bims-auttor Biomed News
on Autophagy and mTOR
Issue of 2020‒03‒15
33 papers selected by
Viktor Korolchuk, Newcastle University

  1. Cell Chem Biol. 2020 Mar 03. pii: S2451-9456(20)30069-6. [Epub ahead of print]
      Macroautophagy (hereafter referred to as autophagy) is an evolutionarily conserved cellular process capable of degrading various biological molecules (e.g., protein, glycogen, lipids, DNA, and RNA) and organelles (e.g., mitochondria, endoplasmic reticulum [ER] ribosomes, lysosomes, and micronuclei) via the lysosomal pathway. Ferroptosis is a type of oxidative stress-dependent regulated cell death associated with iron accumulation and lipid peroxidation. The recently discovered role of autophagy, especially selective types of autophagy (e.g., ferritinophagy, lipophagy, clockophagy, and chaperone-mediated autophagy), in driving cells toward ferroptotic death motivated us to explore the functional interactions between metabolism, immunity, and cell death. Here, we describe types of selective autophagy and discuss the regulatory mechanisms and signaling pathways of autophagy-dependent ferroptosis. We also summarize chemical modulators that are currently available for triggering or blocking autophagy-dependent ferroptosis and that may be developed for therapeutic interventions in human diseases.
    Keywords:  autophagy; ferroptosis; network; regulated cell death; selective autophagy
  2. Cell. 2020 Mar 03. pii: S0092-8674(20)30161-6. [Epub ahead of print]
      Selective autophagy of organelles is critical for cellular differentiation, homeostasis, and organismal health. Autophagy of the ER (ER-phagy) is implicated in human neuropathy but is poorly understood beyond a few autophagosomal receptors and remodelers. By using an ER-phagy reporter and genome-wide CRISPRi screening, we identified 200 high-confidence human ER-phagy factors. Two pathways were unexpectedly required for ER-phagy. First, reduced mitochondrial metabolism represses ER-phagy, which is opposite of general autophagy and is independent of AMPK. Second, ER-localized UFMylation is required for ER-phagy to repress the unfolded protein response via IRE1α. The UFL1 ligase is brought to the ER surface by DDRGK1 to UFMylate RPN1 and RPL26 and preferentially targets ER sheets for degradation, analogous to PINK1-Parkin regulation during mitophagy. Our data provide insight into the cellular logic of ER-phagy, reveal parallels between organelle autophagies, and provide an entry point to the relatively unexplored process of degrading the ER network.
    Keywords:  CRISPR; ER-phagy; UFMylation; autophagy; endoplasmic reticulum; genome-wide screen; organelle turnover; oxidative phosphorylation; post-translational modification
  3. Nat Commun. 2020 Mar 13. 11(1): 1386
      Microglia maintain brain homeostasis by removing neuron-derived components such as myelin and cell debris. The evidence linking microglia to neurodegenerative diseases is growing; however, the precise mechanisms remain poorly understood. Herein, we report a neuroprotective role for microglia in the clearance of neuron-released α-synuclein. Neuronal α-synuclein activates microglia, which in turn engulf α-synuclein into autophagosomes for degradation via selective autophagy (termed synucleinphagy). Synucleinphagy requires the presence of microglial Toll-like receptor 4 (TLR4), which induces transcriptional upregulation of p62/SQSTM1 through the NF-κB signaling pathway. Induction of p62, an autophagy receptor, is necessary for the formation of α-synuclein/ubiquitin-positive puncta that are degraded by autophagy. Finally, disruption of microglial autophagy in mice expressing human α-synuclein promotes the accumulation of misfolded α-synuclein and causes midbrain dopaminergic neuron degeneration. Our study thus identifies a neuroprotective function of microglia in the clearance of α-synuclein via TLR4-NF-κB-p62 mediated synucleinphagy.
  4. Autophagy. 2020 Mar 13. 1-2
      The role protein aggregates play in the pathogenesis of neurodegenerative diseases has been a question since their initial observation. In this autophagic punctum, we discuss our recent findings of how the selectivity scaffold/adaptor WDFY3/Alfy is required for the turnover of aggregated mutant HTT (huntingtin; mHTT) in the adult brain, and how it confers resistance to Huntington disease (HD)-like symptoms. Depletion of WDFY3 in a mouse model of HD accelerates mHTT accumulation, and this is accompanied by an accelerated onset of motoric and neuropathological phenotypes, indicating that WDFY3 levels and the rate of aggregate accumulation can modify disease pathogenesis. Given that the accelerated accumulation is also recapitulated in medium spiny neurons created via direct conversion from human HD fibroblasts, we propose that WDFY3 is a genetic modifier of HD and suggest that it may also influence aging and the pathogenesis of other neurological disorders.
