bims-mibica Biomed News
on Mitochondrial bioenergetics in cancer
Issue of 2020–04–26
forty-six papers selected by
Kelsey Fisher-Wellman, East Carolina University
  1. α-ketobutyrate links alterations in cystine metabolism to glucose oxidation in mtDNA mutant cells.
  2. Mitochondria-targeted 1,4-naphthoquinone (SkQN) is a powerful prooxidant and cytotoxic agent.
  3. Myotonic Dystrophy type 1 cells display impaired metabolism and mitochondrial dysfunction that are reversed by metformin.
  4. An in situ atlas of mitochondrial DNA in mammalian tissues reveals high content in stem/progenitor cells.
  5. Deactivation of mitochondrial NADH:ubiquinone oxidoreductase (respiratory complex I): Extrinsically affecting factors.
  6. The photodynamic and direct actions of methylene blue on mitochondrial energy metabolism: A balance of the useful and harmful effects of this photosensitizer.
  7. Downregulation of MiD49 contributes to tumor growth and metastasis of human pancreatic cancer.
  8. 20,000 picometers under the OMM: diving into the vastness of mitochondrial metabolite transport.
  9. Mitochondrial lipoylation integrates age-associated decline in brown fat thermogenesis.
  10. Methylmalonyl acidemia: from mitochondrial metabolism to defective mitophagy and disease.
  11. Inefficient thermogenic mitochondrial respiration due to futile proton leak in a mouse model of fragile X syndrome.
  12. A procedure to extract functional isolated mitochondria from small-sized human atrial samples. Application to obesity with a partial characterisation of the organelles.
  13. Diazoxide affects mitochondrial bioenergetics by the opening of mKATP channel on submicromolar scale.
  14. Endocrine-Exocrine Signaling Drives Obesity-Associated Pancreatic Ductal Adenocarcinoma.
  15. Metformin in combination with JS-K inhibits growth of renal cell carcinoma cells via reactive oxygen species activation and inducing DNA breaks.
  16. Dissecting the Roles of Mitochondrial Complex I Intermediate Assembly Complex Factors in the Biogenesis of Complex I.
  17. The Mitochondrial Protein VDAC1 at the Crossroads of Cancer Cell Metabolism: The Epigenetic Link.
  18. Regional metabolic signatures in the Ndufs4(KO) mouse brain implicate defective glutamate/α-ketoglutarate metabolism in mitochondrial disease.
  19. Mitochondrial pyruvate and fatty acid flux modulate MICU1-dependent control of MCU activity.
  20. Transient receptor potential ion channel TRPM2 promotes AML proliferation and survival through modulation of mitochondrial function, ROS, and autophagy.
  21. Mitochondrial nucleoid remodeling and biogenesis are regulated by the p53-p21WAF1-PKCζ pathway in p16INK4a-silenced cells.
  22. Chronic treatment with the complex I inhibitor MPP+ depletes endogenous PTEN-induced kinase 1(PINK1) via upregulation of BCL-2-associated athanogene 6.
  23. HDAC inhibitors elicit metabolic reprogramming by targeting super-enhancers in glioblastoma models.
  24. Mitochondrial adaptations in liver and skeletal muscle to pro-longevity nutritional and genetic interventions: the crosstalk between calorie restriction and CYB5R3 overexpression in transgenic mice.
  25. MitoTEMPO provides an antiarrhythmic effect in aged-rats through attenuation of mitochondrial reactive oxygen species.
  26. Para-Halogenation of Amphetamine and Methcathinone Increases the Mitochondrial Toxicity in Undifferentiated and Differentiated SH-SY5Y Cells.
  27. Sirt3 regulates the level of mitochondrial DNA repair activity through deacetylation of NEIL1, NEIL2, OGG1, MUTYH, APE1 and LIG3 in colorectal cancer.
  28. HIGD2A is required for assembly of the COX3 module of human mitochondrial complex IV.
  29. The road to the structure of the mitochondrial respiratory chain supercomplex.
  30. A new brain mitochondrial sodium-sensitive potassium channel: effect of sodium ions on respiratory chain activity.
  31. 1,25-Dihydroxyvitamin D3 Prevents Deleterious Effects of Erythromycin on Mitochondrial Function in Rat Heart Isolated Mitochondria.
  32. Age- and Organ-Specific Differences in Mitochondrial Bioenergetics in Brown Norway Rats.
  33. Developmental and Tumor Angiogenesis Requires the Mitochondria-Shaping Protein Opa1.
  34. IMCA Induces Ferroptosis Mediated by SLC7A11 through the AMPK/mTOR Pathway in Colorectal Cancer.
  35. AMPK activation by metformin promotes survival of dormant ER+ breast cancer cells.
  36. Pentathiepins: a novel class of glutathione peroxidase 1 inhibitors that induce oxidative stress, loss of mitochondrial membrane potential and apoptosis in human cancer cells.
  37. Quantifying mitochondrial respiration in human lymphocytes and monocytes challenged with hydrogen peroxide.
  38. Utilizing Synergistic Potential of Mitochondria-Targeting Drugs for Leukemia Therapy.
  39. Targeting glutamine metabolism enhances tumor specific immunity by modulating suppressive myeloid cells.
  40. Sample multiplexing for targeted pathway proteomics in aging mice.
  41. Rational Design of Allosteric and Selective Inhibitors of the Molecular Chaperone TRAP1.
  42. Tankyrase inhibition ameliorates lipid disorder via suppression of PGC-1α PARylation in db/db mice.
  43. Plumbagin Enhances the Anticancer Efficacy of Cisplatin by Increasing Intracellular ROS in Human Tongue Squamous Cell Carcinoma.
  44. Nicotinamide riboside supplementation alters body composition and skeletal muscle acetylcarnitine concentrations in healthy obese humans.
  45. m.3243A > G-Induced Mitochondrial Dysfunction Impairs Human Neuronal Development and Reduces Neuronal Network Activity and Synchronicity.
  46. CD38 is involved in cell energy metabolism via activating the PI3K/AKT/mTOR signaling pathway in cervical cancer cells.
  1. Metab Eng. 2020 Apr 21. pii: S1096-7176(20)30065-3. [Epub ahead of print]
    α-ketobutyrate links alterations in cystine metabolism to glucose oxidation in mtDNA mutant cells.
    Nicholas P Lesner, Amrita Gokhale, Kalyani Kota, Ralph J DeBerardinis, Prashant Mishra.
      Pathogenic mutations in the mitochondrial genome (mtDNA) impair organellar ATP production, requiring mutant cells to activate metabolic adaptations for survival. Understanding how metabolism adapts to clinically relevant mtDNA mutations may provide insight into cellular strategies for metabolic flexibility. In this study, we use 13C isotope tracing and metabolic flux analysis to investigate central carbon and amino acid metabolic reprogramming in isogenic cells containing mtDNA mutations. We identify alterations in glutamine and cystine transport which indirectly regulate mitochondrial metabolism and electron transport chain function. Metabolism of cystine can promote glucose oxidation through the transsulfuration pathway and the production of α-ketobutyrate. Intriguingly, activating or inhibiting α-ketobutyrate production is sufficient to modulate both glucose oxidation and mitochondrial respiration in mtDNA mutant cells. Thus, cystine-stimulated transsulfuration serves as an adaptive mechanism linking glucose oxidation and amino acid metabolism in the setting of mtDNA mutations.
    Keywords:  Metabolic reprogramming; Transsulfuration; mtDNA mutations
    DOI:  https://doi.org/10.1016/j.ymben.2020.03.010
  2. Biochim Biophys Acta Bioenerg. 2020 Apr 16. pii: S0005-2728(20)30060-8. [Epub ahead of print] 148210
    Mitochondria-targeted 1,4-naphthoquinone (SkQN) is a powerful prooxidant and cytotoxic agent.
    Tatyana N Goleva, Konstantin G Lyamzaev, Anton G Rogov, Ljudmila S Khailova, Khoren K Epremyan, Galina P Shumakovich, Lidia V Domnina, Olga Yu Ivanova, Natalia V Marmiy, Tatiana V Zinevich, Dmitry S Esipov, Renata A Zvyagilskaya, Vladimir P Skulachev, Boris V Chernyak.
      An increase in the production of reactive oxygen species (ROS) in mitochondria due to targeted delivery of redox active compounds may be useful in studies of modulation of cell functions by mitochondrial ROS. Recently, the mitochondria-targeted derivative of menadione (MitoK3) was synthesized. However, MitoK3 did not induce mitochondrial ROS production and lipid peroxidation while exerting significant cytotoxic action. Here we synthesized 1,4-naphthoquinone conjugated with alkyltriphenylphosphonium (SkQN) as a prototype of mitochondria-targeted prooxidant, and its redox properties, interactions with isolated mitochondria, yeast cells and various human cell lines were investigated. According to electrochemical measurements, SkQN was more active redox agent and, due to the absence of methyl group in the naphthoquinone ring, more reactive as electrophile than MitoK3. SkQN (but not MitoK3) stimulated hydrogen peroxide production in isolated mitochondria. At low concentrations, SkQN stimulated state 4 respiration in mitochondria, decreased membrane potential, and blocked ATP synthesis, being more efficient uncoupler of oxidative phosphorylation than MitoK3. In yeast cells, SkQN decreased cell viability and induced oxidative stress and mitochondrial fragmentation. SkQN killed various tumor cells much more efficiently than MitoK3. Since many tumors are characterized by increased oxidative stress, the use of new mitochondria-targeted prooxidants may be a promising strategy for anticancer therapy.
    Keywords:  Cytotoxicity; Mitochondria; Prooxidant; [10-(1,4-Dioxo-1,4-dihydronaphthalen-2-yl)decyl] triphenylphosphonium (SkQN)
    DOI:  https://doi.org/10.1016/j.bbabio.2020.148210
  3. Aging (Albany NY). 2020 Apr 08. 12(7): 6260-6275
    Myotonic Dystrophy type 1 cells display impaired metabolism and mitochondrial dysfunction that are reversed by metformin.
    Mikel García-Puga, Ander Saenz-Antoñanzas, Roberto Fernández-Torrón, Adolfo Lopez de Munain, Ander Matheu.
      Myotonic dystrophy type 1 (DM1; MIM #160900) is an autosomal dominant disorder, clinically characterized by progressive muscular weakness and multisystem degeneration. The broad phenotypes observed in patients with DM1 resemble the appearance of a multisystem accelerated aging process. However, the molecular mechanisms underlying these phenotypes remain largely unknown. In this study, we characterized the impact of metabolism and mitochondria on fibroblasts and peripheral blood mononuclear cells (PBMCs) derived from patients with DM1 and healthy individuals. Our results revealed a decrease in oxidative phosphorylation system (OXPHOS) activity, oxygen consumption rate (OCR), ATP production, energy metabolism, and mitochondrial dynamics in DM1 fibroblasts, as well as increased accumulation of reactive oxygen species (ROS). PBMCs of DM1 patients also displayed reduced mitochondrial dynamics and energy metabolism. Moreover, treatment with metformin reversed the metabolic and mitochondrial defects as well as additional accelerated aging phenotypes, such as impaired proliferation, in DM1-derived fibroblasts. Our results identify impaired cell metabolism and mitochondrial dysfunction as important drivers of DM1 pathophysiology and, therefore, reveal the efficacy of metformin treatment in a pre-clinical setting.
    Keywords:  aging; metabolism; metformin; mitochondria; myotonic dystrophy type 1; neuromuscular disease
    DOI:  https://doi.org/10.18632/aging.103022
  4. Am J Pathol. 2020 Apr 15. pii: S0002-9440(20)30194-2. [Epub ahead of print]
    An in situ atlas of mitochondrial DNA in mammalian tissues reveals high content in stem/progenitor cells.
    Jiayu Chen, Qizhi Zheng, Lauren B Peiffer, Jessica L Hicks, Michael C Haffner, Avi Z Rosenberg, Moshe Levi, Xiaoxin X Wang, Busra Ozbek, Javier Baena-Del Valle, Srinivasan Yegnasubramanian, Angelo M De Marzo.
