bims-mibica Biomed News
on Mitochondrial bioenergetics in cancer
Issue of 2020‒04‒12
forty-nine papers selected by
Kelsey Fisher-Wellman
East Carolina University


  1. Carcinogenesis. 2020 Apr 07. pii: bgaa032. [Epub ahead of print]
    Haumann S, Boix J, Knuever J, Bieling A, Vila Sanjurjo A, Elson JL, Blakely EL, Taylor RW, Riet N, Abken H, Kashkar H, Hornig-Do HT, Wiesner RJ.
      Functioning mitochondria are crucial for cancer metabolism, but aerobic glycolysis is still considered to be an important pathway for energy production in many tumor cells. Here we show that two well established, classic Hodgkin lymphoma cell lines (cHL) harbor deleterious variants within mitochondrial DNA (mtDNA) and thus exhibit reduced steady state levels of respiratory chain complexes. However, instead of resulting in the expected bioenergetic defect, these mtDNA variants evoke a retrograde signaling response that induces mitochondrial biogenesis and ultimately results in increased mitochondrial mass as well as function and enhances proliferation in vitro as well as tumor growth in mice in vivo. When complex I assembly was impaired by knock-down of one of its subunits, this led to further increased mitochondrial mass and function and, consequently, further accelerated tumor growth in vivo. In contrast, inhibition of mitochondrial respiration in vivo by the mitochondrial complex I inhibitor metformin efficiently slowed down growth. We conclude that, as a new mechanism, mildly deleterious mtDNA variants in cHL cancer cells cause an increase of mitochondrial mass and enhanced function as a compensatory effect using a retrograde signaling pathway, which provides an obvious advantage for tumor growth.
    DOI:  https://doi.org/10.1093/carcin/bgaa032
  2. Biochem J. 2020 Apr 09. pii: BCJ20200042. [Epub ahead of print]
    Bayev AY, Angelova PR, Abramov AY.
      Inorganic polyphosphate (polyP) is a polymer present in all living organisms. Although polyP is found to be involved in a variety of functions in cells of higher organisms, the enzyme responsible for polyP production and consumption has not yet been identified. Here we studied the effect of polyP on mitochondrial respiration, oxidative phosphorylation and activity of F0F1-ATPsynthase. We have found that polyP activates mitochondrial respiration which does not coupled with ATP production (V2) but inhibits ADP dependent respiration (V3). Moreover, PolyP can stimulate F0F1-ATPase activity in the presence of ATP and, importantly, can be hydrolysed in this enzyme instead of ATP. Furthermore, PolyP can be produced in mitochondria in the presence of substrates for respiration and phosphate by the F0F1-ATPsynthase. Thus, polyP is an energy molecule in mammalian cells which can be produced and hydrolysed in the mitochondrial F0F1-ATPsynthase.
    Keywords:  F0-F1-ATPase; bioenergetics; inorganic polyphosphates; mitochondria
    DOI:  https://doi.org/10.1042/BCJ20200042
  3. J Cell Mol Med. 2020 Apr 06.
    Alimujiang M, Yu XY, Yu MY, Hou WL, Yan ZH, Yang Y, Bao YQ, Yin J.
      Mitochondrial function is critical in energy metabolism. To fully capture how the mitochondrial function changes in metabolic disorders, we investigated mitochondrial function in liver and muscle of animal models mimicking different types and stages of diabetes. Type 1 diabetic mice were induced by streptozotocin (STZ) injection. The db/db mice were used as type 2 diabetic model. High-fat diet-induced obese mice represented pre-diabetic stage of type 2 diabetes. Oxidative phosphorylation (OXPHOS) of isolated mitochondria was measured with Clark-type oxygen electrode. Both in early and late stages of type 1 diabetes, liver mitochondrial OXPHOS increased markedly with complex IV-dependent OXPHOS being the most prominent. However, ATP, ADP and AMP contents in the tissue did not change. In pre-diabetes and early stage of type 2 diabetes, liver mitochondrial complex I and II-dependent OXPHOS increased greatly then declined to almost normal at late stage of type 2 diabetes, among which alteration of complex I-dependent OXPHOS was the most significant. In contrast, muscle mitochondrial OXPHOS in HFD, early-stage type 1 and 2 diabetic mice, did not change. In vitro, among inhibitors to each complex, only complex I inhibitor rotenone decreased glucose output in primary hepatocytes without cytotoxicity both in the absence and presence of oleic acid (OA). Rotenone affected cellular energy state and had no effects on cellular and mitochondrial reactive oxygen species production. Taken together, the mitochondrial OXPHOS of liver but not muscle increased in obesity and diabetes, and only complex I inhibition may ameliorate hyperglycaemia via lowering hepatic glucose production.
    Keywords:  NAFLD; ROS; diabetes; electron transport chain; insulin resistance; liver steatosis; obesity
    DOI:  https://doi.org/10.1111/jcmm.15238
  4. Cancers (Basel). 2020 Apr 08. pii: E913. [Epub ahead of print]12(4):
    Min HY, Pei H, Hyun SY, Boo HJ, Jang HJ, Cho J, Kim JH, Son J, Lee HY.
      Metabolic rewiring to utilize aerobic glycolysis is a hallmark of cancer. However, recent findings suggest the role of mitochondria in energy generation in cancer cells and the metabolic switch to oxidative phosphorylation (OXPHOS) in response to the blockade of glycolysis. We previously demonstrated that the antitumor effect of gracillin occurs through the inhibition of mitochondrial complex II-mediated energy production. Here, we investigated the potential of gracillin as an anticancer agent targeting both glycolysis and OXPHOS in breast and lung cancer cells. Along with the reduction in adenosine triphosphate (ATP) production, gracillin markedly suppresses the production of several glycolysis-associated metabolites. A docking analysis and enzyme assay suggested phosphoglycerate kinase 1 (PGK1) is a potential target for the antiglycolytic effect of gracillin. Gracillin reduced the viability and colony formation ability of breast cancer cells by inducing apoptosis. Gracillin displayed efficacious antitumor effects in mice bearing breast cancer cell line or breast cancer patient-derived tumor xenografts with no overt changes in body weight. An analysis of publicly available datasets further suggested that PGK1 expression is associated with metastasis status and poor prognosis in patients with breast cancer. These results suggest that gracillin is a natural anticancer agent that inhibits both glycolysis and mitochondria-mediated bioenergetics.
    Keywords:  breast cancer; glycolysis; gracillin; lung cancer; oxidative phosphorylation; phosphoglycerate kinase 1
    DOI:  https://doi.org/10.3390/cancers12040913
  5. Biosci Biotechnol Biochem. 2020 Apr 07. 1-10
    Murai M.
      NADH-quinone oxidoreductase (respiratory complex I) is a key player in mitochondrial energy metabolism. The enzyme couples electron transfer from NADH to quinone with the translocation of protons across the membrane, providing a major proton-motive force that drives ATP synthesis. Recently, X-ray crystallography and cryo-electron microscopy provided further insights into the structure and functions of the enzyme. However, little is known about the mechanism of quinone reduction, which is a crucial step in the energy coupling process. A variety of complex I inhibitors targeting the quinone-binding site have been indispensable tools for mechanistic studies on the enzyme. Using biorationally designed inhibitor probes, the author has accumulated a large amount of experimental data characterizing the actions of complex I inhibitors. On the basis of comprehensive interpretations of the data, the author reviews the structural features of the binding pocket of quinone/inhibitors in bovine mitochondrial complex I.Abbreviations: ATP: adenosine triphosphate; BODIPY: boron dipyrromethene; complex I: proton-translocating NADH-quinone oxidoreductase; DIBO: dibenzocyclooctyne; EM: electron microscopy; FeS: iron-sulfur; FMN: flavin adenine mononucleotide; LDT: ligand-directed tosylate; NADH: nicotinamide adenine dinucleotide; ROS: reactive oxygen species; SMP: submitochondrial particle; TAMRA: 6-carboxy-N,N,N',N'-tetramethylrhodamine; THF: tetrahydrofuran; TMH: transmembrane helix.
    Keywords:  Respiratory complex I; chemical biology; inhibitor; mitochondria; ubiquinone
    DOI:  https://doi.org/10.1080/09168451.2020.1747974
  6. Cells. 2020 Apr 04. pii: E884. [Epub ahead of print]9(4):
    Kuffner K, Triebelhorn J, Meindl K, Benner C, Manook A, Sudria-Lopez D, Siebert R, Nothdurfter C, Baghai TC, Drexler K, Berneburg M, Rupprecht R, Milenkovic VM, Wetzel CH.
      Mitochondrial malfunction is supposed to be involved in the etiology and pathology of major depressive disorder (MDD). Here, we aimed to identify and characterize the molecular pathomechanisms related to mitochondrial dysfunction in adult human skin fibroblasts, which were derived from MDD patients or non-depressive control subjects. We found that MDD fibroblasts showed significantly impaired mitochondrial functioning: basal and maximal respiration, spare respiratory capacity, non-mitochondrial respiration and adenosine triphosphate (ATP)-related oxygen consumption was lower. Moreover, MDD fibroblasts harbor lower ATP levels and showed hyperpolarized mitochondrial membrane potential. To investigate cellular resilience, we challenged both groups of fibroblasts with hormonal (dexamethasone) or metabolic (galactose) stress for one week, and found that both stressors increased oxygen consumption but lowered ATP content in MDD as well as in non-depressive control fibroblasts. Interestingly, the bioenergetic differences between fibroblasts from MDD or non-depressed subjects, which were observed under non-treated conditions, could not be detected after stress. Our findings support the hypothesis that altered mitochondrial function causes a bioenergetic imbalance, which is associated with the molecular pathophysiology of MDD. The observed alterations in the oxidative phosphorylation system (OXPHOS) and other mitochondria-related properties represent a basis for further investigations of pathophysiological mechanisms and might open new ways to gain insight into antidepressant signaling pathways.
