bims-auttor Biomed News
on Autophagy and mTOR
Issue of 2023–07–02
fifty-six papers selected by
Viktor Korolchuk, Newcastle University
  1. Microglial cytokines poison neuronal autophagy via CCR5, a druggable target.
  2. The Pathological Mechanism of Neuronal Autophagy-Lysosome Dysfunction After Ischemic Stroke.
  3. Cellular Recycling Gone Wrong: The Role of Dysregulated Autophagy and Hyperactive mTORC1 in the Pathogenesis of Sarcoidosis.
  4. mLST8 is essential for coronavirus replication and regulates its replication through the mTORC1 pathway.
  5. Autophagy in BRAF-mutant cutaneous melanoma: recent advances and therapeutic perspective.
  6. A Novel Bioactive Peptide, T14, Selectively Activates mTORC1 Signalling: Therapeutic Implications for Neurodegeneration and Other Rapamycin-Sensitive Applications.
  7. An ATG12-ATG5-TECPR1 E3-like complex regulates unconventional LC3 lipidation at damaged lysosomes.
  8. The new insights into autophagy in thyroid cancer progression.
  9. The N-degron pathway mediates lipophagy: The chemical modulation of lipophagy in obesity and NAFLD.
  10. SHIP1 modulates antimalarial immunity by bridging the crosstalk between type I IFN signaling and autophagy.
  11. mTOR in programmed cell death and its therapeutic implications.
  12. TRIM16-mediated lysophagy suppresses high-gluocse-accumulated neuronal Aβ.
  13. Starvation Protects Hepatocytes from Inflammatory Damage through Paradoxical mTORC1 Signaling.
  14. Inhibition of autophagy; an opportunity for the treatment of cancer resistance.
  15. A conserved STRIPAK complex is required for autophagy in muscle tissue.
  16. Targeted degradation of ⍺-synuclein aggregates in Parkinson's disease using the AUTOTAC technology.
  17. Comprehensive analysis of autophagic functions of WIPI family proteins and their implications for the pathogenesis of β-propeller associated neurodegeneration.
  18. Lysosomal LAMP proteins regulate lysosomal pH by direct inhibition of the TMEM175 channel.
  19. Targeting SIRT1-regulated autophagic cell death as a novel therapeutic avenue for cancer prevention.
  20. ULK phosphorylation of STX17 controls autophagosome maturation via FLNA.
  21. The landscape of mitophagy in sepsis reveals PHB1 as an NLRP3 inflammasome inhibitor.
  22. Autophagy, a double-edged sword for oral tissue regeneration.
  23. The ACSL4 Network Regulates Cell Death and Autophagy in Diseases.
  24. An Expansion of the Endoplasmic Reticulum that Halts Autophagy is Permissive to Genome Instability.
  25. α-Synuclein Interactions in Mitochondria-ER Contacts: A Possible Role in Parkinson's Disease.
  26. Alterations of Autophagy Modify Lipids in Epidermal Keratinocytes.
  27. Unveiling the impact of lysosomal ion channels: balancing ion signaling and disease pathogenesis.
  28. Transcriptional and Post-translational Regulation of Plant Autophagy.
  29. A selective autophagy receptor VISP1 induces symptom recovery by targeting viral silencing suppressors.
  30. The Role of Retinal Pigment Epithelial Cells in Age-Related Macular Degeneration: Phagocytosis and Autophagy.
  31. Autotaxin promotes the degradation of the mucus layer by inhibiting autophagy in mouse colitis.
  32. Differences in the Intracellular Localization of Methylated β-Cyclodextrins-Threaded Polyrotaxanes Lead to Different Cellular States.
  33. Biophysical Aspects of Neurodegenerative and Neurodevelopmental Disorders Involving Endo-/Lysosomal CLC Cl-/H+ Antiporters.
  34. The Janus-Faced Role of Lipid Droplets in Aging: Insights from the Cellular Perspective.
  35. PINK1: From Parkinson's disease to mitophagy and back again.
  36. Analysis of LC3-Associated Phagocytosis and Antigen Presentation in Macrophages and B Cells.
  37. Apilimod activates the NLRP3 inflammasome through lysosome-mediated mitochondrial damage.
  38. Polyamines and Physical Activity in Musculoskeletal Diseases: A Potential Therapeutic Challenge.
  39. Oxidative Stress-Involved Mitophagy of Retinal Pigment Epithelium and Retinal Degenerative Diseases.
  40. mTOR Inhibition Is Effective against Growth, Survival and Migration, but Not against Microglia Activation in Preclinical Glioma Models.
  41. Essential amino acid starvation induces cell cycle arrest, autophagy, and inhibits osteogenic differentiation in murine osteoblast.
  42. Toxoplasma gondii inhibits the expression of autophagy-related genes through AKT-dependent inactivation of the transcription factor FOXO3a.
  43. Follicular lymphoma-associated mutations in the V-ATPase chaperone Vma21 activate autophagy by dysfunctional V-ATPase assembly.
  44. A Yeast Mitotic Tale for the Nucleus and the Vacuoles to Embrace.
  45. mTORC2-NDRG1-CDC42 axis couples fasting to mitochondrial fission.
  46. Mitochondrial dysfunction and mitophagy defects in LRRK2-R1441C Parkinson's disease models.
  47. Elevated levels of FMRP-target MAP1B impair human and mouse neuronal development and mouse social behaviors via autophagy pathway.
  48. Novel phosphorylation of Rab29 that regulates its localization and lysosomal stress response in concert with LRRK2.
  49. Autophagy Activation Promoted by Pulses of Light and Phytochemicals Counteracting Oxidative Stress during Age-Related Macular Degeneration.
  50. Lactiplantibacillus plantarum OLL2712 Induces Autophagy via MYD88 and Strengthens Tight Junction Integrity to Promote the Barrier Function in Intestinal Epithelial Cells.
  51. UHRF2 promotes the malignancy of hepatocellular carcinoma by PARP1 mediated autophagy.
  52. Using Zebrafish to Dissect the Interaction of Mycobacteria with the Autophagic Machinery in Macrophages.
  53. Acyl coenzyme A binding protein (ACBP): An aging- and disease-relevant "autophagy checkpoint".
  54. Reprogramming tumour-associated macrophages to outcompete cancer cells.
  55. The active zone protein CLA-1 (Clarinet) bridges two subsynaptic domains to regulate presynaptic sorting of ATG-9.
  56. A Novel Small NPC1 Promoter Enhances AAV-Mediated Gene Therapy in Mouse Models of Niemann-Pick Type C1 Disease.
  1. Autophagy. 2023 Jun 26. 1-3
    Microglial cytokines poison neuronal autophagy via CCR5, a druggable target.
    Beatrice Paola Festa, Farah H Siddiqi, Maria Jimenez-Sanchez, David C Rubinsztein.
      In the prodromal phase of neurodegenerative diseases, microglia switch to an activated state resulting in increased secretion of pro-inflammatory factors. We reported that C - C chemokine ligand 3 (CCL3), C - C chemokine ligand 4 (CCL4) and C - C chemokine ligand 5 (CCL5) contained in the secretome of activated microglia inhibit neuronal autophagy via a non-cell autonomous mechanism. These chemokines bind and activate neuronal C - C chemokine receptor type 5 (CCR5), which, in turn, promotes phosphoinositide 3-kinase (PI3K) - protein kinase B (PKB, or AKT) - mammalian target of rapamycin complex 1 (mTORC1) pathway activation, which inhibits autophagy, thus causing the accumulation of aggregate-prone proteins in the cytoplasm of neurons. The levels of CCR5 and its chemokine ligands are increased in the brains of pre-manifesting Huntington disease (HD) and tauopathy mouse models. CCR5 accumulation might be due to a self-amplifying mechanism, since CCR5 is a substrate of autophagy and CCL5-CCR5-mediated autophagy inhibition impairs CCR5 degradation. Furthermore, pharmacological, or genetic inhibition of CCR5 rescues mTORC1-autophagy dysfunction and improves neurodegeneration in HD and tauopathy mouse models, suggesting that CCR5 hyperactivation is a pathogenic signal driving the progression of these diseases.
    Keywords:  CCR5; Huntington disease; Tau; autophagy; maraviroc; microglia
    DOI:  https://doi.org/10.1080/15548627.2023.2221921
  2. Cell Mol Neurobiol. 2023 Jun 29.
    The Pathological Mechanism of Neuronal Autophagy-Lysosome Dysfunction After Ischemic Stroke.
    Guang-Sen Shi, Qi-Lin Qin, Cheng Huang, Zi-Rong Li, Zi-Han Wang, Yong-Yan Wang, Xiu-Ying He, Xiao-Ming Zhao.
      The abnormal initiation of autophagy flux in neurons after ischemic stroke caused dysfunction of autophagy-lysosome, which not only led to autophagy flux blockage, but also resulted in autophagic death of neurons. However, the pathological mechanism of neuronal autophagy-lysosome dysfunction did not form a unified viewpoint until now. In this review, taking the autophagy lysosomal dysfunction of neurons as a starting point, we summarized the molecular mechanisms that led to neuronal autophagy lysosomal dysfunction after ischemic stroke, which would provide theoretical basis for the clinical treatment of ischemic stroke.
    Keywords:  Autophagy flux; Ischemic stroke; Neuronal autophagy-lysosome dysfunction
    DOI:  https://doi.org/10.1007/s10571-023-01382-0
  3. Sarcoidosis Vasc Diffuse Lung Dis. 2023 Jun 29. 40(2): e2023016
    Cellular Recycling Gone Wrong: The Role of Dysregulated Autophagy and Hyperactive mTORC1 in the Pathogenesis of Sarcoidosis.
    Jennifer Adouli, Aaron Fried, Rachel Swier, Andrew Ghio, Irina Petrache, Stephen Tilley.
       BACKGROUND AND AIMS: Autophagy is a highly regulated, complex intracellular recycling process that is vital to maintaining cellular homeostasis in response to diverse conditions and stressors. Despite the presence of robust regulatory pathways, the intricate and multi-step nature of autophagy creates opportunity for dysregulation. Errors in autophagy have been implicated in the development of a broad range of clinical pathologies including granulomatous disease. Specifically, activation of the mTORC1 pathway has been identified as a key negative regulator of autophagic flux, prompting the study of dysregulated mTORC1 signaling in the pathogenesis of sarcoidosis. Our review: We conducted a thorough search of the extant literature to identify the regulatory pathways of autophagy, and more specifically the implication of upregulated mTORC1 pathways in the pathogenesis of sarcoidosis. We review data showing spontaneous granuloma formation in animal models with upregulate mTORC1 signaling, human genetic studies showing mutation in autophagy genes in sarcoidosis patients, and clinical data showing that targeting autophagy regulatory molecules like mTORC1 may provide new therapeutic approaches for sarcoidosis.
    CONCLUSIONS: Given the incomplete understanding of sarcoidosis pathogenesis and the toxicities of current treatments, a more complete understanding of sarcoidosis pathogenesis is crucial for the development of more effective and safer therapies. In this review, we propose a strong molecular pathway driving sarcoidosis pathogenesis at which autophagy is at the center. A more complete understanding of autophagy and its regulatory molecules, like mTORC1, may provide a window into new therapeutic approaches for sarcoidosis.
    DOI:  https://doi.org/10.36141/svdld.v40i2.13498
  4. mBio. 2023 Jun 28. e0089923
    mLST8 is essential for coronavirus replication and regulates its replication through the mTORC1 pathway.
    Yanan Fu, Zhen Fu, Zhelin Su, Lisha Li, Yilin Yang, Yubei Tan, Yixin Xiang, Yuejun Shi, Shengsong Xie, Limeng Sun, Guiqing Peng.
      Coronaviruses (CoVs), which pose a serious threat to human and animal health worldwide, need to hijack host factors to complete their replicative cycles. However, the current study of host factors involved in CoV replication remains unknown. Here, we identified a novel host factor, mammalian lethal with sec-13 protein 8 (mLST8), which is a common subunit of mTOR complex 1 (mTORC1) and mTOR complex 2 (mTORC2), and is critical for CoV replication. Inhibitor and knockout (KO) experiments revealed that mTORC1, but not mTORC2, is essential for transmissible gastroenteritis virus replication. Furthermore, mLST8 KO reduced the phosphorylation of unc-51-like kinase 1 (ULK1), a factor downstream of the mTORC1 signaling pathway, and mechanistic studies revealed that decreased phosphorylation of the mTORC1 downstream factor ULK1 promoted the activation of autophagy, which is responsible for antiviral replication in mLST8 KO cells. Then, transmission electron microscopy indicated that both mLST8 KO and autophagy activator inhibited the formation of double-membrane vesicles in early viral replication. Finally, mLST8 KO and autophagy activator treatment could also inhibit the replication of other CoVs, indicating a conserved relationship between autophagy activation and CoV replication. In summary, our work reveals that mLST8 is a novel host regulator of CoV replication, which provides new insights into the mechanism of CoV replication and can facilitate the development of broad-spectrum antiviral drugs. IMPORTANCE CoVs are highly variable, and existing CoV vaccines are still limited in their ability to address mutations in CoVs. Therefore, the need to improve our understanding of the interaction of CoVs with the host during viral replication and to find targets for drugs against CoVs is urgent. Here, we found that a novel host factor, mLST8, is critical for CoV infection. Further studies showed that mLST8 KO inhibited the mTORC1 signaling pathway, and we found that autophagy activation downstream of mTORC1 was the main cause of antiviral replication in mLST8 KO cells. Autophagy activation impaired the formation of DMVs and inhibited early viral replication. These findings deepen our understanding of the CoV replication process and provide insights into potential therapeutic applications.
