bims-auttor Biomed News
on Autophagy and mTOR
Issue of 2024‒08‒04
sixty-six papers selected by
Viktor Korolchuk, Newcastle University
  1. mTORC1 activity oscillates throughout the cell cycle, promoting mitotic entry and differentially influencing autophagy induction.
  2. TMEM9 activates Rab9-dependent alternative autophagy through interaction with Beclin1.
  3. Immune suppressive drugs negatively regulate the CD8+T cells function by acetyltransferase p300 induced canonical and non-canonical autophagy.
  4. Autophagy of Glycogen Is Non-Selective in Komagataella phaffii.
  5. ZDHHC7-mediated S-palmitoylation of ATG16L1 facilitates LC3 lipidation and autophagosome formation.
  6. HaloTag as a substrate-based macroautophagy reporter.
  7. PRRSV utilizes MALT1-regulated autophagy flux to switch virus spread and reserve.
  8. The cellular adaptor GULP1 interacts with ATG14 to potentiate autophagy and APP processing.
  9. Sestrin2 drives ER-phagy in response to protein misfolding.
  10. Research Progress on the Mechanism of Lysosome in Myocardial Ischemia-Reperfusion Injury Based on Autophagy.
  11. Human β-defensin-1 activates autophagy in human colon cancer cells via regulation of long non-coding RNA TCONS_00014506.
  12. Human iPSC-derived myelinating organoids and globoid cells to study Krabbe Disease.
  13. A fluorescent reporter for rapid assessment of autophagic flux reveals unique autophagy signatures during C. elegans post-embryonic development and identifies compounds that modulate autophagy.
  14. The V-ATPase/ATG16L1 axis is controlled by the V1H subunit.
  15. The role of autophagy in the progression of HIV infected cardiomyopathy.
  16. Huntingtin contains an ubiquitin-binding domain and regulates lysosomal targeting of mitochondrial and RNA-binding proteins.
  17. Mechanisms of lysosomal tubulation and sorting driven by LRRK2.
  18. Antiretroviral Drugs Impact Autophagy: Opportunities for Drug Repurposing.
  19. Natural autophagy modulators in non-communicable diseases: from autophagy mechanisms to therapeutic potential.
  20. More on the interplay between gut microbiota, autophagy, and inflammatory bowel disease is needed.
  21. Modulations of the mTORC2-GATA3 axis by an isorhamnetin activated endosomal-lysosomal system of the J774.1 macrophage-like cell line.
  22. Autophagy promotes replication of Bombyx mori Nucleopolyhedrovirus in insect cells.
  23. Functional OmpA of Salmonella Typhimurium provides protection from lysosomal degradation and inhibits autophagic processes in macrophages.
  24. Baculovirus-mediated endostatin and angiostatin activation of autophagy through the AMPK/AKT/mTOR pathway inhibits angiogenesis in hepatocellular carcinoma.
  25. Mitophagy Promotes Hair Regeneration by Activating Glutathione Metabolism.
  26. Targeting Autophagy in Atrial Fibrillation.
  27. miR-93-5p impairs autophagy-lysosomal pathway via TET3 after subarachnoid hemorrhage.
  28. AUF1-mediated inhibition of autophagic lysosomal degradation contributes to CagA stability and Helicobacter pylori-induced inflammation.
  29. TDP-43 proteinopathy in frontotemporal lobar degeneration and amyotrophic lateral sclerosis: From pathomechanisms to therapeutic strategies.
  30. The Roles of Endocytosis and Autophagy at the Cellular Level During Influenza Virus Infection: A Mini-Review.
  31. Mitochondrial function in oral health and disease.
  32. Axonal mitophagy in retinal ganglion cells.
  33. Heat-killed probiotic Levilactobacillus brevis MKAK9 and its exopolysaccharide promote longevity by modulating aging hallmarks and enhancing immune responses in Caenorhabditis elegans.
  34. Residual Microglia Following Short-term PLX5622 Treatment in 5xFAD Mice Exhibit Diminished NLRP3 Inflammasome and mTOR Signaling, and Enhanced Autophagy.
  35. Autophagy in drusen biogenesis secondary to age-related macular degeneration.
  36. mTORC1 activation in presumed classical monocytes: observed correlation with human size variation and neuropsychiatric disease.
  37. Glutamine deprivation in glioblastoma stem cells triggers autophagic SIRT3 degradation to epigenetically restrict CD133 expression and stemness.
  38. Fibroinase plays a vital role in silk gland degeneration by regulating autophagy and apoptosis in the silkworm, Bombyx mori.
  39. Endoplasmic reticulum stress and autophagy in cerebral ischemia/reperfusion injury: PERK as a potential target for intervention.
  40. ESCRT-III: a versatile membrane remodeling machinery and its implications in cellular processes and diseases.
  41. Age-related mitophagy regulates orthodontic tooth movement by affecting PDLSCs mitochondrial function and RANKL/OPG.
  42. Context-dependent roles for autophagy in myeloid cells in tumor progression.
  43. TRPML1 activation ameliorates lysosomal phenotypes in CLN3 deficient retinal pigment epithelial cells.
  44. From Defense to Dysfunction: Autophagy's Dual Role in Disease Pathophysiology.
  45. State-of-the-art Nrf2 Inhibitors: Therapeutic Opportunities in Non-cancer Diseases.
  46. The role of calcium-calpain pathway in hyperthermia.
  47. Autophagic degradation of CDK4 is responsible for G0/G1 cell cycle arrest in NVP-BEZ235-treated neuroblastoma.
  48. Differential roles of lysosomal cholesterol transporters in the development of C. elegans NMJs.
  49. The parkin V380L variant is a genetic modifier of Machado-Joseph disease with impact on mitophagy.
  50. V-ATPase Disassembly at the Yeast Lysosome-Like Vacuole Is a Phenotypic Driver of Lysosome Dysfunction in Replicative Aging.
  51. Autophagy-mediated activation of the AIM2 inflammasome enhances M1 polarization of microglia and exacerbates retinal neovascularization.
  52. Novel insights into autophagy in gastrointestinal pathologies, mechanisms in metabolic dysfunction-associated fatty liver disease and acute liver failure.
  53. Advances in Ubiquitination and Proteostasis in Retinal Degeneration.
  54. Lactate contributes to remote ischemic preconditioning-mediated protection against myocardial ischemia reperfusion injury by facilitating autophagy via AMPK-mTOR-TFEB-CX43 axis.
  55. Calreticulin from Apostichopus japonicus relieves endoplasmic reticulum stress induced by Vibrio splendidus through autophagy.
  56. Mitochondrial dynamics and psychiatric disorders: The missing link.
  57. SNX32 Regulates Sorting and Trafficking of Activated EGFR to the Lysosomal Degradation Pathway.
  58. Arf1 promotes porcine intestinal epithelial cell proliferation via the mTORC1 signaling pathway.
  59. Targeting the immunoproteasome in hypothalamic neurons as a novel therapeutic strategy for high-fat diet-induced obesity and metabolic dysregulation.
  60. Molecular mechanisms of PTEN in atherosclerosis: A comprehensive review.
  61. FOXM1 mediates methotrexate resistance in osteosarcoma cells by promoting autophagy.
  62. Loss of Tuberous Sclerosis Complex 2 confers inflammation via dysregulation of Nuclear factor kappa-light-chain-enhancer of activated B cells.
  63. SS-31 Mitigates Oxidative Stress and Restores Mitochondrial Function in Cigarette Smoke-Damaged Oral Epithelial Cells via PINK1-mediated Mitophagy.
  64. Targeted dephosphorylation of TFEB promotes its nuclear translocation.
  65. Capturing the catalytic intermediates of parkin ubiquitination.
  66. Metformin mitigates osteoarthritis progression by modulating the PI3K/AKT/mTOR signaling pathway and enhancing chondrocyte autophagy.
  1. Cell Rep. 2024 Jul 26. pii: S2211-1247(24)00872-6. [Epub ahead of print]43(8): 114543
    mTORC1 activity oscillates throughout the cell cycle, promoting mitotic entry and differentially influencing autophagy induction.
    Jay N Joshi, Ariel D Lerner, Frank Scallo, Alexandra N Grumet, Paul Matteson, James H Millonig, Alexander J Valvezan.
      Mechanistic Target of Rapamycin Complex 1 (mTORC1) is a master metabolic regulator that is active in nearly all proliferating eukaryotic cells; however, it is unclear whether mTORC1 activity changes throughout the cell cycle. We find that mTORC1 activity oscillates from lowest in mitosis/G1 to highest in S/G2. The interphase oscillation is mediated through the TSC complex but is independent of major known regulatory inputs, including Akt and Mek/Erk signaling. By contrast, suppression of mTORC1 activity in mitosis does not require the TSC complex. mTORC1 has long been known to promote progression through G1. We find that mTORC1 also promotes progression through S and G2 and is important for satisfying the Chk1/Wee1-dependent G2/M checkpoint to allow entry into mitosis. We also find that low mTORC1 activity in G1 sensitizes cells to autophagy induction in response to partial mTORC1 inhibition or reduced nutrient levels. Together, these findings demonstrate that mTORC1 is differentially regulated throughout the cell cycle, with important phase-specific consequences for proliferating cells.
    Keywords:  CDK1; CP: Cell biology; G2/M checkpoint; TSC complex; TSC2; autophagy; cell cycle; mTOR; mTORC1; mitosis
    DOI:  https://doi.org/10.1016/j.celrep.2024.114543
  2. Cell Mol Life Sci. 2024 Jul 30. 81(1): 322
    TMEM9 activates Rab9-dependent alternative autophagy through interaction with Beclin1.
    Sohyeon Baek, Jae-Woong Chang, Seung-Min Yoo, JeongRim Choo, Sunmin Jung, Jihoon Nah, Yong-Keun Jung.
      Transmembrane protein 9 (TMEM9) is a transmembrane protein that regulates lysosomal acidification by interacting with the v-type ATPase complex. However, the role of TMEM9 in the lysosome-dependent autophagy machinery has yet to be identified. In this study, we demonstrate that the lysosomal protein TMEM9, which is involved in vesicle acidification, regulates Rab9-dependent alternative autophagy through its interaction with Beclin1. The cytosolic domain of TMEM9 interacts with Beclin1 via its Bcl-2-binding domain. This interaction between TMEM9 and Beclin1 dissociates Bcl-2, an autophagy-inhibiting partner, from Beclin1, thereby activating LC3-independent and Rab9-dependent alternative autophagy. Late endosomal and lysosomal TMEM9 apparently colocalizes with Rab9 but not with LC3. Furthermore, we show that multiple glycosylation of TMEM9, essential for lysosomal localization, is essential for its interaction with Beclin1 and the activation of Rab9-dependent alternative autophagy. These findings reveal that TMEM9 recruits and activates the Beclin1 complex at the site of Rab9-dependent autophagosome to induce alternative autophagy.
    Keywords:  Alternative autophagy; Beclin1; Rab9; TMEM9
    DOI:  https://doi.org/10.1007/s00018-024-05366-1
  3. Heliyon. 2024 Jul 15. 10(13): e33755
    Immune suppressive drugs negatively regulate the CD8+T cells function by acetyltransferase p300 induced canonical and non-canonical autophagy.
    Runfeng Ni, Liwei Fan, Haijin Wang, Quan Zhang, Le Zhang, Aidi Wang, Baoshan Liu.
      Macroautophagy, the mainly regulated form of autophagy, maintains the cellular homeostasis and degrades the transported cargoes. It is initiated by the protein kinase complex regulating by two signals pathway Mammalian target of rapamycin complex 1 (mTORC1)-Adenosine 5' monophosphate activated protein kinase (AMPK)-Unc 51 like kinase 1(ULK1) and ULK1-PI3K- phosphatidylinositol 3-phosphate (PI3P). Currently, autolysosomes are accumulated during the aging process of CD8+T cells in vitro and may participate in inducing death sensitization of senescent cells. The main mechanism of aplastic anemia, a hyperimmune disease, is the T cells subsets imbalance such as CD8+T cells abnormal activation and hyperfunction. Therefore, the role of autophagy in the CD8+T cells and supposed whether some immunosuppress drugs induced the cells autophagic death to treat the hyperimmune diseases were focused. It was decided found that the acetyltransferase p300 obviously increased in the aplastic anemia patients and was related with the severity of disease. Previous studies have reported that canonical autophagy is regulated by the mTORC1-p300 axis. p300 is a critical bridge in the p300-VPS34 axis mediated non-canonical autophagy. There is the deficiency of autophagy and acetylation in the CD8+T cells. The expression of p300 also decreased notably after the immunosuppressive drugs therapy. Our findings provide a framework for understanding how immunosuppressive drugs effect on the AA autophagy deficiency mechanism and proved that immunosuppressive drugs negatively regulated the function of CD8+T cells by p300-mediated canonical autophagy pathway and non-canonical autophagy pathway.
    Keywords:  Aplastic anemia; CD8+T cells; Canonical/non-canonical autophagy; Cyclosporin A; Dioscin; mTORC1; p300
    DOI:  https://doi.org/10.1016/j.heliyon.2024.e33755
  4. Autophagy Rep. 2024 ;pii: 2382659. [Epub ahead of print]3(1):
    Autophagy of Glycogen Is Non-Selective in Komagataella phaffii.
    Taras Y Nazarko.
      Glycogen is an important reserve polysaccharide from bacteria to human. It is organized in glycogen granules that also contain several proteins involved in their metabolism. Glycogen granules can be mobilized in mammalian lysosomes and yeast vacuoles. They are delivered to these organelles by macroautophagy (hereafter autophagy). However, whether this is a selective or a non-selective process remains a matter of debate. It was proposed to be selective and called "glycophagy" (for selective autophagy of glycogen) in the mouse liver. However, the evidence of this selectivity is lacking in other glycogen-rich organs, such as the heart and skeletal muscle, which both are heavily impacted by the aberrant lysosomal accumulation of glycogen in Pompe disease. We recently developed the Komagataella phaffii yeast as a simple model to study the relationship of glycogen and autophagy. Using this model, we showed that cytosolic glycogen granules are delivered to the vacuole by non-selective autophagy, at least during nitrogen starvation. We speculate that this type of autophagy might be responsible for the lysosomal glycogen turnover in non-hepatic mammalian tissues.
    Keywords:  Autophagy; Glg1; Komagataella phaffii; glycogen; glycophagy; yeast
    DOI:  https://doi.org/10.1080/27694127.2024.2382659
  5. Autophagy. 2024 Aug 01.
    ZDHHC7-mediated S-palmitoylation of ATG16L1 facilitates LC3 lipidation and autophagosome formation.
    Fujing Wei, Yu Wang, Jia Yao, Ligang Mei, Xue Huang, Hesheng Kong, Jing Chen, Xiaorong Chen, Lu Liu, Zhuolin Wang, Jiaxin Wang, Jiong Song, Eryan Kong, Aimin Yang.
      Macroautophagy/autophagy is a fundamental cellular catabolic process that delivers cytoplasmic components into double-membrane vesicles called autophagosomes, which then fuse with lysosomes and their contents are degraded. Autophagy recycles cytoplasmic components, including misfolded proteins, dysfunctional organelles and even microbial invaders, thereby playing an essential role in development, immunity and cell death. Autophagosome formation is the main step in autophagy, which is governed by a set of ATG (autophagy related) proteins. ATG16L1 interacts with ATG12-ATG5 conjugate to form an ATG12-ATG5-ATG16L1 complex. The complex acts as a ubiquitin-like E3 ligase that catalyzes the lipidation of MAP1LC3/LC3 (microtubule associated protein 1 light chain 3), which is crucial for autophagosome formation. In the present study, we found that ATG16L1 was subject to S-palmitoylation on cysteine 153, which was catalyzed by ZDHHC7 (zinc finger DHHC-type palmitoyltransferase 7). We observed that re-expressing ATG16L1 but not the S-palmitoylation-deficient mutant ATG16L1C153S rescued a defect in the lipidation of LC3 and the formation of autophagosomes in ATG16L1-KO (knockout) HeLa cells. Furthermore, increasing ATG16L1 S-palmitoylation by ZDHHC7 expression promoted the production of LC3-II, whereas reducing ATG16L1 S-palmitoylation by ZDHHC7 deletion inhibited the LC3 lipidation process and autophagosome formation. Mechanistically, the addition of a hydrophobic 16-carbon palmitoyl group on Cys153 residue of ATG16L1 enhances the formation of ATG16L1-WIPI2B complex and ATG16L1-RAB33B complex on phagophore, thereby facilitating the LC3 lipidation process and autophagosome formation. In conclusion, S-palmitoylation of ATG16L1 is essential for the lipidation process of LC3 and the formation of autophagosomes. Our research uncovers a new regulatory mechanism of ATG16L1 function in autophagy.
