bims-auttor Biomed News
on Autophagy and mTOR
Issue of 2022‒09‒18
67 papers selected by
Viktor Korolchuk, Newcastle University
  1. Structural basis for FLCN RagC GAP activation in MiT-TFE substrate-selective mTORC1 regulation.
  2. Brain-enriched RagB isoforms regulate the dynamics of mTORC1 activity through GATOR1 inhibition.
  3. A Rag GTPase dimer code defines the regulation of mTORC1 by amino acids.
  4. Cytoskeletal vimentin regulates cell size and autophagy through mTORC1 signaling.
  5. A HaloTag-based reporter processing assay to monitor autophagic flux.
  6. The emerging mechanisms and functions of microautophagy.
  7. Sequential conversion of PtdIns3P to PtdIns(3,5)P2 via endosome maturation couples nutrient signaling to lysosome reformation and basal autophagy.
  8. TRIM27 is an autophagy substrate facilitating mitochondria clustering and mitophagy via phosphorylated TBK1.
  9. Identification of CUL4A-DDB1-WDFY1 as an E3 ubiquitin ligase complex involved in initiation of lysophagy.
  10. Protein expression in exocrine pancreatic diseases. Focus on VMP1 mediated autophagy.
  11. The hallmark domain of the oldest autophagy receptor family is a cargo-binding module.
  12. The RNA polymerase II subunit Rpb9 activates ATG1 transcription and autophagy.
  13. Modulation of autophagy as a therapeutic strategy for Toxoplasma gondii infection.
  14. Autophagy regulated by the HIF/REDD1/mTORC1 signaling is progressively increased during erythroid differentiation under hypoxia.
  15. RIPOR2-mediated autophagy dysfunction is critical for aminoglycoside-induced hearing loss.
  16. Mitochondrial autophagy in the sleeping brain.
  17. Recent advances of the mammalian target of rapamycin signaling in mesenchymal stem cells.
  18. NCOA4: More than a receptor for ferritinophagy.
  19. Conjugation of the ubiquitin family proteins to phospholipids.
  20. Recent advances and limitations of mTOR inhibitors in the treatment of cancer.
  21. A plant virus hijacks phosphatidylinositol-3,5-bisphosphate to escape autophagic degradation in its insect vector.
  22. Calcium homeostasis in the control of mitophagy.
  23. Autophagy in renal fibrosis: Protection or promotion?
  24. Plant autophagy and intracellular trafficking.
  25. Novel Bcl-2 Inhibitors Selectively Disrupt the Autophagy-Specific Bcl-2-Beclin 1 Protein-Protein Interaction.
  26. Slingshot homolog-1 mediates the secretion of small extracellular vesicles containing misfolded proteins by regulating autophagy cargo receptors and actin dynamics.
  27. Targeting the ATG5-ATG16L1 Protein-Protein Interaction with a Hydrocarbon-Stapled Peptide Derived from ATG16L1 for Autophagy Inhibition.
  28. Reversing BCG-mediated autophagy inhibition and mycobacterial survival to improve vaccine efficacy.
  29. Protective and deleterious effects of autophagy in the setting of myocardial ischemia/reperfusion injury: an overview.
  30. MCM7 supports the stemness of bladder cancer stem-like cells by enhancing autophagic flux.
  31. Apicoplast biogenesis mediated by ATG8 requires the ATG12-ATG5-ATG16L and SNAP29 complexes in Toxoplasma gondii.
  32. Physiological media advance cell culture experiments.
  33. Structure and mechanism of human cystine exporter cystinosin.
  34. Natural phytochemicals that affect autophagy in the treatment of oral diseases and infections: A review.
  35. Membrane estrogen receptor ERα activation improves tau clearance via autophagy induction in a tauopathy cell model.
  36. cAMP-Protein Kinase A and Stress-Activated MAP Kinase signaling mediate transcriptional control of autophagy in fission yeast during glucose limitation or starvation.
  37. Structure of the HOPS tethering complex, a lysosomal membrane fusion machinery.
  38. ER stress induces myocardial dysfunction and cardiac autophagy in Sestrin2 knockout mice.
  39. Functional and structural consequences of TBK1 missense variants in frontotemporal lobar degeneration and amyotrophic lateral sclerosis.
  40. Sensing local energetics to acutely regulate mitophagy in skeletal muscle.
  41. Tissue-restricted inhibition of mTOR using chemical genetics.
  42. The core autophagy protein ATG5 controls the polarity of the Golgi apparatus and insulin secretion of pancreatic beta cells.
  43. The role of autophagy-lysosomal pathway in motor neuron diseases.
  44. Aberrant autophagy in lysosomal storage disorders marked by a lysosomal SNARE protein shortage due to suppression of endocytosis.
  45. The disturbance of protein synthesis/degradation homeostasis is a common trait of age-related neurodegenerative disorders.
  46. The relationship of alpha-synuclein to mitochondrial dynamics and quality control.
  47. Proteomic examination of Cornus officinalis stimulated 1.1B4 human pancreatic cells reveals activation of autophagy and Keap1/Nrf2 pathway.
  48. Secretory autophagy promotes RAB37-mediated insulin secretion under glucose stimulation both in vitro and in vivo.
  49. Drug repurposing in cancer neuroscience: From the viewpoint of the autophagy-mediated innervated niche.
  50. PSMG2-controlled proteasome-autophagy balance mediates the tolerance for MEK-targeted therapy in triple-negative breast cancer.
  51. Stabilization of DEPTOR sensitizes hypopharyngeal cancer to radiotherapy via targeting degradation.
  52. Coordinated control of the ADP-heptose/ALPK1 signalling network by the E3 ligases TRAF6, TRAF2/c-IAP1 and LUBAC.
  53. A biochemical reconstitution approach to identify autophagy receptors for aggrephagy in mammalian cells.
  54. Brain-restricted mTOR inhibition with binary pharmacology.
  55. Pathological and Therapeutic Advances in Parkinson's Disease: Mitochondria in the Interplay.
  56. GCAF(TMEM251) regulates lysosome biogenesis by activating the mannose-6-phosphate pathway.
  57. ULK1 inhibition attenuates telomerase activity in hepatic cells.
  58. The critical role of dysregulated autophagy in the progression of diabetic kidney disease.
  59. Optineurin-mediated mitophagy as a potential therapeutic target for intervertebral disc degeneration.
  60. ER as master regulator of membrane trafficking and organelle function.
  61. Advanced oxidation protein products mediate human keratinocytes apoptosis by inducing cell autophagy through the mTOR-Beclin-1 pathway.
  62. Exogenous hydrogen sulfide inhibits the senescence of cardiomyocytes through modulating mitophagy in rats.
  63. The emergent role of mitochondrial surveillance in cellular health.
  64. mTOR-regulated mitochondrial metabolism limits mycobacterium-induced cytotoxicity.
  65. Structure of an MHC I-tapasin-ERp57 editing complex defines chaperone promiscuity.
  66. Evaluation of mTORC1 signaling in mouse atherosclerotic macrophages by flow cytometry and immunofluorescence.
  67. Regulation of Archease by the mTOR-vATPase axis.
  1. Sci Adv. 2022 Sep 16. 8(37): eadd2926
    Structural basis for FLCN RagC GAP activation in MiT-TFE substrate-selective mTORC1 regulation.
    Rachel M Jansen, Roberta Peruzzo, Simon A Fromm, Adam L Yokom, Roberto Zoncu, James H Hurley.
      The mechanistic target of rapamycin complex 1 (mTORC1) regulates cell growth and catabolism in response to nutrients through phosphorylation of key substrates. The tumor suppressor folliculin (FLCN) is a RagC/D guanosine triphosphatase (GTPase)-activating protein (GAP) that regulates mTORC1 phosphorylation of MiT-TFE transcription factors, controlling lysosome biogenesis and autophagy. We determined the cryo-electron microscopy structure of the active FLCN complex (AFC) containing FLCN, FNIP2, the N-terminal tail of SLC38A9, the RagAGDP:RagCGDP.BeFx- GTPase dimer, and the Ragulator scaffold. Relative to the inactive lysosomal FLCN complex structure, FLCN reorients by 90°, breaks contact with RagA, and makes previously unseen contacts with RagC that position its Arg164 finger for catalysis. Disruption of the AFC-specific interfaces of FLCN and FNIP2 with RagC eliminated GAP activity and led to nuclear retention of TFE3, with no effect on mTORC1 substrates S6K or 4E-BP1. The structure provides a basis for regulation of an mTORC1 substrate-specific pathway and a roadmap to discover MiT-TFE family selective mTORC1 antagonists.
    DOI:  https://doi.org/10.1126/sciadv.add2926
  2. Nat Cell Biol. 2022 Sep;24(9): 1407-1421
    Brain-enriched RagB isoforms regulate the dynamics of mTORC1 activity through GATOR1 inhibition.
    Gianluca Figlia, Sandra Müller, Anna M Hagenston, Susanne Kleber, Mykola Roiuk, Jan-Philipp Quast, Nora Ten Bosch, Damian Carvajal Ibañez, Daniela Mauceri, Ana Martin-Villalba, Aurelio A Teleman.
      Mechanistic target of rapamycin complex 1 (mTORC1) senses nutrient availability to appropriately regulate cellular anabolism and catabolism. During nutrient restriction, different organs in an animal do not respond equally, with vital organs being relatively spared. This raises the possibility that mTORC1 is differentially regulated in different cell types, yet little is known about this mechanistically. The Rag GTPases, RagA or RagB bound to RagC or RagD, tether mTORC1 in a nutrient-dependent manner to lysosomes where mTORC1 becomes activated. Although the RagA and B paralogues were assumed to be functionally equivalent, we find here that the RagB isoforms, which are highly expressed in neurons, impart mTORC1 with resistance to nutrient starvation by inhibiting the RagA/B GTPase-activating protein GATOR1. We further show that high expression of RagB isoforms is observed in some tumours, revealing an alternative strategy by which cancer cells can retain elevated mTORC1 upon low nutrient availability.
    DOI:  https://doi.org/10.1038/s41556-022-00977-x
  3. Nat Cell Biol. 2022 Sep;24(9): 1394-1406
    A Rag GTPase dimer code defines the regulation of mTORC1 by amino acids.
    Peter Gollwitzer, Nina Grützmacher, Sabine Wilhelm, Daniel Kümmel, Constantinos Demetriades.
      Amino acid availability controls mTORC1 activity via a heterodimeric Rag GTPase complex that functions as a scaffold at the lysosomal surface, bringing together mTORC1 with its activators and effectors. Mammalian cells express four Rag proteins (RagA-D) that form dimers composed of RagA/B bound to RagC/D. Traditionally, the Rag paralogue pairs (RagA/B and RagC/D) are referred to as functionally redundant, with the four dimer combinations used interchangeably in most studies. Here, by using genetically modified cell lines that express single Rag heterodimers, we uncover a Rag dimer code that determines how amino acids regulate mTORC1. First, RagC/D differentially define the substrate specificity downstream of mTORC1, with RagD promoting phosphorylation of its lysosomal substrates TFEB/TFE3, while both Rags are involved in the phosphorylation of non-lysosomal substrates such as S6K. Mechanistically, RagD recruits mTORC1 more potently to lysosomes through increased affinity to the anchoring LAMTOR complex. Furthermore, RagA/B specify the signalling response to amino acid removal, with RagB-expressing cells maintaining lysosomal and active mTORC1 even upon starvation. Overall, our findings reveal key qualitative differences between Rag paralogues in the regulation of mTORC1, and underscore Rag gene duplication and diversification as a potentially impactful event in mammalian evolution.
    DOI:  https://doi.org/10.1038/s41556-022-00976-y
  4. PLoS Biol. 2022 Sep;20(9): e3001737
    Cytoskeletal vimentin regulates cell size and autophagy through mTORC1 signaling.
    Ponnuswamy Mohanasundaram, Leila S Coelho-Rato, Mayank Kumar Modi, Marta Urbanska, Franziska Lautenschläger, Fang Cheng, John E Eriksson.
      The nutrient-activated mTORC1 (mechanistic target of rapamycin kinase complex 1) signaling pathway determines cell size by controlling mRNA translation, ribosome biogenesis, protein synthesis, and autophagy. Here, we show that vimentin, a cytoskeletal intermediate filament protein that we have known to be important for wound healing and cancer progression, determines cell size through mTORC1 signaling, an effect that is also manifested at the organism level in mice. This vimentin-mediated regulation is manifested at all levels of mTOR downstream target activation and protein synthesis. We found that vimentin maintains normal cell size by supporting mTORC1 translocation and activation by regulating the activity of amino acid sensing Rag GTPase. We also show that vimentin inhibits the autophagic flux in the absence of growth factors and/or critical nutrients, demonstrating growth factor-independent inhibition of autophagy at the level of mTORC1. Our findings establish that vimentin couples cell size and autophagy through modulating Rag GTPase activity of the mTORC1 signaling pathway.
    DOI:  https://doi.org/10.1371/journal.pbio.3001737
  5. Autophagy. 2022 Sep 12.
    A HaloTag-based reporter processing assay to monitor autophagic flux.
    Willa Wen-You Yim, Hayashi Yamamoto, Noboru Mizushima.
      Monitoring mammalian macroautophagic/autophagic flux is necessary in most autophagy studies but has generally been difficult to do. Here, we discuss our recent report of a HaloTag-based processing method that offers a straightforward readout for autophagic flux. We found that the self-labeling protein HaloTag becomes resistant to proteolysis when labeled with its ligand. Fusing HaloTag to an autophagy protein such as LC3 results in a reporter that is completely degraded when delivered into lysosomes but, when pulse-labeled with HaloTag ligand, releases free HaloTagligand when processed by lysosomal enzymes. The quantifiable amount of free HaloTagligand, observed by immunoblotting or in-gel fluorescence detection, reflects autophagic flux. Besides being compatible with fluorescence microscopy and flow cytometry applications, this quantitative assay can be readily adapted to monitor most autophagy pathways or the autophagic degradation of a protein of interest.