    Keywords:  Neurodegeneration; Wdfy3/Alfy; neurodegenerative disease; protein aggregation; selective autophagy
  5. Autophagy. 2020 Mar 09.
      In budding yeast, macroautophagy/autophagy is required for cells to enter into the meiotic divisions. Our recent publication showed that autophagy is also required for meiotic exit. Inhibition of autophagy as cells enter into the meiotic divisions results in additional rounds of spindle formation, spindle elongation, and aberrant chromosome segregation leading to cell death. Under these conditions, the meiosis II-specific cyclin Clb3 is absent, and two substrates of the anaphase-promoting complex/cyclosome (APC/C) persist into the additional divisions instead of being degraded after meiosis II. We found that the translational repressor Rim4 is a substrate of autophagy, which could explain these observations through its known role in repressing synthesis of Clb3 and the meiosis-specific co-activator of the APC/C, Ama1. Combined, these results provide new mechanistic insight into the control of meiotic exit through timed autophagic degradation of a master regulator of gene expression.
    Keywords:  APC/C; Atg1; Clb3; Rim4; autophagy; cyclins; meiosis
  6. Autophagy. 2020 Mar 13.
      AKT/PKB is downregulated by the ubiquitin-proteasome system (UPS), which plays a key role in cell survival and tumor progression in various types of cancer. The objective of this study was to determine the relationship between the sequential ubiquitination of lysine residues K284 to K214 in AKT and R-HSPA5 (the arginylated form of HSPA5), which contribute to the autophagic/lysosomal degradation of AKT when impaired proteasomal activity induces cellular stress. Results show that proteasome inhibitors (PIs) increased ATE1 (arginyltransferase 1)-mediated R-HSPA5 levels in a reactive oxygen species (ROS)-dependent manner. Further, binding of fully ubiquitinated AKT with R-HSPA5 induced AKT degradation via the autophagy-lysosome pathway. Specifically, the K48 (Lys48)-linked ubiquitinated form of AKT was selectively degraded in the lysosome with R-HSPA5. The deubiquitinase, USP7 (ubiquitin specific peptidase 7), prevented AKT degradation by inhibiting AKT ubiquitination via interaction with AKT. MUL1 (mitochondrial ubiquitin ligase activator of NFKB 1) also played a vital role in the lysosomal degradation of AKT by sequentially ubiquitinating AKT residues K284 to K214 for R-HSPA5-mediated autophagy. Consistent with this finding, despite HSPA5 arginylation, AKT was not degraded in mul1 KO cells. These results suggest that MUL1-mediated sequential ubiquitination of K284 to K214 may serve as a novel mechanism by which AKT is designated for lysosomal degradation. Moreover, binding of R-HSPA5 with fully ubiquitinated AKT is required for the autophagic/lysosomal degradation of AKT. Thus, modulating the MUL1-mediated non-proteasomal proteolysis mechanisms, such as sequential ubiquitination, may prove to be a novel therapeutic approach for cancer treatment.
    Keywords:  AKT; HSPA5; MUL1; autophagy; lysosome; sequential ubiquitination
  7. EMBO J. 2020 Mar 09. e102731
      Mitochondria house anabolic and catabolic processes that must be balanced and adjusted to meet cellular demands. The RNA-binding protein CLUH (clustered mitochondria homolog) binds mRNAs of nuclear-encoded mitochondrial proteins and is highly expressed in the liver, where it regulates metabolic plasticity. Here, we show that in primary hepatocytes, CLUH coalesces in specific ribonucleoprotein particles that define the translational fate of target mRNAs, such as Pcx, Hadha, and Hmgcs2, to match nutrient availability. Moreover, CLUH granules play signaling roles, by recruiting mTOR kinase and the RNA-binding proteins G3BP1 and G3BP2. Upon starvation, CLUH regulates translation of Hmgcs2, involved in ketogenesis, inhibits mTORC1 activation and mitochondrial anabolic pathways, and promotes mitochondrial turnover, thus allowing efficient reprograming of metabolic function. In the absence of CLUH, a mitophagy block causes mitochondrial clustering that is rescued by rapamycin treatment or depletion of G3BP1 and G3BP2. Our data demonstrate that metabolic adaptation of liver mitochondria to nutrient availability depends on a compartmentalized CLUH-dependent post-transcriptional mechanism that controls both mTORC1 and G3BP signaling and ensures survival.
    Keywords:  CLUH; G3BP; RNA metabolism; mTORC1; mitochondria
  8. Cells. 2020 Mar 11. pii: E687. [Epub ahead of print]9(3):
      AMP-activated protein kinase (AMPK) is activated by vascular endothelial growth factor (VEGF) in endothelial cells and it is significantly involved in VEGF-induced angiogenesis. This study investigates whether the VEGF/AMPK pathway regulates autophagy in endothelial cells and whether this is linked to its pro-angiogenic role. We show that VEGF leads to AMPKα1-dependent phosphorylation of Unc-51-like kinase 1 (ULK1) at its serine residue 556 and to the subsequent phosphorylation of the ULK1 substrate ATG14. This triggers initiation of autophagy as shown by phosphorylation of ATG16L1 and conjugation of the microtubule-associated protein light chain 3B, which indicates autophagosome formation; this is followed by increased autophagic flux measured in the presence of bafilomycin A1 and by reduced expression of the autophagy substrate p62. VEGF-induced autophagy is transient and probably terminated by mechanistic target of rapamycin (mTOR), which is activated by VEGF in a delayed manner. We show that functional autophagy is required for VEGF-induced angiogenesis and may have specific functions in addition to maintaining homeostasis. In line with this, inhibition of autophagy impaired VEGF-mediated formation of the Notch intracellular domain, a critical regulator of angiogenesis. Our study characterizes autophagy induction as a pro-angiogenic function of the VEGF/AMPK pathway and suggests that timely activation of autophagy-initiating pathways may help to initiate angiogenesis.