    Keywords:  ATG16L1; LC3-II; ZDHHC7; autophagosome; autophagy; s-palmitoylation
    DOI:  https://doi.org/10.1080/15548627.2024.2386915
  6. Proc Natl Acad Sci U S A. 2024 Aug 06. 121(32): e2322500121
    HaloTag as a substrate-based macroautophagy reporter.
    Qiang Xiao, Gabrielle Cruz, Rachel Botham, Susan G Fox, Anan Yu, Seth Allen, Richard I Morimoto, Jeffery W Kelly.
      Macroautophagy is a conserved cellular degradation pathway that, upon upregulation, confers resilience toward various stress conditions, including protection against proteotoxicity associated with neurodegenerative diseases, leading to cell survival. Monitoring autophagy regulation in living cells is important to understand its role in physiology and pathology, which remains challenging. Here, we report that when HaloTag is expressed within a cell of interest and reacts with tetramethylrhodamine (TMR; its ligand attached to a fluorophore), the rate of fluorescent TMR-HaloTag conjugate accumulation in autophagosomes and lysosomes, observed by fluorescence microscopy, reflects the rate of autophagy. Notably, we found that TMR-HaloTag conjugates were mainly degraded by the proteasome (~95%) under basal conditions, while lysosomal degradation (~10% upon pharmacological autophagy activation) was slow and incomplete, forming a degraded product that remained fluorescent within a SDS-PAGE gel, in agreement with previous reports that HaloTag is resistant to lysosomal degradation when fused to proteins of interest. Autophagy activation is distinguished from autophagy inhibition by the increased production of the degraded TMR-HaloTag band relative to the full-length TMR-HaloTag band as assessed by SDS-PAGE and by a faster rate of TMR-HaloTag conjugate lysosomal puncta accumulation as observed by fluorescence microscopy. Pharmacological proteasome inhibition leads to accumulation of TMR-HaloTag in lysosomes, indicating possible cross talk between autophagy and proteasomal degradation.
    Keywords:  HaloTag; autophagy; lysosome; macroautophagy; reporter
    DOI:  https://doi.org/10.1073/pnas.2322500121
  7. Autophagy. 2024 Jul 30.
    PRRSV utilizes MALT1-regulated autophagy flux to switch virus spread and reserve.
    Han Gu, He Qiu, Haotian Yang, Zhuofan Deng, Shengkun Zhang, Liuyang Du, Fang He.
      Porcine reproductive and respiratory syndrome virus (PRRSV) is a major swine pathogen, which can survive host antiviral immunity with various mechanisms. PRRSV infection induces macroautophagy/autophagy, facilitating virus replication. MALT1, a central immune regulator, was manipulated by PRRSV to optimize viral infection at different stages of the virus cycle. In this study, the key role of MALT1 in autophagy regulation during PRRSV infection was characterized, enlightening the role of autophagy flux in favor of virus spread and persistent infection. PRRSV-induced autophagy was confirmed to facilitate virus proliferation. Furthermore, autophagic fusion was dynamically regulated during PRRSV infection. Importantly, PRRSV-induced MALT1 facilitated autophagosome-lysosome fusion and autolysosome formation, thus contributing to autophagy flux and virus proliferation. Mechanically, MALT1 regulated autophagy via mediating MTOR-ULK1 and -TFEB signaling and affecting lysosomal homeostasis. MALT1 inhibition by inhibitor Mi-2 or RNAi induced lysosomal membrane permeabilization (LMP), leading to the block of autophagic fusion. Further, MALT1 overexpression alleviated PRRSV-induced LMP via inhibiting ROS generation. In addition, blocking autophagy flux suppressed virus release significantly, indicating that MALT1-maintained complete autophagy flux during PRRSV infection favors successful virus spread and its proliferation. In contrast, autophagosome accumulation upon MALT1 inhibition promoted PRRSV reserve for future virus proliferation once the autophagy flux recovers. Taken together, for the first time, these findings elucidate that MALT1 was utilized by PRRSV to regulate host autophagy flux, to determine the fate of virus for either proliferation or reserve.
    Keywords:  Autophagy flux; LMP; MALT1; MTOR; PRRSV; lysosome homeostasis
    DOI:  https://doi.org/10.1080/15548627.2024.2386195
  8. Cell Mol Life Sci. 2024 Jul 30. 81(1): 323
    The cellular adaptor GULP1 interacts with ATG14 to potentiate autophagy and APP processing.
    Dennis Dik-Long Chau, Zhicheng Yu, Wai Wa Ray Chan, Zhai Yuqi, Raymond Chuen Chung Chang, Jacky Chi Ki Ngo, Ho Yin Edwin Chan, Kwok-Fai Lau.
      Autophagy is a highly conserved catabolic mechanism by which unnecessary or dysfunctional cellular components are removed. The dysregulation of autophagy has been implicated in various neurodegenerative diseases, including Alzheimer's disease (AD). Understanding the molecular mechanism(s)/molecules that influence autophagy may provide important insights into developing therapeutic strategies against AD and other neurodegenerative disorders. Engulfment adaptor phosphotyrosine-binding domain-containing protein 1 (GULP1) is an adaptor that interacts with amyloid precursor protein (APP) to promote amyloid-β peptide production via an unidentified mechanism. Emerging evidence suggests that GULP1 has a role in autophagy. Here, we show that GULP1 is involved in autophagy through an interaction with autophagy-related 14 (ATG14), which is a regulator of autophagosome formation. GULP1 potentiated the stimulatory effect of ATG14 on autophagy by modulating class III phosphatidylinositol 3-kinase complex 1 (PI3KC3-C1) activity. The effect of GULP1 is attenuated by a GULP1 mutation (GULP1m) that disrupts the GULP1-ATG14 interaction. Conversely, PI3KC3-C1 activity is enhanced in cells expressing APP but not in those expressing an APP mutant that does not bind GULP1, which suggests a role of GULP1-APP in regulating PI3KC3-C1 activity. Notably, GULP1 facilitates the targeting of ATG14 to the endoplasmic reticulum (ER). Moreover, the levels of both ATG14 and APP are elevated in the autophagic vacuoles (AVs) of cells expressing GULP1, but not in those expressing GULP1m. APP processing is markedly enhanced in cells co-expressing GULP1 and ATG14. Hence, GULP1 alters APP processing by promoting the entry of APP into AVs. In summary, we unveil a novel role of GULP1 in enhancing the targeting of ATG14 to the ER to stimulate autophagy and, consequently, APP processing.
    Keywords:  Amyloid precursor protein; Autophagy-related 14; GULP1; LC3; Macroautophagy
    DOI:  https://doi.org/10.1007/s00018-024-05351-8
  9. Dev Cell. 2024 Jul 30. pii: S1534-5807(24)00441-6. [Epub ahead of print]
    Sestrin2 drives ER-phagy in response to protein misfolding.
    Chiara De Leonibus, Marianna Maddaluno, Rosa Ferriero, Roberta Besio, Laura Cinque, Pei Jin Lim, Alessandro Palma, Rossella De Cegli, Salvatore Gagliotta, Sandro Montefusco, Maria Iavazzo, Marianne Rohrbach, Cecilia Giunta, Elena Polishchuk, Diego Louis Medina, Diego Di Bernardo, Antonella Forlino, Pasquale Piccolo, Carmine Settembre.
      Protein biogenesis within the endoplasmic reticulum (ER) is crucial for organismal function. Errors during protein folding necessitate the removal of faulty products. ER-associated protein degradation and ER-phagy target misfolded proteins for proteasomal and lysosomal degradation. The mechanisms initiating ER-phagy in response to ER proteostasis defects are not well understood. By studying mouse primary cells and patient samples as a model of ER storage disorders (ERSDs), we show that accumulation of faulty products within the ER triggers a response involving SESTRIN2, a nutrient sensor controlling mTORC1 signaling. SESTRIN2 induction by XBP1 inhibits mTORC1's phosphorylation of TFEB/TFE3, allowing these transcription factors to enter the nucleus and upregulate the ER-phagy receptor FAM134B along with lysosomal genes. This response promotes ER-phagy of misfolded proteins via FAM134B-Calnexin complex. Pharmacological induction of FAM134B improves clearance of misfolded proteins in ERSDs. Our study identifies the interplay between nutrient signaling and ER quality control, suggesting therapeutic strategies for ERSDs.
    Keywords:  ER storage disorders; ER-phagy; FAM134B; TFEB; alpha(1)-antitrypsin Z (alpha(1)-ATZ); autophagy; collagen; endoplasmic reticulum; mTORC1; quality control
    DOI:  https://doi.org/10.1016/j.devcel.2024.07.004
  10. Rev Cardiovasc Med. 2024 Apr;25(4): 113
    Research Progress on the Mechanism of Lysosome in Myocardial Ischemia-Reperfusion Injury Based on Autophagy.
    Yi Li, Hui Wu, Songlin Zhang, Gang Zhou, Dong Zhang, Qingzhuo Yang, Yanfang Liu, Xiaoli Huang.
      In recent years, the interaction of intracellular organelles such as mitochondria and lysosomal functions has attracted increasing attention. Recent evidence suggests that mitochondrion-lysosomal contact plays a key role in regulating lysosomal biogenesis and maintaining cellular homeostasis. Myocardial ischemia and reperfusion will lead to corresponding changes in the autophagy flux in cardiomyocytes, and lysosomes are a key link in the process of autophagy, and the fusion of lysosomes and autophagosomes is an essential link in the occurrence of autophagy. Therefore, the function and homeostasis of lysosomes also undergo different changes during myocardial ischemia and reperfusion. Lysosomal-related biological factors and membrane proteins also play different roles. This article will review the mechanism of lysosomes in myocardial ischemia-reperfusion injury and the research progress of lysosomal-related proteins.
    Keywords:  autophagy; lysosomal membrane protein; lysosome; myocardial ischemia reperfusion injury
    DOI:  https://doi.org/10.31083/j.rcm2504113
  11. World J Gastrointest Oncol. 2024 Jul 15. 16(7): 2894-2901
    Human β-defensin-1 activates autophagy in human colon cancer cells via regulation of long non-coding RNA TCONS_00014506.
    Nabil Eid, Fabian Davamani.
      Macroautophagy (hereafter referred to as autophagy) is a prosurvival mechanism for the clearance of damaged cellular components, specifically related to exposure to various stressors such as starvation, excessive ethanol intake, and chemotherapy. This editorial reviews and comments on an article by Zhao et al, to be published in World J Gastrointestinal Oncology in 2024. Based on various molecular biology methodologies, they found that human β-defensin-1 reduced the proliferation of colon cancer cells, which was associated with the inhibition of the mammalian target of rapamycin, resulting in autophagy activation. The activation of autophagy is evidenced by increased levels of Beclin1 and LC3II/I proteins and mediated by the upregulation of long non-coding RNA TCONS_00014506. Our study discusses the impact of autophagy activation and mechanisms of autophagy, including autophagic flux, on cancer cells. Additionally, we emphasize the importance of describing the detailed methods for isolating long noncoding RNAs TCONS_00014506. Our review will benefit the scientific community and improve the overall clarity of the paper.
    Keywords:  Autophagy; Autophagy flux; Beclin-1; Colon cancer; Human β-defensin-1; LC3-II; Long noncoding RNA; Mammalian target of rapamycin
    DOI:  https://doi.org/10.4251/wjgo.v16.i7.2894
  12. bioRxiv. 2024 Jul 23. pii: 2024.07.19.604372. [Epub ahead of print]
    Human iPSC-derived myelinating organoids and globoid cells to study Krabbe Disease.
    Lisa Marie P Evans, Joseph Gawron, Fraser J Sim, M Laura Feltri, Leandro N Marziali.
      Krabbe disease (Kd) is a lysosomal storage disorder (LSD) caused by the deficiency of the lysosomal galactosylceramidase (GALC) which cleaves the myelin enriched lipid galactosylceramide (GalCer). Accumulated GalCer is catabolized into the cytotoxic lipid psychosine that causes myelinating cells death and demyelination which recruits microglia/macrophages that fail to digest myelin debris and become globoid cells. Here, to understand the pathological mechanisms of Kd, we used induced pluripotent stem cells (iPSCs) from Kd patients to produce myelinating organoids and microglia. We show that Kd organoids have no obvious defects in neurogenesis, astrogenesis, and oligodendrogenesis but manifest early myelination defects. Specifically, Kd organoids showed shorter but a similar number of myelin internodes than Controls at the peak of myelination and a reduced number and shorter internodes at a later time point. Interestingly, myelin is affected in the absence of autophagy and mTOR pathway dysregulation, suggesting lack of lysosomal dysfunction which makes this organoid model a very valuable tool to study the early events that drive demyelination in Kd. Kd iPSC-derived microglia show a marginal rate of globoid cell formation under normal culture conditions that is drastically increased upon GalCer feeding. Under normal culture conditions, Kd microglia show a minor LAMP1 content decrease and a slight increase in the autophagy protein LC3B. Upon GalCer feeding, Kd cells show accumulation of autophagy proteins and strong LAMP1 reduction that at a later time point are reverted showing the compensatory capabilities of globoid cells. Altogether, this supports the value of our cultures as tools to study the mechanisms that drive globoid cell formation and the compensatory mechanism in play to overcome GalCer accumulation in Kd.
    DOI:  https://doi.org/10.1101/2024.07.19.604372
  13. Autophagy Rep. 2024 ;pii: 2371736. [Epub ahead of print]3(1):
    A fluorescent reporter for rapid assessment of autophagic flux reveals unique autophagy signatures during C. elegans post-embryonic development and identifies compounds that modulate autophagy.
    Zachary D Dawson, Hemalatha Sundaramoorthi, Suk Regmi, Bo Zhang, Stephanie Morrison, Sara M Fielder, Jessie R Zhang, Hieu Hoang, David H Perlmutter, Cliff J Luke, Gary A Silverman, Stephen C Pak.
      Autophagy is important for many physiological processes; and disordered autophagy can contribute to the pathogenesis of a broad range of systemic disorders. C. elegans is a useful model organism for studying the genetics of autophagy, however, current methods for studying autophagy are labor-intensive and not readily amenable to high-throughput procedures. Here we describe a fluorescent reporter, GFP::LGG-1::mKate2, which is useful for monitoring autophagic flux in live animals. In the intestine, the fusion protein is processed by endogenous ATG-4 to generate GFP::LGG-1 and mKate2 proteins. We provide data indicating that the GFP:mKate ratio is a suitable readout for measuring cellular autophagic flux. Using this reporter, we measured autophagic flux in L1 larvae to day 7 adult animals. We show that basal autophagic flux is relatively low during larval development but increases markedly in reproductive adults before decreasing with age. Furthermore, we show that wild-type, eat-2, and daf-2 mutant animals have distinct autophagic flux profiles through post-embryonic development. Finally, we demonstrate the utility of this reporter by performing a high-content small molecule screen to identify compounds that alter autophagic flux in C. elegans.
    Keywords:  Biomarker; LC3; LGG-1; high-content screening; probe; small molecule
    DOI:  https://doi.org/10.1080/27694127.2024.2371736
  14. Mol Cell. 2024 Jul 24. pii: S1097-2765(24)00579-3. [Epub ahead of print]
    The V-ATPase/ATG16L1 axis is controlled by the V1H subunit.
    Lewis Timimi, Antoni G Wrobel, George N Chiduza, Sarah L Maslen, Antonio Torres-Méndez, Beatriz Montaner, Colin Davis, Taylor Minckley, Katriona L Hole, Andrea Serio, Michael J Devine, J Mark Skehel, John L Rubinstein, Anne Schreiber, Rupert Beale.