    Keywords:  HaloTag; autophagic activity; autophagic flux; autophagy; lysosomal degradation; lysosome; processing assay; protein degradation; protein turnover; pulse-labeling
    DOI:  https://doi.org/10.1080/15548627.2022.2123638
  6. Nat Rev Mol Cell Biol. 2022 Sep 12.
    The emerging mechanisms and functions of microautophagy.
    Liming Wang, Daniel J Klionsky, Han-Ming Shen.
      'Autophagy' refers to an evolutionarily conserved process through which cellular contents, such as damaged organelles and protein aggregates, are delivered to lysosomes for degradation. Different forms of autophagy have been described on the basis of the nature of the cargoes and the means used to deliver them to lysosomes. At present, the prevailing categories of autophagy in mammalian cells are macroautophagy, microautophagy and chaperone-mediated autophagy. The molecular mechanisms and biological functions of macroautophagy and chaperone-mediated autophagy have been extensively studied, but microautophagy has received much less attention. In recent years, there has been a growth in research on microautophagy, first in yeast and then in mammalian cells. Here we review this form of autophagy, focusing on selective forms of microautophagy. We also discuss the upstream regulatory mechanisms, the crosstalk between macroautophagy and microautophagy, and the functional implications of microautophagy in diseases such as cancer and neurodegenerative disorders in humans. Future research into microautophagy will provide opportunities to develop novel interventional strategies for autophagy- and lysosome-related diseases.
    DOI:  https://doi.org/10.1038/s41580-022-00529-z
  7. Autophagy. 2022 Sep 14.
    Sequential conversion of PtdIns3P to PtdIns(3,5)P2 via endosome maturation couples nutrient signaling to lysosome reformation and basal autophagy.
    Samuel J Rodgers, Emily I Jones, Christina A Mitchell, Meagan J McGrath.
      Macroautophagy/autophagy occurs basally under nutrient-rich conditions in most mammalian cells, contributing to protein and organelle quality control, and protection against ageing and neurodegeneration. During autophagy, lysosomes are heavily utilized via their fusion with autophagosomes and must be repopulated to maintain autophagic degradative capacity. During starvation-induced autophagy, lysosomes are generated via de novo biogenesis under the control of TFEB (transcription factor EB), or by the recycling of autolysosome membranes via autophagic lysosome reformation (ALR). However, these lysosome repopulation processes do not operate under nutrient-rich conditions. In our recent study, we identify a sequential phosphoinositide conversion pathway that enables lysosome repopulation under nutrient-rich conditions to facilitate basal autophagy. Phosphatidylinositol-3,4-bisphosphate (PtdIns[3,4]P2) signals generated downstream of phosphoinositide 3-kinase alpha (PI3Kα) during growth factor stimulation are converted to phosphatidylinositol-3-phosphate (PtdIns3P) on endosomes by INPP4B (inositol polyphosphate-4-phosphatase type II B). We show that PtdIns3P is retained as endosomes mature into endolysosomes, and serves as a substrate for PIKFYVE (phosphoinositide kinase, FYVE-type zinc finger containing) to generate phosphatidylinositol-3,5-bisphosphate (PtdIns[3,5]P2) to promote SNX2-dependent lysosome reformation, basal autophagic flux and protein aggregate degradation. Therefore, endosome maturation couples nutrient signaling to lysosome repopulation during basal autophagy by delivering PI3Kα-derived PtdIns3P to endolysosomes for PtdIns(3,5)P2-dependent lysosome reformation.
    Keywords:  INPP4B; PI3Kα; PIKFYVE; endosome; lysosome; phosphoinositides; proteostasis
    DOI:  https://doi.org/10.1080/15548627.2022.2124499
  8. FEBS J. 2022 Sep 16.
    TRIM27 is an autophagy substrate facilitating mitochondria clustering and mitophagy via phosphorylated TBK1.
    Juncal Garcia-Garcia, Anne Kristin McLaren Berge, Katrine Stange Overå, Kenneth Bowitz Larsen, Zambarlal Bhujabal, Andreas Brech, Yakubu Princely Abudu, Trond Lamark, Terje Johansen, Eva Sjøttem.
      Tripartite motif containing protein 27 (TRIM27/also called RFP) is a multifunctional ubiquitin E3 ligase involved in numerous cellular functions, such as proliferation, apoptosis, regulation of the NF-kB pathway, endosomal recycling, and the innate immune response. TRIM27 interacts directly with TANK-binding kinase 1 (TBK1) and regulates its stability. TBK1 in complex with autophagy receptors are recruited to ubiquitin chains assembled on the mitochondrial outer membrane promoting mitophagy. Here we identify TRIM27 as an autophagy substrate, depending on ATG7, ATG9 and autophagy receptors for its lysosomal degradation. We show that TRIM27 forms ubiquitylated cytoplasmic bodies that colocalize with autophagy receptors. Surprisingly, we observed that induced expression of EGFP-TRIM27 in HEK293 FlpIn TRIM27 Knock-Out cells mediates mitochondrial clustering. TRIM27 interacts with autophagy receptor SQSTM1/p62, and the TRIM27 mediated mitochondrial clustering is facilitated by SQSTM/p62. We show that phosphorylated TBK1 is recruited to the clustered mitochondria. Moreover, induced mitophagy activity is reduced in HEK293 FlpIn TRIM27 Knock Out cells, while re-introduction of EGFP-TRIM27 completely restores the mitophagy activity. Inhibition of TBK1 reduces mitophagy in HEK293 FlpIn cells and in the reconstituted EGFP-TRIM27 expressing cells, but not in HEK293 FlpIn TRIM27 Knock Out cells. Altogether, these data reveal novel roles for TRIM27 in mitophagy, facilitating mitochondrial clustering via SQSTM1/p62 and mitophagy via stabilization of phosphorylated TBK1 on mitochondria.
    DOI:  https://doi.org/10.1111/febs.16628
  9. Cell Rep. 2022 Sep 13. pii: S2211-1247(22)01177-9. [Epub ahead of print]40(11): 111349
    Identification of CUL4A-DDB1-WDFY1 as an E3 ubiquitin ligase complex involved in initiation of lysophagy.
    Hirofumi Teranishi, Keisuke Tabata, Marika Saeki, Tetsuo Umemoto, Tomohisa Hatta, Takanobu Otomo, Kentaro Yamamoto, Toru Natsume, Tamotsu Yoshimori, Maho Hamasaki.
      Macroautophagy is a bulk degradation system in which double membrane-bound structures called autophagosomes to deliver cytosolic materials to lysosomes. Autophagy promotes cellular homeostasis by selectively recognizing and sequestering specific targets, such as damaged organelles, protein aggregates, and invading bacteria, termed selective autophagy. We previously reported a type of selective autophagy, lysophagy, which helps clear damaged lysosomes. Damaged lysosomes become ubiquitinated and recruit autophagic machinery. Proteomic studies using transfection reagent-coated beads and further evaluations reveal that a CUL4A-DDB1-WDFY1 E3 ubiquitin ligase complex is essential to initiate lysophagy and clear damaged lysosomes. Moreover, we show that LAMP2 is ubiquitinated by the CUL4A E3 ligase complex as a substrate on damaged lysosomes. These results reveal how cells selectively tag damaged lysosomes to initiate autophagy for the clearance of lysosomes.
    Keywords:  CP: Molecular biology; CUL4A; LAMP2; autophagy; lysophagy; lysosomal membrane damage; ubiquitination
    DOI:  https://doi.org/10.1016/j.celrep.2022.111349
  10. Adv Protein Chem Struct Biol. 2022 ;pii: S1876-1623(22)00056-6. [Epub ahead of print]132 175-197
    Protein expression in exocrine pancreatic diseases. Focus on VMP1 mediated autophagy.
    Maria I Vaccaro, Florencia Mitchell, Francisco Rivera, Claudio D Gonzalez.
      The exocrine pancreas produces enzymes involved in the digestive process whereas endocrine pancreas mainly regulates glucose metabolism. Diseases of the exocrine pancreas are characterized by high morbidity and mortality. Acute pancreatitis is a painful disease in which pancreatic secretory proteins are prematurely activated causing the digestion of the gland. Pancreatic adenocarcinoma is one of the most malignant cancers due to its resistance to treatment, its late diagnosis and high capacity for metastasis. Autophagy is a catabolic process that aims at degrading cytoplasmic contents and damaged organelles, to preserve cell viability and homeostasis. VMP1 is a transmembrane protein that plays a key role in triggering autophagy and being part of the autophagosome membrane. A specific type of selective autophagy pathway called zymophagy protects the pancreas against self-digestion in the setting of acute pancreatitis by sequestering intracellularly activated zymogen granules. Mitophagy is also responsible for maintaining pancreatitis as a mild disease by preserving mitochondrial function. Dysregulation of these selective autophagic processes by pancreatitis itself constitutes a risk factor for development of severe disease. In pancreatic adenocarcinoma, VMP1 mediated autophagy promotes cancer cell survival and resistance to chemotherapy. Therefore, it is relevant to highlight a role for controlling VMP1 expression and targeting VMP1 molecular pathways to improve exocrine pancreatic diseases prognosis.
    Keywords:  Acute pancreatitis; Autophagy; Chemotherapy; Chronic pancreatitis; Kras; Mitophagy; Pancreatic cancer; VMP1; Zymophagy
    DOI:  https://doi.org/10.1016/bs.apcsb.2022.07.001
  11. Autophagy. 2022 Sep 12.
    The hallmark domain of the oldest autophagy receptor family is a cargo-binding module.
    Zhao-Qian Pan, Keqiong Ye, Li-Lin Du.
      In selective macroautophagy/autophagy, autophagy receptors are key molecules that determine cargo specificity. Most known autophagy receptors only exist in some but not all eukaryotic lineages. The exception is Nbr1 proteins, which are conserved across eukaryotes. The four-tryptophan (FW) domain is the hallmark of Nbr1 proteins, but its function has been unknown. Our recent study found that the FW domain in the Nbr1 protein of the filamentous fungus Chaetomium thermophilum binds the α-mannosidase Ams1, a known selective autophagy cargo in budding yeast and fission yeast. Furthermore, we showed that when C. thermophilum Nbr1 and Ams1 are expressed heterologously in fission yeast, FW domain-mediated binding can promote autophagic delivery of Ams1 into vacuoles. We solved the structure of the FW-Ams1 complex and revealed the structural mechanism underlying Ams1 recognition by the FW domain. The N-terminal di-glycine peptide of Ams1 fits into a conserved pocket of the FW domain. We propose that this cargo-binding mechanism may also be employed by Nbr1 proteins in other eukaryotes.
    Keywords:  Ams1; FW domain; Nbr1; autophagy receptor; selective autophagy
    DOI:  https://doi.org/10.1080/15548627.2022.2123636
  12. EMBO Rep. 2022 Sep 14. e54993
    The RNA polymerase II subunit Rpb9 activates ATG1 transcription and autophagy.
    Ting Huang, Gaoyue Jiang, Yabin Zhang, Yuqing Lei, Shiyan Liu, Huihui Li, Kefeng Lu.
      Macroautophagy/autophagy is a conserved process in eukaryotic cells that mediates the degradation and recycling of intracellular substrates. Proteins encoded by autophagy-related (ATG) genes are essentially involved in the autophagy process and must be tightly regulated in response to various circumstances, such as nutrient-rich and starvation conditions. However, crucial transcriptional activators of ATG genes have remained obscure. Here, we identify the RNA polymerase II subunit Rpb9 as an essential regulator of autophagy by a high-throughput screen of a Saccharomyces cerevisiae gene knockout library. Rpb9 plays a crucial and specific role in upregulating ATG1 transcription, and its deficiency decreases autophagic activities. Rpb9 promotes ATG1 transcription by binding to its promoter region, which is mediated by Gcn4. Furthermore, the function of Rpb9 in autophagy and its regulation of ATG1/ULK1 transcription are conserved in mammalian cells. Together, our results indicate that Rpb9 specifically activates ATG1 transcription and thus positively regulates the autophagy process.
    Keywords:  ATG1; Rpb9; autophagy; starvation; transcription
    DOI:  https://doi.org/10.15252/embr.202254993
  13. Front Cell Infect Microbiol. 2022 ;12 902428
    Modulation of autophagy as a therapeutic strategy for Toxoplasma gondii infection.
    Ao Cheng, Huanan Zhang, Baike Chen, Shengyao Zheng, Hongyi Wang, Yijia Shi, Siyao You, Ming Li, Liping Jiang.
      Toxoplasma gondii infection is a severe health threat that endangers billions of people worldwide. T. gondii utilizes the host cell membrane to form a parasitophorous vacuole (PV), thereby fully isolating itself from the host cell cytoplasm and making intracellular clearance difficult. PV can be targeted and destroyed by autophagy. Autophagic targeting results in T. gondii killing via the fusion of autophagosomes and lysosomes. However, T. gondii has developed many strategies to suppress autophagic targeting. Accordingly, the interplay between host cell autophagy and T. gondii is an emerging area with important practical implications. By promoting the canonical autophagy pathway or attenuating the suppression of autophagic targeting, autophagy can be effectively utilized in the development of novel therapeutic strategies against T gondii. Here, we have illustrated the complex interplay between host cell mediated autophagy and T. gondii. Different strategies to promote autophagy in order to target the parasite have been elucidated. Besides, we have analyzed some potential new drug molecules from the DrugBank database using bioinformatics tools, which can modulate autophagy. Various challenges and opportunities focusing autophagy mediated T. gondii clearance have been discussed, which will provide new insights for the development of novel drugs against the parasite.
    Keywords:  AMPK; CD40; IFN-γ; Toxoplasma gondii; autophagy; mTOR
    DOI:  https://doi.org/10.3389/fcimb.2022.902428
  14. Front Cell Dev Biol. 2022 ;10 896893
    Autophagy regulated by the HIF/REDD1/mTORC1 signaling is progressively increased during erythroid differentiation under hypoxia.
    Jian Li, Cheng Quan, Yun-Ling He, Yan Cao, Ying Chen, Yu-Fei Wang, Li-Ying Wu.