    Keywords:  adenosine triphosphate; bioenergetics; calcium imaging; major depression; mitochondria; mitochondrial DNA copy number; mitochondrial membrane potential; oxidative phosphorylation; skin fibroblasts
    DOI:  https://doi.org/10.3390/cells9040884
  7. Sci Rep. 2020 Apr 08. 10(1): 6095
    Zhao H, Lewellen BM, Wilson RJ, Cui D, Drake JC, Zhang M, Yan Z.
      The common clinical symptoms of Friedreich's ataxia (FRDA) include ataxia, muscle weakness, type 2 diabetes and heart failure, which are caused by impaired mitochondrial function due to the loss of frataxin (FXN) expression. Endurance exercise is the most powerful intervention for promoting mitochondrial function; however, its impact on FRDA has not been studied. Here we found that mice with genetic knockout and knock-in of the Fxn gene (KIKO mice) developed exercise intolerance, glucose intolerance and moderate cardiac dysfunction at 6 months of age. These abnormalities were associated with impaired mitochondrial respiratory function concurrent with reduced iron regulatory protein 1 (Irp1) expression as well as increased oxidative stress, which were not due to loss of mitochondrial content and antioxidant enzyme expression. Importantly, long-term (4 months) voluntary running in KIKO mice starting at a young age (2 months) completely prevented the functional abnormalities along with restored Irp1 expression, improved mitochondrial function and reduced oxidative stress in skeletal muscle without restoring Fxn expression. We conclude that endurance exercise training prevents symptomatic onset of FRDA in mice associated with improved mitochondrial function and reduced oxidative stress. These preclinical findings may pave the way for clinical studies of the impact of endurance exercise in FRDA patients.
    DOI:  https://doi.org/10.1038/s41598-020-62952-6
  8. Biochim Biophys Acta Bioenerg. 2020 Apr 07. pii: S0005-2728(20)30052-9. [Epub ahead of print] 148202
    Sánchez-Caballero L, Elurbe DM, Baertling F, Guerrero-Castillo S, van den Brand M, van Strien J, van Dam TJP, Rodenburg R, Brandt U, Huynen MA, Nijtmans LGJ.
      Protein complexes from the oxidative phosphorylation (OXPHOS) system are assembled with the help of proteins called assembly factors. We here delineate the function of the inner mitochondrial membrane protein TMEM70, in which mutations have been linked to OXPHOS deficiencies, using a combination of BioID, complexome profiling and coevolution analyses. TMEM70 interacts with complex I and V and for both complexes the loss of TMEM70 results in the accumulation of an assembly intermediate followed by a reduction of the next assembly intermediate in the pathway. This indicates that TMEM70 has a role in the stability of membrane-bound subassemblies or in the membrane recruitment of subunits into the forming complex. Independent evidence for a role of TMEM70 in OXPHOS assembly comes from evolutionary analyses. The TMEM70/TMEM186/TMEM223 protein family, of which we show that TMEM186 and TMEM223 are mitochondrial in human as well, only occurs in species with OXPHOS complexes. Our results validate the use of combining complexome profiling with BioID and evolutionary analyses in elucidating congenital defects in protein complex assembly.
    Keywords:  Assembly factor; Complex I; Complex V; Oxidative phosphorylation
    DOI:  https://doi.org/10.1016/j.bbabio.2020.148202
  9. J Biol Chem. 2020 Apr 10. pii: jbc.RA119.011110. [Epub ahead of print]
    Miyamoto S, Zhang G, Hall D, Oates PJ, Maity S, Madesh M, Han X, Sharma K.
      Exposure to chronic hyperglycemia due to diabetes mellitus can lead to the development and progression of diabetic kidney disease (DKD). We have recently reported that reduced superoxide production is associated with mitochondrial dysfunction in the kidneys of mouse models of type 1 DKD. We also demonstrated that humans with DKD have significantly reduced levels of mitochondrion-derived metabolites in their urine. Here, we examined renal superoxide production in a type 2 diabetes animal model, the db/db mouse, and the role of a mitochondrial-protectant, MTP-131 (also called elamipretide, SS-31, or Bendavia) in restoring renal superoxide production and ameliorating DKD. We found that18-week-old db/db mice have reduced renal and cardiac superoxide levels, as measured by dihydroethidium oxidation, and increased levels of albuminuria, mesangial matrix accumulation, and urinary hydrogen peroxide (H2O2). Administration of MTP-131 significantly inhibited the increases in albuminuria, urinary H2O2, and mesangial matrix accumulation in the db/db mice, and fully preserved levels of renal superoxide production in these mice. MTP-131 also reduced total renal lysocardiolipin (lysoCL) and major lysoCL subspecies, and preserved lysocardiolipin acyltransferase 1 (LCLAT1) expression in the db/db mice. These results indicate that in type 2 diabetes, DKD is associated with reduced renal and cardiac superoxide levels and that MTP-131 protects against DKD and preserves physiological superoxide levels possibly by regulating cardiolipin remodeling.
    Keywords:  MTP-131; cardiac metabolism; cardiolipin; diabetic nephropathy; elamipretide; kidney; mitochondria; reactive oxygen species (ROS); shotgun lipidomics; superoxide ion
    DOI:  https://doi.org/10.1074/jbc.RA119.011110
  10. Hum Mol Genet. 2020 Apr 10. pii: ddaa063. [Epub ahead of print]
    Yang TC, Yarmishyn AA, Yang YP, Lu PC, Chou SJ, Wang ML, Lin TC, Hwang DK, Chou YB, Chen SJ, Yu WK, Wang AG, Hsu CC, Chiou SH.
      The mutations in the genes encoding the subunits of complex I of the mitochondrial electron transport chain are the most common cause of Leber's hereditary optic neuropathy (LHON), a maternal hereditary disease characterized by retinal ganglion cell (RGC) degeneration. The characteristics of incomplete penetrance indicate that nuclear genetic and environmental factors also determine phenotypic expression of LHON. Therefore, the role of mutant mitochondrial NADH dehydrogenase subunit proteins and nuclear genetic factors/environmental effects in the aetiology of LHON needs further understanding. In this study, we generated human induced pluripotent stem cells (hiPSCs) from healthy control, unaffected LHON mutation carrier, and affected LHON patient. hiPSC-derived RGCs were used to study the differences between affected and unaffected carriers of mitochondrial DNA point mutation m.11778G > A in the MT-ND4 gene. We found that both mutated cell lines were characterized by increase in ROS production, however, only affected cell line had increased levels of apoptotic cells. We found a significant increase in retrograde mitochondria and a decrease in stationary mitochondria in the affected RGC axons. In addition, the mRNA and protein levels of KIF5A in the LHON-affected RGCs were significantly reduced. Antioxidant NAC could restore the expression of KIF5A and the normal pattern of mitochondrial movement in the affected RGCs. To conclude, we found essential differences in the mutually dependent processes of oxidative stress, mitochondrial transport and apoptosis between two LHON-specific mutation carrier RGC cell lines, asymptomatic carrier and disease-affected, and identified KIF5A as a central modulator of these differences.
    DOI:  https://doi.org/10.1093/hmg/ddaa063
  11. Int J Mol Sci. 2020 Apr 08. pii: E2589. [Epub ahead of print]21(7):
    Knottnerus SJG, Mengarelli I, Wüst RCI, Baartscheer A, Bleeker JC, Coronel R, Ferdinandusse S, Guan K, IJlst L, Li W, Luo X, Portero VM, Ulbricht Y, Visser G, Wanders RJA, Wijburg FA, Verkerk AO, Houtkooper RH, Bezzina CR.
      Patients with very long-chain acyl-CoA dehydrogenase deficiency (VLCADD) can present with life-threatening cardiac arrhythmias. The pathophysiological mechanism is unknown. We reprogrammed fibroblasts from one mildly and one severely affected VLCADD patient, into human induced pluripotent stem cells (hiPSCs) and differentiated these into cardiomyocytes (VLCADD-CMs). VLCADD-CMs displayed shorter action potentials (APs), more delayed afterdepolarizations (DADs) and higher systolic and diastolic intracellular Ca2+ concentration ([Ca2+]i) than control CMs. The mitochondrial booster resveratrol mitigated the biochemical, electrophysiological and [Ca2+]i changes in the mild but not in the severe VLCADD-CMs. Accumulation of potentially toxic intermediates of fatty acid oxidation was blocked by substrate reduction with etomoxir. Incubation with etomoxir led to marked prolongation of AP duration and reduced DADs and [Ca2+]i in both VLCADD-CMs. These results provide compelling evidence that reduced accumulation of fatty acid oxidation intermediates, either by enhanced fatty acid oxidation flux through increased mitochondria biogenesis (resveratrol) or by inhibition of fatty acid transport into the mitochondria (etomoxir), rescues pro-arrhythmia defects in VLCADD-CMs and open doors for new treatments.
    Keywords:  VLCADD; acylcarnitines; arrhythmias; hiPSC
    DOI:  https://doi.org/10.3390/ijms21072589
  12. Mech Ageing Dev. 2020 Apr 06. pii: S0047-6374(20)30033-6. [Epub ahead of print] 111238
    Alica SM, Katja M, Daniel S, Barbara H, Jörg B.
