bims-camemi Biomed News
on Mitochondrial metabolism in cancer
Issue of 2019‒06‒30
27 papers selected by
Christian Frezza
University of Cambridge, MRC Cancer Unit


  1. J Biol Chem. 2019 Jun 24. pii: jbc.RA119.008680. [Epub ahead of print]
    Wang Y, Palmfeldt J, Gregersen N, Makhov AM, Conway JF, Wang M, McCalley SP, Basu S, Alharbi H, St Croix C, Calderon MJ, Watkins S, Vockley J.
      Three mitochondrial metabolic pathways are required for efficient energy production in eukaryotic cells - the electron transfer chain (ETC), fatty acid β-oxidation (FAO), and the tricarboxylic acid (TCA) cycle. The ETC is organized into inner mitochondrial membrane supercomplexes that promote substrate channeling and catalytic efficiency. Although previous studies have suggested functional interaction between FAO and the ETC, their physical interaction has never been demonstrated. In this study, using blue native gel and two-dimensional electrophoreses, nano LC-MS/MS, immunogold EM, and stimulated emission depletion microscopy, we show that FAO enzymes physically interact with ETC supercomplexes at two points. We found that the FAO trifunctional protein (TFP) interacts with the NADH-binding domain of complex I of the ETC, whereas the electron transfer enzyme flavoprotein dehydrogenase interacts with ETC complex III. Moreover, the FAO enzyme very-long-chain acyl-CoA dehydrogenase (VLCAD) physically interacted with TFP, thereby creating a multifunctional energy protein complex. These findings provide a first view of an integrated molecular architecture for the major energy-generating pathways in mitochondria that ensures the safe transfer of unstable reducing equivalents from FAO to the ETC. They also offer insight into clinical ramifications for individuals with genetic defects in these pathways.
    Keywords:  electron microscopy (EM); fatty acid oxidation; mitochondrial metabolism; mitochondrial respiratory chain complex; primary metabolism; protein structure; proteomics; stimulated emission depletion microscopy (STED); supercomplex; trifunctional protein (TFP)
    DOI:  https://doi.org/10.1074/jbc.RA119.008680
  2. Nature. 2019 Jun 26.
    Wang X, Liu R, Zhu W, Chu H, Yu H, Wei P, Wu X, Zhu H, Gao H, Liang J, Li G, Yang W.
      Cancer metastasis is the primary cause of morbidity and mortality, and accounts for up to 95% of cancer-related deaths1. Cancer cells often reprogram their metabolism to efficiently support cell proliferation and survival2,3. However, whether and how those metabolic alterations contribute to the migration of tumour cells remain largely unknown. UDP-glucose 6-dehydrogenase (UGDH) is a key enzyme in the uronic acid pathway, and converts UDP-glucose to UDP-glucuronic acid4. Here we show that, after activation of EGFR, UGDH is phosphorylated at tyrosine 473 in human lung cancer cells. Phosphorylated UGDH interacts with Hu antigen R (HuR) and converts UDP-glucose to UDP-glucuronic acid, which attenuates the UDP-glucose-mediated inhibition of the association of HuR with SNAI1 mRNA and therefore enhances the stability of SNAI1 mRNA. Increased production of SNAIL initiates the epithelial-mesenchymal transition, thus promoting the migration of tumour cells and lung cancer metastasis. In addition, phosphorylation of UGDH at tyrosine 473 correlates with metastatic recurrence and poor prognosis of patients with lung cancer. Our findings reveal a tumour-suppressive role of UDP-glucose in lung cancer metastasis and uncover a mechanism by which UGDH promotes tumour metastasis by increasing the stability of SNAI1 mRNA.
    DOI:  https://doi.org/10.1038/s41586-019-1340-y
  3. Redox Biol. 2019 Jun 15. pii: S2213-2317(19)30522-1. [Epub ahead of print]26 101253
    Zhang Y, Bharathi SS, Beck ME, Goetzman ES.
