bims-midmar Biomed News
on Mitochondrial DNA maintenance and replication
Issue of 2021–09–05
nineteen papers selected by
Flavia Söllner, Ludwig-Maximilians University
  1. High levels of TFAM repress mammalian mitochondrial DNA transcription in vivo.
  2. Purification and Cryo-electron Microscopy Analysis of Plant Mitochondrial Ribosomes.
  3. Protocol for assessing real-time changes in mitochondrial morphology, fission and fusion events in live cells using confocal microscopy.
  4. Through DNA sensors and hidden mitochondrial effects of SARS-CoV-2.
  5. Purification of Mitochondrial Ribosomes with the Translocase Oxa1L from HEK Cells.
  6. Potential biomarkers and targets of mitochondrial dynamics.
  7. Cell reprogramming shapes the mitochondrial DNA landscape.
  8. Mechanisms of SSBP1 variants in mitochondrial disease: Molecular dynamics simulations reveal stable tetramers with altered DNA binding surfaces.
  9. Proteomic analysis demonstrates the role of the quality control protease LONP1 in mitochondrial protein aggregation.
  10. Validation of Low Coverage Whole Genome Sequencing for Mitochondrial DNA Variants Suggests Mitochondrial DNA as a Genetic Cause of Preterm Birth.
  11. From OCR and ECAR to energy: perspectives on the design and interpretation of bioenergetics studies.
  12. A globally diverse reference alignment and panel for imputation of mitochondrial DNA variants.
  13. MitoCellPhe reveals mitochondrial morphologies in single fibroblasts and clustered stem cells.
  14. Short-form OPA1 is a molecular chaperone in mitochondrial intermembrane space.
  15. Significance of mitochondrial activity in neurogenesis and neurodegenerative diseases.
  16. Optimization of mito-roGFP protocol to measure mitochondrial oxidative status in human coronary artery endothelial cells.
  17. Leigh Syndrome: A Tale of Two Genomes.
  18. Mitochondrial links between brain aging and Alzheimer's disease.
  19. Effects of Telmisartan, an AT1 receptor antagonist, on mitochondria-specific genes expression in a mouse MPTP model of Parkinsonism.
  1. Life Sci Alliance. 2021 Nov;pii: e202101034. [Epub ahead of print]4(11):
    High levels of TFAM repress mammalian mitochondrial DNA transcription in vivo.
    Nina A Bonekamp, Min Jiang, Elisa Motori, Rodolfo Garcia Villegas, Camilla Koolmeister, Ilian Atanassov, Andrea Mesaros, Chan Bae Park, Nils-Göran Larsson.
      Mitochondrial transcription factor A (TFAM) is compacting mitochondrial DNA (dmtDNA) into nucleoids and directly controls mtDNA copy number. Here, we show that the TFAM-to-mtDNA ratio is critical for maintaining normal mtDNA expression in different mouse tissues. Moderately increased TFAM protein levels increase mtDNA copy number but a normal TFAM-to-mtDNA ratio is maintained resulting in unaltered mtDNA expression and normal whole animal metabolism. Mice ubiquitously expressing very high TFAM levels develop pathology leading to deficient oxidative phosphorylation (OXPHOS) and early postnatal lethality. The TFAM-to-mtDNA ratio varies widely between tissues in these mice and is very high in skeletal muscle leading to strong repression of mtDNA expression and OXPHOS deficiency. In the heart, increased mtDNA copy number results in a near normal TFAM-to-mtDNA ratio and maintained OXPHOS capacity. In liver, induction of LONP1 protease and mitochondrial RNA polymerase expression counteracts the silencing effect of high TFAM levels. TFAM thus acts as a general repressor of mtDNA expression and this effect can be counterbalanced by tissue-specific expression of regulatory factors.
    DOI:  https://doi.org/10.26508/lsa.202101034
  2. Bio Protoc. 2021 Aug 05. 11(15): e4111
    Purification and Cryo-electron Microscopy Analysis of Plant Mitochondrial Ribosomes.
    Florent Waltz, Philippe Giegé, Yaser Hashem.
