasting via increasing autophagic activity.
    Keywords:  Adipocytes; NCOA1; NCOA2; RUBCN; autophagy; fasting
    DOI:  https://doi.org/10.1080/15548627.2022.2047341
  3. Front Plant Sci. 2022 ;13 817251
    ROOT HAIR DEFECTIVE3 Is a Receptor for Selective Autophagy of the Endoplasmic Reticulum in Arabidopsis.
    Jiaqi Sun, Weina Wang, Huanquan Zheng.
      ROOT HAIR DEFECTIVE3 (RHD3) is a plant member of atlastin GTPases, which belong to an evolutionally conserved family of proteins that mediate the homotypic fusion of the endoplasmic reticulum (ER). An atlastin in mammalian cells has recently been shown to act as an ER-phagy receptor for selective autophagy of the ER (ER-phagy) during nutrient starvation. Although RHD3 has been indicated to play a role in ER stress response, it is not very clear how RHD3 is involved in the process. In this study, we showed that the rhd3 mutant is hyposensitive to ER as well as salt stress. We employed an YFP-tagged ER membrane marker YFP-TMC to monitor the efficiency of ER-phagy microscopically and biochemically. We found that rhd3 is defective in ER-phagy under ER stress. Furthermore, there is an increased association of YFP-RHD3 with ATG8e-marked autophagosomes. YFP-RHD3 is also visible with ATG8e in the vacuole, and there is a breakdown of YFP-RHD3 under ER stress. RHD3 has two putative ATG8 interaction motifs (AIM1-2). We revealed that RHD3 but not RHD3(ΔAIM1) physically interacts with ATG8, a core autophagosomal component that interacts with various receptor proteins to recruit cargos for degradation by selective autophagy. Furthermore, their interaction is enhanced under ER stress. We thus propose that RHD3 acts as an ER-phagy receptor under ER stress to promote ER-phagy in Arabidopsis.
    Keywords:  Atg8; ER; ER stress; ER-phagy; RHD3; autophagosomes
    DOI:  https://doi.org/10.3389/fpls.2022.817251
  4. Cell Chem Biol. 2022 Mar 04. pii: S2451-9456(22)00087-3. [Epub ahead of print]
    Rheb regulates nuclear mTORC1 activity independent of farnesylation.
    Yanghao Zhong, Xin Zhou, Kun-Liang Guan, Jin Zhang.
      The small GTPase Ras homolog enriched in brain (Rheb) plays a critical role in activating the mechanistic target of rapamycin complex 1 (mTORC1), a signaling hub that regulates various cellular functions. We recently observed nuclear mTORC1 activity, raising an intriguing question as to how Rheb, which is known to be farnesylated and localized to intracellular membranes, regulates nuclear mTORC1. In this study, we found that active Rheb is present in the nucleus and required for nuclear mTORC1 activity. We showed that inhibition of farnesyltransferase reduced cytosolic, but not nuclear, mTORC1 activity. Furthermore, a farnesylation-deficient Rheb mutant, with preferential nuclear localization and specific lysosome tethering, enables nuclear and cytosolic mTORC1 activities, respectively. These data suggest that non-farnesylated Rheb is capable of interacting with and activating mTORC1, providing mechanistic insights into the molecular functioning of Rheb as well as regulation of the recently observed, active pool of nuclear mTORC1.
    Keywords:  Compartmentation; PTM; TSC; biosensor; lipid modification; mTOR; small GTPase
    DOI:  https://doi.org/10.1016/j.chembiol.2022.02.006
  5. Life Sci Alliance. 2022 Jun;pii: e202101169. [Epub ahead of print]5(6):
    Single serine on TSC2 exerts biased control over mTORC1 activation mediated by ERK1/2 but not Akt.
    Brittany L Dunkerly-Eyring, Shi Pan, Miguel Pinilla-Vera, Desirae McKoy, Sumita Mishra, Maria I Grajeda Martinez, Christian U Oeing, Mark J Ranek, David A Kass.
      Tuberous sclerosis complex-2 (TSC2) negatively regulates mammalian target of rapamycin complex 1 (mTORC1), and its activity is reduced by protein kinase B (Akt) and extracellular response kinase (ERK1/2) phosphorylation to activate mTORC1. Serine 1364 (human) on TSC2 bidirectionally modifies mTORC1 activation by pathological growth factors or hemodynamic stress but has no impact on resting activity. We now show this modification biases to ERK1/2 but not Akt-dependent TSC2-mTORC1 activation. Endothelin-1-stimulated mTORC1 requires ERK1/2 activation and is bidirectionally modified by phospho-mimetic (S1364E) or phospho-silenced (S1364A) mutations. However, mTORC1 activation by Akt-dependent stimuli (insulin or PDGF) is unaltered by S1364 modification. Thrombin stimulates both pathways, yet only the ERK1/2 component is modulated by S1364. S1364 also has negligible impact on mTORC1 regulation by energy or nutrient status. In vivo, diet-induced obesity, diabetes, and fatty liver couple to Akt activation and are also unaltered by TSC2 S1364 mutations. This contrasts to prior reports showing a marked impact of both on pathological pressure-stress. Thus, S1364 provides ERK1/2-selective mTORC1 control and a genetic means to modify pathological versus physiological mTOR stimuli.
    DOI:  https://doi.org/10.26508/lsa.202101169
  6. Nat Commun. 2022 Mar 17. 13(1): 1436
    LC3B is an RNA-binding protein to trigger rapid mRNA degradation during autophagy.
    Hyun Jung Hwang, Hongseok Ha, Ban Seok Lee, Bong Heon Kim, Hyun Kyu Song, Yoon Ki Kim.
      LC3/ATG8 has long been appreciated to play a central role in autophagy, by which a variety of cytoplasmic materials are delivered to lysosomes and eventually degraded. However, information on the molecular functions of LC3 in RNA biology is very limited. Here, we show that LC3B is an RNA-binding protein that directly binds to mRNAs with a preference for a consensus AAUAAA motif corresponding to a polyadenylation sequence. Autophagic activation promotes an association between LC3B and target mRNAs and triggers rapid degradation of target mRNAs in a CCR4-NOT-dependent manner before autolysosome formation. Furthermore, our transcriptome-wide analysis reveals that PRMT1 mRNA, which encodes a negative regulator of autophagy, is one of the major substrates. Rapid degradation of PRMT1 mRNA by LC3B facilitates autophagy. Collectively, we demonstrate that LC3B acts as an RNA-binding protein and an mRNA decay factor necessary for efficient autophagy.
    DOI:  https://doi.org/10.1038/s41467-022-29139-1
  7. Transl Res. 2022 Mar 11. pii: S1931-5244(22)00047-0. [Epub ahead of print]
    Cell Type-Specific Mechanistic Target of Rapamycin-Dependent Distortion of Autophagy Pathways in Lupus Nephritis.
    Tiffany Caza, Chathura Wijewardena, Laith Al-Rabadi, Andras Perl.
      Pro-inflammatory immune system development, metabolomic defects, and deregulation of autophagy play interconnected roles in driving the pathogenesis of systemic lupus erythematosus (SLE). Lupus nephritis (LN) is a leading cause of morbidity and mortality in SLE. While the causes of SLE have not been clearly delineated, skewing of T and B cell differentiation, activation of antigen-presenting cells, production of antinuclear autoantibodies and pro-inflammatory cytokines are known to contribute to disease development. Underlying this process are defects in autophagy and mitophagy that cause the accumulation of oxidative stress-generating mitochondria which promote necrotic cell death. Autophagy is generally inhibited by the activation of the mammalian target of rapamycin (mTOR), a large protein kinase that underlies abnormal immune cell lineage specification in SLE. Importantly, several autophagy-regulating genes, including ATG5 and ATG7, as well as mitophagy-regulating HRES-1/Rab4A have been linked to lupus susceptibility and molecular pathogenesis. Moreover, genetically-driven mTOR activation has been associated with fulminant lupus nephritis. mTOR activation and diminished autophagy promote the expansion of pro-inflammatory Th17, Tfh and CD3+CD4-CD8- double-negative (DN) T cells at the expense of CD8+ effector memory T (EMT) cells and CD4+ regulatory T cells (Tregs). mTOR activation and aberrant autophagy also involve renal podocytes, mesangial cells, endothelial cells, and tubular epithelial cells that may compromise end-organ resistance in LN. Activation of mTOR complexes 1 (mTORC1) and 2 (mTORC2) has been identified as biomarkers of disease activation and predictors of disease flares and prognosis in SLE patients with and without LN. This review highlights recent advances in molecular pathogenesis of LN with a focus on immuno-metabolic checkpoints of autophagy and their roles in pathogenesis, prognosis and selection of targets for treatment in SLE.
    Keywords:  Autophagy; B cells; CD3(+)CD4(−)CD8(−) double-negative T cells; CD4(+) T cells; CD8(+) T cells; Rab4A; Treg; antinuclear antibodies; biomarkers; dendritic cells; endosome traffic; glomerulonephritis; interferon; lupus nephritis; lysosome; mTOR; macrophages; mechanistic target of rapamycin; metabolomics; mitochondrial dysfunction; mitophagy; netosis; plasma cells; rapamycin; sirolimus; systemic lupus erythematosus
    DOI:  https://doi.org/10.1016/j.trsl.2022.03.004
  8. FEBS J. 2022 Mar 14.
    Regulation of basal autophagy by calmodulin availability.