      Fatty acid oxidation (FAO)-driven H2O2 has been shown to be a major source of oxidative stress in several tissues and disease states. Here, we established that the mitochondrial flavoprotein long-chain acyl-CoA dehydrogenase (LCAD), which catalyzes a key step in mitochondrial FAO, directly produces H2O2in vitro by leaking electrons to oxygen. Kinetic analysis of recombinant human LCAD showed that it produces H2O2 15-fold faster than the related mitochondrial enzyme very long-chain acyl-CoA dehydrogenase (VLCAD), but 50-fold slower than a bona fide peroxisomal acyl-CoA oxidase. The rate of H2O2 formation by human LCAD is slow compared to its activity as a dehydrogenase (about 1%). However, expression of hLCAD in HepG2 cells is sufficient to significantly increase H2O2 in the presence of fatty acids. Liver mitochondria from LCAD-/- mice, but not VLCAD-/- mice, produce significantly less H2O2 during incubation with fatty acids. Finally, we observe highest LCAD expression in human liver, followed by kidney, lung, and pancreas. Based on our data, we propose that the presence of LCAD drives H2O2 formation in response to fatty acids in these tissues.
    Keywords:  Hydrogen peroxide/mitochondria/fatty acid oxidation/acyl-CoA dehydrogenase
    DOI:  https://doi.org/10.1016/j.redox.2019.101253
  4. J Biol Chem. 2019 Jun 26. pii: jbc.RA119.008488. [Epub ahead of print]
    Shum M, Houde VP, Bellemare V, Moreira RJ, Bellmann K, St-Pierre P, Viollet B, Foretz M, Marette A.
      mTOR complex 1 (mTORC1) and p70 S6 kinase (S6K1) are both involved in the development of obesity-linked insulin resistance. Recently, we showed that the S6K1 inhibitor PF-4708671 (PF) increases insulin sensitivity. However, we also reported that PF can increase glucose metabolism even in the absence of insulin in muscle and hepatic cells. Here, we further explored the potential mechanisms by which PF increases glucose metabolism in muscle and liver cells independently of insulin. Time-course experiments revealed that PF induces AMP-activated protein kinase (AMPK) activation before inhibiting S6K1. However, PF-induced glucose uptake was not prevented in primary muscle cells from AMPK α1/2 double-KO (dKO) mice. Moreover, PF-mediated suppression of hepatic glucose production was maintained in hepatocytes derived from the AMPK α1/2-dKO mice. Remarkably, PF could still reduce glucose production and activate AMPK in hepatocytes from S6K1/2-dKO mice. Mechanistically, bioenergetics experiments revealed that PF reduces mitochondrial complex I activity in both muscle and hepatic cells. The stimulatory effect of PF on glucose uptake was partially reduced by expression of the Saccharomyces cerevisiae NADH: ubiquinone oxidoreductase in L6 cells. These results indicate that PF-mediated S6K1 inhibition is not required for its effect on insulin-independent glucose metabolism and AMPK activation. We conclude that although PF rapidly activates AMPK, its ability to acutely increase glucose uptake and suppress glucose production does not require AMPK activation. Unexpectedly, PF rapidly inhibits mitochondrial complex I activity, a mechanism that partially underlies PF's effect on glucose metabolism.
    Keywords:  AMPK-activated protein kinase (AMPK); PF-4708671; diabetes; gluconeogenesis; glucose homeostasis; glucose metabolism; mammalian target of rapamycin (mTOR); mitochondrial complex I; obesity; p70 S6 kinase (S6K1)
    DOI:  https://doi.org/10.1074/jbc.RA119.008488
  5. Redox Biol. 2019 Jun 11. pii: S2213-2317(19)30298-8. [Epub ahead of print]26 101254
    Lin Q, Li S, Jiang N, Shao X, Zhang M, Jin H, Zhang Z, Shen J, Zhou Y, Zhou W, Gu L, Lu R, Ni Z.