      For hematopoietic stem and progenitor cells (HSPCs), hypoxia is a specific microenvironment known as the hypoxic niche. How hypoxia regulates erythroid differentiation of HSPCs remains unclear. In this study, we show that hypoxia evidently accelerates erythroid differentiation, and autophagy plays a pivotal role in this process. We further determine that mTORC1 signaling is suppressed by hypoxia to relieve its inhibition of autophagy, and with the process of erythroid differentiation, mTORC1 activity gradually decreases and autophagy activity increases accordingly. Moreover, we provide evidence that the HIF-1 target gene REDD1 is upregulated to suppress mTORC1 signaling and enhance autophagy, thereby promoting erythroid differentiation under hypoxia. Together, our study identifies that the enhanced autophagy by hypoxia favors erythroid maturation and elucidates a new regulatory pattern whereby autophagy is progressively increased during erythroid differentiation, which is driven by the HIF-1/REDD1/mTORC1 signaling in a hypoxic niche.
    Keywords:  HIF-1; HSPCs; K562; REDD1; autophagy; erythroid differentiation; hypoxia
    DOI:  https://doi.org/10.3389/fcell.2022.896893
  15. Dev Cell. 2022 Sep 07. pii: S1534-5807(22)00597-4. [Epub ahead of print]
    RIPOR2-mediated autophagy dysfunction is critical for aminoglycoside-induced hearing loss.
    Jinan Li, Chang Liu, Ulrich Müller, Bo Zhao.
      Aminoglycosides (AGs) are potent antibiotics that are capable of treating a wide variety of life-threatening infections; however, they are ototoxic and cause irreversible damage to cochlear hair cells. Despite substantial progress, little is known about the molecular pathways critical for hair cell function and survival that are affected by AG exposure. We demonstrate here that gentamicin, a representative AG antibiotic, binds to and within minutes triggers translocation of RIPOR2 in murine hair cells from stereocilia to the pericuticular area. Then, by interacting with a central autophagy component, GABARAP, RIPOR2 affects autophagy activation. Reducing the expression of RIPOR2 or GABARAP completely prevents AG-induced hair cell death and subsequent hearing loss in mice. Additionally, abolishing the expression of PINK1 or Parkin, two key mitochondrial autophagy proteins, prevents hair cell death and subsequent hearing loss caused by AG. In summary, our study demonstrates that RIPOR2-mediated autophagic dysfunction is essential for AG-induced hearing loss.
    Keywords:  GABARAP; RIPOR2; aminoglycoside; autophagy; hair cell; hearing loss; mitophagy; ototoxicity
    DOI:  https://doi.org/10.1016/j.devcel.2022.08.011
  16. Front Cell Dev Biol. 2022 ;10 956394
    Mitochondrial autophagy in the sleeping brain.
    Sofia Mauri, Mariavittoria Favaro, Greta Bernardo, Gabriella M Mazzotta, Elena Ziviani.
      A significant percentage of the mitochondrial mass is replaced on a daily basis via mechanisms of mitochondrial quality control. Through mitophagy (a selective type of autophagy that promotes mitochondrial proteostasis) cells keep a healthy pool of mitochondria, and prevent oxidative stress and inflammation. Furthermore, mitophagy helps adapting to the metabolic demand of the cells, which changes on a daily basis. Core components of the mitophagy process are PINK1 and Parkin, which mutations are linked to Parkinson's Disease. The crucial role of PINK1/Parkin pathway during stress-induced mitophagy has been extensively studied in vitro in different cell types. However, recent advances in the field allowed discovering that mitophagy seems to be only slightly affected in PINK1 KO mice and flies, putting into question the physiological relevance of this pathway in vivo in the whole organism. Indeed, several cell-specific PINK1/Parkin-independent mitophagy pathways have been recently discovered, which appear to be activated under physiological conditions such as those that promote mitochondrial proteome remodeling during differentiation or in response to specific physiological stimuli. In this Mini Review we want to summarize the recent advances in the field, and add another level of complexity by focusing attention on a potentially important aspect of mitophagy regulation: the implication of the circadian clock. Recent works showed that the circadian clock controls many aspects of mitochondrial physiology, including mitochondrial morphology and dynamic, respiratory activity, and ATP synthesis. Furthermore, one of the essential functions of sleep, which is controlled by the clock, is the clearance of toxic metabolic compounds from the brain, including ROS, via mechanisms of proteostasis. Very little is known about a potential role of the clock in the quality control mechanisms that maintain the mitochondrial repertoire healthy during sleep/wake cycles. More importantly, it remains completely unexplored whether (dys)function of mitochondrial proteostasis feedbacks to the circadian clockwork.
    Keywords:  Parkinson’s disease; animal models; circadian rhythms; mitophagy; proteostasis
    DOI:  https://doi.org/10.3389/fcell.2022.956394
  17. Front Genet. 2022 ;13 970699
    Recent advances of the mammalian target of rapamycin signaling in mesenchymal stem cells.
    Huarui Cai, Zhongze Wang, Wenhan Tang, Xiaoxue Ke, Erhu Zhao.
      Mammalian target of rapamycin (mTOR) is a serine/threonine kinase involved in a variety of cellular functions, such as cell proliferation, metabolism, autophagy, survival and cytoskeletal organization. Furthermore, mTOR is made up of three multisubunit complexes, mTOR complex 1, mTOR complex 2, and putative mTOR complex 3. In recent years, increasing evidence has suggested that mTOR plays important roles in the differentiation and immune responses of mesenchymal stem cells (MSCs). In addition, mTOR is a vital regulator of pivotal cellular and physiological functions, such as cell metabolism, survival and ageing, where it has emerged as a novel therapeutic target for ageing-related diseases. Therefore, the mTOR signaling may develop a large impact on the treatment of ageing-related diseases with MSCs. In this review, we discuss prospects for future research in this field.
    Keywords:  ageing-related diseases; differentiation; immune response; mTOR; mesenchymal stem cells; therapeutic target
    DOI:  https://doi.org/10.3389/fgene.2022.970699
  18. J Cell Biol. 2022 Oct 03. pii: e202209004. [Epub ahead of print]221(10):
    NCOA4: More than a receptor for ferritinophagy.
    Zheng Wang, Hong Zhang.
      Liquid-liquid phase separation (LLPS) triages protein cargoes for autophagic degradation. In this issue, Ohshima et al. (2022. J. Cell Biol.https://doi.org/10.1083/jcb.202203102) demonstrate that the autophagy receptor NCOA4 interacts with ferritin particles to form liquid-like condensates via LLPS. The NCOA4-ferritin condensates are delivered to lysosomes for degradation via either canonical macroautophagy or endosomal microautophagy to maintain intracellular iron homeostasis.
    DOI:  https://doi.org/10.1083/jcb.202209004
  19. Autophagy. 2022 Sep 12.
    Conjugation of the ubiquitin family proteins to phospholipids.
    Jun-Ichi Sakamaki, Noboru Mizushima.
      Conjugation of Atg8-family proteins to phosphatidylethanolamine (PE) is important for autophagosome formation. PE conjugation has been thought to be specific to Atg8 among the ubiquitin-family proteins. However, this dogma has not been experimentally verified. Our recent study revealed that ubiquitin is also conjugated to PE on endosomes and the vacuole (or lysosomes). Other ubiquitin-like proteins, such as NEDD8 and ISG15, also covalently bind to phospholipids. We propose that conjugation to phospholipids could be a common feature of the ubiquitin family.
    Keywords:  Atg8; Doa4; Tul1; endosome; lysosome; phosphatidylethanolamine; phospholipids; ubiquitin; ubiquitin-like proteins; vacuole
    DOI:  https://doi.org/10.1080/15548627.2022.2123637
  20. Cancer Cell Int. 2022 Sep 15. 22(1): 284
    Recent advances and limitations of mTOR inhibitors in the treatment of cancer.
    Eunus S Ali, Kangkana Mitra, Shamima Akter, Sarker Ramproshad, Banani Mondal, Ishaq N Khan, Muhammad Torequl Islam, Javad Sharifi-Rad, Daniela Calina, William C Cho.
      The PI3K-Akt-mechanistic (formerly mammalian) target of the rapamycin (mTOR) signaling pathway is important in a variety of biological activities, including cellular proliferation, survival, metabolism, autophagy, and immunity. Abnormal PI3K-Akt-mTOR signalling activation can promote transformation by creating a cellular environment conducive to it. Deregulation of such a system in terms of genetic mutations and amplification has been related to several human cancers. Consequently, mTOR has been recognized as a key target for the treatment of cancer, especially for treating cancers with elevated mTOR signaling due to genetic or metabolic disorders. In vitro and in vivo, rapamycin which is an immunosuppressant agent actively suppresses the activity of mTOR and reduces cancer cell growth. As a result, various sirolimus-derived compounds have now been established as therapies for cancer, and now these medications are being investigated in clinical studies. In this updated review, we discuss the usage of sirolimus-derived compounds and other drugs in several preclinical or clinical studies as well as explain some of the challenges involved in targeting mTOR for treating various human cancers.
    Keywords:  Cancer; Rapamycin; Targeted therapy; mTOR inhibitors; mTOR pathway; mTORC1; mTORC2
    DOI:  https://doi.org/10.1186/s12935-022-02706-8
  21. Autophagy. 2022 Sep 10. 1-16
    A plant virus hijacks phosphatidylinositol-3,5-bisphosphate to escape autophagic degradation in its insect vector.
    Haitao Wang, Jianhua Zhang, Haoqiu Liu, Man Wang, Yan Dong, Yijun Zhou, Sek-Man Wong, Kai Xu, Qiufang Xu.
      Hosts can initiate macroautophagy/autophagy as an antiviral defense response, while viruses have developed multiple ways to evade the host autophagic degradation. However, little is known as to whether viruses can target lipids to subvert autophagic degradation. Here, we show that a low abundant signaling lipid, phosphatidylinositol 3,5-bisphosphate (PtdIns(3,5)P2), is required for rice black-streaked dwarf virus (RBSDV) to evade the autophagic degradation in the insect vector Laodelphax striatellus. RBSDV binds to PtdIns(3,5)P2 and elevates its level through its main capsid protein P10, leading to inhibited autophagy and promoted virus propagation. Furthermore, we show that PtdIns(3,5)P2 inhibits the autophagy pathway by preventing the fusion of autophagosomes and lysosomes through activation of Trpml (transient receptor potential cation channel, mucolipin), an effector of PtdIns(3,5)P2. These findings uncover a strategy whereby a plant virus hijacks PtdIns(3,5)P2 via its viral capsid protein to evade autophagic degradation and promote its survival in insects.
    Keywords:  Autophagy; PtdIns(3,5)P2; RBSDV; lysosome-autophagosome fusion; trpml
    DOI:  https://doi.org/10.1080/15548627.2022.2116676
  22. Antioxid Redox Signal. 2022 Sep 16.
    Calcium homeostasis in the control of mitophagy.
    Mariasole Perrone, Simone Patergnani, Tommaso Di Mambro, Laura Palumbo, Mariusz Więckowski, Carlotta Giorgi, Paolo Pinton.
      SIGNIFICANCE: Maintenance of mitochondrial quality is essential for cellular homeostasis. Among processes responsible for preserving healthy mitochondria, mitophagy selectively eliminates dysfunctional mitochondria by targeting them to the autophagosome for degradation. Alterations in mitophagy lead to the accumulation of damaged mitochondria, which plays an essential role in several diseases like carcinogenesis and tumor progression, neurodegenerative disorders, and autoimmune and cardiovascular pathologies.RECENT ADVANCES: Calcium (Ca2+) plays a fundamental role in cell life, modulating several pathways, such as gene expression, proliferation, differentiation, metabolism, cell death, and survival. Indeed, because it is involved in all these events, Ca2+ is the most versatile intracellular second messenger. Being a process that limits cellular degeneration, mitophagy participates in cellular fate decisions. Several mitochondrial parameters, such as membrane potential, structure, and reactive oxygen species, can trigger the activation of mitophagic machinery. These parameters regulate not only mitophagy but also the mitochondrial Ca2+ uptake.
    CRITICAL ISSUES: Ca2+ handling is fundamental in regulating ATP production by mitochondria and mitochondrial quality control processes. Despite the growing literature about the link between Ca2+ and mitophagy, the mechanism by which Ca2+ homeostasis regulates mitophagy is still debated.
    FUTURE DIRECTIONS: Several studies have revealed that excessive mitophagy together with altered mitochondrial Ca2+ uptake leads to different dysfunctions in numerous diseases. Thus, therapeutic modulation of these pathways is considered promising treatments.
    DOI:  https://doi.org/10.1089/ars.2022.0122
  23. Front Pharmacol. 2022 ;13 963920
    Autophagy in renal fibrosis: Protection or promotion?
    Rong Dai, Lei Zhang, Hua Jin, Dong Wang, Meng Cheng, Tian Sang, Chuyi Peng, Yue Li, Yiping Wang.
      Autophagy is a process that degrades endogenous cellular protein aggregates and damaged organelles via the lysosomal pathway to maintain cellular homeostasis and energy production. Baseline autophagy in the kidney, which serves as a quality control system, is essential for cellular metabolism and organelle homeostasis. Renal fibrosis is the ultimate pathological manifestation of progressive chronic kidney disease. In several experimental models of renal fibrosis, different time points, stimulus intensities, factors, and molecular mechanisms mediating the upregulation or downregulation of autophagy may have different effects on renal fibrosis. Autophagy occurring in a single lesion may also exert several distinct biological effects on renal fibrosis. Thus, whether autophagy prevents or facilitates renal fibrosis remains a complex and challenging question. This review explores the different effects of the dual regulatory function of autophagy on renal fibrosis in different renal fibrosis models, providing ideas for future work in related basic and clinical research.
    Keywords:  autophagy; dual regulation; experimental model; fibrosis; renal fibrosis
    DOI:  https://doi.org/10.3389/fphar.2022.963920
  24. FEBS Lett. 2022 Sep;596(17): 2089-2092
    Plant autophagy and intracellular trafficking.