      Defects in organellar acidification indicate compromised or infected compartments. Recruitment of the autophagy-related ATG16L1 complex to pathologically neutralized organelles targets ubiquitin-like ATG8 molecules to perturbed membranes. How this process is coupled to proton gradient disruption is unclear. Here, we reveal that the V1H subunit of the vacuolar ATPase (V-ATPase) proton pump binds directly to ATG16L1. The V1H/ATG16L1 interaction only occurs within fully assembled V-ATPases, allowing ATG16L1 recruitment to be coupled to increased V-ATPase assembly following organelle neutralization. Cells lacking V1H fail to target ATG8s during influenza infection or after activation of the immune receptor stimulator of interferon genes (STING). We identify a loop within V1H that mediates ATG16L1 binding. A neuronal V1H isoform lacks this loop and is associated with attenuated ATG8 targeting in response to ionophores in primary murine and human iPSC-derived neurons. Thus, V1H controls ATG16L1 recruitment following proton gradient dissipation, suggesting that the V-ATPase acts as a cell-intrinsic damage sensor.
    Keywords:  ATG16L1; ATP6V1H; CASM; STING; V-ATPase; VAIL; autophagy; influenza; non-canonical autophagy; vacuolar ATPase
    DOI:  https://doi.org/10.1016/j.molcel.2024.07.003
  15. Front Cell Dev Biol. 2024 ;12 1372573
    The role of autophagy in the progression of HIV infected cardiomyopathy.
    Yuting Sun, Mengmeng Xu, Qinchun Duan, Joseph L Bryant, Xuehong Xu.
      Although highly active antiretroviral therapy (HAART) has changed infection with human immunodeficiency virus (HIV) from a diagnosis with imminent mortality to a chronic illness, HIV positive patients who do not develop acquired immunodeficiency syndrome (AIDs) still suffer from a high rate of cardiac dysfunction and fibrosis. Regardless of viral load and CD count, HIV-associated cardiomyopathy (HIVAC) still causes a high rate of mortality and morbidity amongst HIV patients. While this is a well characterized clinical phenomena, the molecular mechanism of HIVAC is not well understood. In this review, we consolidate, analyze, and discuss current research on the intersection between autophagy and HIVAC. Multiple studies have linked dysregulation in various regulators and functional components of autophagy to HIV infection regardless of mode of viral entry, i.e., coronary, cardiac chamber, or pericardial space. HIV proteins, including negative regulatory factor (Nef), glycoprotein 120 (gp120), and transactivator (Tat), have been shown to interact with type II microtubule-associated protein-1 β light chain (LC3-II), Rubiquitin, SQSTM1/p62, Rab7, autophagy-specific gene 7 (ATG7), and lysosomal-associated membrane protein 1 (LAMP1), all molecules critical to normal autophagy. HIV infection can also induce dysregulation of mitochondrial bioenergetics by altering production and equilibrium of adenosine triphosphate (ATP), mitochondrial reactive oxygen species (ROS), and calcium. These changes alter mitochondrial mass and morphology, which normally trigger autophagy to clear away dysfunctional organelles. However, with HIV infection also triggering autophagy dysfunction, these abnormal mitochondria accumulate and contribute to myocardial dysfunction. Likewise, use of HAART, azidothymidine and Abacavir, have been shown to induce cardiac dysfunction and fibrosis by inducing abnormal autophagy during antiretroviral therapy. Conversely, studies have shown that increasing autophagy can reduce the accumulation of dysfunctional mitochondria and restore cardiomyocyte function. Interestingly, Rapamycin, a mammalian target of rapamycin (mTOR) inhibitor, has also been shown to reduce HIV-induced cytotoxicity by regulating autophagy-related proteins, making it a non-antiviral agent with the potential to treat HIVAC. In this review, we synthesize these findings to provide a better understanding of the role autophagy plays in HIVAC and discuss the potential pharmacologic targets unveiled by this research.
    Keywords:  acquired immunodeficiency syndrome (AIDS); autophagy; cardiomyopathy; highly active antiretroviral therapy (HAART); human immunodeficiency virus (HIV); mTOR; mitochondria; rapamycin
    DOI:  https://doi.org/10.3389/fcell.2024.1372573
  16. Proc Natl Acad Sci U S A. 2024 Aug 06. 121(32): e2319091121
    Huntingtin contains an ubiquitin-binding domain and regulates lysosomal targeting of mitochondrial and RNA-binding proteins.
    Gianna M Fote, Vinay V Eapen, Ryan G Lim, Clinton Yu, Lisa Salazar, Nicolette R McClure, Jharrayne McKnight, Thai B Nguyen, Marie C Heath, Alice L Lau, Mark A Villamil, Ricardo Miramontes, Ian H Kratter, Steven Finkbeiner, Jack C Reidling, Joao A Paulo, Peter Kaiser, Lan Huang, David E Housman, Leslie M Thompson, Joan S Steffan.
      Understanding the normal function of the Huntingtin (HTT) protein is of significance in the design and implementation of therapeutic strategies for Huntington's disease (HD). Expansion of the CAG repeat in the HTT gene, encoding an expanded polyglutamine (polyQ) repeat within the HTT protein, causes HD and may compromise HTT's normal activity contributing to HD pathology. Here, we investigated the previously defined role of HTT in autophagy specifically through studying HTT's association with ubiquitin. We find that HTT interacts directly with ubiquitin in vitro. Tandem affinity purification was used to identify ubiquitinated and ubiquitin-associated proteins that copurify with a HTT N-terminal fragment under basal conditions. Copurification is enhanced by HTT polyQ expansion and reduced by mimicking HTT serine 421 phosphorylation. The identified HTT-interacting proteins include RNA-binding proteins (RBPs) involved in mRNA translation, proteins enriched in stress granules, the nuclear proteome, the defective ribosomal products (DRiPs) proteome and the brain-derived autophagosomal proteome. To determine whether the proteins interacting with HTT are autophagic targets, HTT knockout (KO) cells and immunoprecipitation of lysosomes were used to investigate autophagy in the absence of HTT. HTT KO was associated with reduced abundance of mitochondrial proteins in the lysosome, indicating a potential compromise in basal mitophagy, and increased lysosomal abundance of RBPs which may result from compensatory up-regulation of starvation-induced macroautophagy. We suggest HTT is critical for appropriate basal clearance of mitochondrial proteins and RBPs, hence reduced HTT proteostatic function with mutation may contribute to the neuropathology of HD.
    Keywords:  Huntingtin; RNA-binding proteins; autophagy; ubiquitin; ubiquitin-binding domain
    DOI:  https://doi.org/10.1073/pnas.2319091121
  17. Biochem Soc Trans. 2024 Jul 31. pii: BST20240087. [Epub ahead of print]
    Mechanisms of lysosomal tubulation and sorting driven by LRRK2.
    Luis Bonet-Ponce, Jillian H Kluss, Mark R Cookson.
      Lysosomes are dynamic cellular structures that adaptively remodel their membrane in response to stimuli, including membrane damage. Lysosomal dysfunction plays a central role in the pathobiology of Parkinson's disease (PD). Gain-of-function mutations in Leucine-rich repeat kinase 2 (LRRK2) cause familial PD and genetic variations in its locus increase the risk of developing the sporadic form of the disease. We previously uncovered a process we term LYTL (LYsosomal Tubulation/sorting driven by LRRK2), wherein membrane-damaged lysosomes generate tubules sorted into mobile vesicles. Subsequently, these vesicles interact with healthy lysosomes. LYTL is orchestrated by LRRK2 kinase activity, via the recruitment and phosphorylation of a subset of RAB GTPases. Here, we summarize the current understanding of LYTL and its regulation, as well as the unknown aspects of this process.
    Keywords:  Parkinson's disease; RAB proteins; leucine-rich repeat kinase
    DOI:  https://doi.org/10.1042/BST20240087
  18. Front Biosci (Landmark Ed). 2024 Jul 02. 29(7): 242
    Antiretroviral Drugs Impact Autophagy: Opportunities for Drug Repurposing.
    Laura Cheney, John M Barbaro, Grace McDermott, Joan W Berman.
      Autophagy is an evolutionarily conserved process in which intracellular macromolecules are degraded in a lysosomal-dependent manner. It is central to cellular energy homeostasis and to quality control of intracellular components. A decline in autophagic activity is associated with aging, and contributes to the development of various age-associated pathologies, including cancer. There is an ongoing need to develop chemotherapeutic agents to improve morbidity and mortality for those diagnosed with cancer, as well as to decrease the cost of cancer care. Autophagic programs are altered in cancer cells to support survival in genetically and metabolically unstable environments, making autophagy an attractive target for new chemotherapy. Antiretroviral drugs, which have dramatically increased the life- and health spans of people with human immunodeficiency virus (HIV) (PWH), have offered promise in the treatment of cancer. One mechanism underlying the antineoplastic effects of antiretroviral drugs is the alteration of cancer cell autophagy that can potentiate cell death. Antiretroviral drugs could be repurposed into the cancer chemotherapy arsenal. A more complete understanding of the impact of antiretroviral drugs on autophagy is essential for effective repurposing. This review summarizes our knowledge of the effects of antiretroviral drugs on autophagy as potential adjunctive chemotherapeutic agents, and highlights gaps to be addressed to reposition antiretroviral drugs into the antineoplastic arsenal successfully.
    Keywords:  HIV; LC3; age-related diseases; antiretroviral drug; autophagy; cancer; cell death; chemotherapy; drug repositioning; p62
    DOI:  https://doi.org/10.31083/j.fbl2907242
  19. Acta Pharmacol Sin. 2024 Aug 01.
    Natural autophagy modulators in non-communicable diseases: from autophagy mechanisms to therapeutic potential.
    Ting-Ting Xu, Ying-Yi Deng, Xi-Yong Yu, Min Li, Yuan-Yuan Fu.
      Non-communicable diseases (NCDs) are defined as a kind of diseases closely related to bad behaviors and lifestyles, e.g., cardiovascular diseases, cancer, and diabetes. Driven by population growth and aging, NCDs have become the biggest disease burden in the world, and it is urgent to prevent and control these chronic diseases. Autophagy is an evolutionarily conserved process that degrade cellular senescent or malfunctioning organelles in lysosomes. Mounting evidence has demonstrated a major role of autophagy in the pathogenesis of cardiovascular diseases, cancer, and other major human diseases, suggesting that autophagy could be a candidate therapeutic target for NCDs. Natural products/phytochemicals are important resources for drugs against a wide variety of diseases. Recently, compounds from natural plants, such as resveratrol, curcumin, and ursolic acid, have been recognized as promising autophagy modulators. In this review, we address recent advances and the current status of the development of natural autophagy modulators in NCDs and provide an update of the latest in vitro and in vivo experiments that pave the way to clinical studies. Specifically, we focus on the relationship between natural autophagy modulators and NCDs, with an intent to identify natural autophagy modulators with therapeutic potential.
    Keywords:  autophagy modulators; cancer; cardiovascular diseases; diabetes; natural products; non-communicable diseases
    DOI:  https://doi.org/10.1038/s41401-024-01356-y
  20. World J Gastroenterol. 2024 Jul 21. 30(27): 3356-3360
    More on the interplay between gut microbiota, autophagy, and inflammatory bowel disease is needed.
    Arunkumar Subramanian, Afrarahamed Jahabardeen, Tamilanban Thamaraikani, Chitra Vellapandian.
      The concept of inflammatory bowel disease (IBD), which encompasses Crohn's disease and ulcerative colitis, represents a complex and growing global health concern resulting from a multifactorial etiology. Both dysfunctional autophagy and dysbiosis contribute to IBD, with their combined effects exacerbating the related inflammatory condition. As a result, the existing interconnection between gut microbiota, autophagy, and the host's immune system is a decisive factor in the occurrence of IBD. The factors that influence the gut microbiota and their impact are another important point in this regard. Based on this initial perspective, this manuscript briefly highlighted the intricate interplay between the gut microbiota, autophagy, and IBD pathogenesis. In addition, it also addressed the potential targeting of the microbiota and modulating autophagic pathways for IBD therapy and proposed suggestions for future research within a more specific and expanded context. Further studies are warranted to explore restoring microbial balance and regulating autophagy mechanisms, which may offer new therapeutic avenues for IBD management and to delve into personalized treatment to alleviate the related burden.
    Keywords:  Autophagy; Crohn’s disease; Gut microbiota; Inflammatory bowel disease; Ulcerative colitis
    DOI:  https://doi.org/10.3748/wjg.v30.i27.3356
  21. J Clin Biochem Nutr. 2024 Jul;75(1): 24-32
    Modulations of the mTORC2-GATA3 axis by an isorhamnetin activated endosomal-lysosomal system of the J774.1 macrophage-like cell line.
    Maiko Sakai, Kohta Ohnishi, Masashi Masuda, Erika Harumoto, Teppei Fukuda, Aika Ohnishi, Shunsuke Ishii, Hirokazu Ohminami, Hisami Yamanaka-Okumura, Kazuto Ohashi, Eisuke Itakura, Kazuki Horikawa, Shigenobu Yonemura, Taichi Hara, Yutaka Taketani.
      The endosomal-lysosomal system represents a crucial degradation pathway for various extracellular substances, and its dysfunction is linked to cardiovascular and neurodegenerative diseases. This degradation process involves multiple steps: (1) the uptake of extracellular molecules, (2) transport of cargos to lysosomes, and (3) digestion by lysosomal enzymes. While cellular uptake and lysosomal function are reportedly regulated by the mTORC1-TFEB axis, the key regulatory signal for cargo transport remains unclear. Notably, our previous study discovered that isorhamnetin, a dietary flavonoid, enhances endosomal-lysosomal proteolysis in the J774.1 cell line independently of the mTORC1-TFEB axis. This finding suggests the involvement of another signal in the mechanism of isorhamnetin. This study analyzes the molecular mechanism of isorhamnetin using transcriptome analysis and reveals that the transcription factor GATA3 plays a critical role in enhanced endosomal-lysosomal degradation. Our data also demonstrate that mTORC2 regulates GATA3 nuclear translocation, and the mTORC2-GATA3 axis alters endosomal formation and maturation, facilitating the efficient transport of cargos to lysosomes. This study suggests that the mTORC2-GATA3 axis might be a novel target for the degradation of abnormal substances.
    Keywords:  GATA3; endocytosis; isorhamnetin; mTORC2; protein degradation
    DOI:  https://doi.org/10.3164/jcbn.24-22
  22. Int J Biol Macromol. 2024 Jul 30. pii: S0141-8130(24)05130-4. [Epub ahead of print] 134325
    Autophagy promotes replication of Bombyx mori Nucleopolyhedrovirus in insect cells.
    Shigang Fei, Junming Xia, Nasir Mehmood, Yeyuan Wang, Min Feng, Jingchen Sun.
      BmNPV is a pathogen that infects silkworms exclusively. Although the interaction between BmNPV and the silkworm has been widely noticed and studied, its specific mechanism has still not been elucidated. In this study, we investigated whether BmNPV infection induces the onset of host cell autophagy to enhance viral replication. We observed a significant increase in double- or single-membrane vesicles and an accumulation of enhanced green fluorescent protein eGFP-ATG8 spots in virus-infected cells 72 h after BmNPV infection, accompanied by a conversion of ATG8 to ATG8-PE. In addition, we observed changes in the mitochondrial morphology of BmN cells after BmNPV infection by transmission electron microscopy. By detecting the mitochondrial membrane potential, we found that BmNPV infection resulted in the decrease of mitochondrial membrane potential, and that eGFP-ATG8 was able to co-localise with mitochondria after virus infection of the cells. Moreover, the use of drugs to regulate the occurrence of autophagy affects the replication of cellular BmNPV. Our data demonstrates that BmNPV infection induces host cell autophagy and leads to cellular mitochondrial damage, which in turn may lead to mitochondrial autophagy, and that BmNPV-induced host autophagy promotes its replication in cells. These findings will provide clues for further understanding of host-virus interactions.