    Jennifer Giles, Vanessa Lopez, Elizabeth McConnaha, Matthew Hayden, Caleb Kragenbring, David Carli, Eric Wauson, Quang-Kim Tran.
      Macroautophagy (hereafter autophagy) is a process that degrades cellular components to maintain homeostasis. The Ca2+ sensor calmodulin (CaM) regulates numerous cell functions but is a limiting factor due to its insufficient availability for all target proteins. However, evidence that CaM availability regulates basal autophagy is lacking. Here, we have tested this hypothesis. CaM antagonists W-7, trifluoperazine and CGS9343b cause autophagosome accumulation and inhibit basal autophagic flux in the same manner as does chloroquine. These reagents promote the activity of AMP-activated protein kinase (AMPK) but not that of the mechanistic target of rapamycin (mTOR). Competitive binding assays using CaM sensors with different Ca2+ dependencies showed that chloroquine directly binds CaM in a Ca2+ -dependent fashion. The CaM antagonists have disparate effects on cytoplasmic Ca2+ , triggering from none to robust signals, indicating that their consistent inhibition of autophagy is due to inhibition of CaM and not Ca2+ . Chelating intracellular Ca2+ reduces the effect of the CaM antagonists to accumulate LC3-II, indicating that they do so by inhibiting CaM-dependent activities at basal Ca2+ level. The CaM antagonists cause lysosomal alkalinisation. Consistently, buffering CaM with a high-affinity CaM-binding protein that binds CaM at resting Ca2+ level increases lysosomal pH. Enhanced CaM buffering using a chimeric protein that contains two high-affinity CaM-binding sites that can collectively bind CaM at a large range of Ca2+ further increases lysosomal pH and increases LC3-II accumulation and AMPK activity, but not that of mTOR. These data demonstrate that CaM availability is required for basal autophagy.
    Keywords:  autophagy; calmodulin; calmodulin antagonists; chloroquine; lysosomal acidification
    DOI:  https://doi.org/10.1111/febs.16432
  9. Stem Cells. 2022 Mar 14. pii: sxac007. [Epub ahead of print]
    Asymmetric Pericentrosomal CD133 Endosomes Induce the Unequal Autophagic Activity During Cytokinesis in CD133-Positive Human Neuroblastoma Cells.
    Hideki Izumi, Yuanyuan Li, Michio Yasunami, Seiji Sato, Takao Mae, Yasuhiko Kaneko, Akira Nakagawara.
      CD133 is a transmembrane protein that mainly localizes to the plasma membrane in hematopoietic/neural stem cells and cancer stem cells. Although CD133 also localizes to the cytoplasm and is degraded through autophagy, the precise mechanisms responsible for the autophagic degradation of endosomal CD133 currently remain unknown. We demonstrated that endosomal CD133 has unique properties for cell homeostasis. Endosomal CD133 is degraded through p62/SQSTM1-mediated selective autophagy. However, in low basal autophagic cells, such as SK-N-DZ and SH-SY5Y cells, endosomal CD133 accumulates at the pericentrosomal region and conversely suppresses autophagy. Endosomal CD133 also asymmetrically distributes to the pericentrosomal region and induces unequal autophagic activity between 2 daughter cells during cytokinesis in SK-N-DZ and TGW cells. In addition, the asymmetric distribution of pericentrosomal CD133 endosomes and nuclear β-catenin cooperatively suppresses autophagic activity against p62 in SK-N-DZ cells. Thus, the present study suggests that the asymmetric distribution of pericentrosomal CD133 endosomes induces the symmetry breaking of autophagic activity during cytokinesis in cooperation with nuclear β-catenin.
    Keywords:  CD133; asymmetric cell division; autophagy; neuroblastoma; p62/SQSTM1; β-catenin
    DOI:  https://doi.org/10.1093/stmcls/sxac007
  10. Nat Commun. 2022 Mar 14. 13(1): 1300
    Lysosomal Ca2+-mediated TFEB activation modulates mitophagy and functional adaptation of pancreatic β-cells to metabolic stress.
    Kihyoun Park, Hyejin Lim, Jinyoung Kim, Yeseong Hwang, Yu Seol Lee, Soo Han Bae, Hyeongseok Kim, Hail Kim, Shin-Wook Kang, Joo Young Kim, Myung-Shik Lee.
      Although autophagy is critical for pancreatic β-cell function, the role and mechanism of mitophagy in β-cells are unclear. We studied the role of lysosomal Ca2+ in TFEB activation by mitochondrial or metabolic stress and that of TFEB-mediated mitophagy in β-cell function. Mitochondrial or metabolic stress induced mitophagy through lysosomal Ca2+ release, increased cytosolic Ca2+ and TFEB activation. Lysosomal Ca2+ replenishment by ER- > lysosome Ca2+ refilling was essential for mitophagy. β-cell-specific Tfeb knockout (TfebΔβ-cell) abrogated high-fat diet (HFD)-induced mitophagy, accompanied by increased ROS and reduced mitochondrial cytochrome c oxidase activity or O2 consumption. TfebΔβ-cell mice showed aggravation of HFD-induced glucose intolerance and impaired insulin release. Metabolic or mitochondrial stress induced TFEB-dependent expression of mitophagy receptors including Ndp52 and Optn, contributing to the increased mitophagy. These results suggest crucial roles of lysosomal Ca2+ release coupled with ER- > lysosome Ca2+ refilling and TFEB activation in mitophagy and maintenance of pancreatic β-cell function during metabolic stress.
    DOI:  https://doi.org/10.1038/s41467-022-28874-9
  11. Autophagy. 2022 Mar 16. 1-15
    Mitochondrial depolarization after acute ethanol treatment drives mitophagy in living mice.
    Devadoss J Samuvel, Li Li, Yasodha Krishnasamy, Monika Gooz, Kenji Takemoto, Patrick M Woster, John J Lemasters, Zhi Zhong.
      Ethanol increases hepatic mitophagy driven by unknown mechanisms. Type 1 mitophagy sequesters polarized mitochondria for nutrient recovery and cytoplasmic remodeling. In Type 2, mitochondrial depolarization (mtDepo) initiates mitophagy to remove the damaged organelles. Previously, we showed that acute ethanol administration produces reversible hepatic mtDepo. Here, we tested the hypothesis that ethanol-induced mtDepo initiates Type 2 mitophagy. GFP-LC3 transgenic mice were gavaged with ethanol (2-6 g/kg) with and without pre-treatment with agents that decrease or increase mtDepo-Alda-1, tacrolimus, or disulfiram. Without ethanol, virtually all hepatocytes contained polarized mitochondria with infrequent autophagic GFP-LC3 puncta visualized by intravital microscopy. At ~4 h after ethanol treatment, mtDepo occurred in an all-or-none fashion within individual hepatocytes, which increased dose dependently. GFP-LC3 puncta increased in parallel, predominantly in hepatocytes with mtDepo. Mitochondrial PINK1 and PRKN/parkin also increased. After covalent labeling of mitochondria with MitoTracker Red (MTR), GFP-LC3 puncta encircled MTR-labeled mitochondria after ethanol treatment, directly demonstrating mitophagy. GFP-LC3 puncta did not associate with fat droplets visualized with BODIPY558/568, indicating that increased autophagy was not due to lipophagy. Before ethanol administration, rhodamine-dextran (RhDex)-labeled lysosomes showed little association with GFP-LC3. After ethanol treatment, TFEB (transcription factor EB) translocated to nuclei, and lysosomal mass increased. Many GFP-LC3 puncta merged with RhDex-labeled lysosomes, showing autophagosomal processing into lysosomes. After ethanol treatment, disulfiram increased, whereas Alda-1 and tacrolimus decreased mtDepo, and mitophagy changed proportionately. In conclusion, mtDepo after acute ethanol treatment induces mitophagic sequestration and subsequent lysosomal processing.Abbreviations : AcAld, acetaldehyde; ADH, alcohol dehydrogenase; ALDH, aldehyde dehydrogenase; ALD, alcoholic liver disease; Alda-1, N-(1,3-benzodioxol-5-ylmethyl)-2,6-dichlorobenzamide; GAPDH, glyceraldehyde-3-phosphate dehydrogenase; GFP, green fluorescent protein; LAMP1, lysosomal-associated membrane protein 1; LMNB1, lamin B1; MAA, malondialdehyde-acetaldehyde adducts; MAP1LC3/LC3, microtubule-associated protein 1 light chain 3; MPT, mitochondrial permeability transition; mtDAMPS, mitochondrial damage-associated molecular patterns; mtDepo, mitochondrial depolarization; mtDNA, mitochondrial DNA; MTR, MitoTracker Red; PI, propidium iodide; PINK1, PTEN induced putative kinase 1; PRKN, parkin; RhDex, rhodamine dextran; TFEB, transcription factor EB; Tg, transgenic; TMRM, tetramethylrhodamine methylester; TOMM20, translocase of outer mitochondrial membrane 20; VDAC, voltage-dependent anion channel.
    Keywords:  Acetaldehyde; Alda-1; alcoholic liver disease; mitochondrial depolarization; mitophagy; tacrolimus
    DOI:  https://doi.org/10.1080/15548627.2022.2046457
  12. Autophagy. 2022 Mar 15. 1-15
    Phosphatidic acid suppresses autophagy through competitive inhibition by binding GAPC (glyceraldehyde-3-phosphate dehydrogenase) and PGK (phosphoglycerate kinase) proteins.
    Bin Guan, Yu-Tong Jiang, De-Li Lin, Wen-Hui Lin, Hong-Wei Xue.