      Caloric reduction (CR) is considered as the most reasonable intervention to delay aging and age-related diseases. Numerous studies in various model organisms provide the main basis for this hypothesis. Human studies exist, but they differ widely in study design, characteristics of test persons and study outcome. In this study we investigated CR in humans on a molecular level to gain a better understanding in these processes. For that purpose, we analyzed human peripheral blood mononuclear cells of healthy people fasting according to F. X. Mayr. In a previous study our group could show a significantly improved DNA repair capacity after fasting. Here we were able to confirm these findings despite a slightly modified fasting therapy. Furthermore, the function of the mitochondrial respiratory chain and the mRNA levels of the mitochondria-associated genes SIRT3 and NDUFS1 were significantly affected by CR. However, these changes were only detectable in people who exhibited no improvement in DNA repair capacity. In contrast to that we could not observe any changes in ROS levels, mitochondrial DNA copy number and non-mitochondrial respiration. Altogether our results reveal that CR in form of F. X. Mayr therapy is able to positively influence several cellular parameters and especially mitochondrial function. Mitochondria Human peripheral blood mononuclear cells.
    Keywords:  Caloric reduction; DNA repair capacity; Respiratory chain
    DOI:  https://doi.org/10.1016/j.mad.2020.111238
  13. Aging Dis. 2020 Apr;11(2): 269-285
    Sun Y, Lu F, Yu X, Wang B, Chen J, Lu F, Peng S, Sun X, Yu M, Chen H, Wang Y, Zhang L, Liu N, Du H, Zhao D, Zhang W.
      Hydrogen sulfide (H2S), an important gasotransmitter, regulates cardiovascular functions. Mitochondrial damage induced by the overproduction of reactive oxygen species (ROS) results in myocardial injury with a diabetic state. The purpose of this study was to investigate the effects of exogenous H2S on mitophagy formation in diabetic cardiomyopathy. In this study, we found that exogenous H2S could improve cardiac functions, reduce mitochondrial fragments and ROS levels, enhance mitochondrial respiration chain activities and inhibit mitochondrial apoptosis in the hearts of db/db mice. Our results showed that exogenous H2S facilitated parkin translocation into mitochondria and promoted mitophagy formation in the hearts of db/db mice. Our studies further revealed that the ubiquitination level of cytosolic parkin was increased and the expression of USP8, a deubiquitinating enzyme, was decreased in db/db cardiac tissues. S-sulfhydration is a novel posttranslational modification of specific cysteine residues on target proteins by H2S. Our results showed that the S-sulfhydration level of USP8 was obviously decreased in vivo and in vitro under hyperglycemia and hyperlipidemia, however, exogenous H2S could reverse this effect and promote USP8/parkin interaction. Dithiothreitol, a reducing agent that reverses sulfhydration-mediated covalent modification, increased the ubiquitylation level of parkin, abolished the effects of exogenous H2S on USP8 deubiquitylation and suppressed the interaction of USP8 with parkin in neonatal rat cardiomyocytes treated with high glucose, oleate and palmitate. Our findings suggested that H2S promoted mitophagy formation by increasing S-sulfhydration of USP8, which enhanced deubiquitination of parkin through the recruitment of parkin in mitochondria.
    Keywords:  Parkin; USP8; diabetic cardiomyopathy; hydrogen sulfide; mitophagy
    DOI:  https://doi.org/10.14336/AD.2019.0524
  14. Glycoconj J. 2020 Apr 08.
    Fazzari M, Audano M, Lunghi G, Di Biase E, Loberto N, Mauri L, Mitro N, Sonnino S, Chiricozzi E.
      The crucial role of ganglioside GM1 in the regulation of neural homeostasis has been assessed by several studies. Recently we shed new light on the molecular basis underlying GM1 effects demonstrating that GM1 oligosaccharide directly binds TrkA receptor and triggers MAPK pathway activation leading to neuronal differentiation and protection. Following its exogenous administration, proteomic analysis revealed an increased expression of proteins involved in several biochemical mechanisms, including mitochondrial bioenergetics. Based on these data, we investigated the possible effect of GM1 oligosaccharide administration on mitochondrial function. We show that wild-type Neuro2a cells exposed to GM1 oligosaccharide displayed an increased mitochondrial density and an enhanced mitochondrial activity together with reduced reactive oxygen species levels. Interestingly, using a Neuro2a model of mitochondrial dysfunction, we found an increased mitochondrial oxygen consumption rate as well as increased complex I and II activities upon GM1 oligosaccharide administration. Taken together, our data identify GM1 oligosaccharide as a mitochondrial regulator that by acting at the plasma membrane level triggers biochemical signaling pathway inducing mitochondriogenesis and increasing mitochondrial activity. Although further studies are necessary, the capability to enhance the function of impaired mitochondria points to the therapeutic potential of the GM1 oligosaccharide for the treatment of pathologies where these organelles are compromised, including Parkinson's disease.
    Keywords:  GM1 ganglioside; GM1 oligosaccharide; Mitochondria dysfunctions; Mitochondrial biogenesis; Mitochondrial function; Plasma membrane signaling
    DOI:  https://doi.org/10.1007/s10719-020-09920-4
  15. Integr Cancer Ther. 2020 Jan-Dec;19:19 1534735420911437
    Sharma P, Singh S.
      Dichloroacetate (DCA) is a metabolic modulator that inhibits pyruvate dehydrogenase activity and promotes the influx of pyruvate into the tricarboxylic acid cycle for complete oxidation of glucose. DCA stimulates oxidative phosphorylation (OXPHOS) more than glycolysis by altering the morphology of the mitochondria and supports mitochondrial apoptosis. As a consequence, DCA induces apoptosis in cancer cells and inhibits the proliferation of cancer cells. Recently, the role of miRNAs has been reported in regulating gene expression at the transcriptional level and also in reprogramming energy metabolism. In this article, we indicate that DCA treatment leads to the upregulation of let-7a expression, but DCA-induced cancer cell death is independent of let-7a. We observed that the combined effect of DCA and let-7a induces apoptosis, reduces reactive oxygen species generation and autophagy, and stimulates mitochondrial biogenesis. This was later accompanied by stimulation of OXPHOS in combined treatment and was thus involved in metabolic reprogramming of MDA-MB-231 cells.
    Keywords:  DCA; OXPHOS; breast cancer; metabolic reprogramming; miRNA; mitogenesis
    DOI:  https://doi.org/10.1177/1534735420911437
  16. J Gerontol A Biol Sci Med Sci. 2020 Apr 08. pii: glaa078. [Epub ahead of print]
    Turnell BR, Reinhardt K.
      Oxidative damage increases with age in a variety of cell types, including sperm, which are particularly susceptible to attack by reactive oxygen species (ROS). While mitochondrial respiration is the main source of cellular ROS, the relationship between the rates of aerobic metabolism and ROS production, and how this relationship may be affected by age, both in sperm and in other cell types, is unclear. Here, we investigate in Drosophila melanogaster sperm the effects of male age on i) the level of hydrogen peroxide in the mitochondria, using a transgenic H2O2 reporter line; ii) the in-situ rate of non-H2O2 ROS production, using a novel biophysical method; and iii) metabolic rate, using fluorescent lifetime imaging microscopy. Sperm from older males had higher mitochondrial ROS levels and a higher metabolic rate but produced ROS at a lower rate. In comparison, a somatic tissue, the gut epithelium, also showed an age-related increase in mitochondrial ROS levels but a decrease in metabolic rate. These results support the idea of a tissue-specific optimal rate of aerobic respiration balancing the production and removal of ROS, with aging causing a shift away from this optimum and leading to increased ROS accumulation. Our findings also support the view that pathways of germline and somatic ageing can be uncoupled, which may have implications for male infertility treatments.
    Keywords:  autofluorescence; reactive oxygen species; redox balance; roGFP; sperm metabolism
    DOI:  https://doi.org/10.1093/gerona/glaa078
  17. Front Cell Dev Biol. 2020 ;8 200
    Chen G, Kroemer G, Kepp O.
      Mitochondrial dysfunction constitutes one of the hallmarks of aging and is characterized by irregular mitochondrial morphology, insufficient ATP production, accumulation of mitochondrial DNA (mtDNA) mutations, increased production of mitochondrial reactive oxygen species (ROS) and the consequent oxidative damage to nucleic acids, proteins and lipids. Mitophagy, a mitochondrial quality control mechanism enabling the degradation of damaged and superfluous mitochondria, prevents such detrimental effects and reinstates cellular homeostasis in response to stress. To date, there is increasing evidence that mitophagy is significantly impaired in several human pathologies including aging and age-related diseases such as neurodegenerative disorders, cardiovascular pathologies and cancer. Therapeutic interventions aiming at the induction of mitophagy may have the potency to ameliorate these dysfunctions. In this review, we summarize recent findings on mechanisms controlling mitophagy and its role in aging and the development of human pathologies.
    Keywords:  ROS; aging; caloric restriction; mitochondria; mitophagy
    DOI:  https://doi.org/10.3389/fcell.2020.00200
  18. Nat Commun. 2020 Apr 08. 11(1): 1740
    Wei W, Pagnamenta AT, Gleadall N, Sanchis-Juan A, Stephens J, Broxholme J, Tuna S, Odhams CA, , , Fratter C, Turro E, Caulfield MJ, Taylor JC, Rahman S, Chinnery PF.
      Several strands of evidence question the dogma that human mitochondrial DNA (mtDNA) is inherited exclusively down the maternal line, most recently in three families where several individuals harbored a 'heteroplasmic haplotype' consistent with biparental transmission. Here we report a similar genetic signature in 7 of 11,035 trios, with allelic fractions of 5-25%, implying biparental inheritance of mtDNA in 0.06% of offspring. However, analysing the nuclear whole genome sequence, we observe likely large rare or unique nuclear-mitochondrial DNA segments (mega-NUMTs) transmitted from the father in all 7 families. Independently detecting mega-NUMTs in 0.13% of fathers, we see autosomal transmission of the haplotype. Finally, we show the haplotype allele fraction can be explained by complex concatenated mtDNA-derived sequences rearranged within the nuclear genome. We conclude that rare cryptic mega-NUMTs can resemble paternally mtDNA heteroplasmy, but find no evidence of paternal transmission of mtDNA in humans.