      Contrast-induced acute kidney injury (CI-AKI) occurs in more than 30% of patients after intravenous iodinated contrast media and causes serious complications, including renal failure and mortality. Recent research has demonstrated that routine antioxidant and alkaline therapy failed to show benefits in CI-AKI patients with high risk for renal complications. Mitophagy is a mechanism of selective autophagy, which controls mitochondrial quality and mitochondrial reactive oxygen species (ROS) through degradation of damaged mitochondria. The role of mitophagy and its regulation of apoptosis in CI-AKI are poorly understood. In this study, we demonstrated that mitophagy was induced in renal tubular epithelial cells (RTECs) during CI-AKI, both in vivo and in vitro. Meanwhile, contrast media-induced mitophagy was abolished when silencing PINK1 or PARK2 (Parkin), indicating a dominant role of the PINK1-Parkin pathway in mitophagy. Moreover, mitochondrial damage, mitochondrial ROS, RTEC apoptosis, and renal injury under contrast exposure were more severe in PINK1- or PARK2-deficient cells and mice than in wild-type groups. Functionally, PINK1-Parkin-mediated mitophagy prevented RTEC apoptosis and tissue damage in CI-AKI through reducing mitochondrial ROS and subsequent NLRP3 inflammasome activation. These results demonstrated that PINK1-Parkin-mediated mitophagy played a protective role in CI-AKI by reducing NLRP3 inflammasome activation.
    Keywords:  Acute kidney injury; Apoptosis; Contrast media; Mitochondrial ROS; Mitophagy; NLRP3 inflammasome
    DOI:  https://doi.org/10.1016/j.redox.2019.101254
  6. Cell Mol Life Sci. 2019 Jun 24.
    Moosavi B, Berry EA, Zhu XL, Yang WC, Yang GF.
      Succinate dehydrogenase (SDH) also known as complex II or succinate:quinone oxidoreductase is an enzyme involved in both oxidative phosphorylation and tricarboxylic acid cycle; the processes that generate energy. SDH is a multi-subunit enzyme which requires a series of proteins for its proper assembly at several steps. This enzyme has medical significance as there is a broad range of human diseases from cancers to neurodegeneration related to SDH malfunction. Some of these disorders have recently been linked to defective assembly factors, reinvigorating further research in this area. Apart from that this enzyme has agricultural importance as many fungicides have been/will be designed targeting specifically this enzyme in plant fungal pathogens. In addition, we speculate it might be possible to design novel fungicides specifically targeting fungal assembly factors. Considering the medical and agricultural implications of SDH, the aim of this review is an overview of the SDH assembly factors and critical analysis of controversial issues around them.
    Keywords:  Complex II; Fumarate reductase; Mitochondria; Oxidative phosphorylation; Succinate:quinone oxidoreductases; Tricarboxylic acid cycle
    DOI:  https://doi.org/10.1007/s00018-019-03200-7
  7. Dev Cell. 2019 Jun 06. pii: S1534-5807(19)30446-0. [Epub ahead of print]
    Wong YC, Peng W, Krainc D.
      Properly regulated mitochondrial networks are essential for cellular function and implicated in multiple diseases. Mitochondria undergo fission and fusion events, but the dynamics and regulation of a third event of inter-mitochondrial contact formation remain unclear. Using super-resolution imaging, we demonstrate that inter-mitochondrial contacts frequently form and play a fundamental role in mitochondrial networks by restricting mitochondrial motility. Inter-mitochondrial contact untethering events are marked and regulated by mitochondria-lysosome contacts, which are modulated by RAB7 GTP hydrolysis. Moreover, inter-mitochondrial contact formation and untethering are further regulated by Mfn1/2 and Drp1 GTP hydrolysis, respectively. Surprisingly, endoplasmic reticulum tubules are also present at inter-mitochondrial contact untethering events, in addition to mitochondrial fission and fusion events. Importantly, we find that multiple Charcot-Marie-Tooth type 2 disease-linked mutations in Mfn2 (CMT2A), RAB7 (CMT2B), and TRPV4 (CMT2C) converge on prolonged inter-mitochondrial contacts and defective mitochondrial motility, highlighting a role for inter-mitochondrial contacts in mitochondrial network regulation and disease.