      Macroautophagy/autophagy is a finely-regulated process in which cytoplasm encapsulated within transient organelles termed autophagosomes is delivered to lysosomes or vacuoles for degradation. Phospholipids, particularly phosphatidic acid (PA) that functions as a second messenger, play crucial and differential roles in autophagosome formation; however, the underlying mechanism remains largely unknown. Here we demonstrated that PA inhibits autophagy through competitive inhibition of the formation of ATG3 (autophagy-related)-ATG8e and ATG6-VPS34 (vacuolar protein sorting 34) complexes. PA bound to GAPC (glyceraldehyde-3-phosphate dehydrogenase) or PGK (phosphoglycerate kinase) and promoted their interaction with ATG3 or ATG6, which further attenuated the interactions of ATG3-ATG8e or ATG6-VPS34, respectively. Structural and mutational analyses revealed the mechanism of PA binding with GAPCs and PGK3, and that GAPCs or ATG8e competitively interacted with ATG3, and PGK3 or VPS34 competitively interacted with ATG6, at the same binding interface. These results elucidate the molecular mechanism of how PA inhibits autophagy through binding GAPC or PGK3 proteins and expand the understanding of the functional mode of PA, demonstrating the importance of phospholipids in plant autophagy and providing a new perspective for autophagy regulation by phospholipids.Abbreviation: ATG: autophagy-related; BiFC: bimolecular fluorescence complementation; co-IP: co-immunoprecipitation; Con A: concanamycin A; ER: endoplasmic reticulum; EZ: elongation zone; FRET-FLIM: fluorescence resonance energy transfer with fluorescence lifetime imaging microscopy; GAPDH: glyceraldehyde-3-phosphate dehydrogenase; GST: glutathione S-transferase; MDC: monodansylcadaverine; MZ: meristem zone; PA: phosphatidic acid; PAS: phagophore assembly site; PC: phosphatidylcholine; PE: phosphatidylethanolamine; PGK3: phosphoglycerate kinase; PtdIns3K: phosphatidylinositol 3-kinase; PLD: phospholipase D; TEM: transmission electron microscopy; TOR: target of rapamycin; VPS34: vacuolar protein sorting 34; WT: wild type; Y2H: yeast two-hybrid.
    Keywords:  Autophagy; autophagy-related protein; competitive inhibition; glyceraldehyde-3-phosphate dehydrogenase; phosphatidic acid; phosphoglycerate kinase
    DOI:  https://doi.org/10.1080/15548627.2022.2046449
  13. EMBO J. 2022 Mar 14. e110057
    The needs of a synapse-How local organelles serve synaptic proteostasis.
    Katarzyna M Grochowska, Maria Andres-Alonso, Anna Karpova, Michael R Kreutz.
      Synaptic function crucially relies on the constant supply and removal of neuronal membranes. The morphological complexity of neurons poses a significant challenge for neuronal protein transport since the machineries for protein synthesis and degradation are mainly localized in the cell soma. In response to this unique challenge, local micro-secretory systems have evolved that are adapted to the requirements of neuronal membrane protein proteostasis. However, our knowledge of how neuronal proteins are synthesized, trafficked to membranes, and eventually replaced and degraded remains scarce. Here, we review recent insights into membrane trafficking at synaptic sites and into the contribution of local organelles and micro-secretory pathways to synaptic function. We describe the role of endoplasmic reticulum specializations in neurons, Golgi-related organelles, and protein complexes like retromer in the synthesis and trafficking of synaptic transmembrane proteins. We discuss the contribution of autophagy and of proteasome-mediated and endo-lysosomal degradation to presynaptic proteostasis and synaptic function, as well as nondegradative roles of autophagosomes and lysosomes in signaling and synapse remodeling. We conclude that the complexity of neuronal cyto-architecture necessitates long-distance protein transport that combines degradation with signaling functions.
    Keywords:  Golgi satellites; autophagy; lysosomes; secretory trafficking
    DOI:  https://doi.org/10.15252/embj.2021110057
  14. Methods Mol Biol. 2022 ;2474 83-89
    GFP-LC3 High-Content Assay for Screening Autophagy Modulators.
    Li Zhang, Jinghua Zhao, Wen-Xing Ding, Menghang Xia.
      Autophagy plays an important role in maintaining cellular homeostasis. Defects in autophagy have been linked to various human diseases, such as cancer, neurodegenerative diseases, and cardiovascular diseases. Therefore, it is useful to develop an assay that can measure the functions of autophagy and also be used to identify autophagy modulators by screening a large number of compounds. This chapter describes a cell-based high content green fluorescent protein (GFP)-LC3 assay using mouse embryonic fibroblasts (MEF) stably expressing GFP-LC3.
    Keywords:  Autophagy; GFP-LC3 cell line; High-content assay
    DOI:  https://doi.org/10.1007/978-1-0716-2213-1_9
  15. Life Sci. 2022 Mar 15. pii: S0024-3205(22)00181-3. [Epub ahead of print] 120481
    MAP kinase-dependent autophagy controls phorbol myristate acetate-induced macrophage differentiation of HL-60 leukemia cells.
    Milos Mandic, Maja Misirkic-Marjanovic, Ljubica Vucicevic, Maja Jovanovic, Mihajlo Bosnjak, Vladimir Perovic, Biljana Ristic, Darko Ciric, Ljubica Harhaji-Trajkovic, Vladimir Trajkovic.
      We investigated the mechanisms and the role of autophagy in the differentiation of HL-60 human acute myeloid leukemia cells induced by protein kinase C (PKC) activator phorbol myristate acetate (PMA). PMA-triggered differentiation of HL-60 cells into macrophage-like cells was confirmed by cell-cycle arrest accompanied by elevated expression of macrophage markers CD11b, CD13, CD14, CD45, EGR1, CSF1R, and IL-8. The induction of autophagy was demonstrated by the increase in intracellular acidification, accumulation/punctuation of autophagosome marker LC3-II, and the increase in autophagic flux. PMA also increased nuclear translocation of autophagy transcription factors TFEB, FOXO1, and FOXO3, as well as the expression of several autophagy-related (ATG) genes in HL-60 cells. PMA failed to activate autophagy inducer AMP-activated protein kinase (AMPK) and inhibit autophagy suppressor mechanistic target of rapamycin complex 1 (mTORC1). On the other hand, it readily stimulated the phosphorylation of mitogen-activated protein (MAP) kinases extracellular signal-regulated kinase (ERK) and c-Jun N-terminal kinase (JNK) via a protein kinase C-dependent mechanism. Pharmacological or genetic inhibition of ERK or JNK suppressed PMA-triggered nuclear translocation of TFEB and FOXO1/3, ATG expression, dissociation of pro-autophagic beclin-1 from its inhibitor BCL2, autophagy induction, and differentiation of HL-60 cells into macrophage-like cells. Pharmacological or genetic inhibition of autophagy also blocked PMA-induced macrophage differentiation of HL-60 cells. Therefore, MAP kinases ERK and JNK control PMA-induced macrophage differentiation of HL-60 leukemia cells through AMPK/mTORC1-independent, TFEB/FOXO-mediated transcriptional and beclin-1-dependent post-translational activation of autophagy.
    Keywords:  Autophagy; Beclin-1; Differentiation; ERK; JNK; Leukemia
    DOI:  https://doi.org/10.1016/j.lfs.2022.120481
  16. J Cell Biol. 2022 May 02. pii: e202112024. [Epub ahead of print]221(5):
    Material properties of phase-separated TFEB condensates regulate the autophagy-lysosome pathway.
    Zheng Wang, Di Chen, Dongshi Guan, Xiaobo Liang, Jianfeng Xue, Hongyu Zhao, Guangtao Song, Jizhong Lou, Yan He, Hong Zhang.
      Very little is known about how the material properties of protein condensates assembled via liquid-liquid phase separation (LLPS) are maintained and affect physiological functions. Here we show that liquid-like condensates of the transcription factor TFEB exhibit low fusion propensity in vitro and in living cells. We directly measured the attraction force between droplets, and we characterized the interfacial tension, viscosity, and elasticity of TFEB condensates. TFEB condensates contain rigid interfacial boundaries that govern their interaction behaviors. Several small molecules, including Ro-3306, modify the material properties of TFEB condensates, increasing their size and fusion propensity. These compounds promote lysosomal biogenesis and function in a TFEB-dependent manner without changing its cytoplasmic-nuclear translocation. Ro-3306 promotes autophagy activity, facilitating degradation of toxic protein aggregates. Our study helps explain how protein condensates are maintained as physically separate entities and reveals that the material properties of TFEB condensates can be harnessed to modulate TFEB activity.
    DOI:  https://doi.org/10.1083/jcb.202112024
  17. Oncogene. 2022 Mar 12.
    eIF3a regulation of mTOR signaling and translational control via HuR in cellular response to DNA damage.
    Shijie Ma, Zizheng Dong, Yanfei Huang, Jing-Yuan Liu, Jian-Ting Zhang.
      eIF3a (eukaryotic translation initiation factor 3a), a subunit of the eIF3 complex, has been suggested to play a regulatory role in protein synthesis and in cellular response to DNA-damaging treatments. S6K1 is an effector and a mediator of mTOR complex 1 (mTORC1) in regulating protein synthesis and integrating diverse signals into control of cell growth and response to stress. Here, we show that eIF3a regulates S6K1 activity by inhibiting mTORC1 kinase via regulating Raptor synthesis. The regulation of Raptor synthesis is via eIF3a interaction with HuR (human antigen R) and binding of the eIF3a-HuR complex to the 5'-UTR of Raptor mRNA. Furthermore, mTORC1 may mediate eIF3a function in cellular response to cisplatin by regulating synthesis of NER proteins and NER activity. Taken together, we conclude that the mTOR signaling pathway may also be regulated by translational control and mediate eIF3a regulation of cancer cell response to cisplatin by regulating NER protein synthesis.
