bims-auttor Biomed News
on Autophagy and mTOR
Issue of 2023‒01‒15
seventy papers selected by
Viktor Korolchuk, Newcastle University
  1. The Importance of mTORC1-Autophagy Axis for Skeletal Muscle Diseases.
  2. GATOR2 rings GATOR1 to speak to mTORC1.
  3. Glutamine Produces Ammonium to Tune Lysosomal pH and Regulate Lysosomal Function.
  4. GTP energy dependence of endocytosis and autophagy in the aging brain and Alzheimer's disease.
  5. Trilateral association of autophagy, mTOR and Alzheimer's disease: Potential pathway in the development for Alzheimer's disease therapy.
  6. LYSET/TMEM251/GCAF is critical for autophagy and lysosomal function by regulating the mannose-6-phosphate (M6P) pathway.
  7. High-content phenotypic screen to identify small molecule enhancers of Parkin-dependent ubiquitination and mitophagy.
  8. Cellular mitophagy: Mechanism, roles in diseases and small molecule pharmacological regulation.
  9. The Consequences of GBA Deficiency in the Autophagy-Lysosome System in Parkinson's Disease Associated with GBA.
  10. A gating mutation in ryanodine receptor type 2 rescues phenotypes of Alzheimer's disease mouse models by upregulating neuronal autophagy.
  11. Loss of TMEM106B exacerbates C9ALS/FTD DPR pathology by disrupting autophagosome maturation.
  12. YWHAE/14-3-3ε crotonylation regulates leucine deprivation-induced autophagy.
  13. Autophagy genes in biology and disease.
  14. A central role for regulated protein stability in the control of TFE3 and MITF by nutrients.
  15. Slingshot homolog-1 amplifies mitochondrial abnormalities by distinctly impairing health and clearance of mitochondria.
  16. Editorial: Highlights in Autophagy-From Basic Mechanisms to Human Disorder Treatments.
  17. Regulation of transferrin receptor trafficking by optineurin and its disease-associated mutants.
  18. Consecutive functions of small GTPases guide HOPS-mediated tethering of late endosomes and lysosomes.
  19. TTR (transthyretin) leads the autophagy disaster relief team against TARDBP/TDP-43 proteinopathy.
  20. Sleep-Disturbance-Induced Microglial Activation Involves CRH-Mediated Galectin 3 and Autophagy Dysregulation.
  21. A novel antimicrobial peptide M1-8 targets the lysosomal pathway to inhibit autolysosome formation and promote apoptosis in liver cancer cells.
  22. RIPosomes are targets of IRGM-SQSTM1-dependent autophagy.
  23. Sensitive imaging of Endoplasmic reticulum (ER) autophagy with an acidity-reporting ER-Tracker.
  24. Bortezomib abrogates temozolomide-induced autophagic flux through an ATG5 dependent pathway.
  25. Atg1 modulates mitochondrial dynamics to promote germline stem cell maintenance in Drosophila.
  26. A small molecule improves diabetes in mice expressing human islet amyloid polypeptide.
  27. Down-regulation of autophagy proteins is associated with higher mTOR expression in the placenta of pregnant women with preeclampsia.
  28. Relationship between Mitochondrial Quality Control Markers, Lower Extremity Tissue Composition, and Physical Performance in Physically Inactive Older Adults.
  29. OGT controls mammalian cell viability by regulating the proteasome/mTOR/ mitochondrial axis.
  30. LYSET/TMEM251- a novel key component of the mannose-6-phosphate pathway.
  31. Nalfurafine Hydrochloride, a κ-Opioid Receptor Agonist, Induces Melanophagy via PKA Inhibition in B16F1 Cells.
  32. Kaposi's sarcoma-associated herpesvirus (KSHV) utilizes the NDP52/CALCOCO2 selective autophagy receptor to disassemble processing bodies.
  33. Copper-dependent autophagic degradation of GPX4 drives ferroptosis.
  34. Microglial phagocytosis dysfunction in stroke is driven by energy depletion and induction of autophagy.
  35. Structure-Function Relationship for a Divergent Atg8 Protein Required for a Nonautophagic Function in Apicomplexan Parasites.
  36. Novel Insights into The Roles of N6-methyladenosine (m6A) Modification and Autophagy in Human Diseases.
  37. Adaptive autophagy reprogramming in Schwann cells during peripheral demyelination.
  38. The mitophagy receptor BNIP3 is critical for the regulation of metabolic homeostasis and mitochondrial function in the nucleus pulposus cells of the intervertebral disc.
  39. Aspirin alleviates pulmonary fibrosis through PI3K/AKT/mTOR-mediated autophagy pathway.
  40. Sesn2 Serves as a Regulator between Mitochondrial Unfolded Protein Response and Mitophagy in Intervertebral Disc Degeneration.
  41. Withaferin A Enhances Mitochondrial Biogenesis and BNIP3-Mediated Mitophagy to Promote Rapid Adaptation to Extreme Hypoxia.
  42. Targeting mTORC1 Activity to Improve Efficacy of Radioligand Therapy in Cancer.
  43. Modulation of protease expression by the transcription factor Ptx1/PITX regulates protein quality control during aging.
  44. Diabetes Influences the Fusion of Autophagosomes With Lysosomes in SH-SY5Y Cells and Induces Aβ Deposition and Cognitive Dysfunction in STZ-induced Diabetic Rats.
  45. L-Theanine Regulates the Abundance of Amino Acid Transporters in Mice Duodenum and Jejunum via the mTOR Signaling Pathway.
  46. E-cigarettes induce dysregulation of autophagy leading to endothelial dysfunction in pulmonary arterial hypertension.
  47. WWOX activates autophagy to alleviate lipopolysaccharide-induced acute lung injury by regulating mTOR.
  48. Cartilage-specific deficiency of clock gene Bmal1 accelerated articular cartilage degeneration in osteoarthritis by up-regulation of mTORC1 signaling.
  49. Mechanism of cystogenesis by Cd79a-driven, conditional mTOR activation in developing mouse nephrons.
  50. A role for BCL2L13 and autophagy in germline purifying selection of mtDNA.
  51. A tale of two pathways: Regulation of proteostasis by UPRmt and MDPs.
  52. The Greatwall-Endosulfine Switch Accelerates Autophagic Flux during the Cell Divisions Leading to G1 Arrest and Entry into Quiescence in Fission Yeast.
  53. PIK3C3/VPS34 keeps body fats healthy.
  54. Where is the field of autophagy research heading?
  55. Lipid Peroxidation and Iron Metabolism: Two Corner Stones in the Homeostasis Control of Ferroptosis.
  56. Chaperone-mediated autophagy promotes PCa survival during ARPI through selective proteome remodeling.
  57. Ubiquilin-2 regulates pathological alpha-synuclein.
  58. Synonymous Codon Variant Analysis for Autophagic Genes Dysregulated in Neurodegeneration.
  59. Metformin and Its Immune-Mediated Effects in Various Diseases.
  60. Nucleophagy contributes to genome stability through degradation of type II topoisomerases A and B and nucleolar components.
  61. Recent Advances in the Role of Autophagy in Endocrine-Dependent Tumors.
  62. mTORC1 implicated in Still's disease and MAS.
  63. Mucopolysaccharidoses: Cellular Consequences of Glycosaminoglycans Accumulation and Potential Targets.
  64. Inhibiting Cardiac Autophagy Suppresses Zika Virus Replication.
  65. Cryptosporidium parvum maintains intracellular survival by activating the host cellular EGFR-PI3K/Akt signaling pathway.
  66. SLC7A14 imports GABA to lysosomes and impairs hepatic insulin sensitivity via inhibiting mTORC2.
  67. Cigarette smoke extract-induced inflammatory response via inhibition of the TFEB-mediated autophagy in NR8383 cells.
  68. The CNC-family transcription factor Nrf3 coordinates the melanogenesis cascade through macropinocytosis and autophagy regulation.
  69. RIP3 impedes Mycobacterium tuberculosis survival and promotes p62-mediated autophagy.
  70. Why is rapamycin not a rapalog?
  1. Int J Mol Sci. 2022 Dec 24. pii: 297. [Epub ahead of print]24(1):
    The Importance of mTORC1-Autophagy Axis for Skeletal Muscle Diseases.
    Xujun Han, Kah Yong Goh, Wen Xing Lee, Sze Mun Choy, Hong-Wen Tang.
      The mechanistic target of rapamycin (mTOR) complex 1, mTORC1, integrates nutrient and growth factor signals with cellular responses and plays critical roles in regulating cell growth, proliferation, and lifespan. mTORC1 signaling has been reported as a central regulator of autophagy by modulating almost all aspects of the autophagic process, including initiation, expansion, and termination. An increasing number of studies suggest that mTORC1 and autophagy are critical for the physiological function of skeletal muscle and are involved in diverse muscle diseases. Here, we review recent insights into the essential roles of mTORC1 and autophagy in skeletal muscles and their implications in human muscle diseases. Multiple inhibitors targeting mTORC1 or autophagy have already been clinically approved, while others are under development. These chemical modulators that target the mTORC1/autophagy pathways represent promising potentials to cure muscle diseases.
    Keywords:  autophagy; mTORC1; muscle diseases
    DOI:  https://doi.org/10.3390/ijms24010297
  2. Mol Cell. 2023 Jan 05. pii: S1097-2765(22)01168-6. [Epub ahead of print]83(1): 6-8
    GATOR2 rings GATOR1 to speak to mTORC1.
    Umakant Sahu, Issam Ben-Sahra.
      The mechanistic target of rapamycin complex 1 (mTORC1) senses cellular leucine levels through the GATOR1/2-Rag axis. Jiang et al. show that the Ring domains of GATOR2 subunits maintain the integrity of the complex and promote ubiquitination and inhibition of GATOR1, thereby leading to mTORC1 activation.
    DOI:  https://doi.org/10.1016/j.molcel.2022.12.011
  3. Cells. 2022 Dec 24. pii: 80. [Epub ahead of print]12(1):
    Glutamine Produces Ammonium to Tune Lysosomal pH and Regulate Lysosomal Function.
    Jian Xiong, Thi Thu Trang Luu, Kartik Venkatachalam, Guangwei Du, Michael X Zhu.
      Glutamine is one of the most abundant amino acids in the cell. In mitochondria, glutaminases 1 and 2 (GLS1/2) hydrolyze glutamine to glutamate, which serves as the precursor of multiple metabolites. Here, we show that ammonium generated during GLS1/2-mediated glutaminolysis regulates lysosomal pH and in turn lysosomal degradation. In primary human skin fibroblasts BJ cells and mouse embryonic fibroblasts, deprivation of total amino acids for 1 h increased lysosomal degradation capacity as shown by the increased turnover of lipidated microtubule-associated proteins 1A/1B light chain 3B (LC3-II), several autophagic receptors, and endocytosed DQ-BSA. Removal of glutamine but not any other amino acids from the culture medium enhanced lysosomal degradation similarly as total amino acid starvation. The presence of glutamine in regular culture media increased lysosomal pH by >0.5 pH unit and the removal of glutamine caused lysosomal acidification. GLS1/2 knockdown, GLS1 antagonist, or ammonium scavengers reduced lysosomal pH in the presence of glutamine. The addition of glutamine or NH4Cl prevented the increase in lysosomal degradation and curtailed the extension of mTORC1 function during the early time period of amino acid starvation. Our findings suggest that glutamine tunes lysosomal pH by producing ammonium, which regulates lysosomal degradation to meet the demands of cellular activities. During the early stage of amino acid starvation, the glutamine-dependent mechanism allows more efficient use of internal reserves and endocytosed proteins to extend mTORC1 activation such that the normal anabolism is not easily interrupted by a brief disruption of the amino acid supply.
    Keywords:  amino acid starvation; autophagosome; autophagy; glutaminase; glutamine; lysosomal pH; mTORC1 activation
    DOI:  https://doi.org/10.3390/cells12010080
  4. Geroscience. 2023 Jan 09.
    GTP energy dependence of endocytosis and autophagy in the aging brain and Alzheimer's disease.
    Ricardo A Santana Martínez, Priyanka D Pinky, Benjamin A Harlan, Gregory J Brewer.
      Increased interest in the aging and Alzheimer's disease (AD)-related impairments in autophagy in the brain raise important questions about regulation and treatment. Since many steps in endocytosis and autophagy depend on GTPases, new measures of cellular GTP levels are needed to evaluate energy regulation in aging and AD. The recent development of ratiometric GTP sensors (GEVALS) and findings that GTP levels are not homogenous inside cells raise new issues of regulation of GTPases by the local availability of GTP. In this review, we highlight the metabolism of GTP in relation to the Rab GTPases involved in formation of early endosomes, late endosomes, and lysosomal transport to execute the autophagic degradation of damaged cargo. Specific GTPases control macroautophagy (mitophagy), microautophagy, and chaperone-mediated autophagy (CMA). By inference, local GTP levels would control autophagy, if not in excess. Additional levels of control are imposed by the redox state of the cell, including thioredoxin involvement. Throughout this review, we emphasize the age-related changes that could contribute to deficits in GTP and AD. We conclude with prospects for boosting GTP levels and reversing age-related oxidative redox shift to restore autophagy. Therefore, GTP levels could regulate the numerous GTPases involved in endocytosis, autophagy, and vesicular trafficking. In aging, metabolic adaptation to a sedentary lifestyle could impair mitochondrial function generating less GTP and redox energy for healthy management of amyloid and tau proteostasis, synaptic function, and inflammation.
    Keywords:  Aging; Alzheimer’s; Autophagy; Endocytosis; Energetics; GTP; Lysosomes; Mitophagy
    DOI:  https://doi.org/10.1007/s11357-022-00717-x
  5. Front Pharmacol. 2022 ;13 1094351
    Trilateral association of autophagy, mTOR and Alzheimer's disease: Potential pathway in the development for Alzheimer's disease therapy.
    Arunkumar Subramanian, T Tamilanban, Abdulrhman Alsayari, Gobinath Ramachawolran, Ling Shing Wong, Mahendran Sekar, Siew Hua Gan, Vetriselvan Subramaniyan, Suresh V Chinni, Nur Najihah Izzati Mat Rani, Nagaraja Suryadevara, Shadma Wahab.
      The primary and considerable weakening event affecting elderly individuals is age-dependent cognitive decline and dementia. Alzheimer's disease (AD) is the chief cause of progressive dementia, and it is characterized by irreparable loss of cognitive abilities, forming senile plaques having Amyloid Beta (Aβ) aggregates and neurofibrillary tangles with considerable amounts of tau in affected hippocampus and cortex regions of human brains. AD affects millions of people worldwide, and the count is showing an increasing trend. Therefore, it is crucial to understand the underlying mechanisms at molecular levels to generate novel insights into the pathogenesis of AD and other cognitive deficits. A growing body of evidence elicits the regulatory relationship between the mammalian target of rapamycin (mTOR) signaling pathway and AD. In addition, the role of autophagy, a systematic degradation, and recycling of cellular components like accumulated proteins and damaged organelles in AD, is also pivotal. The present review describes different mechanisms and signaling regulations highlighting the trilateral association of autophagy, the mTOR pathway, and AD with a description of inhibiting drugs/molecules of mTOR, a strategic target in AD. Downregulation of mTOR signaling triggers autophagy activation, degrading the misfolded proteins and preventing the further accumulation of misfolded proteins that inhibit the progression of AD. Other target mechanisms such as autophagosome maturation, and autophagy-lysosomal pathway, may initiate a faulty autophagy process resulting in senile plaques due to defective lysosomal acidification and alteration in lysosomal pH. Hence, the strong link between mTOR and autophagy can be explored further as a potential mechanism for AD therapy.
    Keywords:  Alzheimer’s disease; autophagy; dementia; mTOR pathway; tau protein
    DOI:  https://doi.org/10.3389/fphar.2022.1094351
  6. Autophagy. 2023 Jan 12.
    LYSET/TMEM251/GCAF is critical for autophagy and lysosomal function by regulating the mannose-6-phosphate (M6P) pathway.
    Bokai Zhang, Xi Yang, Ming Li.