    DOI:  https://doi.org/10.1038/s41467-020-15336-3
  19. Cell Commun Signal. 2020 Apr 07. 18(1): 58
    Zhang Q, Cheng G, Pan J, Zielonka J, Xiong D, Myers CR, Feng L, Shin SS, Kim YH, Bui D, Hu M, Bennett B, Schmainda K, Wang Y, Kalyanaraman B, You M.
      BACKGROUND: Magnolia extract (ME) is known to inhibit cancer growth and metastasis in several cell types in vitro and in animal models. However, there is no detailed study on the preventive efficacy of ME for oral cancer, and the key components in ME and their exact mechanisms of action are not clear. The overall goal of this study is to characterize ME preclinically as a potent oral cancer chemopreventive agent and to determine the key components and their molecular mechanism(s) that underlie its chemopreventive efficacy.METHODS: The antitumor efficacy of ME in oral cancer was investigated in a 4-nitroquinoline-1-oxide (4NQO)-induced mouse model and in two oral cancer orthotopic models. The effects of ME on mitochondrial electron transport chain activity and ROS production in mouse oral tumors was also investigated.
    RESULTS: ME did not cause detectable side effects indicating that it is a promising and safe chemopreventive agent for oral cancer. Three major key active compounds in ME (honokiol, magnolol and 4-O-methylhonokiol) contribute to its chemopreventive effects. ME inhibits mitochondrial respiration at complex I of the electron transport chain, oxidizes peroxiredoxins, activates AMPK, and inhibits STAT3 phosphorylation, resulting in inhibition of the growth and proliferation of oral cancer cells.
    CONCLUSION: Our data using highly relevant preclinical oral cancer models, which share histopathological features seen in human oral carcinogenesis, suggest a novel signaling and regulatory role for mitochondria-generated superoxide and hydrogen peroxide in suppressing oral cancer cell proliferation, progression, and metastasis. Video abstract.
    Keywords:  AMPK; Complex I; Oral cancer; STAT3
    DOI:  https://doi.org/10.1186/s12964-020-0524-2
  20. Biosci Rep. 2020 Apr 07. pii: BSR20194404. [Epub ahead of print]
    Chen YH, Wu YJ, Chen WC, Lee TS, Tsou TC, Chang HC, Lo SW, Chen SL.
      Di (2-ethylhexyl) phthalate (DEHP) is a plasticizer frequently leached out from polyvinyl chloride (PVC) products and is quickly metabolized to its monoester equivalent mono(2-ethylhexyl) phthalate (MEHP) once enters organisms.  Exposure to DEHP/MEHP through food chain intake has been shown to modified metabolism but its effect on the development of metabolic myopathy of skeletal muscle (SKM) has not been revealed so far.  Here we found that MEHP repressed myogenic terminal differentiation of proliferating myoblasts (PMB) and confluent myoblasts (CMB) but had weak effect on this process once it had been initiated.  The transition of mitochondrion (MITO) morphology from high efficient filamentary network to low efficient vesicles was triggered by MEHP, implying its negative effects on MITO functions.  The impaired MITO functions was further demonstrated by reduced MITO DNA (mtDNA) level and SDH enzyme activity as well as highly increased reactive oxygen species (ROS) in cells after MEHP treatment.  The expression of metabolic genes, including PDK4, CPT1b, UCP2, and HO1, was highly increased by MEHP and the promoters of PDK4 and CPT1b were also activated by MEHP.  Additionally, the stability of some subunits in the oxidative phosphorylation system (OXPHOS) complexes was found to be reduced by MEHP, implying defective oxidative metabolism in MITO and which was confirmed by repressed palmitic acid oxidation in MEHP treated cells. Besides, MEHP also blocked insulin induced glucose uptake.  Taken together, our results suggest that MEHP is inhibitory to myogenesis and is harmful to MITO functions in SKM so its exposure should be avoided or limited.
    Keywords:  dehp; fatty acid oxidation; mehp; mitochondria; muscle; oxidative phosphorylation
    DOI:  https://doi.org/10.1042/BSR20194404
  21. Sci Transl Med. 2020 Apr 08. pii: eaaz8264. [Epub ahead of print]12(538):
    Mirali S, Botham A, Voisin V, Xu C, St-Germain J, Sharon D, Hoff FW, Qiu Y, Hurren R, Gronda M, Jitkova Y, Nachmias B, MacLean N, Wang X, Arruda A, Minden MD, Horton TM, Kornblau SM, Chan SM, Bader GD, Raught B, Schimmer AD.
      Neurolysin (NLN) is a zinc metallopeptidase whose mitochondrial function is unclear. We found that NLN was overexpressed in almost half of patients with acute myeloid leukemia (AML), and inhibition of NLN was selectively cytotoxic to AML cells and stem cells while sparing normal hematopoietic cells. Mechanistically, NLN interacted with the mitochondrial respiratory chain. Genetic and chemical inhibition of NLN impaired oxidative metabolism and disrupted the formation of respiratory chain supercomplexes (RCS). Furthermore, NLN interacted with the known RCS regulator, LETM1, and inhibition of NLN disrupted LETM1 complex formation. RCS were increased in patients with AML and positively correlated with NLN expression. These findings demonstrate that inhibiting RCS formation selectively targets AML cells and stem cells and highlights the therapeutic potential of pharmacologically targeting NLN in AML.
    DOI:  https://doi.org/10.1126/scitranslmed.aaz8264
  22. Cell Stress Chaperones. 2020 Apr 06.
    Tezgin D, Giardina C, Perdrizet GA, Hightower LE.
      We present new data on the effects of HBOT on human kidney (HK-2) cell metabolism using a SeaHorse XF Analyzer to evaluate separately the state of mitochondrial and glycolytic energy metabolism. The data are discussed in the context of the concept of cellular caloristasis networks. The information on the changes in cellular energy metabolism stimulated by HBOT presented here provides new insights into the cellular energy state and mitochondrial environment in which sHSPs function. These data will be useful in forming testable hypotheses about the functions of translocated sHSPs in human mitochondria responding to stressors.
    Keywords:  Caloristasis; Cytoprotection; Diabetic kidney disease; HBOT; Hyperbaric oxygen therapy; Mitochondria; Oxidative stress; Proteostasis; Small HSPs
    DOI:  https://doi.org/10.1007/s12192-020-01100-5
  23. Dev Cell. 2020 Apr 07. pii: S1534-5807(20)30192-1. [Epub ahead of print]
    De Henau S, Pagès-Gallego M, Pannekoek WJ, Dansen TB.
      Symmetry breaking is an essential step in cell differentiation and early embryonic development. However, the molecular cues that trigger symmetry breaking remain largely unknown. Here, we show that mitochondrial H2O2 acts as a symmetry-breaking cue in the C. elegans zygote. We find that symmetry breaking is marked by a local H2O2 increase and coincides with a relocation of mitochondria to the cell cortex. Lowering endogenous H2O2 levels delays the onset of symmetry breaking, while artificially targeting mitochondria to the cellular cortex using a light-induced heterodimerization technique is sufficient to initiate symmetry breaking in a H2O2-dependent manner. In wild-type development, both sperm and maternal mitochondria contribute to symmetry breaking. Our findings reveal that mitochondrial H2O2-signaling promotes the onset of polarization, a fundamental process in development and cell differentiation, and this is achieved by both mitochondrial redistribution and differential H2O2-production.
    Keywords:  C. elegans; H(2)O(2); cell polarity; embryonic development; mitochondria; redox signaling; symmetry breaking
    DOI:  https://doi.org/10.1016/j.devcel.2020.03.008
  24. J Biol Chem. 2020 Apr 07. pii: jbc.RA119.012213. [Epub ahead of print]
    Yamaguchi I, Yoshimura SH, Katoh H.
      The the cystine/glutamate transporter system xc- consists of the light-chain subunit xCT (SLC7A11) and the heavy-chain subunit CD98 (4F2hc or SLC3A2) and exchanges extracellular cystine for intracellular glutamate at the plasma membrane. The imported cystine is reduced to cysteine and used for synthesis of glutathione, which is one of the most important antioxidants in cancer cells. Because cancer cells have increased levels of reactive oxygen species (ROS), xCT, being responsible for cystine-glutamate exchange, is overexpressed in many cancers, including glioblastoma. However, under glucose-limited conditions, xCT overexpression induces ROS accumulation and cell death. Here, we report that cell survival under glucose deprivation depends on cell density. We found that a high cell density (HD) down-regulates xCT levels and increases cell viability under glucose deprivation. We also found that growth of glioblastoma cells at HD inactivates mTOR, and that treatment with the mTOR inhibitor Torin 1 of cells grown at low density (LD) down-regulates xCT and inhibits glucose deprivation-induced cell death. The lysosome inhibitor bafilomycin A1 (BafA1) suppressed xCT down-regulation in HD-cultured glioblastoma cells and in Torin 1-treated cells grown at LD. Additionally, BafA1 exposure or ectopic xCT expression restored glucose deprivation-induced cell death at HD. These results suggest that HD inactivates mTOR and promotes lysosomal degradation of xCT, leading to improved glioblastoma cell viability under glucose-limited conditions. Our findings provide evidence that the control of xCT protein expression via lysosomal degradation is an important mechanism for metabolic adaptation in glioblastoma cells.