    DOI:  https://doi.org/10.1038/s41388-022-02262-5
  18. Front Neurosci. 2022 ;16 846584
    Rapamycin Improved Retinal Function and Morphology in a Mouse Model of Retinal Degeneration.
    Meng Zhao, Houting Lv, Na Yang, Guang-Hua Peng.
      The retina is an important visual organ, which is responsible for receiving light signals and transmitting them to the optic nerve center step by step. The retina contains a variety of cells, among which photoreceptor cells receive light signals and convert them into nerve signals, and are mainly responsible for light and dark vision. Retinal degeneration is mainly the degeneration of photoreceptor cells, and retinitis pigmentosa (RP) is characterized by rod degeneration followed by cone degeneration. So far, there is still a lack of effective drugs to treat RP. Here, we established a stable RP model by tail vein injection of methyl methanesulfonate to study the mechanism of retinal photoreceptor degeneration. Mechanistic target of rapamycin (mTOR) is located in the central pathway of growth and energy metabolism and changes in a variety of diseases in response to pathological changes. We found that the mTOR was activated in this model. Therefore, the inhibitor of mTOR, rapamycin was used to suppress the expression of mTOR and interfere with photoreceptor degeneration. Electroretinogram assay showed that the function of mice retina was improved. Hematoxylin and eosin staining results displayed that retinal photoreceptor thickness and morphology were improved. Also, the autophagy in rapamycin group was activated, which revealed that rapamycin may protect the retinal photoreceptor by inhibiting mTOR and then activating autophagy.
    Keywords:  MTOR; autophagy; photoreceptor; rapamycin; retinal degeneration
    DOI:  https://doi.org/10.3389/fnins.2022.846584
  19. Biochim Biophys Acta Mol Basis Dis. 2022 Mar 15. pii: S0925-4439(22)00054-0. [Epub ahead of print] 166391
    The unfolded protein response transducer IRE1α promotes reticulophagy in podocytes.
    José R Navarro-Betancourt, Joan Papillon, Julie Guillemette, Chen-Fang Chung, Takao Iwawaki, Andrey V Cybulsky.
      Glomerular diseases involving podocyte/glomerular epithelial cell (GEC) injury feature protein misfolding and endoplasmic reticulum (ER) stress. Inositol-requiring enzyme 1α (IRE1α) mediates chaperone production and autophagy during ER stress. We examined the role of IRE1α in selective autophagy of the ER (reticulophagy). Control and IRE1α knockout (KO) GECs were incubated with tunicamycin to induce ER stress and subjected to proteomic analysis. This showed IRE1α-dependent upregulation of secretory pathway mediators, including the coat protein complex II component Sec23B. Tunicamycin enhanced expression of Sec23B and the reticulophagy adaptor reticulon-3-long (RTN3L) in control, but not IRE1α KO GECs. Knockdown of Sec23B reduced autophagosome formation in response to ER stress. Tunicamycin stimulated colocalization of autophagosomes with Sec23B and RTN3L in an IRE1α-dependent manner. Similarly, during ER stress, glomerular α5 collagen IV colocalized with RTN3L and autophagosomes. Degradation of RTN3L and collagen IV increased in response to tunicamycin, and the turnover was blocked by deletion of IRE1α; thus, the IRE1α pathway promotes RTN3L-mediated reticulophagy and collagen IV may be an IRE1α-dependent reticulophagy substrate. In experimental glomerulonephritis, expression of Sec23B, RTN3L, and LC3-II increased in glomeruli of control mice, but not in podocyte-specific IRE1α KO littermates. In conclusion, during ER stress, IRE1α redirects a subset of Sec23B-positive vesicles to deliver RTN3L-coated ER fragments to autophagosomes. Reticulophagy is a novel outcome of the IRE1α pathway in podocytes and may play a cytoprotective role in glomerular diseases.
    Keywords:  Autophagy; Collagen IV; ERphagy; Endoplasmic reticulum stress; Reticulon-3; Sec23B
    DOI:  https://doi.org/10.1016/j.bbadis.2022.166391
  20. Mol Biol Cell. 2022 Mar 16. mbcE21120611
    Identification of defined structural elements within TOR2 kinase required for TOR Complex 2 assembly and function in S. cerevisiae.
    Jennifer Tsverov, Kristina Yegorov, Ted Powers.
      mTOR is a large protein kinase that assembles into two multi-subunit protein complexes, mTORC1 and mTORC2, to regulate cell growth in eukaryotic cells. While significant progress has been made in our understanding of the composition and structure of these complexes, important questions remain regarding the role of specific sequences within mTOR important for complex formation and activity. To address these issues, we have used a molecular genetic approach to explore TOR Complex assembly in budding yeast, where two closely related TOR paralogs, TOR1 and TOR2, partition preferentially into TORC1 versus TORC2, respectively. We previously identified a ∼500 amino acid segment within the N-terminal half of each protein, termed the Major Assembly Specificity (MAS) Domain, which can govern specificity in formation of each complex. In this study, we have extended the use of chimeric TOR1-TOR2 genes as a "sensitized" genetic system to identify specific subdomains rendered essential for TORC2 function, using synthetic lethal interaction analyses. Our findings reveal important design principles underlying the dimeric assembly of TORC2, as well as identify specific segments within the MAS domain critical for TORC2 function, to a level approaching single amino acid resolution. Together these findings highlight the complex and cooperative nature of TOR Complex assembly and function.
    DOI:  https://doi.org/10.1091/mbc.E21-12-0611
  21. Cell Rep. 2022 Mar 15. pii: S2211-1247(22)00276-5. [Epub ahead of print]38(11): 110535
    A trio of ubiquitin ligases sequentially drives ubiquitylation and autophagic degradation of dysfunctional yeast proteasomes.
    Richard S Marshall, Richard D Vierstra.
      As central effectors of ubiquitin (Ub)-mediated proteolysis, proteasomes are regulated at multiple levels, including degradation of unwanted or dysfunctional particles via autophagy (termed proteaphagy). In yeast, inactive proteasomes are exported from the nucleus, sequestered into cytoplasmic aggresomes via the Hsp42 chaperone, extensively ubiquitylated, and then tethered to the expanding phagophore by the autophagy receptor Cue5. Here, we demonstrate the need for ubiquitylation driven by the trio of Ub ligases (E3s), San1, Rsp5, and Hul5, which together with their corresponding E2s work sequentially to promote nuclear export and Cue5 recognition. Whereas San1 functions prior to nuclear export, Rsp5 and Hul5 likely decorate aggresome-localized proteasomes in concert. Ultimately, topologically complex Ub chain(s) containing both K48 and K63 Ub-Ub linkages are assembled, mainly on the regulatory particle, to generate autophagy-competent substrates. Because San1, Rsp5, Hul5, Hsp42, and Cue5 also participate in general proteostasis, proteaphagy likely engages a fundamental mechanism for eliminating inactive/misfolded proteins.
    Keywords:  CP: Molecular Biology; aggresome; autophagy; core protease; proteaphagy; proteasome; regulatory particle; ubiquitin; ubiquitin ligase; yeast
    DOI:  https://doi.org/10.1016/j.celrep.2022.110535
  22. Front Cell Dev Biol. 2022 ;10 848214
    Mitochondrial Dynamics, Mitophagy, and Mitochondria-Endoplasmic Reticulum Contact Sites Crosstalk Under Hypoxia.
    Shuying Wang, Jin Tan, Yuyang Miao, Qiang Zhang.
      Mitochondria are double membrane organelles within eukaryotic cells, which act as cellular power houses, depending on the continuous availability of oxygen. Nevertheless, under hypoxia, metabolic disorders disturb the steady-state of mitochondrial network, which leads to dysfunction of mitochondria, producing a large amount of reactive oxygen species that cause further damage to cells. Compelling evidence suggests that the dysfunction of mitochondria under hypoxia is linked to a wide spectrum of human diseases, including obstructive sleep apnea, diabetes, cancer and cardiovascular disorders. The functional dichotomy of mitochondria instructs the necessity of a quality-control mechanism to ensure a requisite number of functional mitochondria that are present to fit cell needs. Mitochondrial dynamics plays a central role in monitoring the condition of mitochondrial quality. The fission-fusion cycle is regulated to attain a dynamic equilibrium under normal conditions, however, it is disrupted under hypoxia, resulting in mitochondrial fission and selective removal of impaired mitochondria by mitophagy. Current researches suggest that the molecular machinery underlying these well-orchestrated processes are coordinated at mitochondria-endoplasmic reticulum contact sites. Here, we establish a holistic understanding of how mitochondrial dynamics and mitophagy are regulated at mitochondria-endoplasmic reticulum contact sites under hypoxia.
    Keywords:  hypoxia; mitochondria; mitochondria-endoplasmic reticulum contact sites; mitochondrial dynamics; mitophagy
    DOI:  https://doi.org/10.3389/fcell.2022.848214
  23. Hypertension. 2022 Mar 16. HYPERTENSIONAHA12118643
    Acute Increase of Renal Perfusion Pressure Causes Rapid Activation of mTORC1 (Mechanistic Target Of Rapamycin Complex 1) and Leukocyte Infiltration.