    Keywords:  autophagy; coronavirus; mLST8; mTORC1; replication
    DOI:  https://doi.org/10.1128/mbio.00899-23
  5. Cell Death Discov. 2023 Jun 29. 9(1): 202
    Autophagy in BRAF-mutant cutaneous melanoma: recent advances and therapeutic perspective.
    Elisabetta Fratta, Giorgio Giurato, Roberto Guerrieri, Francesca Colizzi, Jessica Dal Col, Alessandro Weisz, Agostino Steffan, Barbara Montico.
      Macroautophagy, hereafter referred to as autophagy, represents a highly conserved catabolic process that maintains cellular homeostasis. At present, the role of autophagy in cutaneous melanoma (CM) is still controversial, since it appears to be tumor-suppressive at early stages of malignant transformation and cancer-promoting during disease progression. Interestingly, autophagy has been found to be often increased in CM harboring BRAF mutation and to impair the response to targeted therapy. In addition to autophagy, numerous studies have recently conducted in cancer to elucidate the molecular mechanisms of mitophagy, a selective form of mitochondria autophagy, and secretory autophagy, a process that facilitates unconventional cellular secretion. Although several aspects of mitophagy and secretory autophagy have been investigated in depth, their involvement in BRAF-mutant CM biology has only recently emerged. In this review, we aim to overview autophagy dysregulation in BRAF-mutant CM, along with the therapeutic advantages that may arise from combining autophagy inhibitors with targeted therapy. In addition, the recent advances on mitophagy and secretory autophagy involvement in BRAF-mutant CM will be also discussed. Finally, since a number of autophagy-related non-coding RNAs (ncRNAs) have been identified so far, we will briefly discussed recent advances linking ncRNAs to autophagy regulation in BRAF-mutant CM.
    DOI:  https://doi.org/10.1038/s41420-023-01496-w
  6. Int J Mol Sci. 2023 Jun 09. pii: 9961. [Epub ahead of print]24(12):
    A Novel Bioactive Peptide, T14, Selectively Activates mTORC1 Signalling: Therapeutic Implications for Neurodegeneration and Other Rapamycin-Sensitive Applications.
    Sanskar Ranglani, Anna Ashton, Kashif Mahfooz, Joanna Komorowska, Alexandru Graur, Nadine Kabbani, Sara Garcia-Rates, Susan Greenfield.
      T14 modulates calcium influx via the α-7 nicotinic acetylcholine receptor to regulate cell growth. Inappropriate triggering of this process has been implicated in Alzheimer's disease (AD) and cancer, whereas T14 blockade has proven therapeutic potential in in vitro, ex vivo and in vivo models of these pathologies. Mammalian target of rapamycin complex 1 (mTORC1) is critical for growth, however its hyperactivation is implicated in AD and cancer. T14 is a product of the longer 30mer-T30. Recent work shows that T30 drives neurite growth in the human SH-SY5Y cell line via the mTOR pathway. Here, we demonstrate that T30 induces an increase in mTORC1 in PC12 cells, and ex vivo rat brain slices containing substantia nigra, but not mTORC2. The increase in mTORC1 by T30 in PC12 cells is attenuated by its blocker, NBP14. Moreover, in post-mortem human midbrain, T14 levels correlate significantly with mTORC1. Silencing mTORC1 reverses the effects of T30 on PC12 cells measured via AChE release in undifferentiated PC12 cells, whilst silencing mTORC2 does not. This suggests that T14 acts selectively via mTORC1. T14 blockade offers a preferable alternative to currently available blockers of mTOR as it would enable selective blockade of mTORC1, thereby reducing side effects associated with generalised mTOR blockade.
    Keywords:  Alzheimer’s disease; NBP14; T14; T30; acetylcholinesterase; cancer; mTORC1; rapamycin
    DOI:  https://doi.org/10.3390/ijms24129961
  7. EMBO Rep. 2023 Jun 29. e56841
    An ATG12-ATG5-TECPR1 E3-like complex regulates unconventional LC3 lipidation at damaged lysosomes.
    Dale P Corkery, Sergio Castro-Gonzalez, Anastasia Knyazeva, Laura K Herzog, Yao-Wen Wu.
      Lysosomal membrane damage represents a threat to cell viability. As such, cells have evolved sophisticated mechanisms to maintain lysosomal integrity. Small membrane lesions are detected and repaired by the endosomal sorting complex required for transport (ESCRT) machinery while more extensively damaged lysosomes are cleared by a galectin-dependent selective macroautophagic pathway (lysophagy). In this study, we identify a novel role for the autophagosome-lysosome tethering factor, TECPR1, in lysosomal membrane repair. Lysosomal damage promotes TECPR1 recruitment to damaged membranes via its N-terminal dysferlin domain. This recruitment occurs upstream of galectin and precedes the induction of lysophagy. At the damaged membrane, TECPR1 forms an alternative E3-like conjugation complex with the ATG12-ATG5 conjugate to regulate ATG16L1-independent unconventional LC3 lipidation. Abolishment of LC3 lipidation via ATG16L1/TECPR1 double knockout impairs lysosomal recovery following damage.
    Keywords:  TECPR1; autophagy; lysophagy; lysosome; membrane repair
    DOI:  https://doi.org/10.15252/embr.202356841
  8. J Transl Med. 2023 Jun 24. 21(1): 413
    The new insights into autophagy in thyroid cancer progression.
    Yu-Bo Shi, Shu-Yuan Chen, Ren-Bin Liu.
      In recent decades, the incidence of thyroid cancer keeps growing at a shocking rate, which has aroused increasing concerns worldwide. Autophagy is a fundamental and ubiquitous biological event conserved in mammals including humans. Basically, autophagy is a catabolic process that cellular components including small molecules and damaged organelles are degraded for recycle to meet the energy needs, especially under the extreme conditions. The dysregulated autophagy has indicated to be involved in thyroid cancer progression. The enhancement of autophagy can lead to autophagic cell death during the degradation while the produced energies can be utilized by the rest of the cancerous tissue, thus this influence could be bidirectional, which plays either a tumor-suppressive or oncogenic role. Accordingly, autophagy can be suppressed by therapeutic agents and is thus regarded as a drug target for thyroid cancer treatments. In the present review, a brief description of autophagy and roles of autophagy in tumor context are given. We have addressed summary of the mechanisms and functions of autophagy in thyroid cancer. Some potential autophagy-targeted treatments are also summarized. The aim of the review is linking autophagy to thyroid cancer, so as to develop novel approaches to better control cancer progression.
    Keywords:  Autophagy; Autophagy inhibitor; Cancer progression; Targeted therapy; Thyroid cancer
    DOI:  https://doi.org/10.1186/s12967-023-04265-6
  9. Metabolism. 2023 Jun 27. pii: S0026-0495(23)00248-2. [Epub ahead of print] 155644
    The N-degron pathway mediates lipophagy: The chemical modulation of lipophagy in obesity and NAFLD.
    Eui Jung Jung, Ki Woon Sung, Tae Hyun Bae, Hee-Yeon Kim, Ha Rim Choi, Sung Hyun Kim, Chan Hoon Jung, Su Ran Mun, Yeon Sung Son, Shin Kim, Young Ho Suh, Anna Kashina, Joo-Won Park, Yong Tae Kwon.
       BACKGROUND AND AIMS: Central to the pathogenesis of nonalcoholic fatty liver disease (NAFLD) is the accumulation of lipids in the liver and various fat tissues. We aimed to elucidate the mechanisms by which lipid droplets (LDs) in the liver and adipocytes are degraded by the autophagy-lysosome system and develop therapeutic means to modulate lipophagy, i.e., autophagic degradation of LDs.
    METHODS: We monitored the process in which LDs are pinched off by autophagic membranes and degraded by lysosomal hydrolases in cultured cells and mice. The autophagic receptor p62/SQSTM-1/Sequestosome-1 was identified as a key regulator and used as a target to develop drugs to induce lipophagy. The efficacy of p62 agonists was validated in mice to treat hepatosteatosis and obesity.
    RESULTS: We found that the N-degron pathway modulates lipophagy. This autophagic degradation initiates when the molecular chaperones including BiP/GRP78, retro-translocated from the endoplasmic reticulum, is N-terminally (Nt-) arginylated by ATE1 R-transferase. The resulting Nt-arginine (Nt-Arg) binds the ZZ domain of p62 associated with LDs. Upon binding to Nt-Arg, p62 undergoes self-polymerization and recruits LC3+ phagophores to the site of lipophagy, leading to lysosomal degradation. Liver-specific Ate1 conditional knockout mice under high fat diet developed severe NAFLD. The Nt-Arg was modified into small molecule agonists to p62 that facilitate lipophagy in mice and exerted therapeutic efficacy in obesity and hepatosteatosis of wild-type but not p62 knockout mice.
    CONCLUSIONS: Our results show that the N-degron pathway modulates lipophagy and provide p62 as a drug target to treat NAFLD and other diseases related with metabolic syndrome.
    Keywords:  Hepatosteatosis; Lipid droplet; N-terminal arginylation; Obesity; The autophagy-lysosome system; p62/SQSTM1/Sequestosome-1
    DOI:  https://doi.org/10.1016/j.metabol.2023.155644
  10. mBio. 2023 Jun 27. e0351222
    SHIP1 modulates antimalarial immunity by bridging the crosstalk between type I IFN signaling and autophagy.
    Hongyu Li, Shuai Yang, Ke Zeng, Jiayin Guo, Jian Wu, Huaji Jiang, Yingchao Xie, Zhiqiang Hu, Jiansen Lu, Jianwu Yang, Xin-Zhuan Su, Jun Cui, Xiao Yu.
      Stringent control of the type I interferon (IFN-I) signaling is critical for host immune defense against infectious diseases, yet the molecular mechanisms that regulate this pathway remain elusive. Here, we show that Src homology 2 containing inositol phosphatase 1 (SHIP1) suppresses IFN-I signaling by promoting IRF3 degradation during malaria infection. Genetic ablation of Ship1 in mice leads to high levels of IFN-I and confers resistance to Plasmodium yoelii nigeriensis (P.y.) N67 infection. Mechanistically, SHIP1 promotes the selective autophagic degradation of IRF3 by enhancing K63-linked ubiquitination of IRF3 at lysine 313, which serves as a recognition signal for NDP52-mediated selective autophagic degradation. In addition, SHIP1 is downregulated by IFN-I-induced miR-155-5p upon P.y. N67 infection and severs as a feedback loop of the signaling crosstalk. This study reveals a regulatory mechanism between IFN-I signaling and autophagy, and verifies SHIP1 can be a potential target for therapeutic intervention against malaria and other infectious diseases. IMPORTANCE Malaria remains a serious disease affecting millions of people worldwide. Malaria parasite infection triggers tightly controlled type I interferon (IFN-I) signaling that plays a critical role in host innate immunity; however, the molecular mechanisms underlying the immune responses are still elusive. Here, we discover a host gene [Src homology 2-containing inositol phosphatase 1 (SHIP1)] that can regulate IFN-I signaling by modulating NDP52-mediated selective autophagic degradation of IRF3 and significantly affect parasitemia and resistance of Plasmodium-infected mice. This study identifies SHIP1 as a potential target for immunotherapies in malaria and highlights the crosstalk between IFN-I signaling and autophagy in preventing related infectious diseases. SHIP1 functions as a negative regulator during malaria infection by targeting IRF3 for autophagic degradation.
    Keywords:  IRF3; Plasmodium; SHIP1; autophagy; type I interferon
    DOI:  https://doi.org/10.1128/mbio.03512-22
  11. Cytokine Growth Factor Rev. 2023 Jun 24. pii: S1359-6101(23)00025-4. [Epub ahead of print]
    mTOR in programmed cell death and its therapeutic implications.
    Yawen Xie, Xianli Lei, Guoyu Zhao, Ran Guo, Na Cui.
