bims-mikwok Biomed News
on Mitochondrial quality control
Issue of 2020‒12‒06
33 papers selected by
Avinash N. Mukkala
University of Toronto


  1. Front Cell Dev Biol. 2020 ;8 594203
    Xie Y, Liu J, Kang R, Tang D.
      Mitochondria are multifunctional organelles that regulate cancer biology by synthesizing macromolecules, producing energy, and regulating cell death. The understanding of mitochondrial morphology, function, biogenesis, fission and fusion kinetics, and degradation is important for the development of new anticancer strategies. Mitophagy is a type of selective autophagy that can degrade damaged mitochondria under various environmental stresses, especially oxidative damage and hypoxia. The key regulator of mitophagy is the autophagy receptor, which recognizes damaged mitochondria and allows them to enter autophagosomes by binding to MAP1LC3 or GABARAP, and then undergo lysosomal-dependent degradation. Many components of mitochondria, including mitochondrial membrane proteins (e.g., PINK1, BNIP3L, BNIP3, FUNDC1, NIPSNAP1, NIPSNAP2, BCL2L13, PHB2, and FKBP8) and lipids (e.g., cardiolipin and ceramides), act as mitophagy receptors in a context-dependent manner. Dysfunctional mitophagy not only inhibits, but also promotes, tumorigenesis. Similarly, mitophagy plays a dual role in chemotherapy, radiotherapy, and immunotherapy. In this review, we summarize the latest advances in the mechanisms of mitophagy and highlight the pathological role of mitophagy receptors in tumorigenesis and treatment.
    Keywords:  autophagy; cancer; cell death; mitochondria; mitophagy
    DOI:  https://doi.org/10.3389/fcell.2020.594203
  2. Front Physiol. 2020 ;11 541040
    Glancy B, Kim Y, Katti P, Willingham TB.
      Mitochondria are key determinants of cellular health. However, the functional role of mitochondria varies from cell to cell depending on the relative demands for energy distribution, metabolite biosynthesis, and/or signaling. In order to support the specific needs of different cell types, mitochondrial functional capacity can be optimized in part by modulating mitochondrial structure across several different spatial scales. Here we discuss the functional implications of altering mitochondrial structure with an emphasis on the physiological trade-offs associated with different mitochondrial configurations. Within a mitochondrion, increasing the amount of cristae in the inner membrane improves capacity for energy conversion and free radical-mediated signaling but may come at the expense of matrix space where enzymes critical for metabolite biosynthesis and signaling reside. Electrically isolating individual cristae could provide a protective mechanism to limit the spread of dysfunction within a mitochondrion but may also slow the response time to an increase in cellular energy demand. For individual mitochondria, those with relatively greater surface areas can facilitate interactions with the cytosol or other organelles but may be more costly to remove through mitophagy due to the need for larger phagophore membranes. At the network scale, a large, stable mitochondrial reticulum can provide a structural pathway for energy distribution and communication across long distances yet also enable rapid spreading of localized dysfunction. Highly dynamic mitochondrial networks allow for frequent content mixing and communication but require constant cellular remodeling to accommodate the movement of mitochondria. The formation of contact sites between mitochondria and several other organelles provides a mechanism for specialized communication and direct content transfer between organelles. However, increasing the number of contact sites between mitochondria and any given organelle reduces the mitochondrial surface area available for contact sites with other organelles as well as for metabolite exchange with cytosol. Though the precise mechanisms guiding the coordinated multi-scale mitochondrial configurations observed in different cell types have yet to be elucidated, it is clear that mitochondrial structure is tailored at every level to optimize mitochondrial function to meet specific cellular demands.
    Keywords:  cristae; energetics; mitochondria; mitochondrial dynamics; mitochondrial networks; organelle interaction
    DOI:  https://doi.org/10.3389/fphys.2020.541040
  3. Cells. 2020 Nov 25. pii: E2542. [Epub ahead of print]9(12):
    Gouriou Y, Alam MR, Harhous Z, Crola Da Silva C, Baetz DB, Badawi S, Lefai E, Rieusset J, Durand A, Harisseh R, Gharib A, Ovize M, Bidaux G.
      Following a prolonged exposure to hypoxia-reoxygenation, a partial disruption of the ER-mitochondria tethering by mitofusin 2 (MFN2) knock-down decreases the Ca2+ transfer between the two organelles limits mitochondrial Ca2+ overload and prevents the Ca2+-dependent opening of the mitochondrial permeability transition pore, i.e., limits cardiomyocyte cell death. The impact of the metabolic changes resulting from the alteration of this Ca2+crosstalk on the tolerance to hypoxia-reoxygenation injury remains partial and fragmented between different field of expertise. >In this study, we report that MFN2 loss of function results in a metabolic switch driven by major modifications in energy production by mitochondria. During hypoxia, mitochondria maintain their ATP concentration and, concomitantly, the inner membrane potential by importing cytosolic ATP into mitochondria through an overexpressed ANT2 protein and by decreasing the expression and activity of the ATP hydrolase via IF1. This adaptation further blunts the detrimental hyperpolarisation of the inner mitochondrial membrane (IMM) upon re-oxygenation. These metabolic changes play an important role to attenuate cell death during a prolonged hypoxia-reoxygenation challenge.
    Keywords:  ANT2; ATP; ATP synthase; IF1; bioenergetics; hypoxia; metabolism; mitochondria-associated membranes; mitochondrial membrane potential; mitofusin 2
    DOI:  https://doi.org/10.3390/cells9122542
  4. Cell Calcium. 2020 Nov 22. pii: S0143-4160(20)30164-0. [Epub ahead of print]93 102322
    Feno S, Rizzuto R, Raffaello A, Vecellio Reane D.
      The role of mitochondria in regulating cellular Ca2+ homeostasis is crucial for the understanding of different cellular functions in physiological and pathological conditions. Nevertheless, the study of this aspect was severely limited by the lack of the molecular identity of the proteins responsible for mitochondrial Ca2+ uptake. In 2011, the discovery of the gene encoding for the Mitochondrial Calcium Uniporter (MCU), the selective channel responsible for mitochondrial Ca2+ uptake, gave rise to an explosion of studies aimed to characterize the composition, the regulation of the channel and its pathophysiological roles. Here, we summarize the recent discoveries on the molecular structure and composition of the MCU complex by providing new insights into the mechanisms that regulate MCU channel activity.
    Keywords:  Calcium homeostasis; Mitochondria; Mitochondrial Calcium Uniporter
    DOI:  https://doi.org/10.1016/j.ceca.2020.102322
  5. Cells. 2020 Nov 26. pii: E2544. [Epub ahead of print]9(12):
    Shuler KT, Wilson BE, Muñoz ER, Mitchell AD, Selsby JT, Hudson MB.