    Satoshi Shimada, Chun Yang, Vikash Kumar, David L Mattson, Allen W Cowley.
      BACKGROUND: The present study in Sprague-Dawley rats determined the effects of a rapid rise of renal perfusion pressure (RPP) upon the activation of mTOR (mechanistic target of rapamycin), and the effects upon the infiltration of CD68-positive macrophages/monocytes and CD3-positive T lymphocytes into the kidneys.METHODS: RPP was elevated by 40 mm Hg for 30 minutes in male Sprague-Dawley rats while measuring renal blood flow and urine flow rate. Sham rats were studied in the same way, but RPP was not changed. Since initial studies found that the acute increase of RPP resulted in activation of mTORC1 (phosphorylation of S6S235/236), the effects of inhibition of mTORC1 with rapamycin pretreatment were then determined.
    RESULTS: It was found that a 30-minute increase of RPP (≈40 mm Hg) resulted in an 8-fold increase of renal sodium excretion which was blunted by rapamycin treatment. Renal blood flow was not affected by the elevation of RPP. Activation of mTORC1 was observed. Significant increases in CD68-positive macrophages were found in both the cortex (intraglomerular and periglomerular regions) and in the outer medullary interstitial regions of the kidney and prevented by rapamycin treatment. Increases in CD3-positive T lymphocytes were observed exclusively in the periglomerular regions and prevented by rapamycin treatment. Upregulation of several proinflammatory markers was observed.
    CONCLUSIONS: We conclude that elevation of RPP rapidly activates mTORC1 resulting in infiltration of immune cells into the kidney.
    Keywords:  kidney; macrophages; mechanistic target of rapamycin complex; perfusion; rats, Sprague Dawley
    DOI:  https://doi.org/10.1161/HYPERTENSIONAHA.121.18643
  24. Pharmacol Res. 2022 Mar 15. pii: S1043-6618(22)00089-5. [Epub ahead of print] 106144
    Glutamate delta 1 receptor regulates autophagy mechanisms and affects excitatory synapse maturation in the somatosensory cortex.
    Dinesh Y Gawande, Kishore Kumar S Narasimhan, Jay M Bhatt, Ratnamala Pavuluri, Varun Kesherwani, Pratyush S Suryavanshi, Gajanan P Shelkar, Shashank M Dravid.
      The glutamate delta family of receptors is composed of GluD1 and GluD2 and serve as synaptic organizers. We have previously demonstrated several autism-like molecular and behavioral phenotypes including an increase in dendritic spines in GluD1 knockout mice. Based on previous reports we evaluated whether disruption of autophagy mechanisms may account for these phenotypes. Mouse model with conditional deletion of GluD1 from excitatory neurons in the corticolimbic regions was utilized. GluD1 loss led to overactive Akt-mTOR pathway, higher p62 and a lower LC3-II/LC3-I ratio in the somatosensory cortex suggesting reduced autophagy. Excitatory elements were increased in number but had immature phenotype based on puncta size, lower AMPA subunit GluA1 expression and impaired development switch from predominantly GluN2B to mixed GluN2A/GluN2B subunit expression. Overactive Akt-mTOR signaling and impaired autophagy was also observed in dorsal striatum upon conditional ablation of GluD1 and in the prefrontal cortex and hippocampus in constitutive knockout. Finally, cognitive deficits in novel object recognition test and fear conditioning were observed in mice with conditional ablation of GluD1 from the corticolimbic regions. Together, these results demonstrate a novel function of GluD1 in the regulation of autophagy pathway which may underlie autism phenotypes and is relevant to the genetic association of GluD1 coding, GRID1 gene with autism and other developmental disorders.
    Keywords:  GRID1; GluD1; autism; autophagy; somatosensory cortex
    DOI:  https://doi.org/10.1016/j.phrs.2022.106144
  25. Autophagy. 2022 Mar 15.
    Recombinant pro-CTSD (cathepsin D) enhances SNCA/α-Synuclein degradation in α-Synucleinopathy models.
    Susy Prieto Huarcaya, Alice Drobny, André R A Marques, Alessandro Di Spiezio, Jan Philipp Dobert, Denise Balta, Christian Werner, Tania Rizo, Lisa Gallwitz, Simon Bub, Iva Stojkovska, Nandkishore R Belur, Jens Fogh, Joseph R Mazzulli, Wei Xiang, Amitkumar Fulzele, Mario Dejung, Markus Sauer, Beate Winner, Stefan Rose-John, Philipp Arnold, Paul Saftig, Friederike Zunke.
      Parkinson disease (PD) is a neurodegenerative disorder characterized by the abnormal intracellular accumulation of SNCA/α-synuclein. While the exact mechanisms underlying SNCA pathology are not fully understood, increasing evidence suggests the involvement of autophagic as well as lysosomal deficiencies. Because CTSD (cathepsin D) has been proposed to be the major lysosomal protease involved in SNCA degradation, its deficiency has been linked to the presence of insoluble SNCA conformers in the brain of mice and humans as well as to the transcellular transmission of SNCA aggregates. We here postulate that SNCA degradation can be enhanced by the application of the recombinant human proform of CTSD (rHsCTSD). Our results reveal that rHsCTSD is efficiently endocytosed by neuronal cells, correctly targeted to lysosomes and matured to an enzymatically active protease. In dopaminergic neurons derived from induced pluripotent stem cells (iPSC) of PD patients harboring the A53T mutation within the SNCA gene, we confirm the reduction of insoluble SNCA after treatment with rHsCTSD. Moreover, we demonstrate a decrease of pathological SNCA conformers in the brain and within primary neurons of a CTSD-deficient mouse model after dosing with rHsCTSD. Boosting lysosomal CTSD activity not only enhanced SNCA clearance, but also restored endo-lysosome and autophagy function in human and murine neurons as well as tissue. Our findings indicate that CTSD is critical for SNCA clearance and function. Thus, enzyme replacement strategies utilizing CTSD may also be of therapeutic interest for the treatment of PD and other synucleinopathies aiming to decrease the SNCA burden.
    Keywords:  Parkinson disease; cathepsin D; lysosomal degradation; lysosomal storage disorders; synucleinopathies; α-synuclein
    DOI:  https://doi.org/10.1080/15548627.2022.2045534
  26. J Nanobiotechnology. 2022 Mar 15. 20(1): 131
    Maternal exposure to CeO2NPs derails placental development through trophoblast dysfunction mediated by excessive autophagy activation.
    Zhuxiu Chen, Yanqing Geng, Rufei Gao, Hangtian Zhong, Jun Chen, Xinyi Mu, Xuemei Chen, Yan Zhang, Fangfang Li, Junlin He.
      BACKGROUND: The increasing use of cerium dioxide nanoparticles (CeO2NPs) in biomedical field has attracted substantial attention about their potential risks to human health. Recent studies have shown that nanoparticles can induce placental dysfunction and even fetal abortion, but a more detailed mechanism of nanoparticles affecting placental development remains elusive.RESULTS: Here, we constructed a mouse exposure model with different doses of CeO2NPs (2.5, 4, 5, 7.5, and 10 mg kg-1 day-1, average particle size 3-5 nm), finding that intravenous exposure to pregnant mice with CeO2NPs could cause abnormal placental development. Deposited nanoparticles were able to be observed in the placental trophoblast at doses of 5 and 7.5 mg kg-1 day-1. Diving into molecular mechanisms indicated that CeO2NPs exposure could lead to autophagy activation in placental trophoblast. At the cellular level, exposure to CeO2NPs inhibited the migration and invasion of HTR-8/SVneo and activated the autophagy through mammalian target of rapamycin complex1 (mTORC1) signaling pathway. Furthermore, inhibition of autophagy initiation by 3-Methyladenine (3-MA) partially restored the function of HTR-8/SVneo, while blocking autophagic flow by Chloroquine (CQ) aggravated the functional damage.
    CONCLUSIONS: Maternal exposure to CeO2NPs impairs placental development through trophoblast dysfunction mediated by excessive autophagy activation. These results suggested that autophagy dysfunction may be a potential mechanism for the impairment of trophoblast by CeO2NPs exposure. As above, our findings provide insights into the toxicity mechanism to the reproductive system induced by rare-earth nanoparticles exposure.
    Keywords:  Autophagy; CeO2NPs; HTR-8/SVneo; Placenta; Trophoblast
    DOI:  https://doi.org/10.1186/s12951-022-01334-8
  27. Mol Psychiatry. 2022 Mar 17.
    Autistic-like behavior and cerebellar dysfunction in Bmal1 mutant mice ameliorated by mTORC1 inhibition.
    Dong Liu, Carmen Nanclares, Konstanze Simbriger, Kun Fang, Ethan Lorsung, Nam Le, Inês Silva Amorim, Kleanthi Chalkiadaki, Salil Saurav Pathak, Jin Li, Jonathan C Gewirtz, Victor X Jin, Paulo Kofuji, Alfonso Araque, Harry T Orr, Christos G Gkogkas, Ruifeng Cao.
