bims-mikwok Biomed News
on Mitochondrial quality control
Issue of 2021‒02‒14
thirty-nine papers selected by
Avinash N. Mukkala
University of Toronto


  1. EMBO Rep. 2021 Feb 08. e50629
    Liu L, Li Y, Wang J, Zhang D, Wu H, Li W, Wei H, Ta N, Fan Y, Liu Y, Wang X, Wang J, Pan X, Liao X, Zhu Y, Chen Q.
      Mitophagy is an essential cellular autophagic process that selectively removes superfluous and damaged mitochondria, and it is coordinated with mitochondrial biogenesis to fine tune the quantity and quality of mitochondria. Coordination between these two opposing processes to maintain the functional mitochondrial network is of paramount importance for normal cellular and organismal metabolism. However, the underlying mechanism is not completely understood. Here we report that PGC-1α and nuclear respiratory factor 1 (NRF1), master regulators of mitochondrial biogenesis and metabolic adaptation, also transcriptionally upregulate the gene encoding FUNDC1, a previously characterized mitophagy receptor, in response to cold stress in brown fat tissue. NRF1 binds to the classic consensus site in the promoter of Fundc1 to upregulate its expression and to enhance mitophagy through its interaction with LC3. Specific knockout of Fundc1 in BAT results in reduced mitochondrial turnover and accumulation of functionally compromised mitochondria, leading to impaired adaptive thermogenesis. Our results demonstrate that FUNDC1-dependent mitophagy is directly coupled with mitochondrial biogenesis through the PGC-1α/NRF1 pathway, which dictates mitochondrial quantity, quality, and turnover and contributes to adaptive thermogenesis.
    Keywords:  adaptive thermogenesis; brown adipose tissue; mitochondrial biogenesis; mitophagy
    DOI:  https://doi.org/10.15252/embr.202050629
  2. Rev Neurosci. 2021 Feb 23. 32(2): 203-217
    Espino De la Fuente-Muñoz C, Arias C.
      Mitochondrial activity is essential to support neural functions, and changes in the integrity and activity of the mitochondria can contribute to synaptic damage and neuronal death, especially in degenerative diseases associated with age, such as Alzheimer's and Parkinson's disease. Currently, different approaches are used to treat these conditions, and one strategy under research is mitochondrial transplantation. For years, mitochondria have been shown to be transferred between cells of different tissues. This process has allowed several attempts to develop transplantation schemes by isolating functional mitochondria and introducing them into damaged tissue in particular to counteract the harmful effects of myocardial ischemia. Recently, mitochondrial transfer between brain cells has also been reported, and thus, mitochondrial transplantation for disorders of the nervous system has begun to be investigated. In this review, we focus on the relevance of mitochondria in the nervous system, as well as some mitochondrial alterations that occur in neurodegenerative diseases associated with age. In addition, we describe studies that have performed mitochondrial transplantation in various tissues, and we emphasize the advances in mitochondrial transplantation aimed at treating diseases of the nervous system.
    Keywords:  bioenergetic restoration; mitochondrial transplant; neurodegenerative diseases; neuronal survival
    DOI:  https://doi.org/10.1515/revneuro-2020-0068
  3. Redox Biol. 2021 Jan 30. pii: S2213-2317(21)00032-X. [Epub ahead of print]41 101884
    De Lazzari F, Prag HA, Gruszczyk AV, Whitworth AJ, Bisaglia M.
      DJ-1 is a multifaceted protein with pleiotropic functions that has been implicated in multiple diseases, ranging from neurodegeneration to cancer and ischemia-reperfusion injury. Ischemia is a complex pathological state arising when tissues and organs do not receive adequate levels of oxygen and nutrients. When the blood flow is restored, significant damage occurs over and above that of ischemia alone and is termed ischemia-reperfusion injury. Despite great efforts in the scientific community to ameliorate this pathology, its complex nature has rendered it challenging to obtain satisfactory treatments that translate to the clinic. In this review, we will describe the recent findings on the participation of the protein DJ-1 in the pathophysiology of ischemia-reperfusion injury, firstly introducing the features and functions of DJ-1 and, successively highlighting the therapeutic potential of the protein.
    Keywords:  DJ-1; Glucose homeostasis; Ionic balance; Ischemia-reperfusion injury; Mitochondrial homeostasis; Reactive oxygen species
    DOI:  https://doi.org/10.1016/j.redox.2021.101884
  4. EMBO Rep. 2021 Feb 10. e49097
    Shiiba I, Takeda K, Nagashima S, Ito N, Tokuyama T, Yamashita SI, Kanki T, Komatsu T, Urano Y, Fujikawa Y, Inatome R, Yanagi S.
      Parkin promotes cell survival by removing damaged mitochondria via mitophagy. However, although some studies have suggested that Parkin induces cell death, the regulatory mechanism underlying the dual role of Parkin remains unknown. Herein, we report that mitochondrial ubiquitin ligase (MITOL/MARCH5) regulates Parkin-mediated cell death through the FKBP38-dependent dynamic translocation from the mitochondria to the ER during mitophagy. Mechanistically, MITOL mediates ubiquitination of Parkin at lysine 220 residue, which promotes its proteasomal degradation, and thereby fine-tunes mitophagy by controlling the quantity of Parkin. Deletion of MITOL leads to accumulation of the phosphorylated active form of Parkin in the ER, resulting in FKBP38 degradation and enhanced cell death. Thus, we have shown that MITOL blocks Parkin-induced cell death, at least partially, by protecting FKBP38 from Parkin. Our findings unveil the regulation of the dual function of Parkin and provide a novel perspective on the pathogenesis of PD.
    Keywords:  E3 ubiquitin ligase; MITOL/MARCH5; Parkin; mitochondria; mitophagy
    DOI:  https://doi.org/10.15252/embr.201949097
  5. Arch Biochem Biophys. 2021 Feb 04. pii: S0003-9861(21)00040-0. [Epub ahead of print]700 108790
    Crivellari I, Pecorelli A, Cordone V, Marchi S, Pinton P, Hayek J, Cervellati C, Valacchi G.
      Rett Syndrome (RTT) is a rare neurodevelopmental disorder caused in the 95% of cases by mutations in the X-linked MECP2 gene, affecting almost exclusively females. While the genetic basis of RTT is known, the exact pathogenic mechanisms that lead to the broad spectrum of symptoms still remain enigmatic. Alterations in the redox homeostasis have been proposed among the contributing factors to the development and progression of the syndrome. Mitochondria appears to play a central role in RTT oxidative damage and a plethora of mitochondrial defects has already been recognized. However, mitochondrial dynamics and mitophagy, which represent critical pathways in regulating mitochondrial quality control (QC), have not yet been investigated in RTT. The present work showed that RTT fibroblasts have networks of hyperfused mitochondria with morphological abnormalities and increased mitochondrial volume. Moreover, analysis of mitophagic flux revealed an impaired PINK1/Parkin-mediated mitochondrial removal associated with an increase of mitochondrial fusion proteins Mitofusins 1 and 2 (MFN1 and 2) and a decrease of fission mediators including Dynamin related protein 1 (DRP1) and Mitochondrial fission 1 protein (FIS1). Finally, challenging RTT fibroblasts with FCCP and 2,4-DNP did not trigger a proper apoptotic cell death due to a defective caspase 3/7 activation. Altogether, our findings shed light on new aspects of mitochondrial dysfunction in RTT that are represented by defective mitochondrial QC pathways, also providing new potential targets for a therapeutic intervention aimed at slowing down clinical course and manifestations in the affected patients.
    Keywords:  Apoptosis; Mitochondrial quality control; Mitophagy; Oxidative stress; RTT Syndrome
    DOI:  https://doi.org/10.1016/j.abb.2021.108790
  6. Biomedicines. 2021 Feb 09. pii: 169. [Epub ahead of print]9(2):
    Carinci M, Vezzani B, Patergnani S, Ludewig P, Lessmann K, Magnus T, Casetta I, Pugliatti M, Pinton P, Giorgi C.