    Keywords:  Autophagy; BmNPV; Mitochondrial membrane; Silkworm
    DOI:  https://doi.org/10.1016/j.ijbiomac.2024.134325
  23. J Infect Dis. 2024 Jul 30. pii: jiae376. [Epub ahead of print]
    Functional OmpA of Salmonella Typhimurium provides protection from lysosomal degradation and inhibits autophagic processes in macrophages.
    Atish Roy Chowdhury, Dipasree Hajra, Debapriya Mukherjee, Abhilash Vijay Nair, Dipshikha Chakravortty.
      Our previous study showed that OmpA-deficient Salmonella Typhimurium (STM) failed to retain LAMP-1, quit Salmonella-containing vacuole (SCV) and escaped to the host cytosol. Here we show that the cytosolic population of STM ΔompA sequestered autophagic markers, syntaxin17 and LC3B in a sseL-dependent manner and initiated lysosomal fusion. Moreover, inhibition of autophagy using bafilomycinA1 restored its intracellular proliferation. Ectopic overexpression of OmpA in STM ΔsifA restored its vacuolar niche and increased interaction of LAMP-1, suggesting a sifA-independent role of OmpA in maintaining an intact SCV. The OmpA extracellular loops impaired the LAMP-1 recruitment to SCV and caused bacterial release into the cytosol of macrophages, but unlike STM ΔompA, they retained their outer membrane stability and didn't activate the lysosomal degradation pathway aiding in their intra-macrophage survival. Finally, OmpA extracellular loop mutations protected the cytosolic STM ΔsifA from the lysosomal surveillance, revealing a unique OmpA-dependent strategy of STM for its intracellular survival.
    Keywords:   Salmonella-containing vacuole (SCV); EEA-1; LAMP-1; LC3B; OmpA extracellular loops; lysosomes; outer membrane protein A (OmpA); syntaxin 17
    DOI:  https://doi.org/10.1093/infdis/jiae376
  24. Open Life Sci. 2024 ;19(1): 20220914
    Baculovirus-mediated endostatin and angiostatin activation of autophagy through the AMPK/AKT/mTOR pathway inhibits angiogenesis in hepatocellular carcinoma.
    Tingting Wei, Jiajie Cheng, Yonggan Ji, Xue Cao, Shuqin Ding, Quanxia Liu, Zhisheng Wang.
      Hepatocellular carcinoma (HCC) is a highly vascularized carcinoma, and targeting its neovascularization represents an effective therapeutic approach. Our previous study demonstrated that the baculovirus-mediated endostatin and angiostatin fusion protein (BDS-hEA) effectively inhibits the angiogenesis of vascular endothelial cells and the growth of HCC tumors. However, the mechanism underlying its anti-angiogenic effect remains unclear. Increasing evidence suggests that autophagy has a significant impact on the function of vascular endothelial cells and response to cancer therapy. Hence, the objective of this research was to investigate the correlation between BDS-hEA-induced angiogenesis inhibition and autophagy, along with potential regulatory mechanisms. Our results demonstrated that BDS-hEA induced autophagy in EA.hy926 cells, as evidenced by the increasing number of autophagosomes and reactive oxygen species, accompanied by an upregulation of Beclin-1, LC3-II/LC3-I, and p62 protein expression. Suppression of autophagy using 3-methyladenine attenuated the functions of BDS-hEA-induced EA.hy926 cells, including the viability, proliferation, invasion, migration, and angiogenesis. Moreover, BDS-hEA induced autophagy by downregulating the expression of CD31, VEGF, and VEGFR2, as well as phosphorylated protein kinase B (p-AKT) and phosphorylated mammalian target of rapamycin (p-mTOR), while concurrently upregulating phosphorylated AMP-activated protein kinase (p-AMPK). The in vivo results further indicated that inhibition of autophagy by chloroquine significantly impeded the ability of BDS-hEA to suppress HCC tumor growth in mice. Mechanistically, BDS-hEA prominently facilitated autophagic apoptosis in tumor tissues and decreased the levels of ki67, CD31, VEGF, MMP-9, p-AKT, and p-mTOR while simultaneously enhancing the p-AMPK expression. In conclusion, our findings suggest that BDS-hEA induces autophagy as a cytotoxic response by modulating the AMPK/AKT/mTOR signaling pathway, thereby exerting anti-angiogenic effects against HCC.
    Keywords:  anti-angiogenesis; autophagy; baculovirus; endostatin-angiostatin; hepatocellular carcinoma
    DOI:  https://doi.org/10.1515/biol-2022-0914
  25. Research (Wash D C). 2024 ;7 0433
    Mitophagy Promotes Hair Regeneration by Activating Glutathione Metabolism.
    Dehuan Wang, Jingwei Jiang, Mengyue Wang, Ke Li, Huan Liang, Nian'ou Wang, Weiwei Liu, Miaomiao Wang, Siyi Zhou, Man Zhang, Yang Xiao, Xinyu Shen, Zeming Li, Wang Wu, Xia Lin, Xiao Xiang, Qiaoli Xie, Wanqian Liu, Xun Zhou, Qu Tang, Wei Zhou, Li Yang, Cheng-Ming Chuong, Mingxing Lei.
      Mitophagy maintains tissue homeostasis by self-eliminating defective mitochondria through autophagy. How mitophagy regulates stem cell activity during hair regeneration remains unclear. Here, we found that mitophagy promotes the proliferation of hair germ (HG) cells by regulating glutathione (GSH) metabolism. First, single-cell RNA sequencing, mitochondrial probe, transmission electron microscopy, and immunofluorescence staining showed stronger mitochondrial activity and increased mitophagy-related gene especially Prohibitin 2 (Phb2) expression at early-anagen HG compared to the telogen HG. Mitochondrial inner membrane receptor protein PHB2 binds to LC3 to initiate mitophagy. Second, molecular docking and functional studies revealed that PHB2-LC3 activates mitophagy to eliminate the damaged mitochondria in HG. RNA-seq, single-cell metabolism, immunofluorescence staining, and functional validation discovered that LC3 promotes GSH metabolism to supply energy for promoting HG proliferation. Third, transcriptomics analysis and immunofluorescence staining indicated that mitophagy was down-regulated in the aged compared to young-mouse HG. Activating mitophagy and GSH pathways through small-molecule administration can reactivate HG cell proliferation followed by hair regeneration in aged hair follicles. Our findings open up a new avenue for exploring autophagy that promotes hair regeneration and emphasizes the role of the self-elimination effect of mitophagy in controlling the proliferation of HG cells by regulating GSH metabolism.
    DOI:  https://doi.org/10.34133/research.0433
  26. Rev Cardiovasc Med. 2023 Oct;24(10): 288
    Targeting Autophagy in Atrial Fibrillation.
    Qiang Ye, Wen Chen, Hengsong Fu, Yanling Ding, Yuling Jing, Jingsong Shen, Ziyang Yuan, Kelan Zha.
      Atrial fibrillation (AF) is the most common type of arrhythmia in clinical practice, and its incidence is positively correlated with risk factors that include advanced age, hypertension, diabetes, and heart failure. Although our understanding of the mechanisms that govern the occurrence and persistence of AF has been increasing rapidly, the exact mechanism of AF is still not fully understood. Autophagy is an evolutionarily highly conserved and specific physiological process in cells that has been suggested as a potential therapeutic target for several cardiovascular diseases including the pathophysiology of AF. The present article provides an updated review of the fast-progressing field of research surrounding autophagy in AF, and how regulating autophagy might be a therapeutic target to reduce the incidence of AF.
    Keywords:  atrial anatomical remodeling; atrial electrical remodeling; atrial fibrillation; autophagy; energy metabolism remodeling
    DOI:  https://doi.org/10.31083/j.rcm2410288
  27. Exp Neurol. 2024 Jul 31. pii: S0014-4886(24)00230-9. [Epub ahead of print] 114904
    miR-93-5p impairs autophagy-lysosomal pathway via TET3 after subarachnoid hemorrhage.
    Peng-Fei Ding, Xun-Zhi Liu, Zheng Peng, Yue Cui, Yang Liu, Jia-Tong Zhang, Qi Zhu, Jie Wang, Yan Zhou, Yong-Yue Gao, Chun-Hua Hang, Wei Li.
      Intact autophagy-lysosomal pathway (ALP) in neuronal survival is crucial. However, it remains unclear whether ALP is intact after subarachnoid hemorrhage (SAH). Ten-eleven translocation (TET) 3 primarily regulates genes related to autophagy in neurons in neurodegenerative diseases. This study aims to investigate the role of TET3 in the ALP following SAH. The results indicate that the ALP is impaired after SAH, with suppressed autophagic flux and an increase in autophagosomes. This is accompanied by a decrease in TET3 expression. Activation of TET3 by α-KG can improve ALP function and neural function to some extent. Silencing TET3 in neurons significantly inhibited the ALP function and increased apoptosis. Inhibition of miR-93-5p, which is elevated after SAH, promotes TET3 expression. This suggests that the downregulation of TET3 after SAH is, at least in part, due to elevated miR-93-5p. This study clarifies the key role of TET3 in the functional impairment of the ALP after SAH. The preliminary exploration revealed that miR-93-5p could lead to the downregulation of TET3, which could be a new target for neuroprotective therapy after SAH.
    Keywords:  Autophagy-lysosomal pathway; Neuron; Subarachnoid hemorrhage; TET3; miR-93-5p
    DOI:  https://doi.org/10.1016/j.expneurol.2024.114904
  28. Gut Microbes. 2024 Jan-Dec;16(1):16(1): 2382766
    AUF1-mediated inhibition of autophagic lysosomal degradation contributes to CagA stability and Helicobacter pylori-induced inflammation.
    Huiling Zheng, Ting Zhang, Jing Zhang, Jing Ning, Weiwei Fu, Ye Wang, Yanyan Shi, Guochao Wei, Jing Zhang, Xiangmei Chen, Shigang Ding.
      CagA, a virulence factor of Helicobacter pylori (H. pylori), is known to drive inflammation in gastric epithelial cells and is typically degraded through autophagy. However, the molecular mechanism by which CagA evades autophagy-mediated degradation remains elusive. This study found that H. pylori inhibits autophagic flux by upregulating the expression of AU-rich element RNA-binding factor 1 (AUF1). We confirmed that AUF1 does not affect autophagy initiation but instead hampers lysosomal clearance, as evidenced by treatments with 3-MA, CQ and BafA1. Upregulated AUF1 stabilizes CagA protein levels by inhibiting the autolysosomal degradation of intracellular CagA in H. pylori-infected gastric epithelial cells. Knocking down AUF1 promotes CagA degradation, an effect that can be reversed by the lysosome inhibitor BafA1 and CQ. Transcriptome analysis of AUF1-knockdown gastric epithelial cells infected with H. pylori indicated that AUF1 regulates the expression of lysosomal-associated hydrolase genes, specifically CTSD, to inhibit autolysosomal degradation. Moreover, we observed that knockdown of AUF1 enhanced the stability of CTSD mRNA and identified AUF1 binding to the 3'UTR region of CTSD mRNA. AUF1-mediated downregulation of CTSD expression contributes to CagA stability, and AUF1 overexpression leads to an increase in CagA levels in exosomes, thus promoting extracellular inflammation. In clinical gastric mucosa, the expression of AUF1 and its cytoplasmic translocation are associated with H. pylori-associated gastritis, with CagA being necessary for the translocation of AUF1 into the cytoplasm. Our findings suggest that AUF1 is a novel host-positive regulator of CagA, and dysregulation of AUF1 expression increases the risk of H. pylori-associated gastritis.
    Keywords:  AUF1; CagA degradation; Helicobacter pylori-gastritis
    DOI:  https://doi.org/10.1080/19490976.2024.2382766
  29. Ageing Res Rev. 2024 Jul 27. pii: S1568-1637(24)00259-9. [Epub ahead of print]100 102441
    TDP-43 proteinopathy in frontotemporal lobar degeneration and amyotrophic lateral sclerosis: From pathomechanisms to therapeutic strategies.
    Pei-Chuan Ho, Tsung-Chi Hsieh, Kuen-Jer Tsai.
      Proteostasis failure is a common pathological characteristic in neurodegenerative diseases. Revitalizing clearance systems could effectively mitigate these diseases. The transactivation response (TAR) DNA-binding protein 43 (TDP-43) plays a critical role as an RNA/DNA-binding protein in RNA metabolism and synaptic function. Accumulation of TDP-43 aggregates in the central nervous system is a hallmark of frontotemporal lobar degeneration (FTLD) and amyotrophic lateral sclerosis (ALS). Autophagy, a major and highly conserved degradation pathway, holds the potential for degrading aggregated TDP-43 and alleviating FTLD/ALS. This review explores the causes of TDP-43 aggregation, FTLD/ALS-related genes, key autophagy factors, and autophagy-based therapeutic strategies targeting TDP-43 proteinopathy. Understanding the underlying pathological mechanisms of TDP-43 proteinopathy can facilitate therapeutic interventions.
    Keywords:  Amyotrophic lateral sclerosis; Autophagy; Frontotemporal lobar degeneration; TDP-43
    DOI:  https://doi.org/10.1016/j.arr.2024.102441
  30. Infect Drug Resist. 2024 ;17 3199-3208
    The Roles of Endocytosis and Autophagy at the Cellular Level During Influenza Virus Infection: A Mini-Review.
    Sulpiana, Riezki Amalia, Nur Atik.
      Acute respiratory infections contribute to morbidity and mortality worldwide. The common cause of this deadly disease is a virus, and one of the most commonly found is the influenza virus. Influenza viruses have several capabilities in infection, including utilizing the host's machinery to survive within cells and replicate safely. This review aims to examine the literature on how influenza viruses use host machinery, including endocytosis and autophagy, for their internalization and replication within cells. This review method involves a literature search by examining articles published in the PubMed and Scopus databases. The keywords used were "Endocytosis" OR "Autophagy" AND "Influenza Virus". Eighteen articles were included due to inclusion and exclusion criteria. GTPases switch, and V-ATPase plays a key role in the endocytic machinery hijacked by influenza viruses to enter host cells. On the other hand, LC3 and Atg5 facilitate influenza-induced apoptosis via the autophagic pathway. In conclusion, influenza viruses primarily use clathrin-mediated endocytosis to enter cells and avoid degradation during endosomal maturation by exiting endosomes for transfer to the nucleus for replication. It also uses autophagy to induce apoptosis to continue replication. The capability of the influenza viruses to hijack endocytosis and autophagy mechanisms could be critical points for further research. Therefore, we discuss how the influenza virus utilizes both endocytosis and autophagy and the approach for a new strategic therapy targeting those mechanisms.
    Keywords:  Autophagy; endocytosis; influenza virus; virus internalization
    DOI:  https://doi.org/10.2147/IDR.S471204
  31. J Immunol Methods. 2024 Jul 25. pii: S0022-1759(24)00114-5. [Epub ahead of print]532 113729
    Mitochondrial function in oral health and disease.
    Ana Carolina Morandini, Erivan S Ramos-Junior.
      Monitoring mitochondrial function and mitochondrial quality control in tissues is a crucial aspect of understanding cellular health and dysfunction, which may inform about the pathogenesis of several conditions associated with aging, including chronic inflammatory conditions, neurodegenerative disorders and metabolic diseases. This process involves assessing the functionality, integrity, and abundance of mitochondria within cells. Several lines of evidence have explored techniques and methods for monitoring mitochondrial quality control in tissues. In this review, we summarize and provide our perspective considering the latest evidence in mitochondrial function and mitochondrial quality control in oral health and disease with a particular focus in periodontal inflammation. This research is significant for gaining insights into cellular health and the pathophysiology of periodontal disease, a dysbiosis-related, immune mediated and age-associated chronic condition representing a significant burden to US elderly population. Approaches for assessing mitochondrial health status reviewed here include assessing mitochondrial dynamics, mitophagy, mitochondrial biogenesis, oxidative stress, electron transport chain function and metabolomics. Such assessments help researchers comprehend the role of mitochondrial function in cellular homeostasis and its implications for oral diseases.
    Keywords:  Mitochondrial biogenesis; Mitochondrion; Mitophagy; Oral health; Oxidative phosphorylation; Periodontitis
    DOI:  https://doi.org/10.1016/j.jim.2024.113729
  32. Cell Commun Signal. 2024 Jul 29. 22(1): 382
    Axonal mitophagy in retinal ganglion cells.