      Mechanistic target of rapamycin (mTOR), a highly conserved serine/threonine kinase, is involved in cellular metabolism, protein synthesis, and cell death. Programmed cell death (PCD) assists in eliminating aging, damaged, or neoplastic cells, and is indispensable for sustaining normal growth, fighting pathogenic microorganisms, and maintaining body homeostasis. mTOR has crucial functions in the intricate signaling pathway network of multiple forms of PCD. mTOR can inhibit autophagy, which is part of PCD regulation. Cell survival is affected by mTOR through autophagy to control reactive oxygen species production and the degradation of pertinent proteins. Additionally, mTOR can regulate PCD in an autophagy-independent manner by affecting the expression levels of related genes and phosphorylating proteins. Therefore, mTOR acts through both autophagy-dependent and -independent pathways to regulate PCD. It is conceivable that mTOR exerts bidirectional regulation of PCD, such as ferroptosis, according to the complexity of signaling pathway networks, but the underlying mechanisms have not been fully explained. This review summarizes the recent advances in understanding mTOR-mediated regulatory mechanisms in PCD. Rigorous investigations into PCD-related signaling pathways have provided prospective therapeutic targets that may be clinically beneficial for treating various diseases.
    Keywords:  Apoptosis; Autophagy; Ferroptosis; Necroptosis; Pyroptosis; mTOR
    DOI:  https://doi.org/10.1016/j.cytogfr.2023.06.002
  12. Autophagy. 2023 Jun 25.
    TRIM16-mediated lysophagy suppresses high-gluocse-accumulated neuronal Aβ.
    Chang Woo Chae, Jee Hyeon Yoon, Jae Ryong Lim, Ji Yong Park, Ji Hyeon Cho, Young Hyun Jung, Gee Euhn Choi, Hyun Jik Lee, Ho Jae Han.
      Lysosomal dysfunction is a pathogenic link that may explain the causal relationship between diabetes and Alzheimer disease; however, there is no information about the regulation of hyperglycemia in neuronal lysophagy modulating lysosomal function. We examined the effect and related mechanisms of action of high glucose on lysophagy impairment and subsequent Aβ accumulation in human induced pluripotent stem cell (hiPSC)-derived neurons, mouse hippocampal neurons, and streptozotocin (STZ)-induced diabetic mice. High-glucose-induced neuronal lysosomal dysfunction through reactive oxygen species-mediated lysosomal membrane permeabilization and lysophagy impairment. Among lysophagy-related factors, the expression of TRIM16 (tripartite motif containing 16) was reduced in high-glucose-treated neuronal cells and the diabetic hippocampus through MTOR (mechanistic target of rapamycin kinase) complex 1 (MTORC1)-mediated inhibition of TFEB (transcription factor EB) activity. TRIM16 overexpression recovered lysophagy through the recruitment of MAP1LC3/LC3 (microtubule associated protein 1 light chain 3), SQSTM1/p62, and ubiquitin to damaged lysosomes, which inhibited the high-glucose-induced accumulation of Aβ and p-MAPT/tau. In the diabetic mice model, TFEB enhancer recovered lysophagy in the hippocampus, resulting in the amelioration of cognitive impairment. In conclusion, TRIM16-mediated lysophagy is a promising candidate for the inhibition of diabetes-associated Alzheimer disease pathogenesis.
    Keywords:  Amyloid β; TFEB; TRIM16; autophagy; diabetes; hippocampal neuron; lysophagy
    DOI:  https://doi.org/10.1080/15548627.2023.2229659
  13. Cells. 2023 06 20. pii: 1668. [Epub ahead of print]12(12):
    Starvation Protects Hepatocytes from Inflammatory Damage through Paradoxical mTORC1 Signaling.
    Iqra Hussain, Harini K Sureshkumar, Michael Bauer, Ignacio Rubio.
      Background and aims: Sepsis-related liver failure is associated with a particularly unfavorable clinical outcome. Calorie restriction is a well-established factor that can increase tissue resilience, protect against liver failure and improve outcome in preclinical models of bacterial sepsis. However, the underlying molecular basis is difficult to investigate in animal studies and remains largely unknown.
    METHODS: We have used an immortalized hepatocyte line as a model of the liver parenchyma to uncover the role of caloric restriction in the resilience of hepatocytes to inflammatory cell damage. In addition, we applied genetic and pharmacological approaches to investigate the contribution of the three major intracellular nutrient/energy sensor systems, AMPK, mTORC1 and mTORC2, in this context.
    RESULTS: We demonstrate that starvation reliably protects hepatocytes from cellular damage caused by pro-inflammatory cytokines. While the major nutrient- and energy-related signaling pathways AMPK, mTORC2/Akt and mTORC1 responded to caloric restriction as expected, mTORC1 was paradoxically activated by inflammatory stress in starved, energy-deprived hepatocytes. Pharmacological inhibition of mTORC1 or genetic silencing of the mTORC1 scaffold Raptor, but not its mTORC2 counterpart Rictor, abrogated the protective effect of starvation and exacerbated inflammation-induced cell death. Remarkably, mTORC1 activation in starved hepatocytes was uncoupled from the regulation of autophagy, but crucial for sustained protein synthesis in starved resistant cells.
    CONCLUSIONS: AMPK engagement and paradoxical mTORC1 activation and signaling mediate protection against pro-inflammatory stress exerted by caloric restriction in hepatocytes.
    Keywords:  AMPK; calorie restriction; mTORC2; sepsis
    DOI:  https://doi.org/10.3390/cells12121668
  14. Front Cell Dev Biol. 2023 ;11 1177440
    Inhibition of autophagy; an opportunity for the treatment of cancer resistance.
    Asha Tonkin-Reeves, Charlett M Giuliani, John T Price.
      The process of macroautophagy plays a pivotal role in the degradation of long-lived, superfluous, and damaged proteins and organelles, which are later recycled for cellular use. Normal cells rely on autophagy to combat various stressors and insults to ensure survival. However, autophagy is often upregulated in cancer cells, promoting a more aggressive phenotype that allows mutated cells to evade death after exposure to therapeutic treatments. As a result, autophagy has emerged as a significant factor in therapeutic resistance across many cancer types, with underlying mechanisms such as DNA damage, cell cycle arrest, and immune evasion. This review provides a comprehensive summary of the role of autophagy in therapeutic resistance and the limitations of available autophagic inhibitors in cancer treatment. It also highlights the urgent need to explore new inhibitors that can synergize with existing therapies to achieve better patient treatment outcomes. Advancing research in this field is crucial for developing more effective treatments that can help improve the lives of cancer patients.
    Keywords:  autophagy; autophagy mechanisms; cancer; drug resisitance; therapeutic resistance
    DOI:  https://doi.org/10.3389/fcell.2023.1177440
  15. Mol Biol Cell. 2023 Jun 28. mbcE23010006
    A conserved STRIPAK complex is required for autophagy in muscle tissue.
    Yungui Guo, Qiling Zeng, David Brooks, Erika R Geisbrecht.
      Autophagy is important for cellular homeostasis and to prevent the abnormal accumulation of proteins. While many proteins that comprise the canonical autophagy pathway have been characterized, the identification of new regulators may help understand tissue and/or stress-specific responses. Using an in silico approach, we identified Striatin interacting protein (Strip), MOB kinase activator 4 (Mob4), and Fibroblast growth factor receptor 1 oncogene partner 2 (Fgop2) as conserved mediators of muscle tissue maintenance. We performed affinity purification mass spectrometry (AP-MS) experiments with Drosophila melanogaster (D. melanogaster) Strip as a bait protein and co-purified additional Striatin Interacting Phosphatase and Kinase (STRIPAK) complex members from larval muscle tissue. NUAK and Starvin (Stv) also emerged as Strip-binding proteins and these physical interactions were verified in vivo using Proximity Ligation Assays (PLA). To understand the functional significance of the STRIPAK-NUAK-Stv complex, we employed a sensitized genetic assay combined with RNA interference (RNAi) to demonstrate that both NUAK and stv function in the same biological process with genes that encode for STRIPAK complex proteins. RNAi-directed knockdown of Strip in muscle tissue led to the accumulation of ubiquitinated cargo, p62, and Atg8a, consistent with a block in autophagy. Indeed, autophagic flux was decreased in Strip RNAi muscles, while lysosome biogenesis and activity were unaffected. Our results support a model whereby the STRIPAK-NUAK-Stv complex coordinately regulates autophagy in muscle tissue.
    DOI:  https://doi.org/10.1091/mbc.E23-01-0006
  16. Mol Neurodegener. 2023 Jun 24. 18(1): 41
    Targeted degradation of ⍺-synuclein aggregates in Parkinson's disease using the AUTOTAC technology.
    Jihoon Lee, Ki Woon Sung, Eun-Jin Bae, Dabin Yoon, Dasarang Kim, Jin Saem Lee, Da-Ha Park, Daniel Youngjae Park, Su Ran Mun, Soon Chul Kwon, Hye Yeon Kim, Joo-Ok Min, Seung-Jae Lee, Young Ho Suh, Yong Tae Kwon.
       BACKGROUND: There are currently no disease-modifying therapeutics for Parkinson's disease (PD). Although extensive efforts were undertaken to develop therapeutic approaches to delay the symptoms of PD, untreated α-synuclein (α-syn) aggregates cause cellular toxicity and stimulate further disease progression. PROTAC (Proteolysis-Targeting Chimera) has drawn attention as a therapeutic modality to target α-syn. However, no PROTACs have yet shown to selectively degrade α-syn aggregates mainly owing to the limited capacity of the proteasome to degrade aggregates, necessitating the development of novel approaches to fundamentally eliminate α-syn aggregates.
    METHODS: We employed AUTOTAC (Autophagy-Targeting Chimera), a macroautophagy-based targeted protein degradation (TPD) platform developed in our earlier studies. A series of AUTOTAC chemicals was synthesized as chimeras that bind both α-syn aggregates and p62/SQSTM1/Sequestosome-1, an autophagic receptor. The efficacy of Autotacs was evaluated to target α-syn aggregates to phagophores and subsequently lysosomes for hydrolysis via p62-dependent macroautophagy. The target engagement was monitored by oligomerization and localization of p62 and autophagic markers. The therapeutic efficacy to rescue PD symptoms was characterized in cultured cells and mice. The PK/PD (pharmacokinetics/pharmacodynamics) profiles were investigated to develop an oral drug for PD.
    RESULTS: ATC161 induced selective degradation of α-syn aggregates at DC50 of ~ 100 nM. No apparent degradation was observed with monomeric α-syn. ATC161 mediated the targeting of α-syn aggregates to p62 by binding the ZZ domain and accelerating p62 self-polymerization. These p62-cargo complexes were delivered to autophagic membranes for lysosomal degradation. In PD cellular models, ATC161 exhibited therapeutic efficacy to reduce cell-to-cell transmission of α-syn and to rescue cells from the damages in DNA and mitochondria. In PD mice established by injecting α-syn preformed fibrils (PFFs) into brain striata via stereotaxic surgery, oral administration of ATC161 at 10 mg/kg induced the degradation of α-syn aggregates and reduced their propagation. ATC161 also mitigated the associated glial inflammatory response and improved muscle strength and locomotive activity.
    CONCLUSION: AUTOTAC provides a platform to develop drugs for PD. ATC161, an oral drug with excellent PK/PD profiles, induces selective degradation of α-syn aggregates in vitro and in vivo. We suggest that ATC161 is a disease-modifying drug that degrades the pathogenic cause of PD.
    Keywords:  Lysosome; Macroautophagy; Targeted protein degradation; The N-degron pathway; The autophagy-lysosome system; p62/SQSTM1/Sequestosome-1
    DOI:  https://doi.org/10.1186/s13024-023-00630-7
  17. Hum Mol Genet. 2023 Jun 26. pii: ddad096. [Epub ahead of print]
    Comprehensive analysis of autophagic functions of WIPI family proteins and their implications for the pathogenesis of β-propeller associated neurodegeneration.
    Takahiro Shimizu, Norito Tamura, Taki Nishimura, Chieko Saito, Hayashi Yamamoto, Noboru Mizushima.
      β-propellers that bind polyphosphoinositides (PROPPINs) are an autophagy-related protein family conserved throughout eukaryotes. The PROPPIN family includes Atg18, Atg21, and Hsv2 in yeast and WD-repeat protein interacting with phosphoinositides (WIPI)1-4 in mammals. Mutations in the WIPI genes are associated with human neuronal diseases, including β-propeller associated neurodegeneration (BPAN) caused by mutations in WDR45 (encoding WIPI4). In contrast to yeast PROPPINs, the functions of mammalian WIPI1-WIPI4 have not been systematically investigated. Although the involvement of WIPI2 in autophagy has been clearly shown, the functions of WIPI1, WIPI3, and WIPI4 in autophagy remain poorly understood. In this study, we comprehensively analyzed the roles of WIPI proteins by using WIPI-knockout (single, double, and quadruple knockout) HEK293T cells and recently developed HaloTag-based reporters, which enable us to monitor autophagic flux sensitively and quantitatively. We found that WIPI2 was nearly essential for autophagy. Autophagic flux was unaffected or only slightly reduced by single deletion of WIPI3 (encoded by WDR45B) or WIPI4 but was profoundly reduced by double deletion of WIPI3 and WIPI4. Furthermore, we revealed variable effects of BPAN-related missense mutations on the autophagic activity of WIPI4. BPAN is characterized by neurodevelopmental and neurodegenerative abnormalities, and we found a possible association between the magnitude of the defect of the autophagic activity of WIPI4 mutants and the severity of neurodevelopmental symptoms. However, some of the BPAN-related missense mutations, which produce neurodegenerative signs, showed almost normal autophagic activity, suggesting that non-autophagic functions of WIPI4 may be related to neurodegeneration in BPAN.