      Plants make up by far the largest part of biomass on Earth. They are the primary source of food and the basis of most drugs used for medicinal purposes. Similarly to all eukaryotes, plant cells also use mitochondria for energy production. Among mitochondrial gene expression processes, translation is the least understood; although, recent advances have revealed the specificities of its main component, the mitochondrial ribosome (mitoribosome). Here, we present a detailed protocol to extract highly pure cauliflower mitochondria by differential centrifugation for the purification of mitochondrial ribosomes using a sucrose gradient and the preparation of cryo-electron microscopy (cryo-EM) grids. Finally, the specific bioinformatics pipeline used for image acquisition, the processing steps, and the data analysis used for cryo-EM of the plant mitoribosome are described. This protocol will be used for further analysis of the critical steps of mitochondrial translation, such as its initiation and regulation.
    Keywords:  Cryo-EM; Electron microscopy image processing; Mitoribosome; Plant mitochondria; Sucrose density gradient
    DOI:  https://doi.org/10.21769/BioProtoc.4111
  3. STAR Protoc. 2021 Sep 17. 2(3): 100767
    Protocol for assessing real-time changes in mitochondrial morphology, fission and fusion events in live cells using confocal microscopy.
    Irene Bertolini, Frederick Keeney, Dario C Altieri.
      Changes in mitochondrial size, shape, and subcellular position, a process collectively known as mitochondrial dynamics, are exploited for various cancer traits. Modulation of subcellular mitochondrial trafficking and accumulation at the cortical cytoskeleton has been linked to the machinery of cell movements, fueling cell invasion and metastatic spreading. Here, we detail a technique to track changes in mitochondrial volume using a commercial CellLight™ Mitochondria-RFP/GFP reporter and live confocal microscopy. This allows a real-time study of mitochondrial dynamics in live cells. For complete details on the use and execution of this protocol, please refer to Bertolini et al. (2020).
    Keywords:  Cancer; Microscopy; Molecular/Chemical Probes
    DOI:  https://doi.org/10.1016/j.xpro.2021.100767
  4. J Venom Anim Toxins Incl Trop Dis. 2021 ;27 e20200183
    Through DNA sensors and hidden mitochondrial effects of SARS-CoV-2.
    Vitor Pedro Targhetta, Mariana Abrantes Amaral, Niels Olsen Saraiva Camara.
      The COVID-19 pandemic brought attention to studies about viral infections and their impact on the cell machinery. SARS-CoV-2, for example, invades the host cells by ACE2 interaction and possibly hijacks the mitochondria. To better understand the disease and to propose novel treatments, crucial aspects of SARS-CoV-2 enrolment with host mitochondria must be studied. The replicative process of the virus leads to consequences in mitochondrial function, and cell metabolism. The hijacking of mitochondria, on the other hand, can drive the extrusion of mitochondrial DNA (mtDNA) to the cytosol. Extracellular mtDNA evoke robust proinflammatory responses once detected, that may act in different pathways, eliciting important immune responses. However, few receptors are validated and are able to detect and respond to mtDNA. In this review, we propose that the mtDNA and its detection might be important in the immune process generated by SARS-CoV-2 and that this mechanism might be important in the lung pathogenesis seen in clinical symptoms. Therefore, investigating the mtDNA receptors and their signaling pathways might provide important clues for therapeutic interventions.
    Keywords:  Cytokine storm; Innate receptors; Mitochondria; SARS-CoV-2
    DOI:  https://doi.org/10.1590/1678-9199-JVATITD-2020-0183
  5. Bio Protoc. 2021 Aug 05. 11(15): e4110
    Purification of Mitochondrial Ribosomes with the Translocase Oxa1L from HEK Cells.
    Hanting Yang, Nirupa Desai.
      Mitochondrial ribosomes (mitoribosomes) perform protein synthesis inside mitochondria, the organelles responsible for energy conversion and adenosine triphosphate (ATP) production in eukaryotic cells. To investigate their functions and structures, large-scale purification of intact mitoribosomes from mitochondria-rich animal tissues or HEK cells have been developed. However, the fast purification of mitoribosomes anchored to the mitochondrial inner membrane in complex with the Oxa1L translocase remains particularly challenging. Herein, we present a protocol recently developed and modified in our lab that provides details for the efficient isolation of intact mitoribosomes with its translocase Oxa1L. We combined the cell culture of PDE12-/- or wild-type HEK293 cell lines with the isolation of mitochondria and the purification steps used for the biochemical and structural studies of mitoribosomes and Oxa1L. Graphic abstract: Schematic procedure for the purification of mitoribosomes from HEK cells. The protocol described herein includes two main sections: 1) isolation of mitochondria from HEK cells; and 2) purification of mitoribosome-Oxa1L from mitochondria. RB: Resuspension Buffer (see Recipes) (Created with BioRender.com).
