bims-apauto Biomed News
on Apoptosis and autophagy
Issue of 2023‒01‒29
twelve papers selected by
Su Hyun Lee
Harvard University

  1. Autophagy. 2023 Jan 24. 1-18
      Eukaryotic stress granules (SGs) are highly dynamic assemblies of untranslated mRNAs and proteins that form through liquid-liquid phase separation (LLPS) under cellular stress. SG formation and elimination process is a conserved cellular strategy to promote cell survival, although the precise regulation of this process is poorly understood. Here, we screened six E3 ubiquitin ligases present in SGs and identified TRIM21 (tripartite motif containing 21) as a central regulator of SG homeostasis that is highly enriched in SGs of cells under arsenite-induced oxidative stress. Knockdown of TRIM21 promotes SG formation whereas overexpression of TRIM21 inhibits the formation of physiological and pathological SGs associated with neurodegenerative diseases. TRIM21 catalyzes K63-linked ubiquitination of the SG core protein, G3BP1 (G3BP stress granule assembly factor 1), and G3BP1 ubiquitination can effectively inhibit LLPS, in vitro. Recent reports suggested the involvement of macroautophagy/autophagy, as a stress response pathway, in the regulation of SG homeostasis. We systematically investigated well-defined autophagy receptors and identified SQSTM1/p62 (sequestosome 1) and CALCOCO2/NDP52 (calcium binding and coiled-coil domain 2) as the primary receptors that directly interact with G3BP1 during arsenite-induced stress. Endogenous SQSTM1 and CALCOCO2 localize to the periphery of SGs under oxidative stress and mediate SG elimination, as single knockout of each receptor causes accumulation of physiological and pathological SGs. Collectively, our study broadens the understanding in the regulation of SG homeostasis by showing that TRIM21 and autophagy receptors modulate SG formation and elimination respectively, suggesting the possibility of clinical targeting of these molecules in therapeutic strategies for neurodegenerative diseases.Abbreviations: ACTB: actin beta; ALS: amyotrophic lateral sclerosis; BafA1: bafilomycin A1; BECN1: beclin 1; C9orf72: C9orf72-SMCR8 complex subunit; CALCOCO2/NDP52: calcium binding and coiled-coil domain 2; Co-IP: co-immunoprecipitation; DAPI: 4',6-diamidino-2-phenylindole; FTD: frontotemporal dementia; FUS: FUS RNA binding protein; G3BP1: G3BP stress granule assembly factor 1; GFP: green fluorescent protein; LLPS: liquid-liquid phase separation; MAP1LC3/LC3: microtubule associated protein 1 light chain 3; NBR1: NBR1 autophagy cargo receptor; NES: nuclear export signal; OPTN: optineurin; RFP: red fluorescent protein; SQSTM1/p62: sequestosome 1; SG: stress granule; TAX1BP1: Tax1 binding protein 1; TOLLIP: toll interacting protein; TRIM21: tripartite motif containing 21; TRIM56: tripartite motif containing 56; UB: ubiquitin; ULK1: unc-51 like autophagy activating kinase 1; WT: wild-type.
    Keywords:  Autophagy receptor; CALCOCO2; G3BP1; SQSTM1; TRIM21; stress granule; ubiquitination
  2. Cell Rep. 2023 Jan 25. pii: S2211-1247(23)00048-7. [Epub ahead of print]42(2): 112037
      In response to lysosomal damage, cells engage several quality-control mechanisms, including the selective isolation and degradation of damaged lysosomes by lysophagy. Here, we report that the selective autophagy adaptor SQSTM1/p62 is recruited to damaged lysosomes in both HeLa cells and neurons and is required for lysophagic flux. The Phox and Bem1p (PB1) domain of p62 mediates oligomerization and is specifically required for lysophagy. Consistent with this observation, we find that p62 forms condensates on damaged lysosomes. These condensates are precisely tuned by the small heat shock protein HSP27, which is phosphorylated in response to lysosomal injury and maintains the liquidity of p62 condensates, facilitating autophagosome formation. Mutations in p62 have been identified in patients with amyotrophic lateral sclerosis (ALS); ALS-associated mutations in p62 impair lysophagy, suggesting that deficits in this pathway may contribute to neurodegeneration. Thus, p62 condensates regulated by HSP27 promote lysophagy by forming platforms for autophagosome biogenesis at damaged lysosomes.