      Although circadian and sleep disorders are frequently associated with autism spectrum disorders (ASD), it remains elusive whether clock gene disruption can lead to autistic-like phenotypes in animals. The essential clock gene Bmal1 has been associated with human sociability and its missense mutations are identified in ASD. Here we report that global Bmal1 deletion led to significant social impairments, excessive stereotyped and repetitive behaviors, as well as motor learning disabilities in mice, all of which resemble core behavioral deficits in ASD. Furthermore, aberrant cell density and immature morphology of dendritic spines were identified in the cerebellar Purkinje cells (PCs) of Bmal1 knockout (KO) mice. Electrophysiological recordings uncovered enhanced excitatory and inhibitory synaptic transmission and reduced firing rates in the PCs of Bmal1 KO mice. Differential expression of ASD- and ataxia-associated genes (Ntng2, Mfrp, Nr4a2, Thbs1, Atxn1, and Atxn3) and dysregulated pathways of translational control, including hyperactivated mammalian target of rapamycin complex 1 (mTORC1) signaling, were identified in the cerebellum of Bmal1 KO mice. Interestingly, the antidiabetic drug metformin reversed mTORC1 hyperactivation and alleviated major behavioral and PC deficits in Bmal1 KO mice. Importantly, conditional Bmal1 deletion only in cerebellar PCs was sufficient to recapitulate autistic-like behavioral and cellular changes akin to those identified in Bmal1 KO mice. Together, these results unveil a previously unidentified role for Bmal1 disruption in cerebellar dysfunction and autistic-like behaviors. Our findings provide experimental evidence supporting a putative role for dysregulation of circadian clock gene expression in the pathogenesis of ASD.
    DOI:  https://doi.org/10.1038/s41380-022-01499-6
  28. Toxicol Appl Pharmacol. 2022 Mar 10. pii: S0041-008X(22)00118-1. [Epub ahead of print]441 115973
    F-box protein FBXO41 plays vital role in arsenic trioxide-mediated autophagic death of cancer cells.
    Yashika Agrawal, Kaustubh S Nadkarni, Neha A Gupta, Rajesh Kumar Manne, Manas Kumar Santra.
      Arsenic trioxide (ATO), a potent anti-neoplastic drug, is known to prevent cancer cell growth through induction of autophagic cell death. However, importance of cellular factors in ATO-mediated autophagic cell death is poorly understood. In this study, using biochemical and immunofluorescence techniques, we show that F-box protein FBXO41 plays a critical role in anti-proliferative activity of ATO. Our study reveals the importance of FBXO41 in induction of autophagic death of cancer cells by ATO. Further, we show that the autophagic cell death induced by FBXO41 is distinct and independent of apoptosis and necrosis, showing that FBXO41 may play vital role in inducing autophagic death of apoptosis resistant cancer cells. Overall, our study elucidates the importance of FBXO41 in ATO induced autophagic cell death to prevent cancer progression, which could be explored to develop promising cancer therapeutic strategy.
    Keywords:  ATG7; Apoptosis independent autophagy; Autophagic cell death; FBXO41; Tumor suppressor; mTOR independent autophagy
    DOI:  https://doi.org/10.1016/j.taap.2022.115973
  29. Mol Neurobiol. 2022 Mar 15.
    The Role of Zinc in Axon Formation via the mTORC1 Pathway.
    Seunghyuk Choi, Donghyeon Kang, Jieun Kang, Dae Ki Hong, Beom Seok Kang, A Ra Kho, Bo Young Choi, Sung-Oh Huh, Sang Won Suh.
      Zinc is an essential micronutrient required for proper function during neuronal development because it can modulate neuronal function and structure. A fully functional description of zinc in axonal processing in the central nervous system remains elusive. Here, we define the role of intracellular zinc in axon formation and elongation, involving the mammalian target of rapamycin complex 1 (mTORC1). To investigate the involvement of zinc in axon growth, we performed an ex vivo culture of mouse hippocampal neurons and administrated ZnCl2 as a media supplement. At 2 days in vitro, the administration of zinc induced the formation of multiple and elongated axons in the ex vivo culture system. A similar outcome was witnessed in callosal projection neurons in a developing mouse brain. Treatment with extracellular zinc activated the mTORC1 signaling pathway in mouse hippocampal neuronal cultures. The zinc-dependent enhancement of neuronal processing was inhibited either by the deactivation of mTORC1 with RAPTOR shRNA or by mTOR-insensitive 4EBP1 mutants. Additionally, zinc-dependent mTORC1 activation enhanced the axonal translation of TC10 and Par3 may be responsible for axonal growth. We identified a promising role of zinc in controlling axonogenesis in the developing brain, which, in turn, may indicate a novel structural role of zinc in the cytoskeleton and developing neurons.
    Keywords:  Axon; Primary neuron; Zinc; mTORC1
    DOI:  https://doi.org/10.1007/s12035-022-02785-8
  30. Am J Respir Cell Mol Biol. 2022 Mar 16.
    Muscle Wasting in COPD: Not Enough Autophagy?
    Jean-Philippe Leduc-Gaudet, Sabah N A Hussain.
      
    Keywords:  Autophagy; COPD; Myogenesis; Satellite Cells; Skeletal muscle
    DOI:  https://doi.org/10.1165/rcmb.2022-0064ED
  31. Pharmacol Ther. 2022 Mar 15. pii: S0163-7258(22)00065-1. [Epub ahead of print] 108171
    Enhancing autophagy in Alzheimer's disease through drug repositioning.
    Mehdi Eshraghi, Mazaher Ahmadi, Saeid Afshar, Shahrokh Lorzadeh, Aida Adlimoghaddam, Nahid Rezvani Jalal, Ryan West, Sanaz Dastghaib, Somayeh Igder, Seyed Reza Naeimi Torshizi, Amir Mahmoudzadeh, Pooneh Mokarram, Tayyebeh Madrakian, Benedict C Albensi, Marek J Los, Saeid Ghavami, Stevan Pecic.
      Alzheimer's disease (AD) is one of the biggest human health threats due to increases in aging of the global population. Unfortunately, drugs for treating AD have been largely ineffective. Interestingly, downregulation of macroautophagy (autophagy) plays an essential role in AD pathogenesis. Therefore, targeting autophagy has drawn considerable attention as a therapeutic approach for the treatment of AD. However, developing new therapeutics is time-consuming and requires huge investments. One of the strategies currently under consideration for many diseases is "drug repositioning" or "drug repurposing". In this comprehensive review, we have provided an overview of the impact of autophagy on AD pathophysiology, reviewed the therapeutics that upregulate autophagy and are currently used in the treatment of other diseases, including cancers, and evaluated their repurposing as a possible treatment option for AD. In addition, we discussed the potential of applying nano-drug delivery to neurodegenerative diseases, such as AD, to overcome the challenge of crossing the blood brain barrier and specifically target molecules/pathways of interest with minimal side effects.
    Keywords:  Antimetabolites; Autophagy induction; Simvastatin; Tyrosine kinase inhibitors; mTOR inhibitors
    DOI:  https://doi.org/10.1016/j.pharmthera.2022.108171
  32. Biofactors. 2022 Mar 14.
    Autophagy flux inhibition mediated by lysosomal dysfunction participates in the cadmium exposure-induced cardiotoxicity in swine.
    Xia Zhao, Xu Shi, Yujie Yao, Xiaojing Li, Shiwen Xu.
      Cadmium (Cd), a common toxic heavy metal, is believed as a risk factor for the induction and progression of cardiovascular disease. Autophagy is a highly ordered intracellular lysosomal-mediated degradation pathway that is crucial for protein and organelle quality control. Autophagy dysfunction could develop exacerbated cardiac dysfunction. However, the role of autophagy in Cd exposure-induced cardiotoxicity remains largely unknown. In this study, the Cd-induced swine cardiotoxicity model was established by feeding with a CdCl2 suppled diet (20 mg Cd/kg diet). The results showed that Cd exposure increased the expression of endoplasmic reticulum stress-related genes (GRP78, GRP94, IRE1, XBP1, PERK, ATF4, and ATF6), increased the expression of Ca2+ release channels IP3R and RYR1 and decreased the expression of Ca2+ uptake pump SERCA1. Cd exposure upregulated the expression of autophagy-related genes (CAMKKII, AMPK, ATG5, ATG7, ATG12, Beclin1, LC3-II, and P62) and downregulated mTOR expression. Cd exposure inhibited the expression of V-ATPase and cathepsins (CTSB and CTSD), and increased the expression of cathepsins in cytoplasm. Cd exposure decreased the colocalization of autophagosome and lysosome. This study revealed that autophagy flux inhibition caused by lysosomal dysfunction participates in the cardiotoxicity induced by Cd exposure in swine.
    Keywords:  autophagy flux; cadmium; endoplasmic reticulum stress; lysosome; myocardium
    DOI:  https://doi.org/10.1002/biof.1834
  33. Autophagy. 2022 Mar 14. 1-14
    Targeting lipophagy in macrophages improves repair in multiple sclerosis.
    Mansour Haidar, Melanie Loix, Sam Vanherle, Tess Dierckx, Tim Vangansewinkel, Pascal Gervois, Esther Wolfs, Ivo Lambrichts, Jeroen F J Bogie, Jerome J A Hendriks.
      Foamy macrophages containing abundant intracellular myelin remnants are an important pathological hallmark of multiple sclerosis. Reducing the intracellular lipid burden in foamy macrophages is considered a promising therapeutic strategy to induce a phagocyte phenotype that promotes central nervous system repair. Recent research from our group showed that sustained intracellular accumulation of myelin-derived lipids skews these phagocytes toward a disease-promoting and more inflammatory phenotype. Our data now demonstrate that disturbed lipophagy, a selective form of autophagy that helps with the degradation of lipid droplets, contributes to the induction of this phenotype. Stimulating autophagy using the natural disaccharide trehalose reduced the lipid load and inflammatory phenotype of myelin-laden macrophages. Importantly, trehalose was able to boost remyelination in the ex vivo brain slice model and the in vivo cuprizone-induced demyelination model. In summary, our results provide a molecular rationale for impaired metabolism of myelin-derived lipids in macrophages, and identify lipophagy induction as a promising treatment strategy to promote remyelination.