    Keywords:  amino acid transport; cancer biology; cell biology; cell death; lysosome
    DOI:  https://doi.org/10.1074/jbc.RA119.012213
  25. J Proteomics. 2020 Apr 02. pii: S1874-3919(20)30133-0. [Epub ahead of print] 103765
    Martín-Cano Fe FE, Gaitskell-Phillips G, Ortiz-Rodríguez JM, Silva-Rodríguez A, Román Á, Rojo-Domínguez P, Alonso-Rodríguez E, Tapia JA, Gil MC, Ortega-Ferrusola C, Peña FJ.
      Proteomic technologies allow the detection of thousands of proteins at the same time, being a powerful technique to reveal molecular regulatory mechanisms in spermatozoa and also sperm damage linked to low fertility or specific biotechnologies. Modifications induced by the cryopreservation in the stallion sperm proteome were studied using HPLC/MS/MS. Ejaculates from fertile stallions were collected and split in two subsamples, one was investigated as fresh (control) samples, and the other aliquot frozen and thawed using standard procedures and investigated as frozen thawed subsamples. UHPLC/MS/MS was used to study the sperm proteome under these two distinct conditions and bioinformatic enrichment analysis conducted. Gene Ontology (GO) and pathway enrichment analysis were performed revealing dramatic changes as consequence of cryopreservation. The terms oxidative phosphorylation, mitochondrial ATP synthesis coupled electron transport and electron transport chain were significantly enriched in fresh samples (P = 5.50 × 10-12, 4.26 × 10-8 and 7.26 × 10-8, respectively), while were not significantly enriched in frozen thawed samples (P = 1). The GO terms oxidation reduction process and oxidoreductase activity were enriched in fresh samples and the enrichment was reduced in frozen thawed samples (1.40 × 10-8, 1.69 × 10-6 versus 1.13 × 10-2 and 2-86 × 10-2 respectively). Reactome pathways (using human orthologs) significantly enriched in fresh sperm were TCA cycle and respiratory electron transport (P = 1.867 × 10-8), Respiratory electron transport ATP synthesis by chemiosmosis coupling (P = 2.124 × 10-5), Citric acid cycle (TCA cycle)(P = 8.395 × 10-4) Pyruvate metabolism and TCA cycle (P = 3.380 × 10-3), Respiratory electron transport (P = 2.764 × 10-2) and Beta oxidation of laurolyl-CoA to decanoyl CoA-CoA (P = 1.854 × 10-2) none of these pathways were enriched in thawed samples (P = 1). We have provided the first detailed study on how the cryopreservation process impacts the stallion sperm proteome. Our findings identify the metabolic proteome and redoxome as the two key groups of proteins affected by the procedure. SIGNIFICANCE: In the present manuscript we investigated how the cryopreservation of stallion spermatozoa impacts the proteome of these cells. This procedure is routinely used in horse breeding and has a major impact in the industry, facilitating the trade of genetic material. This is still a suboptimal biotechnology, with numerous unresolved problems. The limited knowledge of the molecular insults occurring during cryopreservation is behind these problems. The application and development of proteomics to the spermatozoa, allow to obtain valuable information of the specific mechanisms affected by the procedure. In this paper, we report that cryopreservation impacts numerous proteins involved in metabolism regulation (mainly mitochondrial proteins involved in the TCA cycle, and oxidative phosphorylation) and also affect proteins with oxidoreductase activity. Moreover, specific proteins involved in the sperm- oocyte interaction are also affected by the procedure. The information gathered in this study, opens interesting questions and offer new lines of research for the improvement of the technology focusing the targets here identified, and the specific steps in the procedure (cooling, toxicity of antioxidants etc..) to be modified to reduce the damage.
    Keywords:  Cryopreservation; Horses; Proteomics; Spermatozoa
    DOI:  https://doi.org/10.1016/j.jprot.2020.103765
  26. Mol Biol Rep. 2020 Apr 04.
    Karanam G, Arumugam MK.
      Cyclic dipeptides are increasingly gaining importance as considering its significant biological and pharmacological activities. This study was aimed to investigate the anticancer activity of a dipeptide Cyclo(-Pro-Tyr) (DP) identified from marine sponge Callyspongia fistularis symbiont Bacillus pumilus AMK1 and the underlying apoptotic mechanisms in the liver cancer HepG2 cell lines. MTT assay was done to demonstrate the cytotoxic effect of DP in HepG2 cells and mouse Fibroblast McCoy cells. Initially, apoptosis inducing activity of DP was identified using propidium iodide (PI) and acridine orange/ethidium bromide (AO/EB) dual staining, then it was confirmed by DNA fragmentation assay and western blotting analysis of apoptosis related markers Bax, Bcl-2, cytochrome c, caspase-3 and cleaved poly (ADP-ribose) polymerase (PARP). Rhodamine 123 staining was performed to observe DP effects on the mitochondrial membrane potential (MMP) and DCFH-DA (Dichloro-dihydro-fluorescein diacetate) staining was done to measure the intracellular reactive oxygen species (ROS) levels. The MTT results revealed that DP initiated dose-dependent cytotoxicity in HepG2 cells, but no significant toxicity in mouse Fibroblast McCoy cells treated with DP at the specified concentrations. DP induced apoptosis, which is confirmed by the appearance of apoptotic bodies with PI and AO/EB dual staining, and DNA fragmentation. DP significantly elevated the Bax/Bcl-2 ratio, disrupted the mitochondrial membrane potential (MMP), enhanced cytochrome c release from mitochondria, increased caspase-3 activation, the cleavage of PARP and increased intracellular reactive oxygen species (ROS) levels. Besides this, DP successfully inhibited the phosphorylation of PI3K, AKT and increased PTEN expression. These results suggested DP might have anti-cancer effect by initiating apoptosis through mitochondrial dysfunction and downregulating PI3K/Akt signaling pathway in HepG2 cells with no toxicity effect on normal fibroblast cells. Therefore, DP may be developed as a potential alternative therapeutic agent for treating hepatocellular carcinoma.
    Keywords:  Apoptosis; Bax/Bcl2 ratio; Cyclo(-pro-tyr); MMP; PI3K/AKT; ROS
    DOI:  https://doi.org/10.1007/s11033-020-05407-5
  27. Saudi J Gastroenterol. 2020 Apr 06.
    Zhao Y, Zhang X, Liu Y, Ma Y, Kong P, Bai T, Han M, Li B.
      Background/Aims: Colorectal cancer (CRC) is the third most common malignant tumour worldwide and the second leading cause of cancer-related deaths. Commonly, 5'-aminolevulinic acid synthase1 (ALAS1) is the rate-limiting enzyme for haem biosynthesis. Recent studies have shown that ALAS1 is involved in a number of cellular functions and has significant effects on non-small cell lung cancer (NSCLC). However, current concepts of disease pathogenesis fail to fully explain the role of ALAS1 expression and biological functions in CRC.Materials and Methods: A total of 67 paired tumour tissues and adjacent colorectal tissues were used to detect ALAS1 levels and further analyse the correlation between ALAS1 expression levels and clinical features. Using HCT116 cell lines, we studied the impact of ALAS1 on biological function by knocking down or inhibiting ALAS1.
    Results: We found an increase in the levels of ALAS1 in cancer tissues compared to adjacent colorectal tissues. The increase in ALAS1 expression was closely related to the invasion depth, N staging and tumour size of CRC patients. The proliferation and metastasis of CRC cells could be inhibited by suppressing ALAS1.
    Conclusions: The abnormal expression of ALAS1 is closely related to the proliferation and metastasis of CRC cells, suggesting that ALAS1 may be a novel therapeutic target for the treatment of CRC.
    Keywords:  Cell proliferation; colorectal neoplasms; neoplasm metastasis
    DOI:  https://doi.org/10.4103/sjg.SJG_477_19
  28. Antioxid Redox Signal. 2020 Apr 07.
    Su X, Wang W, Fang C, Ni C, Zhou J, Wang X, Zhang L, Xu X, Cao R, Lang H, Wang F.
      AIMS: High-fat diet (HFD)-induced insulin resistance (IR) impairs skeletal muscle mitochondrial biogenesis and functions, adversely affecting human health and lifespan. Vitamin K2 (VK2) has a beneficial role in improving insulin sensitivity and glucose metabolism. However, the underlying molecular mechanisms of VK2 on insulin sensitivity have not been well established. We investigated VK2's modulation of mitochondrial function to protect against IR in mice and cell models.RESULTS: VK2 supplementation could effectively ameliorate the development of IR by improving mitochondrial function in both HFD-fed mice and PA-exposed cells. We revealed for the first time that HFD-caused mitochondrial dysfunction could be reversed by VK2 treatment. VK2 enhanced the mitochondrial function by improving mitochondrial respiratory capacity, increasing mitochondrial biogenesis and the enzymatic activities of mitochondrial complexes through SIRT1 signaling. The benefits of VK2 were abrogated in C2C12 transfected with SIRT1 siRNA but not in C2C12 transfected with AMPK siRNA. VK2 and SRT1720, a specific agonist of SIRT1, had the same effect on improving mitochondrial function via SIRT1 signaling. Thus, SIRT1 is required for VK2 improvement skeletal muscle. Furthermore, the beneficial effects of VK2 and GGOH both contribute to inhibited IR in skeletal muscle via SIRT1.
    INNOVATION: These studies demonstrated a previously undiscovered mechanism by which VK2 alleviates IR in skeletal muscle by improving mitochondrial function via SIRT1.
    CONCLUSION: Naturally occurring VK2 prevents IR by improving mitochondrial function through SIRT1 signaling. These results could provide a foundation to identify new VK2-based preventive and therapeutic strategies for IR.