    Keywords:  AMPK; ULK1; VEGF; angiogenesis; autophagy; mTOR
  9. Curr Genet. 2020 Mar 10.
      Parkinson's disease (PD) is a progressive neurodegenerative condition characterized by a gradual loss of a specific group of dopaminergic neurons in the substantia nigra. Importantly, current treatments only address the symptoms of PD, yet not the underlying molecular causes. Concomitantly, the function of genes that cause inherited forms of PD point to mitochondrial dysfunction as a major contributor in the etiology of PD. An inherent challenge that mitochondria face is the continuous exposure to diverse stresses including high levels of reactive oxygen species and protein misfolding, which increase their likelihood of dysregulation. In response, eukaryotic cells have evolved sophisticated quality control mechanisms to identify, repair and/or eliminate abnormal dysfunctional mitochondria. One such mechanism is mitophagy, a process which involves PTEN-induced putative kinase 1 (PINK1), a mitochondrial Ser/Thr kinase and Parkin, an E3 ubiquitin ligase, each encoded by genes responsible for early-onset autosomal recessive familial PD. Over 100 loss-of-function mutations in the PTEN-induced putative kinase 1 (PINK1) gene have been reported to cause autosomal recessive early-onset PD. PINK1 acts upstream of Parkin and is essential for the mitochondrial localization and activation of Parkin. Upon mitochondrial damage, PINK1 builds up on the outer mitochondrial membrane (OMM) and mediates the activation of Parkin. Activated Parkin then ubiquitinates numerous OMM proteins, eliciting mitochondrial autophagy (mitophagy). As a result, damaged mitochondrial components can be selectively eliminated. Thus, PINK1 acts a sensor of damage via fine-tuning of its levels on mitochondria, where it activates Parkin to orchestrate the clearance of unhealthy mitochondria. Previous work has unveiled that the Arg-N-end rule degradation pathway (Arg-N-degron pathway) mediates the degradation of PINK1, and thus fine-tune PINK1-dependent mitochondrial quality control pathway. Herein, we briefly discuss the interconnection between N-end rule degradation pathways and mitophagy in the context of N-degron mediated degradation of mitochondrial kinase PINK1 and highlight some of the future prospects.
    Keywords:  Mitochondrial quality control; N-degron; N-end rule; PINK1; Parkinson’s disease; Proteasome; Protein degradation; Ubiquitin
  10. J Neurochem. 2020 Mar 09. e15002
      In autophagy long-lived proteins, protein aggregates or damaged organelles are engulfed by vesicles called autophagosomes prior to lysosomal degradation. Autophagy dysfunction is a hallmark of several neurodegenerative diseases in which misfolded proteins or dysfunctional mitochondria accumulate. Excessive autophagy can also exacerbate brain injury under certain conditions. In this review, we provide specific examples to illustrate the critical role played by autophagy in pathological conditions affecting the brain and discuss potential therapeutic implications. We show how a singular type of autophagy-dependent cell death termed autosis has attracted attention as a promising target for improving outcomes in perinatal asphyxia and hypoxic-ischaemic injury to the immature brain. We provide evidence that autophagy inhibition may be protective against radiotherapy-induced damage to the young brain. We describe a specialized form of macroautophagy of therapeutic relevance for motoneuron and neuromuscular diseases, known as chaperone-assisted selective autophagy, in which heat shock protein B8 is used to deliver aberrant proteins to autophagosomes. We summarize studies pinpointing mitophagy mediated by the serine/threonine kinase PINK1 and the ubiquitin-protein ligase Parkin as a mechanism potentially relevant to Parkinson's disease, despite debate over the physiological conditions in which it is activated in organisms. Finally, with the example of the autophagy-inducing agent rilmenidine and its discrepant effects in cell culture and mouse models of motor neuron disorders, we illustrate the importance of considering aspects such a disease stage and aggressiveness, type of insult and load of damaged or toxic cellular components, when choosing the appropriate drug, timepoint and duration of treatment.
    Keywords:  Parkinsonism; autosis; chaperone; hypoxia-ischaemia; mitophagy; motor neuron
  11. Mol Cell Oncol. 2020 ;7(2): 1719021
      Although centrosome abnormalities are frequent in cancer, the mechanisms responsible for their accumulation are poorly understood. Here we comment on our recent publication identifying a new type of selective autophagy, named doryphagy, which preserves centrosome organization through targeting Centriolar Satellites (CS). Thus, doryphagy prevents inaccurate mitosis and genomic instability.