    Keywords:  Charcot-Marie-Tooth type 2; Mfn2; RAB7; TRPV4; endoplasmic reticulum; inter-mitochondrial contact; lysosome; mitochondria; mitochondria-lysosome contact; super-resolution imaging
    DOI:  https://doi.org/10.1016/j.devcel.2019.05.033
  8. Autophagy. 2019 Jun 25.
    Zhou Y, Long Q, Wu H, Li W, Qi J, Wu Y, Xiang G, Tang H, Yang L, Chen K, Li L, Bao F, Li H, Wang Y, Li M, Liu X.
      Selective elimination of mitochondria by autophagy is a critical strategy for a variety of physiological processes, including development, cell-fate determination and stress response. Although several mechanisms have been identified as responsible for selective degradation of mitochondria, such as the PINK1-PRKN/PARKIN- and receptor-dependent pathways, aspects of the mechanisms and particularly the principles underlying the selection process of mitochondria remain obscure. Here, we addressed a new selection strategy in which the selective elimination of mitochondria is dependent on organellar topology. We found that populations of mitochondria undergo different topological transformations under serum starvation, either swelling or forming donut shapes. Swollen mitochondria are associated with mitochondrial membrane potential dissipation and PRKN recruitment, which promote their selective elimination, while the donut topology maintains mitochondrial membrane potential and helps mitochondria resist autophagy. Mechanistic studies show that donuts resist autophagy even after depolarization through preventing recruitment of autophagosome receptors CALCOCO2/NDP52 and OPTN even after PRKN recruitment. Our results demonstrate topology-dependent, bifurcated mitochondrial recycling under starvation, that is swollen mitochondria undergo removal by autophagy, while donut mitochondria undergo fission and fusion cycles for reintegration. This study reveals a novel morphological selection for control of mitochondrial quality and quantity under starvation.
    Keywords:  PINK1-PRKN/PARKIN; mitochondrial membrane potential; mitochondrial topology; mitophagy; starvation
    DOI:  https://doi.org/10.1080/15548627.2019.1634944
  9. J Stem Cell Res Ther. 2018 ;pii: 436. [Epub ahead of print]8(9):
    Tung B, Xia S.
      Mitochondria are vital organelles that supply ATP and other energy metabolites to meet the bioenergetics demands of the cell. In environments of stress or increased energy requirement, mitochondria are highly dynamic and can undergo biogenesis, fusion/fission, or autophagy. The transcription factor family, Kruppel-Like Factor (KLF), is necessary to carry out normal cellular processes from proliferation to differentiation. Recently, its importance in metabolic homeostasis in various tissue types has gained much attention. A handful of evidence supports KLF4's involvement in regulating mitochondrial homeostasis in both healthy and cancer cells. In this review, we aim to summarize the available literature that demonstrates KLF4's ability to modulate the mitochondrial life cycle in: Cardiac tissue, in which KLF4-knockdown subsequently leads to Heart Failure (HF), andGlioblastoma (GBM), where its expression promotes extensive mitochondrial fusion and offers mild cell protection under serum-deprivation.
    Keywords:  Glioblastoma; Heart failure; Kruppel-like factor 4; Mitochondria; Mitochondrial homeostasis; Stress
    DOI:  https://doi.org/10.4172/2157-7633.1000436
  10. Mitochondrion. 2019 Jun 19. pii: S1567-7249(19)30049-2. [Epub ahead of print]
    Whitley BN, Engelhart EA, Hoppins S.
      Mitochondrial dynamics shape the mitochondrial network and contribute to mitochondrial function and quality control. Mitochondrial fusion and division are integrated into diverse cellular functions and respond to changes in cell physiology. Imbalanced mitochondrial dynamics are associated with a range of diseases that are broadly characterized by impaired mitochondrial function and increased cell death. In various disease models, modulating mitochondrial fusion and division with either small molecules or genetic approaches has improved function. Although additional mechanistic understanding of mitochondrial fusion and division will be critical to inform further therapeutic approaches, mitochondrial dynamics represent a powerful therapeutic target in a wide range of human diseases.