      Muscle stem cells (MuSCs) hold great potential as a regenerative therapeutic but have met numerous challenges in treating systemic muscle diseases. Muscle stem cell-derived extracellular vesicles (MuSC-EVs) may overcome these limitations. We assessed the number and size distribution of extracellular vesicles (EVs) released by MuSCs ex vivo, determined the extent to which MuSC-EVs deliver molecular cargo to myotubes in vitro, and quantified MuSC-EV-mediated restoration of mitochondrial function following oxidative injury. MuSCs released an abundance of EVs in culture. MuSC-EVs delivered protein cargo into myotubes within 2 h of incubation. Fluorescent labeling of intracellular mitochondria showed co-localization of delivered protein and mitochondria. Oxidatively injured myotubes demonstrated a significant decline in maximal oxygen consumption rate and spare respiratory capacity relative to untreated myotubes. Remarkably, subsequent treatment with MuSC-EVs significantly improved maximal oxygen consumption rate and spare respiratory capacity relative to the myotubes that were damaged but received no subsequent treatment. Surprisingly, MuSC-EVs did not affect mitochondrial function in undamaged myotubes, suggesting the cargo delivered is able to repair but does not expand the existing mitochondrial network. These data demonstrate that MuSC-EVs rapidly deliver proteins into myotubes, a portion of which co-localizes with mitochondria, and reverses mitochondria dysfunction in oxidatively-damaged myotubes.
    Keywords:  cachexia; extracellular vesicles; mitochondria; muscle stem cells; muscular dystrophy; oxidative stress; skeletal muscle
    DOI:  https://doi.org/10.3390/cells9122544
  6. Acta Physiol (Oxf). 2020 Dec 03. e13590
    Zhu H, Toan S, Mui D, Zhou H.
      Myocardial infarction (MI) is a leading cause of morbidity and mortality worldwide. Because mitochondrial dysfunction critically contributes to the pathogenesis of MI, intensive research is focused on the development of therapeutic strategies targeting mitochondrial homeostasis. Mitochondria possess a quality control system which maintains and restores their structure and function by regulating mitochondrial fission, fusion, biogenesis, degradation, and death. In response to slight damage such as transient hypoxia or mild oxidative stress, mitochondrial metabolism shifts from oxidative phosphorylation to glycolysis, in order to reduce oxygen consumption and maintain ATP output. Mitochondrial dynamics are also activated to modify mitochondrial shape and structure, in order to meet cardiomyocyte energy requirements through augmenting or reducing mitochondrial mass. When damaged mitochondria cannot be repaired, poorly structured mitochondria will be degraded through mitophagy, a process which is often accompanied by mitochondrial biogenesis. Once the insult is severe enough to induce lethal damage in the mitochondria and the cell, mitochondrial death pathway activation is an inevitable consequence, and the cardiomyocyte apoptosis or necrosis program will be initiated to remove damaged cells. Mitochondrial quality surveillance is a hierarchical system preserving mitochondrial function and defending cardiomyocytes against stress. A failure of this system has been regarded as one of the potential pathologies underlying MI. In this review, we discuss the recent findings focusing on the role of mitochondrial quality surveillance in MI, and highlight the available therapeutic approaches targeting mitochondrial quality surveillance during MI.
    Keywords:  Mitochondrial quality surveillance; fission; fusion; mitochondria-dependent cell death; mitophagy; myocardial infarction
    DOI:  https://doi.org/10.1111/apha.13590
  7. Biol Open. 2020 Dec 02. pii: bio054262. [Epub ahead of print]9(12):
    Wang LJ, Hsu T, Lin HL, Fu CY.
      The mitochondrial contact site and cristae organizing system (MICOS) is a multi-protein interaction hub that helps define mitochondrial ultrastructure. While the functional importance of MICOS is mostly characterized in yeast and mammalian cells in culture, the contributions of MICOS to tissue homeostasis in vivo remain further elucidation. In this study, we examined how knocking down expression of Drosophila MICOS genes affects mitochondrial function and muscle tissue homeostasis. We found that CG5903/MIC26-MIC27 colocalizes and functions with Mitofilin/MIC60 and QIL1/MIC13 as a Drosophila MICOS component; knocking down expression of any of these three genes predictably altered mitochondrial morphology, causing loss of cristae junctions, and disruption of cristae packing. Furthermore, the knockdown flies exhibited low mitochondrial membrane potential, fusion/fission imbalances, increased mitophagy, and limited cell death. Reductions in climbing ability indicated deficits in muscle function. Knocking down MICOS genes also caused reduced mtDNA content and fragmented mitochondrial nucleoid structure in Drosophila Together, our data demonstrate an essential role of Drosophila MICOS in maintaining proper homeostasis of mitochondrial structure and function to promote the function of muscle tissue.
    Keywords:  Drosophila; MICOS; Mitochondria
    DOI:  https://doi.org/10.1242/bio.054262
  8. PLoS One. 2020 ;15(12): e0231064
    Oikawa Y, Izumi R, Koide M, Hagiwara Y, Kanzaki M, Suzuki N, Kikuchi K, Matsuhashi T, Akiyama Y, Ichijo M, Watanabe S, Toyohara T, Suzuki T, Mishima E, Akiyama Y, Ogata Y, Suzuki C, Hayashi H, Kodama EN, Hayashi KI, Itoi E, Aoki M, Kure S, Abe T.
      Sporadic inclusion body myositis (sIBM) is the most common idiopathic inflammatory myopathy, and several reports have suggested that mitochondrial abnormalities are involved in its etiology. We recruited 9 sIBM patients and found significant histological changes and an elevation of growth differential factor 15 (GDF15), a marker of mitochondrial disease, strongly suggesting the involvement of mitochondrial dysfunction. Bioenergetic analysis of sIBM patient myoblasts revealed impaired mitochondrial function. Decreased ATP production, reduced mitochondrial size and reduced mitochondrial dynamics were also observed in sIBM myoblasts. Cell vulnerability to oxidative stress also suggested the existence of mitochondrial dysfunction. Mitochonic acid-5 (MA-5) increased the cellular ATP level, reduced mitochondrial ROS, and provided protection against sIBM myoblast death. MA-5 also improved the survival of sIBM skin fibroblasts as well as mitochondrial morphology and dynamics in these cells. The reduction in the gene expression levels of Opa1 and Drp1 was also reversed by MA-5, suggesting the modification of the fusion/fission process. These data suggest that MA-5 may provide an alternative therapeutic strategy for treating not only mitochondrial diseases but also sIBM.
    DOI:  https://doi.org/10.1371/journal.pone.0231064
  9. Mitochondrion. 2020 Nov 27. pii: S1567-7249(20)30219-1. [Epub ahead of print]
    Bhansali S, Sohi K, Dhawan V.
      BACKGROUND: Vascular remodeling plays a pivotal role in regulating hypoxia-mediated pulmonary and systemic hypertension via the phenotypic modulation of smooth muscle cells (SMCs) of pulmonary and systemic arteries. Mitochondria serve as putative oxygen (O2) sensors, and consequently, adaptations to hypoxia are mediated via HIF (hypoxia-inducible factors) activation, which impinges on mitochondrial function by suppressing the mitochondrial activity. Therefore, we explored the implication of hypoxia-mediated mitochondrial stress in pulmonary and systemic arterial remodeling.METHODS: The hypoxic (10% O2) effect on human pulmonary artery and aortic SMCs was examined in vitro by cell viability assay, proliferation index, autophagy, and comet assays. Mitochondrial ROS (mtROS), membrane potential (MMP), and mitochondrial morphology were assessed using mitochondrial-selective fluorescent probes. Further, the cell cycle distribution was analyzed by flow cytometry using propidium iodide staining.