      Vertebrate cells rely on mannose-6-phosphate (M6P) modifications to deliver most lumenal hydrolases to the lysosome. As a critical trafficking signal for lysosomal enzymes, the M6P biosynthetic pathway has been thoroughly investigated. However, its regulatory mechanism is largely unknown. Here, we summarize three recent studies that independently discovered LYSET/TMEM251/GCAF as a key regulator of the M6P pathway. LYSET/TMEM251 directly interacts with GNPT, the enzyme that catalyzes the transfer of M6P, and is critical for its activity and stability. Deleting LYSET/TMEM251 impairs the GNPT function and M6P modifications. Consequently, lysosomal enzymes are mistargeted for secretion. Defective lysosomes fail to degrade cargoes such as endocytic vesicles and autophagosomes, leading to a newly identified lysosomal storage disease in humans. These discoveries open up a new direction in the regulation of the M6P biosynthetic pathway.
    Keywords:  Autophagy; GNPT; M6P; TMEM251; lysosomal enzymes; lysosomal storage disease
    DOI:  https://doi.org/10.1080/15548627.2023.2167375
  7. SLAS Discov. 2023 Jan 03. pii: S2472-5552(22)13716-0. [Epub ahead of print]
    High-content phenotypic screen to identify small molecule enhancers of Parkin-dependent ubiquitination and mitophagy.
    Roberta Tufi, Emily H Clark, Tamaki Hoshikawa, Christiana Tsagkaraki, Jack Stanley, Kunitoshi Takeda, James M Staddon, Thomas Briston.
      Mitochondrial dysfunction and aberrant mitochondrial homeostasis are key aspects of Parkinson's disease (PD) pathophysiology. Mutations in PINK1 and Parkin proteins lead to autosomal recessive PD, suggesting that defective mitochondrial clearance via mitophagy is key in PD etiology. Accelerating the identification and/or removal of dysfunctional mitochondria could therefore provide a disease-modifying approach to treatment. To that end, we performed a high-content phenotypic screen (HCS) of ∼125,000 small molecules to identify compounds that positively modulate mitochondrial accumulation of the PINK1-Parkin-dependent mitophagy initiation marker p-Ser65-Ub in Parkin haploinsufficiency (Parkin +/R275W) human fibroblasts. Following confirmatory counter-screening and orthogonal assays, we selected compounds of interest that enhance mitophagy-related biochemical and functional endpoints in patient-derived fibroblasts. Identification of inhibitors of the ubiquitin-specific peptidase and negative regulator of mitophagy USP30 within our hits further validated our approach. The compounds identified provide a novel starting point for further investigation and optimisation.
    Keywords:  PINK1; Parkin; Parkinson's disease; USP30; high-content screening; mitophagy
    DOI:  https://doi.org/10.1016/j.slasd.2022.12.004
  8. Theranostics. 2023 ;13(2): 736-766
    Cellular mitophagy: Mechanism, roles in diseases and small molecule pharmacological regulation.
    Yingying Lu, Zhijia Li, Shuangqian Zhang, Tongtong Zhang, Yanjun Liu, Lan Zhang.
      Cellular mitophagy means that cells selectively wrap and degrade damaged mitochondria through an autophagy mechanism, thus maintaining mitochondria and intracellular homeostasis. In recent years, mitophagy has received increasing attention as a research hotspot related to the pathogenesis of clinical diseases, such as neurodegenerative diseases, cardiovascular diseases, cancer, metabolic diseases, and so on. It has been found that the regulation of mitophagy may become a new direction for the treatment of some diseases. In addition, numerous small molecule modulators of mitophagy have also been reported, which provides new opportunities to comprehend the procedure and potential of therapeutic development. Taken together, in this review, we summarize current understanding of the mechanism of mitophagy, discuss the roles of mitophagy and its relationship with diseases, introduce the existing small-molecule pharmacological modulators of mitophagy and further highlight the significance of their development.
    Keywords:  Diseases; Mechanism; Mitophagy; Small molecule modulators
    DOI:  https://doi.org/10.7150/thno.79876
  9. Cells. 2023 Jan 03. pii: 191. [Epub ahead of print]12(1):
    The Consequences of GBA Deficiency in the Autophagy-Lysosome System in Parkinson's Disease Associated with GBA.
    Eddie Pradas, Marta Martinez-Vicente.
      GBA gene variants were the first genetic risk factor for Parkinson's disease. GBA encodes the lysosomal enzyme glucocerebrosidase (GBA), which is involved in sphingolipid metabolism. GBA exhibits a complex physiological function that includes not only the degradation of its substrate glucosylceramide but also the metabolism of other sphingolipids and additional lipids such as cholesterol, particularly when glucocerebrosidase activity is deficient. In the context of Parkinson's disease associated with GBA, the loss of GBA activity has been associated with the accumulation of α-synuclein species. In recent years, several hypotheses have proposed alternative and complementary pathological mechanisms to explain why lysosomal enzyme mutations lead to α-synuclein accumulation and become important risk factors in Parkinson's disease etiology. Classically, loss of GBA activity has been linked to a dysfunctional autophagy-lysosome system and to a subsequent decrease in autophagy-dependent α-synuclein turnover; however, several other pathological mechanisms underlying GBA-associated parkinsonism have been proposed. This review summarizes and discusses the different hypotheses with a special focus on autophagy-dependent mechanisms, as well as autophagy-independent mechanisms, where the role of other players such as sphingolipids, cholesterol and other GBA-related proteins make important contributions to Parkinson's disease pathogenesis.
    Keywords:  GBA; Parkinson’s disease; alpha-synuclein; autophagy; lysosome
    DOI:  https://doi.org/10.3390/cells12010191
  10. J Neurosci. 2023 Jan 09. pii: JN-RM-1820-22. [Epub ahead of print]
    A gating mutation in ryanodine receptor type 2 rescues phenotypes of Alzheimer's disease mouse models by upregulating neuronal autophagy.
    Hua Zhang, Caitlynn Knight, S R Wayne Chen, Ilya Bezprozvanny.
      It is well established that Ryanodine receptors (RyanR) are overactive in Alzheimer's disease (AD) and it has been suggested that inhibition of RyanR is potentially beneficial for AD treatment. In the present study we explored a potential connection between basal RyanR activity and autophagy in neurons. Autophagy plays an important role in clearing damaged organelles and long-lived protein aggregates, and autophagy dysregulation occurs in both AD patients and AD animal models. Autophagy is known to be regulated by intracellular calcium (Ca2+) signals, and our results indicated that basal RyanR2 activity in hippocampal neurons inhibited autophagy through activation of calcineurin (CaN) and resulting inhibition of AMPK-ULK1 pathway. Thus, we hypothesized that increased basal RyanR2 activity in AD may lead to inhibition of neuronal autophagy and accumulation of β-amyloid. To test this hypothesis, we took advantage of the RyanR2-E4872Q knock-in mouse model (EQ) in which basal RyanR2 activity is reduced due to shortened channel open time. We discovered that crossing EQ mice with the APPKI and APPPS1 mouse models of AD (both males and females) rescued amyloid accumulation and LTP impairment in these mice. Our results revealed that reduced basal activity of RyanR2-EQ channels disinhibited the autophagic pathway and led to increased amyloid clearance in these models. These data indicated a potential pathogenic outcome of RyanR2 overactivation in AD and also provided additional targets for therapeutic intervention in AD.One Sentence Summary:Basal activity of ryanodine receptors controls neuronal autophagy and contributes to development of AD phenotypeSignificance Statement:It is well established that neuronal autophagy is impaired in Alzheimer's disease (AD). Our results suggest that supranormal calcium (Ca2+) release from endoplasmic reticulum contributes to inhibition of autophagy in AD and that reduction in basal activity of type 2 ryanodine receptors (RyanR2) disinhibits the neuronal autophagic pathway and led to increased amyloid clearance in AD models. Our findings directly link neuronal Ca2+ dysregulation with autophagy dysfunction in AD point to additional targets for therapeutic intervention.
    DOI:  https://doi.org/10.1523/JNEUROSCI.1820-22.2022
  11. Front Cell Neurosci. 2022 ;16 1061559
    Loss of TMEM106B exacerbates C9ALS/FTD DPR pathology by disrupting autophagosome maturation.
    Claudia S Bauer, Christopher P Webster, Allan C Shaw, Jannigje R Kok, Lydia M Castelli, Ya-Hui Lin, Emma F Smith, Francisco Illanes-Álvarez, Adrian Higginbottom, Pamela J Shaw, Mimoun Azzouz, Laura Ferraiuolo, Guillaume M Hautbergue, Andrew J Grierson, Kurt J De Vos.
      Disruption to protein homeostasis caused by lysosomal dysfunction and associated impairment of autophagy is a prominent pathology in amyotrophic lateral sclerosis and frontotemporal dementia (ALS/FTD). The most common genetic cause of ALS/FTD is a G4C2 hexanucleotide repeat expansion in C9orf72 (C9ALS/FTD). Repeat-associated non-AUG (RAN) translation of G4C2 repeat transcripts gives rise to dipeptide repeat (DPR) proteins that have been shown to be toxic and may contribute to disease etiology. Genetic variants in TMEM106B have been associated with frontotemporal lobar degeneration with TDP-43 pathology and disease progression in C9ALS/FTD. TMEM106B encodes a lysosomal transmembrane protein of unknown function that is involved in various aspects of lysosomal biology. How TMEM106B variants affect C9ALS/FTD is not well understood but has been linked to changes in TMEM106B protein levels. Here, we investigated TMEM106B function in the context of C9ALS/FTD DPR pathology. We report that knockdown of TMEM106B expression exacerbates the accumulation of C9ALS/FTD-associated cytotoxic DPR proteins in cell models expressing RAN-translated or AUG-driven DPRs as well as in C9ALS/FTD-derived iAstrocytes with an endogenous G4C2 expansion by impairing autophagy. Loss of TMEM106B caused a block late in autophagy by disrupting autophagosome to autolysosome maturation which coincided with impaired lysosomal acidification, reduced cathepsin activity, and juxtanuclear clustering of lysosomes. Lysosomal clustering required Rab7A and coincided with reduced Arl8b-mediated anterograde transport of lysosomes to the cell periphery. Increasing Arl8b activity in TMEM106B-deficient cells not only restored the distribution of lysosomes, but also fully rescued autophagy and DPR protein accumulation. Thus, we identified a novel function of TMEM106B in autophagosome maturation via Arl8b. Our findings indicate that TMEM106B variants may modify C9ALS/FTD by regulating autophagic clearance of DPR proteins. Caution should therefore be taken when considering modifying TMEM106B expression levels as a therapeutic approach in ALS/FTD.
    Keywords:  ALS/FTD; C9orf72; DPR; TMEM106B; autophagy
    DOI:  https://doi.org/10.3389/fncel.2022.1061559
  12. Autophagy. 2023 Jan 10.
    YWHAE/14-3-3ε crotonylation regulates leucine deprivation-induced autophagy.
    Zilong Zheng, Qing Zhong, Xianghua Yan.
      Macroautophagy/autophagy is an important process responsible for protein turnover and cell survival in amino acid-deprived conditions, especially for leucine (Leu). With the dramatic advances in mass spectrometry, many new post-translational modifications (PTMs) have been identified. However, whether these PTMs regulate autophagy remains unclear. Here we found global lysine crotonylation levels are significantly upregulated during Leu-deprivation-induced autophagy. A comprehensive crotonylome profiling showed that YWHA/14-3-3 proteins are significantly enriched in the Leu regulated-crotonylome. The inhibition of YWHAE/14-3-3ε crotonylation by mutating two crotonylated sites to arginine, K73R K78R, significantly attenuates autophagy induced by Leu deprivation. Molecular dynamics suggest that YWHAE K73 and K78 crotonylations decrease protein conformation and thermodynamic stability. Moreover, we found crotonylation of YWHAE releases PPM1B to dephosphorylate ULK1 and consequently activate autophagy. Decrotonylation of YWHAE is mediated by HDAC7 whose activity is inhibited significantly by Leu deprivation. Taken together, our finding reveals a critical role of YWHAE crotonylation in Leu deprivation-induced autophagy.
    Keywords:  14-3-3ε; HDAC7; PPM1B; autophagy; crotonylation; leucine deprivation
    DOI:  https://doi.org/10.1080/15548627.2023.2166276
  13. Nat Rev Genet. 2023 Jan 12.
    Autophagy genes in biology and disease.
    Hayashi Yamamoto, Sidi Zhang, Noboru Mizushima.
      Macroautophagy and microautophagy are highly conserved eukaryotic cellular processes that degrade cytoplasmic material in lysosomes. Both pathways involve characteristic membrane dynamics regulated by autophagy-related proteins and other molecules, some of which are shared between the two pathways. Over the past few years, the application of new technologies, such as cryo-electron microscopy, coevolution-based structural prediction and in vitro reconstitution, has revealed the functions of individual autophagy gene products, especially in autophagy induction, membrane reorganization and cargo recognition. Concomitantly, mutations in autophagy genes have been linked to human disorders, particularly neurodegenerative diseases, emphasizing the potential pathogenic implications of autophagy defects. Accumulating genome data have also illuminated the evolution of autophagy genes within eukaryotes as well as their transition from possible ancestral elements in prokaryotes.
    DOI:  https://doi.org/10.1038/s41576-022-00562-w
  14. Mol Cell. 2023 Jan 05. pii: S1097-2765(22)01170-4. [Epub ahead of print]83(1): 57-73.e9
    A central role for regulated protein stability in the control of TFE3 and MITF by nutrients.
    Christopher Nardone, Brad A Palanski, Daniel C Scott, Richard T Timms, Karl W Barber, Xin Gu, Aoyue Mao, Yumei Leng, Emma V Watson, Brenda A Schulman, Philip A Cole, Stephen J Elledge.
      The TFE3 and MITF master transcription factors maintain metabolic homeostasis by regulating lysosomal, melanocytic, and autophagy genes. Previous studies posited that their cytosolic retention by 14-3-3, mediated by the Rag GTPases-mTORC1, was key for suppressing transcriptional activity in the presence of nutrients. Here, we demonstrate using mammalian cells that regulated protein stability plays a fundamental role in their control. Amino acids promote the recruitment of TFE3 and MITF to the lysosomal surface via the Rag GTPases, activating an evolutionarily conserved phospho-degron and leading to ubiquitination by CUL1β-TrCP and degradation. Elucidation of the minimal functional degron revealed a conserved alpha-helix required for interaction with RagA, illuminating the molecular basis for a severe neurodevelopmental syndrome caused by missense mutations in TFE3 within the RagA-TFE3 interface. Additionally, the phospho-degron is recurrently lost in TFE3 genomic translocations that cause kidney cancer. Therefore, two divergent pathologies converge on the loss of protein stability regulation by nutrients.
    Keywords:  MITF; Rag GTPases; TFE3; kidney cancer; lysosomes; mTORC1; neurodevelopment; nutrient-sensing; phospho-degron; ubiquitin; β-TrCP
    DOI:  https://doi.org/10.1016/j.molcel.2022.12.013
  15. Hum Mol Genet. 2023 Jan 13. pii: ddad006. [Epub ahead of print]
    Slingshot homolog-1 amplifies mitochondrial abnormalities by distinctly impairing health and clearance of mitochondria.
    Sara Cazzaro, Xingyu Zhao, Victoria K Zhao, Yenna K Kim, Jung-A A Woo.
      Accumulating toxic protein assemblies, including Aβ and tau, and dysfunctional mitochondria are associated with synaptic and neuronal loss in Alzheimer's disease (ad). Such accumulations are thought to be due to clearance defects in the autophagy-lysosome pathway. Mitochondrial dysfunction is evident in ad brains and animal models at multiple levels, such as mitochondrial genomic mutations, disrupted bioenergetics, deregulated mitochondrial dynamics, and impaired clearance of damaged mitochondria (mitophagy). Slingshot Homolog-1 (SSH1) is a phosphatase activated by oxidative stress, high intracellular levels of Ca+2, and Aβ42 oligomers (Aβ42O), known for its function to dephosphorylate/activate cofilin through the N-terminal region. SSH1-mediated cofilin dephosphorylation results in Ab42O-induced severing of F-actin and translocation of cofilin to mitochondria, which promotes mitochondria-mediated apoptosis, synaptic loss, and synaptic deficits. On the other hand, SSH1-mediated dephosphorylation/deactivation of the autophagy-cargo receptor p62 (SQSTM1), through its C-terminal region, inhibits p62 autophagy flux. However, the interplay between these two different activities of SSH1 in Aβ42O-induced mitochondrial toxicity remains unclear. In this study, we assessed the role of endogenous SSH1 and different regions of SSH1 in regulating mitochondrial health, mitochondrial respiration, clearance of damaged mitochondria, and synaptic integrity in vitro and in vivo. Our results indicate that SSH1 suppresses mitochondrial health and respiration through the cofilin-binding N-terminal region, whereas SSH1 impairs mitophagy through a newly identified ~ 100 residue p62-binding domain in the C-terminal region. These results indicate that both N-terminal and C-terminal regions negatively impact mitochondria by distinct and independent modalities to amplify mitochondrial abnormalities, making SSH1 an excellent target to mitigate ad pathogenesis.
