bims-mikwok Biomed News
on Mitochondrial quality control
Issue of 2021‒01‒10
eighteen papers selected by
Avinash N. Mukkala
University of Toronto

  1. J Biol Chem. 2020 Dec 02. pii: S0021-9258(20)00036-8. [Epub ahead of print]296 100050
      Large cytosolic protein aggregates are removed by two main cellular processes, autophagy and the ubiquitin-proteasome system, and defective clearance of these protein aggregates results in proteotoxicity and cell death. Recently, we found that the eIF2α kinase heme-regulated inhibitory (HRI) induced a cytosolic unfolded protein response to prevent aggregation of innate immune signalosomes, but whether HRI acts as a general sensor of proteotoxicity in the cytosol remains unclear. Here we show that HRI controls autophagy to clear cytosolic protein aggregates when the ubiquitin-proteasome system is inhibited. We further report that silencing the expression of HRI resulted in decreased levels of BAG3 and HSPB8, two proteins involved in chaperone-assisted selective autophagy, suggesting that HRI may control proteostasis in the cytosol at least in part through chaperone-assisted selective autophagy. Moreover, knocking down the expression of HRI resulted in cytotoxic accumulation of overexpressed α-synuclein, a protein known to aggregate in Parkinson's disease, dementia with Lewy bodies, and multiple system atrophy. In agreement with these data, protein aggregate accumulation and microglia activation were observed in the spinal cord white matter of 7-month-old Hri-/- mice as compared with Hri+/+ littermates. Moreover, aged Hri-/- mice showed accumulation of misfolded α-synuclein in the lateral collateral pathway, a region of the sacral spinal cord horn that receives visceral sensory afferents from the bladder and distal colon, a pathological feature common to α-synucleinopathies in humans. Together, these results suggest that HRI contributes to a general cytosolic unfolded protein response that could be leveraged to bolster the clearance of cytotoxic protein aggregates.
    Keywords:  BAG3–HSPB8 complex; CASA; HRI; Parkinson’s disease; alpha-synuclein; autophagy; integrated stress response; proteasome; protein aggregation; protein misfolding
  2. J Nanobiotechnology. 2021 Jan 06. 19(1): 6
      BACKGROUND: Mitochondrial dysfunction is a critical factor in the onset and progression of neurodegenerative diseases. Recently, mitochondrial transplantation has been advised as an innovative and attractive strategy to transfer and replace damaged mitochondria. Here we propose, for the first time, to use rat brain extracted synaptosomes, a subcellular fraction of isolated synaptic terminal that contains mitochondria, as mitochondrial delivery systems.RESULTS: Synaptosome preparation was validated by the presence of Synaptophysin and PSD95. Synaptosomes were characterized in terms of dimension, zeta potential, polydispersity index and number of particles/ml. Nile Red or CTX-FITCH labeled synaptosomes were internalized in LAN5 recipient cells by a mechanism involving specific protein-protein interaction, as demonstrated by loss of fusion ability after trypsin treatment and using different cell lines. The loading and release ability of the synaptosomes was proved by the presence of curcumin both into synaptosomes and LAN5 cells. The vitality of mitochondria transferred by Synaptosomes was demonstrated by the presence of Opa1, Fis1 and TOM40 mitochondrial proteins and JC-1 measurements. Further, synaptosomes deliver vital mitochondria into the cytoplasm of neuronal cells as demonstrated by microscopic images, increase of TOM 40, cytochrome c, Hexokinase II mitochondrial proteins, and presence of rat mitochondrial DNA. Finally, by using synaptosomes as a vehicle, healthy mitochondria restored mitochondrial function in cells containing rotenone or CCCp damaged mitochondria.
    CONCLUSIONS: Taken together these results suggest that synaptosomes can be a natural vehicle for the delivery of molecules and organelles to neuronal cells. Further, the replacement of affected mitochondria with healthy ones could be a potential therapy for treating neuronal mitochondrial dysfunction-related diseases.