    RESULTS: Our data indicate no significant alterations in cell viability and active proliferation of hypoxic PASMCs; however, an excessive rise in mtROS production and disrupted MMP, accompanied by enhanced DNA damage and reduced autophagy was observed, highlighting the 'apoptosis resistance' phenotype in these cells. Conversely, in hypoxia-treated hASMCs, a modest rise in mtROS levels was associated with reduced DNA damage; followed by upregulated autophagy; increased S-phase DNA content and cell viability, depicting the cytoprotective effect of hypoxia-induced autophagy against mitochondrial damage in hASMCs.
    CONCLUSION: Our findings suggest that differential impact of mtROS on proliferative capacity may contribute to the variable hypoxic responses in pulmonary and systemic vasculature. Therefore, targeting mtROS may serve as an effective therapeutic strategy to prevent hypoxia-induced hypertension.
    Keywords:  Pulmonary vasculature; hypoxia; mitochondrial reactive oxygen species (mtROS); systemic vasculature; vascular remodeling
    DOI:  https://doi.org/10.1016/j.mito.2020.11.012
  10. Results Probl Cell Differ. 2020 ;69 253-280
    Kaczanowski S.
      The progress of evolutionary biology has revealed that symbiosis played a basic role in the evolution of complex eukaryotic organisms, including humans. Mitochondria are actually simplified endosymbiotic bacteria currently playing the role of cellular organelles. Mitochondrial domestication occurred at the very beginning of eukaryotic evolution. Mitochondria have two different basic functions: they produce energy using oxidative respiration, and they initiate different forms of apoptotic programmed/regulated cell death. Apoptotic programmed cell death may have different cytological forms. Mechanisms of apoptotic programmed cell death exist even in the unicellular organisms, and they play a basic role in the development of complex multicellular organisms, such as fungi, green plants, and animals. Multicellularity was independently established many times among eukaryotes. There are indications that apoptotic programmed cell death is a trait required for the establishment of multicellularity. Regulated cell death is initiated by many different parallel biochemical pathways. It is generally accepted that apoptosis evolved during mitochondrial domestication. However, there are different hypothetical models of the origin of apoptosis. The phylogenetic studies of my group indicate that apoptosis probably evolved during an evolutionary arms race between host ancestral eukaryotic predators and ancestral prey mitochondria (named protomitochondria). Protomitochondrial prey produced many different toxins as a defense against predators. From these toxins evolved extant apoptotic factors. There are indications that aerobic respiration and apoptosis co-evolved and are functionally linked in extant organisms. Perturbations of apoptosis and oxidative respiration are frequently observed during neoplastic transition. Our group showed that perturbations of apoptosis in yeasts also cause perturbations of oxidative respiration.
    DOI:  https://doi.org/10.1007/978-3-030-51849-3_10
  11. FEBS Lett. 2020 Nov 28.
    Yuan Q, Chen M, Yang W, Xiao B.
      The morphological structure and metabolic activity of mitochondria are coordinately regulated by circadian mechanisms. However, the mechanistic interplay between circadian mechanisms and mitochondrial architecture remains poorly understood. Here, we demonstrate circadian rhythmicity of Rheb protein in liver, in line with that of Per2. Using genetic mouse models, we show that Rheb, a small GTPase that binds mTOR, is critical for circadian oscillation of mTORC1 activity in liver. Disruption of Rheb oscillation in hepatocytes by persistent expression of Rheb transgene interrupted mTORC1 oscillation. We further show that Rheb-regulated mTORC1 altered mitochondrial fission factor DRP1 in liver, leading to altered mitochondrial dynamics. Our results suggest that Rheb/mTORC1 regulated DRP1 oscillation involves ubiquitin-mediated proteolysis. This study identifies Rheb as a nodal point that couples circadian clock and mitochondrial architecture for optimal mitochondrial metabolism.
    Keywords:  Circadian clock; DRP1; Rheb/mTORC1; mitochondrial dynamics
    DOI:  https://doi.org/10.1002/1873-3468.14009
  12. Int J Mol Sci. 2020 Nov 28. pii: E9066. [Epub ahead of print]21(23):
    Moras M, Hattab C, Gonzalez-Menendez P, Martino S, Larghero J, Le Van Kim C, Kinet S, Taylor N, Lefevre SD, Ostuni MA.
      Translocator protein (TSPO) and voltage dependent anion channels (VDAC) are two proteins forming a macromolecular complex in the outer mitochondrial membrane that is involved in pleiotropic functions. Specifically, these proteins were described to regulate the clearance of damaged mitochondria by selective mitophagy in non-erythroid immortalized cell lines. Although it is well established that erythroblast maturation in mammals depends on organelle clearance, less is known about mechanisms regulating this clearance throughout terminal erythropoiesis. Here, we studied the effect of TSPO1 downregulation and the action of Ro5-4864, a drug ligand known to bind to the TSPO/VDAC complex interface, in ex vivo human terminal erythropoiesis. We found that both treatments delay mitochondrial clearance, a process associated with reduced levels of the PINK1 protein, which is a key protein triggering canonical mitophagy. We also observed that TSPO1 downregulation blocks erythroblast maturation at the orthochromatic stage, decreases the enucleation rate, and increases cell death. Interestingly, TSPO1 downregulation does not modify reactive oxygen species (ROS) production nor intracellular adenosine triphosphate (ATP) levels. Ro5-4864 treatment recapitulates these phenotypes, strongly suggesting an active role of the TSPO/VDAC complex in selective mitophagy throughout human erythropoiesis. The present study links the function of the TSPO/VDAC complex to the PINK1/Parkin-dependent mitophagy induction during terminal erythropoiesis, leading to the proper completion of erythroid maturation.
    Keywords:  PINK1; TSPO1; VDAC; enucleation; erythropoiesis; mitophagy
    DOI:  https://doi.org/10.3390/ijms21239066
  13. PLoS One. 2020 ;15(11): e0242695
    Ramakrishnan RK, Bajbouj K, Hachim MY, Mogas AK, Mahboub B, Olivenstein R, Hamoudi R, Halwani R, Hamid Q.
      BACKGROUND: Sub-epithelial fibrosis is a characteristic feature of airway remodeling in asthma which correlates with disease severity. Current asthma medications are ineffective in treating fibrosis. In this study, we aimed to investigate the mitochondrial phenotype in fibroblasts isolated from airway biopsies of non-asthmatic and severe asthmatic subjects by examining mitophagy as a mechanism contributing to fibroblast persistence and thereby, fibrosis in severe asthma.METHODS: Bioinformatics analysis of publicly available transcriptomic data was performed to identify the top enriched pathways in asthmatic fibroblasts. Endogenous expression of mitophagy markers in severe asthmatic and non-asthmatic fibroblasts was determined using qRT-PCR, western blot and immunofluorescence. Mitophagy flux was examined by using lysosomal protease inhibitors, E64d and pepstatin A. Mitochondrial membrane potential and metabolic activity were also evaluated using JC-1 assay and MTT assay, respectively.