    Keywords:  Lipid droplets; lipophagy; multiple sclerosis; phagocyte; remyelination
    DOI:  https://doi.org/10.1080/15548627.2022.2047343
  34. Biochem Biophys Res Commun. 2022 Mar 02. pii: S0006-291X(22)00325-4. [Epub ahead of print]603 130-137
    Autophagy inhibitors enhance biomolecular delivery efficiency of extracellular vesicles.
    Yuanyuan Zhang, Meijuan Huang, Lijun Jiang, Tongjuan Li, Jue Wang, Lei Zhao, Jianfeng Zhou.
      In recent years, extracellular vesicles (EVs) were loaded with therapeutic molecules to be delivered to recipient cells, so the possibility of utilizing EVs for drug delivery has been investigated in various models. Nonetheless, most EVs are degraded through the autophagy pathway after being up-taken by recipient cells, resulting in a low delivery efficiency. Here we introduced a strategy to overcome inefficient delivery of EVs. We demonstrated that autophagy inhibitors, used for reducing lysosomal degradation of EVs, enhanced the protein or plasmid DNA delivery efficiency of EVs in recipient cells without influencing the uptake of EVs by recipient cells. Moreover, autophagy inhibitors could also improve gene-editing efficiency of EV-loaded CRISPR/Cas9 system.
    Keywords:  Autophagy inhibitor; CRISPR/Cas9 system; Delivery efficiency; Extracellular vesicle
    DOI:  https://doi.org/10.1016/j.bbrc.2022.03.006
  35. Front Oncol. 2022 ;12 837373
    A Novel Copper(II) Indenoisoquinoline Complex Inhibits Topoisomerase I, Induces G2 Phase Arrest, and Autophagy in Three Adenocarcinomas.
    Caroline Molinaro, Nathalie Wambang, Till Bousquet, Anne-Sophie Vercoutter-Edouart, Lydie Pélinski, Katia Cailliau, Alain Martoriati.
      Topoisomerases, targets of inhibitors used in chemotherapy, induce DNA breaks accumulation leading to cancer cell death. A newly synthesized copper(II) indenoisoquinoline complex WN197 exhibits a cytotoxic effect below 0.5 µM, on MDA-MB-231, HeLa, and HT-29 cells. At low doses, WN197 inhibits topoisomerase I. At higher doses, it inhibits topoisomerase IIα and IIβ, and displays DNA intercalation properties. DNA damage is detected by the presence of γH2AX. The activation of the DNA Damage Response (DDR) occurs through the phosphorylation of ATM/ATR, Chk1/2 kinases, and the increase of p21, a p53 target. WN197 induces a G2 phase arrest characterized by the unphosphorylated form of histone H3, the accumulation of phosphorylated Cdk1, and an association of Cdc25C with 14.3.3. Cancer cells die by autophagy with Beclin-1 accumulation, LC3-II formation, p62 degradation, and RAPTOR phosphorylation in the mTOR complex. Finally, WN197 by inhibiting topoisomerase I at low concentration with high efficiency is a promising agent for the development of future DNA damaging chemotherapies.
    Keywords:  adenocarcinoma; autophagy; cell cycle; copper(II) complex; indenoisoquinoline; topoisomerase
    DOI:  https://doi.org/10.3389/fonc.2022.837373
  36. Curr Biol. 2022 Mar 08. pii: S0960-9822(22)00328-1. [Epub ahead of print]
    Parallel kinase pathways stimulate actin polymerization at depolarized mitochondria.
    Tak Shun Fung, Rajarshi Chakrabarti, Jana Kollasser, Klemens Rottner, Theresia E B Stradal, Frieda Kage, Henry N Higgs.
      Mitochondrial damage (MtD) represents a dramatic change in cellular homeostasis, necessitating metabolic changes and stimulating mitophagy. One rapid response to MtD is a rapid peri-mitochondrial actin polymerization termed ADA (acute damage-induced actin). The activation mechanism for ADA is unknown. Here, we use mitochondrial depolarization or the complex I inhibitor metformin to induce ADA. We show that two parallel signaling pathways are required for ADA. In one pathway, increased cytosolic calcium in turn activates PKC-β, Rac, WAVE regulatory complex, and Arp2/3 complex. In the other pathway, a drop in cellular ATP in turn activates AMPK (through LKB1), Cdc42, and FMNL formins. We also identify putative guanine nucleotide exchange factors for Rac and Cdc42, Trio and Fgd1, respectively, whose phosphorylation states increase upon mitochondrial depolarization and whose suppression inhibits ADA. The depolarization-induced calcium increase is dependent on the mitochondrial sodium-calcium exchanger NCLX, suggesting initial mitochondrial calcium efflux. We also show that ADA inhibition results in enhanced mitochondrial shape changes upon mitochondrial depolarization, suggesting that ADA inhibits these shape changes. These depolarization-induced shape changes are not fragmentation but a circularization of the inner mitochondrial membrane, which is dependent on the inner mitochondrial membrane protease Oma1. ADA inhibition increases the proteolytic processing of an Oma1 substrate, the dynamin GTPase Opa1. These results show that ADA requires the combined action of the Arp2/3 complex and formin proteins to polymerize a network of actin filaments around mitochondria and that the ADA network inhibits the rapid mitochondrial shape changes that occur upon mitochondrial depolarization.
    Keywords:  AMPK; Arp2/3 complex; CCCP; FMNL formins; OMA1; OPA1; PKCβ; actin; calcium; mitochondrial depolarization
    DOI:  https://doi.org/10.1016/j.cub.2022.02.058
  37. Cell Death Dis. 2022 Mar 14. 13(3): 233
    Autophagy-associated circRNA circATG7 facilitates autophagy and promotes pancreatic cancer progression.
    Zhiwei He, Kun Cai, Zhirui Zeng, Shan Lei, Wenpeng Cao, Xiaowu Li.
      Dysregulation of autophagy and circular RNAs (circRNAs) are involved in the pancreatic cancer (PC) progression. However, the regulatory network between circRNAs, autophagy, and PC progression remains unknown. Herein, we demonstrated that autophagy-associated circRNA circ-autophagy related 7 (circATG7) was elevated in PC tissues compared to adjacent tissues, and in PC cells treated with EBSS and hypoxia. circATG7 expression was positively associated with tumor diameter and lymph node invasion in patients with PC. circATG7 overexpression promoted PC cell proliferation, mobility, and autophagy in vitro, while circATG7 knockdown induced the opposite effects. ATG7 inhibition attenuated the effects of circATG7 on the biological functions of PC cells. CircATG7 is located in the cell cytoplasm and nucleus. Cytoplasmic circATG7 sponged miR-766-5p and decreased its expression, and increased the expression of ATG7, a target gene of miR-766-5p. Nuclear circATG7 acted as a scaffold to increase the interaction between the human antigen R protein and ATG7 mRNA and enhanced ATG mRNA stability. Furthermore, we demonstrated that circATG7 regulates PC cell proliferation and metastasis in vivo via ATG7-dependent autophagy. In conclusion, our results demonstrated that circATG7 accelerates PC progression via miR-766-5p/ATG7 and that HUR/ATG7 depends on autophagic flux. Thus, circATG7 may be a potential therapeutic target for PC.
    DOI:  https://doi.org/10.1038/s41419-022-04677-0
  38. J Integr Plant Biol. 2022 Mar 16.
    A molecular link between autophagy and circadian rhythm in plants.
    Weijun Chen, Zhaotun Hu, MengTing Yu, Sirui Zhu, Junjie Xing, Limei Song, Wenxuan Pu, Feng Yu.
      Extremely high or low autophagy levels disrupt plant survival under nutrient starvation. Recently, autophagy has been reported to display rhythms in animals. However, the mechanism of circadian regulation of autophagy is still unclear. Here, we observed that autophagy has a robust rhythm and that various autophagy-related genes (ATGs) are rhythmically expressed in Arabidopsis. Chromatin immunoprecipitation (ChIP) and dual-luciferase (LUC) analyses showed that the core oscillator gene TIMING OF CAB EXPRESSION 1 (TOC1) directly binds to the promoters of ATG (ATG1a, ATG2, and ATG8d) and negatively regulates autophagy activities under nutritional stress. Furthermore, autophagy defects might affect endogenous rhythms by reducing the rhythm amplitude of TOC1 and shortening the rhythm period of CIRCADIAN CLOCK-ASSOCIATED 1 (CCA1). Autophagy is essential for the circadian clock pattern in seedling development and plant sensitivity to nutritional deficiencies. Taken together, our studies reveal a plant strategy in which the TOC1-ATG axis involved in autophagy-rhythm crosstalk to fine-tune the intensity of autophagy. This article is protected by copyright. All rights reserved.
    Keywords:   ATG8d ; Circadian clock; TOC1; autophagy activities; nutritional stress
    DOI:  https://doi.org/10.1111/jipb.13250
  39. Food Chem Toxicol. 2022 Mar 12. pii: S0278-6915(22)00107-7. [Epub ahead of print]163 112909
    Ferritinophagy is involved in Bisphenol A-induced ferroptosis of renal tubular epithelial cells through the activation of the AMPK-mTOR-ULK1 pathway.