    DOI:  https://doi.org/10.1093/hmg/ddad096
  18. Mol Cell. 2023 Jun 27. pii: S1097-2765(23)00426-4. [Epub ahead of print]
    Lysosomal LAMP proteins regulate lysosomal pH by direct inhibition of the TMEM175 channel.
    Jiyuan Zhang, Weizhong Zeng, Yan Han, Wan-Ru Lee, Jen Liou, Youxing Jiang.
      Maintaining a highly acidic lysosomal pH is central to cellular physiology. Here, we use functional proteomics, single-particle cryo-EM, electrophysiology, and in vivo imaging to unravel a key biological function of human lysosome-associated membrane proteins (LAMP-1 and LAMP-2) in regulating lysosomal pH homeostasis. Despite being widely used as a lysosomal marker, the physiological functions of the LAMP proteins have long been overlooked. We show that LAMP-1 and LAMP-2 directly interact with and inhibit the activity of the lysosomal cation channel TMEM175, a key player in lysosomal pH homeostasis implicated in Parkinson's disease. This LAMP inhibition mitigates the proton conduction of TMEM175 and facilitates lysosomal acidification to a lower pH environment crucial for optimal hydrolase activity. Disrupting the LAMP-TMEM175 interaction alkalinizes the lysosomal pH and compromises the lysosomal hydrolytic function. In light of the ever-increasing importance of lysosomes to cellular physiology and diseases, our data have widespread implications for lysosomal biology.
    Keywords:  LAMP-1 and LAMP-2; TMEM175; lysosomal LAMP proteins; lysosomal hydrolytic function; lysosomal pH homeostasis; lysosome acidification; risk factor for Parkinson’s disease
    DOI:  https://doi.org/10.1016/j.molcel.2023.06.004
  19. Drug Discov Today. 2023 Jun 26. pii: S1359-6446(23)00208-8. [Epub ahead of print] 103692
    Targeting SIRT1-regulated autophagic cell death as a novel therapeutic avenue for cancer prevention.
    Srimanta Patra, Prakash P Praharaj, Amruta Singh, Sujit K Bhutia.
      Cellular localization and deacetylation activity of Sirtuin 1 (SIRT1) has a significant role in cancer regulation. The multifactorial role of SIRT1 in autophagy regulates several cancer-associated cellular phenotypes, aiding cellular survival and cell death induction. SIRT1-mediated deacetylation of autophagy-related genes (ATGs) and associated signaling mediators control carcinogenesis. The hyperactivation of bulk autophagy, disrupted lysosomal and mitochondrial biogenesis, and excessive mitophagy is a key mechanism for SIRT1-mediated autophagic cell death (ACD). In terms of the SIRT1-ACD nexus, identifying SIRT1-activating small molecules and understanding the possible mechanism triggering ACD could be a potential therapeutic avenue for cancer prevention. In this review, we provide an update on the structural and functional intricacy of SIRT1 and SIRT1-mediated autophagy activation as an alternative cell death modality for cancer prevention.
    Keywords:  SIRT1; apoptosis; autophagic cell death; autophagy; cancer
    DOI:  https://doi.org/10.1016/j.drudis.2023.103692
  20. J Cell Biol. 2023 Aug 07. pii: e202211025. [Epub ahead of print]222(8):
    ULK phosphorylation of STX17 controls autophagosome maturation via FLNA.
    Yufen Wang, Huilin Que, ChuangPeng Li, Zhe Wu, Fenglei Jian, Yuan Zhao, Haohao Tang, Yang Chen, Shuaixin Gao, Catherine C L Wong, Ying Li, Chongchong Zhao, Yueguang Rong.
      Autophagy is a conserved and tightly regulated intracellular quality control pathway. ULK is a key kinase in autophagy initiation, but whether ULK kinase activity also participates in the late stages of autophagy remains unknown. Here, we found that the autophagosomal SNARE protein, STX17, is phosphorylated by ULK at residue S289, beyond which it localizes specifically to autophagosomes. Inhibition of STX17 phosphorylation prevents such autophagosome localization. FLNA was then identified as a linker between ATG8 family proteins (ATG8s) and STX17 with essential involvement in STX17 recruitment to autophagosomes. Phosphorylation of STX17 S289 promotes its interaction with FLNA, activating its recruitment to autophagosomes and facilitating autophagosome-lysosome fusion. Disease-causative mutations around the ATG8s- and STX17-binding regions of FLNA disrupt its interactions with ATG8s and STX17, inhibiting STX17 recruitment and autophagosome-lysosome fusion. Cumulatively, our study reveals an unexpected role of ULK in autophagosome maturation, uncovers its regulatory mechanism in STX17 recruitment, and highlights a potential association between autophagy and FLNA.
    DOI:  https://doi.org/10.1083/jcb.202211025
  21. Front Immunol. 2023 ;14 1188482
    The landscape of mitophagy in sepsis reveals PHB1 as an NLRP3 inflammasome inhibitor.
    Shipeng Chen, Jinqi Ma, Ping Yin, Fang Liang.
      Mitophagy is a selective autophagy targeting damaged and potential cytotoxic mitochondria, which can effectively prevent excessive cytotoxic production from damaged mitochondria and alleviate the inflammatory response. However, the potential role of mitophagy in sepsis remains poorly explored. Here, we studied the role of mitophagy in sepsis and its immune heterogeneity. By performing mitophagy-related typing on 348 sepsis samples, three clusters (A, B, and C) were obtained. Cluster A had the highest degree of mitophagy accompanied by lowest disease severity, while cluster C had the lowest degree of mitophagy with the highest disease severity. The three clusters had unique immune characteristics. We further revealed that the expression of PHB1 in these three clusters was significantly different and negatively correlated with the severity of sepsis, suggesting that PHB1 was involved in the development of sepsis. It has been reported that impaired mitophagy leads to the over-activation of inflammasomes, which promotes sepsis development. Further analysis showed that the expressions of NLRP3 inflammasomes core genes in cluster C were significantly up-regulated and negatively correlated with PHB1. Next, we verified whether PHB1 downregulation caused the activation of inflammasomes and found that the PHB1 knockdown increased the levels of mtDNA in the cytoplasm and enhanced the activation of NLRP3 inflammasomes. In addition, mitophagy inhibitor treatment abolished PHB1 knockdown-mediated activation of NLRP3 inflammasomes, suggesting that PHB1 inhibited the activation of inflammasomes through mitophagy. In conclusion, this study reveals that a high degree of mitophagy may predict a good outcome of sepsis, and PHB1 is a key NLRP3 inflammasome regulator via mitophagy in inflammatory diseases such as sepsis.
    Keywords:  NLRP3 inflammasome; PHB1; immunity; mitophagy; sepsis
    DOI:  https://doi.org/10.3389/fimmu.2023.1188482
  22. J Adv Res. 2023 Jun 23. pii: S2090-1232(23)00172-8. [Epub ahead of print]
    Autophagy, a double-edged sword for oral tissue regeneration.
    Xinyue Xu, Jia Wang, Yunlong Xia, Yuan Yin, Tianxiao Zhu, Faming Chen, Chunxu Hai.
       BACKGROUND: Oral health is of fundamental importance to maintain systemic health in humans. Stem cell-based oral tissue regeneration is a promising strategy to achieve the recovery of impaired oral tissue. As a highly conserved process of lysosomal degradation, autophagy induction regulates stem cell function physiologically and pathologically. Autophagy activation can serve as a cytoprotective mechanism in stressful environments, while insufficient or over-activation may also lead to cell function dysregulation and cell death.
    AIM OF REVIEW: This review focuses on the effects of autophagy on stem cell function and oral tissue regeneration, with particular emphasis on diverse roles of autophagy in different oral tissues, including periodontal tissue, bone tissue, dentin pulp tissue, oral mucosa, salivary gland, maxillofacial muscle, temporomandibular joint, etc. Additionally, this review introduces the molecular mechanisms involved in autophagy during the regeneration of different parts of oral tissue, and how autophagy can be regulated by small molecule drugs, biomaterials, exosomes/RNAs or other specific treatments. Finally, this review discusses new perspectives for autophagy manipulation and oral tissue regeneration.
    KEY SCIENTIFIC CONCEPTS OF REVIEW: Overall, this review emphasizes the contribution of autophagy to oral tissue regeneration and highlights the possible approaches for regulating autophagy to promote the regeneration of human oral tissue.
    Keywords:  Autophagy; Autophagy modulators; Oral disease; Oral tissue regeneration; Stem cell
    DOI:  https://doi.org/10.1016/j.jare.2023.06.010
  23. Biology (Basel). 2023 Jun 15. pii: 864. [Epub ahead of print]12(6):
    The ACSL4 Network Regulates Cell Death and Autophagy in Diseases.
    Fangquan Chen, Rui Kang, Jiao Liu, Daolin Tang.
      Lipid metabolism, cell death, and autophagy are interconnected processes in cells. Dysregulation of lipid metabolism can lead to cell death, such as via ferroptosis and apoptosis, while lipids also play a crucial role in the regulation of autophagosome formation. An increased autophagic response not only promotes cell survival but also causes cell death depending on the context, especially when selectively degrading antioxidant proteins or organelles that promote ferroptosis. ACSL4 is an enzyme that catalyzes the formation of long-chain acyl-CoA molecules, which are important intermediates in the biosynthesis of various types of lipids. ACSL4 is found in many tissues and is particularly abundant in the brain, liver, and adipose tissue. Dysregulation of ACSL4 is linked to a variety of diseases, including cancer, neurodegenerative disorders, cardiovascular disease, acute kidney injury, and metabolic disorders (such as obesity and non-alcoholic fatty liver disease). In this review, we introduce the structure, function, and regulation of ACSL4; discuss its role in apoptosis, ferroptosis, and autophagy; summarize its pathological function; and explore the potential implications of targeting ACSL4 in the treatment of various diseases.
    Keywords:  ACSL4; apoptosis; autophagy; cancer; ferroptosis
    DOI:  https://doi.org/10.3390/biology12060864
  24. Contact (Thousand Oaks). 2023 Jan-Dec;6:6 25152564231157706
    An Expansion of the Endoplasmic Reticulum that Halts Autophagy is Permissive to Genome Instability.
    Eliana Lara-Barba, Alba Torán-Vilarrubias, María Moriel-Carretero.
      The links between autophagy and genome stability, and whether they are important for lifespan and health, are not fully understood. We undertook a study to explore this notion at the molecular level using Saccharomyces cerevisiae. On the one hand, we triggered autophagy using rapamycin, to which we exposed mutants defective in preserving genome integrity, then assessed their viability, their ability to induce autophagy and the link between these two parameters. On the other hand, we searched for molecules derived from plant extracts known to have powerful benefits on human health to try to rescue the negative effects rapamycin had against some of these mutants. We uncover that autophagy execution is lethal for mutants unable to repair DNA double strand breaks, while the extract from Silybum marianum seeds induces an expansion of the endoplasmic reticulum (ER) that blocks autophagy and protects them. Our data uncover a connection between genome integrity and homeostasis of the ER whereby ER stress-like scenarios render cells tolerant to sub-optimal genome integrity conditions.
    Keywords:  DNA DSBs; Silybum marianum; autophagy; endoplasmic reticulum; genome instability; homologous recombination
    DOI:  https://doi.org/10.1177/25152564231157706
  25. Contact (Thousand Oaks). 2022 Jan-Dec;5:5 25152564221119347
    α-Synuclein Interactions in Mitochondria-ER Contacts: A Possible Role in Parkinson's Disease.
    Adolfo Garcia Erustes, Gabriel Cicolin Guarache, Erika da Cruz Guedes, Anderson Henrique França Figueredo Leão, Gustavo José da Silva Pereira, Soraya Soubhi Smaili.
      Endoplasmic reticulum-mitochondria contact sites regulate various biological processes, such as mitochondrial dynamics, calcium homeostasis, autophagy and lipid metabolism. Notably, dysfunctions in these contact sites are closely related to neurodegenerative diseases, including Parkinson's disease, Alzheimer's disease and amyotrophic lateral sclerosis. However, details about the role of endoplasmic reticulum-mitochondria contact sites in neurodegenerative diseases remain unknown. In Parkinson's disease, interactions between α-synuclein in the contact sites and components of tether complexes that connect organelles can lead to various dysfunctions, especially with regards to calcium homeostasis. This review will summarize the main tether complexes present in endoplasmic reticulum-mitochondria contact sites, and their roles in calcium homeostasis and trafficking. We will discuss the impact of α-synuclein accumulation, its interaction with tethering complex components and the implications in Parkinson's disease pathology.