      Mitochondrial dysfunctions are among the main hallmarks of several brain diseases, including ischemic stroke. An insufficient supply of oxygen and glucose in brain cells, primarily neurons, triggers a cascade of events in which mitochondria are the leading characters. Mitochondrial calcium overload, reactive oxygen species (ROS) overproduction, mitochondrial permeability transition pore (mPTP) opening, and damage-associated molecular pattern (DAMP) release place mitochondria in the center of an intricate series of chance interactions. Depending on the degree to which mitochondria are affected, they promote different pathways, ranging from inflammatory response pathways to cell death pathways. In this review, we will explore the principal mitochondrial molecular mechanisms compromised during ischemic and reperfusion injury, and we will delineate potential neuroprotective strategies targeting mitochondrial dysfunction and mitochondrial homeostasis.
    Keywords:  cell death; inflammation; ischemic reperfusion; ischemic stroke; mitochondrial fission; mitochondrial fusion; mitochondrial transfer; mitophagy
    DOI:  https://doi.org/10.3390/biomedicines9020169
  7. Genes (Basel). 2021 Feb 10. pii: 247. [Epub ahead of print]12(2):
    Di Nottia M, Verrigni D, Torraco A, Rizza T, Bertini E, Carrozzo R.
      Mitochondria do not exist as individual entities in the cell-conversely, they constitute an interconnected community governed by the constant and opposite process of fission and fusion. The mitochondrial fission leads to the formation of smaller mitochondria, promoting the biogenesis of new organelles. On the other hand, following the fusion process, mitochondria appear as longer and interconnected tubules, which enhance the communication with other organelles. Both fission and fusion are carried out by a small number of highly conserved guanosine triphosphatase proteins and their interactors. Disruption of this equilibrium has been associated with several pathological conditions, ranging from cancer to neurodegeneration, and mutations in genes involved in mitochondrial fission and fusion have been reported to be the cause of a subset of neurogenetic disorders.
    Keywords:  mitochondrial diseases; mitochondrial dynamics; therapeutic approaches
    DOI:  https://doi.org/10.3390/genes12020247
  8. Cell Calcium. 2021 Jan 30. pii: S0143-4160(21)00011-7. [Epub ahead of print]94 102357
    De Mario A, Gherardi G, Rizzuto R, Mammucari C.
      Mitochondrial activity warrants energy supply to oxidative myofibres to sustain endurance workload. The maintenance of mitochondrial homeostasis is ensured by the control of fission and fusion processes and by the mitophagic removal of aberrant organelles. Many diseases are due to or characterized by dysfunctional mitochondria, and altered mitochondrial dynamics or turnover trigger myopathy per se. In this review, we will tackle the role of mitochondrial dynamics, turnover and metabolism in skeletal muscle, both in health and disease.
    Keywords:  Fusion and fission machinery; Mitochondrial calcium uptake; Mitochondrial myopathies; Mitophagy; Skeletal muscle
    DOI:  https://doi.org/10.1016/j.ceca.2021.102357
  9. Autophagy. 2021 Feb 11.
    Terešak P, Lapao A, Subic N, Boya P, Elazar Z, Simonsen A.
      Mitochondria are dynamic, multifunctional cellular organelles that play a fundamental role in maintaining cellular homeostasis. Keeping the quality of mitochondria in check is of essential importance for functioning and survival of the cells. Selective autophagic clearance of flawed mitochondria, a process termed mitophagy, is one of the most prominent mechanisms through which cells maintain a healthy mitochondrial pool. The best-studied pathway through which mitophagy is exerted is the PINK1-PRKN pathway. However, an increasing number of studies have shown an existence of alternative pathways, where different proteins and lipids are able to recruit autophagic machinery independently of PINK1 and PRKN. The significance of PRKN-independent mitophagy pathways is reflected in various physiological and pathophysiological processes, but many questions regarding the regulation and the interplay between these pathways remain open. Here we review the current knowledge and recent progress made in the field of PRKN-independent mitophagy. Particularly we focus on the regulation of various receptors that participate in targeting impaired mitochondria to autophagosomes independently of PRKN.
    Keywords:  autophagy receptors; mitochondria; mitochondrial dysfunction; mitophagy; selective autophagy
    DOI:  https://doi.org/10.1080/15548627.2021.1888244
  10. Cells. 2021 Feb 10. pii: 369. [Epub ahead of print]10(2):
    Čunátová K, Reguera DP, Vrbacký M, Fernández-Vizarra E, Ding S, Fearnley IM, Zeviani M, Houštěk J, Mráček T, Pecina P.
      The oxidative phosphorylation (OXPHOS) system localized in the inner mitochondrial membrane secures production of the majority of ATP in mammalian organisms. Individual OXPHOS complexes form supramolecular assemblies termed supercomplexes. The complexes are linked not only by their function but also by interdependency of individual complex biogenesis or maintenance. For instance, cytochrome c oxidase (cIV) or cytochrome bc1 complex (cIII) deficiencies affect the level of fully assembled NADH dehydrogenase (cI) in monomeric as well as supercomplex forms. It was hypothesized that cI is affected at the level of enzyme assembly as well as at the level of cI stability and maintenance. However, the true nature of interdependency between cI and cIV is not fully understood yet. We used a HEK293 cellular model where the COX4 subunit was completely knocked out, serving as an ideal system to study interdependency of cI and cIV, as early phases of cIV assembly process were disrupted. Total absence of cIV was accompanied by profound deficiency of cI, documented by decrease in the levels of cI subunits and significantly reduced amount of assembled cI. Supercomplexes assembled from cI, cIII, and cIV were missing in COX4I1 knock-out (KO) due to loss of cIV and decrease in cI amount. Pulse-chase metabolic labeling of mitochondrial DNA (mtDNA)-encoded proteins uncovered a decrease in the translation of cIV and cI subunits. Moreover, partial impairment of mitochondrial protein synthesis correlated with decreased content of mitochondrial ribosomal proteins. In addition, complexome profiling revealed accumulation of cI assembly intermediates, indicating that cI biogenesis, rather than stability, was affected. We propose that attenuation of mitochondrial protein synthesis caused by cIV deficiency represents one of the mechanisms, which may impair biogenesis of cI.
    Keywords:  COX; COX4; OXPHOS; biogenesis interdependency; cI; cIV; cIV assembly; complex I; complexome profiling; knock-out; mitochondria; mitochondrial protein synthesis
    DOI:  https://doi.org/10.3390/cells10020369
  11. Mol Cell Oncol. 2021 ;8(1): 1839341
    Silva-Pavez E, Ahumada-Castro U, Lovy A, Cárdenas JC.
      The inositol 1,4,5-triphosphate receptor (InsP3R)-mediated calcium (Ca2+) transfer to mitochondria is important to maintain mitochondrial respiration and bioenergetics in normal and cancer cells, even though cancer cells have defective oxidative phosphorylation (OXPHOS). Here, we discuss how tumor mitochondria could become a feasible therapeutic target to treat tumors that depend on reductive carboxylation.
    Keywords:  Calcium; OXPHOS; autophagy; cancer; cell survival
    DOI:  https://doi.org/10.1080/23723556.2020.1839341
  12. Cell Commun Signal. 2021 Feb 11. 19(1): 14
    Dong L, Li W, Lin T, Liu B, Hong Y, Zhang X, Li X.