    DOI:  https://doi.org/10.1089/ars.2019.7908
  29. Biomed Res Int. 2020 ;2020 1068671
    Peng J, Jing X, Wu J, Hong D, Hu X, Wang Q, Hu H, Cai X.
      The effect of metformin on human esophageal normal and carcinoma cells remains poorly understood. We aim to investigate the different antiproliferation effects and underlying distinct molecular mechanisms between these two types of cells. Human esophageal squamous cell carcinoma cell line, EC109, and normal esophageal epithelial cell line, HEEC, were used in the experiment. The cell survival rate was determined by cell counting kit-8 (CCK-8). Cell apoptosis was analyzed by flow cytometry. The mRNA and protein levels of signal transducer and activator of transcription 3 (Stat3) were detected by real-time quantitative PCR and western blot. Interleukin-6 (IL-6) was added to activate Stat3. The level of intracellular reactive oxygen species (ROS) was assessed by a DCFH-DA fluorescent probe. Metformin had more significant inhibitory effects on cell proliferation in EC109 cells than HEECs. Metformin induced apoptosis of EC109 cells in a dose-dependent manner instead of HEECs. The expression of Stat3 in both mRNA and protein levels was higher in EC109 cells than HEECs. Further study revealed that metformin may attenuate the phosphorylation of the Stat3 and the Bcl-2 expression, which was restored by IL-6 partly in EC109 cells but not HEECs. On the contrary, metformin increased the level of ROS in both the cell lines, but this intracellular ROS variation had no effect on apoptosis. Metformin has different functional roles on the apoptosis in esophageal carcinoma cells and normal esophageal cells. Therefore, the Stat3/Bcl-2 pathway-mediated apoptosis underlies the cell-type-specific drug sensitivity, suggesting metformin possesses a therapeutic activity and selectivity on esophageal cancer.
    DOI:  https://doi.org/10.1155/2020/1068671
  30. J Alzheimers Dis. 2020 Apr 02.
    Quintana DD, Garcia JA, Anantula Y, Rellick SL, Engler-Chiurazzi EB, Sarkar SN, Brown CM, Simpkins JW.
      Cerebrovascular pathology is pervasive in Alzheimer's disease (AD), yet it is unknown whether cerebrovascular dysfunction contributes to the progression or etiology of AD. In human subjects and in animal models of AD, cerebral hypoperfusion and hypometabolism are reported to manifest during the early stages of the disease and persist for its duration. Amyloid-β is known to cause cellular injury in both neurons and endothelial cells by inducing the production of reactive oxygen species and disrupting intracellular Ca2+ homeostasis. We present a mechanism for mitochondrial degeneration caused by the production of mitochondrial superoxide, which is driven by increased mitochondrial Ca2+ uptake. We found that persistent superoxide production injures mitochondria and disrupts electron transport in cerebrovascular endothelial cells. These observations provide a mechanism for the mitochondrial deficits that contribute to cerebrovascular dysfunction in patients with AD.
    Keywords:  Amyloid-β; calcium; mitochondria; superoxide; vascular endothelial cells
    DOI:  https://doi.org/10.3233/JAD-190964
  31. Sci Rep. 2020 Apr 09. 10(1): 6115
    Lee HM, Sia APE, Li L, Sathasivam HP, Chan MSA, Rajadurai P, Tsang CM, Tsao SW, Murray PG, Tao Q, Paterson IC, Yap LF.
      Nasopharyngeal carcinoma (NPC) is a highly metastatic cancer that is consistently associated with Epstein-Barr virus (EBV) infection. In this study, we identify for the first time a role for monoamine oxidase A (MAOA) in NPC. MAOA is a mitochondrial enzyme that catalyzes oxidative deamination of neurotransmitters and dietary amines. Depending on the cancer type, MAOA can either have a tumour-promoting or tumour-suppressive role. We show that MAOA is down-regulated in primary NPC tissues and its down-regulation enhances the migration of NPC cells. In addition, we found that EBV infection can down-regulate MAOA expression in both pre-malignant and malignant nasopharyngeal epithelial (NPE) cells. We further demonstrate that MAOA is down-regulated as a result of IL-6/IL-6R/STAT3 signalling and epigenetic mechanisms, effects that might be attributed to EBV infection in NPE cells. Taken together, our data point to a central role for EBV in mediating the tumour suppressive effects of MAOA and that loss of MAOA could be an important step in the pathogenesis of NPC.
    DOI:  https://doi.org/10.1038/s41598-020-63150-0
  32. Exp Gerontol. 2020 Apr 01. pii: S0531-5565(20)30172-8. [Epub ahead of print] 110940
    Palee S, Jaiwongkam T, Kerdphoo S, Pratchayasakul W, Chattipakorn SC, Chattipakorn N.
      Obesity and menopause are known as a major risk factor in the development of left ventricular (LV) dysfunction. Calorie restriction (CR) or exercise (Ex) improved metabolic status and LV function. This study aims to investigate the combined effects of Ex and CR on the cardiometabolic status, and cardiac calcium ([Ca2+]i) regulation in estrogen-deprivation, obese prediabetic rats. Female rats were fed with either a high-fat diet (HFD) or a normal diet for 13 weeks. The HFD rats were ovariectomized (HFO), and subjected to 1) vehicle (HFOV); 2) calorie restriction (HFOCR); 3) exercise (HFOEx); 4) combined therapy (HFOCB); or 5) estrogen (HFOE2). After six weeks of interventions the cardiometabolic status, cardiac [Ca2+]i transients, mitochondrial function and dynamics were determined. HFD-fed rats developed insulin resistance as indicated by increased plasma insulin and HOMA index. Although rats in the HFOV groups had markedly reduced %LVFS which indicated impaired LV function, impaired [Ca2+]i homeostasis, cardiac mitochondrial function and their dynamics, all interventions attenuated these impairments. Interestingly, HFOCB rats were observed to have the greatest cardiometabolic improvement. The combination of calorie restriction and exercise exerted greater efficacy in attenuating LV dysfunction through an improved metabolic status, cardiac function, mitochondrial function, and cardiac [Ca2+]i homeostasis than Ex or CR monotherapy in ovariectomized obese prediabetic rats.
    Keywords:  Calorie restriction; Exercise; Heart; Mitochondria; Obesity; Prediabetic
    DOI:  https://doi.org/10.1016/j.exger.2020.110940
  33. Protein Cell. 2020 Apr 10.
    Li E, Li X, Huang J, Xu C, Liang Q, Ren K, Bai A, Lu C, Qian R, Sun N.
      Dysregulation of circadian rhythms associates with cardiovascular disorders. It is known that deletion of the core circadian gene Bmal1 in mice causes dilated cardiomyopathy. However, the biological rhythm regulation system in mouse is very different from that of humans. Whether BMAL1 plays a role in regulating human heart function remains unclear. Here we generated a BMAL1 knockout human embryonic stem cell (hESC) model and further derived human BMAL1 deficient cardiomyocytes. We show that BMAL1 deficient hESC-derived cardiomyocytes exhibited typical phenotypes of dilated cardiomyopathy including attenuated contractility, calcium dysregulation, and disorganized myofilaments. In addition, mitochondrial fission and mitophagy were suppressed in BMAL1 deficient hESC-cardiomyocytes, which resulted in significantly attenuated mitochondrial oxidative phosphorylation and compromised cardiomyocyte function. We also found that BMAL1 binds to the E-box element in the promoter region of BNIP3 gene and specifically controls BNIP3 protein expression. BMAL1 knockout directly reduced BNIP3 protein level, causing compromised mitophagy and mitochondria dysfunction and thereby leading to compromised cardiomyocyte function. Our data indicated that the core circadian gene BMAL1 is critical for normal mitochondria activities and cardiac function. Circadian rhythm disruption may directly link to compromised heart function and dilated cardiomyopathy in humans.
    Keywords:  cardiomyocytes; cell differentiation; circadian gene BMAL1; dilated cardiomyopathy; human embryonic stem cells; mitochondria
    DOI:  https://doi.org/10.1007/s13238-020-00713-x
  34. J Am Heart Assoc. 2020 Apr 07. 9(7): e014366
    Ponce JM, Coen G, Spitler KM, Dragisic N, Martins I, Hinton A, Mungai M, Tadinada SM, Zhang H, Oudit GY, Song LS, Li N, Sicinski P, Strack S, Abel ED, Mitchell C, Hall DD, Grueter CE.
      Background Nuclear-to-mitochondrial communication regulating gene expression and mitochondrial function is a critical process following cardiac ischemic injury. In this study, we determined that cyclin C, a component of the Mediator complex, regulates cardiac and mitochondrial function in part by modifying mitochondrial fission. We tested the hypothesis that cyclin C functions as a transcriptional cofactor in the nucleus and a signaling molecule stimulating mitochondrial fission in response to stimuli such as cardiac ischemia. Methods and Results We utilized gain- and loss-of-function mouse models in which the CCNC (cyclin C) gene was constitutively expressed (transgenic, CycC cTg) or deleted (knockout, CycC cKO) in cardiomyocytes. The knockout and transgenic mice exhibited decreased cardiac function and altered mitochondria morphology. The hearts of knockout mice had enlarged mitochondria with increased length and area, whereas mitochondria from the hearts of transgenic mice were significantly smaller, demonstrating a role for cyclin C in regulating mitochondrial dynamics in vivo. Hearts from knockout mice displayed altered gene transcription and metabolic function, suggesting that cyclin C is essential for maintaining normal cardiac function. In vitro and in vivo studies revealed that cyclin C translocates to the cytoplasm, enhancing mitochondria fission following stress. We demonstrated that cyclin C interacts with Cdk1 (cyclin-dependent kinase 1) in vivo following ischemia/reperfusion injury and that, consequently, pretreatment with a Cdk1 inhibitor results in reduced mitochondrial fission. This finding suggests a potential therapeutic target to regulate mitochondrial dynamics in response to stress. Conclusions Our study revealed that cyclin C acts as a nuclear-to-mitochondrial signaling factor that regulates both cardiac hypertrophic gene expression and mitochondrial fission. This finding provides new insights into the regulation of cardiac energy metabolism following acute ischemic injury.