    Keywords:  Biochemistry; Cryo-EM; Mitochondria; Mitoribosome; Oxa1L; Ribosome purification
    DOI:  https://doi.org/10.21769/BioProtoc.4110
  6. Clin Transl Med. 2021 Aug;11(8): e529
    Potential biomarkers and targets of mitochondrial dynamics.
    Liyang Li, Ruixue Qi, Linlin Zhang, Yuexin Yu, Jiayun Hou, Yutong Gu, Dongli Song, Xiangdong Wang.
      Mitochondrial dysfunction contributes to the imbalance of cellular homeostasis and the development of diseases, which is regulated by mitochondria-associated factors. The present review aims to explore the process of the mitochondrial quality control system as a new source of the potential diagnostic biomarkers and/or therapeutic targets for diseases, including mitophagy, mitochondrial dynamics, interactions between mitochondria and other organelles (lipid droplets, endoplasmic reticulum, endosomes, and lysosomes), as well as the regulation and posttranscriptional modifications of mitochondrial DNA/RNA (mtDNA/mtRNA). The direct and indirect influencing factors were especially illustrated in understanding the interactions among regulators of mitochondrial dynamics. In addition, mtDNA/mtRNAs and proteomic profiles of mitochondria in various lung diseases were also discussed as an example. Thus, alternations of mitochondria-associated regulators can be a new category of biomarkers and targets for disease diagnosis and therapy.
    Keywords:  lung diseases; mitochondria; mitochondrial dynamics; mtDNA
    DOI:  https://doi.org/10.1002/ctm2.529
  7. Nat Commun. 2021 Sep 02. 12(1): 5241
    Cell reprogramming shapes the mitochondrial DNA landscape.
    Wei Wei, Daniel J Gaffney, Patrick F Chinnery.
      Individual induced pluripotent stem cells (iPSCs) show considerable phenotypic heterogeneity, but the reasons for this are not fully understood. Comprehensively analysing the mitochondrial genome (mtDNA) in 146 iPSC and fibroblast lines from 151 donors, we show that most age-related fibroblast mtDNA mutations are lost during reprogramming. However, iPSC-specific mutations are seen in 76.6% (108/141) of iPSC lines at a mutation rate of 8.62 × 10-5/base pair. The mutations observed in iPSC lines affect a higher proportion of mtDNA molecules, favouring non-synonymous protein-coding and tRNA variants, including known disease-causing mutations. Analysing 11,538 single cells shows stable heteroplasmy in sub-clones derived from the original donor during differentiation, with mtDNA variants influencing the expression of key genes involved in mitochondrial metabolism and epidermal cell differentiation. Thus, the dynamic mtDNA landscape contributes to the heterogeneity of human iPSCs and should be considered when using reprogrammed cells experimentally or as a therapy.
    DOI:  https://doi.org/10.1038/s41467-021-25482-x
  8. DNA Repair (Amst). 2021 Aug 17. pii: S1568-7864(21)00168-3. [Epub ahead of print]107 103212
    Mechanisms of SSBP1 variants in mitochondrial disease: Molecular dynamics simulations reveal stable tetramers with altered DNA binding surfaces.
    Margaret A Gustafson, Lalith Perera, Min Shi, William C Copeland.
      Several mutations in the gene for the mitochondrial single stranded DNA binding protein (SSBP1) have recently been implicated in human disease, but initial reports are insufficient to explain the molecular mechanism of disease, including the possible role of SSBP1 heterotetramers in heterozygous patients. Here we employed molecular simulations to model the dynamics of wild type and 31 variant SSBP1 tetramer systems, including 7 variant homotetramer and 24 representative heterotetramer systems. Our simulations indicate that all variants are stable and most have stronger intermonomer interactions, reduced solvent accessible surface areas, and a net loss of positive surface charge. We then used structural alignments and phosphate binding simulations to predict DNA binding surfaces on SSBP1. Our models suggest that nearly the entire surface of SSBP1, excluding flexible loops and protruding helices, is available for DNA binding, and we observed several potential DNA binding hotspots. Changes to the protein surface in variant SSBP1 tetramers potentially alter anchor points or wrapping paths, rather than abolishing binding altogether. Overall, our findings disqualify tetramer destabilization or gross disruption of DNA binding as mechanisms of disease. Instead, they are consistent with subtle changes to DNA binding, wrapping, or release that cause rare but consequential failures of mtDNA maintenance, which, in turn, are consistent with the late onset of disease in most of the reported SSBP1 cases.