      Mitochondria regulate ATP production, metabolism and cell death. Alterations in mitochondrial DNA (mtDNA) sequence and copy number are implicated in aging and organ dysfunction in diverse inherited and sporadic diseases. Since most measurements of mtDNA use homogenates of complex tissues, little is known about cell type-specific mtDNA copy number heterogeneity in normal physiology, aging and disease. Thus, the precise cell types whose loss of mitochondrial activity and altered mtDNA copy number that result in organ dysfunction in aging and disease have often not been clarified. Here, we validated an in situ hybridization approach to generate a single cell resolution atlas of mtDNA content in mammalian tissues. In hierarchically organized self-renewing tissues, higher levels of mtDNA were observed in stem/proliferative compartments compared to differentiated compartments. Striking zonal patterns of mtDNA levels in the liver reflected the known oxygen tension gradient. In the kidney, proximal and distal tubules had markedly higher mtDNA levels compared to cells within glomeruli and collecting duct epithelial cells. In mice, decreased mtDNA levels were visualized in renal tubules as a function of aging, which was prevented by calorie restriction. We provide a novel approach for quantifying species- and cell type-specific mtDNA copy number and dynamics in any normal or diseased tissue that can be used for monitoring the effects of interventions in animal and human studies.
    DOI:  https://doi.org/10.1016/j.ajpath.2020.03.018
  5. Biochim Biophys Acta Bioenerg. 2020 Apr 18. pii: S0005-2728(20)30057-8. [Epub ahead of print] 148207
    Deactivation of mitochondrial NADH:ubiquinone oxidoreductase (respiratory complex I): Extrinsically affecting factors.
    Vera G Grivennikova, Grigory V Gladyshev, Andrei D Vinogradov.
      Mitochondrial NADH:ubiquinone oxidoreductase (proton translocating respiratory complex I) serves several essential functions in cell metabolism: it maintains the intramitochondrial NADH/NAD+ ratio, contributes to generation of the proton-motive force, and participates in physiological and/or pathophysiological production of so-called reactive oxygen species. A characteristic feature of complex I is a slow, compared with its catalytic turnover, transformation to its inactive (deactivated) state, a phenomenon operationally called A/D transition. Here we report data on several extrinsic factors affecting deactivation as observed in coupled or uncoupled bovine heart submitochondrial particles. The time course of the strongly temperature-dependent deactivation deviates from first-order kinetics, and this deviation is abolished in the presence of an SH-group-specific reagent. The residual fraction of activity attained upon extensive deactivation shows the same kinetics of NADH oxidation as the fully active enzyme does. The rate of complex I deactivation is only slightly pH dependent within the range of 7.0-8.5 and significantly increases at higher pH. ATP∙(Mg) decreases the rate of complex I deactivation in coupled SMP, and this effect is abolished if the proton-motive force generating ATPase activity of Fo∙F1 is precluded. Taken together, the data show that an equilibrium between the A and D forms of complex I exists. Possible mechanistic aspects of the deactivation process are discussed.
    Keywords:  A/D transition; Energy transduction; NADH:ubiquinone oxidoreductase; Respiratory chain
    DOI:  https://doi.org/10.1016/j.bbabio.2020.148207
  6. Free Radic Biol Med. 2020 Apr 18. pii: S0891-5849(20)30345-2. [Epub ahead of print]
    The photodynamic and direct actions of methylene blue on mitochondrial energy metabolism: A balance of the useful and harmful effects of this photosensitizer.
    Eduardo Makiyama Klosowski, Byanca Thais Lima de Souza, Marcio Shigueaki Mito, Renato Polimeni Constantin, Gislaine Cristiane Mantovanelli, Juliana Morais Mewes, Paulo Francisco Veiga Bizerra, Paulo Vinicius Moreira da Costa Menezes, Eduardo Hideo Gilglioni, Karina Sayuri Utsunomiya, Rogério Marchiosi, Wanderley Dantas Dos Santos, Osvaldo Ferrarese Filho, Wilker Caetano, Paulo Cesar de Souza Pereira, Renato Sonchini Gonçalves, Jorgete Constantin, Emy Luiza Ishii-Iwamoto, Rodrigo Polimeni Constantin.
      According to the literature, methylene blue (MB) is a photosensitizer (PS) with a high affinity for mitochondria. Therefore, several studies have explored this feature to evaluate its photodynamic effects on the mitochondrial apoptotic pathway under normoxic conditions. We are aware only of limited reports regarding MB's photodynamic effects on mitochondrial energy metabolism, especially under hypoxic conditions. Thus, the purposes of this study were to determine the direct and photodynamic acute effects of MB on the energy metabolism of rat liver mitochondria under hypoxic conditions and its direct acute effects on several parameters linked to energy metabolism in the isolated perfused rat liver. MB presented a high affinity for mitochondria, irrespective of photostimulation or proton gradient formation. Upon photostimulation, MB demonstrated high in vitro oxidizing species generation ability. Consequently, MB damaged the mitochondrial macromolecules, as could be evidenced by the elevated levels of lipid peroxidation and protein carbonyls. In addition to generating a pro-oxidant environment, MB also led to a deficient antioxidant defence system, as indicated by the reduced glutathione (GSH) depletion. Bioenergetically, MB caused uncoupling of oxidative phosphorylation and led to photodynamic inactivation of complex I, complex II, and F1FO-ATP synthase complex, thus decreasing mitochondrial ATP generation. Contrary to what is expected for an ideal PS, MB displayed appreciable dark toxicity on mitochondrial energy metabolism. The results indicated that MB acted via at least three mechanisms: direct damage to the inner mitochondrial membrane; uncoupling of oxidative phosphorylation; and inhibition of electron transfer. Confirming the impairment of mitochondrial energy metabolism, MB also strongly inhibited mitochondrial ATP production. In the perfused rat liver, MB stimulated oxygen consumption, decreased the ATP/ADP ratio, inhibited gluconeogenesis and ureogenesis, and stimulated glycogenolysis, glycolysis, and ammoniagenesis, fully corroborating its uncoupling action in intact cells, as well. It can be concluded that even under hypoxic conditions, MB is a PS with potential for photodynamic effect-induced mitochondrial dysfunction. However, MB disrupts the mitochondrial energy metabolism even in the dark, causing energy-linked liver metabolic changes that could be harmful in specific circumstances.
    Keywords:  Isolated mitochondria; Liver toxicity; Mitochondrial energy metabolism; Photodynamic effect; Uncoupling
    DOI:  https://doi.org/10.1016/j.freeradbiomed.2020.04.015
  7. Oncol Rep. 2020 Feb 12.
    Downregulation of MiD49 contributes to tumor growth and metastasis of human pancreatic cancer.
    Lu Bai, Jing Liang, Lihong Li, Enxiao Li.
      Changes in mitochondrial morphology by dysregulated mitochondrial fission‑fusion proteins have been increasingly recognized as a hallmark of cancer. MiD49 (mitochondrial dynamics protein of 49 kDa) is a newly identified mitochondrial fission protein involved in the dynamic regulation of mitochondrial morphology. However, the expression pattern and biological functions of MiD49 in human cancers remain largely unexplored, especially in pancreatic cancer (PC). In the present study, the expression and clinical significance of MiD49 was firstly determined by RT‑qPCR and western blot analyses in PC cell lines and tumor tissues. In addition, the biologic functions of MiD49 in PC cell growth and metastasis were investigated using gain‑ and loss‑of‑function assays both in vitro and in vivo. Moreover, the underlying mechanisms by which MiD49 regulates PC cell growth and metastasis were further explored. Our results showed that MiD49 was markedly downregulated in both PC cell lines and human PC specimens. Forced expression of MiD49 suppressed PC cell growth and metastasis both in vitro and in vivo, while knockdown of MiD49 exhibited the opposite effect. Mechanistic exploration demonstrated that the tumor‑suppressive effect of MiD49 was mediated by decreased mitochondrial fission and subsequent reduced ROS production in PC cells. Our findings suggest a critical tumor‑suppressive role played by MiD49 in pancreatic cancer.
    DOI:  https://doi.org/10.3892/or.2020.7499
  8. EMBO Rep. 2020 Apr 23. e50071
    20,000 picometers under the OMM: diving into the vastness of mitochondrial metabolite transport.
    Corey N Cunningham, Jared Rutter.
      The metabolic compartmentalization enabled by mitochondria is key feature of many cellular processes such as energy conversion to ATP production, redox balance, and the biosynthesis of heme, urea, nucleotides, lipids, and others. For a majority of these functions, metabolites need to be transported across the impermeable inner mitochondrial membrane by dedicated carrier proteins. Here, we examine the substrates, structural features, and human health implications of four mitochondrial metabolite carrier families: the SLC25A family, the mitochondrial ABCB transporters, the mitochondrial pyruvate carrier (MPC), and the sideroflexin proteins.
    Keywords:  metabolism; metabolite carriers; mitochondria; transporters
    DOI:  https://doi.org/10.15252/embr.202050071
  9. Nat Metab. 2019 Sep;1(9): 886-898
    Mitochondrial lipoylation integrates age-associated decline in brown fat thermogenesis.
    Kazuki Tajima, Kenji Ikeda, Hsin-Yi Chang, Chih-Hsiang Chang, Takeshi Yoneshiro, Yasuo Oguri, Heejin Jun, Jun Wu, Yasushi Ishihama, Shingo Kajimura.
      Thermogenesis in brown adipose tissue (BAT) declines with age; however, what regulates this process remains poorly understood. Here, we identify mitochondria lipoylation as a previously unappreciated molecular hallmark of aged BAT in mice. Using mitochondrial proteomics, we show that mitochondrial lipoylation is disproportionally reduced in aged BAT through a post-transcriptional decrease in the iron-sulfur (Fe-S) cluster formation pathway. A defect in the Fe-S cluster formation by the fat-specific deletion of Bola3 significantly reduces mitochondrial lipoylation and fuel oxidation in BAT, leading to glucose intolerance and obesity. In turn, enhanced mitochondrial lipoylation by α-lipoic acid supplementation effectively restores BAT function in old mice, thereby preventing age-associated obesity and glucose intolerance. The effect of α-lipoic acids requires mitochondrial lipoylation via the Bola3 pathway and does not depend on the anti-oxidant activity of α-lipoic acid. These results open up the possibility to alleviate the age-associated decline in energy expenditure by enhancing the mitochondrial lipoylation pathway.
    Keywords:  Brown adipose tissue; Glucose metabolism; Mitochondria; Thermogenesis
    DOI:  https://doi.org/10.1038/s42255-019-0106-z
  10. Autophagy. 2020 Apr 22. 1-3
    Methylmalonyl acidemia: from mitochondrial metabolism to defective mitophagy and disease.
    Alessandro Luciani, Olivier Devuyst.
      Methylmalonic acidemia (MMA) is an autosomal recessive inborn error of metabolism due to the deficiency of mitochondrial MMUT (methylmalonyl-CoA mutase) - an enzyme that mediates the cellular breakdown of certain amino acids and lipids. The loss of MMUT leads to the accumulation of toxic organic acids causing severe organ dysfunctions and life-threatening complications. The mechanisms linking MMUT deficiency, mitochondrial alterations and cell toxicity remain uncharacterized. Using cell and animal-based models, we recently unveiled that MMUT deficiency impedes the PINK1-induced translocation of PRKN/Parkin to MMA-damaged mitochondria, thereby halting their delivery and subsequent degradation by macroautophagy/autophagy-lysosome systems. In turn, this defective mitophagy process instigates the accumulation of dysfunctional mitochondria that spark epithelial distress and tissue damage. Correction of PINK1-directed mitophagy defects or mitochondrial dysfunctions rescues epithelial distress in MMA cells and alleviates disease-relevant phenotypes in mmut‒deficient zebrafish. Our findings suggest a link between primary MMUT deficiency and diseased mitochondria, mitophagy dysfunction and cell distress, offering potential therapeutic perspectives for MMA and other metabolic diseases.
    Keywords:  Cell damage; inherited metabolic disorders; kidney tubule; metabolism; mitochondria; mitophagy; oxidative stress
    DOI:  https://doi.org/10.1080/15548627.2020.1753927
  11. FASEB J. 2020 Apr 20.
    Inefficient thermogenic mitochondrial respiration due to futile proton leak in a mouse model of fragile X syndrome.