    DOI:  https://doi.org/10.1093/hmg/ddad006
  16. Cells. 2023 Jan 03. pii: 188. [Epub ahead of print]12(1):
    Editorial: Highlights in Autophagy-From Basic Mechanisms to Human Disorder Treatments.
    Pei-Hui Lin, Lydie Combaret.
      Autophagy is an evolutionarily conserved catabolic process and represents a field of research that is constantly growing [...].
    DOI:  https://doi.org/10.3390/cells12010188
  17. Prog Mol Biol Transl Sci. 2023 ;pii: S1877-1173(22)00081-3. [Epub ahead of print]194 67-78
    Regulation of transferrin receptor trafficking by optineurin and its disease-associated mutants.
    Shivranjani C Moharir, Kapil Sirohi, Ghanshyam Swarup.
      Transferrin receptor (TFRC) is a transmembrane protein that plays a crucial role in mediating homeostasis of iron in the cell. The binding of transferrin (that is bound to iron) to TFRC at the cell membrane generally starts endocytosis of TFRC-transferrin complex, which leads to formation of vesicles that are positive for TFRC. These vesicles travel to the early endosomes and later to the endocytic recycling compartment. Release of iron occurs in the early endosomes because of acidic pH. Major fraction of the transferrin and TFRC is transported back to the cell membrane; however, a minor fraction of it is transported to lysosomes through the process of autophagy. Optineurin (OPTN) is a multi-functional adaptor protein that plays a pivotal role in the control of TFRC trafficking, recycling and autophagy dependent degradation. Optineurin also plays a role in cargo-selective and non-selective autophagy. Here, we review our understanding of the function of OPTN in regulating TFRC trafficking, recycling and autophagy dependent degradation. We also discuss the mechanisms by which certain disease-associated mutations of OPTN alter these processes.
    Keywords:  Autophagosome; Membrane delivery; Membrane trafficking; Optineurin; Recycling endosome; Transferrin receptor
    DOI:  https://doi.org/10.1016/bs.pmbts.2022.06.019
  18. Cell Rep. 2023 Jan 04. pii: S2211-1247(22)01873-3. [Epub ahead of print]42(1): 111969
    Consecutive functions of small GTPases guide HOPS-mediated tethering of late endosomes and lysosomes.
    Ariane Schleinitz, Lara-Alina Pöttgen, Tal Keren-Kaplan, Jing Pu, Paul Saftig, Juan S Bonifacino, Albert Haas, Andreas Jeschke.
      The transfer of endocytosed cargoes to lysosomes (LYSs) requires HOPS, a multiprotein complex that tethers late endosomes (LEs) to LYSs before fusion. Many proteins interact with HOPS on LEs/LYSs. However, it is not clear whether these HOPS interactors localize to LEs or LYSs or how they participate in tethering. Here, we biochemically characterized endosomes purified from untreated or experimentally manipulated cells to put HOPS and interacting proteins in order and to establish their functional interdependence. Our results assign Rab2a and Rab7 to LEs and Arl8 and BORC to LYSs and show that HOPS drives LE-LYS fusion by bridging late endosomal Rab2a with lysosomal BORC-anchored Arl8. We further show that Rab7 is absent from sites of HOPS-dependent tethering but promotes fusion by moving LEs toward LYSs via dynein. Thus, our study identifies the topology of the machinery for LE-LYS tethering and elucidates the role of different small GTPases in the process.
    Keywords:  CP: Cell biology; autophagosome; autophagy; dynein; endolysosome; lysosome reformation; membrane fusion; membrane tethering; phagocytosis; phosphoinositides; protein trafficking
    DOI:  https://doi.org/10.1016/j.celrep.2022.111969
  19. Autophagy. 2023 Jan 12.
    TTR (transthyretin) leads the autophagy disaster relief team against TARDBP/TDP-43 proteinopathy.
    Yuan-Ping Chu, Pei-Chuan Ho, Kuen-Jer Tsai.
      TTR (transthyretin) strikes a neuroprotective function in the prevention of amyloid-β (Aβ) deposition in Alzheimer disease (AD). Perturbation of the stringently controlled TARDBP/TDP-43 (TAR DNA binding protein) expression gives rise to cytoplasmic aggregation, characterized by TARDBP proteinopathy affiliated with several neurological disorders, including frontotemporal lobar degeneration with TARDBP pathology (FTLD-TDP) and amyotrophic lateral sclerosis/ALS. Proposedly, TTR can maintain cellular proteostasis susceptible to TARDBP aggregates and initiate its removal. Herein, we disclose that TTR upregulated in response to excessive TARDBP causes TARDBP aggregation in FTLD-TDP and co-accumulates with it. Moreover, TTR expression increases with age in FTLD-TDP but shows a downward decline in the elderly. TTR promotes macroautophagy/autophagy activity and facilitates aggregated TARDBP degradation via autophagy. Compellingly, TTR binds to ATF4 and boosts its nuclear import for autophagy upregulation. Therefore, TTR directs autophagy teamwork in bi-directional regulation through enhancing autophagy activity via ATF4 and chaperoning aggregated TARDBP to phagophores for degradation.
    Keywords:  ATF4; FTLD; TDP-43; TTR; autophagy; proteinopathy
    DOI:  https://doi.org/10.1080/15548627.2023.2167690
  20. Cells. 2022 Dec 30. pii: 160. [Epub ahead of print]12(1):
    Sleep-Disturbance-Induced Microglial Activation Involves CRH-Mediated Galectin 3 and Autophagy Dysregulation.
    Liyang Guo, Kirstin M Reed, Ashley Carter, Yan Cheng, Soheil Kazemi Roodsari, Damian Martinez Pineda, Laurie L Wellman, Larry D Sanford, Ming-Lei Guo.
      Chronic sleep disturbances (CSDs) including insomnia, insufficient sleep time, and poor sleep quality are major public health concerns around the world, especially in developed countries. CSDs are major health risk factors linked to multiple neurodegenerative and neuropsychological diseases. It has been suggested that CSDs could activate microglia (Mg) leading to increased neuroinflammation levels, which ultimately lead to neuronal dysfunction. However, the detailed mechanisms underlying CSD-mediated microglial activation remain mostly unexplored. In this study, we used mice with three-weeks of sleep fragmentation (SF) to explore the underlying pathways responsible for Mg activation. Our results revealed that SF activates Mg in the hippocampus (HP) but not in the striatum and prefrontal cortex (PFc). SF increased the levels of corticotropin-releasing hormone (CRH) in the HP. In vitro mechanism studies revealed that CRH activation of Mg involves galectin 3 (Gal3) upregulation and autophagy dysregulation. CRH could disrupt lysosome membrane integrity resulting in lysosomal cathepsins leakage. CRHR2 blockage mitigated CRH-mediated effects on microglia in vitro. SF mice also show increased Gal3 levels and autophagy dysregulation in the HP compared to controls. Taken together, our results show that SF-mediated hippocampal Mg activation involves CRH mediated galectin 3 and autophagy dysregulation. These findings suggest that targeting the hippocampal CRH system might be a novel therapeutic approach to ameliorate CSD-mediated neuroinflammation and neurodegenerative diseases.
    Keywords:  CRH; autophagy; microglia; neuroinflammation; sleep fragmentation
    DOI:  https://doi.org/10.3390/cells12010160
  21. J Cell Mol Med. 2023 Jan 11.
    A novel antimicrobial peptide M1-8 targets the lysosomal pathway to inhibit autolysosome formation and promote apoptosis in liver cancer cells.
    Jiali Zeng, Jian Wang, Jibin Wu, Rui Deng, Lun Zhang, Qingru Chen, Jie Wang, Xiaobao Jin, Shuiqing Gui, Yinghua Xu, Xuemei Lu.
      Lysosomes, a central regulator of autophagy, play a critical role in tumour growth. Lysosomal protease cathepsin D can initiate apoptosis when released from lysosomes into the cytosol. In this study, we observed that Musca domestica cecropin (Mdc) 1-8 (M1-8), a small anti-tumour peptide derived from Mdc, inhibits hepatoma cell growth by blocking autophagy-lysosome fusion. This effect is likely achieved by targeting lysosomes to activate lysosomal protease D. Additionally, we examined whether lysosomal content and cathepsin D release were involved in M1-8-induced apoptosis. After exposure to M1-8, human hepatoma HepG2 cells rapidly co-localized with lysosomes, disrupted lysosomal integrity, caused leakage of lysosomal protease cathepsin D, caspase activation and mitochondrial membrane potential changes; and promoted cell apoptosis. Interestingly, in M1-8-treated HepG2 cells, autophagic protein content increased and the lysosome-autophagosome fusion was inhibited, suggesting that M1-8 can cause apoptosis through autophagy and lysosomes. This result indicates that a small accumulation of autophagy and autolysosome inhibition in cells can cause cell death. Taken together, these data suggest a novel insight into the regulatory mechanisms of M1-8 in autophagy and lysosomes, which may facilitate the development of M1-8 as a potential cancer therapeutic agent.
    Keywords:  antimicrobial peptides; autophagy; liver cancer; lysosomes
    DOI:  https://doi.org/10.1111/jcmm.17644
  22. Autophagy. 2023 Jan 10.
    RIPosomes are targets of IRGM-SQSTM1-dependent autophagy.
    Subhash Mehto, Soumya Kundu, Swati Chauhan, Santosh Chauhan.
      The NOD1-NOD2-RIPK2-NFKB/NF-κB pro-inflammatory axis plays a significant role in regulating the immune response to bacterial infection. However, an excess of NFKB-dependent cytokine response can be detrimental and, thus, should be kept under control to maintain the innate immune balance. In our recent study, first, we showed that bacterial infection induces the biogenesis of RIPK2 oligomers (RIPosomes) that are recruited around the bacteria to enhance an NFKB-dependent pro-inflammatory response. Next, we showed that SQSTM1- and IRGM-dependent selective macroautophagy/autophagy degrades RIPosomes and their components to limit NOD1-NOD2-RIPK2-NFKB pro-inflammatory signaling. Consistently, depletion of IRGM results in an augmented RIPK2-dependent pro-inflammatory cytokine response induced by Shigella flexneri and Salmonella typhimurium. Further, bacterial infection- and DSS-induced gut inflammation in irgm1KO mice is dampened upon therapeutic inhibition of RIPK2. Taken together, we showed that autophagy selectively degrades RIPosomes to suppress inflammation and maintain innate immune homeostasis.
    Keywords:  Autophagy; IRGM; NFKB; NOD1; NOD2; RIPK2; RIPosomes
    DOI:  https://doi.org/10.1080/15548627.2023.2166724
  23. Autophagy. 2023 Jan 12. 1-11
    Sensitive imaging of Endoplasmic reticulum (ER) autophagy with an acidity-reporting ER-Tracker.
    Yilong Shi, Xiaoxue Zou, Xianghui Zheng, Yimin Wu, Jiahuai Han, Shoufa Han.
      Macroautophagic/autophagic turnover of endoplasmic reticulum (reticulophagy) is critical for cell health. Herein we reported a sensitive fluorescence-on imaging of reticulophagy using a small molecule probe (ER-proRed) comprised of green-emissive fluorinated rhodol for ER targeting and nonfluorescent rhodamine-lactam prone to lysosome-triggered red fluorescence. Partitioned in ER to exhibit green fluorescence, ER-proRed gives intense red fluorescence upon co-delivery with ER into acidic lysosomes. Serving as the signal of reticulophagy, the turning on of red fluorescence enables discernment of reticulophagy induced by starvation, varied levels of reticulophagic receptors, and chemical agents such as etoposide and sodium butyrate. These results show ER probes optically activatable in lysosomes, such as ER-proRed, offer a sensitive and simplified tool for studying reticulophagy in biology and diseases.Abbreviations: Baf-A1, bafilomycin A1; CCCP, carbonyl cyanide m-chlorophenyl hydrazone; CQ, chloroquine diphosphate; ER, endoplasmic reticulum; FHR, fluorinated hydrophobic rhodol; GFP, green fluorescent protein; Reticulophagy, selective autophagy of ER; RFP, red fluorescent protein; ROX, X-rhodamine; UPR, unfolded protein response.
    Keywords:  Autophagy imaging; endoplasmic reticulum; fluorescence-on; lysosomal acidity; reticulophagy
    DOI:  https://doi.org/10.1080/15548627.2023.2165759
  24. Front Cell Dev Biol. 2022 ;10 1022191
    Bortezomib abrogates temozolomide-induced autophagic flux through an ATG5 dependent pathway.
    Mohummad Aminur Rahman, Agnete S T Engelsen, Shahin Sarowar, Christian Bindesbøll, Even Birkeland, Dorota Goplen, Maria L Lotsberg, Stian Knappskog, Anne Simonsen, Martha Chekenya.
      Introduction: Glioblastoma (GBM) is invariably resistant to temozolomide (TMZ) chemotherapy. Inhibiting the proteasomal pathway is an emerging strategy to accumulate damaged proteins and inhibit their lysosomal degradation. We hypothesized that pre-treatment of glioblastoma with bortezomib (BTZ) might sensitize glioblastoma to temozolomide by abolishing autophagy survival signals to augment DNA damage and apoptosis. Methods: P3 patient-derived glioblastoma cells, as well as the tumour cell lines U87, HF66, A172, and T98G were investigated for clonogenic survival after single or combined treatment with temozolomide and bortezomib in vitro. We investigated the requirement of functional autophagy machinery by utilizing pharmacological inhibitors or CRISPR-Cas9 knockout (KO) of autophagy-related genes -5 and -7 (ATG5 and ATG7) in glioblastoma cells and monitored changes in autophagic flux after temozolomide and/or bortezomib treatments. P3 wild-type and P3 ATG5-/- (ATG5 KO) cells were implanted orthotopically into NOD-SCID mice to assess the efficacy of bortezomib and temozolomide combination therapy with and without functional autophagy machinery. Results: The chemo-resistant glioblastoma cells increased autophagic flux during temozolomide treatment as indicated by increased degradation of long-lived proteins, diminished expression of autophagy markers LC3A/B-II and p62 (SQSTM1), increased co-localisation of LC3A/B-II with STX17, augmented and no induction of apoptosis. In contrast, bortezomib treatment abrogated autophagic flux indicated by the accumulation of LC3A/B-II and p62 (SQSTM1) positive autophagosomes that did not fuse with lysosomes and thus reduced the degradation of long-lived proteins. Bortezomib synergistically enhanced temozolomide efficacy by attenuating cell proliferation, increased DNA double-strand breaks, and apoptosis in an autophagy-dependent manner. Abolishing autophagy in ATG5 KOs reversed the bortezomib-induced toxicity, rescued glioblastoma cell death and reduced animal survival. Discussion: We conclude that bortezomib abrogates temozolomide induced autophagy flux through an ATG5 dependent pathway.
    Keywords:  apoptosis; autophagy flux; cell cycle kinetics; chemosensitization; glioblastoma (GBM); proteasome inhibitor bortezomib; secretome; temozolomide chemotherapy
    DOI:  https://doi.org/10.3389/fcell.2022.1022191
  25. Biochem Biophys Res Commun. 2022 Nov 30. pii: S0006-291X(22)01633-3. [Epub ahead of print]643 192-202
    Atg1 modulates mitochondrial dynamics to promote germline stem cell maintenance in Drosophila.
    Minal S Ayachit, Bhupendra V Shravage.