    Diane C Bassham.
      
    DOI:  https://doi.org/10.1002/1873-3468.14466
  25. ACS Med Chem Lett. 2022 Sep 08. 13(9): 1510-1516
    Novel Bcl-2 Inhibitors Selectively Disrupt the Autophagy-Specific Bcl-2-Beclin 1 Protein-Protein Interaction.
    Xiaonan Dong, Qiren Liang, Yun-Zu Pan, Xiaoyu Wang, Yi-Chun Kuo, Wei-Chung Chiang, Xuewu Zhang, Noelle S Williams, Josep Rizo, Beth Levine, Jef K De Brabander.
      Autophagy plays essential roles in a wide variety of physiological processes, such as cellular homeostasis, metabolism, development, differentiation, and immunity. Selective pharmacological modulation of autophagy is considered a valuable potential therapeutic approach to treat diverse human diseases. However, development of such therapies has been greatly impeded by the lack of specific small molecule autophagy modulators. Here, we performed structure-activity relationship studies on a previously discovered weak Bcl-2 inhibitor SW076956, and developed a panel of small molecule compounds that selectively released Bcl-2-mediated inhibition of autophagy-related Beclin 1 compared to apoptosis-related Bax at nanomolar concentration. Our NMR analysis showed that compound 35 directly binds Bcl-2 and specifically inhibits the interaction between the Bcl-2 and Beclin 1 BH3 domains without disruption of the Bcl-2-Bax BH3 interaction. More broadly, this proof-of-concept study demonstrates that targeting protein-protein interactions of the intrinsic autophagy regulatory network can serve as a valuable strategy for the development of autophagy-based therapeutics.
    DOI:  https://doi.org/10.1021/acsmedchemlett.2c00309
  26. Front Aging Neurosci. 2022 ;14 933979
    Slingshot homolog-1 mediates the secretion of small extracellular vesicles containing misfolded proteins by regulating autophagy cargo receptors and actin dynamics.
    Sara Cazzaro, Cenxiao Fang, Hirah Khan, Richard Witas, Teresa R Kee, Jung-A A Woo, David E Kang.
      Increasing evidence indicates that the accumulation misfolded proteins in Alzheimer's disease (AD) arises from clearance defects in the autophagy-lysosome pathway. Misfolded proteins such as Aβ and tau are secreted in small extracellular vesicles (i.e., exosomes) and are propagated from cell to cell in part through secreted small extracellular vesicles (sEVs). Recent studies suggest that autophagic activity and exosome secretion are coregulated events, and multiple autophagy-related proteins are found in sEVs, including the cargo receptors Sqstm1/p62 and optineurin. However, whether and how autophagy cargo receptors per se regulate the secretion of sEVs is unknown. Moreover, despite the prominent role of actin dynamics in secretory vesicle release, its role in EV secretion is unknown. In this study, we leveraged the dual axes of Slingshot Homolog-1 (SSH1), which inhibits Sqstm1/p62-mediated autophagy and activates cofilin-mediated actin dynamics, to study the regulation of sEV secretion. Here we show that cargo receptors Sqstm1/p62 and optineurin inhibit sEV secretion, an activity that requires their ability to bind ubiquitinated cargo. Conversely, SSH1 increases sEV secretion by dephosphorylating Sqstm1/p62 at pSer403, the phospho-residue that allows Sqstm1/p62 to bind ubiquitinated cargo. In addition, increasing actin dynamics through the SSH1-cofilin activation pathway also increases sEV secretion, which is mimicked by latrunculin B treatment. Finally, Aβ42 oligomers and mutant tau increase sEV secretion and are physically associated with secreted sEVs. These findings suggest that increasing cargo receptor engagement with autophagic cargo and reducing actin dynamics (i.e., SSH1 inhibition) represents an attractive strategy to promote misfolded protein degradation while reducing sEV-mediated cell to cell spread of pathology.
    Keywords:  SSH1; actin dynamics; amyloid; autophagy; exosome; optineurin; p62; tau
    DOI:  https://doi.org/10.3389/fnagi.2022.933979
  27. J Am Chem Soc. 2022 Sep 15.
    Targeting the ATG5-ATG16L1 Protein-Protein Interaction with a Hydrocarbon-Stapled Peptide Derived from ATG16L1 for Autophagy Inhibition.
    Jin Cui, Yuta Ogasawara, Ikuko Kurata, Kazuaki Matoba, Yuko Fujioka, Nobuo N Noda, Masakatsu Shibasaki, Takumi Watanabe.
      Selective modulation of autophagy is a promising therapeutic strategy, especially for cancer treatment. However, the lack of specific autophagy inhibitors limits this strategy. The formation of the ATG12-ATG5-ATG16L1 complex is essential for targeting the ATG12-ATG5 conjugate to proper membranes and to generate LC3-II for the progression of autophagy. Thus, targeting ATG5-ATG16L1 protein-protein interactions (PPIs) might inhibit early stage autophagy with high specificity. In this paper, we report that a stapled peptide derived from ATG16L1 exhibits potent binding affinity to ATG5, striking resistance to proteolysis, and significant autophagy inhibition activities in cells.
    DOI:  https://doi.org/10.1021/jacs.2c07648
  28. BMC Immunol. 2022 Sep 14. 23(1): 43
    Reversing BCG-mediated autophagy inhibition and mycobacterial survival to improve vaccine efficacy.
    Maria Gonzalez-Orozco, Emily J Strong, Ruchi Paroha, Sunhee Lee.
      BACKGROUND: Autophagy is an important mechanism for promoting Mycobacterium clearance from macrophages. Pathogenic and non-pathogenic mycobacterium can activate the mTOR pathway while simultaneously inducing autophagy. M. tuberculosis and M. bovis BCG inhibit autophagy and favor intracellular bacteria survival.RESULTS: We observed that pre-infection of live or heat-killed BCG could prevent autophagy induced by pharmacological activators or M. smegmatis, a strong autophagy-inducing mycobacterium. BCG-derived lipids are responsible for autophagy inhibition. However, post-infection with BCG could not stop the autophagy initiated by M. smegmatis, which increases further autophagy induction and mycobacteria clearance. Coinfection with BCG and heat killed M. smegmatis enhanced antigen specific CD4+ T cell responses and reduced mycobacterial survival.
    CONCLUSION: These results suggest that autophagy-inducing M. smegmatis could be used to promote better innate and consequential adaptive immune responses, improving BCG vaccine efficacy.
    Keywords:  Autophagy; BCG; Innate immunity; Lipids; Mycobacterium smegmatis; Mycobacterium tuberculosis
    DOI:  https://doi.org/10.1186/s12865-022-00518-z
  29. Mol Biol Rep. 2022 Sep 15.
    Protective and deleterious effects of autophagy in the setting of myocardial ischemia/reperfusion injury: an overview.
    Behnaz Mokhtari, Reza Badalzadeh.
      INTRODUCTION:  Autophagy is known as a conserved mechanism in order to preserve cell survival under various stress conditions via maintaining cellular homeostasis. Although autophagy is active in the heart at baseline and plays a critical role in cell survival, inappropriate activation of autophagy following ischemia/reperfusion (I/R) injury leads to cell death.MAIN TEXT: The distinct functions of autophagy in myocardial I/R injury condition have been examined in numerous studies, however, contradicting results have been achieved in this field. These studies have documented that autophagy acts as a double-edged sword in myocardial I/R injury. Clarifying the exact role of autophagy in determining the health or death of cardiomyocytes under I/R injury is very helpful to achieve better cardioprotection in prospective clinical studies. Thus, autophagy may be an interesting target for the treatment or prevention of myocardial I/R injury. But before considering this matter, it is necessary to address the gaps in our knowledge about the complex role of autophagy in myocardial I/R injury.
    CONCLUSION: In this review, by providing updated data about the role of autophagy in the heart during ischemia and reperfusion, we tried to provide more insights in this context and encourage scientists to pay special attention towards manipulating autophagy as an intriguing and powerful approach in cardioprotection.
    Keywords:  Cardioprotection; Deleterious autophagy; Myocardial ischemia; Myocardial reperfusion; Protective autophagy
    DOI:  https://doi.org/10.1007/s11033-022-07837-9
  30. iScience. 2022 Sep 16. 25(9): 105029
    MCM7 supports the stemness of bladder cancer stem-like cells by enhancing autophagic flux.
    Lijun Mo, Bijia Su, Lili Xu, Zhiming Hu, Hongwei Li, Hongyan Du, Jinlong Li.
      Autophagy plays critical roles in the pluripotent stemness of cancer stem cells (CSCs). However, how CSCs maintain the elevated autophagy to support stemness remains elusive. Here, we demonstrate that bladder cancer stem-like cells (BCSLCs) are at slow-cycling state with enhanced autophagy and mitophagy. In these slow-cycling BCSLCs, the DNA replication initiator MCM7 is required for autophagy and stemness. MCM7 knockdown inhibits autophagic flux and reduces the stemness of BCSLCs. MCM7 can facilitate autolysosome formation through binding with dynein to promote autophagic flux. The enhanced autophagy/mitophagy helps BCSLCs to maintain mitochondrial respiration, thus inhibiting AMPK activation. AMPK activation can trigger switch from autophagy to apoptosis, through increasing BCL2/BECLIN1 interaction and inducing P53 accumulation. In summary, we find that MCM7 can promote autophagic flux to support.
    Keywords:  Biological sciences; Cancer; Stem cells research; Systems biology
    DOI:  https://doi.org/10.1016/j.isci.2022.105029
  31. Autophagy. 2022 Sep 12.
    Apicoplast biogenesis mediated by ATG8 requires the ATG12-ATG5-ATG16L and SNAP29 complexes in Toxoplasma gondii.
    Jiawen Fu, Lin Zhao, Yu Pang, Heming Chen, Hayashi Yamamoto, Yuntong Chen, Zhaoran Li, Noboru Mizushima, Honglin Jia.
      In apicomplexan parasites, the macroautophagy/autophagy machinery is repurposed to maintain the plastid-like organelle apicoplast. Previously, we showed that in Toxoplasma and Plasmodium, ATG12 interacts with ATG5 in a non-covalent manner, in contrast to the covalent interaction in most organisms. However, it remained unknown whether apicomplexan parasites have functional orthologs of ATG16L1, a protein that is essential for the function of the covalent ATG12-ATG5 complex in vivo in other organisms. Furthermore, the mechanism used by the autophagy machinery to maintain the apicoplast is unclear. We report that the ATG12-ATG5-ATG16L complex exists in Toxoplasma gondii (Tg). This complex is localized on isolated structures at the periphery of the apicoplast dependent on TgATG16L. Inducible depletion of TgATG12, TgATG5, or TgATG16L caused loss of the apicoplast and affected parasite growth. We found that a putative soluble N-ethylmaleimide sensitive factor attachment protein receptor (SNARE) protein, synaptosomal-associated protein 29 (TgSNAP29, Qbc SNARE), is required to maintain the apicoplast in T. gondii. TgSNAP29 depletion disrupted TgATG8 localization at the apicoplast. Additionally, we identified a putative ubiquitin-interacting motif-docking site (UDS) of TgATG8. Mutation of the UDS site abolished TgATG8 localization on the apicoplast but not lipidation. These findings suggest that the TgATG12-TgATG5-TgATG16L complex is required for biogenesis of the apicoplast, in which TgATG8 is translocated to the apicoplast via vesicles in a SNARE -dependent manner in T. gondii.
    Keywords:  Apicoplast; T. gondii; TgATG12–TgATG5-TgATG16L; TgATG8; TgSNAP29; ubiquitin-interacting motif
    DOI:  https://doi.org/10.1080/15548627.2022.2123639
  32. Trends Biochem Sci. 2022 Sep 13. pii: S0968-0004(22)00232-8. [Epub ahead of print]
    Physiological media advance cell culture experiments.
    Martin Fischer.
      The metabolism plays a fundamental role in cellular signaling pathways, but commonly used cell culture media do not reflect physiological metabolite concentrations. The metabolic control hub mammalian target of rapamycin complex 1 (mTORC1) kinase is an illuminating example that it is about time to advance our cell culture to become more physiological and relevant.
    Keywords:  RFX7; cancer research; cell culture media; mTOR; metabolism; p53
    DOI:  https://doi.org/10.1016/j.tibs.2022.08.007
  33. Cell. 2022 Sep 09. pii: S0092-8674(22)01114-X. [Epub ahead of print]
    Structure and mechanism of human cystine exporter cystinosin.
    Xue Guo, Philip Schmiege, Tufa E Assafa, Rong Wang, Yan Xu, Linda Donnelly, Michael Fine, Xiaodan Ni, Jiansen Jiang, Glenn Millhauser, Liang Feng, Xiaochun Li.
      Lysosomal amino acid efflux by proton-driven transporters is essential for lysosomal homeostasis, amino acid recycling, mTOR signaling, and maintaining lysosomal pH. To unravel the mechanisms of these transporters, we focus on cystinosin, a prototypical lysosomal amino acid transporter that exports cystine to the cytosol, where its reduction to cysteine supplies this limiting amino acid for diverse fundamental processes and controlling nutrient adaptation. Cystinosin mutations cause cystinosis, a devastating lysosomal storage disease. Here, we present structures of human cystinosin in lumen-open, cytosol-open, and cystine-bound states, which uncover the cystine recognition mechanism and capture the key conformational states of the transport cycle. Our structures, along with functional studies and double electron-electron resonance spectroscopic investigations, reveal the molecular basis for the transporter's conformational transitions and protonation switch, show conformation-dependent Ragulator-Rag complex engagement, and demonstrate an unexpected activation mechanism. These findings provide molecular insights into lysosomal amino acid efflux and a potential therapeutic strategy.
    Keywords:  DEER; Keywords; Ragulator-Rag complex; X-ray crystallography; cryo-EM; cystinosin; cystinosis; fast adaptation; lysosomal storage disease; lysosomal transporter; membrane protein dynamics
    DOI:  https://doi.org/10.1016/j.cell.2022.08.020
  34. Front Pharmacol. 2022 ;13 970596
    Natural phytochemicals that affect autophagy in the treatment of oral diseases and infections: A review.