    DOI:  https://doi.org/10.1016/j.mito.2019.06.002
  11. Semin Cell Dev Biol. 2019 Jun 22. pii: S1084-9521(18)30186-1. [Epub ahead of print]
    Majem B, Nadal E, Muñoz-Pinedo C.
      Lung cancer is the main cause of cancer death worldwide. Non-Small Cell Lung Carcinoma (NSCLC) is the most common subtype of lung cancer, and the prognosis of NSCLC patients in advanced stages is still very poor. Given the need for new therapies, the metabolism of NSCLC has been widely studied in the past two decades to identify vulnerabilities that could be translated into novel anti-metabolic therapeutic approaches. A number of studies have highlighted the role of glucose and mitochondrial metabolism in the development of NSCLC. The metabolic properties of lung tumors have been characterized in detail in vivo, and they include high glucose and lactate use and high heterogeneity regarding the use of nutrients and mitochondrial pathways. This heterogeneity has also been observed in patients infused with labeled nutrients. We will summarize here the knowledge about the use of amino acids, fatty acids and carbohydrates in NSCLC that could lead to new combination treatments.
    Keywords:  Non-Small Cell Lung Carcinoma; cancer metabolism; glucose; mitochondria
    DOI:  https://doi.org/10.1016/j.semcdb.2019.06.004
  12. Proteomics. 2019 Jun 24. e1800451
    Udupa S, Nguyen S, Hoang G, Nguyen T, Quinones A, Pham K, Asaka R, Nguyen K, Elgogary A, Jung JG, Xu Q, Fu J, Thomas AG, Tsukamoto T, Hanes J, Slusher BS, Cooper AJL, Le A.
      The targeting of glutamine metabolism specifically via pharmacological inhibition of glutaminase 1 (GLS1) has been translated into clinical trials as a novel therapy for several cancers. The results, though encouraging, show room for improvement in terms of tumor reduction. Here, we demonstrate that pancreatic tumors upregulate the production of glutamate from glutamine via the glutaminase II pathway upon GLS1 inhibition. Moreover, genetic suppression of glutamine transaminase K (GTK), a key enzyme of the glutaminase II pathway, leads to complete inhibition of pancreatic tumorigenesis in vivo unveiling GTK as a new metabolic target for cancer therapy. These results suggest that current trials using GLS1 inhibition as a therapeutic approach targeting glutamine metabolism in cancer should take into account the upregulation of other metabolic pathways that can lead to glutamate production, one such pathway is the glutaminase II pathway via GTK. This article is protected by copyright. All rights reserved.
    Keywords:  Glutaminase 1 Inhibition; Glutaminase II Pathway; Glutamine transaminase K
    DOI:  https://doi.org/10.1002/pmic.201800451
  13. Front Oncol. 2019 ;9 480
    Sarmiento-Salinas FL, Delgado-Magallón A, Montes-Alvarado JB, Ramírez-Ramírez D, Flores-Alonso JC, Cortés-Hernández P, Reyes-Leyva J, Herrera-Camacho I, Anaya-Ruiz M, Pelayo R, Millán-Pérez-Peña L, Maycotte P.
      Due to their crucial role in cell metabolism and homeostasis, alterations in mitochondrial biology and function have been related to the progression of diverse diseases including cancer. One of the consequences associated to mitochondrial dysfunction is the production of reactive oxygen species (ROS). ROS are known to have a controversial role during cancer initiation and progression and although several studies have tried to manipulate intracellular ROS levels using antioxidants or pro-oxidation conditions, it is not yet clear how to target oxidation for cancer therapy. In this study, we found differences in mitochondrial morphology in breast cancer cells when compared to a non-tumorigenic cell line and differences in mitochondrial function among breast cancer subtypes when exploring gene-expression data from the TCGA tumor dataset. Interestingly, we found increased ROS levels in triple negative breast cancer (TNBC) cell lines and a dependency on ROS for survival since antioxidant treatment induced cell death in TNBC cells but not in an estrogen receptor positive (ER+) cell line. Moreover, we identified the mitochondria as the main source of ROS in TNBC cell lines. Our results indicate a potential use for ROS as a target for therapy in the TNBC subtype which currently has the worst prognosis among all breast cancers and remains as the only breast cancer subtype which lacks a targeted therapy.