    Keywords:  Delivery system; Mitochondria; Mitochondrial transplantation; Neurodegeneration; Synaptosomes
  3. Ageing Res Rev. 2020 Dec 31. pii: S1568-1637(20)30385-8. [Epub ahead of print]66 101250
      Myocardial infarction (MI) is the irreversible death of cardiomyocyte secondary to prolonged lack of oxygen or fresh blood supply. Historically considered as merely cardiomyocyte powerhouse that manufactures ATP and other metabolites, mitochondrion is recently being identified as a signal regulator that is implicated in the crosstalk and signal integration of cardiomyocyte contraction, metabolism, inflammation, and death. Mitochondria quality surveillance is an integrated network system modifying mitochondrial structure and function through the coordination of various processes including mitochondrial fission, fusion, biogenesis, bioenergetics, proteostasis, and degradation via mitophagy. Mitochondrial fission favors the elimination of depolarized mitochondria through mitophagy, whereas mitochondrial fusion preserves the mitochondrial network upon stress through integration of two or more small mitochondria into an interconnected phenotype. Mitochondrial biogenesis represents a regenerative program to replace old and damaged mitochondria with new and healthy ones. Mitochondrial bioenergetics is regulated by a metabolic switch between glucose and fatty acid usage, depending on oxygen availability. To maintain the diversity and function of mitochondrial proteins, a specialized protein quality control machinery regulates protein dynamics and function through the activity of chaperones and proteases, and induction of the mitochondrial unfolded protein response. In this review, we provide an overview of the molecular mechanisms governing mitochondrial quality surveillance and highlight the most recent preclinical and clinical therapeutic approaches to restore mitochondrial fitness during both MI and post-MI heart failure.
    Keywords:  Fission; Fusion; Mitochondrial quality surveillance; Mitophagy; Myocardial infarction; Proteostasis
  4. Autophagy. 2021 Jan 08.
      Mitochondrial quality control (MQC) balances organelle adaptation and elimination, and mechanistic crosstalk between the underlying molecular processes affects subsequent stress outcomes. FUNDC1 (FUN14 domain containing 1) is a mammalian mitophagy receptor that responds to hypoxia-reoxygenation (HR) stress. Here, we provide evidence that FNDC-1 is the C. elegans ortholog of FUNDC1, and that its loss protects against injury in a worm model of HR. This protection depends upon ATFS-1, a transcription factor that is central to the mitochondrial unfolded protein response (UPRmt). Global mRNA and metabolite profiling suggest that atfs-1-dependent stress responses and metabolic remodeling occur in response to the loss of fndc-1. These data support a role for FNDC-1 in non-hypoxic MQC, and further suggest that these changes are prophylactic in relation to subsequent HR. Our results highlight functional coordination between mitochondrial adaptation and elimination that organizes stress responses and metabolic rewiring to protect against HR injury.
    Keywords:   C. elegans ; hypoxia-reoxygenation (HR); metabolism; mitochondrial unfolded protein response (UPRmt); mitophagy
  5. Redox Biol. 2020 Dec 28. pii: S2213-2317(20)31057-0. [Epub ahead of print]40 101852
      In this paper, we describe an assay to analyze simultaneously the oxygen consumption rate (OCR) and superoxide production in a biological system. The analytical set-up uses electron paramagnetic resonance (EPR) spectroscopy with two different isotopically-labelled sensors: 15N-PDT (4-oxo-2,2,6,6-tetramethylpiperidine-d16-15N-1-oxyl) as oxygen-sensing probe and 14N-CMH (1-hydroxy-3-methoxycarbonyl-2,2,5,5-tetramethylpyrrolidine, a cyclic hydroxylamine, as sensor of reactive oxygen species (ROS). The superoxide contribution to CMH oxidation is assessed using SOD or PEGSOD as controls. Because the EPR spectra are not superimposable, the variation of EPR linewidth of 15N-PDT (linked to OCR) and the formation of the nitroxide from 14N-CMH (linked to superoxide production) can be recorded simultaneously over time on a single preparation. The EPR toolbox was qualified in biological systems of increasing complexity. First, we used an enzymatic assay based on the hypoxanthine (HX)/xanthine oxidase (XO) which is a well described model of oxygen consumption and superoxide production. Second, we used a cellular model of superoxide production using macrophages exposed to phorbol 12-myristate 13-acetate (PMA) which stimulates the NADPH oxidase (NOX) to consume oxygen and produce superoxide. Finally, we exposed isolated mitochondria to established inhibitors of the electron transport chain (rotenone and metformin) in order to assess their impact on OCR and superoxide production. This EPR toolbox has the potential to screen the effect of intoxicants or drugs targeting the mitochondrial function.