      Mitochondrial dynamics (fusion and fission) are necessary for stem cell maintenance and differentiation. However, the relationship between mitophagy, mitochondrial dynamics and stem cell exhaustion needs to be clearly understood. Here we report the multifaceted role of Atg1 in mitophagy, mitochondrial dynamics and stem cell maintenance in female germline stem cells (GSCs) in Drosophila. We found that depletion of Atg1 in GSCs leads to impaired autophagy and mitophagy as measured by reduced formation of autophagosomes, increased accumulation of p62/Ref (2)P and accumulation of damaged mitochondria. Disrupting Atg1 function led to mitochondrial fusion in developing cysts. The fusion resulted from an increase in Marf levels in both GSCs and cysts, and the fusion phenotype could be rescued by overexpression of Drp1 or by depleting Marf via RNAi in Atg1-depleted cyst cells. Interestingly, double knockdown of both Atg1:Drp1 led to the significant loss of germ cells (GCs) as compared to Atg1KD and Drp1KD. Strikingly, Atg1:Marf double knockdown leads to a dramatic loss of GSCs, GCs and a total loss of vitellogenic stages, suggesting a block in oogenesis. Overall, our results demonstrate that Drp1, Marf and Atg1 function together to influence female GSC maintenance, their differentiation into cysts and oogenesis in Drosophila.
    Keywords:  Atg1; Drp1; Homeostasis; Marf; Mitochondria; Mitophagy; Oogenesis
    DOI:  https://doi.org/10.1016/j.bbrc.2022.11.076
  26. Islets. 2023 Dec 31. 15(1): 12-15
    A small molecule improves diabetes in mice expressing human islet amyloid polypeptide.
    Vriti Bhagat, C Bruce Verchere.
      In recent years, the number of studies on islet and beta cell autophagy have substantially increased due to growing interest in the role of autophagy in maintaining cellular homeostasis in diabetes. In type 2 diabetes, human islet amyloid polypeptide (hIAPP) aggregates to form higher structure oligomers and fibrils that are toxic to beta cells and induce islet inflammation. The primary mechanism of oligomer and fibril clearance in beta cells is through the autophagic pathway, a process that is impaired in type 2 diabetes. Thus, toxic oligomeric and fibrillar forms of hIAPP accumulate in type 2 diabetic islets. Recently, Kim et al. characterized the ability of a small molecule autophagy enhancer, MSL-7, to clear hIAPP oligomers in mice expressing hIAPP. Herein, we outline the primary findings of the study, limitations, and future directions to further investigate the therapeutic potential of autophagy enhancers to treat diabetes.
    Keywords:  Islets; amylin; amyloid; autophagy; beta cells; islet amyloid polypeptide; small molecule
    DOI:  https://doi.org/10.1080/19382014.2022.2163829
  27. Braz J Med Biol Res. 2023 ;pii: S0100-879X2022000100683. [Epub ahead of print]55 e12283
    Down-regulation of autophagy proteins is associated with higher mTOR expression in the placenta of pregnant women with preeclampsia.
    I C Weel, V R Ribeiro, M Romão-Veiga, E G Fioratti, J C Peraçoli, M T S Peraçoli.
      Autophagy is a lysosomal degradation pathway that removes protein aggregates and damaged organelles maintaining cellular integrity. It seems to be essential for cell survival during stress, starvation, hypoxia, and consequently to the placenta implantation and development. Preeclampsia (PE) is a multisystemic disorder characterized by the onset of hypertension associated or not with proteinuria and other maternal complications. Considering that the placenta seems to play an important role in the pathogenesis of PE, the objective of the present study was to evaluate protein levels of light chain protein (LC3), beclin-1, and the mammalian target of rapamycin (mTOR) in the placenta of pregnant women with PE. Placental tissues collected from 20 women with PE and 20 normotensive (NT) pregnant women were evaluated for LC3, beclin-1, and mTOR expression by qPCR and immunohistochemistry. The mRNA for LC3 and beclin-1 were significantly lower, while mTOR gene expression was significantly higher in the placenta of pregnant women with PE than in the NT group. Placentas of PE women showed significantly decreased protein expression of LC3-II and beclin-1, whereas mTOR was significantly increased compared with the NT pregnant women. There was a negative correlation between protein expression of mTOR and LC3-II in the placental tissue of PE women. In conclusion, the results showed autophagy deficiency suggesting that failure in this degradation process may contribute to the pathogenesis of PE; however, new studies involving cross-talk between autophagy and inflammatory molecular mechanisms might help to better understand the autophagy process in this obstetric pathology.
    DOI:  https://doi.org/10.1590/1414-431X2022e12283
  28. Cells. 2023 Jan 02. pii: 183. [Epub ahead of print]12(1):
    Relationship between Mitochondrial Quality Control Markers, Lower Extremity Tissue Composition, and Physical Performance in Physically Inactive Older Adults.
    Anna Picca, Matthew Triolo, Stephanie E Wohlgemuth, Matthew S Martenson, Robert T Mankowski, Stephen D Anton, Emanuele Marzetti, Christiaan Leeuwenburgh, David A Hood.
      Altered mitochondrial quality and function in muscle may be involved in age-related physical function decline. The role played by the autophagy-lysosome system, a major component of mitochondrial quality control (MQC), is incompletely understood. This study was undertaken to obtain initial indications on the relationship between autophagy, mitophagy, and lysosomal markers in muscle and measures of physical performance and lower extremity tissue composition in young and older adults. Twenty-three participants were enrolled, nine young (mean age: 24.3 ± 4.3 years) and 14 older adults (mean age: 77.9 ± 6.3 years). Lower extremity tissue composition was quantified volumetrically by magnetic resonance imaging and a tissue composition index was calculated as the ratio between muscle and intermuscular adipose tissue volume. Physical performance in older participants was assessed via the Short Physical Performance Battery (SPPB). Protein levels of the autophagy marker p62, the mitophagy mediator BCL2/adenovirus E1B 19 kDa protein-interacting protein 3 (BNIP3), the lysosomal markers transcription factor EB, vacuolar-type ATPase, and lysosomal-associated membrane protein 1 were measured by Western immunoblotting in vastus lateralis muscle biopsies. Older adults had smaller muscle volume and lower tissue composition index than young participants. The protein content of p62 and BNIP3 was higher in older adults. A negative correlation was detected between p62 and BNIP3 and the tissue composition index. p62 and BNIP3 were also related to the performance on the 5-time sit-to-stand test of the SPPB. Our results suggest that an altered expression of markers of the autophagy/mitophagy-lysosomal system is related to deterioration of lower extremity tissue composition and muscle dysfunction. Additional studies are needed to clarify the role of defective MQC in human muscle aging and identify novel biological targets for drug development.
    Keywords:  Short Physical Performance Battery (SPPB); aging; functional decline; intermuscular adipose tissue (IMAT); lysosomes; mitochondrial dysfunction; mitochondrion; muscle aging; physical performance; sarcopenia
    DOI:  https://doi.org/10.3390/cells12010183
  29. Proc Natl Acad Sci U S A. 2023 Jan 17. 120(3): e2218332120
    OGT controls mammalian cell viability by regulating the proteasome/mTOR/ mitochondrial axis.
    Xiang Li, Xiaojing Yue, Hugo Sepulveda, Rajan A Burt, David A Scott, Steven A Carr, Samuel A Myers, Anjana Rao.
      O-GlcNAc transferase (OGT) modifies serine and threonine residues on nuclear and cytosolic proteins with O-linked N-acetylglucosamine (GlcNAc). OGT is essential for mammalian cell viability, but the underlying mechanisms are still enigmatic. We performed a genome-wide CRISPR-Cas9 screen in mouse embryonic stem cells (mESCs) to identify candidates whose depletion rescued the block in cell proliferation induced by OGT deficiency. We show that the block in cell proliferation in OGT-deficient cells stems from mitochondrial dysfunction secondary to mTOR (mechanistic target of rapamycin) hyperactivation. In normal cells, OGT maintains low mTOR activity and mitochondrial fitness through suppression of proteasome activity; in the absence of OGT, increased proteasome activity results in increased steady-state amino acid levels, which in turn promote mTOR lysosomal translocation and activation, and increased oxidative phosphorylation. mTOR activation in OGT-deficient mESCs was confirmed by an independent phospho-proteomic screen. Our study highlights a unique series of events whereby OGT regulates the proteasome/ mTOR/ mitochondrial axis in a manner that maintains homeostasis of intracellular amino acid levels, mitochondrial fitness, and cell viability. A similar mechanism operates in CD8+ T cells, indicating its generality across mammalian cell types. Manipulating OGT activity may have therapeutic potential in diseases in which this signaling pathway is impaired.
    Keywords:  OGT; genome-wide CRISPR/Cas9 screen; mTOR; mitochondrion; proteasome
    DOI:  https://doi.org/10.1073/pnas.2218332120
  30. Autophagy. 2023 Jan 12.
    LYSET/TMEM251- a novel key component of the mannose-6-phosphate pathway.
    Wenjie Qiao, Christopher M Richards, Sabrina Jabs.
      ​​Degradation of macromolecules delivered to lysosomes by processes such as autophagy or endocytosis is crucial for cellular function. Lysosomes require more than 60 soluble hydrolases in order to catabolize such macromolecules. These soluble hydrolases are tagged with mannose-6-phosphate (M6P) moieties in sequential reactions by the Golgi-resident GlcNAc-1-phosphotransferase complex and NAGPA/UCE/uncovering enzyme (N-acetylglucosamine-1-phosphodiester alpha-N-acetylglucosaminidase), which allows their delivery to endosomal/lysosomal compartments through trafficking mediated by cation-dependent and -independent mannose-6-phosphate receptors (MPRs). We and others recently identified TMEM251 as a novel regulator of the M6P pathway via independent genome-wide genetic screening strategies. We renamed TMEM251 to LYSET (lysosomal enzyme trafficking factor) to establish nomenclature reflective to this gene's function. LYSET is a Golgi-localized transmembrane protein important for the retention of the GlcNAc-1-phosphotransferase complex in the Golgi-apparatus. The current understanding of LYSET's importance regarding human biology is 3-fold: 1) highly pathogenic viruses that depend on lysosomal hydrolase activity require LYSET for infection. 2) The presence of LYSET is critical for cancer cell proliferation in nutrient-deprived environments in which extracellular proteins must be catabolized. 3) Inherited pathogenic alleles of LYSET can cause a severe inherited disease which resembles GlcNAc-1-phosphotransferase deficiency (i.e., mucolipidosis type II).
    Keywords:  GlcNAc-1-phosphotransferase; Golgi-apparatus; lysosomal enzyme trafficking; lysosome; mannose-6-phosphate; mucolipidosis type II
    DOI:  https://doi.org/10.1080/15548627.2023.2167376
  31. Cells. 2022 Dec 29. pii: 146. [Epub ahead of print]12(1):
    Nalfurafine Hydrochloride, a κ-Opioid Receptor Agonist, Induces Melanophagy via PKA Inhibition in B16F1 Cells.
    Ha Jung Lee, Seong Hyun Kim, Yong Hwan Kim, So Hyun Kim, Gyeong Seok Oh, Ji-Eun Bae, Joon Bum Kim, Na Yeon Park, Kyuhee Park, Eunbyul Yeom, Kwiwan Jeong, Pansoo Kim, Doo Sin Jo, Dong-Hyung Cho.
      Selective autophagy controls cellular homeostasis by degrading unnecessary or damaged cellular components. Melanosomes are specialized organelles that regulate the biogenesis, storage, and transport of melanin in melanocytes. However, the mechanisms underlying melanosomal autophagy, known as the melanophagy pathway, are poorly understood. To better understand the mechanism of melanophagy, we screened an endocrine-hormone chemical library and identified nalfurafine hydrochlorides, a κ-opioid receptor agonist, as a potent inducer of melanophagy. Treatment with nalfurafine hydrochloride increased autophagy and reduced melanin content in alpha-melanocyte-stimulating hormone (α-MSH)-treated cells. Furthermore, inhibition of autophagy blocked melanosomal degradation and reversed the nalfurafine hydrochloride-induced decrease in melanin content in α-MSH-treated cells. Consistently, treatment with other κ-opioid receptor agonists, such as MCOPPB or mianserin, inhibited excessive melanin production but induced autophagy in B16F1 cells. Furthermore, nalfurafine hydrochloride inhibited protein kinase A (PKA) activation, which was notably restored by forskolin, a PKA activator. Additionally, forskolin treatment further suppressed melanosomal degradation as well as the anti-pigmentation activity of nalfurafine hydrochloride in α-MSH-treated cells. Collectively, our data suggest that stimulation of κ-opioid receptors induces melanophagy by inhibiting PKA activation in α-MSH-treated B16F1 cells.
    Keywords:  B16F1 cells; autophagy; melanophagy; nalfurafine hydrochloride; κ-opioid receptor
    DOI:  https://doi.org/10.3390/cells12010146
  32. PLoS Pathog. 2023 Jan 12. 19(1): e1011080
    Kaposi's sarcoma-associated herpesvirus (KSHV) utilizes the NDP52/CALCOCO2 selective autophagy receptor to disassemble processing bodies.
    Carolyn-Ann Robinson, Gillian K Singh, Mariel Kleer, Thalia Katsademas, Elizabeth L Castle, Bre Q Boudreau, Jennifer A Corcoran.
      Kaposi's sarcoma-associated herpesvirus (KSHV) causes the inflammatory and angiogenic endothelial cell neoplasm, Kaposi's sarcoma (KS). We previously demonstrated that the KSHV Kaposin B (KapB) protein promotes inflammation via the disassembly of cytoplasmic ribonucleoprotein granules called processing bodies (PBs). PBs modify gene expression by silencing or degrading labile messenger RNAs (mRNAs), including many transcripts that encode inflammatory or angiogenic proteins associated with KS disease. Although our work implicated PB disassembly as one of the causes of inflammation during KSHV infection, the precise mechanism used by KapB to elicit PB disassembly was unclear. Here we reveal a new connection between the degradative process of autophagy and PB disassembly. We show that both latent KSHV infection and KapB expression enhanced autophagic flux via phosphorylation of the autophagy regulatory protein, Beclin. KapB was necessary for this effect, as infection with a recombinant virus that does not express the KapB protein did not induce Beclin phosphorylation or autophagic flux. Moreover, we showed that PB disassembly mediated by KSHV or KapB, depended on autophagy genes and the selective autophagy receptor NDP52/CALCOCO2 and that the PB scaffolding protein, Pat1b, co-immunoprecipitated with NDP52. These studies reveal a new role for autophagy and the selective autophagy receptor NDP52 in promoting PB turnover and the concomitant synthesis of inflammatory molecules during KSHV infection.
    DOI:  https://doi.org/10.1371/journal.ppat.1011080
  33. Autophagy. 2023 Jan 12. 1-15
    Copper-dependent autophagic degradation of GPX4 drives ferroptosis.
    Qian Xue, Ding Yan, Xi Chen, Xiaofen Li, Rui Kang, Daniel J Klionsky, Guido Kroemer, Xin Chen, Daolin Tang, Jinbao Liu.
      Ferroptosis is a type of iron-dependent regulated cell death characterized by unrestricted lipid peroxidation and membrane damage. Although GPX4 (glutathione peroxidase 4) plays a master role in blocking ferroptosis by eliminating phospholipid hydroperoxides, the regulation of GPX4 remains poorly understood. Here, we report an unexpected role for copper in promoting ferroptotic cell death, but not cuproptosis, by inducing macroautophagic/autophagic degradation of GPX4. Copper chelators reduce ferroptosis sensitivity but do not inhibit other types of cell death, such as apoptosis, necroptosis, and alkaliptosis. Conversely, exogenous copper increases GPX4 ubiquitination and the formation of GPX4 aggregates by directly binding to GPX4 protein cysteines C107 and C148. TAX1BP1 (Tax1 binding protein 1) then acts as an autophagic receptor for GPX4 degradation and subsequent ferroptosis in response to copper stress. Consequently, copper enhances ferroptosis-mediated tumor suppression in a mouse model of pancreatic cancer tumor, whereas copper chelators attenuate experimental acute pancreatitis associated with ferroptosis. Taken together, these findings provide new insights into the link between metal stress and autophagy-dependent cell death.Abbreviations: CALCOCO2, calcium binding and coiled-coil domain 2; GPX4, glutathione peroxidase 4; MAP1LC3A/B, microtubule associated protein 1 light chain 3 alpha/beta; MPO, myeloperoxidase; NCOA4, nuclear receptor coactivator 4; OPTN, optineurin; PDAC, pancreatic ductal adenocarcinoma; RIPK1, receptor interacting serine/threonine kinase 1; ROS, reactive oxygen species; SLC40A1, solute carrier family 40 member 1; SQSTM1, sequestosome 1; TAX1BP1, Tax1 binding protein 1; TEPA, tetraethylenepentamine; TM, tetrathiomolybdate.