    Keywords:  ALS; CP: Cell biology; CP: Neuroscience; HSP27; autophagy; condensates; lysophagy; lysosome; neurodegeneration; neurons; p62/SQSTM1; phase separation
  3. Adv Protein Chem Struct Biol. 2023 ;pii: S1876-1623(22)00088-8. [Epub ahead of print]133 159-180
      Autophagy, a classical cellular degradative catabolic process, also involves a functionally discrete non-degradative role in eukaryotic cells. It imparts critical regulatory function on conventional and unconventional protein secretion (degradative and secretory autophagy with distinct lysosomal degradation and extracellular expulsion, respectively) pathways. The N-amino terminal leader sequence containing proteins follows a conventional secretion pathway, while the leader-less proteins opt for secretory autophagy. The secretory autophagic process ensembles core autophagy machinery proteins, specifically ULK1/2, Beclin 1, LC3, and GABARAP, in coordination with Golgi re-assembly and stacking proteins (GRASPs). The secretory omegasomes fuse with the plasma membrane for the expulsion of cytosolic cargos to the extracellular environment. Alternatively, the secretory omegasomes also fuse with multi-vesicular bodies (MVBs) and harmonize ESCRTs (Complex I; TSG101) and Rab GTPase for their release to extracellular space. Autophagy has been associated with the secretion of diverse proteins involved in cellular signaling, inflammation, and carcinogenesis. Secreted proteins play an essential role in cancer by sustaining cell proliferation, inhibiting apoptosis, enhancing angiogenesis and metastasis, immune cell regulation, modulation of cellular energy metabolism, and resistance to anticancer drugs. The complexity of autophagy regulation during tumorigenesis is dependent on protein secretion pathways. Autophagy-regulated TOR-autophagy spatial coupling compartment complex energizes enhanced secretion of pro-inflammatory cytokines and leaderless proteins such as HMGB1. In conclusion, the chapter reviews the role of autophagy in regulating conventional and unconventional protein secretion pathways and its possible role in cancer.
    Keywords:  Autophagy; Cancer; Protein secretion; Unconventional autophagy
  4. Autophagy. 2023 Jan 24.
      Mitophagy and its variants are considered important salvage pathways to remove dysfunctional mitochondria. Non-canonical mitophagy, independent of autophagosome formation and including endosomal-dependent mitophagy, operate upon specific injury. In a recent paper, we describe a new mechanism where, upon mtDNA damage, mitochondrial nucleoids are eliminated via an endosomal-mitophagy pathway. Using proximity proteomics, we identified the proteins required for elimination of mutated mitochondrial nucleoids from the mitochondrial matrix. Among them, ATAD3 and SAMM50 control cristae architecture and nucleoid interaction, necessary for mtDNA extraction. In the mitochondrial outer membrane, SAMM50 coordinates with the retromer protein VPS35 to sequester mtDNA in endosomes and guide them towards elimination, thus avoiding the activation of an exacerbated immune response. Here, we summarize our findings and examine how this newly described pathway contributes to our understanding of mtDNA quality control.