    Keywords:  Autophagy; GABARAP; GABARAPL2; PCM1; centriolar satellites; centrosome; doryphagy; mitosis; selective autophagy
  12. Autophagy. 2020 Mar 11.
      Macroautophagy/autophagy degrades proteins and organelles to generate macromolecular building blocks. As such, some cancer cells are particularly dependent on autophagy. In a previous paper, we found that even highly autophagy-dependent cancer cells can adapt to circumvent autophagy inhibition. However, it remains unclear if autophagy-dependent cancer cells could survive the complete elimination of autophagosome formation. We extended our previous findings to show that knockout (KO) of both the upstream autophagy regulator RB1CC1/FIP200 and the downstream regulator and mediator of LC3 conjugation, ATG7, strongly inhibits growth in highly autophagy-dependent cells within one week of editing. However, rare clones survived the loss of ATG7 or RB1CC1 and maintained growth even under autophagy-inducing conditions. Autophagy-dependent cells circumvent the complete loss of autophagy that is mediated by RB1CC1 KO, similar to the loss of ATG7, by upregulating NFE2L2/NRF2 signaling. These results indicate that cancer cell lines could adapt to the complete loss of autophagy by changing their biology to adopt alternative ways of dealing with autophagy-mediated cellular functions.
    Keywords:  ATG7; CRISPR-Cas9; NFE2L2/NRF2; ROS; STX17; adaptation
  13. Mol Biol Cell. 2020 Mar 11. mbcE19110622
      Environmental stress elicits well-orchestrated programs that either restore cellular homeostasis or induce cell death depending on the insult. Nutrient starvation triggers the autophagic pathway that requires the induction of several Autophagy (ATG) genes. Cyclin C-Cdk8 is a component of the RNA polymerase II Mediator complex that predominantly represses the transcription of stress-responsive genes in yeast. To relieve this repression following oxidative stress, cyclin C translocates to the mitochondria where it induces organelle fragmentation and promotes cell death prior to its destruction by the ubiquitin-proteasome-system (UPS). Here we report that cyclin C-Cdk8, together with the Ume6-Rpd3 histone deacetylase complex, represses the essential autophagy gene ATG8. Similar to oxidative stress, cyclin C is destroyed by the UPS following nitrogen starvation. Removing this repression is important as deleting CNC1 allows enhanced cell growth under mild starvation. However, unlike oxidative stress, cyclin C is destroyed prior to its cytoplasmic translocation. This is important as targeting cyclin C to the mitochondria induces both mitochondrial fragmentation and cell death following nitrogen starvation. These results indicate that cyclin C destruction pathways are fine-tuned depending on the stress and that its terminal subcellular address influences the decision between initiating cell death or cell survival pathways.
  14. J Mol Biol. 2020 Mar 10. pii: S0022-2836(20)30226-6. [Epub ahead of print]
  15. Autophagy. 2020 Mar 13. 1-18
      Lysosomal Ca2+ contributes to macroautophagy/autophagy, an intracellular process for the degradation of cytoplasmic material and organelles in the lysosomes to protect cells against stress responses. TMBIM6 (transmembrane BAX inhibitor motif containing 6) is a Ca2+ channel-like protein known to regulate ER stress response and apoptosis. In this study, we examined the as yet unknown role of TMBIM6 in regulating lysosomal Ca2+ levels. The Ca2+ efflux from the ER through TMBIM6 was found to increase the resting lysosomal Ca2+ level, in which ITPR-independent regulation of Ca2+ status was observed. Further, TMBIM6 regulated the local release of Ca2+ through lysosomal MCOLN1/TRPML1 channels under nutrient starvation or MTOR inhibition. The local Ca2+ efflux through MCOLN1 channels was found to activate PPP3/calcineurin, triggering TFEB (transcription factor EB) nuclear translocation, autophagy induction, and lysosome biogenesis. Upon genetic inactivation of TMBIM6, lysosomal Ca2+ and the associated TFEB nuclear translocation were decreased. Furthermore, autophagy flux was significantly enhanced in the liver or kidney from starved Tmbim6+/+ mice compared with that in the counter tmbim6-/- mice. Together, our observations indicated that under stress conditions, TMBIM6 increases lysosomal Ca2+ release, leading to PPP3/calcineurin-mediated TFEB activation and subsequently enhanced autophagy. Thus, TMBIM6, an ER membrane protein, is suggested to be a lysosomal Ca2+ modulator that coordinates with autophagy to alleviate metabolism stress.Abbreviations: AVs: autophagic vacuoles; CEPIA: calcium-measuring organelle-entrapped protein indicator; ER: endoplasmic reticulum; GPN: glycyl-L-phenylalanine-beta-naphthylamide; ITPR/IP3R: inositol 1,4,5-trisphosphate receptor; LAMP1: lysosomal associated membrane protein 1; MCOLN/TRPML: mucolipin; MEF: mouse embryonic fibroblast; ML-SA1: mucolipin synthetic agonist 1; MTORC1: mechanistic target of rapamycin kinase complex 1; RPS6KB1: ribosomal protein S6 kinase B1; SQSTM1: sequestosome 1; TFEB: transcription factor EB; TKO: triple knockout; TMBIM6/BI-1: transmembrane BAX inhibitor motif containing 6.