    Keywords:  Autophagy; GPX4; TAX1BP1; copper; cuproptosis; ferroptosis
    DOI:  https://doi.org/10.1080/15548627.2023.2165323
  34. Autophagy. 2023 Jan 09.
    Microglial phagocytosis dysfunction in stroke is driven by energy depletion and induction of autophagy.
    Sol Beccari, Virginia Sierra-Torre, Jorge Valero, Marta Pereira-Iglesias, Mikel García-Zaballa, Federico N Soria, Laura De Las Heras-Garcia, Alejandro Carretero-Guillen, Estibaliz Capetillo-Zarate, Maria Domercq, Paloma R Huguet, David Ramonet, Ahmed Osman, Wei Han, Cecilia Dominguez, Travis E Faust, Omar Touzani, Olatz Pampliega, Patricia Boya, Dorothy Schafer, Guillermo Mariño, Emmanuelle Canet-Soulas, Klas Blomgren, Ainhoa Plaza-Zabala, Amanda Sierra.
      Microglial phagocytosis of apoptotic debris prevents buildup damage of neighbor neurons and inflammatory responses. Whereas microglia are very competent phagocytes under physiological conditions, we report their dysfunction in mouse and preclinical monkey models of stroke (macaques and marmosets) by transient occlusion of the medial cerebral artery (tMCAo). By analyzing recently published bulk and single cell RNA sequencing databases, we show that the phagocytosis dysfunction was not explained by transcriptional changes. In contrast, we demonstrate that the impairment of both engulfment and degradation was related to energy depletion triggered by oxygen and nutrient deprivation (OND), which led to reduced process motility, lysosomal exhaustion, and the induction of a protective macroautophagy/autophagy response in microglia. Basal autophagy, in charge of removing and recycling intracellular elements, was critical to maintain microglial physiology, including survival and phagocytosis, as we determined both in vivo and in vitro using pharmacological and transgenic approaches. Notably, the autophagy inducer rapamycin partially prevented the phagocytosis impairment induced by tMCAo in vivo but not by OND in vitro, where it even had a detrimental effect on microglia, suggesting that modulating microglial autophagy to optimal levels may be a hard to achieve goal. Nonetheless, our results show that pharmacological interventions, acting directly on microglia or indirectly on the brain environment, have the potential to recover phagocytosis efficiency in the diseased brain. We propose that phagocytosis is a therapeutic target yet to be explored in stroke and other brain disorders and provide evidence that it can be modulated in vivo using rapamycin.
    Keywords:  autophagy; ischemia; lysosomes; microglia; phagocytosis; rapamycin; stroke; tMCAo
    DOI:  https://doi.org/10.1080/15548627.2023.2165313
  35. mBio. 2023 Jan 10. e0364221
    Structure-Function Relationship for a Divergent Atg8 Protein Required for a Nonautophagic Function in Apicomplexan Parasites.
    Marta Walczak, Thomas R Meister, Hoa Mai Nguyen, Yili Zhu, Sébastien Besteiro, Ellen Yeh.
      Atg8 family proteins are highly conserved eukaryotic proteins with diverse autophagy and nonautophagic functions in eukaryotes. While the structural features required for conserved autophagy functions of Atg8 are well established, little is known about the molecular changes that facilitated acquisition of divergent, nonautophagic functions of Atg8. The malaria parasite Plasmodium falciparum offers a unique opportunity to study nonautophagic functions of Atg8 family proteins because it encodes a single Atg8 homolog whose only essential function is in the inheritance of an unusual secondary plastid called the apicoplast. Here, we used functional complementation to investigate the structure-function relationship for this divergent Atg8 protein. We showed that the LC3-interacting region (LIR) docking site (LDS), the major interaction interface of the Atg8 protein family, is required for P. falciparum Atg8 (PfAtg8) apicoplast localization and function, likely via Atg8 lipidation. On the other hand, another region previously implicated in canonical Atg8 interactions, the N-terminal helix, is not required for apicoplast-specific PfAtg8 function. Finally, our investigations at the cellular level demonstrate that the unique apicomplexan-specific loop, previously implicated in interaction with membrane conjugation machinery in recombinant protein-based in vitro assays, is not required for membrane conjugation nor for the apicoplast-specific effector function of Atg8 in both P. falciparum and related Apicomplexa member Toxoplasma gondii. These results suggest that the effector function of apicomplexan Atg8 is mediated by structural features distinct from those previously identified for macroautophagy and selective autophagy functions. IMPORTANCE The most extensively studied role of Atg8 proteins is in autophagy. However, it is clear that they have other nonautophagic functions critical to cell function and disease pathogenesis that are so far understudied compared to their canonical role in autophagy. Mammalian cells contain multiple Atg8 paralogs that have diverse, specialized functions. Gaining molecular insight into their nonautophagic functions is difficult because of redundancy between the homologs and their role in both autophagy and nonautophagic pathways. Malaria parasites such as Plasmodium falciparum are a unique system to study a novel, nonautophagic function of Atg8 separate from its role in autophagy: they have only one Atg8 protein whose only essential function is in the inheritance of the apicoplast, a unique secondary plastid organelle. Insights into the molecular basis of PfAtg8's function in apicoplast biogenesis will have important implications for the evolution of diverse nonautophagic functions of the Atg8 protein family.
    Keywords:  Atg8; Plasmodium; Toxoplasma; apicomplexan parasites; apicoplast; malaria; nonautophagic function of Atg8
    DOI:  https://doi.org/10.1128/mbio.03642-21
  36. Int J Biol Sci. 2023 ;19(2): 705-720
    Novel Insights into The Roles of N6-methyladenosine (m6A) Modification and Autophagy in Human Diseases.
    Jiaxin Liang, Jingwen Sun, Wei Zhang, Xiwen Wang, Ying Xu, Yuan Peng, Ling Zhang, Wenqian Xiong, Yi Liu, Hengwei Liu.
      Autophagy is an evolutionarily conserved cellular degradation and recycling process. It is important for maintaining vital cellular function and metabolism. Abnormal autophagy activity can cause the development of various diseases. N6-methyladenosine (m6A) methylation is the most prevalent and abundant internal modification in eukaryotes, affecting almost all aspects of RNA metabolism. The process of m6A modification is dynamic and adjustable. Its regulation depends on the regulation of m6A methyltransferases, m6A demethylases, and m6A binding proteins. m6A methylation and autophagy are two crucial and independent cellular events. Recent studies have shown that m6A modification mediates the transcriptional and post-transcriptional regulation of autophagy-related genes, affecting autophagy regulatory networks in multiple diseases. However, the regulatory effects of m6A regulators on autophagy in human diseases are not adequately acknowledged. In the present review, we summarized the latest knowledge of m6A modification in autophagy and elucidated the molecular regulatory mechanisms underlying m6A modification in autophagy regulatory networks. Moreover, we discuss the potentiality of m6A regulators serving as promising predictive biomarkers for human disease diagnosis and targets for therapy. This review will increase our understanding of the relationship between m6A methylation and autophagy, and provide novel insights to specifically target m6A modification in autophagy-associated therapeutic strategies.
    Keywords:  Autophagy; Biomarkers; N6-methyladenosine (m6A); RNA modification; Therapeutic targets
    DOI:  https://doi.org/10.7150/ijbs.75466
  37. Cell Mol Life Sci. 2023 Jan 09. 80(1): 34
    Adaptive autophagy reprogramming in Schwann cells during peripheral demyelination.
    Young Rae Jo, Yuna Oh, Young Hee Kim, Yoon Kyung Shin, Hye Ran Kim, Hana Go, Jaekyoon Shin, Hye Ji Park, Hyongjong Koh, Jong Kuk Kim, Jung Eun Shin, Kyung Eun Lee, Hwan Tae Park.
      The myelin sheath is an essential structure for the rapid transmission of electrical impulses through axons, and peripheral myelination is a well-programmed postnatal process of Schwann cells (SCs), the myelin-forming peripheral glia. SCs transdifferentiate into demyelinating SCs (DSCs) to remove the myelin sheath during Wallerian degeneration after axonal injury and demyelinating neuropathies, and macrophages are responsible for the degradation of myelin under both conditions. In this study, the mechanism by which DSCs acquire the ability of myelin exocytosis was investigated. Using serial ultrastructural evaluation, we found that autophagy-related gene 7-dependent formation of a "secretory phagophore (SP)" and tubular phagophore was necessary for exocytosis of large myelin chambers by DSCs. DSCs seemed to utilize myelin membranes for SP formation and employed p62/sequestosome-1 (p62) as an autophagy receptor for myelin excretion. In addition, the acquisition of the myelin exocytosis ability of DSCs was associated with the decrease in canonical autolysosomal flux and was demonstrated by p62 secretion. Finally, this SC demyelination mechanism appeared to also function in inflammatory demyelinating neuropathies. Our findings show a novel autophagy-mediated myelin clearance mechanism by DSCs in response to nerve damage.
    Keywords:  Autophagy; Demyelinating neuropathy; Demyelination; SQSTM1/p62; Wallerian degeneration
    DOI:  https://doi.org/10.1007/s00018-022-04683-7
  38. Autophagy. 2023 Jan 10. 1-23
    The mitophagy receptor BNIP3 is critical for the regulation of metabolic homeostasis and mitochondrial function in the nucleus pulposus cells of the intervertebral disc.
    Vedavathi Madhu, Miriam Hernandez-Meadows, Paige K Boneski, Yunping Qiu, Anyonya R Guntur, Irwin J Kurland, Ruteja A Barve, Makarand V Risbud.
      The contribution of mitochondria to the metabolic function of hypoxic NP cells has been overlooked. We have shown that NP cells contain networked mitochondria and that mitochondrial translocation of BNIP3 mediates hypoxia-induced mitophagy. However, whether BNIP3 also plays a role in governing mitochondrial function and metabolism in hypoxic NP cells is not known. BNIP3 knockdown altered mitochondrial morphology, and number, and increased mitophagy. Interestingly, BNIP3 deficiency in NP cells reduced glycolytic capacity reflected by lower production of lactate/H+ and lower ATP production rate. Widely targeted metabolic profiling and flux analysis using 1-2-13C-glucose showed that the BNIP3 loss resulted in redirection of glycolytic flux into pentose phosphate and hexosamine biosynthesis as well as pyruvate resulting in increased TCA flux. An overall reduction in one-carbon metabolism was noted suggesting reduced biosynthesis. U13C-glutamine flux analysis showed preservation of glutamine utilization to maintain TCA intermediates. The transcriptomic analysis of the BNIP3-deficient cells showed dysregulation of cellular functions including membrane and cytoskeletal integrity, ECM-growth factor signaling, and protein quality control with an overall increase in themes related to angiogenesis and innate immune response. Importantly, we observed strong thematic similarities with the transcriptome of a subset of human degenerative samples. Last, we noted increased autophagic flux, decreased disc height index and aberrant COL10A1/collagen X expression, signs of early disc degeneration in young adult bnip3 knockout mice. These results suggested that in addition to mitophagy regulation, BNIP3 plays a role in maintaining mitochondrial function and metabolism, and dysregulation of mitochondrial homeostasis could promote disc degeneration.Abbreviations: ECAR extracellular acidification rate; HIF hypoxia inducible factor; MFA metabolic flux analysis; NP nucleus pulposus; OCR oxygen consumption rate; ShBnip3 short-hairpin Bnip3.
    Keywords:  BNIP3; disc degeneration; hypoxia; intervertebral disc; metabolism; mitochondria; mitophagy; nucleus pulposus
    DOI:  https://doi.org/10.1080/15548627.2022.2162245
  39. Exp Gerontol. 2023 Jan 06. pii: S0531-5565(23)00006-2. [Epub ahead of print] 112085
    Aspirin alleviates pulmonary fibrosis through PI3K/AKT/mTOR-mediated autophagy pathway.
    Jieting Peng, Xun Xiao, Shizhen Li, Xing Lyu, Hui Gong, Shengyu Tan, Lini Dong, Yan Y Sanders, Xiangyu Zhang.
      Idiopathic pulmonary fibrosis (IPF) is a chronic and irreversible lung disease with limited therapeutic options. Aspirin can alleviate liver, kidney, and cardiac fibrosis. However, its role in lung fibrosis is unclear. This study aims to investigate the effects of aspirin on lung fibroblast differentiation and pulmonary fibrosis. TGF-β1-induced human embryonic lung fibroblasts, IPF lung fibroblasts, and bleomycin-induced lung fibrosis mouse model were used in this study. The results showed that aspirin significantly decreased the expression of Collagen 1A1, Fibronectin, Alpha-smooth muscle actin, and equestosome1, and increased the ratio of light chain 3 beta II/I and the number of autophagosome in vivo and in vitro; reduced bleomycin-induced lung fibrosis. Aspirin also decreased the ratios of phosphorylated phosphatidylinositol 3 kinase (p-PI3K)/PI3K, protein kinase B (p-AKT)/AKT, and mechanistic target of rapamycin (p-mTOR)/mTOR in vitro. Autophagy inhibitor 3-methyladenine, bafilomycin-A1, and AKT activator SC-79 abrogated the effects of aspirin. These findings indicate that aspirin ameliorates pulmonary fibrosis through a PI3K/AKT/mTOR-dependent autophagy pathway.
    Keywords:  Aspirin; Autophagy; Idiopathic pulmonary fibrosis; Lung fibroblasts; PI3K/AKT/mTOR
    DOI:  https://doi.org/10.1016/j.exger.2023.112085
  40. Int J Biol Sci. 2023 ;19(2): 571-592
    Sesn2 Serves as a Regulator between Mitochondrial Unfolded Protein Response and Mitophagy in Intervertebral Disc Degeneration.
    Wen-Ning Xu, Chun Liu, Huo-Liang Zheng, Hai-Xia Xu, Run-Ze Yang, Sheng-Dan Jiang, Li-Xin Zhu.
      Mitochondrial unfold protein response (UPRmt) can induce mitophagy to protect cell from unfold protein. However, how UPRmt induces mitophagy to protect cell is not yet clear. Herein, Sesn2 was considered to be a key molecule that communicated UPRmt and mitophagy in the intervertebral disc. Silencing of Sesn2 was able to reverse the protective effects of Nicotinamide riboside (NR) on nucleus pulposus (NP) cells and inhibit mitophagy induced by UPRmt. UPRmt upregulated Sesn2 through Eif2ak4/eIF2α/Atf4, and further induced mitophagy. Sesn2 promoted the translocation of cytosolic Parkin and Sqstm1 to the defective mitochondria respectively, thereby enhancing mitophagy. The translocation of cytosolic Sqstm1 to the defective mitochondria was dependent on Parkin. The two functional domains of Sesn2 were necessary for the interaction of Sesn2 with Parkin and Sqstm1. The cytosolic interaction of Sesn2 between Parkin and Sqstm1 was independent on Pink1 (named as PINK1 in human) but the mitochondrial translocation was dependent on Pink1. Sesn2-/- mice showed a more severe degeneration and NR did not completely alleviate the intervertebral disc degeneration (IVDD) of Sesn2-/- mice. In conclusion, UPRmt could attenuate IVDD by upregulation of Sesn2-induced mitophagy. This study will help to further reveal the mechanism of Sesn2 regulating mitophagy, and open up new ideas for the prevention and treatment of IVDD.
    Keywords:  Intervertebral disc degeneration; Mitochondrial unfold protein response; Mitophagy.; Sesn2
    DOI:  https://doi.org/10.7150/ijbs.70211
  41. Cells. 2022 Dec 25. pii: 85. [Epub ahead of print]12(1):
    Withaferin A Enhances Mitochondrial Biogenesis and BNIP3-Mediated Mitophagy to Promote Rapid Adaptation to Extreme Hypoxia.
    Ruzhou Zhao, Yixin Xu, Xiaobo Wang, Xiang Zhou, Yanqi Liu, Shuai Jiang, Lin Zhang, Zhibin Yu.