    Keywords:  EPR; ESR; ETC; Hydroxylamine; Mitochondria; Nitroxide; Oxygen consumption rate (OCR); Superoxide
  6. Mitochondrion. 2021 Jan 04. pii: S1567-7249(20)30242-7. [Epub ahead of print]
      The puzzling traits related to the evolutionary aspect of mitochondria, still positions the mitochondrion at the center of the research. The theory of endosymbiosis popularized by Lynn Margulis in 1967 gained prominence wherein the mitochondrion is believed to have emerged as a prokaryote and later integrated into the eukaryotic system. This semi-autonomous organelle has bagged two responsible but perilous cellular functions: a) energy metabolism, and b) calcium buffering, though both are interdependent. While most of the mitochondrial functions are saliently regulated by calcium ions, the calcium buffering role of mitochondria decides the cellular fate. Though calcium overload in few mitochondria makes them dysfunctional at the early stage of cellular stress, this doesn't lead to sudden cell death due to critical checkpoints like mitophagy, mitochondrial fusion, etc. Thus, mitochondrion juggles with multiple crucial cellular functions with its calcium buffering skill.
    Keywords:  Cellular homeostasis; MCU, apoptosis; mitochondrial calcium
  7. Cell Metab. 2020 Dec 24. pii: S1550-4131(20)30661-6. [Epub ahead of print]
      Platelets are known to enhance the wound-healing activity of mesenchymal stem cells (MSCs). However, the mechanism by which platelets improve the therapeutic potential of MSCs has not been elucidated. Here, we provide evidence that, upon their activation, platelets transfer respiratory-competent mitochondria to MSCs primarily via dynamin-dependent clathrin-mediated endocytosis. We found that this process enhances the therapeutic efficacy of MSCs following their engraftment in several mouse models of tissue injury, including full-thickness cutaneous wound and dystrophic skeletal muscle. By combining in vitro and in vivo experiments, we demonstrate that platelet-derived mitochondria promote the pro-angiogenic activity of MSCs via their metabolic remodeling. Notably, we show that activation of the de novo fatty acid synthesis pathway is required for increased secretion of pro-angiogenic factors by platelet-preconditioned MSCs. These results reveal a new mechanism by which platelets potentiate MSC properties and underline the importance of testing platelet mitochondria quality prior to their clinical use.
    Keywords:  angiogenesis; cell therapy; citrate; de novo; fatty acid synthesis; intercellular mitochondria transfer; mesenchymal stem cells; metabolism reprogramming; mitochondria; mitochondrial respiration; platelets
  8. Haematologica. 2021 Jan 07.
      Erythroblast maturation in mammals is dependent on organelle clearance throughout terminal erythropoiesis. We studied the role of the outer mitochondrial membrane protein VDAC1 (Voltage-Dependent Anion Channel-1) in human terminal erythropoiesis. We show that shRNA-mediated downregulation of VDAC1 accelerates erythroblast maturation. Thereafter, erythroblasts are blocked at orthochromatic stage, exhibiting a significant decreased level of enucleation, concomitant with an increased cell death. We demonstrate that mitochondria clearance starts at the transition from basophilic to polychromatic erythroblast, and that VDAC1 downregulation induces the mitochondrial retention. In damaged mitochondria from non-erythroid cells, VDAC1 was identified as a target for Parkin-mediated ubiquitination to recruit the phagophore. Here, we showed that VDAC1 is involved in phagophore's membrane recruitment regulating selective mitophagy of still functional mitochondria from human erythroblasts. These findings demonstrate for the first time a crucial role for VDAC1 in human erythroblast terminal differentiation, regulating mitochondria clearance.