    Yang Liang, Yulin Li, Qing Jiao, Muyang Wei, Yan Wang, Aoteng Cui, Zhihui Li, Guangyu Li.
      Neurons, exhibiting unique polarized structures, rely primarily on the mitochondrial production of ATP to maintain their hypermetabolic energy requirements. To maintain a normal energy supply, mitochondria are transported to the distal end of the axon. When mitochondria within the axon are critically damaged beyond their compensatory capacity, they are cleared via autophagosomal phagocytosis, and the degradation products are recycled to replenish energy. When the mitochondria are dysfunctional or their transport processes are blocked, axons become susceptible to degeneration triggered by energy depletion, resulting in neurodegenerative diseases. As the final checkpoint for mitochondrial quality control, axonal mitophagy is vital for neuronal growth, development, injury, and regeneration. Furthermore, abnormal axonal mitophagy is crucial in the pathogenesis of optic nerve-related diseases such as glaucoma. We review recent studies on axonal mitophagy and summarize the progress of research on axonal mitophagy in optic nerve-related diseases to provide insights into diseases associated with axonal damage in optic ganglion cells.
    Keywords:  Axon; Energy; Mitophagy; Optic nerve
    DOI:  https://doi.org/10.1186/s12964-024-01761-0
  33. Immun Ageing. 2024 Aug 02. 21(1): 52
    Heat-killed probiotic Levilactobacillus brevis MKAK9 and its exopolysaccharide promote longevity by modulating aging hallmarks and enhancing immune responses in Caenorhabditis elegans.
    Arun Kumar, Manti Kumar Saha, Vipin Kumar, Anupam Bhattacharya, Sagar Barge, Ashis K Mukherjee, Mohan C Kalita, Mojibur R Khan.
      BACKGROUND: Proteostasis is a critical aging hallmark responsible for removing damaged or misfolded proteins and their aggregates by improving proteasomal degradation through the autophagy-lysosome pathway (ALP) and the ubiquitin-proteasome system (UPS). Research on the impact of heat-killed probiotic bacteria and their structural components on aging hallmarks and innate immune responses is scarce, yet enhancing these effects could potentially delay age-related diseases.RESULTS: This study introduces a novel heat-killed Levilactobacillus brevis strain MKAK9 (HK MKAK9), along with its exopolysaccharide (EPS), demonstrating their ability to extend longevity by improving proteostasis and immune responses in wild-type Caenorhabditis elegans. We elucidate the underlying mechanisms through a comprehensive approach involving mRNA- and small RNA sequencing, proteomic analysis, lifespan assays on loss-of-function mutants, and quantitative RT-PCR. Mechanistically, HK MKAK9 and its EPS resulted in downregulation of the insulin-like signaling pathway in a DAF-16-dependent manner, enhancing protein ubiquitination and subsequent proteasomal degradation through activation of the ALP pathway, which is partially mediated by microRNA mir-243. Importantly, autophagosomes engulf ubiquitinylated proteins, as evidenced by increased expression of the autophagy receptor sqst-3, and subsequently fuse with lysosomes, facilitated by increased levels of the lysosome-associated membrane protein (LAMP) lmp-1, suggesting the formation of autolysosomes for degradation of the selected cargo. Moreover, HK MKAK9 and its EPS activated the p38 MAPK pathway and its downstream SKN-1 transcription factor, which are known to regulate genes involved in innate immune response (thn-1, ilys-1, cnc-2, spp-9, spp-21, clec-47, and clec-266) and antioxidation (sod-3 and gst-44), thereby reducing the accumulation of reactive oxygen species (ROS) at both cellular and mitochondrial levels. Notably, SOD-3 emerged as a transcriptional target of both DAF-16 and SKN-1 transcription factors.
    CONCLUSION: Our research sets a benchmark for future investigations by demonstrating that heat-killed probiotic and its specific cellular component, EPS, can downregulate the insulin-signaling pathway, potentially improving the autophagy-lysosome pathway (ALP) for degrading ubiquitinylated proteins and promoting organismal longevity. Additionally, we discovered that increased expression of microRNA mir-243 regulates insulin-like signaling and its downstream ALP pathway. Our findings also indicate that postbiotic treatment may bolster antioxidative and innate immune responses, offering a promising avenue for interventions in aging-related diseases.
    Keywords:   Lactobacillus ; Autophagy-lysosome pathway; HK MKAK9; Longevity; Probiotics; miRNA; p38 MAPK signalling
    DOI:  https://doi.org/10.1186/s12979-024-00457-w
  34. bioRxiv. 2024 Jul 16. pii: 2024.07.11.603157. [Epub ahead of print]
    Residual Microglia Following Short-term PLX5622 Treatment in 5xFAD Mice Exhibit Diminished NLRP3 Inflammasome and mTOR Signaling, and Enhanced Autophagy.
    Maheedhar Kodali, Leelavathi N Madhu, Yogish Somayaji, Sahithi Attaluri, Charles Huard, Prashanta Kumar Panda, Goutham Shankar, Shama Rao, Bing Shuai, Jenny J Gonzalez, Chris Oake, Catherine Hering, Roshni Sara Babu, Sanya Kotian, Ashok K Shetty.
      Chronic neuroinflammation represents a prominent hallmark of Alzheimer's disease (AD). While moderately activated microglia are pivotal in clearing amyloid beta (Aβ), hyperactivated microglia perpetuate neuroinflammation. Prior investigations have indicated that the elimination of ∼80% of microglia through a month-long inhibition of the colony-stimulating factor 1 receptor (CSF1R) during the advanced stage of neuroinflammation in 5xFamilial AD (5xFAD) mice mitigates synapse loss and neurodegeneration without impacting Aβ levels. Furthermore, prolonged CSF1R inhibition diminished the development of parenchymal plaques. Nonetheless, the immediate effects of short-term CSF1R inhibition during the early stages of neuroinflammation on residual microglial phenotype or metabolic fitness are unknown. Therefore, we investigated the effects of 10-day CSF1R inhibition in three-month-old female 5xFAD mice, a stage characterized by the onset of neuroinflammation and minimal Aβ plaques. We observed ∼65% microglia depletion in the hippocampus and cerebral cortex. The leftover microglia demonstrated a noninflammatory phenotype, with highly branched and ramified processes and reduced NOD-, LRR-, and pyrin domain-containing protein 3 (NLRP3) inflammasome complexes. Moreover, plaque-associated microglia were reduced in number with diminished Clec7a (dectin-1) expression. Additionally, both microglia and neurons displayed reduced mechanistic target of rapamycin (mTOR) signaling and autophagy. Biochemical assays validated the inhibition of NLRP3 inflammasome activation, decreased mTOR signaling, and enhanced autophagy. However, short-term CSF1R inhibition did not influence Aβ plaques, soluble Aβ-42 levels, or hippocampal neurogenesis. Thus, short-term CSF1R inhibition during the early stages of neuroinflammation in 5xFAD mice promotes the retention of homeostatic microglia with diminished inflammasome activation and mTOR signaling, alongside increased autophagy.
    DOI:  https://doi.org/10.1101/2024.07.11.603157
  35. Acta Ophthalmol. 2024 Aug 01.
    Autophagy in drusen biogenesis secondary to age-related macular degeneration.
    Juha M T Hyttinen, Ali Koskela, Janusz Blasiak, Kai Kaarniranta.
      Age-related macular degeneration (AMD) is an emerging cause of blindness in aged people worldwide. One of the key signs of AMD is the degeneration of the retinal pigment epithelium (RPE), which is indispensable for the maintenance of the adjacent photoreceptors. Because of impaired energy metabolism resulting from constant light exposure, hypoxia, and oxidative stress, accumulation of drusen in AMD-affected eyes is observed. Drusen contain damaged cellular proteins, lipoprotein particles, lipids and carbohydrates and they are related to impaired protein clearance, inflammation, and extracellular matrix modification. When autophagy, a major cellular proteostasis pathway, is impaired, the accumulations of intracellular lipofuscin and extracellular drusen are detected. As these aggregates grow over time, they finally cause the disorganisation and destruction of the RPE and photoreceptors leading to visual loss. In this review, the role of autophagy in drusen biogenesis is discussed since impairment in removing cellular waste in RPE cells plays a key role in AMD progression. In the future, means which improve intracellular clearance might be of use in AMD therapy to slow the progression of drusen formation.
    Keywords:  age‐related macular degeneration; degradative autophagy; drusen; lipofuscin; phagocytosis; retinal pigment epithelium; secretory autophagy
    DOI:  https://doi.org/10.1111/aos.16744
  36. Aging (Albany NY). 2024 Jul 26. 16
    mTORC1 activation in presumed classical monocytes: observed correlation with human size variation and neuropsychiatric disease.
    Karl Berner, Naci Oz, Alaattin Kaya, Animesh Acharjee, Jon Berner.
      BACKGROUND: Gain of function disturbances in nutrient sensing are likely the largest component in human age-related disease. Mammalian target of rapamycin complex 1 (mTORC1) activity affects health span and longevity. The drugs ketamine and rapamycin are effective against chronic pain and depression, and both affect mTORC1 activity. Our objective was to measure phosphorylated p70S6K, a marker for mTORC1 activity, in individuals with psychiatric disease to determine whether phosphorylated p70S6K could predict medication response.METHODS: Twenty-seven females provided blood samples in which p70S6K and phosphorylated p70S6K were analyzed. Chart review gathered biometric measurements, clinical phenotypes, and medication response. Questionnaires assessed anxiety, depression, autism traits, and mitochondrial dysfunction, to determine neuropsychiatric disease profiles. Univariate and multivariate statistical analyses were used to identify predictors of medication response.
    RESULTS: mTORC1 activity correlated highly with both classical biometrics (height, macrocephaly, pupil distance) and specific neuropsychiatric disease profiles (anxiety and autism). Across all cases, phosphorylated p70S6K was the best predictor for ketamine response, and also the best predictor for rapamycin response in a single instance.
    CONCLUSIONS: The data illustrate the importance of mTORC1 activity in both observable body structure and medication response. This report suggests that a simple assay may allow cost-effective prediction of medication response.
    Keywords:  ketamine; lithium; mTORC1; monocyte; rapamycin
    DOI:  https://doi.org/10.18632/aging.206033
  37. Apoptosis. 2024 Jul 28.
    Glutamine deprivation in glioblastoma stem cells triggers autophagic SIRT3 degradation to epigenetically restrict CD133 expression and stemness.
    Zhengcao Xing, Xianguo Jiang, Yalan Chen, Tiange Wang, Xiaohe Li, Xiangyun Wei, Qiuju Fan, Jie Yang, Hongmei Wu, Jinke Cheng, Rong Cai.
      Glioblastoma multiforme (GBM) is a highly malignant brain tumor, and glioblastoma stem cells (GSCs) are the primary cause of GBM heterogeneity, invasiveness, and resistance to therapy. Sirtuin 3 (SIRT3) is mainly localized in the mitochondrial matrix and plays an important role in maintaining GSC stemness through cooperative interaction with the chaperone protein tumor necrosis factor receptor-associated protein 1 (TRAP1) to modulate mitochondrial respiration and oxidative stress. The present study aimed to further elucidate the specific mechanisms by which SIRT3 influences GSC stemness, including whether SIRT3 serves as an autophagy substrate and the mechanism of SIRT3 degradation. We first found that SIRT3 is enriched in CD133+ GSCs. Further experiments revealed that in addition to promoting mitochondrial respiration and reducing oxidative stress, SIRT3 maintains GSC stemness by epigenetically regulating CD133 expression via succinate. More importantly, we found that SIRT3 is degraded through the autophagy-lysosome pathway during GSC differentiation into GBM bulk tumor cells. GSCs are highly dependent on glutamine for survival, and in these cells, we found that glutamine deprivation triggers autophagic SIRT3 degradation to restrict CD133 expression, thereby disrupting the stemness of GSCs. Together our results reveal a novel mechanism by which SIRT3 regulates GSC stemness. We propose that glutamine restriction to trigger autophagic SIRT3 degradation offers a strategy to eliminate GSCs, which combined with other treatment methods may overcome GBM resistance to therapy as well as relapse.
    Keywords:  Autophagic degradation; GBM; Glioblastoma stem cell; Glutamine deprivation; SIRT3
    DOI:  https://doi.org/10.1007/s10495-024-02003-x
  38. Int J Biol Macromol. 2024 Jul 29. pii: S0141-8130(24)05117-1. [Epub ahead of print] 134312
    Fibroinase plays a vital role in silk gland degeneration by regulating autophagy and apoptosis in the silkworm, Bombyx mori.
    Zhan Wang, Pengchao Guo, Lan Hu, Guosheng Hua, Yuanyuan Yang, Haogang Zheng, Huan Fang, Qingyou Xia, Ping Zhao.
      The silkworm is an incredibly valuable insect that produces silk through its silk gland. Within this organ, Fibroinase has been identified and named due to its ability to fibroin degradation. The expression of Fibroinase in the silk gland significantly increases during the larval-pupal stage, which might be associated with the degeneration of the silk gland. In this study, Fibroinase was overexpressed and knocked down specifically both in the middle and posterior silk glands, respectively, using transgenic technology. The investigation of silk gland development in these transgenic silkworms showed that Fibroinase plays a direct role in accelerating silk gland degeneration. The staining analyses performed in the silk glands of transgenic silkworms suggest that Fibroinase is involved in the processes of autophagy and apoptosis during silk gland degeneration. Further experiments demonstrated that Fibroinase, acting as a lysosomal regulator, negatively regulates autophagy via the mTOR (mechanistic target of rapamycin) pathway. Moreover, during apoptosis, Fibroinase could activate Caspase3 by increasing the activity of BmCaspase1, ultimately accelerating the apoptosis process. These findings enhance our understanding of the physiological role of Fibroinase in promoting silk gland degeneration, which plays a role in breaking down proteins in the silk gland and coordinating the regulation of autophagy and apoptosis.
    Keywords:  Apoptosis; Autophagy; Degeneration; Silk gland; Transgenic silkworms
    DOI:  https://doi.org/10.1016/j.ijbiomac.2024.134312
  39. Neural Regen Res. 2025 May 01. 20(5): 1455-1466
    Endoplasmic reticulum stress and autophagy in cerebral ischemia/reperfusion injury: PERK as a potential target for intervention.
    Ju Zheng, Yixin Li, Ting Zhang, Yanlin Fu, Peiyan Long, Xiao Gao, Zhengwei Wang, Zhizhong Guan, Xiaolan Qi, Wei Hong, Yan Xiao.
      JOURNAL/nrgr/04.03/01300535-202505000-00028/figure1/v/2024-07-28T173839Z/r/image-tiff Several studies have shown that activation of unfolded protein response and endoplasmic reticulum (ER) stress plays a crucial role in severe cerebral ischemia/reperfusion injury. Autophagy occurs within hours after cerebral ischemia, but the relationship between ER stress and autophagy remains unclear. In this study, we established experimental models using oxygen-glucose deprivation/reoxygenation in PC12 cells and primary neurons to simulate cerebral ischemia/reperfusion injury. We found that prolongation of oxygen-glucose deprivation activated the ER stress pathway protein kinase-like endoplasmic reticulum kinase (PERK)/eukaryotic translation initiation factor 2 subunit alpha (eIF2α)-activating transcription factor 4 (ATF4)-C/EBP homologous protein (CHOP), increased neuronal apoptosis, and induced autophagy. Furthermore, inhibition of ER stress using inhibitors or by siRNA knockdown of the PERK gene significantly attenuated excessive autophagy and neuronal apoptosis, indicating an interaction between autophagy and ER stress and suggesting PERK as an essential target for regulating autophagy. Blocking autophagy with chloroquine exacerbated ER stress-induced apoptosis, indicating that normal levels of autophagy play a protective role in neuronal injury following cerebral ischemia/reperfusion injury. Findings from this study indicate that cerebral ischemia/reperfusion injury can trigger neuronal ER stress and promote autophagy, and suggest that PERK is a possible target for inhibiting excessive autophagy in cerebral ischemia/reperfusion injury.