    Keren K Griffiths, Aili Wang, Lifei Wang, Matthew Tracey, Giulio Kleiner, Catarina M Quinzii, Linlin Sun, Guang Yang, Jose F Perez-Zoghbi, Pawel Licznerski, Mu Yang, Elizabeth A Jonas, Richard J Levy.
      Fragile X syndrome (FXS) is the leading known inherited intellectual disability and the most common genetic cause of autism. The full mutation results in transcriptional silencing of the Fmr1 gene and loss of fragile X mental retardation protein (FMRP) expression. Defects in neuroenergetic capacity are known to cause a variety of neurodevelopmental disorders. Thus, we explored the integrity of forebrain mitochondria in Fmr1 knockout mice during the peak of synaptogenesis. We found inefficient thermogenic respiration due to futile proton leak in Fmr1 KO mitochondria caused by coenzyme Q (CoQ) deficiency and an open cyclosporine-sensitive channel. Repletion of mitochondrial CoQ within the Fmr1 KO forebrain closed the channel, blocked the pathological proton leak, restored rates of protein synthesis during synaptogenesis, and normalized the key phenotypic features later in life. The findings demonstrate that FMRP deficiency results in inefficient oxidative phosphorylation during the neurodevelopment and suggest that dysfunctional mitochondria may contribute to the FXS phenotype.
    Keywords:   Fmr1 ; Fragile X syndrome; coenzyme Q; mitochondria; permeability transition pore; proton leak; synaptogenesis; thermogenic; ubiquinone; uncoupled respiration
    DOI:  https://doi.org/10.1096/fj.202000283RR
  12. Free Radic Biol Med. 2020 Apr 21. pii: S0891-5849(20)30331-2. [Epub ahead of print]
    A procedure to extract functional isolated mitochondria from small-sized human atrial samples. Application to obesity with a partial characterisation of the organelles.
    Thibault Leger, Chrystele Jouve, Veronique Patrac, Valerie Batel, Damien Bouvier, Vincent Sapin, Bruno Miguel, Luc Demaison, Kasra Azarnoush.
      Evaluating the activity of cardiac mitochondria is probably the best way to estimate early cellular damage in chronic pathology. Early diagnosis allows rapid therapeutic intervention thus increasing patient survival rate in a number of diseases. However, data on human cardiac mitochondria are scarce in the international literature. Here, we describe a method to extract and study functional mitochondria from the small-sized right atrial aliquots (minimum of 400 mg) obtained during extracorporeal circulation and usually considered as surgical waste products. The mitochondria were purified through several mechanical processes (fine myocardial cutting, tissue grinding and potter Elvehjem homogenising), an enzymatic proteolytic action (subtilisin) and differential centrifugations. In chronic pathologies, including obesity, early disturbances of mitochondrial function can occur. The effects of obesity on the rate of mitochondrial oxygen consumption and H2O2 release were thus determined with three different substrates (glutamate/malate, succinate/rotenone and palmitoylcarnitine/malate). The human atrial mitochondria were of high quality from a functional viewpoint, compared to rat ventricle organelles, but the extraction yield of the human mitochondria was twice lower than that of rat mitochondria. Tests showed that glutamate/malate-related ADP-stimulated respiration was strongly increased in obese subjects, although the oxidation of the other two substrates was unaffected. Reactive oxygen species (ROS) production by the isolated mitochondria was low in comparison with that of the lean subjects. These results confirm those found in one of our previous studies in the ventricles of rats fed a high-fat diet. In conclusion, the described method is simple, reliable and sensitive. It allows for the description of the impact of obesity on the function of atrial mitochondria while using only a small patient sampling (n = 5 in both the lean and the obese groups).
    Keywords:  Human right atrium; Mitochondria; Obesity; Oxidative phosphorylation; ROS
    DOI:  https://doi.org/10.1016/j.freeradbiomed.2020.04.006
  13. BMC Mol Cell Biol. 2020 Apr 19. 21(1): 31
    Diazoxide affects mitochondrial bioenergetics by the opening of mKATP channel on submicromolar scale.
    Olga Akopova, Liudmila Kolchinskaya, Valentina Nosar, Iryna Mankovska, Vadim Sagach.
       BACKGROUND: Cytoprotection afforded by mitochondrial ATP-sensitive K+-channel (mKATP-channel) opener diazoxide (DZ) largely depends on the activation of potassium cycle with eventual modulation of mitochondrial functions and ROS production. However, generally these effects were studied in the presence of Mg∙ATP known to block K+ transport. Thus, the purpose of our work was the estimation of DZ effects on K+ transport, K+ cycle and ROS production in rat liver mitochondria in the absence of Mg∙ATP.
    RESULTS: Without Mg·ATP, full activation of native mKATP-channel, accompanied by the increase in ATP-insensitive K+ uptake, activation of K+-cycle and respiratory uncoupling, was reached at ≤0.5 μM of DZ,. Higher diazoxide concentrations augmented ATP-insensitive K+ uptake, but not mKATP-channel activity. mKATP-channel was blocked by Mg·ATP, reactivated by DZ, and repeatedly blocked by mKATP-channel blockers glibenclamide and 5-hydroxydecanoate, whereas ATP-insensitive potassium transport was blocked by Mg2+ and was not restored by DZ. High sensitivity of potassium transport to DZ in native mitochondria resulted in suppression of mitochondrial ROS production caused by the activation of K+-cycle on sub-micromolar scale. Based on the oxygen consumption study, the share of mKATP-channel in respiratory uncoupling by DZ was found.
    CONCLUSIONS: The study of mKATP-channel activation by diazoxide in the absence of MgATP discloses novel, not described earlier, aspects of mKATP-channel interaction with this drug. High sensitivity of mKATP-channel to DZ results in the modulation of mitochondrial functions and ROS production by DZ on sub-micromolar concentration scale. Our experiments led us to the hypothesis that under the conditions marked by ATP deficiency affinity of mKATP-channel to DZ can increase, which might contribute to the high effectiveness of this drug in cardio- and neuroprotection.
    Keywords:  Diazoxide; Mild uncoupling; Potassium cycle; Potassium transport; ROS production; mKATP-channel
    DOI:  https://doi.org/10.1186/s12860-020-00275-0
  14. Cell. 2020 Apr 16. pii: S0092-8674(20)30395-0. [Epub ahead of print]
    Endocrine-Exocrine Signaling Drives Obesity-Associated Pancreatic Ductal Adenocarcinoma.
    Katherine Minjee Chung, Jaffarguriqbal Singh, Lauren Lawres, Kimberly Judith Dorans, Cathy Garcia, Daniel B Burkhardt, Rebecca Robbins, Arjun Bhutkar, Rebecca Cardone, Xiaojian Zhao, Ana Babic, Sara A Vayrynen, Andressa Dias Costa, Jonathan A Nowak, Daniel T Chang, Richard F Dunne, Aram F Hezel, Albert C Koong, Joshua J Wilhelm, Melena D Bellin, Vibe Nylander, Anna L Gloyn, Mark I McCarthy, Richard G Kibbey, Smita Krishnaswamy, Brian M Wolpin, Tyler Jacks, Charles S Fuchs, Mandar Deepak Muzumdar.
      Obesity is a major modifiable risk factor for pancreatic ductal adenocarcinoma (PDAC), yet how and when obesity contributes to PDAC progression is not well understood. Leveraging an autochthonous mouse model, we demonstrate a causal and reversible role for obesity in early PDAC progression, showing that obesity markedly enhances tumorigenesis, while genetic or dietary induction of weight loss intercepts cancer development. Molecular analyses of human and murine samples define microenvironmental consequences of obesity that foster tumorigenesis rather than new driver gene mutations, including significant pancreatic islet cell adaptation in obesity-associated tumors. Specifically, we identify aberrant beta cell expression of the peptide hormone cholecystokinin (Cck) in response to obesity and show that islet Cck promotes oncogenic Kras-driven pancreatic ductal tumorigenesis. Our studies argue that PDAC progression is driven by local obesity-associated changes in the tumor microenvironment and implicate endocrine-exocrine signaling beyond insulin in PDAC development.
    Keywords:  beta cells; cholecystokinin; genetically engineered mouse models; leptin; obesity; pancreatic cancer; pancreatic islets; tumor microenvironment
    DOI:  https://doi.org/10.1016/j.cell.2020.03.062
  15. J Cancer. 2020 ;11(13): 3701-3712
    Metformin in combination with JS-K inhibits growth of renal cell carcinoma cells via reactive oxygen species activation and inducing DNA breaks.
    Yuwan Zhao, Qiuming Luo, Jierong Mo, Jianwei Li, Dongcai Ye, Zhixian Ao, Lixin Chen, Jianjun Liu.
      Metformin (MET) is taken as a principal medication for remedying Type 2 diabetes mellitus. Its anti-tumor effect has been reported increasingly, but the precise mechanism of it remains unclear. This study aims to explore the efficacy of MET and MET combined with nitric oxide donor prodrug JS-K on the proliferation, apoptosis, and DNA damage in human renal cell carcinoma (RCC) cells, and investigate the possible molecular mechanism involved. The cell proliferation was tested through methyl-tetrazolium assay and cell apoptosis was ascertained by flow cytometry. The dihydroethidium and JC-1 fluorescent methods were used to detect Reactive oxygen species (ROS) and mitochondrial transmembrane potential (Δψm), respectively. Proteins associated with apoptosis and DNA damage were evaluated by Western blotting. Results showed that MET and JS-K could suppress cell growth, and the inhibition concentration 50 of treatment with MET combined with JS-K (MET + JS-K) showed more toxicity than individual agents on RCC cells. This augmented toxicity was associated with intracellular reactive oxygen species (ROS) level, mitochondrial membrane potential alteration, and induced DNA breaks. The results of Western blotting showed that the expression level of pro-apoptotic proteins, such as Bax, Bak, caspase-3, and caspase-9, was up-regulated, and the anti-apoptotic protein Bcl-2 was down-regulated after treatment using MET alone and MET + JS-K, correspondingly. Moreover, MET + JS-K inhibited the expression of cellular PCNA and Rad51, and immunofluorescence analysis of γH2AX proved that MET + JS-K enhanced DNA damage. In summary, the results of this research indicated that MET and JS-K inhibited RCC cell growth by activating ROS, targeting mitochondria-dependent apoptotic pathways, and inducing DNA breaks.
    Keywords:  DNA breaks; JS-K; ROS; metformin; renal cell carcinoma
    DOI:  https://doi.org/10.7150/jca.36372
  16. Cell Rep. 2020 Apr 21. pii: S2211-1247(20)30441-1. [Epub ahead of print]31(3): 107541
    Dissecting the Roles of Mitochondrial Complex I Intermediate Assembly Complex Factors in the Biogenesis of Complex I.
    Luke E Formosa, Linden Muellner-Wong, Boris Reljic, Alice J Sharpe, Thomas D Jackson, Traude H Beilharz, Diana Stojanovski, Michael Lazarou, David A Stroud, Michael T Ryan.
      Mitochondrial complex I harbors 7 mitochondrial and 38 nuclear-encoded subunits. Its biogenesis requires the assembly and integration of distinct intermediate modules, mediated by numerous assembly factors. The mitochondrial complex I intermediate assembly (MCIA) complex, containing assembly factors NDUFAF1, ECSIT, ACAD9, and TMEM126B, is required for building the intermediate ND2-module. The role of the MCIA complex and the involvement of other proteins in the biogenesis of this module is unclear. Cell knockout studies reveal that while each MCIA component is critical for complex I assembly, a hierarchy of stability exists centered on ACAD9. We also identify TMEM186 and COA1 as bona fide components of the MCIA complex with loss of either resulting in MCIA complex defects and reduced complex I assembly. TMEM186 enriches with newly translated ND3, and COA1 enriches with ND2. Our findings provide new functional insights into the essential nature of the MCIA complex in complex I assembly.
    Keywords:  MCIA complex; NADH-ubiquinone dehydrogenase; assembly factors; complex I; mitochondria; oxidative phosphorylation
    DOI:  https://doi.org/10.1016/j.celrep.2020.107541
  17. Cancers (Basel). 2020 Apr 22. pii: E1031. [Epub ahead of print]12(4):
    The Mitochondrial Protein VDAC1 at the Crossroads of Cancer Cell Metabolism: The Epigenetic Link.