      Rapid adaptation to extreme hypoxia is a challenging problem, and there is no effective scheme to achieve rapid adaptation to extreme hypoxia. In this study, we found that withaferin A (WA) can significantly reduce myocardial damage, maintain cardiac function, and improve survival in rats in extremely hypoxic environments. Mechanistically, WA protects against extreme hypoxia by affecting BCL2-interacting protein 3 (BNIP3)-mediated mitophagy and the peroxisome proliferator-activated receptor γ coactivator 1α (PGC-1α)-mediated mitochondrial biogenesis pathway among mitochondrial quality control mechanisms. On the one hand, enhanced mitophagy eliminates hypoxia-damaged mitochondria and prevents the induction of apoptosis; on the other hand, enhanced mitochondrial biogenesis can supplement functional mitochondria and maintain mitochondrial respiration to ensure mitochondrial ATP production under acute extreme hypoxia. Our study shows that WA can be used as an effective drug to improve tolerance to extreme hypoxia.
    Keywords:  BNIP3; PGC-1α; mitochondrial biogenesis; mitophagy; withaferin A
    DOI:  https://doi.org/10.3390/cells12010085
  42. Cancers (Basel). 2022 Dec 20. pii: 17. [Epub ahead of print]15(1):
    Targeting mTORC1 Activity to Improve Efficacy of Radioligand Therapy in Cancer.
    Michal Grzmil, Fabius Wiesmann, Roger Schibli, Martin Behe.
      Radioligand therapy (RLT) represents an effective strategy to treat malignancy by cancer-selective delivery of radioactivity following systemic application. Despite recent therapeutic successes, cancer radioresistance and insufficient delivery of the radioactive ligands, as well as cytotoxicity to healthy organs, significantly impairs clinical efficacy. To improve disease management while minimizing toxicity, in recent years, the combination of RLT with molecular targeted therapies against cancer signaling networks showed encouraging outcomes. Characterization of the key deregulated oncogenic signaling pathways revealed their convergence to activate the mammalian target of rapamycin (mTOR), in which signaling plays an essential role in the regulation of cancer growth and survival. Therapeutic interference with hyperactivated mTOR pathways was extensively studied and led to the development of mTOR inhibitors for clinical applications. In this review, we outline the regulation and oncogenic role of mTOR signaling, as well as recapitulate and discuss mTOR complex 1 (mTORC1) inhibition to improve the efficacy of RLT in cancer.
    Keywords:  PRRT; RLT; TRT; mTOR; mammalian target of rapamycin; radioligand therapy; radiosensitization; rapalogs; targeted radionuclide therapy
    DOI:  https://doi.org/10.3390/cancers15010017
  43. Cell Rep. 2023 Jan 10. pii: S2211-1247(22)01874-5. [Epub ahead of print]42(1): 111970
    Modulation of protease expression by the transcription factor Ptx1/PITX regulates protein quality control during aging.
    Jianqin Jiao, Michelle Curley, Flavia A Graca, Maricela Robles-Murguia, Abbas Shirinifard, David Finkelstein, Beisi Xu, Yiping Fan, Fabio Demontis.
      Protein quality control is important for healthy aging and is dysregulated in age-related diseases. The autophagy-lysosome and ubiquitin-proteasome are key for proteostasis, but it remains largely unknown whether other proteolytic systems also contribute to maintain proteostasis during aging. Here, we find that expression of proteolytic enzymes (proteases/peptidases) distinct from the autophagy-lysosome and ubiquitin-proteasome systems declines during skeletal muscle aging in Drosophila. Age-dependent protease downregulation undermines proteostasis, as demonstrated by the increase in detergent-insoluble poly-ubiquitinated proteins and pathogenic huntingtin-polyQ levels in response to protease knockdown. Computational analyses identify the transcription factor Ptx1 (homologous to human PITX1/2/3) as a regulator of protease expression. Consistent with this model, Ptx1 protein levels increase with aging, and Ptx1 RNAi counteracts the age-associated downregulation of protease expression. Moreover, Ptx1 RNAi improves muscle protein quality control in a protease-dependent manner and extends lifespan. These findings indicate that proteases and their transcriptional modulator Ptx1 ensure proteostasis during aging.
    Keywords:  CP: Cell biology; CP: Molecular biology; PITX; Ptx1; aging; betaTry; huntingtin; peptidase; protease; protein quality control; proteostasis; skeletal muscle
    DOI:  https://doi.org/10.1016/j.celrep.2022.111970
  44. Behav Brain Res. 2023 Jan 04. pii: S0166-4328(23)00004-9. [Epub ahead of print] 114286
    Diabetes Influences the Fusion of Autophagosomes With Lysosomes in SH-SY5Y Cells and Induces Aβ Deposition and Cognitive Dysfunction in STZ-induced Diabetic Rats.
    Lou-Yan Ma, Song-Fang Liu, Ya-Gang Guo, Zheng-Quan Ma, Ya Li, Shu-Jin Wang, Yu Niu, Mo Li, Jia-Jia Zhai, Su-Hang Shang, Ya-Li Lv, Qiu-Min Qu.
      Diabetes has been regarded as an independent risk factor for Alzheimer's disease (AD). Our previous study found that diabetes activated autophagy, but lysosome function was impaired. Autophagy-lysosome dysfunction may be involved in Aβ deposition in diabetic cognitive impairment. In the present study, we used STZ-induced diabetic rats and SH-SY5Y cells to investigate whether diabetes inhibits autophagosome fusion with lysosomes. We found that in the in vivo study, STZ-induced diabetic rats exhibited cognitive dysfunction, and the lysosome function-related factors CTSL, CTSD, and Rab7 were decreased (P<0.05). In an in vitro study, the mRFP-GFP-LC3 assay showed that the fusion of autophagosomes with lysosomes was partly blocked in SH-SY5Y cells. High glucose treatment downregulated the number of autophagolysosomes, downregulated CTSD, CTSL, and Rab7 expression (P<0.05), and then influenced the function of ACP2 to partly block the fusion of autophagosomes and lysosomes to inhibit Aβ clearance. These findings indicate that high glucose treatment affected lysosome function, interfered with the fusion of autophagosomes with lysosomes, and partly blocked autophagic flux to influence Aβ clearance.
    Keywords:  Aβ; Diabetes; autophagy flux; fusion; lysosome
    DOI:  https://doi.org/10.1016/j.bbr.2023.114286
  45. Nutrients. 2022 Dec 28. pii: 142. [Epub ahead of print]15(1):
    L-Theanine Regulates the Abundance of Amino Acid Transporters in Mice Duodenum and Jejunum via the mTOR Signaling Pathway.
    Kehong Liu, Yingqi Peng, Ling Lin, Zhihua Gong, Wenjun Xiao, Yinhua Li.
      The intestine is a key organ for the absorption of amino acids. L-theanine (LTA) is a structural analog of glutamine and a characteristic non-protein amino acid found in tea (Camellia sinensis) that regulates lipid and protein metabolism. The present study explored the role of LTA in intestinal amino acid absorption, protein synthesis, and its mechanisms. Overall, our findings suggest that LTA supplementation not only affects serum alkaline phosphatase (AKP), total protein (TP), and urea nitrogen (BUN) levels, but it also upregulates the mRNA and protein expression of amino acid transporters (EAAT3, EAAT1, 4F2hc, y+LAT1, CAT1, ASCT2, and B0AT1), and activates the mTOR signaling pathway. The downstream S6 and S6K1 proteins are regulated, and the expression of amino acid transporters is regulated. These findings suggest that LTA increases intestinal AA absorption, promotes protein metabolism, and increases nitrogen utilization by upregulating AAT expression, activating the mTOR signaling pathway, and phosphorylating the mTOR downstream proteins S6 and S6K1.
    Keywords:  L-theanine; S6; S6K1; amino acid transporters; mTOR signaling pathway
    DOI:  https://doi.org/10.3390/nu15010142
  46. Stem Cells. 2023 Jan 14. pii: sxad004. [Epub ahead of print]
    E-cigarettes induce dysregulation of autophagy leading to endothelial dysfunction in pulmonary arterial hypertension.
    Chen-Wei Liu, Hoai Huong Thi Le, Philip Denaro Iii, Zhiyu Dai, Ning-Yi Shao, Sang-Ging Ong, Won Hee Lee.
      Given the increasing popularity of electronic cigarettes (e-cigs), it is imperative to evaluate the potential health risks of e-cigs, especially in users with preexisting health concerns such as pulmonary arterial hypertension (PAH). The aim of the present study was to investigate whether differential susceptibility exists between healthy and PAH patients to e-cig exposure and the molecular mechanisms contributing to it. Patient-specific induced pluripotent stem cell-derived endothelial cells (iPSC-ECs) from healthy individuals and PAH patients were used to investigate whether e-cig contributes to the pathophysiology of PAH and affects EC homeostasis in PAH. Our results showed that PAH iPSC-ECs showed a greater amount of damage than healthy iPSC-ECs upon e-cig exposure. Transcriptomic analyses revealed that differential expression of Akt3 may be responsible for increased autophagic flux impairment in PAH iPSC-ECs, which underlies increased susceptibility upon e-cig exposure. Moreover, knockdown of Akt3 in healthy iPSC-ECs significantly induced autophagic flux impairment and endothelial dysfunction, which further increased with e-cig treatment, thus mimicking the PAH cell phenotype after e-cig exposure. In addition, functional disruption of mTORC2 by knocking down Rictor in PAH iPSC-ECs caused autophagic flux impairment, which was mediated by downregulation of Akt3. Finally, pharmacological induction of autophagy via direct inhibition of mTORC1 and indirect activation of mTORC2 with rapamycin reverses e-cig-induced decreased Akt3 expression, endothelial dysfunction, autophagic flux impairment, and decreased cell viability and migration in PAH iPSC-ECs. Taken together, these data suggest a potential link between autophagy and Akt3-mediated increased susceptibility to e-cig in PAH.
    Keywords:  Akt3; autophagic flux; e-cigarettes; endothelial dysfunction; human induced pluripotent stem cell-derived endothelial cells; pulmonary arterial hypertension
    DOI:  https://doi.org/10.1093/stmcls/sxad004
  47. Int Immunopharmacol. 2023 Jan 06. pii: S1567-5769(22)01156-0. [Epub ahead of print]115 109671
    WWOX activates autophagy to alleviate lipopolysaccharide-induced acute lung injury by regulating mTOR.
    Cheng Wang, Yuting Yang, Chaoqi Zhou, Xianghuang Mei, Jing Liu, Kaihang Luo, Jia Zhou, Cheng Qin, Zhenguo Zeng.
      Acute lung injury (ALI) is characterized by acute systemic inflammatory responses that may lead to severe acute respiratory distress syndrome (ARDS). The clinical course of ALI/ARDS is variable; however, it has been reported that lipopolysaccharides (LPS) play a role in its development. The fragile chromosomal site gene WWOX is highly sensitive to genotoxic stress induced by environmental exposure and is an important candidate gene for exposure-related lung disease research. However, the expression of WWOX and its role in LPS-induced ALI still remain unidentified. This study investigated the expression of WWOX in mouse lung and epithelial cells and explored the role of WWOX in LPS-induced ALI model in vitro and in vivo. In addition, we explored one of the possible mechanisms by which WWOX alleviates ALI from the perspective of autophagy. Here, we observed that LPS stimulation reduced the expression of WWOX and the autophagy marker microtubule-associated protein 1 light chain 3β-II (MAP1LC3B/LC3B) in mouse lung epithelial and human epithelial (H292) cells. Overexpression of WWOX led to the activation of autophagy and inhibited inflammatory responses in LPS-induced ALI cells and mouse model. More importantly, we found that WWOX interacts with mechanistic target of rapamycin [serine/threonine kinase] (mTOR) and regulates mTOR and ULK-1 signaling-mediated autophagy. Thus, reduced WWOX levels were associated with LPS-induced ALI. WWOX can activate autophagy in lung epithelial cells and protect against LPS-induced ALI, which is partly related to the mTOR-ULK1 signaling pathway.
    Keywords:  Acute lung injury (ALI); Autophagy; Lipopolysaccharide (LPS); WWOX; mTOR
    DOI:  https://doi.org/10.1016/j.intimp.2022.109671
  48. Int Immunopharmacol. 2023 Jan 09. pii: S1567-5769(23)00015-2. [Epub ahead of print]115 109692
    Cartilage-specific deficiency of clock gene Bmal1 accelerated articular cartilage degeneration in osteoarthritis by up-regulation of mTORC1 signaling.
    Zhuang Qian, Xin Gao, Xinxin Jin, Xiaomin Kang, Shufang Wu.
      Although a growing body of studies recently demonstrated that circadian clock gene Bmal1 plays an important role in cartilage development and homeostasis, evidence regarding the contribution of Bmal1 in articular cartilage of OA progression is still unclear. In the present study, we investigated the direct role of Bmal1 in articular cartilage homeostasis during OA progression using tamoxifen-induced cartilage-specific knockout mice. We found that the expression of BMAL1 was decreased in OA-damaged and aging cartilage tissues. Cartilage-specific deletion of Bmal1 promoted cartilage degradation and chondrocyte apoptosis, and inhibited chondrocyte anabolism in OA mice, leading to acceleration of articular cartilage degeneration and osteophyte formation during OA progression. Mechanistic study indicated that loss of Bmal1 resulted in hyperactivation of mammalian target of rapamycin complex 1(mTORC1) signaling in OA cartilage, and pharmacological inhibition of mTORC1 signaling pathway by rapamycin alleviated partially Bmal1 ablation-induced cartilage degradation and chondrocyte apoptosis in ex vivo OA model. Therefore, our results provide the evidence of a vital role for Bmal1 in cartilage degradation in post-traumatic OA by partially regulating the mTORC1 signaling.
    Keywords:  Bmal1; Chondrocyte; Osteoarthritis; mTORC1
    DOI:  https://doi.org/10.1016/j.intimp.2023.109692
  49. Sci Rep. 2023 Jan 10. 13(1): 508
    Mechanism of cystogenesis by Cd79a-driven, conditional mTOR activation in developing mouse nephrons.
    Linh Tran Nguyen Truc, Satoshi Matsuda, Akiko Takenouchi, Quynh Tran Thuy Huong, Yui Kotani, Tatsuhiko Miyazaki, Hiroaki Kanda, Katsuhiko Yoshizawa, Hiroyasu Tsukaguchi.
      Polycystic kidney disease (PKD) is a common genetic disorder arising from developmental and postnatal processes. Defects in primary cilia and their signaling (eg, mTOR) underlie the pathogenesis. However, how mTOR regulates tubular integrity remains unclear. The paucity of faithful models has limited our understanding of pathogenesis and, therefore, the refinement of therapeutic targets. To understand the role of mTOR in early cystogenesis, we studied an in-house mouse model, Cd79a-Cre;Tsc1ff. (Cd79a-Tsc1 KO hereafter), recapitulating human autosomal-dominant PKD histology. Cre-mediated Tsc1 depletion driven by the promoter for Cd79a, a known B-cell receptor, activated mTORC1 exclusively along the distal nephron from embryonic day 16 onward. Cysts appeared in the distal nephron at 1 weeks of age and mice developed definite PKD by 4 weeks. Cd79a-Tsc1 KO tubule cells proliferated at a rate comparable to controls after birth but continued to divide even after postnatal day 14 when tubulogenesis is normally completed. Apoptosis occurred only after 9 weeks. During postnatal days 7-11, pre-cystic Cd79a-Tsc1 KO tubule cells showed cilia elongation, aberrant cell intercalation, and mitotic division, suggesting that defective cell planar polarity (PCP) may underlie cystogenesis. mTORC1 was activated in a portion of cyst-lining cells and occasionally even when Tsc1 was not depleted, implying a non-autonomous mechanism. Our results indicate that mTORC1 overactivation in developing distal tubules impairs their postnatal narrowing by disrupting morphogenesis, which orients an actively proliferating cell toward the elongating axis. The interplay between mTOR and cilium signaling, which coordinate cell proliferation with PCP, may be essential for cystogenesis.
    DOI:  https://doi.org/10.1038/s41598-023-27766-2
  50. PLoS Genet. 2023 Jan 06. 19(1): e1010573
    A role for BCL2L13 and autophagy in germline purifying selection of mtDNA.
    Laura S Kremer, Lyuba V Bozhilova, Diana Rubalcava-Gracia, Roberta Filograna, Mamta Upadhyay, Camilla Koolmeister, Patrick F Chinnery, Nils-Göran Larsson.