      BACKGROUND: Abnormal neovascularization is the most common cause of blindness, and hypoxia alters tissue metabolism, function, and morphology. HIF-1α, the transcriptional activator of VEGF, has intricate mechanisms of nuclear translocation and activation, but its signal termination mechanisms remain unclear.METHODS: We investigated the role of polypyrimidine tract-binding protein-associated splicing factor (PSF) in cellular energy production, migration, and proliferation by targeting HIF-1α in vivo and in vitro PSF plasmids were transfected with liposome 2000 transfection reagent. Young C57/BL6J mice were kept in a hyperoxia environment, followed by indoor air, resulting in oxygen-induced retinopathy. Oxygen-induced retinopathy (OIR) animals were randomly divided into three groups: OIR group, OIR + vector group (OIR cubs treated with rAAV vector) and OIR + PSF group (OIR cubs treated with rAAV-PSF). Age-matched C57/BL6J mice were used as controls and exposed to constant normoxic conditions. The animals were executed and their pupils were subjected to subsequent experiments. The metabolic spectrum was analyzed by Seahorse XFe96 flux analyzer, and OCR and extracellular acidification rate were quantified at the same time.
    RESULTS: PSF ameliorated retinal neovascularization and corrected abnormal VEGF expression in mice with oxygen-induced retinopathy and reduced intra-retinal neovascularization in Vldlr - / - mice. PSF reprogrammed mitochondrial bioenergetics and inhibited the transition of endothelial cells after hypoxia, suggesting its involvement in pathological angiogenesis.Ectopic PSF expression inhibited hypoxia-induced HIF-1α activation in the nucleus by recruiting Hakai to the PSF/HIF-1α complex, causing HIF-1α inhibition. PSF knockdown increased hypoxia-stimulated HIF-1α reactions. These hypoxia-dependent processes may play a vital role in cell metabolism, migration, and proliferation. Thus, PSF is a potential treatment target in neovascularization-associated ophthalmopathy.
    CONCLUSION: This is the first study showing that PSF inhibits HIF-1α via recruitment of Hakai, modulates mitochondrial oxidation and glycolysis, and downregulates VEGF expression under hypoxia. We propose a new HIF-1 α/Hakai regulatory mechanism that may play a vital role in the pathogenesis of neovascularization in ophthalmopathy. PSF-Hakai-HIF-1α signaling pathway under hypoxia condition. Schematic diagram showing that the PSF-Hakai-HIF-1α signaling pathway. Under hypoxia condition, PSF-Hakai complex regulate HIF-1α signaling, thus inhibiting downstream target gene VEGF, cell metabolism and angiogenesis eventually. Video Abstract: Detailed information of Materials and Methods.
    Keywords:  HIF1-α; Hakai; Hypoxia; Mitochondrion; Neovascularization; PSF; VEGF
    DOI:  https://doi.org/10.1186/s12964-020-00684-w
  13. Cell Death Discov. 2020 Mar 30. 6(1): 18
    Doccini S, Morani F, Nesti C, Pezzini F, Calza G, Soliymani R, Signore G, Rocchiccioli S, Kanninen KM, Huuskonen MT, Baumann MH, Simonati A, Lalowski MM, Santorelli FM.
      CLN5 disease is a rare form of late-infantile neuronal ceroid lipofuscinosis (NCL) caused by mutations in the CLN5 gene that encodes a protein whose primary function and physiological roles remains unresolved. Emerging lines of evidence point to mitochondrial dysfunction in the onset and progression of several forms of NCL, offering new insights into putative biomarkers and shared biological processes. In this work, we employed cellular and murine models of the disease, in an effort to clarify disease pathways associated with CLN5 depletion. A mitochondria-focused quantitative proteomics approach followed by functional validations using cell biology and immunofluorescence assays revealed an impairment of mitochondrial functions in different CLN5 KO cell models and in Cln5-/- cerebral cortex, which well correlated with disease progression. A visible impairment of autophagy machinery coupled with alterations of key parameters of mitophagy activation process functionally linked CLN5 protein to the process of neuronal injury. The functional link between impaired cellular respiration and activation of mitophagy pathways in the human CLN5 disease condition was corroborated by translating organelle-specific proteome findings to CLN5 patients' fibroblasts. Our study highlights the involvement of CLN5 in activation of mitophagy and mitochondrial homeostasis offering new insights into alternative strategies towards the CLN5 disease treatment.
    DOI:  https://doi.org/10.1038/s41420-020-0250-y
  14. Cell Mol Life Sci. 2021 Feb 13.
    Poole LP, Macleod KF.
      Cells use mitophagy to remove dysfunctional or excess mitochondria, frequently in response to imposed stresses, such as hypoxia and nutrient deprivation. Mitochondrial cargo receptors (MCR) induced by these stresses target mitochondria to autophagosomes through interaction with members of the LC3/GABARAP family. There are a growing number of these MCRs, including BNIP3, BNIP3L, FUNDC1, Bcl2-L-13, FKBP8, Prohibitin-2, and others, in addition to mitochondrial protein targets of PINK1/Parkin phospho-ubiquitination. There is also an emerging link between mitochondrial lipid signaling and mitophagy where ceramide, sphingosine-1-phosphate, and cardiolipin have all been shown to promote mitophagy. Here, we review the upstream signaling mechanisms that regulate mitophagy, including components of the mitochondrial fission machinery, AMPK, ATF4, FoxOs, Sirtuins, and mtDNA release, and address the significance of these pathways for stress responses in tumorigenesis and metastasis. In particular, we focus on how mitophagy modulators intersect with cell cycle control and survival pathways in cancer, including following ECM detachment and during cell migration and metastasis. Finally, we interrogate how mitophagy affects tissue atrophy during cancer cachexia and therapy responses in the clinic.
    Keywords:  AMPK; ATF4; Autophagy; BCL2-L-13; BNIP3/BNIP3L; Cachexia; DRP1; Electron transport chain; FUNDC1; Fission; FoxOs; LC3/GABARAP; Metabolism; Metastasis; Mitochondria; Mitohormesis; Mitophagy; NAD+; PARP; PINK1/Parkin; ROS; Respiration; Sirtuins; UPRmt
    DOI:  https://doi.org/10.1007/s00018-021-03774-1
  15. Sci Rep. 2021 Feb 08. 11(1): 3308
    Sorrells JE, Martin EM, Aksamitiene E, Mukherjee P, Alex A, Chaney EJ, Marjanovic M, Boppart SA.
      The heterogeneous nature of extracellular vesicles (EVs) creates the need for single EV characterization techniques. However, many common biochemical and functional EV analysis techniques lack single EV resolution. Two-photon fluorescence lifetime imaging microscopy (FLIM) is widely used to functionally characterize the reduced form of nicotinamide adenine dinucleotide and nicotinamide adenine dinucleotide phosphate (NAD(P)H) in cells and tissues. Here, we demonstrate that FLIM can also be used to image and characterize NAD(P)H in single isolated EVs. EVs were isolated using standard differential ultracentrifugation techniques from multiple cell lines and imaged using a custom two-photon FLIM system. The presented data show that the NAD(P)H fluorescence lifetimes in isolated cell-derived EVs follow a wide Gaussian distribution, indicating the presence of a range of different protein-bound and free NAD(P)H species. EV NAD(P)H fluorescence lifetime distribution has a larger standard deviation than that of cells and a significantly different fluorescence lifetime distribution than the nuclei, mitochondria, and cytosol of cells. Additionally, changes in the metabolic conditions of cells were reflected in changes in the mean fluorescence lifetime of NAD(P)H in the produced EVs. These data suggest that FLIM of NAD(P)H could be a valuable tool for EV research.
    DOI:  https://doi.org/10.1038/s41598-020-80813-0
  16. Cell Mol Life Sci. 2021 Feb 12.
    Shah M, Chacko LA, Joseph JP, Ananthanarayanan V.
      The ability of a mitochondrion to undergo fission and fusion, and to be transported and localized within a cell are central not just to proper functioning of mitochondria, but also to that of the cell. The cytoskeletal filaments, namely microtubules, F-actin and intermediate filaments, have emerged as prime movers in these dynamic mitochondrial shape and position transitions. In this review, we explore the complex relationship between the cytoskeleton and the mitochondrion, by delving into: (i) how the cytoskeleton helps shape mitochondria via fission and fusion events, (ii) how the cytoskeleton facilitates the translocation and anchoring of mitochondria with the activity of motor proteins, and (iii) how these changes in form and position of mitochondria translate into functioning of the cell.