    Zohar Amsalem, Tasleem Arif, Anna Shteinfer-Kuzmine, Vered Chalifa-Caspi, Varda Shoshan-Barmatz.
      Carcinogenesis is a complicated process that involves the deregulation of epigenetics, resulting in cellular transformational events, such as proliferation, differentiation, and metastasis. Most chromatin-modifying enzymes utilize metabolites as co-factors or substrates and thus are directly dependent on such metabolites as acetyl-coenzyme A, S-adenosylmethionine, and NAD+. Here, we show that using specific siRNA to deplete a tumor of VDAC1 not only led to reprograming of the cancer cell metabolism but also altered several epigenetic-related enzymes and factors. VDAC1, in the outer mitochondrial membrane, controls metabolic cross-talk between the mitochondria and the rest of the cell, thus regulating the metabolic and energetic functions of mitochondria, and has been implicated in apoptotic-relevant events. We previously demonstrated that silencing VDAC1 expression in glioblastoma (GBM) U-87MG cell-derived tumors, resulted in reprogramed metabolism leading to inhibited tumor growth, angiogenesis, epithelial-mesenchymal transition and invasiveness, and elimination of cancer stem cells, while promoting the differentiation of residual tumor cells into neuronal-like cells. These VDAC1 depletion-mediated effects involved alterations in transcription factors regulating signaling pathways associated with cancer hallmarks. As the epigenome is sensitive to cellular metabolism, this study was designed to assess whether depleting VDAC1 affects the metabolism-epigenetics axis. Using DNA microarrays, q-PCR, and specific antibodies, we analyzed the effects of si-VDAC1 treatment of U-87MG-derived tumors on histone modifications and epigenetic-related enzyme expression levels, as well as the methylation and acetylation state, to uncover any alterations in epigenetic properties. Our results demonstrate that metabolic rewiring of GBM via VDAC1 depletion affects epigenetic modifications, and strongly support the presence of an interplay between metabolism and epigenetics.
    Keywords:  VDAC1; cancer; histones epigenetics; metabolism; mitochondria
    DOI:  https://doi.org/10.3390/cancers12041031
  18. Mol Genet Metab. 2020 Apr 03. pii: S1096-7192(20)30081-0. [Epub ahead of print]
    Regional metabolic signatures in the Ndufs4(KO) mouse brain implicate defective glutamate/α-ketoglutarate metabolism in mitochondrial disease.
    Simon C Johnson, Ernst-Bernhard Kayser, Rebecca Bornstein, Julia Stokes, Alessandro Bitto, Kyung Yeon Park, Amanda Pan, Grace Sun, Daniel Raftery, Matt Kaeberlein, Margaret M Sedensky, Philip G Morgan.
      Leigh Syndrome (LS) is a mitochondrial disorder defined by progressive focal neurodegenerative lesions in specific regions of the brain. Defects in NDUFS4, a subunit of complex I of the mitochondrial electron transport chain, cause LS in humans; the Ndufs4 knockout mouse (Ndufs4(KO)) closely resembles the human disease. Here, we probed brain region-specific molecular signatures in pre-symptomatic Ndufs4(KO) to identify factors which underlie focal neurodegeneration. Metabolomics revealed that free amino acid concentrations are broadly different by region, and glucose metabolites are increased in a manner dependent on both region and genotype. We then tested the impact of the mTOR inhibitor rapamycin, which dramatically attenuates LS in Ndufs4(KO), on region specific metabolism. Our data revealed that loss of Ndufs4 drives pathogenic changes to CNS glutamine/glutamate/α-ketoglutarate metabolism which are rescued by mTOR inhibition Finally, restriction of the Ndufs4 deletion to pre-synaptic glutamatergic neurons recapitulated the whole-body knockout. Together, our findings are consistent with mTOR inhibition alleviating disease by increasing availability of α-ketoglutarate, which is both an efficient mitochondrial complex I substrate in Ndufs4(KO) and an important metabolite related to neurotransmitter metabolism in glutamatergic neurons.
    Keywords:  Genetics; Leigh syndrome; Metabolism; Mitochondria; Mouse; Rapamycin; Reactive oxygen species
    DOI:  https://doi.org/10.1016/j.ymgme.2020.03.007
  19. Sci Signal. 2020 Apr 21. pii: eaaz6206. [Epub ahead of print]13(628):
    Mitochondrial pyruvate and fatty acid flux modulate MICU1-dependent control of MCU activity.
    Neeharika Nemani, Zhiwei Dong, Cassidy C Daw, Travis R Madaris, Karthik Ramachandran, Benjamin T Enslow, Cherubina S Rubannelsonkumar, Santhanam Shanmughapriya, Varshini Mallireddigari, Soumya Maity, Pragya SinghMalla, Kalimuthusamy Natarajanseenivasan, Robert Hooper, Christopher E Shannon, Warren G Tourtellotte, Brij B Singh, W Brian Reeves, Kumar Sharma, Luke Norton, Subramanya Srikantan, Jonathan Soboloff, Muniswamy Madesh.
      The tricarboxylic acid (TCA) cycle converts the end products of glycolysis and fatty acid β-oxidation into the reducing equivalents NADH and FADH2 Although mitochondrial matrix uptake of Ca2+ enhances ATP production, it remains unclear whether deprivation of mitochondrial TCA substrates alters mitochondrial Ca2+ flux. We investigated the effect of TCA cycle substrates on MCU-mediated mitochondrial matrix uptake of Ca2+, mitochondrial bioenergetics, and autophagic flux. Inhibition of glycolysis, mitochondrial pyruvate transport, or mitochondrial fatty acid transport triggered expression of the MCU gatekeeper MICU1 but not the MCU core subunit. Knockdown of mitochondrial pyruvate carrier (MPC) isoforms or expression of the dominant negative mutant MPC1R97W resulted in increased MICU1 protein abundance and inhibition of MCU-mediated mitochondrial matrix uptake of Ca2+ We also found that genetic ablation of MPC1 in hepatocytes and mouse embryonic fibroblasts resulted in reduced resting matrix Ca2+, likely because of increased MICU1 expression, but resulted in changes in mitochondrial morphology. TCA cycle substrate-dependent MICU1 expression was mediated by the transcription factor early growth response 1 (EGR1). Blocking mitochondrial pyruvate or fatty acid flux was linked to increased autophagy marker abundance. These studies reveal a mechanism that controls the MCU-mediated Ca2+ flux machinery and that depends on TCA cycle substrate availability. This mechanism generates a metabolic homeostatic circuit that protects cells from bioenergetic crisis and mitochondrial Ca2+ overload during periods of nutrient stress.
    DOI:  https://doi.org/10.1126/scisignal.aaz6206
  20. Cell Death Dis. 2020 Apr 20. 11(4): 247
    Transient receptor potential ion channel TRPM2 promotes AML proliferation and survival through modulation of mitochondrial function, ROS, and autophagy.
    Shu-Jen Chen, Lei Bao, Kerry Keefer, Santhanam Shanmughapriya, Longgui Chen, John Lee, JuFang Wang, Xue-Qian Zhang, Iwona Hirschler-Laszkiewicz, Salim Merali, Carmen Merali, Yuka Imamura, Sinisa Dovat, Muniswamy Madesh, Joseph Y Cheung, Hong-Gang Wang, Barbara A Miller.
      Transient receptor potential melastatin 2 (TRPM2) ion channel has an essential function in maintaining cell survival following oxidant injury. Here, we show that TRPM2 is highly expressed in acute myeloid leukemia (AML). The role of TRPM2 in AML was studied following depletion with CRISPR/Cas9 technology in U937 cells. In in vitro experiments and in xenografts, depletion of TRPM2 in AML inhibited leukemia proliferation, and doxorubicin sensitivity was increased. Mitochondrial function including oxygen consumption rate and ATP production was reduced, impairing cellular bioenergetics. Mitochondrial membrane potential and mitochondrial calcium uptake were significantly decreased in depleted cells. Mitochondrial reactive oxygen species (ROS) were significantly increased, and Nrf2 was decreased, reducing the antioxidant response. In TRPM2-depleted cells, ULK1, Atg7, and Atg5 protein levels were decreased, leading to autophagy inhibition. Consistently, ATF4 and CREB, two master transcription factors for autophagosome biogenesis, were reduced in TRPM2-depleted cells. In addition, Atg13 and FIP200, which are known to stabilize ULK1 protein, were decreased. Reconstitution with TRPM2 fully restored proliferation, viability, and autophagy; ATF4 and CREB fully restored proliferation and viability but only partially restored autophagy. TRPM2 expression reduced the elevated ROS found in depleted cells. These data show that TRPM2 has an important role in AML proliferation and survival through regulation of key transcription factors and target genes involved in mitochondrial function, bioenergetics, the antioxidant response, and autophagy. Targeting TRPM2 may represent a novel therapeutic approach to inhibit myeloid leukemia growth and enhance susceptibility to chemotherapeutic agents through multiple pathways.
    DOI:  https://doi.org/10.1038/s41419-020-2454-8
  21. Aging (Albany NY). 2020 Apr 24. 12
    Mitochondrial nucleoid remodeling and biogenesis are regulated by the p53-p21WAF1-PKCζ pathway in p16INK4a-silenced cells.
    Yun Yeong Lee, Yeon Seung Choi, Do Wan Kim, Jae Youn Cheong, Kye Yong Song, Min Sook Ryu, In Kyoung Lim.
      Mitochondrial dysfunction is linked to age-related senescence phenotypes. We report here the pathway increasing nucleoid remodeling and biogenesis in mitochondria during the senescence of foreskin human diploid fibroblasts (fs-HDF) and WI-38 cells. Replicative senescence in fs-HDF cells increased mitochondrial nucleoid remodeling as indicated by 5-bromo-2'-deoxyuridine (BrdU) incorporation and mitochondrial transcription factor A (TFAM) expression in enlarged and fused mitochondria. Mitochondrial nucleoid remodeling was accompanied by mitochondrial biogenesis in old cells, and the expression levels of OXPHOS complex-I, -IV and -V subunits, PGC-1α and NRF1 were greatly increased compared to young cells. Activated protein kinase C zeta (PKCζ) increased mitochondrial activity and expressed phenotypes of delayed senescence in fs-HDF cells, but not in WI-38 cells. The findings were reproduced in the doxorubicin-induced senescence of young fs-HDF and WI-38 cells via the PKCζ-LKB1-AMPK signaling pathway, which was regulated by the p53-p21WAF1 pathway when p16INK4a was silenced. The signaling enhanced PGC-1α-NRF1-TFAM axis in mitochondria, which was demonstrated by Ingenuity Pathway Analysis of young and old fs-HDF cells. Activation of the p53-p21WAF1 pathway and silencing of p16INK4a are responsible for mitochondrial reprogramming in senescent cells, which may be a compensatory mechanism to promote cell survival under senescence stress.
    Keywords:  mitochondria; nucleoid remodeling; p16INK4a silence; p53-p21-PKCζ activation; senescence
    DOI:  https://doi.org/10.18632/aging.103029
  22. J Biol Chem. 2020 Apr 24. pii: jbc.RA119.010474. [Epub ahead of print]
    Chronic treatment with the complex I inhibitor MPP+ depletes endogenous PTEN-induced kinase 1(PINK1) via upregulation of BCL-2-associated athanogene 6.
    Manish Verma, Jianhui Zhu, Kent Z Q Wang, Charleen T Chu.