    Keywords:  Autophagy; MTORC1; TMBIM6; lysosomal calcium; transcription factor EB (TFEB)
  16. Semin Cell Dev Biol. 2020 Mar 10. pii: S1084-9521(18)30301-X. [Epub ahead of print]
      Lipid droplets (LDs) are not an inert storage of excessive lipids, but play various roles in cellular lipid metabolism. Autophagy involves several mechanisms for the degradation of cellular components, and is related to many aspects of lipid metabolism. LD and autophagic membranes often distribute in proximity, but their relationship is complex. LDs can be degraded by autophagy, but LDs are also generated as a result of autophagy or support the execution of autophagy. Moreover, several proteins crucial for autophagy were shown to affect different aspects of LD formation. This article aims to categorize this multifaceted and seemingly entangled LD-autophagy relationship and to discuss unresolved issues.
    Keywords:  Autophagy; Isolation membrane; Lipid droplet; Lipophagy; ndoplasmic reticulum
  17. EMBO Rep. 2020 Mar 11. e48901
      Recent evidence has linked the lysosomal cholesterol accumulation in Niemann-Pick type C1 with anomalies associated with primary ciliogenesis. Here, we report that perturbed intracellular cholesterol distribution imposed by lysosomal cholesterol accumulation during TMEM135 depletion is closely associated with impaired ciliogenesis. TMEM135 depletion does not affect the formation of the basal body and the ciliary transition zone. TMEM135 depletion severely blunts Rab8 trafficking to the centrioles without affecting the centriolar localization of Rab11 and Rabin8, the upstream regulators of Rab8 activation. Although TMEM135 depletion prevents enhanced IFT20 localization at the centrioles, ciliary vesicle formation is not affected. Furthermore, enhanced IFT20 localization at the centrioles is dependent on Rab8 activation. Supplementation of cholesterol in complex with cyclodextrin rescues Rab8 trafficking to the centrioles and Rab8 activation, thereby recovering primary ciliogenesis in TMEM135-depleted cells. Taken together, our data suggest that TMEM135 depletion prevents ciliary vesicle elongation, a characteristic of impaired Rab8 function. Our study thus reveals a previously uncharacterized effect of erroneous intracellular cholesterol distribution on impairing Rab8 function and primary ciliogenesis.
    Keywords:  IFT20; Rab8; TMEM135; intracellular cholesterol transport; peroxisome; primary cilia
  18. Elife. 2020 Mar 13. pii: e54712. [Epub ahead of print]9
      Ion selectivity is a defining feature of a given ion channel and is considered immutable. Here we show that ion selectivity of the lysosomal ion channel TPC2, which is hotly debated (Calcraft et al., 2009; Guo et al., 2017; Jha et al., 2014; Ruas et al., 2015; Wang et al., 2012), depends on the activating ligand. A high throughput screen identified two structurally distinct TPC2 agonists. One of these evoked robust Ca2+-signals and non-selective cation currents, the other weaker Ca2+-signals and Na+-selective currents. These properties were mirrored by the Ca2+-mobilizing messenger, NAADP and the phosphoinositide, PI(3,5)P2, respectively. Agonist action was differentially inhibited by mutation of a single TPC2 residue and coupled to opposing changes in lysosomal pH and exocytosis. Our findings resolve conflicting reports on the permeability and gating properties of TPC2 and they establish a new paradigm whereby a single ion channel mediates distinct, functionally-relevant ionic signatures on demand.
    Keywords:  biochemistry; chemical biology; mouse
  19. Autophagy. 2020 Mar 11.
      NRBF2, a regulatory subunit of the ATG14-BECN1/Beclin 1-PIK3C3/VPS34 complex, positively regulates macroautophagy/autophagy. In this study, we report that NRBF2 is required for the clearance of apoptotic cells and alleviation of inflammation during colitis in mice. NRBF2-deficient mice displayed much more severe colitis symptoms after the administration of ulcerative colitis inducer, dextran sulfate sodium salt (DSS), accompanied by prominent intestinal inflammation and apoptotic cell accumulation. Interestingly, we found that nrbf2-/- mice and macrophages displayed impaired apoptotic cell clearance capability, while adoptive transfer of Nrbf2+/+ macrophages to nrbf2-/- mice alleviated DSS-induced colitis lesions. Mechanistically, NRBF2 is required for the generation of the active form of RAB7 to promote the fusion between phagosomes containing engulfed apoptotic cells and lysosomes via interacting with the MON1-CCZ1 complex and regulating the guanine nucleotide exchange factor (GEF) activity of the complex. Evidence from clinical samples further reveals the physiological role of NRBF2 in maintaining intestinal homeostasis. In biopsies of UC patient colon, we observed upregulated NRBF2 in the colon macrophages and the engulfment of apoptotic cells by NRBF2-positive cells, suggesting a potential protective role for NRBF2 in UC. To confirm the relationship between apoptotic cell clearance and IBD development, we compared TUNEL-stained cell counts in the UC with UC severity (Mayo Score) and observed a strong correlation between the two indexes, indicating that apoptotic cell population in colon tissue correlates with UC severity. The findings of our study reveal a novel role for NRBF2 in regulating apoptotic cell clearance to restrict intestinal inflammation.