    Xi Cheng, Qianming Chen, Ping Sun.
      Autophagy is a critical factor in eukaryotic evolution. Cells provide nutrition and energy during autophagy by destroying non-essential components, thereby allowing intracellular material conversion and managing temporary survival stress. Autophagy is linked to a variety of oral disorders, including the type and extent of oral malignancies. Furthermore, autophagy is important in lymphocyte formation, innate immunity, and the regulation of acquired immune responses. It is also required for immunological responses in the oral cavity. Knowledge of autophagy has aided in the identification and treatment of common oral disorders, most notably cancers. The involvement of autophagy in the oral immune system may offer a new understanding of the immune mechanism and provide a novel approach to eliminating harmful bacteria in the body. This review focuses on autophagy creation, innate and acquired immunological responses to autophagy, and the status of autophagy in microbial infection research. Recent developments in the regulatory mechanisms of autophagy and therapeutic applications in oral illnesses, particularly oral cancers, are also discussed. Finally, the relationship between various natural substances that may be used as medications and autophagy is investigated.
    Keywords:  autophagy; cancer; microbial infection; natural substances; oral diseases
    DOI:  https://doi.org/10.3389/fphar.2022.970596
  35. Brain Res. 2022 Sep 08. pii: S0006-8993(22)00303-1. [Epub ahead of print]1795 148079
    Membrane estrogen receptor ERα activation improves tau clearance via autophagy induction in a tauopathy cell model.
    Angelica Jardim Costa, Rafaela Brito Oliveira, Patrícia Wachilewski, Michelle Sayuri Nishino, Taysa Bervian Bassani, Roberta Sessa Stilhano, Janete Maria Cerutti, Bruno Nozima, Catarina Segreti Porto, Gustavo Jose da Silva Pereira, Ana Lopez Ramirez, Soraya Soubhi Smaili, Rodrigo Portes Ureshino.
      Alzheimer's disease (AD) is the most prevalent aging-associated neurodegenerative disease, with a higher incidence in women than men. There is evidence that sex hormone replacement therapy, particularly estrogen, reduces memory loss in menopausal women. Neurofibrillary tangles are associated with tau protein aggregation, a characteristic of AD and other tauopathies. In this sense, autophagy is a promising cellular process to remove these protein aggregates. This study evaluated the autophagy mechanisms involved in neuroprotection induced by 17β-estradiol (E2) in a Tet-On inducible expression tauopathy cell model (EGFP-tau WT or with the P301L mutation, 0N4R isoform). The results indicated that 17β-estradiol induces autophagy by activating AMPK in a concentration-dependent manner, independent of mTOR signals. The estrogen receptor α (ERα) agonist, PPT, also induced autophagy, while the ERα antagonist, MPP, substantially attenuated the 17β-estradiol-mediated autophagy induction. Notably, 17β-estradiol increased LC3-II levels and phosphorylated and total tau protein clearance in the EGFP-tau WT cell line but not in EGPF-tau P301L. Similar results were observed with E2-BSA, a plasma membrane-impermeable estrogen, suggesting membrane ERα involvement in non-genomic estrogenic pathway activation. Furthermore, 17β-estradiol-induced autophagy led to EGFP-tau protein clearance. These results demonstrate that modulating autophagy via the estrogenic pathway may represent a new therapeutic target for treating AD.
    Keywords:  17β-estradiol; Alzheimer's Disease; Autophagy; Neuroprotection; Tauopathy
    DOI:  https://doi.org/10.1016/j.brainres.2022.148079
  36. Autophagy. 2022 Sep 15.
    cAMP-Protein Kinase A and Stress-Activated MAP Kinase signaling mediate transcriptional control of autophagy in fission yeast during glucose limitation or starvation.
    Armando Jesús Pérez-Díaz, Beatriz Vázquez-Marín, Jero Vicente-Soler, Francisco Prieto-Ruiz, Teresa Soto, Alejandro Franco, José Cansado, Marisa Madrid.
      Macroautophagy/autophagy is an essential adaptive physiological response in eukaryotes induced during nutrient starvation, including glucose, the primary immediate carbon and energy source for most cells. Although the molecular mechanisms that induce autophagy during glucose starvation have been extensively explored in the budding yeast Saccharomyces cerevisiae, little is known about how this coping response is regulated in the evolutionary distant fission yeast Schizosaccharomyces pombe. Here, we show that S. pombe autophagy in response to glucose limitation relies on mitochondrial respiration and the electron transport chain (ETC), but, in contrast to S. cerevisiae, the AMP-activated protein kinase (AMPK) and DNA damage response pathway components do not modulate fission yeast autophagic flux under these conditions. In the presence of glucose, the cAMP-protein kinase A (PKA) signaling pathway constitutively represses S. pombe autophagy by downregulating the transcription factor Rst2, which promotes the expression of respiratory genes required for autophagy induction under limited glucose availability. Furthermore, the stress-activated protein kinase (SAPK) signaling pathway, and its central mitogen-activated protein kinase (MAPK) Sty1, positively modulate autophagy upon glucose limitation at the transcriptional level through its downstream effector Atf1 and by direct in vivo phosphorylation of Rst2 at S292. Thus, our data indicate that the signaling pathways that govern autophagy during glucose shortage or starvation have evolved differently in S. pombe and uncover the existence of sophisticated and multifaceted mechanisms that control this self-preservation and survival response.
    Keywords:  Autophagy; MAP kinase; Schizosaccharomyces pombe; cAMP-protein kinase A; fermentation; glucose; respiration; transcription
    DOI:  https://doi.org/10.1080/15548627.2022.2125204
  37. Elife. 2022 Sep 13. pii: e80901. [Epub ahead of print]11
    Structure of the HOPS tethering complex, a lysosomal membrane fusion machinery.
    Dmitry Shvarev, Jannis Schoppe, Caroline König, Angela Perz, Nadia Füllbrunn, Stephan Kiontke, Lars Langemeyer, Dovile Januliene, Kilian Schnelle, Daniel Kümmel, Florian Fröhlich, Arne Moeller, Christian Ungermann.
      Lysosomes are essential for cellular recycling, nutrient signaling, autophagy, and pathogenic bacteria and viruses invasion. Lysosomal fusion is fundamental to cell survival and requires HOPS, a conserved heterohexameric tethering complex. On the membranes to be fused, HOPS binds small membrane-associated GTPases and assembles SNAREs for fusion, but how the complex fulfills its function remained speculative. Here, we used cryo-electron microscopy to reveal the structure of HOPS. Unlike previously reported, significant flexibility of HOPS is confined to its extremities, where GTPase binding occurs. The SNARE-binding module is firmly attached to the core, therefore, ideally positioned between the membranes to catalyze fusion. Our data suggest a model for how HOPS fulfills its dual functionality of tethering and fusion and indicate why it is an essential part of the membrane fusion machinery.
    Keywords:  S. cerevisiae; cell biology; molecular biophysics; structural biology
    DOI:  https://doi.org/10.7554/eLife.80901
  38. Am J Transl Res. 2022 ;14(8): 5800-5811
    ER stress induces myocardial dysfunction and cardiac autophagy in Sestrin2 knockout mice.
    Jie Zhang, Linlin Yao, Shaohua Li, Misbahul Ferdous, Peng Zhao.
      OBJECTIVES: Sestrin2 is an essential regulator of the cellular adaptive response against various stresses. The endoplasmic reticulum (ER) is critical in maintaining normal cardiac function by controlling intracellular Ca2+ accumulation, as well as protein folding and processing. Autophagy contributes to stress-associated heart dysfunction. AMP-activated protein kinase (AMPK) is important in energy homeostasis in cardiomyocytes. However, the function of Sestrin2 (Sesn2) in ER stress-induced autophagy that induces myocardial dysfunction has not been clarified. In this study, mice and cardiac tissues were treated with tunicamycin (TN), an inducer of ER stress. We then explored the roles of Sesn2 and the AMPK pathway associated with autophagy in ER stress-induced myocardial dysfunction in mice.METHODS: Echocardiography, contractile function analysis, intracellular Ca2+ status, and immunoblot analysis of AMPK pathway were performed, ER stress and autophagy markers were examined.
    RESULTS: The study revealed that ER stress caused significant heart dysfunction and cardiotoxicity in the mouse heart and cardiomyocytes. Biochemical analysis indicated enhanced cardiac autophagy mediated by ER stress and AMPK/mTOR activation. Sesn2 knockout exacerbated ER stress-related myocardial dysfunction due to the failed response of cardiac autophagy and AMPK/mTOR pathway activation. Further, pharmacological inhibition of AMPK or autophagy worsened TN-induced cardiac dysfunction.
    CONCLUSION: Taken together, loss of the Sesn2 protein exacerbates ER stress-induced cardiac dysfunction through the AMPK/mTOR signaling cascade and loss of autophagy response.
    Keywords:  ER stress; Sestrin2; autophagy; cardiac dysfunction
  39. Neurobiol Dis. 2022 Sep 13. pii: S0969-9961(22)00251-0. [Epub ahead of print] 105859
    Functional and structural consequences of TBK1 missense variants in frontotemporal lobar degeneration and amyotrophic lateral sclerosis.
    Gurfinkel Yuval, Polain Nicole, Sonar Krushna, Nice Penelope, L Mancera Ricardo, Rea Sarah Lyn.
      Mutations in the Tank-binding kinase 1 (TBK1) gene were identified in 2015 in individuals with frontotemporal lobar degeneration (FTLD) and amyotrophic lateral sclerosis (ALS). They account for ~3-4% of cases. To date, over 100 distinct mutations, including missense, nonsense, deletion, insertion, duplication, and splice-site mutations have been reported. While nonsense mutations are predicted to cause disease via haploinsufficiency, the mechanisms underlying disease pathogenesis with missense mutations is not fully elucidated. TBK1 is a kinase involved in neuroinflammation, which is commonly observed in these diseases. TBK1 also phosphorylates key autophagy mediators, thereby regulating proteostasis, a pathway that is dysregulated in ALS-FTLD. Recently, several groups have characterised various missense mutations with respect to their effects on the phosphorylation of known TBK1 substrates, TBK1 homodimerization, interaction with optineurin, and the regulation of autophagy and neuroinflammatory pathways. Further, the effects of either global or conditional heterozygous knock-out of Tbk1, or the heterozygous or homozygous knock-in of ALS-FTLD associated mutations, alone or when crossed with the SOD1G93A classical ALS mouse model or a TDP-43 mouse model, have been reported. In this review we summarise the known functional effects of TBK1 missense mutations. We also present novel modelling data that predicts the structural effects of missense mutations and discuss how they correlate with the known functional effects of these mutations.
    Keywords:  Amyotrophic lateral sclerosis; Autophagy; Frontotemporal lobar degeneration; Mitophagy; Neuroinflammation; OPTN: optineurin; SQSTM1: Sequestosome-1/p62; TBK1: TANK-binding kinase 1
    DOI:  https://doi.org/10.1016/j.nbd.2022.105859
  40. Front Cell Dev Biol. 2022 ;10 987317
    Sensing local energetics to acutely regulate mitophagy in skeletal muscle.
    Anna S Nichenko, Kalyn S Specht, Siobhan M Craige, Joshua C Drake.
      The energetic requirements of skeletal muscle to sustain movement, as during exercise, is met largely by mitochondria, which form an intricate, interconnected reticulum. Maintenance of a healthy mitochondrial reticulum is essential for skeletal muscle function, suggesting quality control pathways are spatially governed. Mitophagy, the process by which damaged and/or dysfunctional regions of the mitochondrial reticulum are removed and degraded, has emerged as an integral part of the molecular response to exercise. Upregulation of mitophagy in response to acute exercise is directly connected to energetic sensing mechanisms through AMPK. In this review, we discuss the connection of mitophagy to muscle energetics and how AMPK may spatially control mitophagy through multiple potential means.
    Keywords:  AMPK; energetic stress; mitochondria; mitophagy; reactive oxygen species
    DOI:  https://doi.org/10.3389/fcell.2022.987317
  41. Proc Natl Acad Sci U S A. 2022 Sep 20. 119(38): e2204083119
    Tissue-restricted inhibition of mTOR using chemical genetics.
    Douglas R Wassarman, Kondalarao Bankapalli, Leo J Pallanck, Kevan M Shokat.
      Mammalian target of rapamycin (mTOR) is a highly conserved eukaryotic protein kinase that coordinates cell growth and metabolism, and plays a critical role in cancer, immunity, and aging. It remains unclear how mTOR signaling in individual tissues contributes to whole-organism processes because mTOR inhibitors, like the natural product rapamycin, are administered systemically and target multiple tissues simultaneously. We developed a chemical-genetic system, termed selecTOR, that restricts the activity of a rapamycin analog to specific cell populations through targeted expression of a mutant FKBP12 protein. This analog has reduced affinity for its obligate binding partner FKBP12, which reduces its ability to inhibit mTOR in wild-type cells and tissues. Expression of the mutant FKBP12, which contains an expanded binding pocket, rescues the activity of this rapamycin analog. Using this system, we show that selective mTOR inhibition can be achieved in Saccharomyces cerevisiae and human cells, and we validate the utility of our system in an intact metazoan model organism by identifying the tissues responsible for a rapamycin-induced developmental delay in Drosophila.
    Keywords:  Drosophila; kinase inhibitor; mTOR; rapamycin; tissue specific
    DOI:  https://doi.org/10.1073/pnas.2204083119
  42. Biochem Biophys Res Commun. 2022 Sep 06. pii: S0006-291X(22)01223-2. [Epub ahead of print]629 26-33
    The core autophagy protein ATG5 controls the polarity of the Golgi apparatus and insulin secretion of pancreatic beta cells.
    Xintong Pei, Huiyu Wang, Pingyong Xu, Kuo Liang, Lin Yuan.