    Keywords:  Cytoskeleton; Microtubules; Mitochondria; Mitochondrial dynamics; Molecular motors
    DOI:  https://doi.org/10.1007/s00018-021-03762-5
  17. PLoS Comput Biol. 2021 Feb 12. 17(2): e1008750
    Preto J, Krimm I.
      The voltage-dependent anion channel (VDAC) is a critical β-barrel membrane protein of the mitochondrial outer membrane, which regulates the transport of ions and ATP between mitochondria and the cytoplasm. In addition, VDAC plays a central role in the control of apoptosis and is therefore of great interest in both cancer and neurodegenerative diseases. Although not fully understood, it is presumed that the gating mechanism of VDAC is governed by its N-terminal region which, in the open state of the channel, exhibits an α-helical structure positioned midway inside the pore and strongly interacting with the β-barrel wall. In the present work, we performed molecular simulations with a recently developed force field for disordered systems to shed new light on known experimental results, showing that the N-terminus of VDAC is an intrinsically disordered region (IDR). First, simulation of the N-terminal segment as a free peptide highlighted its disordered nature and the importance of using an IDR-specific force field to properly sample its conformational landscape. Secondly, accelerated dynamics simulation of a double cysteine VDAC mutant under applied voltage revealed metastable low conducting states of the channel representative of closed states observed experimentally. Related structures were characterized by partial unfolding and rearrangement of the N-terminal tail, that led to steric hindrance of the pore. Our results indicate that the disordered properties of the N-terminus are crucial to properly account for the gating mechanism of VDAC.
    DOI:  https://doi.org/10.1371/journal.pcbi.1008750
  18. J Biol Chem. 2021 Feb 05. pii: S0021-9258(21)00155-1. [Epub ahead of print] 100383
    Lysyk L, Brassard R, Arutyunova E, Siebert V, Jiang Z, Takyi E, Morrison M, Young HS, Lemberg MK, O'Donoghue AJ, Lemieux MJ.
      The rhomboid protease PARL is a critical regulator of mitochondrial homeostasis through its cleavage of substrates such as PINK1, PGAM5, and Smac/Diablo, which have crucial roles in mitochondrial quality control and apoptosis. However, the catalytic properties of PARL, including the effect of lipids on the protease, have never been characterized in vitro. To address this, we isolated human PARL expressed in yeast and used FRET-based kinetic assays to measure proteolytic activity in vitro. We show PARL activity in detergent is enhanced by cardiolipin, a lipid enriched in the mitochondrial inner membrane. Significantly higher turnover rates were observed for PARL reconstituted in proteoliposomes, with Smac/Diablo being cleaved most rapidly at a rate of 1 min-1. In contrast, PGAM5 is cleaved with the highest efficiency (kcat/KM) compared to PINK1 and Smac/Diablo. In proteoliposomes, a truncated β-cleavage form of PARL, a physiological form known to affect mitochondrial fragmentation, is more active than the full-length enzyme for hydrolysis of PINK1, PGAM5 and Smac/Diablo. Multiplex profiling of 228 peptides reveals that PARL prefers substrates with a bulky side chain such as Phe in P1, which is distinct from the preference for small side chain residues typically found with bacterial rhomboid proteases. This study using recombinant PARL provides fundamental insights into its catalytic activity and substrate preferences that enhance our understanding of its role in mitochondrial function and has implications for specific inhibitor design.
    Keywords:  GlpG; PGAM5; PINK1; Smac/Diablo; intramembrane proteolysis; membrane protease; mitochondria; rhomboid protease
    DOI:  https://doi.org/10.1016/j.jbc.2021.100383
  19. Cell Mol Life Sci. 2021 Feb 13.
    Sessions DT, Kashatus DF.
      Many tumors are now understood to be heterogenous cell populations arising from a minority of epithelial-like cancer stem cells (CSCs). CSCs demonstrate distinctive metabolic signatures from the more differentiated surrounding tumor bulk that confer resistance to traditional chemotherapeutic regimens and potential for tumor relapse. Many CSC phenotypes including metabolism, epithelial-to-mesenchymal transition, cellular signaling pathway activity, and others, arise from altered mitochondrial function and turnover, which are regulated by constant cycles of mitochondrial fusion and fission. Further, recycling of mitochondria through mitophagy in CSCs is associated with maintenance of reactive oxygen species levels that dictate gene expression. The protein machinery that drives mitochondrial dynamics is surprisingly simple and may represent attractive new therapeutic avenues to target CSC metabolism and selectively eradicate tumor-generating cells to reduce the risks of metastasis and relapse for a variety of tumor types.
    Keywords:  Cancer stem cells; EMT; Metabolism; Mitochondrial dynamics; Mitochondrial morphology; Signaling; Therapeutic resistance
    DOI:  https://doi.org/10.1007/s00018-021-03773-2
  20. Mol Metab. 2021 Feb 06. pii: S2212-8778(21)00025-9. [Epub ahead of print] 101185
    Kim MJ, Febbraro D, Farkona S, Gillmore T, Son JE, Regeenes R, Chang HH, Pollock-Tahiri E, Yang J, Park YJ, Sivasubramaniyam T, Oh SJ, Saraon P, Stagljar I, Rocheleau JV, Hui CC, Caniggia I, Hao Z, Mak TW, Konvalinka A, Woo M.
      OBJECTIVE: Autophagy is a physiological self-eating process that can promote cell survival or activate cell death in eukaryotic cells. In skeletal muscle, it is important in the maintenance of muscle mass and function that is critical to sustain mobility and regulate metabolism. UV radiation resistance-associated gene (UVRAG) regulates early stages of autophagy and autophagosome maturation, while also playing a key role in endosomal trafficking. This study investigated the essential in vivo role of UVRAG in skeletal muscle biology.METHODS: To determine the role of UVRAG in skeletal muscle in vivo, we generated muscle specific UVRAG knock-out mice using the cre-loxP system driven by Myf6 promoter that is exclusively expressed in skeletal muscle. Myf6- Cre+ UVRAGfl/fl (M-UVRAG-/-) mice were compared to littermate Myf6-Cre+ UVRAG+/+ (M-UVRAG+/+) controls under basal conditions on normal chow diet. Body composition, muscle function and mitochondria morphology were assessed in muscles of WT and KO mice at 24 weeks of age.
    RESULTS: M-UVRAG-/- mice developed accelerated sarcopenia and impaired muscle function compared to M-UVRAG+/+ littermates at 24 weeks of age. Interestingly, these mice displayed improved glucose tolerance and increased energy expenditure likely related to up-regulated Fgf21, a marker of muscle dysfunction. Skeletal muscle of M-UVRAG-/- mice showed altered mitochondrial morphology with increased mitochondrial fission, as well as EGFR accumulation reflecting defects in endosomal trafficking. To determine whether increased EGFR signaling had a causal role in muscle dysfunction, mice were treated with an EGFR inhibitor, gefitinib, which partially restored markers of muscle and mitochondrial deregulation. Conversely, constitutively active EGFR transgenic expression in UVRAG deficient muscle led to further detrimental effects with non-overlapping distinct defects in muscle function, with EGFR activation affecting muscle fiber type whereas UVRAG deficiency impaired mitochondrial homeostasis.
    CONCLUSIONS: Our results show that both UVRAG and EGFR signaling are critical in the maintenance of muscle mass and function with distinct mechanisms in the differentiation pathway.
    Keywords:  EGFR; Fgf21; UVRAG; mitochondrial dynamics; skeletal muscle
    DOI:  https://doi.org/10.1016/j.molmet.2021.101185
  21. J Biol Chem. 2021 Feb 03. pii: S0021-9258(21)00142-3. [Epub ahead of print] 100370
    Chinnarasu S, Alogaili F, Bove KE, Jaeschke A, Hui DY.