    Keywords:  MON1-CCZ1; NRBF2; RAB7; apoptotic cell clearance; inflammatory bowel disease; macrophage
  20. Int J Mol Sci. 2020 Mar 05. pii: E1772. [Epub ahead of print]21(5):
      Endoplasmic reticulum (ER)-mitochondria contact sites are critical structures for cellular function. They are implicated in a plethora of cellular processes, including Ca2+ signalling and mitophagy, the selective degradation of damaged mitochondria. Phosphatase and tensin homolog (PTEN)-induced kinase (PINK) and Parkin proteins, whose mutations are associated with familial forms of Parkinson's disease, are two of the best characterized mitophagy players. They accumulate at ER-mitochondria contact sites and modulate organelles crosstalk. Alterations in ER-mitochondria tethering are a common hallmark of many neurodegenerative diseases including Parkinson's disease. Here, we summarize the current knowledge on the involvement of PINK1 and Parkin at the ER-mitochondria contact sites and their role in the modulation of Ca2+ signalling and mitophagy.
    Keywords:  Ca2+; ER–mitochondria tethering; PINK1; Parkin; mitophagy
  21. EMBO J. 2020 Mar 10. e102468
      Vertebrate vision relies on the daily phagocytosis and lysosomal degradation of photoreceptor outer segments (POS) within the retinal pigment epithelium (RPE). However, how these events are controlled by light is largely unknown. Here, we show that the light-responsive miR-211 controls lysosomal biogenesis at the beginning of light-dark transitions in the RPE by targeting Ezrin, a cytoskeleton-associated protein essential for the regulation of calcium homeostasis. miR-211-mediated down-regulation of Ezrin leads to Ca2+ influx resulting in the activation of calcineurin, which in turn activates TFEB, the master regulator of lysosomal biogenesis. Light-mediated induction of lysosomal biogenesis and function is impaired in the RPE from miR-211-/- mice that show severely compromised vision. Pharmacological restoration of lysosomal biogenesis through Ezrin inhibition rescued the miR-211-/- phenotype, pointing to a new therapeutic target to counteract retinal degeneration associated with lysosomal dysfunction.
    Keywords:  Ezrin; RPE; TFEB; autophagy; miR-211
  22. Front Cell Dev Biol. 2020 ;8 90
      The endoplasmic reticulum (ER) is a dynamic membrane system comprising different and interconnected subdomains. The ER structure changes in response to different stress conditions through the activation of a selective autophagic pathway called ER-phagy. This represents a quality control mechanism for ER turnover and component recycling. Several ER-resident proteins have been indicated as receptors for ER-phagy; among these, there are proteins characterized by the presence of a reticulon homology domain (RHD). RHD-containing proteins promote ER fragmentation by a mechanism that involves LC3 binding and lysosome delivery. Moreover, the presence of a correct RHD structure is closely related to their capability to regulate ER shape and morphology by curvature induction and membrane remodeling. Deregulation of the ER-selective autophagic pathway due to defects in proteins with RHD has been implicated in several human diseases, infectious and neurodegenerative diseases in particular, as well as in cancer development. While the molecular mechanisms and the physiological role of ER-phagy are not yet fully understood, it is quite clear that this process is involved in different cellular signaling pathways and has an impact in several human pathologies.
    Keywords:  autophagy; domain; endoplasmic reticulum; membrane; reticulon
  23. Dev Cell. 2020 Mar 09. pii: S1534-5807(20)30101-5. [Epub ahead of print]52(5): 542-544
      Cancer cells need to acquire specific molecular traits in order to spread to distant organs. In this issue of Developmental Cell, Marsh et al. show that autophagy restricts the outgrowth of breast cancer metastases in contrast to its impact on primary tumor progression.
  24. Nat Commun. 2020 Mar 12. 11(1): 1325
      Brain wiring is remarkably precise, yet most neurons readily form synapses with incorrect partners when given the opportunity. Dynamic axon-dendritic positioning can restrict synaptogenic encounters, but the spatiotemporal interaction kinetics and their regulation remain essentially unknown inside developing brains. Here we show that the kinetics of axonal filopodia restrict synapse formation and partner choice for neurons that are not otherwise prevented from making incorrect synapses. Using 4D imaging in developing Drosophila brains, we show that filopodial kinetics are regulated by autophagy, a prevalent degradation mechanism whose role in brain development remains poorly understood. With surprising specificity, autophagosomes form in synaptogenic filopodia, followed by filopodial collapse. Altered autophagic degradation of synaptic building material quantitatively regulates synapse formation as shown by computational modeling and genetic experiments. Increased filopodial stability enables incorrect synaptic partnerships. Hence, filopodial autophagy restricts inappropriate partner choice through a process of kinetic exclusion that critically contributes to wiring specificity.