    RESULTS: Bioinformatics analysis revealed the enrichment of Pink/Parkin-mediated mitophagy in asthmatic fibroblasts compared to healthy controls. In severe asthmatic fibroblasts, the differential expression of mitophagy genes, PINK1 and PRKN, was accompanied by the accumulation of PINK1, Parkin and other mitophagy proteins at baseline. The further accumulation of endogenous LC3BII, p62 and PINK1 in the presence of E64d and pepstatin A in severe asthmatic fibroblasts reinforced their enhanced mitophagy flux. Significantly reduced mitochondrial membrane potential and metabolic activity were also demonstrated at baseline confirming the impairment in mitochondrial function in severe asthmatic fibroblasts. Interestingly, these fibroblasts displayed neither an apoptotic nor senescent phenotype but a pro-fibrotic phenotype with an adaptive survival mechanism triggered by increased AMPKα phosphorylation and mitochondrial biogenesis.
    CONCLUSIONS: Our results demonstrated a role for mitophagy in the pathogenesis of severe asthma where the enhanced turnover of damaged mitochondria may contribute to fibrosis in severe asthma by promoting the persistence and pro-fibrotic phenotype of fibroblasts.
    DOI:  https://doi.org/10.1371/journal.pone.0242695
  14. Sci Rep. 2020 Dec 03. 10(1): 21045
    Polo CC, Fonseca-Alaniz MH, Chen JH, Ekman A, McDermott G, Meneau F, Krieger JE, Miyakawa AA.
      Mitochondria are dynamic organelles that change morphology to adapt to cellular energetic demands under both physiological and stress conditions. Cardiomyopathies and neuronal disorders are associated with structure-related dysfunction in mitochondria, but three-dimensional characterizations of the organelles are still lacking. In this study, we combined high-resolution imaging and 3D electron density information provided by cryo-soft X-ray tomography to characterize mitochondria cristae morphology isolated from murine. Using the linear attenuation coefficient, the mitochondria were identified (0.247 ± 0.04 µm-1) presenting average dimensions of 0.90 ± 0.20 µm in length and 0.63 ± 0.12 µm in width. The internal mitochondria structure was successfully identified by reaching up the limit of spatial resolution of 35 nm. The internal mitochondrial membranes invagination (cristae) complexity was calculated by the mitochondrial complexity index (MCI) providing quantitative and morphological information of mitochondria larger than 0.90 mm in length. The segmentation to visualize the cristae invaginations into the mitochondrial matrix was possible in mitochondria with MCI ≥ 7. Altogether, we demonstrated that the MCI is a valuable quantitative morphological parameter to evaluate cristae modelling and can be applied to compare healthy and disease state associated to mitochondria morphology.
    DOI:  https://doi.org/10.1038/s41598-020-78150-3
  15. FEBS Lett. 2020 Nov 28.
    Krämer L, Groh C, Herrmann JM.
      Most mitochondrial proteins are synthesized in the cytosol and subsequently translocated as unfolded polypeptides into mitochondria. Cytosolic chaperones maintain precursor proteins in an import-competent state. This post-translational import reaction is under surveillance of the cytosolic ubiquitin-proteasome system, which carries out several distinguishable activities. On the one hand, the proteasome degrades non-productive protein precursors from the cytosol and nucleus, import intermediates that are stuck in mitochondrial translocases, and misfolded or damaged proteins from the outer membrane and the intermembrane space. These surveillance activities of the proteasome are essential for mitochondrial functionality, as well as cellular fitness and survival. On the other hand, the proteasome competes with mitochondria for non-imported cytosolic precursor proteins, which can compromise mitochondrial biogenesis. In order to balance the positive and negative effects of the cytosolic protein quality control system on mitochondria, mitochondrial import efficiency directly regulates the capacity of the proteasome via transcription factor Rpn4 in yeast and nuclear respiratory factor (Nrf) 1 and 2 in animal cells. In this review, we provide a thorough overview of how the proteasome regulates mitochondrial biogenesis.
    Keywords:  Aging; Mitochondria; Mitochondria-Associated Degradation; Mitoprotein-Induced Stress Response; Proteasome; Protein Quality Control; Protein degradation; Rpn4; Ubiquitin
    DOI:  https://doi.org/10.1002/1873-3468.14010
  16. Free Radic Biol Med. 2020 Nov 30. pii: S0891-5849(20)31628-2. [Epub ahead of print]
    Santos JH.
      Epigenetic modifications influence gene expression programs ultimately dictating physiological outcomes. In the past decades, an increasing body of work has demonstrated that the enzymes that deposit and/or remove epigenetic marks on DNA or histones use metabolites as substrates or co-factors, rendering the epigenome sensitive to metabolic changes. In this context, acetyl-CoA and α-ketoglutarate have been recognized as critical for epigenetics, impinging on histone marks and nuclear DNA methylation patterns. Given that these metabolites are primarily generated in the mitochondria through the tricarboxylic acid cycle (TCA), the requirement of proper mitochondrial function for maintenance of the epigenetic landscape seems obvious. Nevertheless, it was not until recently when the epigenomic outcomes of mitochondrial dysfunction were tested, revealing mitochondria's far-reaching impact on epigenetics. This review will focus on data that directly tested the role of mitochondria on the epigenetic landscape, the mechanisms by which mitochondrial dysfunction may dysregulate the epigenome and gene expression, and their potential implications to health and disease.
    DOI:  https://doi.org/10.1016/j.freeradbiomed.2020.11.016
  17. Nucleic Acids Res. 2020 Dec 02. pii: gkaa1132. [Epub ahead of print]
    Lavdovskaia E, Denks K, Nadler F, Steube E, Linden A, Urlaub H, Rodnina MV, Richter-Dennerlein R.
      Translation and ribosome biogenesis in mitochondria require auxiliary factors that ensure rapid and accurate synthesis of mitochondrial proteins. Defects in translation are associated with oxidative phosphorylation deficiency and cause severe human diseases, but the exact roles of mitochondrial translation-associated factors are not known. Here we identify the functions of GTPBP6, a homolog of the bacterial ribosome-recycling factor HflX, in human mitochondria. Similarly to HflX, GTPBP6 facilitates the dissociation of ribosomes in vitro and in vivo. In contrast to HflX, GTPBP6 is also required for the assembly of mitochondrial ribosomes. GTPBP6 ablation leads to accumulation of late assembly intermediate(s) of the large ribosomal subunit containing ribosome biogenesis factors MTERF4, NSUN4, MALSU1 and the GTPases GTPBP5, GTPBP7 and GTPBP10. Our data show that GTPBP6 has a dual function acting in ribosome recycling and biogenesis. These findings contribute to our understanding of large ribosomal subunit assembly as well as ribosome recycling pathway in mitochondria.
    DOI:  https://doi.org/10.1093/nar/gkaa1132
  18. RNA. 2020 Dec 01. pii: rna.077347.120. [Epub ahead of print]
    Scherbakov D, Duscha S, Juskeviciene R, Restelli L, Frank S, Laczko E, Boettger EC.