      Pancreatic beta cells are insulin-producing cells that are structurally and functionally polarized in the islets of Langerhans. The organization and position of the Golgi complex play a key role in maintaining a polarized cell state, but the factors and molecular mechanisms determining the Golgi polarization of pancreatic beta cells are still unknown. In the current study, using pancreatic beta cell-specific Atg5 knockout mice, we found that Atg5, an essential gene for autophagy, plays a pivotal role in regulating Golgi integrity and polarization by affecting the expression of genes involved in vesicle transport. Deletion of Atg5 led to endoplasmic reticulum (ER) stress and impaired the distribution of proinsulin and insulin secretion of pancreatic beta cells, which further exacerbates diabetes. These results contribute to a comprehensive understanding of autophagy-mediated Golgi polarization and its regulation of the function of pancreatic beta cells.
    Keywords:  Autophagy; Diabetes; Golgi apparatus; Polarity
    DOI:  https://doi.org/10.1016/j.bbrc.2022.08.084
  43. Biochem Soc Trans. 2022 Sep 16. pii: BST20220778. [Epub ahead of print]
    The role of autophagy-lysosomal pathway in motor neuron diseases.
    Barbara Tedesco, Veronica Ferrari, Marta Cozzi, Marta Chierichetti, Elena Casarotto, Paola Pramaggiore, Francesco Mina, Margherita Piccolella, Riccardo Cristofani, Valeria Crippa, Paola Rusmini, Mariarita Galbiati, Angelo Poletti.
      Motor neuron diseases (MNDs) include a broad group of diseases in which neurodegeneration mainly affects upper and/or lower motor neurons (MNs). Although the involvement of specific MNs, symptoms, age of onset, and progression differ in MNDs, the main pathogenic mechanism common to most MNDs is represented by proteostasis alteration and proteotoxicity. This pathomechanism may be directly related to mutations in genes encoding proteins involved in the protein quality control system, particularly the autophagy-lysosomal pathway (ALP). Alternatively, proteostasis alteration can be caused by aberrant proteins that tend to misfold and to aggregate, two related processes that, over time, cannot be properly handled by the ALP. Here, we summarize the main ALP features, focusing on different routes utilized to deliver substrates to the lysosome and how the various ALP pathways intersect with the intracellular trafficking of membranes and vesicles. Next, we provide an overview of the mutated genes that have been found associated with MNDs, how these gene products are involved in different steps of ALP and related processes. Finally, we discuss how autophagy can be considered a valid therapeutic target for MNDs treatment focusing on traditional autophagy modulators and on emerging approaches to overcome their limitations.
    Keywords:  HSPB8; autophagy; chaperone-assisted selective autophagy; motorneuron diseases; neurodegeneration; trehalose
    DOI:  https://doi.org/10.1042/BST20220778
  44. J Inherit Metab Dis. 2022 Sep 14.
    Aberrant autophagy in lysosomal storage disorders marked by a lysosomal SNARE protein shortage due to suppression of endocytosis.
    Hiroki Tanaka, Daisuke Tsuji, Ryosuke Watanabe, Yukiya Ohnishi, Shindai Kitaguchi, Ryuto Nakae, Hiromi Teramoto, Jun Tsukimoto, Yuto Horii, Kohji Itoh.
      Lysosomal storage disorders (LSDs) are inherited metabolic diseases caused by genetic defects in lysosomal enzymes or related factors. LSDs are associated with excessive accumulation of natural substrates in lysosomes leading to central nervous system and peripheral tissue damage. Abnormal autophagy is also involved in pathogenesis, although the underlying mechanisms remain unclear. We demonstrated that impairment of lysosome-autophagosome fusion is due to suppressed endocytosis in LSDs. The fusion was reduced in several LSD cells and the brains of LSD model mice, suggesting that the completion of autophagy is suppressed by the accumulation of substrates. In this brain, the expression of the soluble N-ethylmaleimide sensitive factor attachment protein receptor (SNARE) proteins, VAMP8 and Syntaxin7, was decreased on the lysosomal surface but not intracellular. This aberrant autophagy preceded the development of pathological phenotypes in LSD-model mice. Furthermore, the enzyme deficiency leading to the substrate accumulation could suppress endocytosis, and the inhibited endocytosis decreased SNARE proteins localized on lysosomes. These findings suggest that the shortage of SNARE proteins on lysosomes is one of the reasons for the impairment of lysosome-autophagosome fusion in LSD cells. This article is protected by copyright. All rights reserved.
    DOI:  https://doi.org/10.1002/jimd.12558
  45. Adv Protein Chem Struct Biol. 2022 ;pii: S1876-1623(22)00050-5. [Epub ahead of print]132 49-87
    The disturbance of protein synthesis/degradation homeostasis is a common trait of age-related neurodegenerative disorders.
    Fabio Di Domenico, Chiara Lanzillotta.
      Protein homeostasis or "proteostasis" represent the process that regulates the balance of the intracellular functional and "healthy" proteins. Proteostasis is fundamental to preserve physiological metabolic processes in the cell and it allow to respond to any given stimulus as the expression of components of the proteostasis network is customized according to the proteomic demands of different cellular environments. In conditions that promote unfolding/misfolding of proteins chaperones act as signaling molecules inducing extreme measures to either fix the problem or destroy unfolded proteins. When the chaperone machinery fails under pathological insults unfolded proteins induce the endoplasmic reticulum (ER) stress activating the unfolded protein response (UPR) machinery. The activation of the UPR restores ER proteostasis primarily through the transcriptional remodeling of ER protein folding, trafficking, and degradation pathways, such as the ubiquitin proteasome system (UPS). If these mechanisms do not manage to clear the aberrant proteins, proteasome overload and become defective, and misfolded proteins may form aggregates thus extending the UPR mechanism. These aggregates are then attempted to be cleared by macroautophagy. Impaired proteostasis promote the accumulation of misfolded proteins that exacerbate the damage to chaperones, surveillance systems and/or degradative activities. Remarkably, the removal of toxic misfolded proteins is critical for all cells, but it is especially significant in neurons since these cannot be readily replaced. In neurons, the maintenance of efficient proteostasis is essential to healthy aging since the dysregulation of the proteostasis network can lead to neurodegenerative disease. Each of these brain pathologies is characterized by the repeated misfolding of one of more peculiar proteins, which evade both the protein folding machinery and cellular degradation mechanisms and begins to form aggregates that nucleate out into large fibrillar aggregates. In this chapter we describe the mechanisms, associated with faulty proteostasis, that promote the formation of protein aggregates, amyloid fibrils, intracellular, and extracellular inclusions in the most common nondegenerative disorders also referred to as protein misfolding disorders.
    Keywords:  Alzheimer's disease; Amyotrophic lateral sclerosis; Autophagy; Chaperones; Parkinson's disease; Proteostasis; Ubiquitin proteasome system; Unfolded protein response
    DOI:  https://doi.org/10.1016/bs.apcsb.2022.05.008
  46. Front Mol Neurosci. 2022 ;15 947191
    The relationship of alpha-synuclein to mitochondrial dynamics and quality control.
    Naomi J Thorne, David A Tumbarello.
      Maintenance of mitochondrial health is essential for neuronal survival and relies upon dynamic changes in the mitochondrial network and effective mitochondrial quality control mechanisms including the mitochondrial-derived vesicle pathway and mitophagy. Mitochondrial dysfunction has been implicated in driving the pathology of several neurodegenerative diseases, including Parkinson's disease (PD) where dopaminergic neurons in the substantia nigra are selectively degenerated. In addition, many genes with PD-associated mutations have defined functions in organelle quality control, indicating that dysregulation in mitochondrial quality control may represent a key element of pathology. The most well-characterized aspect of PD pathology relates to alpha-synuclein; an aggregation-prone protein that forms intracellular Lewy-body inclusions. Details of how alpha-synuclein exerts its toxicity in PD is not completely known, however, dysfunctional mitochondria have been observed in both PD patients and models of alpha-synuclein pathology. Accordingly, an association between alpha-synuclein and mitochondrial function has been established. This relates to alpha-synuclein's role in mitochondrial transport, dynamics, and quality control. Despite these relationships, there is limited research defining the direct mechanisms linking alpha-synuclein to mitochondrial dynamics and quality control. In this review, we will discuss the current literature addressing this association and provide insight into the proposed mechanisms promoting these functional relationships. We will also consider some of the alternative mechanisms linking alpha-synuclein with mitochondrial dynamics and speculate what the relationship between alpha-synuclein and mitochondria might mean both physiologically and in relation to PD.
    Keywords:  Parkinson’s disease; lysosome; membrane trafficking; mitochondria; mitochondrial quality control; vesicle transport
    DOI:  https://doi.org/10.3389/fnmol.2022.947191
  47. Mol Cell Endocrinol. 2022 Sep 10. pii: S0303-7207(22)00221-0. [Epub ahead of print] 111773
    Proteomic examination of Cornus officinalis stimulated 1.1B4 human pancreatic cells reveals activation of autophagy and Keap1/Nrf2 pathway.
    Arielle Sharp-Tawfik, Justin D Fletcher, Jennifer Guergues, Catherine Marelia-Bennett, Tiara J Wolf, Alexis M Coiner, Y Clare Zhang, Stanley M Stevens, Brant R Burkhardt.
      Type 1 diabetes (T1D) is an autoimmune disease initiated by genetic predisposition and environmental influences culminating in the immunologically mediated destruction of pancreatic β-cells with eventual loss of insulin production. Although T1D can be accurately predicted via autoantibodies, therapies are lacking that can intercede autoimmunity and protect pancreatic β-cells. There are no approved interventional modalities established for this purpose. One such potential source for clinical agents of this use is from the frequently utilized Cornus officinalis (CO) in the field of ethnopharmacology. Studies by our lab and others have demonstrated that CO has robust proliferative, metabolic, and cytokine protective effects on pancreatic β-cells. To identify the molecular mechanism of the biological effects of CO, we performed a proteomic and phosphoproteomic analysis examining the cellular networks impacted by CO application on the 1.1B4 pancreatic β-cell line. Our label-free mass spectrometry approach has demonstrated significant increased phosphorylation of the selective autophagy receptor of p62 (Sequestosome-1/SQSTM1/p62) and predicted activation of the antioxidant Kelch-like ECH-associated protein 1 (Keap1)/Nuclear factor-erythroid factor 2-related factor 2 (Nrf2) pathway. Further validation by immunoblotting and immunofluorescence revealed markers of autophagy such as increased LC3-II and decreased total p62 along with nuclear localization of Nrf2. Both autophagy and the Keap1/Nrf2 pathways have been shown to be impaired in human and animal models of T1D and may serve as an excellent potential therapeutic target stimulated by CO.
    Keywords:  Autophagy; Cornus officinalis; Keap1/Nrf2; Proteomics; Type 1 diabetes; p62
    DOI:  https://doi.org/10.1016/j.mce.2022.111773
  48. Autophagy. 2022 Sep 15. 1-19
    Secretory autophagy promotes RAB37-mediated insulin secretion under glucose stimulation both in vitro and in vivo.
    Shan-Ying Wu, Hung-Tsung Wu, Yi-Ching Wang, Chih-Jen Chang, Yan-Shen Shan, Shang-Rung Wu, Yen-Chi Chiu, Chia-Lang Hsu, Hsueh-Fen Juan, Kai-Ying Lan, Chi-Wen Chu, Ying-Ray Lee, Sheng-Hui Lan, Hsiao-Sheng Liu.
      High blood glucose is one of the risk factors for metabolic disease and INS (insulin) is the key regulatory hormone for glucose homeostasis. Hypoinsulinemia accompanied with hyperglycemia was diagnosed in mice with pancreatic β-cells exhibiting autophagy deficiency; however, the underlying mechanism remains elusive. The role of secretory autophagy in the regulation of metabolic syndrome is gaining more attention. Our data demonstrated that increased macroautophagic/autophagic activity leads to induction of insulin secretion in β-cells both in vivo and in vitro under high-glucose conditions. Moreover, proteomic analysis of purified autophagosomes from β-cells identified a group of vesicular transport proteins participating in insulin secretion, implying that secretory autophagy regulates insulin exocytosis. RAB37, a small GTPase, regulates vesicle biogenesis, trafficking, and cargo release. We demonstrated that the active form of RAB37 increased MAP1LC3/LC3 lipidation (LC3-II) and is essential for the promotion of insulin secretion by autophagy, but these phenomena were not observed in rab37 knockout (rab37-/-) cells and mice. Unbalanced insulin and glucose concentration in the blood was improved by manipulating autophagic activity using a novel autophagy inducer niclosamide (an antihelminthic drug) in a high-fat diet (HFD)-obesity mouse model. In summary, we reveal that secretory autophagy promotes RAB37-mediated insulin secretion to maintain the homeostasis of insulin and glucose both in vitro and in vivo.
    Keywords:  Glucose-stimulated insulin secretion; LC3; RAB37; insulin; secretory autophagy
    DOI:  https://doi.org/10.1080/15548627.2022.2123098
  49. Front Pharmacol. 2022 ;13 990665
    Drug repurposing in cancer neuroscience: From the viewpoint of the autophagy-mediated innervated niche.
    Jiayan Shi, Jia Xu, Yang Li, Bowen Li, Hui Ming, Edouard C Nice, Canhua Huang, Qifu Li, Chuang Wang.
      Based on the bidirectional interactions between neurology and cancer science, the burgeoning field "cancer neuroscience" has been proposed. An important node in the communications between nerves and cancer is the innervated niche, which has physical contact with the cancer parenchyma or nerve located in the proximity of the tumor. In the innervated niche, autophagy has recently been reported to be a double-edged sword that plays a significant role in maintaining homeostasis. Therefore, regulating the innervated niche by targeting the autophagy pathway may represent a novel therapeutic strategy for cancer treatment. Drug repurposing has received considerable attention for its advantages in cost-effectiveness and safety. The utilization of existing drugs that potentially regulate the innervated niche via the autophagy pathway is therefore a promising pharmacological approach for clinical practice and treatment selection in cancer neuroscience. Herein, we present the cancer neuroscience landscape with an emphasis on the crosstalk between the innervated niche and autophagy, while also summarizing the underlying mechanisms of candidate drugs in modulating the autophagy pathway. This review provides a strong rationale for drug repurposing in cancer treatment from the viewpoint of the autophagy-mediated innervated niche.