    DOI:  https://doi.org/10.4103/NRR.NRR-D-23-00794
  40. Anim Cells Syst (Seoul). 2024 ;28(1): 367-380
    ESCRT-III: a versatile membrane remodeling machinery and its implications in cellular processes and diseases.
    Jisoo Park, Jongyoon Kim, Hyungsun Park, Taewan Kim, Seongju Lee.
      The endosomal sorting complexes required for transport (ESCRT) machinery is an evolutionarily conserved cytosolic protein complex that plays a crucial role in membrane remodeling and scission events across eukaryotes. Initially discovered for its function in multivesicular body (MVB) formation, the ESCRT complex has since been implicated in a wide range of membrane-associated processes, including endocytosis, exocytosis, cytokinesis, and autophagy. Recent advances have elucidated the ESCRT assembly pathway and highlighted the distinct functions of the various ESCRT complexes and their associated partners. Among the ESCRT complexes, ESCRT-III stands out as a critical player in membrane remodeling, with its subunits assembled into higher-order multimers capable of bending and severing membranes. This review focuses on the ESCRT-III complex, exploring its diverse functions in cellular processes beyond MVB biogenesis. We delve into the molecular mechanisms underlying ESCRT-III-mediated membrane remodeling and highlight its emerging roles in processes such as viral budding, autophagosome closure, and cytokinetic abscission. We also discuss the implications of ESCRT-III dysregulation in neurodegenerative diseases. The versatile membrane remodeling capabilities of ESCRT-III across diverse cellular processes underscore its importance in maintaining proper cellular function. Furthermore, we highlight the promising potential of ESCRT-III as a therapeutic target for neurodegenerative diseases, offering insights into the treatments of the diseases and the technical applications in related research fields.
    Keywords:  Autophagy; ESCRT-III; MVB; membrane remodeling; neurodegenerative disease
    DOI:  https://doi.org/10.1080/19768354.2024.2380294
  41. FASEB J. 2024 Aug 15. 38(15): e23865
    Age-related mitophagy regulates orthodontic tooth movement by affecting PDLSCs mitochondrial function and RANKL/OPG.
    Tong Yan, Huilin Li, Jiayin Yan, Siyuan Ma, Jiali Tan.
      A thorough comprehension of age-related variances in orthodontic tooth movement (OTM) and bone remodeling response to mechanical force holds significant implications for enhancing orthodontic treatment. Mitophagy plays a crucial role in bone metabolism and various age-related diseases. However, the impact of mitophagy on the bone remodeling process during OTM remains elusive. Using adolescent (6 weeks old) and adult (12 months old) rats, we established OTM models and observed that orthodontic force increased the expression of the mitophagy proteins PTEN-induced putative kinase 1 (PINK1) and Parkin, as well as the number of tartrate-resistant acid phosphatase-positive osteoclasts and osteocalcin-positive osteoblasts. These biological changes were found to be age-related. In vitro, compression force loading promoted PINK1/Parkin-dependent mitophagy in periodontal ligament stem cells (PDLSCs) derived from adolescents (12-16 years old) and adults (25-35 years old). Furthermore, adult PDLSCs exhibited lower levels of mitophagy, impaired mitochondrial function, and a decreased ratio of RANKL/OPG compared to young PDLSCs after compression. Transfection of siRNA confirmed that inhibition of mitophagy in PDLSC resulted in decreased mitochondrial function and reduced RANKL/OPG ratio. Application of mitophagy inducer Urolithin A enhanced bone remodeling and accelerated OTM in rats, while the mitophagy inhibitor Mdivi-1 had the opposite effect. These findings indicate that force-stimulated PDLSC mitophagy contributes to alveolar bone remodeling during OTM, and age-related impairment of mitophagy negatively impacts the PDLSC response to mechanical stimulus. Our findings enhance the understanding of mitochondrial mechanotransduction and offer new targets to tackle current clinical challenges in orthodontic therapy.
    Keywords:  PTEN‐induced putative kinase 1; mechanotransduction; mitophagy; orthodontics; periodontal ligament stem cells
    DOI:  https://doi.org/10.1096/fj.202401280R
  42. bioRxiv. 2024 Jul 16. pii: 2024.07.12.603292. [Epub ahead of print]
    Context-dependent roles for autophagy in myeloid cells in tumor progression.
    Jayoung Choi, Gayoung Park, Steve Seung-Young Lee, Erin Dominici, Lev Becker, Kay F Macleod, Stephen J Kron, Seungmin Hwang.
      Autophagy is known to suppress tumor initiation by removing genotoxic stresses in normal cells. Conversely, autophagy is also known to support tumor progression by alleviating metabolic stresses in neoplastic cells. Centered on this pro-tumor role of autophagy, there have been many clinical trials to treat cancers through systemic blocking of autophagy. Such systemic inhibition affects both tumor cells and non-tumor cells, and the consequence of blocked autophagy in non-tumor cells in the context of tumor microenvironment is relatively understudied. Here, we examined the effect of autophagy-deficient myeloid cells on the progression of autophagy-competent tumors. We found that blocking autophagy only in myeloid cells modulated tumor progression markedly but such effects were context dependent. In a tumor implantation model, the growth of implanted tumor cells was substantially reduced in mice with autophagy-deficient myeloid cells; T cells infiltrated deeper into the tumors and were responsible for the reduced growth of the implanted tumor cells. In an oncogene-driven tumor induction model, however, tumors grew faster and metastasized more in mice with autophagy-deficient myeloid cells. These data demonstrate that the autophagy status of myeloid cells plays a critical role in tumor progression, promoting or suppressing tumor growth depending on the context of tumor-myeloid cell interactions. This study indicates that systemic use of autophagy inhibitors in cancer therapy may have differential effects on rates of tumor progression in patients due to effects on myeloid cells and that this warrants more targeted use of selective autophagy inhibitors in a cancer therapy in a clinical setting.
    Keywords:  Autophagy; macrophage; microenvironment; myeloid cell; tumor
    DOI:  https://doi.org/10.1101/2024.07.12.603292
  43. Sci Rep. 2024 Jul 29. 14(1): 17469
    TRPML1 activation ameliorates lysosomal phenotypes in CLN3 deficient retinal pigment epithelial cells.
    D Wünkhaus, R Tang, K Nyame, N N Laqtom, M Schweizer, A Scotto Rosato, E K Krogsæter, C Wollnik, M Abu-Remaileh, C Grimm, G Hermey, R Kuhn, D Gruber-Schoffnegger, S Markmann.
      Mutations in the lysosomal membrane protein CLN3 cause Juvenile Neuronal Ceroid Lipofuscinosis (JNCL). Activation of the lysosomal ion channel TRPML1 has previously been shown to be beneficial in several neurodegenerative disease models. Here, we tested whether TRPML1 activation rescues disease-associated phenotypes in CLN3-deficient retinal pigment epithelial (ARPE-19 CLN3-KO) cells. ARPE-19 CLN3-KO cells accumulate LAMP1 positive organelles and show lysosomal storage of mitochondrial ATPase subunit C (SubC), globotriaosylceramide (Gb3), and glycerophosphodiesters (GPDs), whereas lysosomal bis(monoacylglycero)phosphate (BMP/LBPA) lipid levels were significantly decreased. Activation of TRPML1 reduced lysosomal storage of Gb3 and SubC but failed to restore BMP levels in CLN3-KO cells. TRPML1-mediated decrease of storage was TFEB-independent, and we identified TRPML1-mediated enhanced lysosomal exocytosis as a likely mechanism for clearing storage including GPDs. Therefore, ARPE-19 CLN3-KO cells represent a human cell model for CLN3 disease showing many of the described core lysosomal deficits, some of which can be improved using TRPML1 agonists.
    DOI:  https://doi.org/10.1038/s41598-024-67479-8
  44. Eur J Pharmacol. 2024 Jul 26. pii: S0014-2999(24)00545-4. [Epub ahead of print] 176856
    From Defense to Dysfunction: Autophagy's Dual Role in Disease Pathophysiology.
    Jonaid Ahmad Malik, Mohammad Adeel Zafar, Sanpreet Singh, Sidhanta Nanda, Hilal Bashir, Deepjyoti Kumar Das, Taruna Lamba, Mohammad Affan Khan, Gurpreet Kaur, Javed N Agrewala.
      Autophagy is a fundamental pillar of cellular resilience, indispensable for maintaining cellular health and vitality. It coordinates the meticulous breakdown of cytoplasmic macromolecules as a guardian of cell metabolism, genomic integrity, and survival. In the complex play of biological warfare, autophagy emerges as a firm defender, bravely confronting various pathogenic, infectious, and cancerous adversaries. Nevertheless, its role transcends mere defense, wielding both protective and harmful effects in the complex landscape of disease pathogenesis. From the onslaught of infectious outbreaks to the devious progression of chronic lifestyle disorders, autophagy emerges as a central protagonist, convolutedly shaping the trajectory of cellular health and disease progression. In this article, we embark on a journey into the complicated web of molecular and immunological mechanisms that govern autophagy's profound influence over disease. Our focus sharpens on dissecting the impact of various autophagy-associated proteins on the kaleidoscope of immune responses, spanning the spectrum from infectious outbreaks to chronic lifestyle ailments. Through this voyage of discovery, we unveil the vast potential of autophagy as a therapeutic linchpin, offering tantalizing prospects for targeted interventions and innovative treatment modalities that promise to transform the landscape of disease management.
    Keywords:  Autophagy; autoimmune diseases; infectious diseases; lifestyle diseases
    DOI:  https://doi.org/10.1016/j.ejphar.2024.176856
  45. ChemMedChem. 2024 Jul 31. e202400377
    State-of-the-art Nrf2 Inhibitors: Therapeutic Opportunities in Non-cancer Diseases.
    Matteo Borgini, Emil G Ayala-Cosme, Deborah Yang, Kyara Vences, Lindsey O Davis.
      Nrf2 is a cytoprotective transcription factor that induces the transcription of genes responsible for the cell's response to oxidative stress. While Nrf2 activation has led to the development of clinically relevant therapeutics, the oncogenic role of Nrf2 in the proliferation of cancer cells has underscored the complex nature of Nrf2 and the necessity for the development of Nrf2 inhibitors. Although the application of Nrf2 inhibitors appears limited as anticancer agents, recent studies have begun to pinpoint the impairment of autophagy in diseases as a cellular marker that shifts Nrf2 from a protective to a deleterious state. Therefore, the cytoplasmic accumulation of Nrf2 can lead to the accumulation of lipid hydroperoxides and, ultimately, to ferroptosis. However, some studies aimed at elucidating the role of Nrf2 in non-cancer diseases have yielded conflicting results, attributed to differences in approaches used to inhibit or activate Nrf2, as well as variations in disease models. Overall, these results highlight the necessity for a deeper evaluation of Nrf2's role in diseases, especially chronic diseases. In this review, we discuss diseases where Nrf2 inhibition holds potential for beneficial therapeutic effects and summarize recently reported Nrf2 inhibitors exploiting medicinal chemistry approaches suitable for targeting transcription factors like Nrf2.
    Keywords:  Nrf2 inhibitors * Nuclear factor erythroid 2-related factor * Autophagy * Ferroptosis * Transcription factor
    DOI:  https://doi.org/10.1002/cmdc.202400377
  46. Front Mol Med. 2022 ;2 1005258
    The role of calcium-calpain pathway in hyperthermia.
    Atsushi Enomoto, Takemichi Fukasawa.
      Hyperthermia is a promising anticancer treatment modality. Heat stress stimulates proteolytic machineries to regulate cellular homeostasis. Calpain, an intracellular calcium (Ca2+)-dependent cysteine protease, is a modulator that governs various cellular functions. Hyperthermia induces an increase in cytosolic Ca2+ levels and triggers calpain activation. Contrastingly, pre-exposure of cells to mild hyperthermia induces thermotolerance due to the presence of cellular homeostatic processes such as heat shock response and autophagy. Recent studies suggest that calpain is a potential key molecule that links autophagy and apoptosis. In this review, we briefly introduce the regulation of intracellular Ca2+ homeostasis, basic features of calpains with their implications in cancer, immune responses, and the roles and cross-talk of calpains in cellular protection and cell death in hyperthermia.
    Keywords:  autophagy; calcium; calpain; cell death; hyperthermia
    DOI:  https://doi.org/10.3389/fmmed.2022.1005258
  47. Cancer Biol Ther. 2024 Dec 31. 25(1): 2385517
    Autophagic degradation of CDK4 is responsible for G0/G1 cell cycle arrest in NVP-BEZ235-treated neuroblastoma.
    Zhen Liu, Xiao-Yang Wang, Han-Wei Wang, Shan-Ling Liu, Chao Zhang, Feng Liu, Ying Guo, Feng-Hou Gao.
      BACKGROUND: CDK4 is highly expressed and associated with poor prognosis and decreased survival in advanced neuroblastoma (NB). Targeting CDK4 degradation presents a potentially promising therapeutic strategy compared to conventional CDK4 inhibitors. However, the autophagic degradation of the CDK4 protein and its anti-proliferation effect in NB cells has not been mentioned.RESULTS: We identified autophagy as a new pathway for the degradation of CDK4. Firstly, autophagic degradation of CDK4 is critical for NVP-BEZ235-induced G0/G1 arrest, as demonstrated by the overexpression of CDK4, autophagy inhibition, and blockade of autophagy-related genes. Secondly, we present the first evidence that p62 binds to CDK4 and then enters the autophagy-lysosome to degrade CDK4 in a CTSB-dependent manner in NVP-BEZ235 treated NB cells. Similar results regarding the interaction between p62 and CDK4 were observed in the NVP-BEZ235 treated NB xenograft mouse model.
    CONCLUSIONS: Autophagic degradation of CDK4 plays a pivotal role in G0/G1 cell cycle arrest in NB cells treated with NVP-BEZ235.
    Keywords:  CDK4; NVP-BEZ235; P62; autophagic degradation; neuroblastoma
    DOI:  https://doi.org/10.1080/15384047.2024.2385517
  48. Life Sci Alliance. 2024 Oct;pii: e202402584. [Epub ahead of print]7(10):
    Differential roles of lysosomal cholesterol transporters in the development of C. elegans NMJs.
    Amin Guo, Qi Wu, Xin Yan, Kanghua Chen, Yuxiang Liu, Dingfa Liang, Yuxiao Yang, Qunfeng Luo, Mingtao Xiong, Yong Yu, Erkang Fei, Fei Chen.
      Cholesterol homeostasis in neurons is critical for synapse formation and maintenance. Neurons with impaired cholesterol uptake undergo progressive synapse loss and eventual degeneration. To investigate the molecular mechanisms of neuronal cholesterol homeostasis and its role during synapse development, we studied motor neurons of Caenorhabditis elegans because these neurons rely on dietary cholesterol. Combining lipidomic analysis, we discovered that NCR-1, a lysosomal cholesterol transporter, promotes cholesterol absorption and synapse development. Loss of ncr-1 causes smaller synapses, and low cholesterol exacerbates the deficits. Moreover, NCR-1 deficiency hinders the increase in synapses under high cholesterol. Unexpectedly, NCR-2, the NCR-1 homolog, increases the use of cholesterol and sphingomyelins and impedes synapse formation. NCR-2 deficiency causes an increase in synapses regardless of cholesterol concentration. Inhibiting the degradation or synthesis of sphingomyelins can induce or suppress the synaptic phenotypes in ncr-2 mutants. Our findings indicate that neuronal cholesterol homeostasis is differentially controlled by two lysosomal cholesterol transporters and highlight the importance of neuronal cholesterol homeostasis in synapse development.
    DOI:  https://doi.org/10.26508/lsa.202402584
  49. Acta Neuropathol. 2024 Aug 01. 148(1): 14
    The parkin V380L variant is a genetic modifier of Machado-Joseph disease with impact on mitophagy.