      We have recently reported on an experimental model of mitochondrial mistranslation conferred by amino acid exchange V338Y in the mitochondrial ribosomal protein MrpS5. Here we used a combination of RNA-Seq and metabolic profiling of homozygous transgenic MrpS5V338Y/V338Y mice to analyze the changes associated with the V338Y mutation in post-mitotic skeletal muscle. Metabolic profiling demonstrated age-dependent metabolic changes in the mutant V338Y animals, which included enhanced levels of age-associated metabolites and which were accompanied by increased glycolysis, lipid desaturation and eicosanoid biosynthesis, and alterations of the pentose phosphate pathway. In addition, transcriptome signatures of aged V338Y mutant muscle pointed to elevated inflammation, likely reflecting the increased levels of bioactive lipids. Our findings indicate that mistranslation-mediated chronic impairment of mitochondrial function affects specific bioenergetic processes in muscle in an age-dependent manner.
    Keywords:  Aging; Metabolome; Misreading; Mitochondria; Skeletal Muscle
    DOI:  https://doi.org/10.1261/rna.077347.120
  19. Liver Transpl. 2020 Dec 01.
    Guo J, Akahoshi T, Mizuta Y, Murata M, Narahara S, Kawano T, Nagao Y, Zhang S, Tomikawa M, Kawanaka H, Hashizume M.
      BACKGROUND AND AIMS: Hepatic ischemia/reperfusion (I/R) injury is a major complication of liver surgery and transplantation, especially in non-alcoholic steatohepatitis (NASH). The mechanism of NASH susceptibility to I/R injury has not been fully clarified. We investigated the role of liver-produced histidine-rich glycoprotein (HRG) in NASH I/R injury.APPROACH AND RESULTS: A NASH mouse model was established using C57Bl/6j mice fed a methionine- and choline-deficient diet (MCD) for 6 weeks. The MCD and standard diet (SD) groups were exposed to 60 min of partial hepatic ischemia with reperfusion. We further evaluated the impact of HRG in this context using HRG knockdown (KD) mice. I/R injury increased HRG expression in the SD group but not in the MCD group after I/R. HRG expression was inversely correlated with neutrophil infiltration and neutrophil extracellular traps (NETs) formation. HRG KD mice showed severe liver injury with neutrophil infiltration and NETs formation. Pre-treatment with supplementary HRG protected against I/R with inhibition of neutrophil infiltration and NETs formation. In vitro, hepatocytes showed that the expression of HRG was upregulated under hypoxia/re-oxygenation conditions, but not in response to oleic acid treated hepatocytes. The decrease in HRG expression in fatty hepatocytes was accompanied by decreased Farnesoid X receptor (FXR) and Hypoxia-inducible factor-2α (HIF-2α) expression.
    CONCLUSIONS: HRG is a hepatoprotective factor during hepatic I/R injury by decreasing neutrophil infiltration and NETs formation. The decrease in HRG is a cause of susceptibility to I/R injury in steatotic livers. Therefore, HRG is a new therapeutic target for minimizing liver damage in NASH patients.
    Keywords:  Nonalcoholic fatty liver disease; histidine-rich glycoprotein; ischemia/reperfusion; neutrophil extracellular-traps
    DOI:  https://doi.org/10.1002/lt.25960
  20. J Cell Sci. 2020 Dec 01. pii: jcs.250241. [Epub ahead of print]
    Chatterjee S, Chakrabarty Y, Banerjee S, Ghosh S, Bhattacharyya SN.
      Defective intracellular trafficking and export of microRNAs have been observed in growth retarded mammalian cells having impaired mitochondrial potential and dynamics. Uncoupling Protein 2 mediated depolarization of mitochondrial membrane also results in progressive sequestration of microRNAs with polysomes and lowered their release via extracellular vesicles. Interestingly, impaired miRNA-trafficking process in growth retarded human cells could be reversed in presence of Genipin an inhibitor of Uncoupling Protein 2. Mitochondrial detethering of endoplasmic reticulum, observed in mitochondria depolarized cells, found to be responsible for defective compartmentalization of translation initiation factor eIF4E to endoplasmic reticulum attached polysomes. It causes retarded translation process accompanied by enhanced retention of miRNAs and target mRNAs with endoplasmic reticulum attached polysomes to restrict extracellular export of miRNAs. Reduced compartment specific activity of mTORC1 complex, the master regulator of protein synthesis, in mitochondria defective or ER- detethered cells, causes reduced phosphorylation of eIF4E-BP1 to prevent eIF-4E targeting to ER attached polysome and microRNA export. These data suggest how mitochondrial membrane potential and dynamics, by affecting mTORC1 activity and compartmentalization, determine sub-cellular localization and export of microRNAs.
    Keywords:  EIF4E and mTORC1; Exosomes; Extracellular vesicles; MiRNA; Mitochondria; P-body; Polysome; Processing bodies
    DOI:  https://doi.org/10.1242/jcs.250241
  21. Cell Death Dis. 2020 Nov 30. 11(11): 1023
    Ploeger C, Huth T, Sugiyanto RN, Pusch S, Goeppert B, Singer S, Tabti R, Hausser I, Schirmacher P, Désaubry L, Roessler S.
      Chromosome 8p is frequently deleted in various cancer entities and has been shown to correlate with poor patient survival. SH2D4A is located on chromosome 8p and prevents the nuclear translocation of the pro-tumorigenic transcription factor STAT3. Here, we investigated the interaction of SH2D4A and STAT3 to shed light on the non-canonical functions of STAT3 in cooperation with the tumor suppressor SH2D4A. Using an immunoprecipitation-mass spectrometry (IP-MS) approach, we identified the mitochondrial scaffold proteins prohibitin 1 (PHB1) and prohibitin 2 (PHB2) among other proteins to potentially bind to SH2D4A. Co-immunoprecipitation and proximity ligation assays confirmed direct interactions of STAT3, PHB1, and SH2D4A in situ and in vitro. In addition, cell fractionation and immunofluorescence staining revealed co-localization of these proteins with mitochondria. These interactions were selectively interrupted by the small molecule and PHB ligand FL3. Furthermore, FL3 led to a reduction of STAT3 protein levels, STAT3 transcriptional activity, and HIF1α protein stabilization upon dimethyloxalylglycine (DMOG) treatment. Besides, mitochondrial fusion and fission markers, L-OPA1, Mfn1, and FIS1, were dysregulated upon FL3 treatment. This dysregulated morphology was accompanied by significant reduction of mitochondrial respiration, thus, FL3 significantly diminished mitochondrial respirational capacity. In contrast, SH2D4A knockout increased mitochondrial respiration, whereas FL3 reversed the effect of SH2D4A knockout. The here described results indicate that the interaction of SH2D4A and PHB1 is involved in the mitochondrial function and integrity. The demonstrated interaction with STAT3, accompanied by its reduction of transcriptional activity, further suggests that SH2D4A is linking STAT3 to its mitochondrial functions, and inhibition of PHB-interaction may have therapeutic effects in tumor cells with STAT3 activation.
    DOI:  https://doi.org/10.1038/s41419-020-03220-3
  22. Aging (Albany NY). 2020 Nov 26. 12
    Yang Y, Du J, Xu R, Shen Y, Yang D, Li , Hu H, Pei H, Yang Y.