    Keywords:  ischemia; mitochondria; signal transduction; transcriptional coactivator; transgenic mice
    DOI:  https://doi.org/10.1161/JAHA.119.014366
  35. Front Immunol. 2020 ;11 521
    Salminen TS, Vale PF.
      Understanding why the response to infection varies between individuals remains one of the major challenges in immunology and infection biology. A substantial proportion of this heterogeneity can be explained by individual genetic differences which result in variable immune responses, and there are many examples of polymorphisms in nuclear-encoded genes that alter immunocompetence. However, how immunity is affected by genetic polymorphism in an additional genome, inherited maternally inside mitochondria (mtDNA), has been relatively understudied. Mitochondria are increasingly recognized as important mediators of innate immune responses, not only because they are the main source of energy required for costly immune responses, but also because by-products of mitochondrial metabolism, such as reactive oxygen species (ROS), may have direct microbicidal action. Yet, it is currently unclear how naturally occurring variation in mtDNA contributes to heterogeneity in infection outcomes. In this review article, we describe potential sources of variation in mitochondrial function that may arise due to mutations in vital nuclear and mitochondrial components of energy production or due to a disruption in mito-nuclear crosstalk. We then highlight how these changes in mitochondrial function can impact immune responses, focusing on their effects on ATP- and ROS-generating pathways, as well as immune signaling. Finally, we outline how being a powerful and genetically tractable model of infection, immunity and mitochondrial genetics makes the fruit fly Drosophila melanogaster ideally suited to dissect mitochondrial effects on innate immune responses to infection.
    Keywords:  Drosophila melanogaster; cybrid; infection; innate immunity; mitochondria; mtDNA; oxidative phosphorylation; reactive oxygen species
    DOI:  https://doi.org/10.3389/fimmu.2020.00521
  36. iScience. 2020 Mar 25. pii: S2589-0042(20)30186-3. [Epub ahead of print]23(4): 101002
    Li W, Wang J.
      Cancer metabolism is critical for understanding the mechanism of tumorigenesis, yet the understanding is still challenging. We studied gene-metabolism regulatory interactions and quantified the global driving forces for cancer-metabolism dynamics as the underlying landscape and probability flux. We uncovered four steady-state attractors: a normal state attractor, a cancer OXPHOS state attractor, a cancer glycolysis state attractor, and an intermediate cancer state attractor. We identified the key regulatory interactions through global sensitivity analysis based on the landscape topography. Different landscape topographies of glycolysis switch between normal cells and cancer cells were identified. We uncovered that the normal state to cancer state transformation is associated with the peaks of the probability flux and the thermodynamic dissipation, giving dynamical and thermodynamic origin of cancer formation. We found that cancer metabolism oscillations consume more energy to support cancer malignancy. This study provides a quantitative understanding of cancer metabolism and suggests a metabolic therapeutic strategy.
    Keywords:  Cancer; Gene Network; Mathematical Biosciences; Metabolic Flux Analysis
    DOI:  https://doi.org/10.1016/j.isci.2020.101002
  37. PLoS One. 2020 ;15(4): e0231173
    Piel S, Chamkha I, Dehlin AK, Ehinger JK, Sjövall F, Elmér E, Hansson MJ.
      Acetaminophen is one of the most common over-the-counter pain medications used worldwide and is considered safe at therapeutic dose. However, intentional and unintentional overdose accounts for up to 70% of acute liver failure cases in the western world. Extensive research has demonstrated that the induction of oxidative stress and mitochondrial dysfunction are central to the development of acetaminophen-induced liver injury. Despite the insight gained on the mechanism of acetaminophen toxicity, there still is only one clinically approved pharmacological treatment option, N-acetylcysteine. N-acetylcysteine increases the cell's antioxidant defense and protects liver cells from further acetaminophen-induced oxidative damage. Because it primarily protects healthy liver cells rather than rescuing the already injured cells alternative treatment strategies that target the latter cell population are warranted. In this study, we investigated mitochondria as therapeutic target for the development of novel treatment strategies for acetaminophen-induced liver injury. Characterization of the mitochondrial toxicity due to acute acetaminophen overdose in vitro in human cells using detailed respirometric analysis revealed that complex I-linked (NADH-dependent) but not complex II-linked (succinate-dependent) mitochondrial respiration is inhibited by acetaminophen. Treatment with a novel cell-permeable succinate prodrug rescues acetaminophen-induced impaired mitochondrial respiration. This suggests cell-permeable succinate prodrugs as a potential alternative treatment strategy to counteract acetaminophen-induced liver injury.
    DOI:  https://doi.org/10.1371/journal.pone.0231173
  38. Cell Metab. 2020 Apr 07. pii: S1550-4131(20)30131-5. [Epub ahead of print]31(4): 662-663
    Chen YR, Jarosz DF.
      Covalent cysteine modification by reactive oxygen species (ROS) has been implicated in regulating diverse biological processes, yet global understanding of this modification has remained fragmentary. Developing new approaches for detecting cysteine modification, Xiao et al. (2020) recently charted a comprehensive map of cysteine oxidation across tissues and life stages.
    DOI:  https://doi.org/10.1016/j.cmet.2020.03.016
  39. Cell Metab. 2020 Apr 06. pii: S1550-4131(20)30132-7. [Epub ahead of print]
    Al Batran R, Gopal K, Capozzi ME, Chahade JJ, Saleme B, Tabatabaei-Dakhili SA, Greenwell AA, Niu J, Almutairi M, Byrne NJ, Masson G, Kim R, Eaton F, Mulvihill EE, Garneau L, Masters AR, Desta Z, Velázquez-Martínez CA, Aguer C, Crawford PA, Sutendra G, Campbell JE, Dyck JRB, Ussher JR.
      Perturbations in carbohydrate, lipid, and protein metabolism contribute to obesity-induced type 2 diabetes (T2D), though whether alterations in ketone body metabolism influence T2D pathology is unknown. We report here that activity of the rate-limiting enzyme for ketone body oxidation, succinyl-CoA:3-ketoacid-CoA transferase (SCOT/Oxct1), is increased in muscles of obese mice. We also found that the diphenylbutylpiperidine pimozide, which is approved to suppress tics in individuals with Tourette syndrome, is a SCOT antagonist. Pimozide treatment reversed obesity-induced hyperglycemia in mice, which was phenocopied in mice with muscle-specific Oxct1/SCOTdeficiency. These actions were dependent on pyruvate dehydrogenase (PDH/Pdha1) activity, the rate-limitingenzyme of glucose oxidation, as pimozide failed to alleviate hyperglycemia in obese mice with a muscle-specific Pdha1/PDH deficiency. This work defines a fundamental contribution of enhanced ketone body oxidation to the pathology of obesity-induced T2D, while suggesting pharmacological SCOT inhibition as a new class of anti-diabetes therapy.
    Keywords:  glycemia; insulin resistance; ketone bodies; ketone body oxidation; obesity; pimozide; type 2 diabetes
    DOI:  https://doi.org/10.1016/j.cmet.2020.03.017
  40. Nat Commun. 2020 Apr 09. 11(1): 1755
    Deng L, Yao P, Li L, Ji F, Zhao S, Xu C, Lan X, Jiang P.
      Asparagine synthetase (ASNS) catalyses the ATP-dependent conversion of aspartate to asparagine. However, both the regulation and biological functions of asparagine in tumour cells remain largely unknown. Here, we report that p53 suppresses asparagine synthesis through the transcriptional downregulation of ASNS expression and disrupts asparagine-aspartate homeostasis, leading to lymphoma and colon tumour growth inhibition in vivo and in vitro. Moreover, the removal of asparagine from culture medium or the inhibition of ASNS impairs cell proliferation and induces p53/p21-dependent senescence and cell cycle arrest. Mechanistically, asparagine and aspartate regulate AMPK-mediated p53 activation by physically binding to LKB1 and oppositely modulating LKB1 activity. Thus, we found that p53 regulates asparagine metabolism and dictates cell survival by generating an auto-amplification loop via asparagine-aspartate-mediated LKB1-AMPK signalling. Our findings highlight a role for LKB1 in sensing asparagine and aspartate and connect asparagine metabolism to the cellular signalling transduction network that modulates cell survival.
    DOI:  https://doi.org/10.1038/s41467-020-15573-6
  41. Metabolites. 2020 Apr 02. pii: E139. [Epub ahead of print]10(4):
    McCann MR, McHugh CE, Kirby M, Jennaro TS, Jones AE, Stringer KA, Puskarich MA.
      BACKGROUND: Sepsis-induced alterations in mitochondrial function contribute to organ dysfunction and mortality. Measuring mitochondrial function in vital organs is neither feasible nor practical, highlighting the need for non-invasive approaches. Mitochondrial function may be reflected in the concentrations of metabolites found in platelets and whole blood (WB) samples. We proposed to use these as alternates to indirectly estimate platelet mitochondrial oxygen consumption rate (mOCR) in sepsis patients.METHODS: We determined the relationships between platelet mOCR and metabolites in both platelets and WB, as measured by quantitative 1H-NMR metabolomics. The associations were identified by building multiple linear regression models with stepwise forward-backward variable selection. We considered the models to be significant with an ANOVA test (p-value ≤ 0.05) and a positive predicted-R2.