    EUROSCA Network
      Machado-Joseph disease (MJD) is an autosomal dominant neurodegenerative spinocerebellar ataxia caused by a polyglutamine-coding CAG repeat expansion in the ATXN3 gene. While the CAG length correlates negatively with the age at onset, it accounts for approximately 50% of its variability only. Despite larger efforts in identifying contributing genetic factors, candidate genes with a robust and plausible impact on the molecular pathogenesis of MJD are scarce. Therefore, we analysed missense single nucleotide polymorphism variants in the PRKN gene encoding the Parkinson's disease-associated E3 ubiquitin ligase parkin, which is a well-described interaction partner of the MJD protein ataxin-3, a deubiquitinase. By performing a correlation analysis in the to-date largest MJD cohort of more than 900 individuals, we identified the V380L variant as a relevant factor, decreasing the age at onset by 3 years in homozygous carriers. Functional analysis in an MJD cell model demonstrated that parkin V380L did not modulate soluble or aggregate levels of ataxin-3 but reduced the interaction of the two proteins. Moreover, the presence of parkin V380L interfered with the execution of mitophagy-the autophagic removal of surplus or damaged mitochondria-thereby compromising cell viability. In summary, we identified the V380L variant in parkin as a genetic modifier of MJD, with negative repercussions on its molecular pathogenesis and disease age at onset.
    Keywords:   PRKN ; Aggregation; Polyglutamine disease; SCA3; SNP; Spinocerebellar ataxia type 3
    DOI:  https://doi.org/10.1007/s00401-024-02762-6
  50. bioRxiv. 2024 Jul 25. pii: 2024.07.23.604825. [Epub ahead of print]
    V-ATPase Disassembly at the Yeast Lysosome-Like Vacuole Is a Phenotypic Driver of Lysosome Dysfunction in Replicative Aging.
    Fiza Hashmi, Patricia M Kane.
      Declines in lysosomal acidification and function with aging are observed in organisms ranging from yeast to humans. V-ATPases play a central role in organelle acidification and V-ATPase activity is regulated by reversible disassembly in many different settings. Using the yeast Saccharomyces cerevisiae as a replicative aging model, we demonstrate that V-ATPases disassemble into their V 1 and V 0 subcomplexes in aging cells, with release of V 1 subunit C (Vma5) from the lysosome-like vacuole into the cytosol. Disassembly is observed after > 5 cell divisions and results in overall vacuole alkalinization. Caloric restriction, an established mechanism for reversing many age-related outcomes, prevents V-ATPase disassembly in older cells and preserves vacuolar pH homeostasis. Reversible disassembly is controlled in part by the activity of two opposing and conserved factors, the RAVE complex and Oxr1. The RAVE complex promotes V-ATPase assembly and a rav1 Δ mutant shortens replicative lifespan; Oxr1 promotes disassembly and an oxr1 Δ mutation extends lifespan. Importantly, the level of Rav2, a key subunit of the RAVE complex, declines in aged cells. These data indicate that reduced V-ATPase assembly contributes to the loss of lysosome acidification with age, which affects replicative lifespan.
    DOI:  https://doi.org/10.1101/2024.07.23.604825
  51. MedComm (2020). 2024 Aug;5(8): e668
    Autophagy-mediated activation of the AIM2 inflammasome enhances M1 polarization of microglia and exacerbates retinal neovascularization.
    Xianyang Liu, Qian Zhou, Jiayu Meng, Hangjia Zuo, Ruonan Li, Rui Zhang, Huiping Lu, Zhi Zhang, Hongshun Li, Shuhao Zeng, Meng Tian, Hong Wang, Ke Hu, Na Li, Liming Mao, Shengping Hou.
      Retinopathy of prematurity (ROP) is a retinal neovascularization (RNV) disease that is characterized by abnormal blood vessel development in the retina. Importantly, the etiology of ROP remains understudied. We re-analyzed previously published single-cell data and discovered a strong correlation between microglia and RNV diseases, particularly ROP. Subsequently, we found that reactive oxygen species reduced autophagy-dependent protein degradation of absent in melanoma 2 (AIM2) in hypoxic BV2 cells, leading to increased AIM2 protein accumulation. Furthermore, we engineered AIM2 knockout mice and observed that the RNV was significantly reduced compared to wild-type mice. In vitro vascular function assays also demonstrated diminished angiogenic capabilities following AIM2 knockdown in hypoxic BV2 cells. Mechanistically, AIM2 enhanced the M1-type polarization of microglia via the ASC/CASP1/IL-1β pathway, resulting in RNV. Notably, the administration of recombinant protein IL-1β exacerbated angiogenesis, while its inhibition ameliorated the condition. Taken together, our study provides a novel therapeutic target for ROP and offers insight into the interaction between pyroptosis and autophagy.
    Keywords:  AIM2 inflammasomes; autophagy; microglia; retinal angiogenesis; retinopathy of prematurity
    DOI:  https://doi.org/10.1002/mco2.668
  52. World J Gastroenterol. 2024 Jul 21. 30(27): 3273-3277
    Novel insights into autophagy in gastrointestinal pathologies, mechanisms in metabolic dysfunction-associated fatty liver disease and acute liver failure.
    Tsvetelina Velikova, Milena Gulinac.
      In this editorial, we comment on three articles published in a recent issue of World Journal of Gastroenterology. There is a pressing need for new research on autophagy's role in gastrointestinal (GI) disorders, and also novel insights into some liver conditions, such as metabolic dysfunction-associated fatty liver disease (MAFLD) and acute liver failure (ALF). Despite advancements, understanding autophagy's intricate mechanisms and implications in these diseases remains incomplete. Moreover, MAFLD's pathogenesis, encompassing hepatic steatosis and metabolic dysregulation, require further elucidation. Similarly, the mechanisms underlying ALF, a severe hepatic dysfunction, are poorly understood. Innovative studies exploring the interplay between autophagy and GI disorders, as well as defined mechanisms of MAFLD and ALF, are crucial for identifying therapeutic targets and enhancing diagnostic and treatment strategies to mitigate the global burden of these diseases.
    Keywords:  Acute liver failure; Autophagy; Gastrointestinal diseases; High-normal alanine aminotransferase level; Metabolic dysfunction-associated fatty liver disease; Silent information regulator sirtuin 1
    DOI:  https://doi.org/10.3748/wjg.v30.i27.3273
  53. Front Biosci (Landmark Ed). 2024 Jul 22. 29(7): 260
    Advances in Ubiquitination and Proteostasis in Retinal Degeneration.
    Jia Wei, Xiaona Chen, Yingyue Xiong, Yali Gao.
      Retinal degeneration (RD) is a group of chronic blinding diseases characterised by progressive retinal cell death. As the disease progresses, vision deteriorates due to retinal cell death and impaired retinal integrity, eventually leading to complete loss of vision. Therefore, the function and environmental homeostasis of the retina have an important impact on the pathogenesis and treatment of RD. Ubiquitination, as a complex post-translational modification process, plays an essential role in maintaining retinal homeostasis and normal function. It covalently combines ubiquitin with protein through a series of enzyme-mediated reactions, and participates in cell processes such as gene transcription, cell cycle process, DNA repair, apoptosis and immune response. At the same time, it plays a central role in protein degradation. There are two major protein degradation systems in eukaryotic cells: the ubiquitin-proteasome system and the autophagy-lysosomal system. The protein degradation pathway maintains retinal protein homeostasis by reducing abnormal protein accumulation in the retina through two modes of degradation. Either dysregulation of ubiquitination or disruption of protein homeostasis may lead to the development of RD. This article aims to comprehensively review recent research progress on ubiquitin-related genes, proteins and protein homeostasis in the pathogenesis of RD, and to summarize the potential targeted therapy strategies for it. The review is expected to provide valuable guidance for further development and application of ubiquitination in RD.
    Keywords:  autophagy; retinal degeneration; ubiquitin-proteasome system; ubiquitination
    DOI:  https://doi.org/10.31083/j.fbl2907260
  54. Am J Pathol. 2024 Jul 26. pii: S0002-9440(24)00247-5. [Epub ahead of print]
    Lactate contributes to remote ischemic preconditioning-mediated protection against myocardial ischemia reperfusion injury by facilitating autophagy via AMPK-mTOR-TFEB-CX43 axis.
    Zhang-Jian Yang, Wei-Fang Zhang, Qing-Qing Jin, Zhi-Rong Wu, Yun-Yan Du, Hao Shi, Zhen-Sheng Qu, Xiao-Jian Han, Li-Ping Jiang.
      Remote ischemic preconditioning (RIPC) exerts a protective role on myocardial ischemia reperfusion (I/R) injury by the release of various humoral factors. Lactate is a common metabolite in ischemic tissues. Nevertheless, little is known about the role lactate plays in myocardial I/R injury and its underlying mechanism. This investigation revealed that RIPC elevated the level of lactate in blood and myocardium. Furthermore, AZD3965, a selective monocarboxylate transporter 1 (MCT1) inhibitor and 2-Deoxy-D-glucose (2-DG), a glycolysis inhibitor, mitigated the effects of RIPC-induced elevated lactate in the myocardium and prevented RIPC against myocardial I/R injury. In an in vitro hypoxia reoxygenation (H/R) model, lactate markedly mitigated H/R-induced cell damage in H9c2 cells. Meanwhile, further studies suggested that lactate contributed to RIPC rescuing I/R-induced autophagy deficiency by promoting TFEB translocation to the nucleus through activating the AMPK-mTOR pathway without influencing the PI3K-Akt pathway, thus reducing cardiomyocytes damage. Interestingly, we also found that lactate upregulated the mRNA and protein expression of CX43 by facilitating the binding of TFEB to CX43 promoter in the myocardium. Functionally, silencing of TFEB attenuated the protective effect of lactate on cell damage, which was reversed by overexpression of CX43. Further mechanistic studies suggested lactate facilitated CX43-regulated autophagy via AMPK-mTOR-TFEB signaling pathway. Collectively, our research demonstrates that RIPC protects against myocardial I/R injury through lactate-mediated myocardial autophagy via AMPK-mTOR-TFEB-CX43 axis.
    Keywords:  autophagy; connexin 43; lactate; myocardial ischemia reperfusion injury; remote ischemic preconditioning
    DOI:  https://doi.org/10.1016/j.ajpath.2024.07.005
  55. Fish Shellfish Immunol. 2024 Jul 29. pii: S1050-4648(24)00443-1. [Epub ahead of print] 109798
    Calreticulin from Apostichopus japonicus relieves endoplasmic reticulum stress induced by Vibrio splendidus through autophagy.
    Weikang Liang, Lincheng Hu, Fa Dai, Yue Shi, Lei Yang, Chenghua Li.
      When organisms are exposed to external stimuli, misfolded proteins accumulate continuously, resulting in endoplasmic reticulum (ER) stress. Autophagy is of great significance for eliminating aggregated proteins and maintaining cellular homeostasis. However, the molecular mechanism of activating autophagy in response to ER stress in sea cucumber is remain unclear. In the current study, we demonstrated that the pathogen Vibrio splendidus can cause ER stress in Apostichopus japonicus coelomocytes and identified a Ca2+ binding partner calreticulin (designated as AjCRT), which increased with the occurrence of ER stress. The nucleotide sequence analysis showed that the open reading frame of AjCRT was 1242 bp and encoded a 413-amino-acid residue polyprotein with calreticulin domains. The spatial expression analysis revealed that AjCRT was ubiquitously expressed in all examined tissues with large magnitude in the coelomocytes and was minimally expressed in muscle. Furthermore, silencing AjCRT in vivo could significantly exacerbate ER stress induced by V. splendidus and resulted in the significant reduction of coelomocyte autophagy. These findings indicate a calreticulin-based mechanism that positively regulates autophagy in response to ER stress induced by pathogen infection. The results will provide a basis for understanding the way of host alleviating ER stress through autophagy, and pharmacological approaches may have potential for managing ER stress induced by pathogen and related cellular disorders.
    Keywords:  Apostichopus japonicus; Vibrio splendidus; autophagy; calreticulin; endoplasmic reticulum
    DOI:  https://doi.org/10.1016/j.fsi.2024.109798
  56. Neurosci Biobehav Rev. 2024 Jul 30. pii: S0149-7634(24)00306-3. [Epub ahead of print] 105837
    Mitochondrial dynamics and psychiatric disorders: The missing link.
    Maria P Papageorgiou, Michaela D Filiou.
      Elucidating the molecular mechanisms of psychopathology is crucial for optimized diagnosis and treatment. Accumulating literature has underlined how mitochondrial bioenergetics affect major psychiatric disorders. However, how mitochondrial dynamics, a term addressing mitochondria quality control, including mitochondrial fission, fusion, biogenesis and mitophagy, is implicated in psychopathologies remains elusive. In this review we summarize the existing literature on mitochondrial dynamics perturbations in psychiatric disorders/neuropsychiatric phenotypes. We include preclinical/clinical literature on mitochondrial dynamics recalibrations in anxiety, depression, post-traumatic stress disorder (PTSD), bipolar disorder and schizophrenia. We discuss alterations in mitochondrial network, morphology and shape; molecular markers of the mitochondrial dynamics machinery and mitochondrial DNA copy number (mtDNAcn) in animal models and human cohorts in brain and peripheral material. By looking for common altered mitochondrial dynamics patterns across diagnoses/phenotypes, we highlight mitophagy and biogenesis as regulators of anxiety and depression pathophysiology, respectively, as well as the fusion mediator dynamin-like 120kDa protein (Opa1) as a molecular hub contributing to psychopathology. Finally, we comment on limitations and future directions in this novel neuropsychiatry field.
    Keywords:  OPA1; anxiety; mitochondria; mitochondrial dynamics; mitochondrial quality; mitophagy; psychiatric disorders
    DOI:  https://doi.org/10.1016/j.neubiorev.2024.105837
  57. Traffic. 2024 Jul;25(7): e12952
    SNX32 Regulates Sorting and Trafficking of Activated EGFR to the Lysosomal Degradation Pathway.
    Dou Wang, Xia Zhao, Panpan Wang, Jia-Jia Liu.
      SNX32 is a member of the evolutionarily conserved Phox (PX) homology domain- and Bin/Amphiphysin/Rvs (BAR) domain- containing sorting nexin (SNX-BAR) family of proteins, which play important roles in sorting and membrane trafficking of endosomal cargoes. Although SNX32 shares the highest amino acid sequence homology with SNX6, and has been believed to function redundantly with SNX5 and SNX6 in retrieval of the cation-independent mannose-6-phosphate receptor (CI-MPR) from endosomes to the trans-Golgi network (TGN), its role(s) in intracellular protein trafficking remains largely unexplored. Here, we report that it functions in parallel with SNX1 in mediating epidermal growth factor (EGF)-stimulated postendocytic trafficking of the epidermal growth factor receptor (EGFR). Moreover, SNX32 interacts directly with EGFR, and recruits SNX5 to promote sorting of EGF-EGFR into multivesicular bodies (MVBs) for lysosomal degradation. Thus, SNX32 functions distinctively from other SNX-BAR proteins to mediate signaling-coupled endolysosomal trafficking of EGFR.
    Keywords:  EGFR; SNX32; SNX5; endolysosomal trafficking; lysosomal degradation; recycling
    DOI:  https://doi.org/10.1111/tra.12952
  58. In Vitro Cell Dev Biol Anim. 2024 Aug 02.
    Arf1 promotes porcine intestinal epithelial cell proliferation via the mTORC1 signaling pathway.
    Yong-Xia Fang, En-Qing Lu, E Xu, Yi-Yu Zhang, Min Zhu.
      The promotion of gut health, a pervasive problem in modern animal husbandry, positively affects organismal health, productivity, and economics. Porcine intestinal epithelial cells (IPEC-J2) continuously proliferate to maintain intestinal homeostasis, including barrier, immune, and absorptive functions. Gut homeostasis is fundamental to organismal health. ADP-ribosylation factor 1 (Arf1), a small GTPase, plays a crucial role in coordinating mTORC1 in response to nutrients, especially amino acid availability in the gut. mTORC1 is the central hub of proliferation. Thus, it seems likely that Arf1 promotes IPEC-J2 cell proliferation. However, the exact role of Arf1 in the porcine gut remains unclear. Therefore, we evaluated the functional role and possible mechanisms of Arf1 in the porcine intestine through Arf1 overexpression and knockdown in IPEC-J2 cells. Arf1 overexpression and knockdown significantly enhanced and inhibited, respectively, IPEC-J2 cell viability, and PCNA expression varied with Arf1 expression. Moreover, the proportion of Ki67-positive cells was significantly greater in the Arf1-overexpressing group than in the control group. These results suggest that Arf1 improves IPEC-J2 cell proliferation. The underlying mechanism was explored by Western blotting. Arf1 overexpression and knockdown significantly enhanced and suppressed, respectively, the levels of p-S6K1 and p-RPS6, which are key downstream targets of the mTORC1 signaling pathway. Collectively, our findings reveal the role of the Arf1-mTORC1 axis in IPEC-J2 cell proliferation and its potential function in regulating intestinal homeostasis and health.