      Mitochondrial calcium uptake 1 (MICU1) is a pivotal molecule in maintaining mitochondrial homeostasis under stress conditions. However, it is unclear whether MICU1 attenuates mitochondrial stress in angiotensin II (Ang-II)-induced cardiac hypertrophy or if it has a role in the function of melatonin. Here, small-interfering RNAs against MICU1 or adenovirus-based plasmids encoding MICU1 were delivered into left ventricles of mice or incubated with neonatal murine ventricular myocytes (NMVMs) for 48 h. MICU1 expression was depressed in hypertrophic myocardia and MICU1 knockdown aggravated Ang-II-induced cardiac hypertrophy in vivo and in vitro. In contrast, MICU1 upregulation decreased cardiomyocyte susceptibility to hypertrophic stress. Ang-II administration, particularly in NMVMs with MICU1 knockdown, led to significantly increased reactive oxygen species (ROS) overload, altered mitochondrial morphology, and suppressed mitochondrial function, all of which were reversed by MICU1 supplementation. Moreover, peroxisome proliferator-activated receptor gamma coactivator 1-α (PGC-1α)/MICU1 expression in hypertrophic myocardia increased with melatonin. Melatonin ameliorated excessive ROS generation, promoted mitochondrial function, and attenuated cardiac hypertrophy in control but not MICU1 knockdown NMVMs or mice. Collectively, our results demonstrate that MICU1 attenuates Ang-II-induced cardiac hypertrophy by inhibiting mitochondria-derived oxidative stress. MICU1 activation may be the mechanism underlying melatonin-induced protection against myocardial hypertrophy.
    Keywords:  MICU1; PGC-1α; ROS; cardiac hypertrophy; melatonin
    DOI:  https://doi.org/10.18632/aging.202159
  23. Stem Cell Rev Rep. 2020 Dec 02.
    Calleri A, Roggio D, Navarro-Tableros V, De Stefano N, Pasquino C, David E, Frigatti G, Rigo F, Antico F, Caropreso P, Patrono D, Bruno S, Romagnoli R.
      Hepatic ischemia-reperfusion injury (IRI) is observed in liver transplantation and hepato-biliary surgery and is associated with an inflammatory response. Human liver stem cell-derived extracellular vesicles (HLSC-EV) have been demonstrated to reduce liver damage in different experimental settings by accelerating regeneration and by modulating inflammation. The aim of the present study was to investigate whether HLSC-EV may protect liver from IRI in a mouse experimental model. Segmental IRI was obtained by selective clamping of intrahepatic pedicles for 90 min followed by 6 h of reperfusion. HLSC-EV were administered intravenously at the end of the ischemic period and histopathological and biochemical alterations were evaluated in comparison with controls injected with vehicle alone. Intra liver localization of labeled HLSC-EV was assessed by in in vivo Imaging System (IVIS) and the internalization into hepatocytes was confirmed by fluorescence analyses. As compared to the control group, administration of 3 × 109 particles (EV1 group) significantly reduced alanine aminotransferase (ALT) and lactate dehydrogenase (LDH) release, necrosis extension and cytokines expression (TNF-α, CCL-2 and CXCL-10). However, the administration of an increased dose of HLSC-EV (7.5 × 109 particles, EV2 group) showed no significant improvement in respect to controls at enzyme and histology levels, despite a significantly lower cytokine expression. In conclusion, this study demonstrated that 3 × 109 HLSC-EV were able to modulate hepatic IRI by preserving tissue integrity and by reducing transaminases release and inflammatory cytokines expression. By contrast, a higher dose was ineffective suggesting a restricted window of biological activity. Graphical abstract.
    Keywords:  Adult stem cells; Hepatic inflammation; Ischemia-reperfusion; Liver regeneration; Microvesicles
    DOI:  https://doi.org/10.1007/s12015-020-10078-7
  24. Redox Biol. 2020 Nov 25. pii: S2213-2317(20)31013-2. [Epub ahead of print]38 101808
    Hegedűs C, Juhász T, Fidrus E, Janka EA, Juhász G, Boros G, Paragh G, Uray K, Emri G, Remenyik É, Bai P.
      Ultraviolet B radiation (UVB) is an environmental complete carcinogen, which induces and promotes keratinocyte carcinomas, the most common human malignancies. UVB induces the formation of cyclobutane pyrimidine dimers (CPDs). Repairing CPDs through nucleotide excision repair is slow and error-prone in placental mammals. In addition to the mutagenic and malignancy-inducing effects, UVB also elicits poorly understood complex metabolic changes in keratinocytes, possibly through CPDs. To determine the effects of CPDs, CPD-photolyase was overexpressed in keratinocytes using an N1-methyl pseudouridine-containing in vitro-transcribed mRNA. CPD-photolyase, which is normally not present in placental mammals, can efficiently and rapidly repair CPDs to block signaling pathways elicited by CPDs. Keratinocytes surviving UVB irradiation turn hypermetabolic. We show that CPD-evoked mitochondrial reactive oxygen species production, followed by the activation of several energy sensor enzymes, including sirtuins, AMPK, mTORC1, mTORC2, p53, and ATM, is responsible for the compensatory metabolic adaptations in keratinocytes surviving UVB irradiation. Compensatory metabolic changes consist of enhanced glycolytic flux, Szent-Györgyi-Krebs cycle, and terminal oxidation. Furthermore, mitochondrial fusion, mitochondrial biogenesis, and lipophagy characterize compensatory hypermetabolism in UVB-exposed keratinocytes. These properties not only support the survival of keratinocytes, but also contribute to UVB-induced differentiation of keratinocytes. Our results indicate that CPD-dependent signaling acutely maintains skin integrity by supporting cellular energy metabolism.
    Keywords:  CPD; Keratinocyte; Mitochondria; Photolyase mRNA; UVB
    DOI:  https://doi.org/10.1016/j.redox.2020.101808
  25. Cells. 2020 Nov 28. pii: E2552. [Epub ahead of print]9(12):
    Leal NS, Dentoni G, Schreiner B, Naia L, Piras A, Graff C, Cattaneo A, Meli G, Hamasaki M, Nilsson P, Ankarcrona M.
      Recent findings have shown that the connectivity and crosstalk between mitochondria and the endoplasmic reticulum (ER) at mitochondria-ER contact sites (MERCS) are altered in Alzheimer's disease (AD) and in AD-related models. MERCS have been related to the initial steps of autophagosome formation as well as regulation of mitochondrial function. Here, the interplay between MERCS, mitochondria ultrastructure and function and autophagy were evaluated in different AD animal models with increased levels of Aβ as well as in primary neurons derived from these animals. We start by showing that the levels of Mitofusin 1, Mitofusin 2 and mitochondrial import receptor subunit TOM70 are decreased in post-mortem brain tissue derived from familial AD. We also show that Aβ increases the juxtaposition between ER and mitochondria both in adult brain of different AD mouse models as well as in primary cultures derived from these animals. In addition, the connectivity between ER and mitochondria are also increased in wild-type neurons exposed to Aβ. This alteration in MERCS affects autophagosome formation, mitochondrial function and ATP formation during starvation. Interestingly, the increment in ER-mitochondria connectivity occurs simultaneously with an increase in mitochondrial activity and is followed by upregulation of autophagosome formation in a clear chronological sequence of events. In summary, we report that Aβ can affect cell homeostasis by modulating MERCS and, consequently, altering mitochondrial activity and autophagosome formation. Our data suggests that MERCS is a potential target for drug discovery in AD.