    Keywords:  autophagy; cancer neuroscience; cancer treatment; drug repurposing; innervated niche
    DOI:  https://doi.org/10.3389/fphar.2022.990665
  50. Cell Rep Med. 2022 Sep 07. pii: S2666-3791(22)00290-7. [Epub ahead of print] 100741
    PSMG2-controlled proteasome-autophagy balance mediates the tolerance for MEK-targeted therapy in triple-negative breast cancer.
    Xueyan Wang, Jing Yu, Xiaowei Liu, Dan Luo, Yanchu Li, Linlin Song, Xian Jiang, Xiaomeng Yin, Yan Wang, Li Chai, Ting Luo, Jing Jing, Hubing Shi.
      Although the MAPK pathway is aberrantly activated in triple-negative breast cancers (TNBCs), the clinical outcome of MEK-targeted therapy is still poor. Through a genome-wide CRISPR-Cas9 library screening, we find that inhibition of PSMG2 sensitizes TNBC cells BT549 and MB468 to the MEK inhibitor AZD6244. Mechanistically, PSMG2 knockdown impairs proteasome function, which in turn activates autophagy-mediated PDPK1 degradation. The PDPK1 degradation significantly enhances AZD6244-induced tumor cell growth inhibition by interrupting the negative feedback signals toward the AKT pathway. Consistently, co-targeting proteasomes and MEK with inhibitors synergistically suppresses tumor cell growth. The autophagy inhibitor chloroquine partially relieves the PDPK1 degradation and reverses the growth inhibition induced by combinatorial inhibition of MEK and proteasome. The combination regimen with the proteasome inhibitor MG132 plus AZD6244 synergistically inhibits tumor growth in a 4T1 xenograft mouse model. In summary, our study not only unravels the mechanism of MEK inhibitor resistance but also provides a combinatorial therapeutic strategy for TNBC in clinics.
    Keywords:  AKT pathway; CRISPR-Cas9; MAPK pathway; PDPK1; PSMG2; autophagy; proteasome; resistance; targeted therapy; triple-negative breast cancer
    DOI:  https://doi.org/10.1016/j.xcrm.2022.100741
  51. Mol Ther Oncolytics. 2022 Sep 15. 26 330-346
    Stabilization of DEPTOR sensitizes hypopharyngeal cancer to radiotherapy via targeting degradation.
    Xuecen Wang, Zhirui Cao, Xin Yue, Tingyu Liu, Gesi Wen, Dongmei Jiang, Weijian Wu, Liyuan Le, Yan Wang, Chengtao Wang, Ziyang Wang, Meng Jin, Meiyan Zhu, Shasha He, Xiaoyue Zhang, Xianzhang Bu, Ran-Yi Liu, Zhenwei Peng, Yong Chen.
      The use of radiotherapy for hypopharyngeal cancer (HC) treatment is increasing, and it is currently the primary treatment option for this cancer. However, radioresistance occurs in a proportion of patients. Here, we found that radiation increased proteasomal gene expression and that proteasome assembly was dependent on the induction of transcription factor NRF1 in HC. Through screening assays, we identified a mechanism by which proteasome-mediated degradation of DEP domain-containing mTOR-interacting protein (DEPTOR) contributes to the elevation of mTORC1 signaling after radiation. Therefore, after treatment with proteasome inhibitors (PIs), stabilization of DEPTOR inhibited mTORC1 signaling elevated by radiation and ultimately sensitized HC to radiotherapy. Mechanically, PIs not only interrupted the deubiquitination and degradation of DEPTOR but also suppressed the ubiquitination of DEPTOR mediated by β-TrCP. Clinically, the high levels of DEPTOR in HC cells were associated with sensitivity to radiotherapy and favorable prognosis. Stabilizing DEPTOR through targeting proteasome-mediated degradation is a potential strategy for sensitizing HC to radiotherapy.
    Keywords:  DEPTOR; hypopharyngeal cancer; mTORC1; proteasome; radioresistance
    DOI:  https://doi.org/10.1016/j.omto.2022.08.002
  52. Biochem J. 2022 Sep 13. pii: BCJ20220401. [Epub ahead of print]
    Coordinated control of the ADP-heptose/ALPK1 signalling network by the E3 ligases TRAF6, TRAF2/c-IAP1 and LUBAC.
    Tom Snelling, Natalia Shpiro, Robert Gourlay, Frederic Lamoliatte, Philip Cohen.
      ADP-heptose activates the protein kinase ALPK1 triggering TIFA phosphorylation at Thr9, the recruitment of TRAF6 and the subsequent production of inflammatory mediators. Here, we demonstrate that ADP-heptose also stimulates the formation of Lys63- and Met1-linked ubiquitin chains to activate the TAK1 and canonical IKK complexes, respectively. We further show that the E3 ligases TRAF6 and c-IAP1 operate redundantly to generate the Lys63-linked ubiquitin chains required for pathway activation, which we demonstrate are attached to TRAF6, TRAF2 and c-IAP1, and that c-IAP1 is recruited to TIFA by TRAF2. ADP-heptose also induces activation of the kinase TBK1 by a TAK1-independent mechanism, which requires TRAF2 and TRAF6. We establish that ALPK1 phosphorylates TIFA directly at Thr177 as well as Thr9 in vitro. Thr177 is located within the TRAF6-binding motif and its mutation to Asp prevents TRAF6 but not TRAF2 binding, indicating a role in restricting ADP-heptose signalling. We conclude that ADP-heptose signalling is controlled by the combined actions of TRAF2/c-IAP1 and TRAF6.
    Keywords:  ADP-heptose; ALPK1; TAK1; TBK1; TIFA; TRAF
    DOI:  https://doi.org/10.1042/BCJ20220401
  53. STAR Protoc. 2022 Sep 08. pii: S2666-1667(22)00542-1. [Epub ahead of print]3(3): 101662
    A biochemical reconstitution approach to identify autophagy receptors for aggrephagy in mammalian cells.
    Xinyu Ma, Wenhao Zhang, Haiteng Deng, Min Zhang, Liang Ge.
      Aggrephagy is a major way to clear protein aggregates. Here, we describe a pipeline of experiments to find autophagy receptors for aggrephagy. Steps include an in vitro reconstitution to recapitulate autophagosome recognizing aggregates and receptor identification steps based on flow cytometry and mass spectrometry. We also describe functional validation steps based on immunofluorescence and immunoblot. The protocol provides a practical way to identify aggrephagy receptors. For complete details on the use and execution of this protocol, please refer to Ma et al. (2022).
    Keywords:  Cell biology; Flow cytometry/Mass cytometry; Protein biochemistry
    DOI:  https://doi.org/10.1016/j.xpro.2022.101662
  54. Nature. 2022 Sep 14.
    Brain-restricted mTOR inhibition with binary pharmacology.
    Ziyang Zhang, Qiwen Fan, Xujun Luo, Kevin Lou, William A Weiss, Kevan M Shokat.
      On-target-off-tissue drug engagement is an important source of adverse effects that constrains the therapeutic window of drug candidates1,2. In diseases of the central nervous system, drugs with brain-restricted pharmacology are highly desirable. Here we report a strategy to achieve inhibition of mammalian target of rapamycin (mTOR) while sparing mTOR activity elsewhere through the use of the brain-permeable mTOR inhibitor RapaLink-1 and the brain-impermeable FKBP12 ligand RapaBlock. We show that this drug combination mitigates the systemic effects of mTOR inhibitors but retains the efficacy of RapaLink-1 in glioblastoma xenografts. We further present a general method to design cell-permeable, FKBP12-dependent kinase inhibitors from known drug scaffolds. These inhibitors are sensitive to deactivation by RapaBlock, enabling the brain-restricted inhibition of their respective kinase targets.
    DOI:  https://doi.org/10.1038/s41586-022-05213-y
  55. J Alzheimers Dis. 2022 Sep 06.
    Pathological and Therapeutic Advances in Parkinson's Disease: Mitochondria in the Interplay.
    Padmashri Naren, Anjali Cholkar, Suchita Kamble, Khan Sabiya Samim, Saurabh Srivastava, Jitender Madan, Neelesh Mehra, Vinod Tiwari, Shashi Bala Singh, Dharmendra Kumar Khatri.
      Parkinson's disease (PD) is the second most common neurodegenerative illness majorly affecting the population between the ages of 55 to 65 years. Progressive dopaminergic neuronal loss and the collective assemblage of misfolded alpha-synuclein in the substantia nigra, remain notable neuro-pathological hallmarks of the disease. Multitudes of mechanistic pathways have been proposed in attempts to unravel the pathogenesis of PD but still, it remains elusive. The convergence of PD pathology is found in organelle dysfunction where mitochondria remain a major contributor. Mitochondrial processes like bioenergetics, mitochondrial dynamics, and mitophagy are under strict regulation by the mitochondrial genome and nuclear genome. These processes aggravate neurodegenerative activities upon alteration through neuroinflammation, oxidative damage, apoptosis, and proteostatic stress. Therefore, the mitochondria have grabbed a central position in the patho-mechanistic exploration of neurodegenerative diseases like PD. The management of PD remains a challenge to physicians to date, due to the variable therapeutic response of patients and the limitation of conventional chemical agents which only offer symptomatic relief with minimal to no disease-modifying effect. This review describes the patho-mechanistic pathways involved in PD not only limited to protein dyshomeostasis and oxidative stress, but explicit attention has been drawn to exploring mechanisms like organelle dysfunction, primarily mitochondria and mitochondrial genome influence, while delineating the newer exploratory targets such as GBA1, GLP, LRRK2, and miRNAs and therapeutic agents targeting them.
    Keywords:  Autophagy; Parkinson’s disease; mitochondrial dysfunction; mitogenome; neuroinflammation; oxidative stress
    DOI:  https://doi.org/10.3233/JAD-220682
  56. Nat Commun. 2022 Sep 12. 13(1): 5351
    GCAF(TMEM251) regulates lysosome biogenesis by activating the mannose-6-phosphate pathway.
    Weichao Zhang, Xi Yang, Yingxiang Li, Linchen Yu, Bokai Zhang, Jianchao Zhang, Woo Jung Cho, Varsha Venkatarangan, Liang Chen, Bala Bharathi Burugula, Sarah Bui, Yanzhuang Wang, Cunming Duan, Jacob O Kitzman, Ming Li.
      The mannose-6-phosphate (M6P) biosynthetic pathway for lysosome biogenesis has been studied for decades and is considered a well-understood topic. However, whether this pathway is regulated remains an open question. In a genome-wide CRISPR/Cas9 knockout screen, we discover TMEM251 as the first regulator of the M6P modification. Deleting TMEM251 causes mistargeting of most lysosomal enzymes due to their loss of M6P modification and accumulation of numerous undigested materials. We further demonstrate that TMEM251 localizes to the Golgi and is required for the cleavage and activity of GNPT, the enzyme that catalyzes M6P modification. In zebrafish, TMEM251 deletion leads to severe developmental defects including heart edema and skeletal dysplasia, which phenocopies Mucolipidosis Type II. Our discovery provides a mechanism for the newly discovered human disease caused by TMEM251 mutations. We name TMEM251 as GNPTAB cleavage and activity factor (GCAF) and its related disease as Mucolipidosis Type V.
    DOI:  https://doi.org/10.1038/s41467-022-33025-1
  57. Biochim Biophys Acta Mol Cell Res. 2022 Sep 13. pii: S0167-4889(22)00147-1. [Epub ahead of print] 119355
    ULK1 inhibition attenuates telomerase activity in hepatic cells.
    Sana Raza, Sangam Rajak, Jyotika Srivastava, Archana Tewari, Pratima Gupta, Bandana Chakravarti, Sujoy Ghosh, Chandra P Chaturvedi, Rohit A Sinha.
      Autophagy and telomere maintenance are two cellular survival processes that show a strong correlation during human ageing and cancer growth, however, their causal relationship remains unclear. In this study, using an unbiased transcriptomics approach, we uncover a novel role of autophagy genes in regulating telomere extension and maintenance pathways. Concomitantly, the pharmacological inhibition of ULK1 (Unc-51 like autophagy activating kinase 1) attenuated human telomerase reverse transcriptase (hTERT) gene expression and telomerase activity in HepG2 cells. Furthermore, the suppression of telomerase activity upon ULK1 inhibition was associated with telomere shortening and onset of cellular senescence in HepG2 cells. These results, thus, demonstrate a direct role of autophagy in maintaining cellular longevity via regulation of telomerase activity, which may have implications in the pathophysiology of ageing and cancers.
    DOI:  https://doi.org/10.1016/j.bbamcr.2022.119355
  58. Front Pharmacol. 2022 ;13 977410
    The critical role of dysregulated autophagy in the progression of diabetic kidney disease.
    Ziwei Zhang, Yuting Sun, Jiaojiao Xue, De Jin, Xiangyan Li, Daqing Zhao, Fengmei Lian, Wenxiu Qi, Xiaolin Tong.
      Diabetic kidney disease (DKD) is one of the major public health problems in society today. It is a renal complication caused by diabetes mellitus with predominantly microangiopathy and is a major cause of end-stage renal disease (ESRD). Autophagy is a metabolic pathway for the intracellular degradation of cytoplasmic products and damaged organelles and plays a vital role in maintaining homeostasis and function of the renal cells. The dysregulation of autophagy in the hyperglycaemic state of diabetes mellitus can lead to the progression of DKD, and the activation or restoration of autophagy through drugs is beneficial to the recovery of renal function. This review summarizes the physiological process of autophagy, illustrates the close link between DKD and autophagy, and discusses the effects of drugs on autophagy and the signaling pathways involved from the perspective of podocytes, renal tubular epithelial cells, and mesangial cells, in the hope that this will be useful for clinical treatment.