  9. Brain Res Bull. 2021 Jan 02. pii: S0361-9230(20)30739-5. [Epub ahead of print]
      Autism spectrum disorders (ASDs) are a group of complex neurodevelopmental disorders, including autistic disorder, Asperger's syndrome, pervasive developmental disorder and childhood disintegrative disorder. Mitochondria not only provide neurons with energy in the form of ATP to sustain neuron growth, proliferation and neurodevelopment, but also regulate neuron apoptosis, intracellular calcium ion (Ca2+) homeostasis, and reactive oxygen species (ROS) clearance. Due to their postmitotic state and high energy-demanded feature, neurons are particularly prone to mitophagy and mitochondrial disfunction. Mitophagy, a selective autophagy, is critical for sustaining mitochondrial turnover and quality control via eliminating unwanted and dysfunctional mitochondria in neurons. Dysfunctional mitochondria and dysregulated mitophagy have been closely associated with the onset of ASDs. In this review, we summarize the mechanism of mitophagy and its role in neurons, and the consequence of mitophagy dysfunction in ASDs. Deeper appreciation of the role of mitophagy in ASDs pathology is required for developing new therapeutic approaches.
    Keywords:  Autism spectrum disorders; Mitochondrial homeostasis; Mitophagy; Neurodevelopmental disease
  10. Mitochondrion. 2021 Jan 04. pii: S1567-7249(20)30244-0. [Epub ahead of print]
      Endothelial mitochondria play important signaling roles critical for the regulation of various cellular processes, including calcium signaling, ROS generation, NO synthesis or inflammatory response. Mitochondrial stress or disturbances in mitochondrial function may participate in the development and/or progression of endothelial dysfunction and could precede vascular diseases. Vascular functions are also strictly regulated by properly functioning degradation machinery, including autophagy and mitophagy, and tightly coordinated by mitochondrial and endoplasmic reticulum responses to stress. Within this review, current knowledge related to the development of cardiovascular disorders and the importance of mitochondria, endoplasmic reticulum and degradation mechanisms in vascular endothelial functions are summarized.
    Keywords:  ER stress; autophagy; endothelium; inflammation; mitochondria; mitochondrial stress; mitophagy
  11. Int J Mol Sci. 2020 Dec 31. pii: E363. [Epub ahead of print]22(1):
      The effects of airway inflammation on airway smooth muscle (ASM) are mediated by pro-inflammatory cytokines such as tumor necrosis factor alpha (TNFα). In this review article, we will provide a unifying hypothesis for a homeostatic response to airway inflammation that mitigates oxidative stress and thereby provides resilience to ASM. Previous studies have shown that acute exposure to TNFα increases ASM force generation in response to muscarinic stimulation (hyper-reactivity) resulting in increased ATP consumption and increased tension cost. To meet this increased energetic demand, mitochondrial O2 consumption and oxidative phosphorylation increases but at the cost of increased reactive oxygen species (ROS) production (oxidative stress). TNFα-induced oxidative stress results in the accumulation of unfolded proteins in the endoplasmic reticulum (ER) and mitochondria of ASM. In the ER, TNFα selectively phosphorylates inositol-requiring enzyme 1 alpha (pIRE1α) triggering downstream splicing of the transcription factor X-box binding protein 1 (XBP1s); thus, activating the pIRE1α/XBP1s ER stress pathway. Protein unfolding in mitochondria also triggers an unfolded protein response (mtUPR). In our conceptual framework, we hypothesize that activation of these pathways is homeostatically directed towards mitochondrial remodeling via an increase in peroxisome proliferator-activated receptor-gamma coactivator 1 alpha (PGC1α) expression, which in turn triggers: (1) mitochondrial fragmentation (increased dynamin-related protein-1 (Drp1) and reduced mitofusin-2 (Mfn2) expression) and mitophagy (activation of the Phosphatase and tensin homolog (PTEN)-induced putative kinase 1 (PINK1)/Parkin mitophagy pathway) to improve mitochondrial quality; (2) reduced Mfn2 also results in a disruption of mitochondrial tethering to the ER and reduced mitochondrial Ca2+ influx; and (3) mitochondrial biogenesis and increased mitochondrial volume density. The homeostatic remodeling of mitochondria results in more efficient O2 consumption and oxidative phosphorylation and reduced ROS formation by individual mitochondrion, while still meeting the increased ATP demand. Thus, the energetic load of hyper-reactivity is shared across the mitochondrial pool within ASM cells.
    Keywords:  Drp1; MCU; Mfn2; asthma; oxidative stress; reactive oxygen species (ROS)
  12. Methods Mol Biol. 2021 ;2261 411-419
      In-depth analysis of the mitochondrial proteome can be greatly improved by analyzing isolated mitochondria instead of whole cells. However, isolation of sufficient amounts of mitochondria from cell culture has proven to be notoriously difficult due to small sample size. Thus, we have developed a reproducible, controllable, and highly customizable method to isolate high microgram to low milligram amounts of intact mitochondria from cell culture samples along with an optional density gradient purification. This chapter provides a methodological update of our approach and underlines the excellent quality and coverage of the mitochondrial proteome of crude and purified mitochondria from cultured liver cancer cell lines.