      The LDL receptor-related protein 1 (LRP1) is a multi-functional transmembrane protein with endocytosis and signal transduction functions. Previous studies have shown that hepatic LRP1 deficiency exacerbates diet-induced steatohepatitis and insulin resistance via mechanisms related to increased lysosome and mitochondria permeability and dysfunction. The current study examined the impact of LRP1 deficiency on mitochondrial function in the liver. Hepatocytes isolated from liver-specific LRP1 knockout (hLrp1-/-) mice showed reduced oxygen consumption compared to control mouse hepatocytes. The mitochondria in hLrp1-/- mouse livers have an abnormal morphology and their membranes contain significantly less anionic phospholipids, including lower levels of phosphatidylethanolamine and cardiolipin that increase mitochondrial fission and impair fusion. Additional studies showed that LRP1 complexes with phosphatidylinositol 4-phosphate 5-kinase like protein-1 (PIP5KL1) and phosphatidylinositol 4-phosphate 5-kinase-1β (PIP5K1β). The absence of LRP1 reduces the levels of both PIP5KL1 and PIP5K1β in the plasma membrane, and also lowers phosphatidylinositol(4,5) bisphosphate (PI(4,5)P2) levels in hepatocytes. These data indicate that LRP1 recruits PIP5KL1 and PIP5K1β to the plasma membrane for PI(4,5)P2 biosynthesis. The lack of LRP1 reduces lipid kinase expression, leading to lower PI(4,5)P2 levels thereby decreasing the availability of this lipid metabolite in the cardiolipin biosynthesis pathway to cause cardiolipin reduction and the impairment in mitochondria homeostasis. Taken together, the current study identifies another signaling mechanism by which LRP1 regulates cell functions: Binding and recruitment of PIP5KL1 and PIP5K1β to the membrane for PI(4,5)P2 synthesis. In addition, it highlights the importance of this mechanism for maintaining the integrity and functions of intracellular organelles.
    Keywords:  cardiolipin; inositol phosphate; lipoprotein receptor; lipoprotein receptor-related protein (LRP); liver metabolism; respiration
    DOI:  https://doi.org/10.1016/j.jbc.2021.100370
  22. Matrix Biol. 2021 Feb 05. pii: S0945-053X(21)00014-7. [Epub ahead of print]
    Mende H, Müller S.
      The nucleolus functions as the cellular hub for the initiation and early steps of ribosome biogenesis. Ribosomes are key components of the translation machinery and, accordingly, their abundance needs to be adjusted to the cellular energy status. Further, to ensure translational fidelity, the integrity and quality of ribosomes needs to be monitored under conditions of cellular stress. Stressful insults, such as nutrient, genotoxic or proteotoxic stress, interfere with ribosome biogenesis and activate a cellular response referred to as nucleolar stress. This nucleolar stress response typically affects nucleolar integrity and is intricately linked to the activation of protein quality control pathways, including (i) the ubiquitin proteasome system (UPS) and (ii) the autophagy machinery, to restore cellular proteostasis. Here we will review some key features of the nucleolar stress response with a particular focus on the role of the UPS and autophagy in this process.
    Keywords:  Autophagy; nucleolar stress; p53; ribosome biogenesis; ubiquitin-proteasome system (UPS)
    DOI:  https://doi.org/10.1016/j.matbio.2021.02.001
  23. Biochim Biophys Acta Gen Subj. 2021 Feb 08. pii: S0304-4165(21)00030-1. [Epub ahead of print] 129871
    Paul S, Pickrell AM.
      PINK1, a serine/threonine ubiquitin kinase, and Parkin, an E3 ubiquitin ligase, work in coordination to target damaged mitochondria to the lysosome in a process called mitophagy. This review will cover what we have learned from PINK1 and Parkin knockout (KO) mice. Systemic PINK1 and Parkin KO mouse models haven't faithfully recapitulated early onset forms of Parkinson's disease found in humans with recessive mutations in these genes. However, the utilization of these mouse models has given us insight into how PINK1 and Parkin contribute to mitochondrial quality control and function in different tissues beyond the brain such as in heart and adipose tissue. Although PINK1 and Parkin KO mice have been generated over a decade ago, these models are still being used today to creatively elucidate cell-type specific functions. Recently, these mouse models have uncovered that these proteins contribute to innate immunity and cancer phenotypes.
    Keywords:  PINK1; Parkin; knockout mouse; mitophagy
    DOI:  https://doi.org/10.1016/j.bbagen.2021.129871
  24. Biomolecules. 2021 Feb 05. pii: 222. [Epub ahead of print]11(2):
    Manganelli V, Capozzi A, Recalchi S, Riitano G, Mattei V, Longo A, Misasi R, Garofalo T, Sorice M.
      Cardiolipin (CL) is a hallmark phospholipid localized within the inner mitochondrial membrane. Upon several mitochondrial stress conditions, CL is translocated to specialized platforms, where it may play a role in signaling events to promote mitophagy and apoptosis. Recent studies characterized the molecular composition of MAM-associated lipid microdomains and their implications in regulating the autophagic process. In this study we analyzed the presence of CL within MAMs following autophagic stimulus and the possible implication of raft-like microdomains enriched in CL as a signaling platform in autophagosome formation. Human 2FTGH fibroblasts and SKNB-E-2 cells were stimulated under nutrient deprivation with HBSS. MAM fraction was obtained by an ultracentrifugation procedure and analyzed by HPTLC immunostaining. CL interactions with mitofusin2 (MFN2), calnexin (CANX) and AMBRA1 were analyzed by scanning confocal microscopy and coimmunoprecipitation. The analysis revealed that CL accumulates in MAMs fractions following autophagic stimulus, where it interacts with MFN2 and CANX. It associates with AMBRA1, which in turn interacts with BECN1 and WIPI1. This study demonstrates that CL is present in MAM fractions following autophagy triggering and interacts with the multimolecular complex (AMBRA1/BECN1/WIPI1) involved in autophagosome formation. It may have both structural and functional implications in the pathophysiology of neurodegenerative disease(s).
    Keywords:  MAMs; autophagosome; cardiolipin; mitochondria
    DOI:  https://doi.org/10.3390/biom11020222
  25. Neurosci Res. 2021 Feb 06. pii: S0168-0102(21)00030-4. [Epub ahead of print]
    Nii T, Eguchi R, Otsuguro KI.
      Hydrogen sulfide (H2S) is a well-known inhibitor of the mitochondrial electron transport chain (ETC). H2S also increases intracellular Ca2+ levels in astrocytes, which are glial cells and that supply lactate as an energy substrate to neurons. Here, we examined the relationship between H2S-induced metabolic changes and Ca2+ responses in spinal cord astrocytes. Na2S (150 μM), an H2S donor, increased the intracellular Ca2+ concentration, which was inhibited by an ETC inhibitor and an uncoupler of mitochondrial oxidative phosphorylation. Na2S also increased the accumulation of extracellular lactate. Na2S alone did not change intracellular ATP content, but decreased it when glycolysis was inhibited. The Na2S-induced Ca2+ increase and accumulation of extracellular lactate were inhibited by emetine, an inhibitor of translocon complex, which mediates Ca2+ leak from the endoplasmic reticulum (ER). Furthermore, an inhibitor of the Ca2+-sensitive NADH shuttle decreased Na2S-mediated accumulation of lactate. We conclude that inhibition of the mitochondrial ETC by H2S induces Ca2+ release from mitochondria and the ER in spinal cord astrocytes, which increases lactate production. H2S may promote glycolysis by activating the Ca2+-sensitive NADH shuttle and facilitating the supply of lactate from astrocytes to neurons.
    Keywords:  ATP; astrocytes; calcium; hydrogen sulfide; lactate
    DOI:  https://doi.org/10.1016/j.neures.2021.01.008
  26. Biochim Biophys Acta Bioenerg. 2021 Feb 04. pii: S0005-2728(21)00026-8. [Epub ahead of print] 148393
    Kuznetsov AV, Javadov S, Margreiter R, Grimm M, Hagenbuchner J, Ausserlechner MJ.