      Mitochondrial dysfunction is implicated in sporadic and familial Parkinson's disease (PD). However, the mechanisms that impair homeostatic responses to mitochondrial dysfunction remain unclear. Previously, we found that chronic, low dose administration of the mitochondrial complex I inhibitor 1-methyl-4-phenylpyridinium (MPP+) dysregulates mitochondrial fission-fusion, mitophagy and mitochondrial biogenesis. Given that PTEN-induced kinase 1 (PINK1) regulates mitochondrial function, dynamics and turnover, we hypothesized that alterations in endogenous PINK1 levels contribute to depletion of mitochondria during chronic complex I injury. Here we found that chronic MPP+ treatment of differentiated SH-SY5Y neuronal cells significantly decreases PINK1 expression prior to reductions in other mitochondrial components. Furthermore, Bcl2 associated athanogene 6 (BAG6, BAT3, or Scythe), a protein involved in protein quality control and degradation, was highly up-regulated during the chronic MPP+ treatment. BAG6 interacted with PINK1, and BAG6 overexpression decreased the half-life of PINK1. Conversely, siRNA-mediated BAG6 knockdown prevented chronic MPP+ stress-induced loss of PINK1, reversed MPP+-provoked mitochondrial changes, increased cell viability and prevented MPP+-induced dendrite shrinkage in primary neurons. These results indicate that BAG6 up-regulation during chronic complex I inhibition contributes to mitochondrial pathology by decreasing the levels of endogenous PINK1. Given that recessive mutations in PINK1 cause familial PD, the finding of accelerated PINK1 degradation in the chronic MPP+ model suggests that PINK1 loss-of-function represents a point of convergence between neurotoxic and genetic causes of PD.
    Keywords:  BAG6; MPP+; PTEN-induced putative kinase 1 (PINK1); Parkinson disease; mitochondria; mitochondrial complex I inhibition; mitochondrial dysfunction; neurodegeneration; neuron injury; protein degradation
    DOI:  https://doi.org/10.1074/jbc.RA119.010474
  23. J Clin Invest. 2020 Apr 21. pii: 129049. [Epub ahead of print]
    HDAC inhibitors elicit metabolic reprogramming by targeting super-enhancers in glioblastoma models.
    Trang Nguyen, Yiru Zhang, Enyuan Shang, Chang Shu, Consuelo Torrini, Junfei Zhao, Elena Bianchetti, Angeliki Mela, Nelson Humala, Aayushi Mahajan, Arif O Harmanci, Zhengdeng Lei, Mark Maienschein-Cline, Catarina Maria Quinzii, Mike-Andrew Westhoff, Georg Karpel-Massler, Jeffrey N Bruce, Peter Canoll, Markus D Siegelin.
      The Warburg effect is a tumor related phenomenon that may be targeted therapeutically. Here, we showed that glioblastoma cultures and patient tumors harbored super-enhancers in several genes related to the Warburg effect. By conducting a transcriptome analysis followed by chromatin immunoprecipitation (CHIP) sequencing coupled with a comprehensive metabolite analysis in GBM models, we unraveled that FDA-approved global (panobinostat, vorinostat) and selective (romidepsin) histone-deacetylase (HDAC) inhibitors elicited metabolic reprogramming in concert with disruption of several Warburg-effect related super-enhancers. Extracellular flux and carbon tracing analyses revealed that HDAC inhibitors blunted glycolysis in a c-Myc dependent manner accompanied by lower ATP levels. This resulted in engagement of oxidative phosphorylation (OXPHOS) driven by elevated fatty acid oxidation (FAO), rendering GBM cells dependent on these pathways. Mechanistically, interference with HDAC1/2 elicited a suppression of c-Myc protein levels and a concomitant increase of two transcriptional drivers of oxidative metabolism, PGC1A and PPARD, suggesting an inverse relationship. Rescue and CHIP experiments indicated that c-Myc bound to the promoter regions of PGC1A and PPARD to counteract their up-regulation driven by HDAC1/2 inhibition. Finally, we demonstrated that the combination treatment of HDAC and FAO inhibitors extended animal survival in patient-derived xenograft (PDX) model systems in vivo more potently than single treatments in the absence of toxicity.
    Keywords:  Intermediary metabolism; Oncology
    DOI:  https://doi.org/10.1172/JCI129049
  24. Geroscience. 2020 Apr 22.
    Mitochondrial adaptations in liver and skeletal muscle to pro-longevity nutritional and genetic interventions: the crosstalk between calorie restriction and CYB5R3 overexpression in transgenic mice.
    Sandra Rodríguez-López, Sara López-Bellón, José A González-Reyes, M Isabel Burón, Rafael de Cabo, José M Villalba.
      Calorie restriction without malnutrition (CR) is considered as the most effective nongenetic nor pharmacological intervention that promotes healthy aging phenotypes and can extend lifespan in most model organisms. Lifelong CR leads to an increase of cytochrome b5 reductase-3 (CYB5R3) expression and activity. Overexpression of CYB5R3 confers some of the salutary effects of CR, although the mechanisms involved might be independent because key aspects of energy metabolism and lipid profiles of tissues go in opposite ways. It is thus important to study if some of the metabolic adaptations induced by CR are affected by CYB5R3 overexpression. CYB5R3 overexpression greatly preserved body and liver weight in mice under CR conditions. In liver, CR did not modify mitochondrial abundance, but lead to increased expression of mitofusin Mfn2 and TFAM, a transcription factor involved in mitochondrial biogenesis. These changes were prevented by CYB5R3 overexpression but resulted in a decreased expression of a different mitochondrial biogenesis-related transcription factor, Nrf1. In skeletal muscle, CR strongly increased mitochondrial mass, mitofusin Mfn1, and Nrf1. However, CYB5R3 mice on CR did not show increase in muscle mitochondrial mass, regardless of a clear increase in expression of TFAM and mitochondrial complexes in this tissue. Our results support that CYB5R3 overexpression significantly modifies the metabolic adaptations of mice to CR.
    Keywords:  Calorie restriction; Cytochrome b5 reductase; Liver; Mitochondria; Skeletal muscle
    DOI:  https://doi.org/10.1007/s11357-020-00187-z
  25. Exp Gerontol. 2020 Apr 20. pii: S0531-5565(20)30309-0. [Epub ahead of print] 110961
    MitoTEMPO provides an antiarrhythmic effect in aged-rats through attenuation of mitochondrial reactive oxygen species.
    Yusuf Olgar, Deniz Billur, Erkan Tuncay, Belma Turan.
      The death prevalence from cardiovascular disease is significantly high in elderly-populations, while mitochondrial-aging plays an important in abnormal function of vital organs through high mitochondrial ROS production. Mitochondria have a unique mode of action by providing ATP production and modulating the cytosolic Ca2+-signaling and maintain the redox status of cardiomyocytes. There is an aging-associated impairment in oxidative phosphorylation which causes a marked dysregulation of mitochondrial biogenesis. Therefore, we aimed to examine whether a mitochondria-targeting antioxidant, MitoTEMPO, can directly provide a cardioprotective effect on ventricular cardiomyocyte function under in vitro conditions. The MitoTEMPO-treatment (0.1 μM for 4-h) of aged-ventricular cardiomyocytes (from 24-mo-old rats), compared to those of the adults (from 8-mo-old rats) markedly augmented not only the depressed biochemical parameters but also the ultrastructure of mitochondria. It also provided marked protective action against increased mitochondrial superoxide formation and Bnip3 overexpression, which both markedly induce depolarized mitochondrial potential, increase reactive oxygen species, mitochondrial swelling and fission, and accelerate mitochondrial turnover via autophagy. Furthermore, it provided marked protection against spontaneous action potentials, via shortening the prolonged action potential duration, at most, through recovery in depressed K+-channel currents. Moreover, we determined significant recovery in the depressed intracellular Ca2+-changes under electrical stimulation in MitoTEMPO-treated the aged-cardiomyocytes. Overall, we provided important information associated with an antiarrhythmic action, thereby controlling cytosolic and mitochondrial Ca2+-handling, implying its possible protective role of mitochondria-targeting antioxidant-treatment during aging.
    Keywords:  Aging-heart; Electrophysiology; Mitochondrial calcium; Mitochondrial reactive oxygen species; Sarcoplasmic reticulum calcium
    DOI:  https://doi.org/10.1016/j.exger.2020.110961
  26. Int J Mol Sci. 2020 Apr 18. pii: E2841. [Epub ahead of print]21(8):
    Para-Halogenation of Amphetamine and Methcathinone Increases the Mitochondrial Toxicity in Undifferentiated and Differentiated SH-SY5Y Cells.
    Xun Zhou, Jamal Bouitbir, Matthias E Liechti, Stephan Krähenbühl, Riccardo V Mancuso.
      Halogenation of amphetamines and methcathinones has become a common method to obtain novel psychoactive substances (NPS) also called "legal highs". The para-halogenated derivatives of amphetamine and methcathinone are available over the internet and have entered the illicit drug market but studies on their potential neurotoxic effects are rare. The primary aim of this study was to explore the neurotoxicity of amphetamine, methcathinone and their para-halogenated derivatives 4-fluoroamphetamine (4-FA), 4-chloroamphetamine (PCA), 4-fluoromethcathinone (4-FMC), and 4-chloromethcathinone (4-CMC) in undifferentiated and differentiated SH-SY5Y cells. We found that 4-FA, PCA, and 4-CMC were cytotoxic (decrease in cellular ATP and plasma membrane damage) for both cell types, whereby differentiated cells were less sensitive. IC50 values for cellular ATP depletion were in the range of 1.4 mM for 4-FA, 0.4 mM for PCA and 1.4 mM for 4-CMC. The rank of cytotoxicity observed for the para-substituents was chloride > fluoride > hydrogen for both amphetamines and cathinones. Each of 4-FA, PCA and 4-CMC decreased the mitochondrial membrane potential in both cell types, and PCA and 4-CMC impaired the function of the electron transport chain of mitochondria in SH-SY5Y cells. 4-FA, PCA, and 4-CMC increased the ROS level and PCA and 4-CMC induced apoptosis by the endogenous pathway. In conclusion, para-halogenation of amphetamine and methcathinone increases their neurotoxic properties due to the impairment of mitochondrial function and induction of apoptosis. Although the cytotoxic concentrations were higher than those needed for pharmacological activity, the current findings may be important regarding the uncontrolled recreational use of these compounds.
    Keywords:  amphetamine; methcathinone; mitochondria; neurotoxicity; para-halogenation
    DOI:  https://doi.org/10.3390/ijms21082841
  27. Pol Przegl Chir. 2019 Nov 04. 92(1): 1-4
    Sirt3 regulates the level of mitochondrial DNA repair activity through deacetylation of NEIL1, NEIL2, OGG1, MUTYH, APE1 and LIG3 in colorectal cancer.
    Jacek Kabziński, Anna Walczak, Michał Mik, Ireneusz Majsterek.
      Colorectal cancer (CRC) is one of the most common malignant tumors. One of the factors increasing the risk of its occurrence may be the reduced efficiency of repairing DNA damage, both nuclear and mitochondrial. The main mechanism for repairing oxidative damage is the BER system (in mitochondria mtBER), whose key proteins NEIL1, NEIL2, OGG1, MUTYH, APE1 and LIG3 obtain full efficiency only at the appropriate level of acetylation. Sirtuin 3 is a key protein for mitochondrial homeostasis, regulating a number of metabolic processes related mainly to the control of the level of reactive oxygen species. Because Sirt3 possesses acetylase activity, it can modulate the level of activity of mtBER proteins by their deacetylation. The conducted study showed that the tested proteins NEIL1, NEIL2, OGG1, MUTYH, APE1 and LIG3 are the substrate for the enzymatic deacetylation activity of Sirt3, which may lead to modulation of the risk of CRC, and in cancer cells may be a potential therapeutic target enhancing the action of cytostatic drugs.
    Keywords:  DNA damage/repair; cancer; colorectal cancer; genetics; proteins
    DOI:  https://doi.org/10.5604/01.3001.0013.5539
  28. Mol Cell Proteomics. 2020 Apr 21. pii: mcp.RA120.002076. [Epub ahead of print]
    HIGD2A is required for assembly of the COX3 module of human mitochondrial complex IV.
    Daniella H Hock, Boris Reljic, Ching-Seng Ang, Linden Muellner-Wong, Hayley S Mountford, Alison G Compton, Michael T Ryan, David R Thorburn, David A Stroud.