    Keywords:  Alzheimer’s disease; Mitochondria-ER contact sites; amyloid β-peptide; autophagy; mitochondria
    DOI:  https://doi.org/10.3390/cells9122552
  26. Sci Rep. 2020 Dec 03. 10(1): 21135
    Borisova MA, Achasova KM, Morozova KN, Andreyeva EN, Litvinova EA, Ogienko AA, Morozova MV, Berkaeva MB, Kiseleva E, Kozhevnikova EN.
      The disruption of the protective intestinal barrier-the 'leaky gut'-is a common complication of the inflammatory bowel disease. There is limited data on the mechanisms of the intestinal barrier disruption upon low-grade inflammation characteristic of patients with inflammatory bowel disease in clinical remission. Thus, animal models that recapitulate the complexity of chronic intestinal inflammation in vivo are of particular interest. In this study, we used Mucin-2 (Muc2) knockout mice predisposed to colitis to study intestinal barrier upon chronic inflammation. We used 4-kDa FITC-Dextran assay and transmission electron microscopy to demonstrate the increased intestinal permeability and morphological defects in intercellular junctions in Muc2 knockout mice. Confocal microscopy revealed the disruption of the apical F-actin cytoskeleton and delocalization of tight junction protein Claudin-3 from the membrane. We further demonstrate mitochondrial damage, impaired oxygen consumption and the reduction of the intestinal ATP content in Muc2 knockout mice. Finally, we show that chemically induced mitochondrial uncoupling in the wild type mice mimics the intestinal barrier disruption in vivo and causes partial loss of F-actin and membrane localization of Claudin-3. We propose that mitochondrial damage and metabolic shifts during chronic inflammation contribute to the leaky gut syndrome in Muc2 knockout animal model of colitis.
    DOI:  https://doi.org/10.1038/s41598-020-78141-4
  27. PLoS One. 2020 ;15(11): e0238754
    Martinez M, Fendley GA, Saxberg AD, Zoghbi ME.
      Heme biosynthesis occurs through a series of reactions that take place within the cytoplasm and mitochondria, so intermediates need to move across these cellular compartments. However, the specific membrane transport mechanisms involved in the process are not yet identified. The ATP-binding cassette protein ABCB10 is essential for normal heme production, as knocking down this transporter in mice is embryonically lethal and accompanied by severe anemia plus oxidative damage. The role of ABCB10 is unknown, but given its location in the inner mitochondrial membrane, it has been proposed as a candidate to export either an early heme precursor or heme. Alternatively, ABCB10 might transport a molecule important for protection against oxidative damage. To help discern between these possibilities, we decided to study the effect of heme analogs, precursors, and antioxidant peptides on purified human ABCB10. Since substrate binding increases the ATP hydrolysis rate of ABC transporters, we have determined the ability of these molecules to activate purified ABCB10 reconstituted in lipid nanodiscs using ATPase measurements. Under our experimental conditions, we found that the only heme analog increasing ABCB10 ATPase activity was Zinc-mesoporphyrin. This activation of almost seventy percent was specific for ABCB10, as the ATPase activity of a negative control bacterial ABC transporter was not affected. The activation was also observed in cysteine-less ABCB10, suggesting that Zinc-mesoporphyrin's effect did not require binding to typical heme regulatory motifs. Furthermore, our data indicate that ABCB10 was not directly activated by neither the early heme precursor delta-aminolevulinic acid nor glutathione, downsizing their relevance as putative substrates for this transporter. Although additional studies are needed to determine the physiological substrate of ABCB10, our findings reveal Zinc-mesoporphyrin as the first tool compound to directly modulate ABCB10 activity and raise the possibility that some actions of Zinc-mesoporphyrin in cellular and animal studies could be mediated by ABCB10.
    DOI:  https://doi.org/10.1371/journal.pone.0238754
  28. Proc Natl Acad Sci U S A. 2020 Nov 30. pii: 202005877. [Epub ahead of print]
    Aras S, Purandare N, Gladyck S, Somayajulu-Nitu M, Zhang K, Wallace DC, Grossman LI.
      MNRR1 (CHCHD2) is a bi-organellar regulator of mitochondrial function that directly activates cytochrome c oxidase in the mitochondria and functions in the nucleus as a transcriptional activator for hundreds of genes. Since MNRR1 depletion contains features of a mitochondrial disease phenotype, we evaluated the effects of forced expression of MNRR1 on the mitochondrial disease MELAS (mitochondrial encephalomyopathy, lactic acidosis and stroke-like episodes) syndrome. MELAS is a multisystem encephalomyopathy disorder that can result from a heteroplasmic mutation in the mitochondrial DNA (mtDNA; m.3243A > G) at heteroplasmy levels of ∼50 to 90%. Since cybrid cell lines with 73% m.3243A > G heteroplasmy (DW7) display a significant reduction in MNRR1 levels compared to the wild type (0% heteroplasmy) (CL9), we evaluated the effects of MNRR1 levels on mitochondrial functioning. Overexpression of MNRR1 in DW7 cells induces the mitochondrial unfolded protein response (UPRmt), autophagy, and mitochondrial biogenesis, thereby rescuing the mitochondrial phenotype. It does so primarily as a transcription activator, revealing this function to be a potential therapeutic target. The role of MNRR1 in stimulating UPRmt, which is blunted in MELAS cells, was surprising and further investigation uncovered that under conditions of stress the import of MNRR1 into the mitochondria was blocked, allowing the protein to accumulate in the nucleus to enhance its transcription function. In the mammalian system, ATF5, has been identified as a mediator of UPRmt MNRR1 knockout cells display an ∼40% reduction in the protein levels of ATF5, suggesting that MNRR1 plays an important role upstream of this known mediator of UPRmt.
    Keywords:  CHCHD2; cytochrome c oxidase; mitochondria; transcription; unfolded protein response
    DOI:  https://doi.org/10.1073/pnas.2005877117
  29. Sci Rep. 2020 Dec 03. 10(1): 21029
    Rahmel T, Marko B, Nowak H, Bergmann L, Thon P, Rump K, Kreimendahl S, Rassow J, Peters J, Singer M, Adamzik M, Koos B.
      Sepsis is characterized by a dysregulated immune response, metabolic derangements and bioenergetic failure. These alterations are closely associated with a profound and persisting mitochondrial dysfunction. This however occurs despite increased expression of the nuclear-encoded transcription factor A (TFAM) that normally supports mitochondrial biogenesis and functional recovery. Since this paradox may relate to an altered intracellular distribution of TFAM in sepsis, we tested the hypothesis that enhanced extramitochondrial TFAM expression does not translate into increased intramitochondrial TFAM abundance. Accordingly, we prospectively analyzed PBMCs both from septic patients (n = 10) and lipopolysaccharide stimulated PBMCs from healthy volunteers (n = 20). Extramitochondrial TFAM protein expression in sepsis patients was 1.8-fold greater compared to controls (p = 0.001), whereas intramitochondrial TFAM abundance was approximate 80% less (p < 0.001). This was accompanied by lower mitochondrial DNA copy numbers (p < 0.001), mtND1 expression (p < 0.001) and cellular ATP content (p < 0.001) in sepsis patients. These findings were mirrored in lipopolysaccharide stimulated PBMCs taken from healthy volunteers. Furthermore, TFAM-TFB2M protein interaction within the human mitochondrial core transcription initiation complex, was 74% lower in septic patients (p < 0.001). In conclusion, our findings, which demonstrate a diminished mitochondrial TFAM abundance in sepsis and endotoxemia, may help to explain the paradox of lacking bioenergetic recovery despite enhanced TFAM expression.