      Cancer cells bioenergetics is more dependent on glycolysis than mitochondrial oxidative phosphorylation, a phenomenon known as the Warburg Effect. It has been proposed that inhibition of glycolysis may selectively affect cancer cells. However, the effects of glycolysis inhibition on mitochondrial function and structure in cancer cells are not completely understood. Here, we investigated the comparative effects of 2-deoxy-D-glucose (2-DG, a glucose analogue, which suppresses cellular glycolysis) on cellular bioenergetics in human colon cancer DLD-1 cells, smooth muscle cells, human umbilical vein endothelial cells and HL-1 cardiomyocytes. In all cells, 2-DG treatment resulted in significant ATP depletion, however, the cell viability remained unchanged. Also, we did not observe the synergistic effects of 2-DG with anticancer drugs doxorubicin and 5-fluorouracil. Instead, after 2-DG treatment and ATP depletion, mitochondrial respiration and membrane potential were significantly enhanced and mitochondrial morphology changed in the direction of more network organization. Analysis of protein expression demonstrated that 2-DG treatment induced an activation of AMPK (elevated pAMPK/AMPK ratio), increased mitochondrial fusion (mitofusins 1 and 2) and decreased fission (Drp1) proteins. In conclusion, this study suggests a strong link between respiratory function and structural organization of mitochondria in the cell. We propose that the functionality of the mitochondrial network is enhanced compared to disconnected mitochondria.
    Keywords:  2-Deoxy-D-glucose; Cancer cells; Cellular ATP; Energy stress; Glucose metabolism; Mitochondria; Mitochondrial dynamics/network; Mitochondrial function; Mitochondrial membrane potential
    DOI:  https://doi.org/10.1016/j.bbabio.2021.148393
  27. Mol Ther Nucleic Acids. 2021 Mar 05. 23 643-656
    Woo HN, Park S, Kim HL, Jung MK, Pack CG, Park J, Cho Y, Jo DG, Kim DK, Mook-Jung I, Kim SW, Lee H.
      Adult hippocampal neurogenesis supports the structural and functional plasticity of the brain, while its decline is associated with neurodegeneration common in Alzheimer's disease (AD). Although the dysregulation of certain microRNAs (miRNAs) in AD have been observed, the effects of miRNAs on hippocampal neurogenesis are largely unknown. In this study, we demonstrated miR-351-5p as a causative factor in hippocampal neural progenitor cell death through modulation of the mitochondrial guanosine triphosphatase (GTPase), Miro2. Downregulation of Miro2 by siMiro2 induced cell death, similar to miR-351-5p, whereas ectopic Miro2 expression using an adenovirus abolished these effects. Excessively fragmented mitochondria and dysfunctional mitochondria were indexed by decreased mitochondrial potential, and increased reactive oxygen species were identified in miR-351-5p-induced cell death. Moreover, subsequent induction of mitophagy via Pink1 and Parkin was observed in the presence of miR-351-5p and siMiro2. The suppression of mitochondrial fission by Mdivi-1 completely inhibited cell death by miR-351-5p. miR-351-5p expression increased whereas the level of Miro2 decreased in the hippocampus of AD model mice, emulating expression in AD patients. Collectively, the data indicate the mitochondrial fission and accompanying mitophagy by miR-351-5p/Miro2 axis as critical in hippocampal neural progenitor cell death, and a potential therapeutic target in AD.
    Keywords:  Alzheimer’s disease; Miro GTPase; autophagy; cell death; hippocampal neural progenitor cells; miRNA-351-5p; mitophagy
    DOI:  https://doi.org/10.1016/j.omtn.2020.12.014
  28. Free Radic Res. 2021 Feb 08. 1-11
    Canizal-García M, Olmos-Orizaba BE, Moreno-Jiménez M, Calderón-Cortés E, Saavedra-Molina A, Cortés-Rojo C.
      Glutathione peroxidase 4 (Gpx4) counteracts mitochondrial lipid peroxidation in mammals. In yeast, Gpx2 is orthologous of Gpx4, is localized in mitochondria, and reduces both inorganic and organic peroxides. However, a phenotype of oxidative stress hypersensitivity has not been observed with gpx2 deletion. We hypothesized that the absence of polyunsaturated fatty acids (PUFA) in yeast membranes may mask an antioxidant role of Gpx2 in mitochondria. Thus, we tested the effects of PUFA on cell viability, mitochondrial function, ROS production, and mitochondrial fatty acid composition of a gpx2Δ mutant subjected to chronological aging. As expected, gpx2Δ mutation did not alter these parameters with respect to wild-type (WT) cells after 30 h growth, even in the presence of linolenic acid (C18:3), except for some activities of the electron transport chain (ETC) complexes. Conversely, aged gpx2Δ cells exhibited lower viability, impaired respiration, decreased ETC activities, and increased ROS generation in comparison to aged WT cells. These effects were exacerbated by C18:3, as gpx2Δ cells displayed residual respiration, full inhibition of ETC complexes, and a burst in ROS production on day 15 that decreased on day 30, although ROS remained several-fold higher than in WT cells. gpx2 was not involved in the preservation of PUFA levels, as no differences in mitochondrial C18:3 content were observed between WT and gpx2Δ cells. These results indicate that gpx2 is a late - acting antioxidant system that decreases mitochondrial ROS production and preserves ETC function, without being involved in the preservation of PUFA levels in mitochondria.
    Keywords:  Yeast; aging; electron transport chain; glutathione peroxidase; polyunsaturated fatty acids
    DOI:  https://doi.org/10.1080/10715762.2021.1882677
  29. Oxid Med Cell Longev. 2021 ;2021 6620677
    Chang X, Zhao Z, Zhang W, Liu D, Ma C, Zhang T, Meng Q, Yan P, Zou L, Zhang M.
      Cardiovascular disease has become one of the main causes of human death. In addition, many cardiovascular diseases are accompanied by a series of irreversible damages that lead to organ and vascular complications. In recent years, the potential therapeutic strategy of natural antioxidants in the treatment of cardiovascular diseases through mitochondrial quality control has received extensive attention. Mitochondria are the main site of energy metabolism in eukaryotic cells, including myocardial and vascular endothelial cells. Mitochondrial quality control processes ensure normal activities of mitochondria and cells by maintaining stable mitochondrial quantity and quality, thus protecting myocardial and endothelial cells against stress. Various stresses can affect mitochondrial morphology and function. Natural antioxidants extracted from plants and natural medicines are becoming increasingly common in the clinical treatment of diseases, especially in the treatment of cardiovascular diseases. Natural antioxidants can effectively protect myocardial and endothelial cells from stress-induced injury by regulating mitochondrial quality control, and their safety and effectiveness have been preliminarily verified. This review summarises the damage mechanisms of various stresses in cardiomyocytes and vascular endothelial cells and the mechanisms of natural antioxidants in improving the vulnerability of these cell types to stress by regulating mitochondrial quality control. This review is aimed at paving the way for novel treatments for cardiovascular diseases and the development of natural antioxidant drugs.
    DOI:  https://doi.org/10.1155/2021/6620677
  30. Org Biomol Chem. 2021 Feb 08.
    Mukherjee T, Soppina V, Ludovic R, Mély Y, Klymchenko AS, Collot M, Kanvah S.
      Visualization of sub-cellular organelles allows the determination of various cellular processes and the underlying mechanisms. Herein, we report a fluorescent probe, bearing push-pull substituents emitting at 600 nm and its application in cellular imaging. The probe shows dual imaging of mitochondria and nucleoli and maps mitochondrial viscosity in live cells under various physiological variations and show minimum cytotoxicity. Nucleolar staining is confirmed by RNAase digestion.
    DOI:  https://doi.org/10.1039/d0ob02378g
  31. Annu Rev Biochem. 2021 Feb 08.
    Ruprecht JJ, Kunji ERS.