    Keywords:  Parkinson's disease; calcium; endoplasmic reticulum; mitochondria; mitochondria-ER contact sites; α-synuclein
    DOI:  https://doi.org/10.1177/25152564221119347
  26. Clin Cosmet Investig Dermatol. 2023 ;16 1569-1581
    Alterations of Autophagy Modify Lipids in Epidermal Keratinocytes.
    Qingwei Geng, Guoqi Wei, Yebei Hu, Jinhui Xu, Xiuzu Song.
       Background: The skin barrier is the first line of defense of the body, while skin lipids play an important role in the skin permeability barrier. Lamellar bodies are also involved in maintaining the stability of the skin permeability barrier. However, the exact origin of lamellar bodies remains unclear. Recent studies have suggested that autophagy may participate in the formation of lamellar bodies.
    Aim: This study aimed to investigate the role of autophagy in the formation of lamellar bodies in keratinocytes and the regulation of keratinocyte lipids.
    Methods: Keratinocytes were incubated with autophagy inducer Rapamycin and autophagy inhibitor Bafilomycin A1. The changes in autophagy flux were detected by Western blot, and the formation of lamellar bodies was observed by transmission electron microscopy. Furthermore, the changes in keratinocytes lipidomics were detected by liquid chromatography-mass spectrometry.
    Results: Our research showed that the autophagy inducer promoted autophagy activation and formation of lamellar bodies in keratinocytes, while the inhibitor inhibited autophagy signals and the formation of lamellar bodies in keratinocytes. In addition, the lipidomics results revealed a significant change in glycerophospholipids after autophagy induction and autophagy inhibition.
    Conclusion: These results demonstrate that autophagy may play an essential role in skin lipids via glycerophospholipids pathway.
    Keywords:  Bafilomycin A1; keratinocytes; lipid; liquid chromatography-mass spectrometry; rapamycin; skin barrier
    DOI:  https://doi.org/10.2147/CCID.S410252
  27. Korean J Physiol Pharmacol. 2023 Jul 01. 27(4): 311-323
    Unveiling the impact of lysosomal ion channels: balancing ion signaling and disease pathogenesis.
    Yoona Jung, Wonjoon Kim, Na Kyoung Shin, Young Min Bae, Jinhong Wie.
      Ion homeostasis, which is regulated by ion channels, is crucial for intracellular signaling. These channels are involved in diverse signaling pathways, including cell proliferation, migration, and intracellular calcium dynamics. Consequently, ion channel dysfunction can lead to various diseases. In addition, these channels are present in the plasma membrane and intracellular organelles. However, our understanding of the function of intracellular organellar ion channels is limited. Recent advancements in electrophysiological techniques have enabled us to record ion channels within intracellular organelles and thus learn more about their functions. Autophagy is a vital process of intracellular protein degradation that facilitates the breakdown of aged, unnecessary, and harmful proteins into their amino acid residues. Lysosomes, which were previously considered protein-degrading garbage boxes, are now recognized as crucial intracellular sensors that play significant roles in normal signaling and disease pathogenesis. Lysosomes participate in various processes, including digestion, recycling, exocytosis, calcium signaling, nutrient sensing, and wound repair, highlighting the importance of ion channels in these signaling pathways. This review focuses on different lysosomal ion channels, including those associated with diseases, and provides insights into their cellular functions. By summarizing the existing knowledge and literature, this review emphasizes the need for further research in this field. Ultimately, this study aims to provide novel perspectives on the regulation of lysosomal ion channels and the significance of ion-associated signaling in intracellular functions to develop innovative therapeutic targets for rare and lysosomal storage diseases.
    Keywords:  Autophagy; Ion channels; Lysosomal storage diseases; Lysosomes
    DOI:  https://doi.org/10.4196/kjpp.2023.27.4.311
  28. J Exp Bot. 2023 Jun 26. pii: erad211. [Epub ahead of print]
    Transcriptional and Post-translational Regulation of Plant Autophagy.
    William Agbemafle, Min May Wong, Diane C Bassham.
      In response to changing environmental conditions, plants activate cellular responses to enable them to adapt to these changes. One such response is autophagy, in which cellular components, for example proteins and organelles, are delivered to the vacuole for degradation. Autophagy is activated by a wide range of conditions, and the regulatory pathways controlling this activation are now being elucidated. However, key insights into how these factors may function together to properly modulate autophagy in response to specific internal or external signals are yet to be discovered. In this review we discuss mechanisms for regulation of autophagy in response to environmental stress and disruptions in cell homeostasis. These pathways include post-translational modification of proteins required for autophagy activation and progression, control of protein stability of the autophagy machinery, and transcriptional regulation, resulting in changes in transcription of genes involved in autophagy. In particular, we highlight potential connections between the roles of key regulators and explore gaps in research, the filling of which can further our understanding of the autophagy regulatory network in plants.
    Keywords:  ATG; Autophagy; gene expression; persulfidation; phosphorylation; post-translational modification; starvation; stress; transcription factors; ubiquitination
    DOI:  https://doi.org/10.1093/jxb/erad211
  29. Nat Commun. 2023 Jun 29. 14(1): 3852
    A selective autophagy receptor VISP1 induces symptom recovery by targeting viral silencing suppressors.
    Xin Tong, Jia-Jia Zhao, Ya-Lan Feng, Jing-Ze Zou, Jian Ye, Junfeng Liu, Chenggui Han, Dawei Li, Xian-Bing Wang.
      Selective autophagy is a double-edged sword in antiviral immunity and regulated by various autophagy receptors. However, it remains unclear how to balance the opposite roles by one autophagy receptor. We previously identified a virus-induced small peptide called VISP1 as a selective autophagy receptor that facilitates virus infections by targeting components of antiviral RNA silencing. However, we show here that VISP1 can also inhibit virus infections by mediating autophagic degradation of viral suppressors of RNA silencing (VSRs). VISP1 targets the cucumber mosaic virus (CMV) 2b protein for degradation and attenuates its suppression activity on RNA silencing. Knockout and overexpression of VISP1 exhibit compromised and enhanced resistance against late infection of CMV, respectively. Consequently, VISP1 induces symptom recovery from CMV infection by triggering 2b turnover. VISP1 also targets the C2/AC2 VSRs of two geminiviruses and enhances antiviral immunity. Together, VISP1 induces symptom recovery from severe infections of plant viruses through controlling VSR accumulation.
    DOI:  https://doi.org/10.1038/s41467-023-39426-0
  30. Biomolecules. 2023 05 29. pii: 901. [Epub ahead of print]13(6):
    The Role of Retinal Pigment Epithelial Cells in Age-Related Macular Degeneration: Phagocytosis and Autophagy.
    Zhibo Si, Yajuan Zheng, Jing Zhao.
      Age-related macular degeneration (AMD) causes vision loss in the elderly population. Dry AMD leads to the formation of Drusen, while wet AMD is characterized by cell proliferation and choroidal angiogenesis. The retinal pigment epithelium (RPE) plays a key role in AMD pathogenesis. In particular, helioreceptor renewal depends on outer segment phagocytosis of RPE cells, while RPE autophagy can protect cells from oxidative stress damage. However, when the oxidative stress burden is too high and homeostasis is disturbed, the phagocytosis and autophagy functions of RPE become damaged, leading to AMD development and progression. Hence, characterizing the roles of RPE cell phagocytosis and autophagy in the pathogenesis of AMD can inform the development of potential therapeutic targets to prevent irreversible RPE and photoreceptor cell death, thus protecting against AMD.
    Keywords:  age-related macular degeneration; autophagy; oxidative stress; phagocytosis; retinal pigment epithelial cell
    DOI:  https://doi.org/10.3390/biom13060901
  31. Mol Immunol. 2023 Jun 23. pii: S0161-5890(23)00123-2. [Epub ahead of print]160 44-54
    Autotaxin promotes the degradation of the mucus layer by inhibiting autophagy in mouse colitis.
    Xiaoyan Chen, Hui Zhang, Xiaojiang Zhou, Yunwu Wang, Wenjie Shi.
      Autotaxin (ATX or ENPP2) is an autocrine enzyme associated with the metabolism of various phospholipids. ATX has recently been identified as a regulatory factor in immune-related and inflammation-associated diseases, such as inflammatory bowel disease, but the exact mechanism is unclear. Here, we treated mice with recombinant ATX protein or an ATX inhibitor to investigate the effect of ATX on colitis in mice and the underlying mechanism. In a mouse model of colitis, ATX expression was increased, autophagy was impaired, and the mucus barrier was disrupted. Recombinant ATX protein promoted intestinal inflammation, inhibited autophagy, and disrupted the mucus barrier, while an ATX inhibitor had the opposite effect. Next, we treated mice that received ATX with an autophagy activator and an adenosine 5'-monophosphate-activated protein kinase (AMPK) agonist. We observed that autophagy activator and AMPK agonist could repair the mucus barrier and alleviate intestinal inflammation in ATX-treated mice. In vitro, we obtained consistent results. Thus, we concluded that ATX could inhibit autophagy through the AMPK pathway, which consequently disordered the mucus barrier and aggravated intestinal inflammation.
    Keywords:  AMPK; Autophagy; Autotaxin; Mucus barrier; Ulcerative colitis
    DOI:  https://doi.org/10.1016/j.molimm.2023.06.002
  32. Biomolecules. 2023 05 29. pii: 903. [Epub ahead of print]13(6):
    Differences in the Intracellular Localization of Methylated β-Cyclodextrins-Threaded Polyrotaxanes Lead to Different Cellular States.
    Yuma Yamada, Shinnosuke Daikuhara, Atsushi Tamura, Kei Nishida, Nobuhiko Yui, Hideyoshi Harashima.
      Activation of autophagy represents a potential therapeutic strategy for the treatment of diseases that are caused by the accumulation of defective proteins and the formation of abnormal organelles. Methylated β-cyclodextrins-threaded polyrotaxane (Me-PRX), a supramolecular structured polymer, induces autophagy by interacting with the endoplasmic reticulum. We previously reported on the successful activation of mitochondria-targeted autophagy by delivering Me-RRX to mitochondria using a MITO-Porter, a mitochondria-targeted nanocarrier. The same level of autophagy induction was achieved at one-twentieth the dosage for the MITO-Porter (Me-PRX) compared to the naked Me-PRX. We report herein on the quantitative evaluation of the intracellular organelle localization of both naked Me-PRX and the MITO-Porter (Me-PRX). Mitochondria, endoplasmic reticulum and lysosomes were selected as target organelles because they would be involved in autophagy induction. In addition, organelle injury and cell viability assays were performed. The results showed that the naked Me-PRX and the MITO-Porter (Me-PRX) were localized in different intracellular organelles, and organelle injury was different, depending on the route of administration, indicating that different organelles contribute to autophagy induction. These findings indicate that the organelle to which the autophagy-inducing molecules are delivered plays an important role in the level of induction of autophagy.
    Keywords:  MITO-Porter; autophagy; cell biology; endoplasmic reticulum; mitochondria; polyrotaxane
    DOI:  https://doi.org/10.3390/biom13060903
  33. Life (Basel). 2023 Jun 02. pii: 1317. [Epub ahead of print]13(6):
    Biophysical Aspects of Neurodegenerative and Neurodevelopmental Disorders Involving Endo-/Lysosomal CLC Cl-/H+ Antiporters.
    Maria Antonietta Coppola, Abraham Tettey-Matey, Paola Imbrici, Paola Gavazzo, Antonella Liantonio, Michael Pusch.
      Endosomes and lysosomes are intracellular vesicular organelles with important roles in cell functions such as protein homeostasis, clearance of extracellular material, and autophagy. Endolysosomes are characterized by an acidic luminal pH that is critical for proper function. Five members of the gene family of voltage-gated ChLoride Channels (CLC proteins) are localized to endolysosomal membranes, carrying out anion/proton exchange activity and thereby regulating pH and chloride concentration. Mutations in these vesicular CLCs cause global developmental delay, intellectual disability, various psychiatric conditions, lysosomal storage diseases, and neurodegeneration, resulting in severe pathologies or even death. Currently, there is no cure for any of these diseases. Here, we review the various diseases in which these proteins are involved and discuss the peculiar biophysical properties of the WT transporter and how these properties are altered in specific neurodegenerative and neurodevelopmental disorders.
    Keywords:  CLC proteins; chloride transport; developmental; endosome; lysosome
    DOI:  https://doi.org/10.3390/life13061317
  34. Biomolecules. 2023 05 30. pii: 912. [Epub ahead of print]13(6):
    The Janus-Faced Role of Lipid Droplets in Aging: Insights from the Cellular Perspective.
    Nikolaus Bresgen, Melanie Kovacs, Angelika Lahnsteiner, Thomas Klaus Felder, Mark Rinnerthaler.