    DOI:  https://doi.org/10.1038/s41598-020-78195-4
  30. Nat Commun. 2020 12 01. 11(1): 6145
    Girardi E, Agrimi G, Goldmann U, Fiume G, Lindinger S, Sedlyarov V, Srndic I, Gürtl B, Agerer B, Kartnig F, Scarcia P, Noia MAD, Liñeiro E, Rebsamen M, Wiedmer T, Bergthaler A, Palmieri L, Superti-Furga G.
      About a thousand genes in the human genome encode for membrane transporters. Among these, several solute carrier proteins (SLCs), representing the largest group of transporters, are still orphan and lack functional characterization. We reasoned that assessing genetic interactions among SLCs may be an efficient way to obtain functional information allowing their deorphanization. Here we describe a network of strong genetic interactions indicating a contribution to mitochondrial respiration and redox metabolism for SLC25A51/MCART1, an uncharacterized member of the SLC25 family of transporters. Through a combination of metabolomics, genomics and genetics approaches, we demonstrate a role for SLC25A51 as enabler of mitochondrial import of NAD, showcasing the potential of genetic interaction-driven functional gene deorphanization.
    DOI:  https://doi.org/10.1038/s41467-020-19871-x
  31. Autophagy. 2020 Nov 29.
    Singh SR, Meyer-Jens M, Alizoti E, Bacon WC, Davis G, Osinska H, Gulick J, Reischmann-Düsener S, Orthey E, McLendon PM, Molkentin JD, Schlossarek S, Robbins J, Carrier L.
      The ubiquitin-proteasome system (UPS) and autophagy-lysosomal pathway (ALP) are two major protein degradation pathways in eukaryotic cells. Initially considered as two independent pathways, there is emerging evidence that they can work in concert. As alterations of UPS and ALP function can contribute to neurodegenerative disorders, cancer and cardiac disease, there is great interest in finding targets that modulate these catabolic processes. We undertook an unbiased, total genome high-throughput screen to identify novel effectors that regulate both the UPS and ALP. We generated a stable HEK293 cell line expressing a UPS reporter (UbG76V-mCherry) and an ALP reporter (GFP-LC3) and screened for genes for which knockdown increased both UbG76V-mCherry intensity and GFP-LC3 puncta. With stringent selection, we isolated 80 candidates, including the transcription factor ZNF418 (ZFP418 in rodents). After screen validation with Zfp418 overexpression in HEK293 cells, we evaluated Zfp418 knockdown and overexpression in neonatal rat ventricular myocytes (NRVMs). Endogenous and overexpressed ZFP418 were localized in the nucleus. Subsequent experiments showed that ZFP418 negatively regulates UPS and positively regulates ALP activity in NRVMs. RNA-seq from Zfp418 knockdown revealed altered gene expression of numerous ubiquitinating and deubiquitinating enzymes, decreased expression of autophagy activators and initiators and increased expression of autophagy inhibitors. We found that ZPF418 activated the promoters of Dapk2 and Fyco1, which are involved in autophagy. RNA-seq from Zfp418 knockdown also revealed accumulation of several genes involved in cardiac development and/or hypertrophy. In conclusion, our study provides evidence that ZNF418 activates the ALP, inhibits the UPS and regulates genes associated with cardiomyocyte structure/function.
    Keywords:  ALP; UPS; ZFP418; ZNF418; autophagy; cardiomyocyte proteasome; protein degradation; screen; ubiquitin
    DOI:  https://doi.org/10.1080/15548627.2020.1856493
  32. Reproduction. 2020 Dec 01. pii: REP-20-0351.R1. [Epub ahead of print]
    Ferreira AF, Soares M, Almeida Reis S, Ramalho-Santos J, Sousa AP, Almeida-Santos T.
      Mitochondrial supplementation was proposed as a complementary treatment to assisted reproductive technologies to improve oocyte competence and support post-fertilization development. This strategy is based on the fact that poor-quality/aged oocytes contain lower and dysfunctional mitochondria. However, the efficacy and safety of mitochondrial supplementation is still controversial. Therefore, this review summarizes the clinical/biological outcomes of mitochondrial supplementation, aiming to improve oocyte competence or explore the safety of this technique, and was based on an online search using PubMed and Web of Science, until September 2019. The studies included reported outcomes related to efficacy and safety of mitochondrial supplementation either in human or animal models (bovine, porcine and mouse). Extracted data were organized according to study objective, the mitochondrial source and the main outcomes: fertilization/pregnancy rates, embryo development and adverse outcomes. Clinical pregnancy was not improved in the only randomized controlled trial published, although an increase was demonstrated in other non-randomized studies. Fertilization rate and embryo development were not different from control groups in the majority of studies, although performed in different contexts and using diverse sources of mitochondria. The safety of mitochondria transfer is still a concern, however, the euploid rate and the absence of reported congenital malformation from the clinical studies are reassuring. In summary, mitochondrial supplementation does not seem to cause harm although the benefit of improving oocyte competence is still unclear due to the diversity of methodological approaches and low-quality of the data available. Analyzed data supports the need to investigate further, in both pre-clinical and clinical contexts.
    DOI:  https://doi.org/10.1530/REP-20-0351
  33. PLoS One. 2020 ;15(12): e0243220
    Feige K, Raupach A, Torregroza C, Muehlenbernd J, Stroethoff M, Bunte S, Hollmann MW, Huhn R.
      Coronary effluent collected from ischemic preconditioning (IPC) treated hearts induces myocardial protection in non-ischemic-preconditioned hearts. So far, little is known about the number of IPC cycles required for the release of cardioprotective factors into the coronary effluent to successfully induce cardioprotection. This study investigated the cardioprotective potency of effluent obtained after various IPC cycles in the rat heart. Experiments were performed on isolated hearts of male Wistar rats, mounted onto a Langendorff system and perfused with Krebs-Henseleit buffer. In a first part, effluent was taken before (Con) and after each IPC cycle (Eff 1, Eff 2, Eff 3). IPC was induced by 3 cycles of 5 min of global myocardial ischemia followed by 5 minutes of reperfusion. In a second part, hearts of male Wistar rats were randomized to four groups (each group n = 4-5) and underwent 33 min of global ischemia followed by 60 min of reperfusion. The previously obtained coronary effluent was administered for 10 minutes before ischemia as a preconditioning stimulus. Infarct size was determined at the end of reperfusion by triphenyltetrazoliumchloride (TTC) staining. Infarct size with control effluent was 54±12%. Effluent obtained after IPC confers a strong infarct size reduction independent of the number of IPC cycles (Eff 1: 27±5%; Eff 2: 35±7%; Eff 3: 35±8%, each P<0.05 vs. Con). Effluent extracted after one cycle IPC is comparably protective as after two or three cycles IPC.
    DOI:  https://doi.org/10.1371/journal.pone.0243220