      Assembly factors play a critical role in the biogenesis of mitochondrial respiratory chain complexes I-IV where they assist in the membrane insertion of subunits, attachment of co-factors, and stabilization of assembly intermediates. The major fraction of complexes I, III and IV are present together in large molecular structures known as respiratory chain supercomplexes. A number of assembly factors have been proposed as required for supercomplex assembly, including the hypoxia inducible gene 1 domain family member HIGD2A. Using gene-edited human cell lines and extensive steady state, translation and affinity enrichment proteomics techniques we show that loss of HIGD2A leads to defects in the de novo biogenesis of mtDNA-encoded COX3, subsequent accumulation of complex IV intermediates and turnover of COX3 partner proteins. Deletion of HIGD2A also leads to defective complex IV activity. The impact of HIGD2A loss on complex IV was not altered by growth under hypoxic conditions, consistent with its role being in basal complex IV assembly. While in the absence of HIGD2A we show that mitochondria do contain an altered supercomplex assembly, we demonstrate it to harbor a crippled complex IV lacking COX3. Our results redefine HIGD2A as a classical assembly factor required for building the COX3 module of complex IV.
    Keywords:  Cytochrome c oxidase; Energy metabolism; Knockouts*; Mitochondria function or biology; Mitochondrial disease; OXPHOS; Translation*; mitochondria; mtDNA translation; respirasome
    DOI:  https://doi.org/10.1074/mcp.RA120.002076
  29. Biochem Soc Trans. 2020 Apr 20. pii: BST20190930. [Epub ahead of print]
    The road to the structure of the mitochondrial respiratory chain supercomplex.
    Nikeisha J Caruana, David A Stroud.
      The four complexes of the mitochondrial respiratory chain are critical for ATP production in most eukaryotic cells. Structural characterisation of these complexes has been critical for understanding the mechanisms underpinning their function. The three proton-pumping complexes, Complexes I, III and IV associate to form stable supercomplexes or respirasomes, the most abundant form containing 80 subunits in mammals. Multiple functions have been proposed for the supercomplexes, including enhancing the diffusion of electron carriers, providing stability for the complexes and protection against reactive oxygen species. Although high-resolution structures for Complexes III and IV were determined by X-ray crystallography in the 1990s, the size of Complex I and the supercomplexes necessitated advances in sample preparation and the development of cryo-electron microscopy techniques. We now enjoy structures for these beautiful complexes isolated from multiple organisms and in multiple states and together they provide important insights into respiratory chain function and the role of the supercomplex. While we as non-structural biologists use these structures for interpreting our own functional data, we need to remind ourselves that they stand on the shoulders of a large body of previous structural studies, many of which are still appropriate for use in understanding our results. In this mini-review, we discuss the history of respiratory chain structural biology studies leading to the structures of the mammalian supercomplexes and beyond.
    Keywords:  mitochondria; oxidative phosphorylation; structural biology
    DOI:  https://doi.org/10.1042/BST20190930
  30. J Cell Sci. 2020 Apr 23. pii: jcs.242446. [Epub ahead of print]
    A new brain mitochondrial sodium-sensitive potassium channel: effect of sodium ions on respiratory chain activity.
    Javad Fahanik-Babaei, Bahareh Rezaee, Maryam Nazari, Nihad Torabi, Reza Saghiri, Remy Sauve, Afsaneh Eliassi.
      We have determined the electropharmacological properties of a new potassium channel from brain mitochondrial membrane by planar lipid bilayer method. Our results showed the presence of a channel with a conductance of 150 pS at potentials between 0 and -60 mV in 200 cis/50 trans mM KCl solutions.The channel was voltage-independent, with an open probability value ∼0.6 at different voltages. ATP did not affect current amplitude and Po at positive and negative voltages. Notably, adding iberiotoxin, charybdotoxin, lidocaine, and margatoxin had no effect on the channel behavior. Similarly, no changes were observed by decreasing the cis-pH to 6. Interestingly, the channel was inhibited by adding sodium in a dose dependent manner. Our results also indicated a significant increase in mitochondrial complex IV activity and membrane potential and decrease in complex I activity and mitochondrial ROS production in the presence of sodium ions.We propose that inhibition of mitochondrial K+ transport by Na ions on K+ channel opening may be important for cell protection and ATP synthesis.
    Keywords:  Brain; Intracellular ion channel; Mitochondria; Mitochondrial respiratory chain; Potassium channels; Single channel
    DOI:  https://doi.org/10.1242/jcs.242446
  31. Clin Exp Pharmacol Physiol. 2020 Apr 20.
    1,25-Dihydroxyvitamin D3 Prevents Deleterious Effects of Erythromycin on Mitochondrial Function in Rat Heart Isolated Mitochondria.
    Amir Mersa, Saman Atashbar, Negar Ahvar, Ahmad Salimi.
      Since erythromycin (ERY) is a risk factor for cardiotoxicity through mitochondria pathway. In the current study, we tested this hypothesis that erythromycin could impair mitochondrial function and oxidative stress and 1,25-dihydroxivitamin D3 (calcitriol) treatment could prevent these effects in rat heart isolated mitochondria. Rat heart mitochondria were isolated with mechanical lysis and differential centrifugation. Then isolated mitochondria were first pretreated with 3 different concentrations of 1,25-dihydroxivitamin D3 (2.5, 5 and 10 µM) for 5 minutes at 37 °C, after which erythromycin (10 µM) was added to promote deleterious effects on mitochondria. During 1 hour of incubation, using by flow cytometry and biochemical evaluations, the parameters of mitochondrial toxicity including: succinate dehydrogenase (SDH) activity, mitochondrial swelling, mitochondrial membrane potential (MMP) collapse, reactive oxygen species (ROS) formation and lipid peroxidation (LP) was evaluated. The results showed that erythromycin (10 µM) caused a significant change in mitochondrial function, ROS formation, mitochondrial swelling, MMP collapse, increasing lipid peroxidation and oxidative stress. 1,25-dihydroxivitamin D3 (10 µM) reverted the effect of erythromycin on the parameters tested. In this study, we showed that erythromycin impairs mitochondrial function and induces mitochondrial toxicity in rat heart isolated mitochondria, which were reverted by calcitriol. These findings suggest that 1,25-dihydroxivitamin D3 may be a preventive/therapeutic strategy for cardiotoxicity complications caused by erythromycin.
    Keywords:  1,25-dihydroxivitamin D3; Calcitriol; Cardiotoxicity; Erythromycin; Mitochondria
    DOI:  https://doi.org/10.1111/1440-1681.13328
  32. J Aging Res. 2020 ;2020 7232614
    Age- and Organ-Specific Differences in Mitochondrial Bioenergetics in Brown Norway Rats.
    Jignesh D Pandya, Matthew Valdez, Joyce E Royland, Robert C MacPhail, Patrick G Sullivan, Prasada Rao S Kodavanti.
      Mitochondria play a central role in energy homeostasis and act as regulatory checkpoints for downstream metabolic responses and cell senescence processes during an entire life span. Acute or chronic environmental toxicant exposures have shown deleterious organ-specific human health issues at various life stages. Since mitochondria are a prime target for ensuing cellular bioenergetics responses and senescence, it is essential to understand mitochondrial bioenergetic responses in different organs over multiple life stages. Therefore, in the present study, we evaluated mitochondrial bioenergetic parameters in the liver, lung, and heart in four diverse age groups (young: 1 month; adult: 4 months; middle-aged: 12 months; old-aged: 24 month) using male Brown Norway rats as a model of aging (n = 5 sample size/organ/age group) and compared them with our previously published results on brain. Real-time mitochondrial bioenergetic parameters (i.e., State III, State IV, and State V) were measured using the Seahorse Extracellular Flux Analyzer. Additionally, mitochondrial enzyme pyruvate dehydrogenase complex (PDHC), Complex I, Complex II, and Complex IV activities were measured using Synergy HT plate reader. Our results indicated that nearly in all parameters, significant age- and organ-specific interactions were observed. We observed age-specific declines in State III (i.e., ATP synthesis rate) responses in both the heart and lung, where opposite was observed in the liver as age advances. Across the age, the heart has highest enzyme activities than the liver and lung. Interestingly, heart and liver mitochondrial bioenergetic rates and enzyme activities remain higher than the lung, which specifies their higher metabolic capabilities than the lung. Amongst all, bioenergetic rates and enzyme activities in the lung remain lowest suggesting the lung may display higher vulnerability and lower resilience to environmental toxicants during aging than other organs tested here. Overall, these age- and organ-specific findings may facilitate a more contextualized understanding of mitochondrial bioenergetic outcomes when considering the interactions of age-related sensitivities with exposure to chemical stressors from the environment.
    DOI:  https://doi.org/10.1155/2020/7232614
  33. Cell Metab. 2020 Apr 17. pii: S1550-4131(20)30189-3. [Epub ahead of print]
    Developmental and Tumor Angiogenesis Requires the Mitochondria-Shaping Protein Opa1.
    Stéphanie Herkenne, Olivier Ek, Margherita Zamberlan, Anna Pellattiero, Maya Chergova, Iñigo Chivite, Eliška Novotná, Giovanni Rigoni, Tiago Branco Fonseca, Dijana Samardzic, Andrielly Agnellini, Camilla Bean, Giulietta Di Benedetto, Natascia Tiso, Francesco Argenton, Antonella Viola, Maria Eugenia Soriano, Marta Giacomello, Elena Ziviani, Gabriele Sales, Marc Claret, Mariona Graupera, Luca Scorrano.
      While endothelial cell (EC) function is influenced by mitochondrial metabolism, the role of mitochondrial dynamics in angiogenesis, the formation of new blood vessels from existing vasculature, is unknown. Here we show that the inner mitochondrial membrane mitochondrial fusion protein optic atrophy 1 (OPA1) is required for angiogenesis. In response to angiogenic stimuli, OPA1 levels rapidly increase to limit nuclear factor kappa-light-chain-enhancer of activated B cell (NFκB) signaling, ultimately allowing angiogenic genes expression and angiogenesis. Endothelial Opa1 is indeed required in an NFκB-dependent pathway essential for developmental and tumor angiogenesis, impacting tumor growth and metastatization. A first-in-class small molecule-specific OPA1 inhibitor confirms that EC Opa1 can be pharmacologically targeted to curtail tumor growth. Our data identify Opa1 as a crucial component of physiological and tumor angiogenesis.
    Keywords:  NFκB; Opa1; angiogenesis; cancer; fusion-fission; lymphangiogenesis; metastasis; mitochondria; mouse; tumor; zebrafish
    DOI:  https://doi.org/10.1016/j.cmet.2020.04.007
  34. Oxid Med Cell Longev. 2020 ;2020 1675613
    IMCA Induces Ferroptosis Mediated by SLC7A11 through the AMPK/mTOR Pathway in Colorectal Cancer.
    Lei Zhang, Wen Liu, Fangyan Liu, Qun Wang, Mengjiao Song, Qi Yu, Kun Tang, Tieshan Teng, Dongdong Wu, Xijing Wang, Wuqi Han, Yanzhang Li.
      Ferroptosis, implicated in several diseases, is a new form of programmed and nonapoptotic cell death triggered by iron-dependent lipid peroxidation after inactivation of the cystine/glutamate antiporter system xc-, which is composed of solute carrier family 7 membrane 11 (SLC7A11) and solute carrier family 3 membrane 2 (SLC3A2). Therefore, inducing ferroptosis through inhibiting the cystine/glutamate antiporter system xc- may be an effective way to treat cancer. In previous screening tests, we found that the benzopyran derivative 2-imino-6-methoxy-2H-chromene-3-carbothioamide (IMCA) significantly inhibited the viability of colorectal cancer cells. However, the impact of IMCA on ferroptosis remains unknown. Hence, this study investigated the effect of IMCA on ferroptosis and elucidated the underlying molecular mechanism. Results showed that IMCA significantly inhibited the cell viability of colorectal cancer cells in vitro and inhibited tumor growth with negligible organ toxicity in vivo. Further studies showed that IMCA significantly induced the ferroptosis of colorectal cancer cells. Mechanistically, IMCA downregulated the expression of SLC7A11 and decreased the contents of cysteine and glutathione, which resulted in reactive oxygen species accumulation and ferroptosis. Furthermore, overexpression of SLC7A11 significantly attenuated the ferroptosis caused by IMCA. In addition, IMCA regulated the activity of the AMPK/mTOR/p70S6k signaling pathway, which is related to the activity of SLC7A11 and ferroptosis. Collectively, our research provided experimental evidences on the activity and mechanism of ferroptosis induced by IMCA and revealed that IMCA might be a promising therapeutic drug for colorectal cancer.