      Mammalian mitochondrial DNA (mtDNA) is inherited uniparentally through the female germline without undergoing recombination. This poses a major problem as deleterious mtDNA mutations must be eliminated to avoid a mutational meltdown over generations. At least two mechanisms that can decrease the mutation load during maternal transmission are operational: a stochastic bottleneck for mtDNA transmission from mother to child, and a directed purifying selection against transmission of deleterious mtDNA mutations. However, the molecular mechanisms controlling these processes remain unknown. In this study, we systematically tested whether decreased autophagy contributes to purifying selection by crossing the C5024T mouse model harbouring a single pathogenic heteroplasmic mutation in the tRNAAla gene of the mtDNA with different autophagy-deficient mouse models, including knockouts of Parkin, Bcl2l13, Ulk1, and Ulk2. Our study reveals a statistically robust effect of knockout of Bcl2l13 on the selection process, and weaker evidence for the effect of Ulk1 and potentially Ulk2, while no statistically significant impact is seen for knockout of Parkin. This points at distinctive roles of these players in germline purifying selection. Overall, our approach provides a framework for investigating the roles of other important factors involved in the enigmatic process of purifying selection and guides further investigations for the role of BCL2L13 in the elimination of non-synonymous mutations in protein-coding genes.
    DOI:  https://doi.org/10.1371/journal.pgen.1010573
  51. Curr Opin Neurobiol. 2023 Jan 06. pii: S0959-4388(22)00167-2. [Epub ahead of print]78 102673
    A tale of two pathways: Regulation of proteostasis by UPRmt and MDPs.
    Angela Johns, Ryo Higuchi-Sanabria, Max A Thorwald, David Vilchez.
      Mitochondrial fitness is critical to organismal health and its impairment is associated with aging and age-related diseases. As such, numerous quality control mechanisms exist to preserve mitochondrial stability, including the unfolded protein response of the mitochondria (UPRmt). The UPRmt is a conserved mechanism that drives the transcriptional activation of mitochondrial chaperones, proteases, autophagy (mitophagy), and metabolism to promote restoration of mitochondrial function under stress conditions. UPRmt has direct ramifications in aging, and its activation is often ascribed to improve health whereas its dysfunction tends to correlate with disease. This review pairs a description of the most recent findings within the field of UPRmt with a more poorly understood field: mitochondria-derived peptides (MDPs). Similar to UPRmt, MDPs are microproteins derived from the mitochondria that can impact organismal health and longevity. We then highlight a tantalizing interconnection between UPRmt and MDPs wherein both mechanisms may be efficiently coordinated to maintain organismal health.
    DOI:  https://doi.org/10.1016/j.conb.2022.102673
  52. Int J Mol Sci. 2022 Dec 21. pii: 148. [Epub ahead of print]24(1):
    The Greatwall-Endosulfine Switch Accelerates Autophagic Flux during the Cell Divisions Leading to G1 Arrest and Entry into Quiescence in Fission Yeast.
    Alicia Vázquez-Bolado, Rafael López-San Segundo, Natalia García-Blanco, Ana Elisa Rozalén, Daniel González-Álvarez, M Belén Suárez, Livia Pérez-Hidalgo, Sergio Moreno.
      Entry into quiescence in the fission yeast Schizosaccharomyces pombe is induced by nitrogen starvation. In the absence of nitrogen, proliferating fission yeast cells divide twice without cell growth and undergo cell cycle arrest in G1 before becoming G0 quiescent cells. Under these conditions, autophagy is induced to produce enough nitrogen for the two successive cell divisions that take place before the G1 arrest. In parallel to the induction of autophagy, the Greatwall-Endosulfine switch is activated upon nitrogen starvation to down-regulate protein phosphatase PP2A/B55 activity, which is essential for cell cycle arrest in G1 and implementation of the quiescent program. Here we show that, although inactivation of PP2A/B55 by the Greatwall-Endosulfine switch is not required to promote autophagy initiation, it increases autophagic flux at least in part by upregulating the expression of a number of autophagy-related genes.
    Keywords:  Endosulfine; Greatwall; PP2A; autophagy; fission yeast; quiescence
    DOI:  https://doi.org/10.3390/ijms24010148
  53. Autophagy. 2023 Jan 11.
    PIK3C3/VPS34 keeps body fats healthy.
    Wenqiang Song, J Luke Postoak, Lan Wu, Luc Van Kaer.
      Adipose tissue, or body fat, plays a critical role in the maintenance of health and the development of metabolic diseases. The pathological expansion of adipose tissue during obesity and the pathological reduction of adipose tissue during lipodystrophy can lead to a similar array of metabolic diseases that include diabetes, but mechanisms remain to be fully defined. In our recent studies, we explored the contribution of the lipid kinase PIK3C3/VPS34 to adipose tissue health and metabolic disease. We found that adipocyte-specific PIK3C3/VPS34 deficiency causes defects in the differentiation, survival and functional properties of adipocytes, resulting in reduced adipose tissue mass, altered blood lipid levels, fatty liver disease, diabetes, and defective body temperature control. These abnormalities mirror those observed in patients with lipodystrophy. These findings identify adipocyte PIK3C3/VPS34 as a potential target for therapeutic intervention in metabolic diseases.
    Keywords:  Adipocytes; PIK3C3/VPS34; autophagy; brown adipose tissue; diabetes; insulin resistance; lipodystrophy; metabolic disease; thermoregulation; white adipose tissue
    DOI:  https://doi.org/10.1080/15548627.2023.2166275
  54. Autophagy. 2023 Jan 10.
    Where is the field of autophagy research heading?
    Hagai Abeliovich, Jayanta Debnath, Wen-Xing Ding, William T Jackson, Do-Hyung Kim, Daniel J Klionsky, Nicholas Ktistakis, Marta Margeta, Christian Münz, Morten Petersen, Junichi Sadoshima, Isabelle Vergne.
      In this editors' corner, the section editors were asked to indicate where they see the autophagy field heading and to suggest what they consider to be key unanswered questions in their specialty area.
    Keywords:  Ideas for your grant proposal; suggestions; the big picture; the grand scheme; thoughts; yada yada yada
    DOI:  https://doi.org/10.1080/15548627.2023.2166301
  55. Int J Mol Sci. 2022 Dec 27. pii: 449. [Epub ahead of print]24(1):
    Lipid Peroxidation and Iron Metabolism: Two Corner Stones in the Homeostasis Control of Ferroptosis.
    Luc Rochette, Geoffrey Dogon, Eve Rigal, Marianne Zeller, Yves Cottin, Catherine Vergely.
      Regulated cell death (RCD) has a significant impact on development, tissue homeostasis, and the occurrence of various diseases. Among different forms of RCD, ferroptosis is considered as a type of reactive oxygen species (ROS)-dependent regulated necrosis. ROS can react with polyunsaturated fatty acids (PUFAs) of the lipid (L) membrane via the formation of a lipid radical L• and induce lipid peroxidation to form L-ROS. Ferroptosis is triggered by an imbalance between lipid hydroperoxide (LOOH) detoxification and iron-dependent L-ROS accumulation. Intracellular iron accumulation and lipid peroxidation are two central biochemical events leading to ferroptosis. Organelles, including mitochondria and lysosomes are involved in the regulation of iron metabolism and redox imbalance in ferroptosis. In this review, we will provide an overview of lipid peroxidation, as well as key components involved in the ferroptotic cascade. The main mechanism that reduces ROS is the redox ability of glutathione (GSH). GSH, a tripeptide that includes glutamic acid, cysteine, and glycine, acts as an antioxidant and is the substrate of glutathione peroxidase 4 (GPX4), which is then converted into oxidized glutathione (GSSG). Increasing the expression of GSH can inhibit ferroptosis. We highlight the role of the xc- GSH-GPX4 pathway as the main pathway to regulate ferroptosis. The system xc-, composed of subunit solute carrier family members (SLC7A11 and SLC3A2), mediates the exchange of cystine and glutamate across the plasma membrane to synthesize GSH. Accumulating evidence indicates that ferroptosis requires the autophagy machinery for its execution. Ferritinophagy is used to describe the removal of the major iron storage protein ferritin by the autophagy machinery. Nuclear receptor coactivator 4 (NCOA4) is a cytosolic autophagy receptor used to bind ferritin for subsequent degradation by ferritinophagy. During ferritinophagy, stored iron released becomes available for biosynthetic pathways. The dysfunctional ferroptotic response is implicated in a variety of pathological conditions. Ferroptosis inducers or inhibitors targeting redox- or iron metabolism-related proteins and signal transduction have been developed. The simultaneous detection of intracellular and extracellular markers may help diagnose and treat diseases related to ferroptotic damage.
    Keywords:  autophagy; ferritin; ferritinophagy; ferroptosis; iron; lipid peroxidation
    DOI:  https://doi.org/10.3390/ijms24010449
  56. Oncogene. 2023 Jan 07.
    Chaperone-mediated autophagy promotes PCa survival during ARPI through selective proteome remodeling.
    Nicholas Nikesitch, Eliana Beraldi, Fan Zhang, Hans Adomat, Robert Bell, Kotaro Suzuki, Ladan Fazli, Sonia Hy Kung, Christopher Wells, Nicholas Pinette, Neetu Saxena, Yuzhuo Wang, Martin Gleave.
      The androgen receptor (AR) plays an important role in PCa metabolism, with androgen receptor pathway inhibition (ARPI) subjecting PCa cells to acute metabolic stress caused by reduced biosynthesis and energy production. Defining acute stress response mechanisms that alleviate ARPI stress and therefore mediate prostate cancer (PCa) treatment resistance will help improve therapeutic outcomes of patients treated with ARPI. We identified the up-regulation of chaperone-mediated autophagy (CMA) in response to acute ARPI stress, which persisted in castration-resistant PCa (CRPC); previously undefined in PCa. CMA is a selective protein degradation pathway and a key stress response mechanism up-regulated under several stress stimuli, including metabolic stress. Through selective protein degradation, CMA orchestrates the cellular stress response by regulating cellular pathways through selective proteome remodeling. Through broad-spectrum proteomic analysis, CMA coordinates metabolic reprogramming of PCa cells to sustain PCa growth and survival during ARPI; through the upregulation of mTORC1 signaling and pathways associated with PCa biosynthesis and energetics. This not only promoted PCa growth during ARPI, but also promoted the emergence of CRPC in-vivo. During CMA inhibition, PCa metabolism is compromised, leading to ATP depletion, resulting in a profound anti-proliferative effect on PCa cells, and is enhanced when combined with ARPI. Furthermore, CMA inhibition prevented in-vivo tumour formation, and also re-sensitized enzalutamide-resistant cell lines in-vitro. The profound anti-proliferative effect of CMA inhibition was attributed to cell cycle arrest mediated through p53 transcriptional repression of E2F target genes. In summary, CMA is an acute ARPI stress response mechanism, essential in alleviating ARPI induced metabolic stress, essential for ensuring PCa growth and survival. CMA plays a critical role in the development of ARPI resistance in PCa.
    DOI:  https://doi.org/10.1038/s41388-022-02573-7
  57. Sci Rep. 2023 Jan 06. 13(1): 293
    Ubiquilin-2 regulates pathological alpha-synuclein.
    Stephanie S Sandoval-Pistorius, Julia E Gerson, Nyjerus Liggans, Jaimie H Ryou, Kulin Oak, Xingli Li, Keyshla Y Negron-Rios, Svetlana Fischer, Henry Barsh, Emily V Crowley, Mary E Skinner, Lisa M Sharkey, Sami J Barmada, Henry L Paulson.
      The key protein implicated in Parkinson's disease and other synucleinopathies is α-synuclein, and a post-translationally modified form of the protein, phosphorylated at serine 129 (pS129), is a principal component in Lewy bodies, a pathological hallmark of PD. While altered proteostasis has been implicated in the etiology of Parkinson's disease, we still have a limited understanding of how α-synuclein is regulated in the nervous system. The protein quality control protein Ubiquilin-2 (UBQLN2) is known to accumulate in synucleinopathies, but whether it directly regulates α-synuclein is unknown. Using cellular and mouse models, we find that UBQLN2 decreases levels of α-synuclein, including the pS129 phosphorylated isoform. Pharmacological inhibition of the proteasome revealed that, while α-synuclein may be cleared by parallel and redundant quality control pathways, UBQLN2 preferentially targets pS129 for proteasomal degradation. Moreover, in brain tissue from human PD and transgenic mice expressing pathogenic α-synuclein (A53T), native UBQLN2 becomes more insoluble. Collectively, our studies support a role for UBQLN2 in directly regulating pathological forms of α-synuclein and indicate that UBQLN2 dysregulation in disease may contribute to α-synuclein-mediated toxicity.
    DOI:  https://doi.org/10.1038/s41598-022-26899-0
  58. Mol Neurobiol. 2023 Jan 13.
    Synonymous Codon Variant Analysis for Autophagic Genes Dysregulated in Neurodegeneration.
    Rekha Khandia, Megha Katare Pandey, Igor Vladimirovich Rzhepakovsky, Azmat Ali Khan, Athanasios Alexiou.
      Neurodegenerative disorders are often a culmination of the accumulation of abnormally folded proteins and defective organelles. Autophagy is a process of removing these defective proteins, organelles, and harmful substances from the body, and it works to maintain homeostasis. If autophagic removal of defective proteins has interfered, it affects neuronal health. Some of the autophagic genes are specifically found to be associated with neurodegenerative phenotypes. Non-functional, mutated, or gene copies having silent mutations, often termed synonymous variants, might explain this. However, these synonymous variant which codes for exactly similar proteins have different translation rates, stability, and gene expression profiling. Hence, it would be interesting to study the pattern of synonymous variant usage. In the study, synonymous variant usage in various transcripts of autophagic genes ATG5, ATG7, ATG8A, ATG16, and ATG17/FIP200 reported to cause neurodegeneration (if dysregulated) is studied. These genes were analyzed for their synonymous variant usage; nucleotide composition; any possible nucleotide skew in a gene; physical properties of autophagic protein including GRAVY and AROMA; hydropathicity; instability index; and frequency of acidic, basic, neutral amino acids; and gene expression level. The study will help understand various evolutionary forces acting on these genes and the possible augmentation of a gene if showing unusual behavior.
    Keywords:  ATG16L1; Atg5; Atg7; Autophagy genes; Codon usage ; Neurodegeneration; Neuronal survival FIP200 (human counterpart Atg17 gene)
    DOI:  https://doi.org/10.1007/s12035-022-03081-1
  59. Int J Mol Sci. 2023 Jan 01. pii: 755. [Epub ahead of print]24(1):
    Metformin and Its Immune-Mediated Effects in Various Diseases.
    Ichiro Nojima, Jun Wada.
      Metformin has been a long-standing prescribed drug for treatment of type 2 diabetes (T2D) and its beneficial effects on virus infection, autoimmune diseases, aging and cancers are also recognized. Metformin modulates the differentiation and activation of various immune-mediated cells such as CD4+ and CD+8 T cells. The activation of adenosine 5'-monophosphate-activated protein kinase (AMPK) and mammalian target of rapamycin complex 1 (mTORC1) pathway may be involved in this process. Recent studies using Extracellular Flux Analyzer demonstrated that metformin alters the activities of glycolysis, oxidative phosphorylation (OXPHOS), lipid oxidation, and glutaminolysis, which tightly link to the modulation of cytokine production in CD4+ and CD+8 T cells in various disease states, such as virus infection, autoimmune diseases, aging and cancers.
    Keywords:  AMPK; CD8 T cells; OXPHOS; aging; autoimmune disease; cancer; mTORC
    DOI:  https://doi.org/10.3390/ijms24010755
  60. J Cell Sci. 2023 Jan 01. pii: jcs260563. [Epub ahead of print]136(1):
    Nucleophagy contributes to genome stability through degradation of type II topoisomerases A and B and nucleolar components.
    Gabriel Muciño-Hernández, Pilar Sarah Acevo-Rodríguez, Sandra Cabrera-Benitez, Adán Oswaldo Guerrero, Horacio Merchant-Larios, Susana Castro-Obregón.