      Members of the mitochondrial carrier family [solute carrier family 25 (SLC25)] transport nucleotides, amino acids, carboxylic acids, fatty acids, inorganic ions, and vitamins across the mitochondrial inner membrane. They are important for many cellular processes, such as oxidative phosphorylation of lipids and sugars, amino acid metabolism, macromolecular synthesis, ion homeostasis, cellular regulation, and differentiation. Here, we describe the functional elements of the transport mechanism of mitochondrial carriers, consisting of one central substrate-binding site and two gates with salt-bridge networks on either side of the carrier. Binding of the substrate during import causes three gate elements to rotate inward, forming the cytoplasmic network and closing access to the substrate-binding site from the intermembrane space. Simultaneously, three core elements rock outward, disrupting the matrix network and opening the substrate-binding site to the matrix side of the membrane. During export, substrate binding triggers conformational changes involving the same elements but operating in reverse. Expected final online publication date for the Annual Review of Biochemistry, Volume 90 is June 2021. Please see http://www.annualreviews.org/page/journal/pubdates for revised estimates.
    DOI:  https://doi.org/10.1146/annurev-biochem-072820-020508
  32. J Biosci. 2021 ;pii: 7. [Epub ahead of print]46
    Wu Y, Gu C, Huang LU, Zhao Y, Tang Y, Zhao H, Wu Q.
      The protective effect of astrocytes on nerves was demonstrated by mitochondrial transfer. The neuroprotective effect of hypoxic pretreatment is widely accepted. The aim of this research is to investigate the role of hypoxic preconditioning on astrocytes mitochondria. Rat neuronal cells and astrocytes were isolated and cultured. A hypoxic preconditioned astrocyte and oxygen glucose deprivation (OGD) neuronal cell co-culture experiment was used to detect the effect of hypoxic preconditioning (HP) on nerve damage. The silencing of proliferatoractivated receptor γ coactivator-1α (PGC-1α) with siRNA was used to explore the role of HP in the repair of nerve damage and biogenesis of mitochondria. HP increased astrocyte viability and promoted neuroprotective factor secretion. The expression levels of antioxidant enzymes, PGC-1α and uncoupling protein2 (UCP2) were up-regulated by HP. In addition, HP improved mitochondrial function and reduced oxidative stress induced by OGD. It was found that HP astrocytes had a greater neuroprotective effect than normal astrocytes cells. Neuronal apoptosis and reactive oxygen species levels were down-regulated by cell co-culture. The PGC-1α siRNA experiment showed that hypoxia treatment promotes mitochondrial biogenesis and plays a neuroprotective role. HP significantly enhanced the efficacy of astrocytes in the treatment of neuronal injury.
  33. Redox Biol. 2021 Jan 26. pii: S2213-2317(21)00026-4. [Epub ahead of print]41 101878
    Wang H, Sun X, Lu Q, Zemskov EA, Yegambaram M, Wu X, Wang T, Tang H, Black SM.
      Acute lung injury (ALI) is a devastating clinical syndrome with no effective therapies. Inflammasome activation has been reported to play a critical role in the initiation and progression of ALI. The molecular mechanisms involved in regulating the activation of inflammasome in ALI remains unresolved, although increases in mitochondrial derived reactive oxygen species (mito-ROS) are involved. Our previous work has shown that the mitochondrial redistribution of uncoupled eNOS impairs mitochondrial bioenergetics and increases mito-ROS generation. Thus, the focus of our study was to determine if lipopolysaccharide (LPS)-mediated inflammasome activation involves the mitochondrial redistribution of uncoupled eNOS. Our data show that the increase in mito-ROS involved in LPS-mediated inflammasome activation is associated with the disruption of mitochondrial bioenergetics in human lung microvascular endothelial cells (HLMVEC) and the mitochondrial redistribution of eNOS. These effects are dependent on RhoA-ROCK signaling and are mediated via increased phosphorylation of eNOS at Threonine (T)-495. A derivative of the mitochondrial targeted Szeto-Schiller peptide (SSP) attached to the antioxidant Tiron (T-SSP), significantly attenuated LPS-mediated mito-ROS generation and inflammasome activation in HLMVEC. Further, T-SSP attenuated mitochondrial superoxide production in a mouse model of sepsis induced ALI. This in turn significantly reduced the inflammatory response and attenuated lung injury. Thus, our findings show that the mitochondrial redistribution of uncoupled eNOS is intimately involved in the activation of the inflammatory response in ALI and implicate attenuating mito-ROS as a therapeutic strategy in humans.
    Keywords:  ALI; Inflammasome activation; Mitochondrial ROS; RhoA; eNOS uncoupling
    DOI:  https://doi.org/10.1016/j.redox.2021.101878
  34. Transplantation. 2021 Feb 11.
    Shi S, Wang L, van der Laan LJW, Pan Q, Verstegen MMA.
      Mitochondria are essential organelles for cellular energy and metabolism. Like with any organ, the liver highly depends on the function of these cellular powerhouses. Hepatotoxic insults often lead to an impairment of mitochondrial activity and an increase in oxidative stress, thereby compromising the metabolic and synthetic functions. Mitochondria play a critical role in ATP synthesis and the production or scavenging of free radicals. Mitochondria orchestrate many cellular signalling pathways involved in the regulation of cell death, metabolism, cell division and progenitor cell differentiation. Mitochondrial dysfunction and oxidative stress are closely associated with ischemia-reperfusion injury during organ transplantation and with different liver diseases, including cholestasis, steatosis, viral hepatitis, and drug-induced liver injury. In order to develop novel mitochondria-targeting therapies or interventions, a better understanding of mitochondrial dysfunction and oxidative stress in hepatic pathogenesis is very much needed. Therapies targeting mitochondria impairment and oxidative imbalance in liver diseases have been extensively studied in preclinical and clinical research. In this review, we provide an overview of how oxidative stress and mitochondrial dysfunction affect liver diseases and liver transplantation. Furthermore, we summarize recent developments of antioxidant and mitochondria-targeted interventions.
    DOI:  https://doi.org/10.1097/TP.0000000000003691
  35. Mol Immunol. 2021 Feb 05. pii: S0161-5890(21)00042-0. [Epub ahead of print]132 82-92
    Wang Z, Han S, Chen X, Li X, Xia N, Pu L.
      Ischemia-reperfusion(IR) injury is one of the main complications of liver transplantation and partial hepatectomy. Innate immunity mediated by kupffer cells plays an important role in it. In this study, we focused on evaluating the intrinsic relationship between the autophagy induction of kupffer cells and the activation of NLRP3 inflammasomes caused by liver ischemia-reperfusion. Pre-depletion of kupffer cells can aggravate inflammation and tissue damage within 24 h after IR.Enhancing the autophagy of kupffer cells can inhibit the activation of NLRP3 caused by IR, and inhibiting autophagy can induce the secretion of IL1β dependent on NLRP3 activation.Eva1a is up-regulated by the inflammatory cascade activated by IR.Knockdown of Eva1a in vivo on the one hand will aggravate IR inflammation, increase the production of TNF-α, IL-1β and inhibit the secretion of IL-10.On the other hand, it will aggravate the liver histological damage. Knockout of Eva1a induces ASC activation and cleavage of caspase1 and IL1β in an NLRP3-dependent manner, which is closely related to the function of blocking Eva1a to promote autophagosome formation.We further found that knockdown of ATG16L1 will reverse the more formation of autophagosomes induced by overexpression of Eva1a, whereas knockdown of ATG16L1 did not further reduce the formation of autophagosomes inhibited by siEva1a. We also found that the addition of siATG7, siATG5 and siATG12 would reverse the IR autophagy of liver induced by overexpression of Eva1a, but inhibition of the Beclin1-Vps34 pathway did not significantly reverse the effect of overexpression of Eva1a.These prove that Eva1a and ATG16L1 may work together in the liver IR model to actively induce the formation of autophagosomes and be independent from the beclin1-vps34-induced autophagy pathway to limit the excessive activation of IR inflammation. Our study provides brand new insights into the mechanism of liver macrophages in the progression of inflammation in the context of liver ischemia-reperfusion injury.