      It is widely accepted that nine hallmarks-including mitochondrial dysfunction, epigenetic alterations, and loss of proteostasis-exist that describe the cellular aging process. Adding to this, a well-described cell organelle in the metabolic context, namely, lipid droplets, also accumulates with increasing age, which can be regarded as a further aging-associated process. Independently of their essential role as fat stores, lipid droplets are also able to control cell integrity by mitigating lipotoxic and proteotoxic insults. As we will show in this review, numerous longevity interventions (such as mTOR inhibition) also lead to strong accumulation of lipid droplets in Saccharomyces cerevisiae, Caenorhabditis elegans, Drosophila melanogaster, and mammalian cells, just to name a few examples. In mammals, due to the variety of different cell types and tissues, the role of lipid droplets during the aging process is much more complex. Using selected diseases associated with aging, such as Alzheimer's disease, Parkinson's disease, type II diabetes, and cardiovascular disease, we show that lipid droplets are "Janus"-faced. In an early phase of the disease, lipid droplets mitigate the toxicity of lipid peroxidation and protein aggregates, but in a later phase of the disease, a strong accumulation of lipid droplets can cause problems for cells and tissues.
    Keywords:  IIS; LDs; aging; autophagy; lifespan; lipid peroxides; mTOR; misfolded proteins; mitochondria; protein aggregates
    DOI:  https://doi.org/10.3390/biom13060912
  35. PLoS Biol. 2023 Jun 29. 21(6): e3002196
    PINK1: From Parkinson's disease to mitophagy and back again.
    Benjamin O'Callaghan, John Hardy, Helene Plun-Favreau.
      The genetics of Parkinson's disease has been key to unravelling the PINK1-dependent mitophagy process. Here, we discuss the implications of a 2010 PLOS Biology paper that shed light on the functional importance of PINK1 in the mitophagy cascade.
    DOI:  https://doi.org/10.1371/journal.pbio.3002196
  36. Methods Mol Biol. 2023 ;2692 311-336
    Analysis of LC3-Associated Phagocytosis and Antigen Presentation in Macrophages and B Cells.
    Svenja Luisa Nopper, Katarina Wendy Schmidt, Laure-Anne Ligeon, Christian Münz.
      Canonical autophagy and the non-canonical autophagy pathway LC3-associated phagocytosis (LAP) play crucial roles in the immune system by processing antigens for major histocompatibility complex (MHC) class II restricted presentation to CD4+ T cells. Recent studies offer insight into the relationship between LAP, autophagy, and antigen processing in macrophages and dendritic cells; however their involvement during antigen processing in B cells is less well understood.In this chapter, we describe how to monitor, manipulate, and understand the role of LAP and classical autophagy during MHC-restricted antigen presentation by human monocyte-derived macrophages as well as in B cell lymphoblastoid cell lines (LCLs). It includes an explanation on how to generate LCLs and monocyte-derived macrophages from primary human cells. Then we describe two different approaches to manipulate the autophagy pathways: silencing of the atg4b gene using CRISPR/Cas9 technology and a lentivirus delivery system for specific ATG4B overexpression. We also propose a method for triggering LAP and measuring different ATG proteins using Western blot and immunofluorescence. Finally, we show an approach to investigate MHC class II antigen presentation by an in vitro co-culture assay that uses the measurement of secreted cytokines, released by activated CD4+ T cells, as readout.
    Keywords:  Autophagy; B cells; CRISPR/Cas9; Confocal microscopy; LC3-associated phagosome
    DOI:  https://doi.org/10.1007/978-1-0716-3338-0_21
  37. Front Immunol. 2023 ;14 1128700
    Apilimod activates the NLRP3 inflammasome through lysosome-mediated mitochondrial damage.
    Yingting Hou, Hongbin He, Ming Ma, Rongbin Zhou.
      NLRP3 is an important innate immune sensor that responses to various signals and forms the inflammasome complex, leading to IL-1β secretion and pyroptosis. Lysosomal damage has been implicated in NLRP3 inflammasome activation in response to crystals or particulates, but the mechanism remains unclear. We developed the small molecule library screening and found that apilimod, a lysosomal disruptor, is a selective and potent NLRP3 agonist. Apilimod promotes the NLRP3 inflammasome activation, IL-1β secretion, and pyroptosis. Mechanismically, while the activation of NLRP3 by apilimod is independent of potassium efflux and directly binding, apilimod triggers mitochondrial damage and lysosomal dysfunction. Furthermore, we found that apilimod induces TRPML1-dependent calcium flux in lysosomes, leading to mitochondrial damage and the NLRP3 inflammasome activation. Thus, our results revealed the pro-inflammasome activity of apilimod and the mechanism of calcium-dependent lysosome-mediated NLRP3 inflammasome activation.
    Keywords:  NLRP3 inflammasome; activation; apilimod; lysosome; mitochondria
    DOI:  https://doi.org/10.3389/fimmu.2023.1128700
  38. Int J Mol Sci. 2023 Jun 06. pii: 9798. [Epub ahead of print]24(12):
    Polyamines and Physical Activity in Musculoskeletal Diseases: A Potential Therapeutic Challenge.
    Letizia Galasso, Annalisa Cappella, Antonino Mulè, Lucia Castelli, Andrea Ciorciari, Alessandra Stacchiotti, Angela Montaruli.
      Autophagy dysregulation is commonplace in the pathogenesis of several invalidating diseases, such as musculoskeletal diseases. Polyamines, as spermidine and spermine, are small aliphatic cations essential for cell growth and differentiation, with multiple antioxidant, anti-inflammatory, and anti-apoptotic effects. Remarkably, they are emerging as natural autophagy regulators with strong anti-aging effects. Polyamine levels were significantly altered in the skeletal muscles of aged animals. Therefore, supplementation of spermine and spermidine may be important to prevent or treat muscle atrophy. Recent in vitro and in vivo experimental studies indicate that spermidine reverses dysfunctional autophagy and stimulates mitophagy in muscles and heart, preventing senescence. Physical exercise, as polyamines, regulates skeletal muscle mass inducing proper autophagy and mitophagy. This narrative review focuses on the latest evidence regarding the efficacy of polyamines and exercise as autophagy inducers, alone or coupled, in alleviating sarcopenia and aging-dependent musculoskeletal diseases. A comprehensive description of overall autophagic steps in muscle, polyamine metabolic pathways, and effects of the role of autophagy inducers played by both polyamines and exercise has been presented. Although literature shows few data in regard to this controversial topic, interesting effects on muscle atrophy in murine models have emerged when the two "autophagy-inducers" were combined. We hope these findings, with caution, can encourage researchers to continue investigating in this direction. In particular, if these novel insights could be confirmed in further in vivo and clinical studies, and the two synergic treatments could be optimized in terms of dose and duration, then polyamine supplementation and physical exercise might have a clinical potential in sarcopenia, and more importantly, implications for a healthy lifestyle in the elderly population.
    Keywords:  aging; autophagy; exercise; mitophagy; physical activity; polyamines; sarcopenia; spermidine; spermine
    DOI:  https://doi.org/10.3390/ijms24129798
  39. Cell Mol Neurobiol. 2023 Jul 01.
    Oxidative Stress-Involved Mitophagy of Retinal Pigment Epithelium and Retinal Degenerative Diseases.
    Si-Ming Zhang, Bin Fan, Yu- Lin Li, Zhao-Yang Zuo, Guang-Yu Li.
      The retinal pigment epithelium (RPE) is a highly specialized and polarized epithelial cell layer that plays an important role in sustaining the structural and functional integrity of photoreceptors. However, the death of RPE is a common pathological feature in various retinal diseases, especially in age-related macular degeneration (AMD) and diabetic retinopathy (DR). Mitophagy, as a programmed self-degradation of dysfunctional mitochondria, is crucial for maintaining cellular homeostasis and cell survival under stress. RPE contains a high density of mitochondria necessary for it to meet energy demands, so severe stimuli can cause mitochondrial dysfunction and the excess generation of intracellular reactive oxygen species (ROS), which can further trigger oxidative stress-involved mitophagy. In this review, we summarize the classical pathways of oxidative stress-involved mitophagy in RPE and investigate its role in the progression of retinal diseases, aiming to provide a new therapeutic strategy for treating retinal degenerative diseases. The role of mitophagy in AMD and DR. In AMD, excessive ROS production promotes mitophagy in the RPE by activating the Nrf2/p62 pathway, while in DR, ROS may suppress mitophagy by the FOXO3-PINK1/parkin signaling pathway or the TXNIP-mitochondria-lysosome-mediated mitophagy.
    Keywords:  Age-related macular degeneration; Diabetic retinopathy; Mitophagy; Oxidative stress; Retinal pigment epithelium
    DOI:  https://doi.org/10.1007/s10571-023-01383-z
  40. Int J Mol Sci. 2023 Jun 07. pii: 9834. [Epub ahead of print]24(12):
    mTOR Inhibition Is Effective against Growth, Survival and Migration, but Not against Microglia Activation in Preclinical Glioma Models.
    Lucia Lisi, Michela Pizzoferrato, Gabriella Maria Pia Ciotti, Maria Martire, Pierluigi Navarra.
      Initially introduced in therapy as immunosuppressants, the selective inhibitors of mTORC1 have been approved for the treatment of solid tumors. Novel non-selective inhibitors of mTOR are currently under preclinical and clinical developments in oncology, attempting to overcome some limitations associated with selective inhibitors, such as the development of tumor resistance. Looking at the possible clinical exploitation in the treatment of glioblastoma multiforme, in this study we used the human glioblastoma cell lines U87MG, T98G and microglia (CHME-5) to compare the effects of a non-selective mTOR inhibitor, sapanisertib, with those of rapamycin in a large array of experimental paradigms, including (i) the expression of factors involved in the mTOR signaling cascade, (ii) cell viability and mortality, (iii) cell migration and autophagy, and (iv) the profile of activation in tumor-associated microglia. We could distinguish between effects of the two compounds that were overlapping or similar, although with differences in potency and or/time-course, and effects that were diverging or even opposite. Among the latter, especially relevant is the difference in the profile of microglia activation, with rapamycin being an overall inhibitor of microglia activation, whereas sapanisertib was found to induce an M2-profile, which is usually associated with poor clinical outcomes.
    Keywords:  glioblastoma; mTOR; microglia; rapamycin; sapanisertib
    DOI:  https://doi.org/10.3390/ijms24129834
  41. Biochem Biophys Res Commun. 2023 Jun 17. pii: S0006-291X(23)00810-0. [Epub ahead of print]672 168-176
    Essential amino acid starvation induces cell cycle arrest, autophagy, and inhibits osteogenic differentiation in murine osteoblast.
    Runbo Li, Hirohito Kato, Takaya Nakata, Isao Yamawaki, Nobuhiro Yamauchi, Kazutaka Imai, Yoichiro Taguchi, Makoto Umeda.
      This study investigates the effects of essential amino acid (EAA) starvation on murine osteoblasts cells and the underlying mechanisms. We performed and observed the cell proliferation, autophagy, and osteogenic differentiation under deprivation of EAA in vitro. The results showed that EAA starvation resulted in cell cycle arrest via phosphorylation of the MAPK signaling pathway, leading to inhibition of cell proliferation and osteogenic differentiation. Additionally, the LKB1-AMPK signaling pathway was also found to be phosphorylated, inducing autophagy. These findings highlight the significant role of EAA in regulating cellular processes. Furthermore, this study contributes to our understanding of the effects of nutrient deprivation on cellular physiology and may aid in the development of novel therapeutic strategies for diseases associated with amino acid metabolism.
    Keywords:  Autophagy; Cell cycle; Essential amino acid; Osteoblast
    DOI:  https://doi.org/10.1016/j.bbrc.2023.06.055
  42. mBio. 2023 Jun 30. e0079523
    Toxoplasma gondii inhibits the expression of autophagy-related genes through AKT-dependent inactivation of the transcription factor FOXO3a.
    Andres Felipe Diez, Louis-Philippe Leroux, Sophie Chagneau, Alexandra Plouffe, Mackenzie Gold, Visnu Chaparro, Maritza Jaramillo.