      The nuclear architecture of mammalian cells can be altered as a consequence of anomalous accumulation of nuclear proteins or genomic alterations. Most of the knowledge about nuclear dynamics comes from studies on cancerous cells. How normal healthy cells maintain genome stability, avoiding accumulation of nuclear damaged material, is less understood. Here, we describe that primary mouse embryonic fibroblasts develop a basal level of nuclear buds and micronuclei, which increase after etoposide-induced DNA double-stranded breaks. Both basal and induced nuclear buds and micronuclei colocalize with the autophagic proteins BECN1 and LC3B (also known as MAP1LC3B) and with acidic vesicles, suggesting their clearance by nucleophagy. Some of the nuclear alterations also contain autophagic proteins and type II DNA topoisomerases (TOP2A and TOP2B), or the nucleolar protein fibrillarin, implying they are also targets of nucleophagy. We propose that basal nucleophagy contributes to genome and nuclear stability, as well as in response to DNA damage.
    Keywords:  Autophagy; DNA damage; Mammalian nucleophagy; Micronuclei; Nucleolus
    DOI:  https://doi.org/10.1242/jcs.260563
  61. Endocr Rev. 2023 Jan 12. pii: bnad001. [Epub ahead of print]
    Recent Advances in the Role of Autophagy in Endocrine-Dependent Tumors.
    Anvita Komarla, Suzanne Dufresne, Christina G Towers.
      Autophagy plays a complex role in several cancer types, including endocrine-dependent cancers, by fueling cellular metabolism and clearing damaged substrates. This conserved recycling process has a dual function across tumor types where it can be tumor suppressive at early stages but tumor promotional in established disease. This review highlights the controversial roles of autophagy in endocrine-dependent tumors regarding cancer initiation, tumorigenesis, metastasis, and treatment response. We summarize clinical trial results thus far and highlight the need for additional mechanistic, pre-clinical and clinical studies in endocrine-dependent tumors particularly in breast cancer and prostate cancer.
    Keywords:  Autophagy; Cancer; Chloroquine; Clinical trials; Endocrine-Dependent Tumors
    DOI:  https://doi.org/10.1210/endrev/bnad001
  62. Nat Rev Rheumatol. 2023 Jan 06.
    mTORC1 implicated in Still's disease and MAS.
    Sarah Onuora.
      
    DOI:  https://doi.org/10.1038/s41584-023-00908-6
  63. Int J Mol Sci. 2022 Dec 28. pii: 477. [Epub ahead of print]24(1):
    Mucopolysaccharidoses: Cellular Consequences of Glycosaminoglycans Accumulation and Potential Targets.
    Andrés Felipe Leal, Eliana Benincore-Flórez, Estera Rintz, Angélica María Herreño-Pachón, Betul Celik, Yasuhiko Ago, Carlos Javier Alméciga-Díaz, Shunji Tomatsu.
      Mucopolysaccharidoses (MPSs) constitute a heterogeneous group of lysosomal storage disorders characterized by the lysosomal accumulation of glycosaminoglycans (GAGs). Although lysosomal dysfunction is mainly affected, several cellular organelles such as mitochondria, endoplasmic reticulum, Golgi apparatus, and their related process are also impaired, leading to the activation of pathophysiological cascades. While supplying missing enzymes is the mainstream for the treatment of MPS, including enzyme replacement therapy (ERT), hematopoietic stem cell transplantation (HSCT), or gene therapy (GT), the use of modulators available to restore affected organelles for recovering cell homeostasis may be a simultaneous approach. This review summarizes the current knowledge about the cellular consequences of the lysosomal GAGs accumulation and discusses the use of potential modulators that can reestablish normal cell function beyond ERT-, HSCT-, or GT-based alternatives.
    Keywords:  endoplasmic reticulum; glycosaminoglycans; lysosome; mitochondria; mucopolysaccharidoses
    DOI:  https://doi.org/10.3390/ijms24010477
  64. J Med Virol. 2023 Jan 10.
    Inhibiting Cardiac Autophagy Suppresses Zika Virus Replication.
    Xin Ma, Yinnong Jia, Jing Yuan, Qiu-Ju Tang, Wen-Cong Gao, Guang-Feng Zhou, Ren-Hua Yang, Wei Pang, Chang-Bo Zheng.
      Zika Virus (ZIKV) infection is a global threat. Other than the congenital neurological disorders it causes, ZIKV infection has been reported to induce cardiac complications. However, the precise treatment plans are unclear. Thus, illustrating the pathogenic mechanism of ZIKV in the heart is critical to providing effective prevention and treatment of ZIKV infection. The mechanism of autophagy has been reported recently in Dengue virus infection. Whether or not autophagy participates in ZIKV infection and its role remains unrevealed. This study successfully established the in vitro cardiomyocytes and in vivo mouse models of ZIKV infection to investigate the involvement of autophagy in ZIKV infection. The results showed that ZIKV infection is both time and gradient-dependent. The key autophagy protein, LC3B, increased remarkably after ZIKV infection. Meanwhile, autophagic flux was detected by immunofluorescence. Applying autophagy inhibitors decreased the LC3B levels. Furthermore, the number of viral copies was quantified to evaluate the influence of autophagy during infection. We found that autophagy was actively involved in the ZIKV infection and the inhibition of autophagy could effectively reduce the viral copies, suggesting a potential intervention strategy for reducing ZIKV infection and the undesired complications caused by ZIKV. This article is protected by copyright. All rights reserved.
    Keywords:  LC3B; ZIKV infection; autophagic flux; cardiac complications; inhibition; viral copies
    DOI:  https://doi.org/10.1002/jmv.28483
  65. Mol Immunol. 2023 Jan 06. pii: S0161-5890(23)00002-0. [Epub ahead of print]154 69-79
    Cryptosporidium parvum maintains intracellular survival by activating the host cellular EGFR-PI3K/Akt signaling pathway.
    Heng Yang, Mengge Zhang, Xiaocen Wang, Pengtao Gong, Nan Zhang, Xichen Zhang, Xin Li, Jianhua Li.
      Autophagy is a critical cellular mechanism in helping infected cells remove intracellular pathogens and is countered by pathogens maintaining intracellular survival by regulating autophagy through the manipulation of the host cellular signal transduction pathway. Cryptosporidium parvum is a zoonotic intracellular but extracytoplasmic protozoon that causes diarrhea in infants and young children worldwide. However, it is still unclear how Cryptosporidium adapts to intracellular survival. In the present study, we demonstrated that C. parvum could activate the EGFR-PI3K/Akt signaling pathway to promote intracellular survival in HCT-8 cells. The western blot results showed that C. parvum induced EGFR and Akt phosphorylation in HCT-8 cells. The EGFR inhibitor AG1478 decreased EGFR and Akt phosphorylation, and the PI3K inhibitor LY294002 impaired Akt phosphorylation induced by C. parvum in HCT-8 cells. Inhibition of EGFR or Akt decreased the number of intracellular parasites. Second, low-dose infection of C. parvum triggered complete autophagy and enhanced autophagic flux in HCT-8 cells. The expressions of mTOR and p62 were decreased, and the expressions of LC3 and Beclin1 were increased in C. parvum-infected HCT-8 cells. Transfection with siBeclin1 or siATG7 reduced LC3 accumulation, while lysosome inhibitor E64d+pepA increased LC3 accumulation induced by C. parvum in HCT-8 cells. Intracellular parasite proliferation was decreased when treated with autophagy inducer rapamycin, whereas autophagy inhibitor 3-MA, E64d+pep A, siBeclin1 or siATG7 increased intracellular parasites. Third, C. parvum inhibited autophagy killing to promote its own intracellular survival by activating EGFR-Akt signaling pathway. The EGFR inhibitor AG1478 enhanced autophagic flux, and Akt inhibitor IV increased LC3 accumulation and inhibited C. parvum proliferation in HCT-8 cells. Akt inhibitor IV-inhibited C. parvum proliferation was attenuated by E64d+pepA. In summary, C. parvum could maintain intracellular survival by inhibiting autophagy via EGFR-PI3K/Akt pathway. These results revealed a new mechanism for the interaction of C. parvum with host cells.
    Keywords:  Autophagy; Cryptosporidium parvum; EGFR-PI3K/Akt signaling pathway; Intracellular survival
    DOI:  https://doi.org/10.1016/j.molimm.2023.01.002
  66. Cell Rep. 2023 Jan 10. pii: S2211-1247(22)01888-5. [Epub ahead of print]42(1): 111984
    SLC7A14 imports GABA to lysosomes and impairs hepatic insulin sensitivity via inhibiting mTORC2.
    Xiaoxue Jiang, Kan Liu, Haizhou Jiang, Hanrui Yin, En-Duo Wang, Hong Cheng, Feixiang Yuan, Fei Xiao, Fenfen Wang, Wei Lu, Bo Peng, Yousheng Shu, Xiaoying Li, Shanghai Chen, Feifan Guo.
      Lysosomal amino acid accumulation is implicated in several diseases, but its role in insulin resistance, the central mechanism to type 2 diabetes and many metabolic diseases, is unclear. In this study, we show the hepatic expression of lysosomal membrane protein solute carrier family 7 member 14 (SLC7A14) is increased in insulin-resistant mice. The promoting effect of SLC7A14 on insulin resistance is demonstrated by loss- and gain-of-function experiments. SLC7A14 is further demonstrated as a transporter resulting in the accumulation of lysosomal γ-aminobutyric acid (GABA), which induces insulin resistance via inhibiting mTOR complex 2 (mTORC2)'s activity. These results establish a causal link between lysosomal amino acids and insulin resistance and suggest that SLC7A14 inhibition may provide a therapeutic strategy in treating insulin resistance-related and GABA-related diseases and may provide insights into the upstream mechanisms for mTORC2, the master regulator in many important processes.
    Keywords:  CP: Metabolism; GABA; SLC7A14; amino acid; insulin resistance; liver; lysosome
    DOI:  https://doi.org/10.1016/j.celrep.2022.111984
  67. Exp Lung Res. 2023 Jan 13. 1-10
    Cigarette smoke extract-induced inflammatory response via inhibition of the TFEB-mediated autophagy in NR8383 cells.
    Shu-Wen Xu, Yu-Jie Zhang, Wen-Mei Liu, Xin-Fang Zhang, Yuan Wang, Shui-Ying Xiang, Jing-Chao Su, Zi-Bing Liu.
      Objective: Chronic pulmonary inflammation caused by long-term smoking is the core pathology of COPD. Alveolar macrophages (AMs) are involved in the pulmonary inflammation of COPD. The accumulation of damaged materials caused by impaired autophagy triggers inflammatory response in macrophages. As a key transcription regulator, transcription factor EB (TFEB) activates the transcription of target genes related autophagy and lysosome by binding to promoters, whereas it is unclarified for the relationship between inflammatory response induced by cigarette smoke extract (CSE) and TFEB-mediated autophagy. Thus, we investigated the role of TFEB-mediated autophagy in inflammatory response induced by CSE in NR8383 cells, and to explore its potential mechanism. Methods: Based on cell viability and autophagy, cells treated with 20% concentration of CSE for 24 h were selected for further studies. Cells were divided into control group, chloroquine (CQ, the autophagy inhibitor) group, CSE group, CSE + rapamycin (the autophagy inducer) group and CSE + fisetin (the TFEB inducer) group. The levels of tumor necrosis factor α (TNF-α), interleukin 1β (IL-1β), and IL-6 in supernatant were detected by ELISA kits. The protein expressions were tested by western blot. The intensity of fluorescence of Lysosome-associated membrane protein 1 (LAMP1) and TFEB was detected by immunofluorescence. Lyso-Tracker Red staining was applied to detect the lysosome environment. Results: CSE inhibited the cell viability, increased the contents of TNF-α, IL-1β, IL-6, the ratio of LC3II/I, and the level of P62 protein. Besides, CSE decreased the fluorescence intensity of LAMP1 protein and Lyso-Tracker Red staining, as well as the ratio of nucleus/cytosol of TFEB protein. Activating autophagy with rapamycin alleviated CSE-induced inflammatory response. The activation of TFEB via fisetin alleviated CSE-induced autophagy impairment and lysosomal dysfunction, thus alleviated inflammatory response in NR8383 cells. Conclusion: CSE-induced inflammatory response in NR8383 cells, which may be related to the inhibition of TFEB-mediated autophagy.
    Keywords:  Alveolar macrophage; autophagy; cigarette smoke extract; inflammation; transcription factor EB
    DOI:  https://doi.org/10.1080/01902148.2022.2164674
  68. Cell Rep. 2022 Dec 30. pii: S2211-1247(22)01805-8. [Epub ahead of print]42(1): 111906
    The CNC-family transcription factor Nrf3 coordinates the melanogenesis cascade through macropinocytosis and autophagy regulation.
    Tsuyoshi Waku, Sota Nakada, Haruka Masuda, Haruna Sumi, Ayaka Wada, Shuuhei Hirose, Iori Aketa, Akira Kobayashi.
      Melanin is a pigment produced from the amino acid L-tyrosine in melanosomes. The CNC-family transcription factor Nrf3 is expressed in the basal layer of the epidermis, where melanocytes reside, but its melanogenic function is unclear. Here, we show that Nrf3 regulates macropinocytosis and autophagy to coordinate melanogenesis cascade. In response to an exogenous inducer of melanin production, forskolin, Nrf3 upregulates the core melanogenic gene circuit, which includes Mitf, Tyr, Tyrp1, Pmel, and Oca2. Furthermore, Nrf3 induces the gene expression of Cln3, an autophagosome-related factor, for melanin precursor uptake by macropinocytosis. Ulk2 and Gabarapl2 are also identified as Nrf3-target autophagosome-related genes for melanosome formation. In parallel, Nrf3 prompts autolysosomal melanosome degradation for melanocyte survival. An endogenous melanogenic inducer αMSH also activates Nrf3-mediated melanin production, whereas it is suppressed by an HIV-1 protease inhibitor, nelfinavir. These findings indicate the significant role of Nrf3 in the melanogenesis and the anti-melanogenic potential of nelfinavir.
    Keywords:  CP: Cell biology; CP: Molecular biology; Nrf3; autophagy; macropinocytosis; melanogenesis; nelfinavir
    DOI:  https://doi.org/10.1016/j.celrep.2022.111906
  69. Int Immunopharmacol. 2023 Jan 11. pii: S1567-5769(23)00019-X. [Epub ahead of print]115 109696
    RIP3 impedes Mycobacterium tuberculosis survival and promotes p62-mediated autophagy.
    Jiamei Zhang, Lu Han, Qinmei Ma, Xiaoping Wang, Jialin Yu, Yanan Xu, Xu Zhang, Xiaoling Wu, Guangcun Deng.
      Macrophage is believed to play a vital role in the fight against Mycobacterium tuberculosis (M.tb) infection by activating autophagy. Recently, receptor-interacting protein kinase-3 (RIP3), an essential kinase for necroptotic cell death signaling, has been demonstrated to be involved in autophagy. However, RIP3's role in fighting against M.tb infection remains elusive. Here we show that a substantial increase in inflammatory cell infiltration and higher bacterial burden are observed in the lungs of RIP3-/- mice with Mycobacterium bovis Bacillus Calmette-Guerin (BCG) infection. Meanwhile, RIP3 ameliorates lung injury and promote autophagy via induce autophagosome and autophagolysosome formation which indicate that RIP3 is indispensable for host clearance of BCG via autophagy. Mechanically, RIP3 enhances p62 binding to ubiquitylated proteins and LC3 by interacting with p62, and RHIM domain is required for RIP3-p62 interaction. Hence, our results conclusively show that RIP3 impedes M.tb survival and promotes p62-mediated autophagy. The findings provide further insight into understanding the mechanism of M.tb immune escape and pathogenesis of tuberculosis.
    Keywords:  Autophagy; BCG; Macrophage; RIP3; p62
    DOI:  https://doi.org/10.1016/j.intimp.2023.109696
  70. Gerontology. 2023 Jan 07.
    Why is rapamycin not a rapalog?
    Ajla Hodzic Kuerec, Andrea B Maier.
      Rapamycin (sirolimus) is an immunosuppressive drug approved by the Food and Drug Administration (FDA). It is also a leading candidate for targeting aging. Rapamycin and its analogs (everolimus, temsirolimus, ridaforolimus) inhibit the mammalian target of rapamycin (mTOR) kinase by binding to FK506-binding proteins (FKBP) and have a similar chemical structure that only differs in the functional group present at carbon-40. Analogs of rapamycin were developed to improve its pharmacological properties, such as low oral bioavailability and a long half-life. The analogs of rapamycin are referred to as 'rapalogs.' Rapamycin is the parent compound and should there with not be called a 'rapalog.'
    DOI:  https://doi.org/10.1159/000528985
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