  25. Autophagy. 2020 Mar 11.
      Trimethyltin chloride (TMT) is widely used as a constituent of fungicides and plastic stabilizers in the industrial and agricultural fields, and is generally acknowledged to have potent neurotoxicity, especially in the hippocampus; however, the mechanism of induction of neurotoxicity by TMT remains elusive. Herein, we exposed Neuro-2a cells to different concentrations of TMT (2, 4, and 8 μM) for 24 h. Proteomic analysis, coupled with bioinformatics analysis, revealed the important role of macroautophagy/autophagy-lysosome machinery in TMT-induced neurotoxicity. Further analyses indicated significant impairment of autophagic flux by TMT via suppressed lysosomal function, such as by inhibiting lysosomal proteolysis and changing the lysosomal pH, thereby contributing to defects in autophagic clearance and subsequently leading to nerve cell death. Mechanistically, molecular interaction networks of Ingenuity Pathway Analysis identified a downregulated molecule, KIF5A (kinesin family member 5A), as a key target in TMT-impaired autophagic flux. TMT decreased KIF5A protein expression, disrupted the interaction between KIF5A and lysosome, and impaired lysosomal axonal transport. Moreover, Kif5a overexpression restored axonal transport, increased lysosomal dysfunction, and antagonized TMT-induced neurotoxicity in vitro. Importantly, in TMT-administered mice with seizure symptoms and histomorphological injury in the hippocampus, TMT inhibited KIF5A expression in the hippocampus. Gene transfer of Kif5a enhanced autophagic clearance in the hippocampus and alleviated TMT-induced neurotoxicity in vivo. Our results are the first to demonstrate KIF5A-dependent axonal transport deficiency to cause autophagic flux impairment via disturbance of lysosomal function in TMT-induced neurotoxicity; manipulation of KIF5A may be a therapeutic approach for antagonizing TMT-induced neurotoxicity.
    Keywords:  Ingenuity Pathway Analysis; KIF5A; autophagy; axonal transport; neurotoxicity; proteomic analysis; trimethyltin
  26. Antioxid Redox Signal. 2020 Mar 13.
      SIGNIFICANCE: Cell senescence was originally defined by an acute loss of replicative capacity and thus thought to be restricted to proliferation-competent cells. More recently, senescence has been recognised as a cellular stress and damage response encompassing multiple pathways or senescence domains, namely DNA Damage Response (DDR), cell cycle arrest, Senescence-Associated Secretory Phenotype (SASP), Senescence-Associated Mitochondrial Dysfunction (SAMD), Autophagy/Mitophagy Dysfunction, Nutrient and Stress Signalling and Epigenetic Reprogramming. Each of these domains are activated during senescence, and all appear to interact with each other. Cell senescence has been identified as an important driver of mammalian ageing. Recent Advances: Activation of all these senescence domains has now also been observed in a wide range of post-mitotic cells, suggesting that senescence as a stress response can occur in non-dividing cells temporally uncoupled from cell cycle arrest. Here, we review recent evidence for post-mitotic cell senescence and speculate about its possible relevance for mammalian ageing.CRITICAL ISSUES: While a majority of senescence domains has been found to be activated in a range of post-mitotic cells during ageing, independent confirmation of these results is still lacking for most of them.
    FUTURE DIRECTIONS: To define whether post-mitotic senescence plays a significant role as driver of ageing phenotypes in tissues like brain, muscle, heart and others.
  27. Cell Rep. 2020 Mar 10. pii: S2211-1247(20)30226-6. [Epub ahead of print]30(10): 3240-3249.e4
      Target of Rapamycin Complex 1 (TORC1) signaling promotes growth and aging. Inhibition of TORC1 leads to reduced protein translation, which promotes longevity. TORC1-dependent post-transcriptional regulation of protein translation has been well studied, while analogous transcriptional regulation is less understood. Here we screen fission yeast mutants for resistance to Torin1, which inhibits TORC1 and cell growth. Cells lacking the GATA factor Gaf1 (gaf1Δ) grow normally even in high doses of Torin1. The gaf1Δ mutation shortens the chronological lifespan of non-dividing cells and diminishes Torin1-mediated longevity. Expression profiling and genome-wide binding experiments show that upon TORC1 inhibition, Gaf1 directly upregulates genes for small-molecule metabolic pathways and indirectly represses genes for protein translation. Surprisingly, Gaf1 binds to and downregulates the tRNA genes, so it also functions as a transcription factor for RNA polymerase III. Thus, Gaf1 controls the transcription of both protein-coding and tRNA genes to inhibit translation and growth downstream of TORC1.
    Keywords:  GATA6; RNA polymerase III; S. pombe; TOR; aging; cell growth; protein translation; tRNA; transcription factor; vacuole
  28. J Mol Biol. 2020 Mar 04. pii: S0022-2836(20)30220-5. [Epub ahead of print]
      Rare genetic diseases affect a limited number of patients, but their etiology is often known, facilitating the development of reliable animal models and giving the opportunity to investigate physiopathology. Lysosomal storage disorders are a group of rare diseases due to primary alteration of lysosome function. These diseases are often associated with neurological symptoms, which highlighted the importance of lysosome in neurodegeneration. Likewise, other groups of rare neurodegenerative diseases also present lysosomal alteration. Lysosomes fuse with autophagosomes and endosomes to allow the degradation of their content thanks to hydrolytic enzymes. It has emerged that alteration of the autophagy-lysosome pathway could play a critical role in neuronal death in many neurodegenerative diseases. Using a repertoire of selected rare neurodegenerative diseases, we highlight that a variety of alterations of the autophagy-lysosomal pathway are associated with neuronal death. Yet, in most cases, it is still unclear why alteration of this pathway can lead to neurodegeneration.