    Keywords:  Human disease variants; Mitochondrial DNA replication; Molecular modeling; Single stranded DNA binding protein; mtSSB
    DOI:  https://doi.org/10.1016/j.dnarep.2021.103212
  9. J Biol Chem. 2021 Aug 27. pii: S0021-9258(21)00935-2. [Epub ahead of print] 101134
    Proteomic analysis demonstrates the role of the quality control protease LONP1 in mitochondrial protein aggregation.
    Karen Pollecker, Marc Sylvester, Wolfgang Voos.
      The mitochondrial matrix protease LONP1 is an essential part of the organellar protein quality control system. LONP1 has been shown to be involved in respiration control and apoptosis. Furthermore, a reduction in LONP1 level correlates with ageing. Up to now, the effects of a LONP1 defect were mostly studied by utilizing transient, siRNA-mediated knockdown approaches. We generated a new cellular model system for studying the impact of LONP1 on mitochondrial protein homeostasis by a CRISPR/Cas-mediated genetic knockdown (gKD). These cells show a stable reduction of LONP1 along with a mild phenotype characterized by absent morphological differences and only small negative effects on mitochondrial functions under normal culture conditions. To assess the consequences of a permanent LONP1 depletion on the mitochondrial proteome, we analyzed the alterations of protein levels by quantitative mass spectrometry, demonstrating small adaptive changes, in particular with respect to mitochondrial protein biogenesis. In an additional proteomic analysis, we determined the temperature-dependent aggregation behavior of mitochondrial proteins and its dependence on a reduction of LONP1 activity, demonstrating the important role of the protease for mitochondrial protein homeostasis in mammalian cells. We identified a significant number of mitochondrial proteins that are affected by LONP1 activity especially with respect to their stress-induced solubility. Taken together, our results suggest a very good applicability of the LONP1 gKD cell line as a model system for human ageing processes.
    Keywords:  Human; LONP1 protease; cell biology; mitochondria; protein aggregation; proteostasis
    DOI:  https://doi.org/10.1016/j.jbc.2021.101134
  10. Hum Mutat. 2021 Sep 01.
    Validation of Low Coverage Whole Genome Sequencing for Mitochondrial DNA Variants Suggests Mitochondrial DNA as a Genetic Cause of Preterm Birth.
    Zeyu Yang, Jesse Slone, Xinjian Wang, Jack Zhan, Yongbo Huang, Bahram Namjou, Kenneth M Kaufman, Michael Pauciulo, John B Harley, Louis J Muglia, Iouri Chepelev, Taosheng Huang.
      Preterm birth (PTB), or birth that occurs earlier than 37 weeks of gestational age, is a major contributor to infant mortality and neonatal hospitalization. Mutations in the mitochondrial genome (mtDNA) have been linked to various rare mitochondrial disorders, and may be a contributing factor in PTB given that maternal genetic factors have been strongly linked to PTB. However, to date, no study has found a conclusive connection between a particular mtDNA variant and PTB. Given the high mtDNA copy number per cell, an automated pipeline was developed for detecting mtDNA variants using low-pass whole genome sequencing (lcWGS) data. The pipeline was first validated against samples of known heteroplasmy, and then applied to 929 samples from a PTB cohort from diverse ethnic backgrounds with an average gestational age of 27.18 weeks (range: 21-30). Our new pipeline successfully identified haplogroups and a large number of mtDNA variants in this large PTB cohort, including 8 samples carrying known pathogenic variants and 47 samples carrying rare mtDNA variants. These results confirm that lcWGS can be utilized to reliably identify mtDNA variants. These mtDNA variants may make a contribution toward preterm birth in a small proportion of live births. This article is protected by copyright. All rights reserved.
    Keywords:  Preterm birth; human genetics; low coverage whole genome sequencing; mitochondrial disease; mitochondrial genome
    DOI:  https://doi.org/10.1002/humu.24279
  11. J Biol Chem. 2021 Aug 27. pii: S0021-9258(21)00941-8. [Epub ahead of print] 101140
    From OCR and ECAR to energy: perspectives on the design and interpretation of bioenergetics studies.
    Cameron A Schmidt, Kelsey H Fisher-Wellman, P Darrell Neufer.