    Keywords:  autophagosome; autophagy; diabetic kidney disease; lysosome; podocytes; renal tubular epithelial cells
    DOI:  https://doi.org/10.3389/fphar.2022.977410
  59. Front Pharmacol. 2022 ;13 893307
    Optineurin-mediated mitophagy as a potential therapeutic target for intervertebral disc degeneration.
    Zhilei Hu, Yu Wang, Xiaoxin Gao, Yuyao Zhang, Chenhao Liu, Yu Zhai, Xian Chang, Haiyin Li, Yueyang Li, Jinhui Lou, Changqing Li.
      Low back pain is thought to be mainly caused by intervertebral disc degeneration (IVDD), and there is a lack of effective treatments. Cellular senescence and matrix degradation are important factors that cause disc degeneration. Mitochondrial dysfunction induced by oxidative stress is an important mechanism of cellular senescence and matrix degradation in the nucleus pulposus (NP), and mitophagy can effectively remove damaged mitochondria, restore mitochondrial homeostasis, and mitigate the damage caused by oxidative stress. Optineurin (OPTN) is a selective mitophagy receptor, and its role in intervertebral disc degeneration remains unclear. Here, we aimed to explore the effect of OPTN on H2O2-induced nucleus pulposus cell (NPCs) senescence and matrix degradation in a rat model of disc degeneration. Western blot analysis showed that OPTN expression was reduced in degenerative human and rat nucleus pulposus tissues and increased in H2O2-induced senescent NPCs. OPTN overexpression significantly inhibited H2O2-induced senescence and increased matrix-associated protein expression in NPCs, but OPTN knockdown showed the opposite effect. As previous reports have suggested that mitophagy significantly reduces mitochondrial damage and reactive oxygen species (ROS) caused by oxidative stress, and we used the mitophagy agonist CCCP, the mitophagy inhibitor cyclosporin A (CsA), and the mitochondrial ROS (mtROS) scavenger mitoTEMPO and confirmed that OPTN attenuated NPCs senescence and matrix degeneration caused by oxidative stress by promoting mitophagy to scavenge damaged mitochondria and excess reactive oxygen species, thereby slowing the progression of IVDD. In conclusion, our research suggests that OPTN is involved in IVDD and exerts beneficial effects against IVDD.
    Keywords:  OPTN; intervertebral disc degeneration; mitophagy; nucleus pulposus cell; senescence
    DOI:  https://doi.org/10.3389/fphar.2022.893307
  60. J Cell Biol. 2022 Oct 03. pii: e202205135. [Epub ahead of print]221(10):
    ER as master regulator of membrane trafficking and organelle function.
    Eva Maria Wenzel, Liv Anker Elfmark, Harald Stenmark, Camilla Raiborg.
      The endoplasmic reticulum (ER), which occupies a large portion of the cytoplasm, is the cell's main site for the biosynthesis of lipids and carbohydrate conjugates, and it is essential for folding, assembly, and biosynthetic transport of secreted proteins and integral membrane proteins. The discovery of abundant membrane contact sites (MCSs) between the ER and other membrane compartments has revealed that, in addition to its biosynthetic and secretory functions, the ER plays key roles in the regulation of organelle dynamics and functions. In this review, we will discuss how the ER regulates endosomes, lysosomes, autophagosomes, mitochondria, peroxisomes, and the Golgi apparatus via MCSs. Such regulation occurs via lipid and Ca2+ transfer and also via control of in trans dephosphorylation reactions and organelle motility, positioning, fusion, and fission. The diverse controls of other organelles via MCSs manifest the ER as master regulator of organelle biology.
    DOI:  https://doi.org/10.1083/jcb.202205135
  61. Cell Biochem Funct. 2022 Sep 16.
    Advanced oxidation protein products mediate human keratinocytes apoptosis by inducing cell autophagy through the mTOR-Beclin-1 pathway.
    Ruoting Ding, Zhongyuan Liu, Jie Tan, Baihui Sun.
      Excessive keratinocyte apoptosis leads to impaired wound healing. Recently, advanced oxidation protein products (AOPP) have been recognized as a marker of oxidative stress and a potent inducer of apoptosis. Previously, we have demonstrated that extracellular AOPP accumulation induced keratinocyte apoptosis, and we discovered that autophagy was involved. To further elucidate the role and mechanism of autophagy in AOPP-induced-apoptosis of keratinocytes, we treated HaCaT cells with increasing concentrations of AOPP-human serum albumin or with AOPP-human serum albumin for increasing durations. Cyto-ID solution staining was used to assess cell autophagy using confocal laser scanning microscopy. Autophagy-related protein interactions were investigated using western blot analysis. Exposure of HaCaT cells to AOPP decreased the expression of mammalian target of rapamycin (mTOR) and increased the expression of autophagy-related proteins Beclin-l and LC3, and eventually led to autophagy. Furthermore, an autophagy agonist significantly decreased the expression of apoptosis-related proteins. Taken together, we showed that accumulation of extracellular AOPP induced autophagy in HaCaT cells via a reactive oxygen species-dependent, mTOR-Beclin-1-mediated pathway, and that excessive autophagy-mediated apoptosis, which resulted in delayed wound healing.
    Keywords:  advanced oxidation protein products; apoptosis; autophagy; impaired wound healing; oxidative stress
    DOI:  https://doi.org/10.1002/cbf.3749
  62. Cell Signal. 2022 Sep 07. pii: S0898-6568(22)00227-3. [Epub ahead of print]100 110465
    Exogenous hydrogen sulfide inhibits the senescence of cardiomyocytes through modulating mitophagy in rats.
    Jinghui Hao, Yuxin Xi, Lijie Jiao, Xin Wen, Ren Wu, Guiquan Chang, Fengqi Sun, Can Wei, Hongzhu Li.
      Hydrogen sulfide (H2S), a gaseous molecule, has been shown to be involved in the regulation of body pathophysiological processes. Aging is related to structural and functional alterations within the heart. There is evidence that diminished mitophagy accelerates the aging process. Studies in recent years have revealed that plasma levels of H2S in humans and old rats decrease with age, and H2S acts as a cytoprotective mediator in the aging process. However, it is unclear whether H2S can delay the senescence of cardiomyocytes by regulating mitophagy. Our present results showed that exogenous H2S inhibited mitochondrial damage, oxidative stress and cell apoptosis, and enhanced mitophagy through upregulating the SIRT1-PINK1-parkin pathway in myocardial tissues of aged rats and cultured aged cardiomyocytes. Furthermore, the effect of exogenous H2S on the above indicators was the same as that of SRT1720 (a SIRT1 agonist) and kinetin (a PINK1 activator). Our findings suggest that exogenous H2S inhibits the senescence of cardiomyocytes by increasing mitophagy via upregulation of the SIRT1-PINK1-parkin pathway in rats.
    Keywords:  Cardiomyocytes; Hydrogen sulfide; Mitophagy; Rats; Senescence
    DOI:  https://doi.org/10.1016/j.cellsig.2022.110465
  63. Aging Cell. 2022 Sep 11. e13710
    The emergent role of mitochondrial surveillance in cellular health.
    Elissa Tjahjono, Daniel R Kirienko, Natalia V Kirienko.
      Mitochondrial dysfunction is one of the primary causatives for many pathologies, including neurodegenerative diseases, cancer, metabolic disorders, and aging. Decline in mitochondrial functions leads to the loss of proteostasis, accumulation of ROS, and mitochondrial DNA damage, which further exacerbates mitochondrial deterioration in a vicious cycle. Surveillance mechanisms, in which mitochondrial functions are closely monitored for any sign of perturbations, exist to anticipate possible havoc within these multifunctional organelles with primitive origin. Various indicators of unhealthy mitochondria, including halted protein import, dissipated membrane potential, and increased loads of oxidative damage, are on the top of the lists for close monitoring. Recent research also indicates a possibility of reductive stress being monitored as part of a mitochondrial surveillance program. Upon detection of mitochondrial stress, multiple mitochondrial stress-responsive pathways are activated to promote the transcription of numerous nuclear genes to ameliorate mitochondrial damage and restore compromised cellular functions. Co-expression occurs through functionalization of transcription factors, allowing their binding to promoter elements to initiate transcription of target genes. This review provides a comprehensive summary of the intricacy of mitochondrial surveillance programs and highlights their roles in our cellular life. Ultimately, a better understanding of these surveillance mechanisms is expected to improve healthspan.
    Keywords:  aging; mitochondria; mitochondrial membrane transport proteins; mitophagy; physiological stress; reactive oxygen species; surveillance
    DOI:  https://doi.org/10.1111/acel.13710
  64. Cell. 2022 Sep 08. pii: S0092-8674(22)01112-6. [Epub ahead of print]
    mTOR-regulated mitochondrial metabolism limits mycobacterium-induced cytotoxicity.
    Antonio J Pagán, Lauren J Lee, Joy Edwards-Hicks, Cecilia B Moens, David M Tobin, Elisabeth M Busch-Nentwich, Erika L Pearce, Lalita Ramakrishnan.
      Necrosis of macrophages in the granuloma, the hallmark immunological structure of tuberculosis, is a major pathogenic event that increases host susceptibility. Through a zebrafish forward genetic screen, we identified the mTOR kinase, a master regulator of metabolism, as an early host resistance factor in tuberculosis. We found that mTOR complex 1 protects macrophages from mycobacterium-induced death by enabling infection-induced increases in mitochondrial energy metabolism fueled by glycolysis. These metabolic adaptations are required to prevent mitochondrial damage and death caused by the secreted mycobacterial virulence determinant ESAT-6. Thus, the host can effectively counter this early critical mycobacterial virulence mechanism simply by regulating energy metabolism, thereby allowing pathogen-specific immune mechanisms time to develop. Our findings may explain why Mycobacterium tuberculosis, albeit humanity's most lethal pathogen, is successful in only a minority of infected individuals.
    Keywords:  ESAT-6 mitotoxicity; Mycobacterium marinum; Mycobacterium tuberculosis; granuloma necrosis; mTOR; macrophage death; mitochondrial metabolism; oxidative phosphorylation; tuberculosis; zebrafish TB model
    DOI:  https://doi.org/10.1016/j.cell.2022.08.018
  65. Nat Commun. 2022 Sep 14. 13(1): 5383
    Structure of an MHC I-tapasin-ERp57 editing complex defines chaperone promiscuity.
    Ines Katharina Müller, Christian Winter, Christoph Thomas, Robbert M Spaapen, Simon Trowitzsch, Robert Tampé.
      Adaptive immunity depends on cell surface presentation of antigenic peptides by major histocompatibility complex class I (MHC I) molecules and on stringent ER quality control in the secretory pathway. The chaperone tapasin in conjunction with the oxidoreductase ERp57 is crucial for MHC I assembly and for shaping the epitope repertoire for high immunogenicity. However, how the tapasin-ERp57 complex engages MHC I clients has not yet been determined at atomic detail. Here, we present the 2.7-Å crystal structure of a tapasin-ERp57 heterodimer in complex with peptide-receptive MHC I. Our study unveils molecular details of client recognition by the multichaperone complex and highlights elements indispensable for peptide proofreading. The structure of this transient ER quality control complex provides the mechanistic basis for the selector function of tapasin and showcases how the numerous MHC I allomorphs are chaperoned during peptide loading and editing.
    DOI:  https://doi.org/10.1038/s41467-022-32841-9
  66. STAR Protoc. 2022 Sep 10. pii: S2666-1667(22)00545-7. [Epub ahead of print]3(4): 101665
    Evaluation of mTORC1 signaling in mouse atherosclerotic macrophages by flow cytometry and immunofluorescence.
    Xiangyu Zhang, Jeremiah Stitham, Astrid Rodriguez-Velez, Se-Jin Jeong, Arick Park, Yu-Sheng Yeh, Divya Kapoor, Bettina Mittendorfer, Babak Razani.
      Previous studies have demonstrated that a high-protein diet leads to increased atherosclerosis in mice, and that this adverse effect is caused by activation of macrophage mTORC1 signaling. Here, we provide a detailed protocol for the evaluation of diet-induced mTORC1 signaling in plaque macrophages in atherosclerosis-prone apolipoprotein E (ApoE) knockout (KO) mice. This protocol includes flow cytometry and immunofluorescence analysis of atherosclerotic macrophages that can be used to study the atherogenic potential of a variety of mTORC1 modulators. For complete details on the use and execution of this protocol, please refer to Zhang et al. (2020).
    Keywords:  Cell isolation; Flow cytometry/Mass cytometry; Immunology
    DOI:  https://doi.org/10.1016/j.xpro.2022.101665
  67. Development. 2022 Sep 16. pii: dev.200908. [Epub ahead of print]
    Regulation of Archease by the mTOR-vATPase axis.
    Deanne Francis, Alondra S Burguete, Amin Ghabrial.
      Larval terminal cells of the Drosophila tracheal system generate extensive branched tubes, requiring a huge increase in apical membrane. We discovered that terminal cells compromised for apical membrane expansion - mTOR-vATPase axis and apical polarity mutants - were invaded by the neighboring stalk cell. The invading cell grows and branches, replacing the original single intercellular junction between stalk and terminal cell with multiple intercellular junctions . Here we characterize disjointed, a mutation in the same phenotypic class. We find that disjointed encodes Drosophila Archease, required for RNA ligase (RtcB) function critical for tRNA maturation and ER stress-regulated nonconventional splicing of Xbp1 mRNA. We show that the steady-state subcellular localization of Archease is principally nuclear, and dependent upon TOR-vATPase activity. In tracheal cells mutant for Rheb or vATPase, Archease localization shifted dramatically from nucleus to cytoplasm. Further, we found that blocking tRNA maturation by knockdown of tRNAse z also induced compensatory branching. Taken together, these data suggest that the TOR-vATPase axis promotes apical membrane growth in part through nuclear localization of Archease, where Archease is required for tRNA maturation.
    Keywords:  Archease; Drosophila; Trachea; mTOR; tRNA maturation; vATPase
    DOI:  https://doi.org/10.1242/dev.200908
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