    Keywords:  ADP-ribosylation factor 1; Cell proliferation; Intestinal development; Intestinal epithelial cells; mTORC1
    DOI:  https://doi.org/10.1007/s11626-024-00942-y
  59. J Neuroinflammation. 2024 Aug 02. 21(1): 191
    Targeting the immunoproteasome in hypothalamic neurons as a novel therapeutic strategy for high-fat diet-induced obesity and metabolic dysregulation.
    Nicolás Albornoz, Javiera Álvarez-Indo, Adely de la Peña, Eloisa Arias-Muñoz, Alanis Coca, Fabián Segovia-Miranda, Bredford Kerr, Mauricio Budini, Alfredo Criollo, María A García-Robles, Eugenia Morselli, Andrea Soza, Patricia V Burgos.
      OBJECTIVE: Obesity represents a significant global health challenge characterized by chronic low-grade inflammation and metabolic dysregulation. The hypothalamus, a key regulator of energy homeostasis, is particularly susceptible to obesity's deleterious effects. This study investigated the role of the immunoproteasome, a specialized proteasomal complex implicated in inflammation and cellular homeostasis, during metabolic diseases.METHODS: The levels of the immunoproteasome β5i subunit were analyzed by immunostaining, western blotting, and proteasome activity assay in mice fed with either a high-fat diet (HFD) or a regular diet (CHOW). We also characterized the impact of autophagy inhibition on the levels of the immunoproteasome β5i subunit and the activation of the AKT pathway. Finally, through confocal microscopy, we analyzed the contribution of β5i subunit inhibition on mitochondrial function by flow cytometry and mitophagy assay.
    RESULTS: Using an HFD-fed obese mouse model, we found increased immunoproteasome levels in hypothalamic POMC neurons. Furthermore, we observed that palmitic acid (PA), a major component of saturated fats found in HFD, increased the levels of the β5i subunit of the immunoproteasome in hypothalamic neuronal cells. Notably, the increase in immunoproteasome expression was associated with decreased autophagy, a critical cellular process in maintaining homeostasis and suppressing inflammation. Functionally, PA disrupted the insulin-glucose axis, leading to reduced AKT phosphorylation and increased intracellular glucose levels in response to insulin due to the upregulation of the immunoproteasome. Mechanistically, we identified that the protein PTEN, a key regulator of insulin signaling, was reduced in an immunoproteasome-dependent manner. To further investigate the potential therapeutic implications of these findings, we used ONX-0914, a specific immunoproteasome inhibitor. We demonstrated that this inhibitor prevents PA-induced insulin-glucose axis imbalance. Given the interplay between mitochondrial dysfunction and metabolic disturbances, we explored the impact of ONX-0914 on mitochondrial function. Notably, ONX-0914 preserved mitochondrial membrane potential and attenuated mitochondrial ROS production in the presence of PA. Moreover, we found that ONX-0914 reduced mitophagy in the presence of PA.
    CONCLUSIONS: Our findings strongly support the pathogenic involvement of the immunoproteasome in hypothalamic neurons in the context of HFD-induced obesity and metabolic disturbances. Targeting the immunoproteasome highlights a promising therapeutic strategy to mitigate the detrimental effects of obesity on the insulin-glucose axis and cellular homeostasis. This study provides valuable insights into the mechanisms driving obesity-related metabolic diseases and offers potential avenues for developing novel therapeutic interventions.
    Keywords:  Autophagy; Hypothalamus; Immunoproteasome; Insulin-glucose axis; Metabolic disturbances; Mitochondrial function; Neurons; Obesity; Redox biology
    DOI:  https://doi.org/10.1186/s12974-024-03154-z
  60. Eur J Pharmacol. 2024 Jul 31. pii: S0014-2999(24)00546-6. [Epub ahead of print] 176857
    Molecular mechanisms of PTEN in atherosclerosis: A comprehensive review.
    Yuan Tian, Yi-Fan Liu, Yan-Yue Wang, Yong-Zhen Li, Wen-Yan Ding, Chi Zhang.
      Atherosclerosis is a chronic inflammatory disease of the arterial wall caused by an imbalance of lipid metabolism and a maladaptive inflammatory response. A variety of harmful cellular changes associated with atherosclerosis include endothelial dysfunction, the migration of circulating inflammatory cells to the arterial wall, the production of proinflammatory cytokines, lipid buildup in the intima, local inflammatory responses in blood vessels, atherosclerosis-associated apoptosis, and autophagy. PTEN inhibits the phosphatidylinositol 3-kinase (PI3K)/protein kinase B (PKB/AKT)/mammalian target of rapamycin (mTOR) pathway through its lipid phosphatase activity. Previous studies have shown that PTEN is closely related to atherosclerosis. This article reviews the role of PTEN in atherosclerosis from the perspectives of autophagy, apoptosis, inflammation, proliferation, and angiogenesis.
    Keywords:  Apoptosis; Autophagy; Inflammatory; PTEN; Proliferation; atherosclerosis
    DOI:  https://doi.org/10.1016/j.ejphar.2024.176857
  61. Acta Biochim Biophys Sin (Shanghai). 2024 Aug 01.
    FOXM1 mediates methotrexate resistance in osteosarcoma cells by promoting autophagy.
    Luoyang Wang, Dongchang Zhai, Lei Tang, Hui Zhang, Xinlong Wang, Ning Ma, Xiaoyue Zhang, Mingguo Cheng, Ruowu Shen.
      Osteosarcoma (OS) is a primary bone cancer mostly found in adolescents and elderly individuals. The treatment of OS is still largely dependent on traditional chemotherapy. However, the high incidence of drug resistance remains one of the greatest impediments to limiting improvements in OS treatment. Recent findings have indicated that the transcription factor FOXM1 plays an important role in various cancer-related events, especially drug resistance. However, the possible role of FOXM1 in the resistance of OS to methotrexate (MTX) remains to be explored. Here, we find that FOXM1, which confers resistance to MTX, is highly expressed in OS tissues and MTX-resistant cells. FOXM1 overexpression promotes MTX resistance by enhancing autophagy in an HMMR/ATG7-dependent manner. Importantly, silencing of FOXM1 or inhibiting autophagy reverses drug resistance. These findings demonstrate a new mechanism for FOXM1-induced MTX resistance and provide a promising target for improving OS chemotherapy outcomes.
    Keywords:  FOXM1; drug resistance; methotrexate; osteosarcoma
    DOI:  https://doi.org/10.3724/abbs.2024084
  62. Res Sq. 2024 Jul 15. pii: rs.3.rs-4569999. [Epub ahead of print]
    Loss of Tuberous Sclerosis Complex 2 confers inflammation via dysregulation of Nuclear factor kappa-light-chain-enhancer of activated B cells.
    Darius K McPhail, Mohammad A M Alzahrani, Katie R Martin, Brian L Calver, Adrian J Harwood, Jeffrey P MacKeigan, David M Davies, Andrew R Tee.
      Background: Aberrant activation of mTORC1 is clearly defined in TSC, causing uncontrolled cell growth. While mTORC1 inhibitors show efficacy to stabilise tumour growth in TSC, they are not fully curative. Disease facets of TSC that are not restored with mTOR inhibitors might involve NF-κB. The study aimed to characterise NF-κB in the context of TSC.Results: Enrichment of NF-κB-regulated genes was observed in TSC patient tumours, SEN/SEGAs, cortical tubers and a TSC tumour-derived cell line (621 - 101). Highlighting an inflammatory component of TSC, TSC cell models showed an elevated level of NF-κB and STAT3 activation. Herein, we report a dysregulated inflammatory phenotype of TSC2-deficient cells where NF-κB promotes autocrine signalling involving IL-6. Of importance, mTORC1 inhibition does not block this inflammatory signal to promote STAT3, while NF-κB inhibition was much more effective. Combined mTORC1 and NF-κB inhibition was potent at preventing anchorage-independent growth of TSC2-deficient cells, and unlike mTORC1 inhibition alone was sufficient to prevent colony regrowth after cessation of treatment.
    Conclusion: This study reveals autocrine signalling crosstalk between NF-κB and STAT3 in TSC cell models. Furthermore, the data presented indicate that NF-κB pathway inhibitors could be a viable adjunct therapy with the current mTOR inhibitors to treat TSC.
    Keywords:  IL-6; NF-κB; STAT3; TSC; inflammation; mTOR; rapamycin
    DOI:  https://doi.org/10.21203/rs.3.rs-4569999/v1
  63. Chem Biol Interact. 2024 Jul 26. pii: S0009-2797(24)00312-0. [Epub ahead of print] 111166
    SS-31 Mitigates Oxidative Stress and Restores Mitochondrial Function in Cigarette Smoke-Damaged Oral Epithelial Cells via PINK1-mediated Mitophagy.
    Pei Ye, Hong Liu, Yao Qin, Zhiyuan Li, Zhuwei Huang, Xiangwen Bu, Qiao Peng, Ning Duan, Wenmei Wang, Xiang Wang.
      Smoking is a well-established risk factor for several oral diseases, including oral cancer, oral leukoplakia and periodontitis, primarily related to reactive oxygen species (ROS). SS-31, a mitochondria-targeting tetrapeptide, has exhibited demonstrable efficacy in medical conditions by attenuating mitochondrial ROS production. However, its potential in the treatment of oral diseases remains underexplored. The aim of this study was to investigate the therapeutic potential of SS-31 in mitigating smoking-induced oral epithelial injury. Through in vitro experiments, our results indicate that SS-31 plays a protective role against cigarette smoke extract (CSE) by reducing oxidative stress, attenuating inflammatory response, and restoring mitochondrial function. Furthermore, we found that mitophagy, regulated by PINK1 (PTEN-induced putative kinase 1)/Parkin (Parkin RBR E3 ubiquitin-protein ligase), was critical for the protective role of SS-31. Our findings offer valuable insights into SS-31's therapeutic potential in mitigating CSE-induced oxidative stress, inflammatory response, and mitochondrial dysfunction in oral epithelial cells. This study provides novel intervention targets for smoking-related oral diseases.
    Keywords:  Cigarette smoke; Mitochondria; Oral epithelial injury; Oxidative stress; PINK1; SS-31
    DOI:  https://doi.org/10.1016/j.cbi.2024.111166
  64. iScience. 2024 Aug 16. 27(8): 110432
    Targeted dephosphorylation of TFEB promotes its nuclear translocation.
    Jin-Feng Zhao, Natalia Shpiro, Gajanan Sathe, Abigail Brewer, Thomas J Macartney, Nicola T Wood, Florentina Negoita, Kei Sakamoto, Gopal P Sapkota.
      Reversible phosphorylation of the transcription factor EB (TFEB) coordinates cellular responses to metabolic and other stresses. During nutrient replete and stressor-free conditions, phosphorylated TFEB is primarily localized to the cytoplasm. Stressor-mediated reduction of TFEB phosphorylation promotes its nuclear translocation and context-dependent transcriptional activity. In this study, we explored targeted dephosphorylation of TFEB as an approach to activate TFEB in the absence of nutrient deprivation or other cellular stress. Through an induction of proximity between TFEB and several phosphatases using the AdPhosphatase system, we demonstrate targeted dephosphorylation of TFEB in cells. Furthermore, by developing a heterobifunctional molecule BDPIC (bromoTAG-dTAG proximity-inducing chimera), we demonstrate targeted dephosphorylation of TFEB-dTAG through induced proximity to bromoTAG-PPP2CA. Targeted dephosphorylation of TFEB-dTAG by bromoTAG-PPP2CA with BDPIC at the endogenous levels is sufficient to induce nuclear translocation and some transcriptional activity of TFEB.
    Keywords:  Health sciences; Medical specialty; Medicine; Precision medicine
    DOI:  https://doi.org/10.1016/j.isci.2024.110432
  65. Proc Natl Acad Sci U S A. 2024 Aug 06. 121(32): e2403114121
    Capturing the catalytic intermediates of parkin ubiquitination.
    Elizabeth M Connelly, Anne C Rintala-Dempsey, Mehmet Gundogdu, E Aisha Freeman, Joanna Koszela, Jacob D Aguirre, Grace Zhu, Outi Kämäräinen, Roya Tadayon, Helen Walden, Gary S Shaw.
      Parkin is an E3 ubiquitin ligase implicated in early-onset forms of Parkinson's disease. It catalyzes a transthiolation reaction by accepting ubiquitin (Ub) from an E2 conjugating enzyme, forming a short-lived thioester intermediate, and transfers Ub to mitochondrial membrane substrates to signal mitophagy. A major impediment to the development of Parkinsonism therapeutics is the lack of structural and mechanistic detail for the essential, short-lived transthiolation intermediate. It is not known how Ub is recognized by the catalytic Rcat domain in parkin that enables Ub transfer from an E2~Ub conjugate to the catalytic site and the structure of the transthiolation complex is undetermined. Here, we capture the catalytic intermediate for the Rcat domain of parkin in complex with ubiquitin (Rcat-Ub) and determine its structure using NMR-based chemical shift perturbation experiments. We show that a previously unidentified α-helical region near the Rcat domain is unmasked as a recognition motif for Ub and guides the C-terminus of Ub toward the parkin catalytic site. Further, we apply a combination of guided AlphaFold modeling, chemical cross-linking, and single turnover assays to establish and validate a model of full-length parkin in complex with UbcH7, its donor Ub, and phosphoubiquitin, trapped in the process of transthiolation. Identification of this catalytic intermediate and orientation of Ub with respect to the Rcat domain provides important structural insights into Ub transfer by this E3 ligase and explains how the previously enigmatic Parkinson's pathogenic mutation T415N alters parkin activity.
    Keywords:  NMR; Parkinson’s disease; catalysis; parkin; protein structure
    DOI:  https://doi.org/10.1073/pnas.2403114121
  66. Open Life Sci. 2024 ;19(1): 20220922
    Metformin mitigates osteoarthritis progression by modulating the PI3K/AKT/mTOR signaling pathway and enhancing chondrocyte autophagy.
    Tianjie Xu, Kainan Liu, Jiaxin Fan, Xiang Jia, Xiaoling Guo, Xingwang Zhao, Yanhua Cao, Hui Zhang, Qian Wang.
      Osteoarthritis (OA) is a chronic degenerative disease characterized by overall joint tissue damage. Metformin (Met) has been shown to inhibit inflammatory reactions, though its potential protective mechanism on cartilage remains unclear. This study investigated Met's potential to protect cartilage in an OA rat model. Various morphological experiments were conducted to assess changes in cartilage tissue morphology before and after Met treatment. Protein and mRNA levels of cartilage-specific genes were measured using western blot, immunohistochemical staining, and RT-qPCR. Additionally, protein levels of autophagy-related and mTOR pathway-related proteins were measured. The results indicate an imbalance in the synthesis and degradation metabolism of chondrocytes, downregulation of cellular autophagy, and activation of the PI3K/Akt/mTOR pathway after surgery. However, treatment with Met could upregulate the expression of synthetic metabolic factors, indicating its contribution to cartilage repair. Furthermore, analysis of autophagy and pathway protein levels indicated that Met effectively attenuated autophagic damage to osteoarthritic cartilage cells and abnormal activation of the PI3K/Akt/mTOR pathway. In conclusion, Met can inhibit the abnormal activation of the PI3K/AKT/mTOR signaling pathway in cartilage tissue, promote the restoration of cartilage cell autophagic function, improve the balance of cartilage cell synthesis and degradation metabolism, and thus exert a protective effect on rat joint cartilage.
    Keywords:  Met; PI3K/AKT/mTOR; autophagy; cartilage; osteoarthritis
    DOI:  https://doi.org/10.1515/biol-2022-0922
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