    Keywords:  Balch homogenizer; Cell culture; Mitochondria; Proteomics
  13. Crit Care. 2021 Jan 07. 25(1): 20
      BACKGROUND: Sepsis has a high mortality rate, but no specific drug has been proven effective, prompting the development of new drugs. Immunologically, sepsis can involve hyperinflammation, immune paralysis, or both, which might pose challenges during drug development. Recently, mitochondrial transplantation has emerged as a treatment modality for various diseases involving mitochondrial dysfunction, but it has never been tested for sepsis.METHODS: We isolated mitochondria from L6 muscle cells and umbilical cord mesenchymal stem cells and tested the quality of the isolated mitochondria. We conducted both in vivo and in vitro sepsis studies. We investigated the effects of intravenous mitochondrial transplantation on cecal slurry model in rats in terms of survival rate, bacterial clearance rate, and the immune response. Furthermore, we observed the effects of mitochondrial transplantation on the immune reaction regarding both hyperinflammation and immune paralysis. To do this, we studied early- and late-phase cytokine production in spleens from cecal slurry model in rats. We also used a lipopolysaccharide (LPS)-stimulated human PBMC monocyte model to confirm the immunological effects of mitochondrial transplantation. Apoptosis and the intrinsic apoptotic pathway were investigated in septic spleens.
    RESULTS: Mitochondrial transplantation improved survival and bacterial clearance. It also mitigated mitochondrial dysfunction and apoptosis in septic spleens and attenuated both hyperinflammation and immune paralysis in the spleens of cecal slurry model in rats. This effect was confirmed with an LPS-stimulated human PBMC study.
    CONCLUSIONS: In rat polymicrobial cecal slurry model, the outcome is improved by mitochondrial transplantation, which might have an immunomodulatory effect.
    Keywords:  Hyperinflammation; Immune modulation; Immune paralysis; Mitochondria dysfunction; Mitochondria transplantation; Sepsis
  14. Cell Biosci. 2021 Jan 07. 11(1): 9
      BACKGROUND: Acute liver failure (ALF) is associated with a high mortality rate, and there are still no effective treatments except liver transplantation and artificial liver therapies. This study aimed to determine the effects, therapeutic window and mechanisms of selonsertib, a selective inhibitor of ASK1, for ALF therapy.RESULTS: Lipopolysaccharide and D-galactosamine (LPS/GalN) were used to simulate ALF. We found that selonsertib pretreatment significantly ameliorated ALF, as determined by reduced hepatic necrosis and serum alanine aminotransferase, aspartate aminotransferase and inflammatory cytokine levels. However, selonsertib is only effective early after LPS/GalN administration, and the limited therapeutic window is related to the activation and mitochondrial translocation of JNK and DRP1. Further experiments revealed that selonsertib could alleviate LPS-induced mitochondrial damage in macrophages by evaluating the mitochondrial membrane potential and mitochondrial permeability transition pore opening in macrophages. Selonsertib also suppressed the release of inflammatory cytokines from macrophages by reducing DRP1-mediated mitochondrial dysfunction, which was confirmed by using mdivi, a specific DRP1 inhibitor.
    CONCLUSIONS: Selonsertib protected against LPS/GalN-induced ALF by attenuating JNK-mediated DRP1 mitochondrial translocation and then rescuing mitochondrial damage in macrophages and may have therapeutic potential for early ALF patients.
    Keywords:  ASK1; Acute liver failure; DRP1; Macrophage; Mitochondria
  15. Trends Cell Biol. 2021 Jan 05. pii: S0962-8924(20)30254-3. [Epub ahead of print]
      Mitoribosomes catalyze essential protein synthesis within mitochondria. Mitoribosome biogenesis is assisted by an increasing number of assembly factors, among which guanosine triphosphate hydrolases (GTPases) are the most abundant class. Here, we review recent progress in our understanding of mitoribosome assembly GTPases. We describe their shared and specific features and mechanisms of action, compare them with their bacterial counterparts, and discuss their possible roles in the assembly of small or large mitoribosomal subunits and the formation of the monosome by establishing quality-control checkpoints during these processes. Furthermore, following the recent unification of the nomenclature for the mitoribosomal proteins, we also propose a unified nomenclature for mitoribosome assembly GTPases.