    Keywords:  - mitophagy; endosomes; mtDNA
  5. Front Physiol. 2022 ;13 1115511
    Keywords:  autophagy; autophagy regulation; autophagy-related clinical practice; autophagy-related diseases; cardiovascular diseases; diabetes; protozoa
  6. Mol Cell. 2023 Jan 14. pii: S1097-2765(22)01217-5. [Epub ahead of print]
      The protection of DNA replication forks under stress is essential for genome maintenance and cancer suppression. One mechanism of fork protection involves an elevation in intracellular Ca2+ ([Ca2+]i), which in turn activates CaMKK2 and AMPK to prevent uncontrolled fork processing by Exo1. How replication stress triggers [Ca2+]i elevation is unclear. Here, we report a role of cytosolic self-DNA (cytosDNA) and the ion channel TRPV2 in [Ca2+]i induction and fork protection. Replication stress leads to the generation of ssDNA and dsDNA species that, upon translocation into cytoplasm, trigger the activation of the sensor protein cGAS and the production of cGAMP. The subsequent binding of cGAMP to STING causes its dissociation from TRPV2, leading to TRPV2 derepression and Ca2+ release from the ER, which in turn activates the downstream signaling cascade to prevent fork degradation. This Ca2+-dependent genome protection pathway is also activated in response to replication stress caused by oncogene activation.
    Keywords:  AMPK; CaMKK2; STING; TREX1; TRPV2; cGAS; cytosolic DNA; fork resection; intracellular Ca(2+); replication stress
  7. Nat Cell Biol. 2023 Jan 23.
      Mitochondria are complex organelles with different compartments, each harbouring their own protein quality control factors. While chaperones of the mitochondrial matrix are well characterized, it is poorly understood which chaperones protect the mitochondrial intermembrane space. Here we show that cytosolic small heat shock proteins are imported under basal conditions into the mitochondrial intermembrane space, where they operate as molecular chaperones. Protein misfolding in the mitochondrial intermembrane space leads to increased recruitment of small heat shock proteins. Depletion of small heat shock proteins leads to mitochondrial swelling and reduced respiration, while aggregation of aggregation-prone substrates is countered in their presence. Charcot-Marie-Tooth disease-causing mutations disturb the mitochondrial function of HSPB1, potentially linking previously observed mitochondrial dysfunction in Charcot-Marie-Tooth type 2F to its role in the mitochondrial intermembrane space. Our results reveal that small heat shock proteins form a chaperone system that operates in the mitochondrial intermembrane space.
  8. Nature. 2023 Jan 25.
      Mitochondria have crucial roles in cellular energetics, metabolism, signalling and quality control1-4. They contain around 1,000 different proteins that often assemble into complexes and supercomplexes such as respiratory complexes and preprotein translocases1,3-7. The composition of the mitochondrial proteome has been characterized1,3,5,6; however, the organization of mitochondrial proteins into stable and dynamic assemblies is poorly understood for major parts of the proteome1,4,7. Here we report quantitative mapping of mitochondrial protein assemblies using high-resolution complexome profiling of more than 90% of the yeast mitochondrial proteome, termed MitCOM. An analysis of the MitCOM dataset resolves >5,200 protein peaks with an average of six peaks per protein and demonstrates a notable complexity of mitochondrial protein assemblies with distinct appearance for respiration, metabolism, biogenesis, dynamics, regulation and redox processes. We detect interactors of the mitochondrial receptor for cytosolic ribosomes, of prohibitin scaffolds and of respiratory complexes. The identification of quality-control factors operating at the mitochondrial protein entry gate reveals pathways for preprotein ubiquitylation, deubiquitylation and degradation. Interactions between the peptidyl-tRNA hydrolase Pth2 and the entry gate led to the elucidation of a constitutive pathway for the removal of preproteins. The MitCOM dataset-which is accessible through an interactive profile viewer-is a comprehensive resource for the identification, organization and interaction of mitochondrial machineries and pathways.
  9. EMBO J. 2023 Jan 24. e112344
      Target of rapamycin complex 1 (TORC1) promotes biogenesis and inhibits the degradation of ribosomes in response to nutrient availability. To ensure a basal supply of ribosomes, cells are known to preserve a small pool of dormant ribosomes under nutrient-limited conditions. However, the regulation of these dormant ribosomes is poorly characterized. Here, we show that upon inhibition of yeast TORC1 by rapamycin or nitrogen starvation, the ribosome preservation factor Stm1 mediates the formation of nontranslating, dormant 80S ribosomes. Furthermore, Stm1-bound 80S ribosomes are protected from proteasomal degradation. Upon nutrient replenishment, TORC1 directly phosphorylates and inhibits Stm1 to reactivate translation. Finally, we find that SERBP1, a mammalian ortholog of Stm1, is likewise required for the formation of dormant 80S ribosomes upon mTORC1 inhibition in mammalian cells. These data suggest that TORC1 regulates ribosomal dormancy in an evolutionarily conserved manner by directly targeting a ribosome preservation factor.