      Biological energy transduction underlies all physiological phenomena in cells. The metabolic systems that support energy transduction have been of great interest due to their association with numerous pathologies including diabetes, cancer, rare genetic diseases, and aberrant cell death. Commercially available bioenergetics technologies (e.g. extracellular flux analysis, high resolution respirometry, fluorescent dye kits, etc.) have made practical assessment of metabolic parameters widely accessible. This has facilitated an explosion in the number of studies exploring, in particular, the biological implications of oxygen consumption rate (OCR) and substrate level phosphorylation via glycolysis (i.e. via extracellular acidification rate (ECAR)). Though these technologies have demonstrated substantial utility and broad applicability to cell biology research, they are also susceptible to historical assumptions, experimental limitations, and other caveats that have led to premature and/or erroneous interpretations. This review enumerates various important considerations for designing and interpreting cellular and mitochondrial bioenergetics experiments, some common challenges and pitfalls in data interpretation, and some potential 'next steps' to be taken that can address these highlighted challenges.
    Keywords:  ATP; Anaerobic glycolysis; Bioenergetics; Cell metabolism; Mitochondria
    DOI:  https://doi.org/10.1016/j.jbc.2021.101140
  12. BMC Bioinformatics. 2021 Sep 01. 22(1): 417
    A globally diverse reference alignment and panel for imputation of mitochondrial DNA variants.
    for the Alzheimer’s Disease Neuroimaging Initiative
       BACKGROUND: Variation in mitochondrial DNA (mtDNA) identified by genotyping microarrays or by sequencing only the hypervariable regions of the genome may be insufficient to reliably assign mitochondrial genomes to phylogenetic lineages or haplogroups. This lack of resolution can limit functional and clinical interpretation of a substantial body of existing mtDNA data. To address this limitation, we developed and evaluated a large, curated reference alignment of complete mtDNA sequences as part of a pipeline for imputing missing mtDNA single nucleotide variants (mtSNVs). We call our reference alignment and pipeline MitoImpute.
    RESULTS: We aligned the sequences of 36,960 complete human mitochondrial genomes downloaded from GenBank, filtered and controlled for quality. These sequences were reformatted for use in imputation software, IMPUTE2. We assessed the imputation accuracy of MitoImpute by measuring haplogroup and genotype concordance in data from the 1000 Genomes Project and the Alzheimer's Disease Neuroimaging Initiative (ADNI). The mean improvement of haplogroup assignment in the 1000 Genomes samples was 42.7% (Matthew's correlation coefficient = 0.64). In the ADNI cohort, we imputed missing single nucleotide variants.
    CONCLUSION: These results show that our reference alignment and panel can be used to impute missing mtSNVs in existing data obtained from using microarrays, thereby broadening the scope of functional and clinical investigation of mtDNA. This improvement may be particularly useful in studies where participants have been recruited over time and mtDNA data obtained using different methods, enabling better integration of early data collected using less accurate methods with more recent sequence data.
    Keywords:  Imputation; Mitochondrial DNA; Reference panel
    DOI:  https://doi.org/10.1186/s12859-021-04337-8
  13. Am J Physiol Cell Physiol. 2021 09 01.
    MitoCellPhe reveals mitochondrial morphologies in single fibroblasts and clustered stem cells.
    Ajibola B Bakare, Fibi Meshrkey, Benjamin Lowe, Carson Molder, Raj R Rao, Justin Zhan, Shilpa Iyer.
      Mitochondria are dynamic organelles that differ significantly in their morphologies across cell types, reflecting specific cellular needs and stages in development. Despite the wide biological significance in disease and health, delineating mitochondrial morphologies in complex systems remains challenging. Here, we present the Mitochondrial Cellular Phenotype (MitoCellPhe) tool developed for quantifying mitochondrial morphologies and demonstrate its utility in delineating differences in mitochondrial morphologies in a human fibroblast and human induced pluripotent stem cell (hiPSC) line. MitoCellPhe generates 24 parameters, allowing for a comprehensive analysis of mitochondrial structures and importantly allows for quantification to be performed on mitochondria in images containing single cells or clusters of cells. With this tool, we were able to validate previous findings that show networks of mitochondria in healthy fibroblast cell lines and a more fragmented morphology in hiPSCs. Using images generated from control and diseased fibroblasts and hiPSCs, we also demonstrate the efficacy of the toolset in delineating differences in morphologies between healthy and the diseased state in both stem cell (hiPSC) and differentiated fibroblast cells. Our results demonstrate that MitoCellPhe enables high-throughput, sensitive, detailed and quantitative mitochondrial morphological assessment and thus enables better biological insights into mitochondrial dynamics in health and disease.