      The intracellular parasite Toxoplasma gondii induces host AKT activation to prevent autophagy-mediated clearance; however, the molecular underpinnings are not fully understood. Autophagy can be negatively regulated through AKT-sensitive phosphorylation and nuclear export of the transcription factor Forkhead box O3a (FOXO3a). Using a combination of pharmacological and genetic approaches, herein we investigated whether T. gondii hinders host autophagy through AKT-dependent inactivation of FOXO3a. We found that infection by type I and II strains of T. gondii promotes gradual and sustained AKT-dependent phosphorylation of FOXO3a at residues S253 and T32 in human foreskin fibroblasts (HFF) and murine 3T3 fibroblasts. Mechanistically, AKT-sensitive phosphorylation of FOXO3a by T. gondii required live infection and the activity of PI3K but was independent of the plasma membrane receptor EGFR and the kinase PKCα. Phosphorylation of FOXO3a at AKT-sensitive residues was paralleled by its nuclear exclusion in T. gondii-infected HFF. Importantly, the parasite was unable to drive cytoplasmic localization of FOXO3a upon pharmacological blockade of AKT or overexpression of an AKT-insensitive mutant form of FOXO3a. Transcription of a subset of bona fide autophagy-related targets of FOXO3a was reduced during T. gondii infection in an AKT-dependent fashion. However, parasite-directed repression of autophagy-related genes was AKT-resistant in cells deficient in FOXO3a. Consistent with this, T. gondii failed to inhibit the recruitment of acidic organelles and LC3, an autophagy marker, to the parasitophorous vacuole upon chemically or genetically induced nuclear retention of FOXO3a. In all, we provide evidence that T. gondii suppresses FOXO3a-regulated transcriptional programs to prevent autophagy-mediated killing. IMPORTANCE The parasite Toxoplasma gondii is the etiological agent of toxoplasmosis, an opportunistic infection commonly transmitted by ingestion of contaminated food or water. To date, no effective vaccines in humans have been developed and no promising drugs are available to treat chronic infection or prevent congenital infection. T. gondii targets numerous host cell processes to establish a favorable replicative niche. Of note, T. gondii activates the host AKT signaling pathway to prevent autophagy-mediated killing. Herein, we report that T. gondii inhibits FOXO3a, a transcription factor that regulates the expression of autophagy-related genes, through AKT-dependent phosphorylation. The parasite's ability to block the recruitment of the autophagy machinery to the parasitophorous vacuole is impeded upon pharmacological inhibition of AKT or overexpression of an AKT-insensitive form of FOXO3a. Thus, our study provides greater granularity in the role of FOXO3a during infection and reinforces the potential of targeting autophagy as a therapeutic strategy against T. gondii.
    Keywords:  AKT; FOXO3a; Toxoplasma gondii; autophagy; host response; host-pathogen interactions; transcriptional regulation
    DOI:  https://doi.org/10.1128/mbio.00795-23
  43. Autophagy Rep. 2022 ;1(1): 226-233
    Follicular lymphoma-associated mutations in the V-ATPase chaperone Vma21 activate autophagy by dysfunctional V-ATPase assembly.
    Ying Yang, Zhihai Zhang, Daniel J Klionsky.
      A significant number of follicular lymphoma patients display recurrent mutations in subunits and regulators of the vacuolar-type H+-translocating ATPase (V-ATPase). Past studies focusing on the role of these mutations highlighted essential functions of macroautophagy/autophagy, amino-acid, and nutrient-sensing pathways in the pathogenesis of this disease. Here, we demonstrate novel results understanding the role of the follicular lymphoma-associated hotspot mutation VMA21p.93X, which corresponds to Vma21[Δ66-77] in S. cerevisiae cells. We find that V-ATPase assembly is affected by the Vma21[Δ66-77] mutation, shown by decreased vacuolar levels of V0 subunits as well as a Vph1 stability assay. In addition, we report that vacuolar levels of histidine, lysine and arginine are significantly reduced in Vma21[Δ66-77] mutant cells. These results deepen the current understanding on the mechanism of how autophagy is activated by these mutations in follicular lymphoma.
    Keywords:  Proton gradient; Vph1; stress; vacuole; yeast
    DOI:  https://doi.org/10.1080/27694127.2022.2077509
  44. Int J Mol Sci. 2023 Jun 06. pii: 9829. [Epub ahead of print]24(12):
    A Yeast Mitotic Tale for the Nucleus and the Vacuoles to Embrace.
    Silvia Santana-Sosa, Emiliano Matos-Perdomo, Jessel Ayra-Plasencia, Félix Machín.
      The morphology of the nucleus is roughly spherical in most eukaryotic cells. However, this organelle shape needs to change as the cell travels through narrow intercellular spaces during cell migration and during cell division in organisms that undergo closed mitosis, i.e., without dismantling the nuclear envelope, such as yeast. In addition, the nuclear morphology is often modified under stress and in pathological conditions, being a hallmark of cancer and senescent cells. Thus, understanding nuclear morphological dynamics is of uttermost importance, as pathways and proteins involved in nuclear shaping can be targeted in anticancer, antiaging, and antifungal therapies. Here, we review how and why the nuclear shape changes during mitotic blocks in yeast, introducing novel data that associate these changes with both the nucleolus and the vacuole. Altogether, these findings suggest a close relationship between the nucleolar domain of the nucleus and the autophagic organelle, which we also discuss here. Encouragingly, recent evidence in tumor cell lines has linked aberrant nuclear morphology to defects in lysosomal function.
    Keywords:  TOR; autophagy; lipid metabolism; lysosome; mitosis; nuclear envelope; nucleolus; nucleophagy; ribosomal DNA (rDNA); vacuole
    DOI:  https://doi.org/10.3390/ijms24129829
  45. Nat Cell Biol. 2023 Jun 29.
    mTORC2-NDRG1-CDC42 axis couples fasting to mitochondrial fission.
    Nuria Martinez-Lopez, Pamela Mattar, Miriam Toledo, Henrietta Bains, Manu Kalyani, Marie Louise Aoun, Mridul Sharma, Laura Beth J McIntire, Leslie Gunther-Cummins, Frank P Macaluso, Jennifer T Aguilan, Simone Sidoli, Mathieu Bourdenx, Rajat Singh.
      Fasting triggers diverse physiological adaptations including increases in circulating fatty acids and mitochondrial respiration to facilitate organismal survival. The mechanisms driving mitochondrial adaptations and respiratory sufficiency during fasting remain incompletely understood. Here we show that fasting or lipid availability stimulates mTORC2 activity. Activation of mTORC2 and phosphorylation of its downstream target NDRG1 at serine 336 sustains mitochondrial fission and respiratory sufficiency. Time-lapse imaging shows that NDRG1, but not the phosphorylation-deficient NDRG1Ser336Ala mutant, engages with mitochondria to facilitate fission in control cells, as well as in those lacking DRP1. Using proteomics, a small interfering RNA screen, and epistasis experiments, we show that mTORC2-phosphorylated NDRG1 cooperates with small GTPase CDC42 and effectors and regulators of CDC42 to orchestrate fission. Accordingly, RictorKO, NDRG1Ser336Ala mutants and Cdc42-deficient cells each display mitochondrial phenotypes reminiscent of fission failure. During nutrient surplus, mTOR complexes perform anabolic functions; however, paradoxical reactivation of mTORC2 during fasting unexpectedly drives mitochondrial fission and respiration.
    DOI:  https://doi.org/10.1038/s41556-023-01163-3
  46. Hum Mol Genet. 2023 Jun 29. pii: ddad102. [Epub ahead of print]
    Mitochondrial dysfunction and mitophagy defects in LRRK2-R1441C Parkinson's disease models.
    Matthew G Williamson, Marta Madureira, William McGuinness, Rachel Heon-Roberts, Elliot D Mock, Kalina Naidoo, Kaitlyn M L Cramb, Maria-Claudia Caiazza, Ana Belen Malpartida, Martha Lavelle, Katrina Savory, Stewart W Humble, Ryan Patterson, John B Davis, Natalie Connor-Robson, Brent J Ryan, Richard Wade-Martins.
      Mutations in the Leucine-Rich Repeat Kinase 2 (LRRK2) gene have been identified as one of the most common genetic causes of Parkinson's disease (PD). The LRRK2 PD-associated mutations LRRK2G2019S and LRRK2R1441C, located in the kinase domain and in the ROC-COR domain, respectively, have been demonstrated to impair mitochondrial function. Here, we sought to further our understanding of mitochondrial health and mitophagy by integrating data from LRRK2R1441C rat primary cortical and human induced pluripotent stem cell-derived dopamine (iPSC-DA) neuronal cultures as models of PD. We found LRRK2R1441C neurons exhibit decreased mitochondrial membrane potential, impaired mitochondrial function and decreased basal mitophagy levels. Mitochondrial morphology was altered in LRRK2R1441C iPSC-DA but not in cortical neuronal cultures or aged striatal tissue, indicating a cell type-specific phenotype. Additionally, LRRK2R1441C but not LRRK2G2019S neurons demonstrated decreased levels of the mitophagy marker pS65Ub in response to mitochondrial damage, which could disrupt degradation of damaged mitochondria. This impaired mitophagy activation and mitochondrial function were not corrected by the LRRK2 inhibitor MLi-2 in LRRK2R1441C iPSC-DA neuronal cultures. Furthermore, we demonstrate LRRK2 interaction with MIRO1, a protein necessary to stabilise and to anchor mitochondria for transport, occurs at mitochondria, in a genotype-independent manner. Despite this, we found that degradation of MIRO1 was impaired in LRRK2R1441C cultures upon induced mitochondrial damage, suggesting a divergent mechanism from the LRRK2G2019S mutation.
    DOI:  https://doi.org/10.1093/hmg/ddad102
  47. Nat Commun. 2023 06 26. 14(1): 3801
    Elevated levels of FMRP-target MAP1B impair human and mouse neuronal development and mouse social behaviors via autophagy pathway.
    Birth Defects Research Laboratory
      Fragile X messenger ribonucleoprotein 1 protein (FMRP) binds many mRNA targets in the brain. The contribution of these targets to fragile X syndrome (FXS) and related autism spectrum disorder (ASD) remains unclear. Here, we show that FMRP deficiency leads to elevated microtubule-associated protein 1B (MAP1B) in developing human and non-human primate cortical neurons. Targeted MAP1B gene activation in healthy human neurons or MAP1B gene triplication in ASD patient-derived neurons inhibit morphological and physiological maturation. Activation of Map1b in adult male mouse prefrontal cortex excitatory neurons impairs social behaviors. We show that elevated MAP1B sequesters components of autophagy and reduces autophagosome formation. Both MAP1B knockdown and autophagy activation rescue deficits of both ASD and FXS patients' neurons and FMRP-deficient neurons in ex vivo human brain tissue. Our study demonstrates conserved FMRP regulation of MAP1B in primate neurons and establishes a causal link between MAP1B elevation and deficits of FXS and ASD.
    DOI:  https://doi.org/10.1038/s41467-023-39337-0
  48. J Cell Sci. 2023 Jun 27. pii: jcs.261003. [Epub ahead of print]
    Novel phosphorylation of Rab29 that regulates its localization and lysosomal stress response in concert with LRRK2.
    Tadayuki Komori, Tomoki Kuwahara, Tetta Fujimoto, Maria Sakurai, Ikuko Koyama-Honda, Mitsunori Fukuda, Takeshi Iwatsubo.
      Rab proteins are small GTPases that regulate a myriad of intracellular membrane trafficking events. Rab29 is one of the Rab proteins phosphorylated by leucine-rich repeat kinase 2 (LRRK2), a Parkinson's disease-associated kinase. Recent studies suggest that Rab29 regulates LRRK2, whereas the mechanism by which Rab29 is regulated remained unclear. Here we report a novel phosphorylation in Rab29 that is not mediated by LRRK2 and occurs under lysosomal overload stress. Mass spectrometry analysis identified the phosphorylation site of Rab29 as Ser185, and cellular expression studies of phosphomimetic mutants of Rab29 at Ser185 unveiled the involvement of this phosphorylation in counteracting lysosomal enlargement. PKCα and PKCδ were deemed to be involved in this phosphorylation and control the lysosomal localization of Rab29 in concert with LRRK2. These results implicate PKCs in the lysosomal stress response pathway comprised of Rab29 and LRRK2, and further underscore the importance of this pathway in the mechanisms underlying lysosomal homeostasis.
    Keywords:  LRRK2; Lysosome; PKC; Phosphorylation; Rab29
    DOI:  https://doi.org/10.1242/jcs.261003
  49. Antioxidants (Basel). 2023 May 30. pii: 1183. [Epub ahead of print]12(6):
    Autophagy Activation Promoted by Pulses of Light and Phytochemicals Counteracting Oxidative Stress during Age-Related Macular Degeneration.
    Roberto Pinelli, Michela Ferrucci, Francesca Biagioni, Caterina Berti, Violet Vakunseth Bumah, Carla Letizia Busceti, Stefano Puglisi-Allegra, Gloria Lazzeri, Alessandro Frati, Francesco Fornai.
      The seminal role of autophagy during age-related macular degeneration (AMD) lies in the clearance of a number of reactive oxidative species that generate dysfunctional mitochondria. In fact, reactive oxygen species (ROS) in the retina generate misfolded proteins, alter lipids and sugars composition, disrupt DNA integrity, damage cell organelles and produce retinal inclusions while causing AMD. This explains why autophagy in the retinal pigment epithelium (RPE), mostly at the macular level, is essential in AMD and even in baseline conditions to provide a powerful and fast replacement of oxidized molecules and ROS-damaged mitochondria. When autophagy is impaired within RPE, the deleterious effects of ROS, which are produced in excess also during baseline conditions, are no longer counteracted, and retinal degeneration may occur. Within RPE, autophagy can be induced by various stimuli, such as light and naturally occurring phytochemicals. Light and phytochemicals, in turn, may synergize to enhance autophagy. This may explain the beneficial effects of light pulses combined with phytochemicals both in improving retinal structure and visual acuity. The ability of light to activate some phytochemicals may further extend such a synergism during retinal degeneration. In this way, photosensitive natural compounds may produce light-dependent beneficial antioxidant effects in AMD.