    Keywords:  ROS; breast cancer; mitochondria; mitochondrial ROS; mitochondrial morphology
    DOI:  https://doi.org/10.3389/fonc.2019.00480
  14. J Chromatogr B Analyt Technol Biomed Life Sci. 2019 Jun 05. pii: S1570-0232(18)31816-6. [Epub ahead of print]1124 247-255
    Kim J, Jonus HC, Zastre JA, Bartlett MG.
      Understanding the mechanisms underlying cancer cell survival is critical toward advancing drug discovery efforts in this field. Supplemental vitamins have been proposed to play a role in cancer cell metabolism because the increased supply of nutrients is thought to provide cofactors supporting the higher metabolic rate of cancer cells. Particularly, the role of thiamine (vitamin B1) in many biochemical pathways that supports cancer cell metabolism has been investigated. Consequently, the analysis of thiamine and its derivatives in a manner that reflects its dynamic response to genetic modification and pathophysiological stimuli is essential. In this work, we developed a mass spectrometry based-analytical method to track metabolites derived from stable isotope tracers for a better understanding of the metabolic fate of thiamine in cancer cells. This method used ion-pair reversed phase liquid chromatography to simultaneously quantify underivatized thiamine, thiamine monophosphate (TMP) and thiamine pyrophosphate (TPP) in cells. Hexylamine was used as an ion-pairing agent. The method was successfully validated for accuracy, precision and selectivity in accordance with U.S. FDA guidance. Furthermore, the method was then applied for the determination of thiamine and its derivatives with stable isotope labeling to explore the metabolic fate of intracellular thiamine in cancer cells. The finding shows that thiamine is rapidly converted to TPP however, the TPP does not return to thiamine. It appears that TPP may be utilized for other purposes rather than simply being an enzyme cofactor, suggesting unexplored roles for thiamine in cancer.
    Keywords:  Hexylamine; Liquid chromatography-mass spectrometry; Stable isotope-labeled thiamine; Thiamine; Thiamine kinetics; Thiamine pyrophosphate
    DOI:  https://doi.org/10.1016/j.jchromb.2019.05.037
  15. Hypertens Res. 2019 Jun 24.
    Zheng X, Chen M, Li X, Yang P, Zhao X, Ouyang Y, Yang Z, Liang M, Hou E, Tian Z.
      Fumarase insufficiencies can increase reactive oxygen species (ROS). This study will further dissect the imbalance of redox metabolism and the mechanism of ROS production using proteomic technology in fumarase knockdown HK-2 cells. The contribution of fumarase was further confirmed by supplementation of fumarate and malate in Dahl salt-sensitive rats. Proteomic analysis indicated that fumarase knockdown in HK-2 cells changed the expression or activity of NADPH oxidase (NOX), mitochondrial respiratory chain Complex I and III, ATP synthase subunits, and α-oxoglutarate dehydrogenase (OGDH). Meanwhile, the activities of key antioxidant enzymes, including glucose-6-phosphate dehydrogenase, 6-phosphogluconate dehydrogenase, glutathione reductase, glutathione peroxidase, and glutathione S-transferase, increased significantly. The apparent activation of antioxidant defense appeared insufficient as the glutathione and GSH/GSSG ratio were decreased significantly. Dahl salt-sensitive rats exhibited changes in redox metabolism similar to HK-2 cells with fumarase knockdown. Supplementation with fumarate and malate increased and decreased, respectively, blood pressure and H2O2 and malondialdehyde in salt-sensitive rats. These results indicated that insufficient fumarase activity increased ROS by regulating NOX, Complex I and III, ATPase alpha, and OGDH and the imbalance of glutathione metabolism, which may be one of the main reasons for salt-sensitive hypertension. Malate may be a potentially effective drug for the prevention and treatment of salt-sensitive hypertension.