    Keywords:  SERBP1; Stm1; TORC1; dormant ribosomes; proteasome
  10. EMBO J. 2023 Jan 27. e112309
      Hundreds of nucleus-encoded mitochondrial precursor proteins are synthesized in the cytosol and imported into mitochondria in a post-translational manner. However, the early processes associated with mitochondrial protein targeting remain poorly understood. Here, we show that in Saccharomyces cerevisiae, the cytosol has the capacity to transiently store mitochondrial matrix-destined precursors in dedicated deposits that we termed MitoStores. Competitive inhibition of mitochondrial protein import via clogging of import sites greatly enhances the formation of MitoStores, but they also form during physiological cell growth on nonfermentable carbon sources. MitoStores are enriched for a specific subset of nucleus-encoded mitochondrial proteins, in particular those containing N-terminal mitochondrial targeting sequences. Our results suggest that MitoStore formation suppresses the toxic potential of aberrantly accumulating mitochondrial precursor proteins and is controlled by the heat shock proteins Hsp42 and Hsp104. Thus, the cytosolic protein quality control system plays an active role during the early stages of mitochondrial protein targeting through the coordinated and localized sequestration of mitochondrial precursor proteins.
    Keywords:  chaperones; mitochondria; proteasome; protein aggregates; protein translocation
  11. Cell Death Discov. 2023 Jan 23. 9(1): 23
      NEDD8-conjugating enzymes, E2s, include the well-studied ubiquitin-conjugating enzyme E2 M (UBE2M) and the poorly characterized ubiquitin-conjugating enzyme E2 F (UBE2F). UBE2M and UBE2F have distinct and prominent roles in catalyzing the neddylation of Cullin or non-Cullin substrates. These enzymes are overexpressed in various malignancies, conferring a worse overall survival. Targeting UBE2M to influence tumor growth by either modulating several biological responses of tumor cells (such as DNA-damage response, apoptosis, or senescence) or regulating the anti-tumor immunity holds strong therapeutic potential. Multiple inhibitors that target the interaction between UBE2M and defective cullin neddylation protein 1 (DCN1), a co-E3 for neddylation, exhibit promising anti-tumor effects. By contrast, the potential benefits of targeting UBE2F are still to be explored. It is currently reported to inhibit apoptosis and then induce cell growth; hence, targeting UBE2F serves as an effective chemo-/radiosensitizing strategy by triggering apoptosis. This review highlights the most recent advances in the roles of UBE2M and UBE2F in tumor progression, indicating these E2s as two promising anti-tumor targets.
  12. Cell Death Dis. 2023 Jan 23. 14(1): 54
      The mitochondrial protein IF1 binds to the catalytic domain of the ATP synthase and inhibits ATP hydrolysis in ischemic tissues. Moreover, IF1 is overexpressed in many tumors and has been shown to act as a pro-oncogenic protein, although its mechanism of action is still debated. Here, we show that ATP5IF1 gene disruption in HeLa cells decreases colony formation in soft agar and tumor mass development in xenografts, underlining the role of IF1 in cancer. Notably, the lack of IF1 does not affect proliferation or oligomycin-sensitive mitochondrial respiration, but it sensitizes the cells to the opening of the permeability transition pore (PTP). Immunoprecipitation and proximity ligation analysis show that IF1 binds to the ATP synthase OSCP subunit in HeLa cells under oxidative phosphorylation conditions. The IF1-OSCP interaction is confirmed by NMR spectroscopy analysis of the recombinant soluble proteins. Overall, our results suggest that the IF1-OSCP interaction protects cancer cells from PTP-dependent apoptosis under normoxic conditions.