    Keywords:  mitochondria; morphology; networks; stem cells; structure
    DOI:  https://doi.org/10.1152/ajpcell.00231.2021
  14. Sci China Life Sci. 2021 Aug 31.
    Short-form OPA1 is a molecular chaperone in mitochondrial intermembrane space.
    Deyang Yao, Yukun Li, Sheng Zeng, Zhifan Li, Zahir Shah, Bigui Song, Jinglei Liu, Yi Wu, Liang Yang, Qi Long, Wenqian Wang, Zhijuan Hu, Haite Tang, Xingguo Liu.
      Mitochondria, double-membrane organelles, are known to participate in a variety of metabolic and signal transduction pathways. The intermembrane space (IMS) of mitochondria is proposed to subject to multiple damages emanating from the respiratory chain. The optic atrophy 1 (OPA1), an important protein for mitochondrial fusion, is cleaved into soluble short-form (S-OPA1) under stresses. Here we report that S-OPA1 could function as a molecular chaperone in IMS. We purified the S-OPA1 (amino acid sequence after OPA1 isoform 5 S1 site) protein and showed it protected substrate proteins from thermally and chemically induced aggregation and strengthened the thermotolerance of Escherichia coli (E. coli). We also showed that S-OPA1 conferred thermotolerance on IMS proteins, e.g., neurolysin. The chaperone activity of S-OPA1 may be required for maintaining IMS homeostasis in mitochondria.
    Keywords:  OPA1; chaperone; heat shock; mitochondria; mitochondrial homeostasis
    DOI:  https://doi.org/10.1007/s11427-021-1962-0
  15. Neural Regen Res. 2022 Apr;17(4): 741-747
    Significance of mitochondrial activity in neurogenesis and neurodegenerative diseases.
    Serra Ozgen, Judith Krigman, Ruohan Zhang, Nuo Sun.
      Mitochondria play a multidimensional role in the function and the vitality of the neurological system. From the generation of neural stem cells to the maintenance of neurons and their ultimate demise, mitochondria play a critical role in regulating our neural pathways' homeostasis, a task that is critical to our cognitive health and neurological well-being. Mitochondria provide energy via oxidative phosphorylation for the neurotransmission and generation of an action potential along the neuron's axon. This paper will first review and examine the molecular subtleties of the mitochondria's role in neurogenesis and neuron vitality, as well as outlining the impact of defective mitochondria in neural aging. The authors will then summarize neurodegenerative diseases related to either neurogenesis or homeostatic dysfunction. Because of the significant detriment neurodegenerative diseases have on the quality of life, it is essential to understand their etiology and ongoing molecular mechanics. The mitochondrial role in neurogenesis and neuron vitality is essential. Dissecting and understanding this organelle's role in the genesis and homeostasis of neurons should assist in finding pharmaceutical targets for neurodegenerative diseases.
    Keywords:  Alzheimer’s disease; PINK1; Parkin; Parkinson’s disease; autophagy; mitochondria; mitophagy; neural stem cells; neurodegenerative diseases; neurogenesis
    DOI:  https://doi.org/10.4103/1673-5374.322429
  16. STAR Protoc. 2021 Sep 17. 2(3): 100753
    Optimization of mito-roGFP protocol to measure mitochondrial oxidative status in human coronary artery endothelial cells.
    Rayane Brinck Teixeira, Catherine Karbasiafshar, Mohamed Sabra, M Ruhul Abid.
      Reactive oxygen species (ROS) are implicated in endothelial dysfunction and cardiovascular disease. Endothelial cells (ECs) produce most ATP through glycolysis rather than oxidative phosphorylation; thus mitochondrial ROS production is lower than in other cell types. This makes quantification of changes in EC mitochondrial oxidative status challenging. Here, we present an optimized protocol using mitochondrial-targeted adenovirus-based redox sensor for ratiometric quantification of specific changes in mitochondrial ROS in live human coronary artery EC. For complete details on the use and execution of this protocol, please refer to Waypa et al. (2010); Liao et al. (2020); Gao et al. (2021).