    Keywords:  Lysosomal storage diseases; autophagy; neuronal death; signaling; trafficking
  29. Nat Med. 2020 Mar;26(3): 398-407
    Dominantly Inherited Alzheimer Network
      Development of tau-based therapies for Alzheimer's disease requires an understanding of the timing of disease-related changes in tau. We quantified the phosphorylation state at multiple sites of the tau protein in cerebrospinal fluid markers across four decades of disease progression in dominantly inherited Alzheimer's disease. We identified a pattern of tau staging where site-specific phosphorylation changes occur at different periods of disease progression and follow distinct trajectories over time. These tau phosphorylation state changes are uniquely associated with structural, metabolic, neurodegenerative and clinical markers of disease, and some (p-tau217 and p-tau181) begin with the initial increases in aggregate amyloid-β as early as two decades before the development of aggregated tau pathology. Others (p-tau205 and t-tau) increase with atrophy and hypometabolism closer to symptom onset. These findings provide insights into the pathways linking tau, amyloid-β and neurodegeneration, and may facilitate clinical trials of tau-based treatments.
  30. Mol Cell Oncol. 2020 ;7(2): 1685840
      "In the field of observation, chance favours only the prepared mind" (Louis Pasteur). This motto seems to have guided our unexpected results published recently in Nature Communications, where we describe an epigenetic rheostat that regulates expression of the constituents of the lysosomal and autophagic systems.
    Keywords:  Acetylation; Epigenetic regulation; Lysosome and autophagy gene transcription; MYC; MiT/TFE; intermittent drainage
  31. Redox Biol. 2020 Feb 07. pii: S2213-2317(19)31460-0. [Epub ahead of print] 101450
      Reactive Oxygen Species (ROS) are essential cellular messengers required for cellular homeostasis and regulate the lifespan of several animal species. The main site of ROS production is the mitochondrion, and within it, respiratory complex I (CI) is the main ROS generator. ROS produced by CI trigger several physiological responses that are essential for the survival of neurons, cardiomyocytes and macrophages. Here, we show that CI produces ROS when electrons flow in either the forward (Forward Electron Transport, FET) or reverse direction (Reverse Electron Transport, RET). We demonstrate that ROS production via RET (ROS-RET) is activated under thermal stress conditions and that interruption of ROS-RET production, through ectopic expression of the alternative oxidase AOX, attenuates the activation of pro-survival pathways in response to stress. Accordingly, we find that both suppressing ROS-RET signalling or decreasing levels of mitochondrial H2O2 by overexpressing mitochondrial catalase (mtCAT), reduces survival dramatically in flies under stress. Our results uncover a specific ROS signalling pathway where hydrogen peroxide (H2O2) generated by CI via RET is required to activate adaptive mechanisms, maximising survival under stress conditions.
    Keywords:  AOX; Alternative oxidase; Complex I; Heat stress; Reactive oxygen species; Reverse electron transport
  32. Sci Rep. 2020 Mar 10. 10(1): 4401
      Mitochondrial dysfunctions belong amongst the most common metabolic diseases but the signalling networks that lead to the manifestation of a disease phenotype are often not well understood. We identified the subunits of respiratory complex I, III and IV as mediators of major signalling changes during Drosophila wing disc development. Their downregulation in larval wing disc leads to robust stimulation of TOR activity, which in turn orchestrates a complex downstream signalling network. Specifically, after downregulation of the complex I subunit ND-49 (mammalian NDUFS2), TOR activates JNK to induce cell death and ROS production essential for the stimulation of compensatory apoptosis-induced proliferation within the tissue. Additionally, TOR upregulates Notch and JAK/STAT signalling and it directs glycolytic switch of the target tissue. Our results highlight the central role of TOR signalling in mediating the complex response to mitochondrial respiratory dysfunction and they provide a rationale why the disease symptoms associated with respiratory dysfunctions are often alleviated by mTOR inhibitors.
  33. Science. 2020 Mar 13. 367(6483): 1230-1234
      How long-lived memories withstand molecular turnover is a fundamental question. Aggregates of a prion-like RNA-binding protein, cytoplasmic polyadenylation element-binding (CPEB) protein, is a putative substrate of long-lasting memories. We isolated aggregated Drosophila CPEB, Orb2, from adult heads and determined its activity and atomic structure, at 2.6-angstrom resolution, using cryo-electron microscopy. Orb2 formed ~75-nanometer-long threefold-symmetric amyloid filaments. Filament formation transformed Orb2 from a translation repressor to an activator and "seed" for further translationally active aggregation. The 31-amino acid protofilament core adopted a cross-β unit with a single hydrophilic hairpin stabilized through interdigitated glutamine packing. Unlike the hydrophobic core of pathogenic amyloids, the hydrophilic core of Orb2 filaments suggests how some neuronal amyloids could be a stable yet regulatable substrate of memory.