    DOI:  https://doi.org/10.1155/2020/1675613
  35. Clin Cancer Res. 2020 Apr 22. pii: clincanres.0269.2020. [Epub ahead of print]
    AMPK activation by metformin promotes survival of dormant ER+ breast cancer cells.
    Riley A Hampsch, Jason D Wells, Nicole A Traphagen, Charlotte F McCleery, Jennifer L Fields, Kevin Shee, Lloye M Dillon, Darcy B Pooler, Lionel D Lewis, Eugene Demidenko, Yina H Huang, Jonathan D Marotti, Abigail E Goen, William B Kinlaw, Todd W Miller.
       PURPOSE: Despite adjuvant endocrine therapy for patients with estrogen receptor alpha (ER)-positive breast cancer, dormant residual disease can persist for years and eventually cause tumor recurrence. We sought to deduce mechanisms underlying the persistence of dormant cancer cells to identify therapeutic strategies.
    EXPERIMENTAL DESIGN: Mimicking the aromatase inhibitor-induced depletion of estrogen levels used to treat patients, we developed preclinical models of dormancy in ER+ breast cancer induced by estrogen withdrawal in mice. We analyzed tumor xenografts and cultured cancer cells for molecular and cellular responses to estrogen withdrawal and drug treatments. Publicly available clinical breast tumor gene expression datasets were analyzed for responses to neoadjuvant endocrine therapy.
    RESULTS: Dormant breast cancer cells exhibited upregulated 5' adenosine monophosphate-activated protein kinase (AMPK) levels and activity, and upregulated fatty acid oxidation. While the anti-diabetes AMPK-activating drug metformin slowed the estrogen-driven growth of cells and tumors, metformin promoted the persistence of estrogen-deprived cells and tumors through increased mitochondrial respiration driven by fatty acid oxidation. Pharmacologic or genetic inhibition of AMPK or fatty acid oxidation promoted clearance of dormant residual disease, while dietary fat increased tumor cell survival.
    CONCLUSIONS: AMPK has context-dependent effects in cancer, cautioning against the widespread use of an AMPK activator across disease settings. The development of therapeutics targeting fat metabolism is warranted in ER+ breast cancer.
    DOI:  https://doi.org/10.1158/1078-0432.CCR-20-0269
  36. ChemMedChem. 2020 Apr 20.
    Pentathiepins: a novel class of glutathione peroxidase 1 inhibitors that induce oxidative stress, loss of mitochondrial membrane potential and apoptosis in human cancer cells.
    Steven Behnisch-Cornwell, Siva Sankar Murthy Bandarub, Martin Napierkowski, Lisa Wolff, Muhammad Zubair, Claudia Urbainsky, Christopher Lillig, Carola Schulzke, Patrick Bednarski.
      A novel class of glutathione peroxidase 1 (GPx1) inhibitors, namely tri- and tetracyclic- pentathiepins, has been identified that is ca. 15 times more potent than the most active known GPx1 inhibitor, mercaptosuccinic acid. Enzyme kinetic studies with bovine erythrocyte GPx1 indicate that pentathiepins reversibly inhibit oxidation of the substrate glutathione (GSH). Moreover, no inhibition of superoxide dismutase, catalase, thioredoxin reductase or glutathione reductase was observed at concentrations that effectively inhibit GPx1. As well as potent enzyme inhibitory activity, the pentathiepins also show strong anticancer activity in various human cancer cell lines, with IC 50 values in a low micro molar range. A representative tetracyclic pentathiepin causes formation of reactive oxygen species in these cells, fragmentation of nuclear DNA and induces apoptosis via the intrinsic pathway. Moreover, this pentathiepin leads to a rapid and strong loss of mitochondrial membrane potential in treated cancer cells. On the other hand, evidence for the induction of ferroptosis as a form of cell death was negative. These new findings show that pentathiepins possess interesting biological activities beyond those originally ascribed to these compounds.
    Keywords:  Pentathiepin; cancer cells; cytotoxicity; glutathione peroxidase; oxidative stress
    DOI:  https://doi.org/10.1002/cmdc.202000160
  37. Free Radic Res. 2020 Apr 24. 1-9
    Quantifying mitochondrial respiration in human lymphocytes and monocytes challenged with hydrogen peroxide.
    Andree G Pearson, Masuma Zawari, John F Pearson, Mark B Hampton.
      Improved technology for the bioenergetic profiling of human blood cells enables population-based screening for alterations in mitochondrial respiration. Mitochondria are sensitive to oxidative stress, and the aim of this study was to quantify mitochondrial respiration in freshly isolated lymphocytes and monocytes challenged with a bolus of H2O2. Mitochondrial reserve capacity, calculated as the difference between basal oxygen consumption and maximal activity after uncoupling of the electron transport chain, was the most sensitive to H2O2. Treatment of lymphocytes with 20 μM H2O2 reduced the reserve capacity by approximately 50%, while monocyte reserve capacity was five times more resistant. Healthy donors of a similar age were tested to determine the variation between individuals, and within the same individuals tested on several different occasions. Lymphocytes obtained from a population of people aged 70-80 years showed a similar inhibition upon challenge with H2O2 as those aged 18-25 years, indicating no decline in resilience with age.
    Keywords:  Oxidative stress; ageing; bioenergetics; hydrogen peroxide; mitochondrial respiration
    DOI:  https://doi.org/10.1080/10715762.2020.1753722
  38. Front Oncol. 2020 ;10 435
    Utilizing Synergistic Potential of Mitochondria-Targeting Drugs for Leukemia Therapy.
    Svetlana B Panina, Jingqi Pei, Natalia Baran, Marina Konopleva, Natalia V Kirienko.
      Acute myeloid leukemia (AML) is an aggressive group of cancers with high mortality rates and significant relapse risks. Current treatments are insufficient, and new therapies are needed. Recent discoveries suggest that AML may be particularly sensitive to chemotherapeutics that target mitochondria. To further investigate this sensitivity, six compounds that target mitochondria [IACS-010759, rotenone, cytarabine, etoposide, ABT-199 (venetoclax), and carbonyl cyanide m-chlorophenylhydrazone] were each paired with six compounds with other activities, including tyrosine kinase inhibitors (midostaurin and dasatinib), glycolytic inhibitors (2-deoxy-D-glucose, 3-bromopyruvate, and lonidamine), and the microtubule destabilizer vinorelbine. The 36 resulting drug combinations were tested for synergistic cytotoxicity against MOLM-13 and OCI-AML2 AML cell lines. Four combinations (IACS-010759 with vinorelbine, rotenone with 2-deoxy-D-glucose, carbonyl cyanide m-chlorophenylhydrazone with dasatinib, and venetoclax with lonidamine) showed synergistic cytotoxicity in both AML cell lines and were selective for tumor cells, as survival of healthy PBMCs was dramatically higher. Among these drug pairs, IACS-010759/vinorelbine decreased ATP level and impaired mitochondrial respiration and coupling efficiency most profoundly. Some of these four treatments were also effective in K-562, KU812 (chronic myelogenous leukemia) and CCRF-CEM, MOLT-4 (acute lymphoblastic leukemia) cells, suggesting that these treatments may have value in treating other forms of leukemia. Finally, two of the four combinations retained high synergy and strong selectivity in primary AML cells from patient samples, supporting the potential of these treatments for patients.
    Keywords:  AML; leukemia treatment; mitocans; mitochondria-targeted drugs; synergistic drug combinations; targeted drug screening
    DOI:  https://doi.org/10.3389/fonc.2020.00435
  39. J Clin Invest. 2020 Apr 23. pii: 131859. [Epub ahead of print]
    Targeting glutamine metabolism enhances tumor specific immunity by modulating suppressive myeloid cells.
    Min-Hee Oh, Im-Hong Sun, Liang Zhao, Robert D Leone, Im-Meng Sun, Wei Xu, Samuel L Collins, Ada J Tam, Richard L Blosser, Chirag H Patel, Judson M Englert, Matthew L Arwood, Jiayu Wen, Yee Chan-Li, Lukáš Tenora, Pavel Majer, Rana Rais, Barbara S Slusher, Maureen R Horton, Jonathan D Powell.
      Myeloid cells comprise a major component of the tumor-microenvironment (TME) promoting tumor growth and immune evasion. By employing a novel small molecule inhibitor of glutamine metabolism, not only were we able to inhibit tumor growth, but we markedly inhibited the generation and recruitment of myeloid-derived suppressor cells (MDSCs). Targeting tumor glutamine metabolism led to a decrease in CSF3 and hence recruitment of MDSCs as well immunogenic cell death leading to an increase in inflammatory tumor-associated macrophages (TAMs). Alternatively, inhibiting glutamine metabolism of the MDSCs themselves led to activation induced cell death and conversion of MDSCs to inflammatory macrophages. Surprisingly, blocking glutamine metabolism also inhibited IDO expression of both the tumor and myeloid derived cells leading to a marked decrease in kynurenine levels. This in turn inhibited the development of metastasis and further enhanced anti-tumor immunity. Indeed, targeting glutamine metabolism rendered checkpoint blockade-resistant tumors susceptible to immunotherapy. Overall, our studies define an intimate interplay between the unique metabolism of tumors and the metabolism of suppressive immune cells.
    Keywords:  Cancer immunotherapy; Immunology; Innate immunity; Oncology
    DOI:  https://doi.org/10.1172/JCI131859
  40. Proc Natl Acad Sci U S A. 2020 Apr 24. pii: 201919410. [Epub ahead of print]
    Sample multiplexing for targeted pathway proteomics in aging mice.
    Qing Yu, Haopeng Xiao, Mark P Jedrychowski, Devin K Schweppe, Jose Navarrete-Perea, Jeffrey Knott, John Rogers, Edward T Chouchani, Steven P Gygi.
      Pathway proteomics strategies measure protein expression changes in specific cellular processes that carry out related functions. Using targeted tandem mass tags-based sample multiplexing, hundreds of proteins can be quantified across 10 or more samples simultaneously. To facilitate these highly complex experiments, we introduce a strategy that provides complete control over targeted sample multiplexing experiments, termed Tomahto, and present its implementation on the Orbitrap Tribrid mass spectrometer platform. Importantly, this software monitors via the external desktop computer to the data stream and inserts optimized MS2 and MS3 scans in real time based on an application programming interface with the mass spectrometer. Hundreds of proteins of interest from diverse biological samples can be targeted and accurately quantified in a sensitive and high-throughput fashion. It achieves sensitivity comparable to, if not better than, deep fractionation and requires minimal total sample input (∼10 µg). As a proof-of-principle experiment, we selected four pathways important in metabolism- and inflammation-related processes (260 proteins/520 peptides) and measured their abundance across 90 samples (nine tissues from five old and five young mice) to explore effects of aging. Tissue-specific aging is presented here and we highlight the role of inflammation- and metabolism-related processes in white adipose tissue. We validated our approach through comparison with a global proteome survey across the tissues, work that we also provide as a general resource for the community.
    Keywords:  Tomahto; isobaric labeling; real-time instrument control; targeted pathway proteomics; tissue-specific aging
    DOI:  https://doi.org/10.1073/pnas.1919410117
  41. Cell Rep. 2020 Apr 21. pii: S2211-1247(20)30431-9. [Epub ahead of print]31(3): 107531
    Rational Design of Allosteric and Selective Inhibitors of the Molecular Chaperone TRAP1.
    Carlos Sanchez-Martin, Elisabetta Moroni, Mariarosaria Ferraro, Claudio Laquatra, Giuseppe Cannino, Ionica Masgras, Alessandro Negro, Paolo Quadrelli, Andrea Rasola, Giorgio Colombo.