    Keywords:  GTPBP; OXPHOS deficiency; mitochondrial diseases; mitochondrial ribosome; mitoribosome assembly GTPase; quality control of mitoribosome maturation
  16. Sci Rep. 2021 Jan 08. 11(1): 3
      SARS-CoV-2 induces a muted innate immune response compared to other respiratory viruses. Mitochondrial dynamics might partially mediate this effect of SARS-CoV-2 on innate immunity. Polypeptides encoded by open reading frames of SARS-CoV and SARS-CoV-2 have been shown to localize to mitochondria and disrupt Mitochondrial Antiviral Signaling (MAVS) protein signaling. Therefore, we hypothesized that SARS-CoV-2 would distinctly regulate the mitochondrial transcriptome. We analyzed multiple publicly available RNASeq data derived from primary cells, cell lines, and clinical samples (i.e., BALF and lung). We report that SARS-CoV-2 did not dramatically regulate (1) mtDNA-encoded gene expression or (2) MAVS expression, and (3) SARS-CoV-2 downregulated nuclear-encoded mitochondrial (NEM) genes related to cellular respiration and Complex I.
  17. Autophagy. 2021 Jan 06.
      Retinal ganglion cell axons are heavily myelinated (98%) and myelin damage in the optic nerve (ON) severely affects vision. Understanding the molecular mechanism of oligodendrocyte progenitor cell (OPC) differentiation into mature oligodendrocytes will be essential for developing new therapeutic approaches for ON demyelinating diseases. To this end, we developed a new method for isolation and culture of ON-derived oligodendrocyte lineage cells and used it to study OPC differentiation. A critical aspect of cellular differentiation is macroautophagy/autophagy, a catabolic process that allows for cell remodeling by degradation of excess or damaged cellular molecules and organelles. Knockdown of ATG9A and BECN1 (pro-autophagic proteins involved in the early stages of autophagosome formation) led to a significant reduction in proliferation and survival of OPCs. We also found that autophagy flux (a measure of autophagic degradation activity) is significantly increased during progression of oligodendrocyte differentiation. Additionally, we demonstrate a significant change in mitochondrial dynamics during oligodendrocyte differentiation, which is associated with a significant increase in programmed mitophagy (selective autophagic clearance of mitochondria). This process is mediated by the mitophagy receptor BNIP3L (BCL2/adenovirus E1B interacting protein 3-like). BNIP3L-mediated mitophagy plays a crucial role in the regulation of mitochondrial network formation, mitochondrial function and the viability of newly differentiated oligodendrocytes. Our studies provide novel evidence that proper mitochondrial dynamics is required for establishment of functional mitochondria in mature oligodendrocytes. These findings are significant because targeting BNIP3L-mediated programmed mitophagy may provide a novel therapeutic approach for stimulating myelin repair in ON demyelinating diseases.
    Keywords:  ATG9A; autophagy; autophagy flux; co-culture; demyelinating diseases; glial cells; mitochondrial dynamics; myelin; oligodendrocyte lineage cells; retinal ganglion cell axons
  18. Mitochondrion. 2021 Jan 04. pii: S1567-7249(20)30247-6. [Epub ahead of print]
      Protein phosphorylation is one of the best-known post-translational modifications occurring in all domains of life. In eukaryotes, protein phosphorylation affects all cellular compartments including mitochondria. High-throughput techniques of mass spectrometry combined with cell fractionation and biochemical methods yielded thousands of phospho-sites on hundreds of mitochondrial proteins. We have compiled the information on mitochondrial protein kinases and phosphatases and their substrates in Saccharomyces cerevisiae and provide the current state-of-the-art overview of mitochondrial protein phosphorylation in this model eukaryote. Using several examples, we describe emerging features of the yeast mitochondrial phosphoproteome and present challenges lying ahead in this exciting field.
    Keywords:  mitochondrial phosphoproteome; post-translational modifications; protein kinase; protein phosphatase; protein phosphorylation; yeast