    Lijuan Bao, Caijun Zhao, Lianjun Feng, Yihong Zhao, Shiyu Duan, Min Qiu, Keyi Wu, Naisheng Zhang, Xiaoyu Hu, Yunhe Fu.
      Bisphenol A (BPA) is a common environmental contaminant, whose exposure is associated with the progression of various kidney diseases. BPA exposure has turned out to be associated with cytotoxicity to renal tubular epithelial cells, but its underlying mechanism remains unknown. Herein, we found that BPA induced ferroptosis in kidney and renal tubular epithelial cells, as showed by increased intracellular iron accumulation, lipid peroxidation and cells death upon BPA exposure. Additionally, utilization of ferrostatin-1 and desferrioxamine, typical ferroptosis inhibitors, can fundamentally diminish cells death. Intriguingly, we discovered that autophagy inhibitor chloroquine can shield renal tubular epithelial cells from BPA-caused ferroptosis. Furthermore, we found that ferritinophagy, a phenomenon that degradation of ferritin and inducing subsequent iron overload, occurred after BPA exposure and excessive iron promoted ferroptosis through Fenton reaction. We next demonstrated that BPA activated the AMPK-mTOR-ULK1 signaling pathway. In turn, AMPK, mTOR, and ULK1 knockdown dramatically mitigated BPA-induced TCMK-1 cells death, and decreased MDA and LC3 levels, but increased FTH protein content. These results indicate that activation of the AMPK-mTOR-ULK1 signaling is involved in BPA-induced ferritinophagy. In conclusion, renal dysfunction and renal tubular epithelial damage induced by BPA are linked to ferroptosis, which depends on the activation of ferritinophagy through AMPK-mTOR-ULK1 axis.
    Keywords:  Autophagy; BPA; Ferritinophagy; Ferroptosis; Renal tubular
    DOI:  https://doi.org/10.1016/j.fct.2022.112909
  40. Poult Sci. 2022 Jan 30. pii: S0032-5791(22)00055-4. [Epub ahead of print]101(5): 101750
    Role of autophagy machinery dysregulation in bacterial chondronecrosis with osteomyelitis.
    Alison Ramser, Elizabeth Greene, Adnan A K Alrubaye, Robert Wideman, Sami Dridi.
      Autophagy is a cell survival and homeostasis mechanism involving lysosomal degradation of cellular components and foreign bodies. It plays a role in bone homeostasis, skeletal diseases, and bacterial infections as both a cell-survival or cell-death pathway. This study sought to determine if autophagy played a role in bacterial chondronecrosis with osteomyelitis (BCO). BCO is a prominent cause of lameness in modern broilers and results from bacterial infection of mechanically stressed leg bone growth plates. The protein and gene expression of key autophagy machinery was analyzed in both normal and BCO-affected broilers using real-time qPCR and immunoblot, respectively. Gene expression showed a significant downregulation of key target signatures involved in every stage of autophagy in BCO-affected bone, such as ATG13, SQSTM1 (p62), ATG9B, ATG16L, ATG12, LC3C, and RAB7A. Additionally, protein expression for LC3 was also significantly lower in BCO. An in vitro study using human fetal osteoblast cells challenged with BCO isolate, Staphylococcus agnetis 908, showed a similar dysregulation of autophagy machinery along with a significant decrease in cell viability. When autophagy was inhibited via 3-methyladenine or chloroquine, comparable decreases in cell viability were seen along with dysregulation of autophagy machinery. Together, these results are the first to implicate autophagy machinery dysregulation in the pathology of BCO.
    Keywords:  autophagy; bacterial chondronecrosis with osteomyelitis; broiler; lameness; osteoblast
    DOI:  https://doi.org/10.1016/j.psj.2022.101750
  41. EMBO J. 2022 Mar 14. e109365
    Airway secretory cell fate conversion via YAP-mTORC1-dependent essential amino acid metabolism.
    Hae Yon Jeon, Jinwook Choi, Lianne Kraaier, Young Hoon Kim, David Eisenbarth, Kijong Yi, Ju-Gyeong Kang, Jin Woo Kim, Hyo Sup Shim, Joo-Hyeon Lee, Dae-Sik Lim.
      Tissue homeostasis requires lineage fidelity of stem cells. Dysregulation of cell fate specification and differentiation leads to various diseases, yet the cellular and molecular mechanisms governing these processes remain elusive. We demonstrate that YAP/TAZ activation reprograms airway secretory cells, which subsequently lose their cellular identity and acquire squamous alveolar type 1 (AT1) fate in the lung. This cell fate conversion is mediated via distinctive transitional cell states of damage-associated transient progenitors (DATPs), recently shown to emerge during injury repair in mouse and human lungs. We further describe a YAP/TAZ signaling cascade to be integral for the fate conversion of secretory cells into AT1 fate, by modulating mTORC1/ATF4-mediated amino acid metabolism in vivo. Importantly, we observed aberrant activation of the YAP/TAZ-mTORC1-ATF4 axis in the altered airway epithelium of bronchiolitis obliterans syndrome, including substantial emergence of DATPs and AT1 cells with severe pulmonary fibrosis. Genetic and pharmacologic inhibition of mTORC1 activity suppresses lineage alteration and subepithelial fibrosis driven by YAP/TAZ activation, proposing a potential therapeutic target for human fibrotic lung diseases.
    Keywords:  Damage-Associated Transient Progenitors; Hippo-YAP signaling; essential amino acid metabolism; mTORC1-ATF4 axis; pulmonary fibrosis and bronchiolitis obliterans
    DOI:  https://doi.org/10.15252/embj.2021109365
  42. Circ Res. 2022 Mar 18. 130(6): 848-850
    Autophagy in Atherosclerosis: Not All Foam Cells Are Created Equal.
    Gordon A Francis, Babak Razani.
      
    Keywords:  Editorials; atherosclerosis; autophagy; cholesterol; inflammation; macrophages; muscle, smooth
    DOI:  https://doi.org/10.1161/CIRCRESAHA.122.320857
  43. Haematologica. 2022 Mar 17.
    A self-assembled leucine polymer sensitizes leukemic stem cells to chemotherapy by inhibiting autophagy in acute myeloid leukemia.
    Xi Xu, Jian Wang, Tong Tong, Wenwen Zhang, Jin Wang, Weiwei Ma, Shunqing Wang, Dunhua Zhou, Jun Wu, Linjia Jiang, Meng Zhao.
      Chemotherapy is the primary treatment option for acute myeloid leukemia (AML), but leukemic stem cells (LSCs) can survive chemotherapy for disease recurrence and refractory. Here, we found that AML cells obtained from relapsed patients had increased autophagy levels than de novo AML cells. Furthermore, Doxorubicin (DOX) treatment stimulated autophagy in LSCs by repressing the mTOR pathway, and pharmaceutical inhibition of autophagy rendered chemoresistant LSCs sensitive to DOX treatment in MLL-AF9 induced murine AML. Moreover, we developed a self-assembled leucine polymer, which activated mTOR to inhibit autophagy in AML cells by releasing leucine. The leucine polymer loaded DOX (Leu-DOX) induced much less autophagy but more robust apoptosis in AML cells than the DOX treatment. Notably, the leucine polymer and Leu-DOX were specifically uptaken by AML cells and LSCs but not by normal hematopoietic cells and hematopoietic stem/progenitor cells in the bone marrow. Consequently, Leu-DOX efficiently reduced LSCs and prolonged the survival of AML mice, with limited myeloablation and tissue damage side effects than DOX treatment. Overall, we proposed that the newly developed Leu-DOX is an effective autophagy inhibitor and an ideal drug to efficiently eliminate LSCs, thus serving as a revolutionary strategy to enhance the chemotherapy efficacy in AML.
    DOI:  https://doi.org/10.3324/haematol.2021.280290
  44. Nat Commun. 2022 Mar 17. 13(1): 1440
    Autophagy impairment in liver CD11c+ cells promotes non-alcoholic fatty liver disease through production of IL-23.
    Lauriane Galle-Treger, Doumet Georges Helou, Christine Quach, Emily Howard, Benjamin P Hurrell, German R Aleman Muench, Pedram Shafiei-Jahani, Jacob D Painter, Andrea Iorga, Lily Dara, Juliet Emamaullee, Lucy Golden-Mason, Hugo R Rosen, Pejman Soroosh, Omid Akbari.
      There has been a global increase in rates of obesity with a parallel epidemic of non-alcoholic fatty liver disease (NAFLD). Autophagy is an essential mechanism involved in the degradation of cellular material and has an important function in the maintenance of liver homeostasis. Here, we explore the effect of Autophagy-related 5 (Atg5) deficiency in liver CD11c+ cells in mice fed HFD. When compared to control mice, Atg5-deficient CD11c+ mice exhibit increased glucose intolerance and decreased insulin sensitivity when fed HFD. This phenotype is associated with the development of NAFLD. We observe that IL-23 secretion is induced in hepatic CD11c+ myeloid cells following HFD feeding. We demonstrate that both therapeutic and preventative IL-23 blockade alleviates glucose intolerance, insulin resistance and protects against NAFLD development. This study provides insights into the function of autophagy and IL-23 production by hepatic CD11c+ cells in NAFLD pathogenesis and suggests potential therapeutic targets.
    DOI:  https://doi.org/10.1038/s41467-022-29174-y
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