      TRAP1 is the mitochondrial paralog of the heat shock protein 90 (HSP90) chaperone family. Its activity as an energy metabolism regulator has important implications in cancer, neurodegeneration, and ischemia. Selective inhibitors of TRAP1 could inform on its mechanisms of action and set the stage for targeted drug development, but their identification was hampered by the similarity among active sites in HSP90 homologs. We use a dynamics-based approach to identify a TRAP1 allosteric pocket distal to its active site that can host drug-like molecules, and we select small molecules with optimal stereochemical features to target the pocket. These leads inhibit TRAP1, but not HSP90, ATPase activity and revert TRAP1-dependent downregulation of succinate dehydrogenase activity in cancer cells and in zebrafish larvae. TRAP1 inhibitors are not toxic per se, but they abolish tumorigenic growth of neoplastic cells. Our results indicate that exploiting conformational dynamics can expand the chemical space of chaperone antagonists to TRAP1-specific inhibitors with wide therapeutic opportunities.
    Keywords:  HSP90; TRAP1; allosteric ligands; anticancer compound; cancer cells; chaperone inhibitors; mitochondria; mitochondrial biology; molecular dynamics; neurofibroma; zebrafish
    DOI:  https://doi.org/10.1016/j.celrep.2020.107531
  42. Int J Obes (Lond). 2020 Apr 21.
    Tankyrase inhibition ameliorates lipid disorder via suppression of PGC-1α PARylation in db/db mice.
    Hong Wang, Sara Kuusela, Rita Rinnankoski-Tuikka, Vincent Dumont, Rim Bouslama, Usama Abo Ramadan, Jo Waaler, Anni-Maija Linden, Nai-Wen Chi, Stefan Krauss, Eija Pirinen, Sanna Lehtonen.
       OBJECTIVE: Human TNKS, encoding tankyrase 1 (TNKS1), localizes to a susceptibility locus for obesity and type 2 diabetes mellitus (T2DM). Here, we addressed the therapeutic potential of G007-LK, a TNKS-specific inhibitor, for obesity and T2DM.
    METHODS: We administered G007-LK to diabetic db/db mice and measured the impact on body weight, abdominal adiposity, and serum metabolites. Muscle, liver, and white adipose tissues were analyzed by quantitative RT-PCR and western blotting to determine TNKS inhibition, lipolysis, beiging, adiponectin level, mitochondrial oxidative metabolism and mass, and gluconeogenesis. Protein interaction and PARylation analyses were carried out by immunoprecipitation, pull-down and in situ proximity ligation assays.
    RESULTS: TNKS inhibition reduced body weight gain, abdominal fat content, serum cholesterol levels, steatosis, and proteins associated with lipolysis in diabetic db/db mice. We discovered that TNKS associates with PGC-1α and that TNKS inhibition attenuates PARylation of PGC-1α, contributing to increased PGC-1α level in WAT and muscle in db/db mice. PGC-1α upregulation apparently modulated transcriptional reprogramming to increase mitochondrial mass and fatty acid oxidative metabolism in muscle, beiging of WAT, and raised circulating adiponectin level in db/db mice. This was in sharp contrast to the liver, where TNKS inhibition in db/db mice had no effect on PGC-1α expression, lipid metabolism, or gluconeogenesis.
    CONCLUSION: Our study unravels a novel molecular mechanism whereby pharmacological inhibition of TNKS in obesity and diabetes enhances oxidative metabolism and ameliorates lipid disorder. This happens via tissue-specific PGC-1α-driven transcriptional reprogramming in muscle and WAT, without affecting liver. This highlights inhibition of TNKS as a potential pharmacotherapy for obesity and T2DM.
    DOI:  https://doi.org/10.1038/s41366-020-0573-z
  43. Oxid Med Cell Longev. 2020 ;2020 5649174
    Plumbagin Enhances the Anticancer Efficacy of Cisplatin by Increasing Intracellular ROS in Human Tongue Squamous Cell Carcinoma.
    Danfeng Xue, Shu-Ting Pan, Xiongming Zhou, Fangfei Ye, Qun Zhou, Fanzhe Shi, Fei He, Hui Yu, Jiaxuan Qiu.
      Cisplatin is widely used in the treatment of tongue squamous cell carcinoma (TSCC), but its clinical efficacy is limited by drug resistance and toxic side effects. Hence, a novel compound capable of enhancing the anticancer effect of cisplatin while reducing the side effects is urgently needed. We have previously shown that plumbagin (PLB), an anticancer phytochemical, is able to inhibit the growth of TSCC in vitro and in vivo. The objective of this study was to investigate the effect of PLB in reversing the resistance of TSCC to cisplatin as well as its molecular mechanisms. Here, we found that PLB enhances cisplatin-induced cytotoxicity, apoptosis, and autophagy in CAL27 and cisplatin-resistant CAL27/CDDP cells. PLB could inhibit the viability and growth of TSCC cells by increasing the production of intracellular reactive oxygen species (ROS). In addition, the combination treatment of PLB and cisplatin resulted in a synergistic inhibition of TSCC viability, apoptosis, and autophagy by increasing intracellular ROS, which may be achieved by activating JNK and inhibiting AKT/mTOR signaling pathways. Finally, the synergistic treatment was also demonstrated in vivo. Therefore, PLB combined with cisplatin is a potential therapeutic strategy against therapy TSCC cisplatin resistance.
    DOI:  https://doi.org/10.1155/2020/5649174
  44. Am J Clin Nutr. 2020 Apr 22. pii: nqaa072. [Epub ahead of print]
    Nicotinamide riboside supplementation alters body composition and skeletal muscle acetylcarnitine concentrations in healthy obese humans.
    Carlijn M E Remie, Kay H M Roumans, Michiel P B Moonen, Niels J Connell, Bas Havekes, Julian Mevenkamp, Lucas Lindeboom, Vera H W de Wit, Tineke van de Weijer, Suzanne A B M Aarts, Esther Lutgens, Bauke V Schomakers, Hyung L Elfrink, Rubén Zapata-Pérez, Riekelt H Houtkooper, Johan Auwerx, Joris Hoeks, Vera B Schrauwen-Hinderling, Esther Phielix, Patrick Schrauwen.
       BACKGROUND: Nicotinamide riboside (NR) is an NAD+ precursor that boosts cellular NAD+ concentrations. Preclinical studies have shown profound metabolic health effects after NR supplementation.
    OBJECTIVES: We aimed to investigate the effects of 6 wk NR supplementation on insulin sensitivity, mitochondrial function, and other metabolic health parameters in overweight and obese volunteers.
    METHODS: A randomized, double-blinded, placebo-controlled, crossover intervention study was conducted in 13 healthy overweight or obese men and women. Participants received 6 wk NR (1000 mg/d) and placebo supplementation, followed by broad metabolic phenotyping, including hyperinsulinemic-euglycemic clamps, magnetic resonance spectroscopy, muscle biopsies, and assessment of ex vivo mitochondrial function and in vivo energy metabolism.
    RESULTS: Markers of increased NAD+ synthesis-nicotinic acid adenine dinucleotide and methyl nicotinamide-were elevated in skeletal muscle after NR compared with placebo. NR increased body fat-free mass (62.65% ± 2.49% compared with 61.32% ± 2.58% in NR and placebo, respectively; change: 1.34% ± 0.50%, P = 0.02) and increased sleeping metabolic rate. Interestingly, acetylcarnitine concentrations in skeletal muscle were increased upon NR (4558 ± 749 compared with 3025 ± 316 pmol/mg dry weight in NR and placebo, respectively; change: 1533 ± 683 pmol/mg dry weight, P = 0.04) and the capacity to form acetylcarnitine upon exercise was higher in NR than in placebo (2.99 ± 0.30 compared with 2.40 ± 0.33 mmol/kg wet weight; change: 0.53 ± 0.21 mmol/kg wet weight, P = 0.01). However, no effects of NR were found on insulin sensitivity, mitochondrial function, hepatic and intramyocellular lipid accumulation, cardiac energy status, cardiac ejection fraction, ambulatory blood pressure, plasma markers of inflammation, or energy metabolism.
    CONCLUSIONS: NR supplementation of 1000 mg/d for 6 wk in healthy overweight or obese men and women increased skeletal muscle NAD+ metabolites, affected skeletal muscle acetylcarnitine metabolism, and induced minor changes in body composition and sleeping metabolic rate. However, no other metabolic health effects were observed.This trial was registered at clinicaltrials.gov as NCT02835664.
    Keywords:  NAD; acetylcarnitine; body composition; human; insulin sensitivity; metabolic health; mitochondrial function; nicotinamide riboside; obesity
    DOI:  https://doi.org/10.1093/ajcn/nqaa072
  45. Cell Rep. 2020 Apr 21. pii: S2211-1247(20)30438-1. [Epub ahead of print]31(3): 107538
    m.3243A > G-Induced Mitochondrial Dysfunction Impairs Human Neuronal Development and Reduces Neuronal Network Activity and Synchronicity.
    Teun M Klein Gunnewiek, Eline J H Van Hugte, Monica Frega, Gemma Solé Guardia, Katharina Foreman, Daan Panneman, Britt Mossink, Katrin Linda, Jason M Keller, Dirk Schubert, David Cassiman, Richard Rodenburg, Noemi Vidal Folch, Devin Oglesbee, Ester Perales-Clemente, Timothy J Nelson, Eva Morava, Nael Nadif Kasri, Tamas Kozicz.
      Epilepsy, intellectual and cortical sensory deficits, and psychiatric manifestations are the most frequent manifestations of mitochondrial diseases. How mitochondrial dysfunction affects neural structure and function remains elusive, mostly because of a lack of proper in vitro neuronal model systems with mitochondrial dysfunction. Leveraging induced pluripotent stem cell technology, we differentiated excitatory cortical neurons (iNeurons) with normal (low heteroplasmy) and impaired (high heteroplasmy) mitochondrial function on an isogenic nuclear DNA background from patients with the common pathogenic m.3243A > G variant of mitochondrial encephalomyopathy, lactic acidosis, and stroke-like episodes (MELAS). iNeurons with high heteroplasmy exhibited mitochondrial dysfunction, delayed neural maturation, reduced dendritic complexity, and fewer excitatory synapses. Micro-electrode array recordings of neuronal networks displayed reduced network activity and decreased synchronous network bursting. Impaired neuronal energy metabolism and compromised structural and functional integrity of neurons and neural networks could be the primary drivers of increased susceptibility to neuropsychiatric manifestations of mitochondrial disease.
    Keywords:  MELAS; induced pluripotent stem cells; m.3243A > G; micro-electrode array; mitochondria; mitochondrial disease; network activity; neurodevelopment; neuron
    DOI:  https://doi.org/10.1016/j.celrep.2020.107538
  46. Int J Oncol. 2020 Apr 08.
    CD38 is involved in cell energy metabolism via activating the PI3K/AKT/mTOR signaling pathway in cervical cancer cells.
    Shan Liao, Lin Liang, Chunxue Yue, Junyu He, Zhengxi He, Xi Jin, Gengqiu Luo, Yanhong Zhou.
      In contrast to normal cells, cancer cells typically undergo metabolic reprogramming. Studies have shown that oncogenes play an important role in this metabolic reprogramming. CD38 is a multifunctional transmembrane protein that is expressed abnormally in a variety of tumor types. To investigate the effect and possible mechanism of CD38 in cervical cancer cells and to provide a new therapeutic target for the treatment of cervical cancer, the present study identified that CD38 is involved in regulating cell metabolism in cervical cancer cells. Liquid chromatography‑tandem mass spectrometry and bioinformatic analyses revealed that differentially abundant proteins in CD38‑overexpressed cervical cancer cells (CaSki‑CD38 and HeLa‑CD38) are predominantly involved in glycolytic pathways, oxidative phosphorylation and the NAD/NADH metabolic process. Further experiments using an ATP test kit and lactate test kit revealed that CD38 promotes glucose consumption, increases lactate accumulation and increases ATP production. In addition, CD38 increases the phosphorylation of phosphatidylserine/threonine kinase (AKT), mechanistic target of rapamycin (mTOR) and phosphatidylinositol‑4,5‑bisphosphate 3‑kinase (PI3K), which play a key role in tumor metabolism. Furthermore, it was found that the energy metabolism of cervical cancer cells was inhibited following treatment with the mTOR inhibitor rapamycin. In conclusion, the results of the present study suggested that CD38 regulates the metabolism of cervical cancer cells by regulating the PI3K/AKT/mTOR pathway, which may be a candidate target for the treatment of cervical cancer.
    DOI:  https://doi.org/10.3892/ijo.2020.5040
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