    Keywords:  amber light; autophagolysosomes; bilberry; blue light; curcumin; drusen; lutein; red light; resveratrol; visual acuity
    DOI:  https://doi.org/10.3390/antiox12061183
  50. Nutrients. 2023 Jun 07. pii: 2655. [Epub ahead of print]15(12):
    Lactiplantibacillus plantarum OLL2712 Induces Autophagy via MYD88 and Strengthens Tight Junction Integrity to Promote the Barrier Function in Intestinal Epithelial Cells.
    Yumiko Watanabe-Yasuoka, Ayako Gotou, Shigeomi Shimizu, Toshihiro Sashihara.
      Autophagy is an important system conserved in eukaryotes that maintains homeostasis by degrading abnormal proteins. Autophagy incompetence in intestinal epithelial cells causes the abnormal function of intestinal stem cells and other cells and damages intestinal barrier function. The disruption of the intestinal barrier causes chronic inflammation throughout the body, followed by impaired glucose and lipid metabolism. Lactiplantibacillus plantarum OLL2712 (OLL2712) is a lactic acid bacterium that induces interleukin-10 production from immune cells, alleviates chronic inflammation, and improves glucose and lipid metabolism. In this study, we hypothesized that OLL2712 exerts anti-inflammatory effects by inducing autophagy and ameliorating intestinal barrier dysfunction, and we investigated its autophagy-inducing activities and functions. Caco-2 cells stimulated with OLL2712 for 24 h showed an increased number of autolysosomes per cell, compared with unstimulated cells. Therefore, the permeability of fluorescein isothiocyanate dextran 4000 (FD-4) was suppressed by inducing autophagy. In contrast, mucin secretion in HT-29-MTX-E12 cells was also increased by OLL2712 but not via autophagy induction. Finally, the signaling pathway involved in autophagy induction by OLL2712 was found to be mediated by myeloid differentiation factor 88 (MYD88). In conclusion, our findings suggest that OLL2712 induces autophagy in intestinal epithelial cells via MYD88, and that mucosal barrier function is strengthened by inducing autophagy.
    Keywords:  autophagy; intestinal barrier; lactic acid bacteria; mucin 2; myeloid differentiation factor 88
    DOI:  https://doi.org/10.3390/nu15122655
  51. Cell Signal. 2023 Jun 24. pii: S0898-6568(23)00196-1. [Epub ahead of print]109 110782
    UHRF2 promotes the malignancy of hepatocellular carcinoma by PARP1 mediated autophagy.
    Yiqi Zhang, Kejia Wu, Yuxin Liu, Shuangling Sun, Yue Shao, Qingxiu Li, Xinying Sui, Changzhu Duan.
      Autophagy have critical implications in the proliferation and metastasis of HCC. In the current study, we aimed to explore the underlying mechanisms of UHRF2 regulates HCC cells autophagy and HCC progression. We initially determined the relationship between UHRF2 and HCC autophagy, oncogenicity and patient survival through GSEA database and TCGA database. We mainly investigated the effect of UHRF2 on HCC development and autophagy through western blot, electron microscopy, and immunofluorescence. Functionally, UHRF2 was positively involved in the autophagy activation. Overexpression of UHRF2 reduced apoptosis in HCC cells, and promoted the malignancy phenotype of HCC both in vitro and in vivo. Mechanistically, PRDX1 bound to UHRF2 and upregulated its protein expression to facilitate the biological function of UHRF2 in HCC. Meanwhile, UHRF2 bound to autophagy-related protein PARP1 and upregulated PARP1 protein level. The results showed that UHRF2 promoted autophagy and contributed to the malignant phenotype of HCC by regulating PARP1 protein level. In summary, a novel interaction between PRDX1, UHRF2, and PARP1 was revealed, suggesting that UHRF2 could inspire a potential biomarker and potential therapeutic target for HCC.
    Keywords:  Autophagy; HCC; PARP1; PRDX1; Pathway; UHRF2
    DOI:  https://doi.org/10.1016/j.cellsig.2023.110782
  52. Biology (Basel). 2023 Jun 04. pii: 817. [Epub ahead of print]12(6):
    Using Zebrafish to Dissect the Interaction of Mycobacteria with the Autophagic Machinery in Macrophages.
    Salomé Muñoz-Sánchez, Mónica Varela, Michiel van der Vaart, Annemarie H Meijer.
      Existing drug treatment against tuberculosis is no match against the increasing number of multi-drug resistant strains of its causative agent, Mycobacterium tuberculosis (Mtb). A better understanding of how mycobacteria subvert the host immune defenses is crucial for developing novel therapeutic strategies. A potential approach is enhancing the activity of the autophagy machinery, which can direct bacteria to autophagolysosomal degradation. However, the interplay specifics between mycobacteria and the autophagy machinery must be better understood. Here, we analyzed live imaging data from the zebrafish model of tuberculosis to characterize mycobacteria-autophagy interactions during the early stages of infection in vivo. For high-resolution imaging, we microinjected fluorescent Mycobacterium marinum (Mm) into the tail fin tissue of zebrafish larvae carrying the GFP-LC3 autophagy reporter. We detected phagocytosed Mm clusters and LC3-positive Mm-containing vesicles within the first hour of infection. LC3 associations with these vesicles were transient and heterogeneous, ranging from simple vesicles to complex compound structures, dynamically changing shape by fusions between Mm-containing and empty vesicles. LC3-Mm-vesicles could adopt elongated shapes during cell migration or alternate between spacious and compact morphologies. LC3-Mm-vesicles were also observed in cells reverse migrating from the infection site, indicating that the autophagy machinery fails to control infection before tissue dissemination.
    Keywords:  autophagy; infection; innate immunity; live imaging; macrophages; zebrafish
    DOI:  https://doi.org/10.3390/biology12060817
  53. Aging Cell. 2023 Jun 26. e13910
    Acyl coenzyme A binding protein (ACBP): An aging- and disease-relevant "autophagy checkpoint".
    Léa Montégut, Mahmoud Abdellatif, Omar Motiño, Frank Madeo, Isabelle Martins, Victor Quesada, Carlos López-Otín, Guido Kroemer.
      Acyl coenzyme A binding protein (ACBP), also known as diazepam-binding inhibitor (DBI), is a phylogenetically ancient protein present in some eubacteria and the entire eukaryotic radiation. In several eukaryotic phyla, ACBP/DBI transcends its intracellular function in fatty acid metabolism because it can be released into the extracellular space. This ACBP/DBI secretion usually occurs in response to nutrient scarcity through an autophagy-dependent pathway. ACBP/DBI and its peptide fragments then act on a range of distinct receptors that diverge among phyla, namely metabotropic G protein-coupled receptor in yeast (and likely in the mammalian central nervous system), a histidine receptor kinase in slime molds, and ionotropic gamma-aminobutyric acid (GABA)A receptors in mammals. Genetic or antibody-mediated inhibition of ACBP/DBI orthologs interferes with nutrient stress-induced adaptations such as sporulation or increased food intake in multiple species, as it enhances lifespan or healthspan in yeast, plant leaves, nematodes, and multiple mouse models. These lifespan and healthspan-extending effects of ACBP/DBI suppression are coupled to the induction of autophagy. Altogether, it appears that neutralization of extracellular ACBP/DBI results in "autophagy checkpoint inhibition" to unleash the anti-aging potential of autophagy. Of note, in humans, ACBP/DBI levels increase in various tissues, as well as in the plasma, in the context of aging, obesity, uncontrolled infection or cardiovascular, inflammatory, neurodegenerative, and malignant diseases.
    Keywords:  aging; autophagy; diazepam-binding inhibitor; endozepin; evolution; metabolism
    DOI:  https://doi.org/10.1111/acel.13910
  54. Nature. 2023 Jun 28.
    Reprogramming tumour-associated macrophages to outcompete cancer cells.
    Xian Zhang, Shun Li, Isha Malik, Mytrang H Do, Liangliang Ji, Chun Chou, Wei Shi, Kristelle J Capistrano, Jing Zhang, Ting-Wei Hsu, Briana G Nixon, Ke Xu, Xinxin Wang, Andrea Ballabio, Laura S Schmidt, W Marston Linehan, Ming O Li.
      In metazoan organisms, cell competition acts as a quality control mechanism to eliminate unfit cells in favour of their more robust neighbours1,2. This mechanism has the potential to be maladapted, promoting the selection of aggressive cancer cells3-6. Tumours are metabolically active and are populated by stroma cells7,8, but how environmental factors affect cancer cell competition remains largely unknown. Here we show that tumour-associated macrophages (TAMs) can be dietarily or genetically reprogrammed to outcompete MYC-overexpressing cancer cells. In a mouse model of breast cancer, MYC overexpression resulted in an mTORC1-dependent 'winner' cancer cell state. A low-protein diet inhibited mTORC1 signalling in cancer cells and reduced tumour growth, owing unexpectedly to activation of the transcription factors TFEB and TFE3 and mTORC1 in TAMs. Diet-derived cytosolic amino acids are sensed by Rag GTPases through the GTPase-activating proteins GATOR1 and FLCN to control Rag GTPase effectors including TFEB and TFE39-14. Depletion of GATOR1 in TAMs suppressed the activation of TFEB, TFE3 and mTORC1 under the low-protein diet condition, causing accelerated tumour growth; conversely, depletion of FLCN or Rag GTPases in TAMs activated TFEB, TFE3 and mTORC1 under the normal protein diet condition, causing decelerated tumour growth. Furthermore, mTORC1 hyperactivation in TAMs and cancer cells and their competitive fitness were dependent on the endolysosomal engulfment regulator PIKfyve. Thus, noncanonical engulfment-mediated Rag GTPase-independent mTORC1 signalling in TAMs controls competition between TAMs and cancer cells, which defines a novel innate immune tumour suppression pathway that could be targeted for cancer therapy.
    DOI:  https://doi.org/10.1038/s41586-023-06256-5
  55. Autophagy. 2023 Jun 30. 1-2
    The active zone protein CLA-1 (Clarinet) bridges two subsynaptic domains to regulate presynaptic sorting of ATG-9.
    Zhao Xuan, Daniel A Colón-Ramos.
      In neuronal synapses, autophagosome biogenesis is coupled with the activity-dependent synaptic vesicle cycle via ATG-9. How vesicles containing ATG-9 are sorted at the presynapse is unknown. We performed forward genetic screens at single synapses of C. elegans neurons for mutants that disrupt ATG-9 presynaptic localization, and identified the long isoform of the active zone protein CLA-1 (Clarinet; CLA-1 L). We find that disrupting CLA-1 L results in abnormal accumulation of ATG-9-containing vesicles enriched with clathrin. The adaptor protein complexes and proteins at the periactive zone genetically interact with CLA-1 L in ATG-9 sorting. Moreover, the phenotype of the ATG-9 protein in cla-1(L) mutants was not observed for integral synaptic vesicle proteins, suggesting distinct mechanisms that regulate sorting of ATG-9-containing vesicles and synaptic vesicles. Our findings reveal novel roles for active zone proteins in the sorting of ATG-9 and in presynaptic macroautophagy/autophagy.
    Keywords:  ATG-9; CLA-1; active zone; adaptor protein complexes; periactive zone; synaptic vesicle cycle; syndapin 1
    DOI:  https://doi.org/10.1080/15548627.2023.2229227
  56. Cells. 2023 06 13. pii: 1619. [Epub ahead of print]12(12):
    A Novel Small NPC1 Promoter Enhances AAV-Mediated Gene Therapy in Mouse Models of Niemann-Pick Type C1 Disease.
    Michael Paul Hughes, Hemanth Ramesh Nelvagal, Oliver Coombe-Tennant, Dave Smith, Claire Smith, Giulia Massaro, Laura Poupon-Bejuit, Frances Mary Platt, Ahad Abdul Rahim.
      Niemann-Pick disease type C1 (NP-C) is a prematurely lethal genetic lysosomal storage disorder with neurological and visceral pathology resulting from mutations in the NPC1 gene encoding the lysosomal transmembrane protein NPC1. There is currently no cure for NP-C, and the only disease modifying treatment, miglustat, slows disease progression but does not significantly attenuate neurological symptoms. AAV-mediated gene therapy is an attractive option for NP-C, but due to the large size of the human NPC1 gene, there may be packaging and truncation issues during vector manufacturing. One option is to reduce the size of DNA regulatory elements that are essential for gene expression, such as the promoter sequence. Here, we describe a novel small truncated endogenous NPC1 promoter that leads to high gene expression both in vitro and in vivo and compare its efficacy to other commonly used promoters. Following neonatal intracerebroventricular (ICV) injection into the CNS, this novel promoter provided optimal therapeutic efficacy compared to all other promoters including increased survival, improved behavioural phenotypes, and attenuated neuropathology in mouse models of NP-C. Taken together, we propose that this novel promoter can be extremely efficient in designing an optimised AAV9 vector for gene therapy for NP-C.
    Keywords:  AAV; Niemann–Pick type C disease; gene therapy; mouse model; promoters
    DOI:  https://doi.org/10.3390/cells12121619
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