    Keywords:  ATG16L1; Autophagy; Eva1a; Kupffer cells; Liver ischemia reperfusion injury; NLRP3
    DOI:  https://doi.org/10.1016/j.molimm.2021.01.028
  36. Inflammation. 2021 Feb 11.
    Wang XP, Zheng WC, Bai Y, Li Y, Xin Y, Wang JZ, Chang YL, Zhang LM.
      Following hepatic ischemia-reperfusion injury, Kupffer cells could be activated by inflammatory factors released from damaged hepatocytes. Carbon monoxide (CO)-releasing molecule (CORM)-3, a water-soluble transition metal carbonyl, exhibits excellent anti-inflammatory and anti-pyroptosis properties. We investigated whether CORM-3 attenuated hemorrhagic shock and resuscitation (HSR)-induced pyroptosis of Kupffer cells through the soluble guanylate-cyclase (sGC)-cyclic guanosine monophosphate (cGMP) signal pathway. NS2028 (10 mg/kg), a blocker of sGC, was administrated at the onset of hemorrhage, but CORM-3 (4 mg/kg) was infused after resuscitation via femoral vein. Serum alanine aminotransferase (ALT), aspartate aminotransferase (AST) levels, tumor necrosis Factor-α (TNF-α), and interleukin-1β (IL-1β) were measured at 3, 6, 12, and 24 h after HSR, respectively. Six hours post-HSR, liver injury, pyroptosis of Kupffer cells, and expressions in total caspase-1, cleaved caspase-1, gasdermin D (GSDMD) N-terminal fragment, IL-1β, and IL-18 were measured by hematoxylin-eosin (H&E), immunofluorescence and western blot assays, respectively (Fig. 1). The rats exposed to HSR exhibited significant upregulated levels of serum ALT, AST, TNF-α, and IL-1β, elevated liver injury score, increased pyroptosis of Kupffer cells, and accumulated expressions of pyroptosis-associated protein including cleaved caspase-1, GSDMD N-terminal fragment, IL-1β, and IL-18 than sham-treated rats. However, CORM-3 administration markedly reduced liver injury and pyroptosis of Kupffer cells, whereas these protective effects could be partially blocked by NS2028. CORM-3 can mitigate pyroptosis of Kupffer cells in a blood loss and re-infusion model of rats via sGC-cGMP signal pathway.
    Keywords:  CORM-3; Ischemia/reperfusion injury; Kupffer cells; Liver; Pyroptosis
    DOI:  https://doi.org/10.1007/s10753-021-01419-w
  37. Int Immunol. 2021 Feb 09. pii: dxab006. [Epub ahead of print]
    Kobayashi T, Nguyen-Tien D, Ohshima D, Karyu H, Demoto-Shimabukuro S, Sugitani-Yoshida R, Toyama-Sorimachi N.
      SLC15A4 is an endolysosome-resident amino acid transporter that regulates innate immune responses, and is genetically associated with inflammatory diseases such as systemic lupus erythematosus (SLE) and colitis. SLC15A4-deficient mice showed the amelioration of symptoms of these model diseases, and thus SLC15A4 is a promising therapeutic target of SLE and colitis. For developing SLC15A4-based therapeutic strategy, understanding human SLC15A4's property is essential. Here we characterized human SLC15A4 and demonstrated that human SLC15A4 possessed pH- and temperature-dependent activity for the transportation of dipeptide or tripeptide. Human SLC15A4 localized in LAMP1 + compartments and constitutively associated with Raptor and LAMTORs. We also investigated SLC15A4's role in inflammatory responses using human plasmacytoid dendritic cell line, CAL-1. Knock-down (KD) of SLC15A4 gene in CAL-1 (SLC15A4-KD CAL1) impaired TLR7/8 or TLR9-triggered type I interferon (IFN-I) production and mTORC1 activity, indicating that human SLC15A4 is critical for TLR7/8/9-mediated inflammatory signaling. We also examined SLC15A4's role in autophagy response since SLC15A4 loss caused the decrease of mTORC1 activity, which greatly influences on autophagy. We found that SLC15A4 was not required for autophagy induction, but was critical for autophagy sustainability. Notably, SLC15A4-KD CAL1 severely decreased mitochondria membrane potential in the starvation condition. Our findings revealed that SLC15A4 plays a key role in mitochondria integrity in human cells, which might benefit immune cells to fulfill their functions in inflammatory milieu.
    DOI:  https://doi.org/10.1093/intimm/dxab006
  38. Cell Death Discov. 2020 Jul 22. 6(1): 62
    Yang Y, Ying G, Wu F, Chen Z.
      Liver failure (LF) is a monocyte/macrophage-mediated liver injury that has been associated with inflammatory mediators. However, the mechanism through which monocytes/macrophages regulate LF has not been fully elucidated. In this study, we investigated the role of soluble T-cell immunoglobulin domain and mucin domain-containing molecule-3 (sTim-3) in inhibition of release of inflammatory mediators. We further assess this role in protection against D-galactosamine (D-GalN)/lipopolysaccharide (LPS)-induced acute liver failure (ALF), via monocyte/macrophage regulation and autophagy induction in mice. Our findings indicate significantly higher plasma sTim-3 in acute-on-chronic liver failure (ACLF) group relative to other groups, with this trend associated with disease progression. Furthermore, infiltrated recombinant sTim-3 inhibited release of various inflammatory mediators, including cytokines and human high-mobility group box-1 (HMGB1), potentially via autophagy induction. Furthermore, H&E staining and the low levels of alanine aminotransferase (ALT) and aspartate aminotransferase (AST) in ALF mice, supported that recombinant sTim-3 effectively alleviated liver injury. Moreover, sTim-3 induced changes in monocyte/macrophage population in mice's liver or blood, which consequently caused a reduction in proinflammatory CD11bhiF4/80lo monocyte-derived macrophages and Ly-6C(+)CD11b(+) monocytes. Conversely, sTim-3 increased autophagy levels of hepatic CD11b(+) monocyte-derived macrophages and decreased apoptosis rate of CD11b (+) monocytes in the blood. Collectively, our findings demonstrated that sTim-3 alleviated inflammatory response and liver injury by promoting autophagy and regulating monocyte/macrophage function. This indicates its potential for future development of novel therapeutic strategies against LF.
    DOI:  https://doi.org/10.1038/s41420-020-00299-7
  39. Blood Adv. 2021 Jan 12. 5(1): 26-38
    Surace L, Doisne JM, Escoll P, Marie S, Dardalhon V, Croft C, Thaller A, Topazio D, Sparaneo A, Cama A, Musumeci O, d'Ecclesia A, Buchrieser C, Taylor N, Di Santo JP.
      Distinct metabolic demands accompany lymphocyte differentiation into short-lived effector and long-lived memory cells. How bioenergetics processes are structured in innate natural killer (NK) cells remains unclear. We demonstrate that circulating human CD56Dim (NKDim) cells have fused mitochondria and enhanced metabolism compared with CD56Br (NKBr) cells. Upon activation, these 2 subsets showed a dichotomous response, with further mitochondrial potentiation in NKBr cells vs paradoxical mitochondrial fission and depolarization in NKDim cells. The latter effect impaired interferon-γ production, but rescue was possible by inhibiting mitochondrial fragmentation, implicating mitochondrial polarization as a central regulator of NK cell function. NKDim cells are heterogeneous, and mitochondrial polarization was associated with enhanced survival and function in mature NKDim cells, including memory-like human cytomegalovirus-dependent CD57+NKG2C+ subsets. In contrast, patients with genetic defects in mitochondrial fusion had a deficiency in adaptive NK cells, which had poor survival in culture. These results support mitochondrial polarization as a central regulator of mature NK cell fitness.
    DOI:  https://doi.org/10.1182/bloodadvances.2020003458