    Keywords:  cell-based assays; metabolism; microscopy; molecular/chemical probes; single cell
    DOI:  https://doi.org/10.1016/j.xpro.2021.100753
  17. Front Physiol. 2021 ;12 693734
    Leigh Syndrome: A Tale of Two Genomes.
    Ajibola B Bakare, Edward J Lesnefsky, Shilpa Iyer.
      Leigh syndrome is a rare, complex, and incurable early onset (typically infant or early childhood) mitochondrial disorder with both phenotypic and genetic heterogeneity. The heterogeneous nature of this disorder, based in part on the complexity of mitochondrial genetics, and the significant interactions between the nuclear and mitochondrial genomes has made it particularly challenging to research and develop therapies. This review article discusses some of the advances that have been made in the field to date. While the prognosis is poor with no current substantial treatment options, multiple studies are underway to understand the etiology, pathogenesis, and pathophysiology of Leigh syndrome. With advances in available research tools leading to a better understanding of the mitochondria in health and disease, there is hope for novel treatment options in the future.
    Keywords:  Leigh syndrome; mitochondria; mitochondrial DNA; mitochondrial genetics; respiratory chain complex
    DOI:  https://doi.org/10.3389/fphys.2021.693734
  18. Transl Neurodegener. 2021 Sep 01. 10(1): 33
    Mitochondrial links between brain aging and Alzheimer's disease.
    Heather M Wilkins, Russell H Swerdlow.
      Advancing age is a major risk factor for Alzheimer's disease (AD). This raises the question of whether AD biology mechanistically diverges from aging biology or alternatively represents exaggerated aging. Correlative and modeling studies can inform this question, but without a firm grasp of what drives aging and AD it is difficult to definitively resolve this quandary. This review speculates over the relevance of a particular hallmark of aging, mitochondrial function, to AD, and further provides background information that is pertinent to and provides perspective on this speculation.
    Keywords:  Aging; Alzheimer’s disease; Mitochondria; Mitochondrial DNA
    DOI:  https://doi.org/10.1186/s40035-021-00261-2
  19. Front Biosci (Landmark Ed). 2021 Aug 30. 26(8): 262-271
    Effects of Telmisartan, an AT1 receptor antagonist, on mitochondria-specific genes expression in a mouse MPTP model of Parkinsonism.
    Bipul Ray, Girish Ramesh, Sudhir Rama Verma, Srinivasan Ramamurthy, Sunanda Tuladhar, Arehally Marappa Mahalakshmi, Musthafa Mohamed Essa, Saravana Babu Chidambaram.
      Background: Mitochondrial dysfunction plays a crucial role in Parkinson's disease (PD) pathogenesis. The present study was undertaken to investigate the effects of Telmisartan (TEL), an angiotensin II type 1 receptor (AT1R) blocker, on the mitochondria-specific genes expression in a mouse model of Parkinsonism. Materials and methods: Mice were divided into 5 groups with 6 in each; Group I received 0.5% CMC (control) + saline, Group II received 0.5% CMC + 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) (positive control), Group III & IV received MPTP + TEL 3 and 10 mg/kg, p.o. respectively, Group V received TEL 10 mg/kg, p.o. (drug control). MPTP was given 80 mg/kg intraperitoneal in two divided doses (40 mg/kg × 2 at 16 h time interval). Vehicle or TEL was administered 1 h before the MPTP injection. Motor function was assessed 48 h after the first dose of MPTP and animals were euthanized to collect brain. Results: Mice intoxicated with MPTP showed locomotor deficits and significant upregulation of α-synuclein (α-syn), downregulation of metastasis-associated protein 1 (MTA1), and Ubiquitin C-terminal hydrolase L1 (UCHL1) in the substantia nigra pars compacta (SNpc) and Striatum (STr) regions of brains. In addition, MPTP intoxication down-regulated mitochondria-specific genes such as DJ-1, PTEN-induced putative kinase 1 (PINK1), Parkin, enriched with leucine repeats kinase 2 (LRRK2) gene expfression. Pre-treatment with TEL restored locomotor functions and upregulated PINK1, Parkin, LRRK2, DJ-1, MTA1 and UCHL1. Conclusion: The present study evidences that TEL has the ability to improve mitochondrial functions in PD.
    Keywords:  DJ-1; LRRK2; MTA1; Mitochondria; PINK1; Parkin; Parkinson’s disease; Telmisartan; UCHL1; α-synuclein
    DOI:  https://doi.org/10.52586/4942
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