    RESULTS: The differences in adjusted-R2 and ANOVA p-values (platelet adj-R2: 0.836 (0.0003), 0.711 (0.0004) vs. WB adj-R2: 0.428 (0.0079)) from the significant models indicate the platelet models were more associated with platelet mOCR.
    CONCLUSIONS: Our data suggest there are groups of metabolites in WB (leucine, acetylcarnitine) and platelets (creatine, ADP, glucose, taurine) that are associated with platelet mOCR. Thus, WB and platelet metabolites could be used to estimate platelet mOCR.
    Keywords:  acetylcarnitine; bioenergetics; metabolism; mitochondria; mitochondrial function; mitochondrial respiration; nuclear magnetic resonance spectroscopy
    DOI:  https://doi.org/10.3390/metabo10040139
  42. Cell Metab. 2020 Apr 07. pii: S1550-4131(20)30126-1. [Epub ahead of print]31(4): 669-678
    Sung AY, Floyd BJ, Pagliarini DJ.
      Defining functions for the full complement of proteins is a grand challenge in the post-genomic era and is essential for our understanding of basic biology and disease pathogenesis. In recent times, this endeavor has benefitted from a combination of modern large-scale and classical reductionist approaches-a process we refer to as "systems biochemistry"-that helps surmount traditional barriers to the characterization of poorly understood proteins. This strategy is proving to be particularly effective for mitochondria, whose well-defined proteome has enabled comprehensive analyses of the full mitochondrial system that can position understudied proteins for fruitful mechanistic investigations. Recent systems biochemistry approaches have accelerated the identification of new disease-related mitochondrial proteins and of long-sought "missing" proteins that fulfill key functions. Collectively, these studies are moving us toward a more complete understanding of mitochondrial activities and providing a molecular framework for the investigation of mitochondrial pathogenesis.
    Keywords:  mitochondria; multi-omics; orphan proteins; rare disease; systems biochemistry
    DOI:  https://doi.org/10.1016/j.cmet.2020.03.011
  43. Cell Metab. 2020 Apr 07. pii: S1550-4131(20)30127-3. [Epub ahead of print]31(4): 660-662
    Maynard AG, Kanarek N.
      In this issue of Cell Metabolism, Yang et al., 2020 report that serine is a source of mitochondrial NADH derived from one-carbon metabolism. Serine becomes a major source of NADH when cellular respiration is inhibited, and the un-utilized, accumulated NADH inhibits the TCA cycle and slows proliferation.
    DOI:  https://doi.org/10.1016/j.cmet.2020.03.012
  44. Cell Metab. 2020 Apr 07. pii: S1550-4131(20)30116-9. [Epub ahead of print]31(4): 666-668
    Lundø K, Trauelsen M, Pedersen SF, Schwartz TW.
      Lactate accumulation in tumors-a hallmark of the Warburg effect-has recently been shown to regulate cancer cell metabolism and survival through autocrine activation of GPR81. Now, Brown et al. (2020) demonstrate that lactate surprisingly also controls immune evasion through paracrine activation of GPR81 on stromal dendritic cells.
    DOI:  https://doi.org/10.1016/j.cmet.2020.03.001
  45. J Thorac Dis. 2020 Mar;12(3): 830-838
    He HH, Fu JH, Hao ZX, Wu HF, Zhong Q, Wang F, Liu HH, Gu XS, Wang B, Huang HD, Li ZY, He JX.
      Background: Diabetes mellitus is a recognized risk factor for esophageal squamous cell carcinomas (ESCC), and metformin is a recognized protective factor for some gastrointestinal tumors. But knowledge is limited regarding the effect of metformin on survival outcome of ESCC patients with type 2 diabetes mellitus (T2DM). We assessed the impact of post-diagnosis metformin use on overall survival (OS) and disease-free survival (DFS) in ESCC with T2DM undergoing surgical resection.Methods: A retrospective analysis was performed on 3,523 patients with ESCC who met the study conditions after surgical resection. Log-rank and Cox regression models were used to evaluate the relationship between metformin and T2DM and ESCC survival rate, and adjusted according to age, gender, BMI, smoking, drinking and staging, et al.
    Results: Among included ESCC patients, 619 were associated with type 2 diabetes, while the remaining 2,904 were not associated with type 2 diabetes. The 5-year OS (28.43%) of patients with T2DM was significantly lower than that of patients without T2DM (32.75%), P=0.037. DFS in 5 years were 27.30% (with T2DM) and 31.75% (without T2DM) (P=0.030), respectively. Compared with patients without T2DM, patients with T2DM presented worse OS [adjusted risk ratio (HRadj) =1.19] and DFS (HRadj =1.17; P<0.001). Among the 619 patients with type 2 diabetes, 485 were treated with metformin and 134 were not treated with metformin. Patients treated with metformin had significantly improved OS [adjusted risk ratio (HRadj) =0.89; P=0.031) and DFS (HRadj =0.90; P=0.013).
    Conclusions: T2DM was again associated with poorer survival in ESCC patients, and metformin may improve the prognosis of these patients.
    Keywords:  Esophageal squamous cell carcinomas (ESCC); disease-free survival (DFS); metformin; overall survival (OS); type 2 diabetes mellitus (T2DM)
    DOI:  https://doi.org/10.21037/jtd.2019.12.98
  46. J Biol Chem. 2020 Apr 09. pii: jbc.RA119.012094. [Epub ahead of print]
    Mitchell W, Ng EA, Tamucci JD, Boyd KJ, Sathappa M, Coscia A, Pan M, Han X, Eddy NA, May ER, Szeto HH, Alder NN.
      Mitochondrial dysfunction underlies many heritable diseases, acquired pathologies, and aging-related declines in health. Szeto-Schiller (SS) peptides comprise a class of amphipathic tetrapeptides that are efficacious towards a wide array of mitochondrial disorders and are believed to target mitochondrial membranes because they are enriched in the anionic phospholipid cardiolipin (CL). However, little is known regarding how SS peptides interact with or alter the physical properties of lipid bilayers. In this study, using biophysical and computational approaches, we have analyzed the interactions of the lead compound SS-31 (elamipretide) with model and mitochondrial membranes. Our results show that this polybasic peptide partitions into the membrane interfacial region with an affinity and a lipid binding density that are directly related to surface charge. We found that SS-31 binding does not destabilize lamellar bilayers even at the highest binding concentrations; however, it did cause saturable alterations in lipid packing. Most notably, SS-31 modulated the surface electrostatics of both model and mitochondrial membranes. We propose nonexclusive mechanisms by which the tuning of surface charge could underpin the mitoprotective properties of SS-31, including alteration of the distribution of ions and basic proteins at the interface, and/or modulation of bilayer physical properties. As a proof of concept, we show that SS-31 alters divalent cation (calcium) distribution within the interfacial region and reduces the energetic burden of calcium stress in mitochondria. The mechanistic details of SS-31 revealed in this study will help inform the development of future compound variants with enhanced efficacy and bioavailability.
    Keywords:  SS-31; cardiolipin; drug action; elamipretide; electrostatics; lipid structure; membrane biophysics; mitochondria; peptide therapeutic; peptides
    DOI:  https://doi.org/10.1074/jbc.RA119.012094
  47. Am J Physiol Endocrinol Metab. 2020 Apr 07.
    Groennebaek T, Nielsen J, Jespersen NR, Botker HE, de Paoli FV, Miller BF, Vissing K.
      The measurement of mitochondrial content is essential for bioenergetic research as it provides a tool to evaluate if changes in mitochondrial function are strictly due to changes in content or other mechanisms that influence function. In this perspective, we argue that commonly used biomarkers of mitochondrial content may possess limited utility for capturing changes in content with physiological intervention. Moreover, we argue that they may not provide reliable estimates of content in certain pathological situations. Finally, we discuss potential solutions to overcome issues related to the utilization of biomarkers of mitochondrial content. Shedding light on this important issue will hopefully aid conclusions about the mitochondrial structure-function relationship.
    Keywords:  Biomarkers; bioenergetics; chronic disease; citrate synthase activity; exercise
    DOI:  https://doi.org/10.1152/ajpendo.00101.2020
  48. Sci Rep. 2020 Apr 10. 10(1): 6225
    Sun P, Bai T, Ma T, Ding J.
      Human NAD-dependent isocitrate dehydrogenase (NAD-IDH) is responsible for the catalytic conversion of isocitrate into α-ketoglutarate in the Krebs cycle. This enzyme exists as the α2βγ heterotetramer composed of the αβ and αγ heterodimers. Our previous biochemical data showed that the αγ heterodimer and the holoenzyme can be activated by low concentrations of ATP but inhibited by high concentrations of ATP; however, the molecular mechanism was unknown. Here, we report the crystal structures of the αγ heterodimer with ATP binding only to the allosteric site (αMgγMg+CIT+ATP) and to both the allosteric site and the active site (αMg+ATPγMg+CIT+ATP). Structural data show that ATP at low concentrations can mimic ADP to bind to the allosteric site, which stabilizes CIT binding and leads the enzyme to adopt an active conformation, revealing why the enzyme can be activated by low concentrations of ATP. On the other hand, at high concentrations ATP is competitive with NAD for binding to the catalytic site. In addition, our biochemical data show that high concentrations of ATP promote the formation of metal ion-ATP chelates. This reduces the concentration of free metal ion available for the catalytic reaction, and thus further inhibits the enzymatic activity. The combination of these two effects accounts for the inhibition of the enzyme at high concentrations of ATP. Taken together, our structural and biochemical data reveal the molecular mechanism for the dual regulatory roles of ATP on the αγ heterodimer of human NAD-IDH.
    DOI:  https://doi.org/10.1038/s41598-020-63425-6