    Keywords:  Dahl salt-sensitive rats; Fumarase; Fumarate; Reactive oxygen species
    DOI:  https://doi.org/10.1038/s41440-019-0290-y
  16. Nat Metab. 2019 Feb;1(2): 177-188
    Intlekofer AM, Finley LWS.
      In contrast to terminally differentiated cells, cancer cells and stem cells retain the ability to re-enter the cell cycle and proliferate. In order to proliferate, cells must increase the uptake and catabolism of nutrients to support anabolic cell growth. Intermediates of central metabolic pathways have emerged as key players that can influence cell differentiation 'decisions', processes relevant for both oncogenesis and normal development. Consequently, how cells rewire metabolic pathways to support proliferation may have profound consequences for cellular identity. Here, we discuss the metabolic programs that support proliferation and explore how metabolic states are intimately entwined with the cell fate decisions that characterize stem cells and cancer cells. By comparing the metabolism of pluripotent stem cells and cancer cells, we hope to illuminate common metabolic strategies as well as distinct metabolic features that may represent specialized adaptations to unique cellular demands.
    DOI:  https://doi.org/10.1038/s42255-019-0032-0
  17. Stem Cells Int. 2019 ;2019 8195614
    Lees JG, Cliff TS, Gammilonghi A, Ryall JG, Dalton S, Gardner DK, Harvey AJ.
      Metabolism has been shown to alter cell fate in human pluripotent stem cells (hPSC). However, current understanding is almost exclusively based on work performed at 20% oxygen (air), with very few studies reporting on hPSC at physiological oxygen (5%). In this study, we integrated metabolic, transcriptomic, and epigenetic data to elucidate the impact of oxygen on hPSC. Using 13C-glucose labeling, we show that 5% oxygen increased the intracellular levels of glycolytic intermediates, glycogen, and the antioxidant response in hPSC. In contrast, 20% oxygen increased metabolite flux through the TCA cycle, activity of mitochondria, and ATP production. Acetylation of H3K9 and H3K27 was elevated at 5% oxygen while H3K27 trimethylation was decreased, conforming to a more open chromatin structure. RNA-seq analysis of 5% oxygen hPSC also indicated increases in glycolysis, lysine demethylases, and glucose-derived carbon metabolism, while increased methyltransferase and cell cycle activity was indicated at 20% oxygen. Our findings show that oxygen drives metabolite flux and specifies carbon fate in hPSC and, although the mechanism remains to be elucidated, oxygen was shown to alter methyltransferase and demethylase activity and the global epigenetic landscape.
    DOI:  https://doi.org/10.1155/2019/8195614
  18. Biophys Chem. 2019 Jun 17. pii: S0301-4622(19)30188-7. [Epub ahead of print]252 106208
    Nath S.
      The central aspects of the energy economics of living cells revolve around the synthesis and utilization of molecules of adenosine triphosphate (ATP). Current descriptions of cell metabolism and its regulation in most textbooks of biochemistry assume that enzymes and transporters behave in the same way in isolation and in a cell. Calculations of the mechanistic or maximal P/O ratios in oxidative phosphorylation by mammalian cells generally consider only the supply side of the problem without linking to ATP-demand processes. The purpose of this article is to calculate the mechanistic P/O ratio by integration of the supply and demand sides of ATP reactions. The mechanistic stoichiometry calculated from an integrated approach is compared with that obtained from the standard model that considers only ATP supply. After accounting for leaks, slips, and other losses, the actual or operative P/O calculated by the integrated method is found to be in good agreement with the experimental values of the P/O ratio determined in mitochondria for both succinate and NADH-linked respiratory substrates. The thermodynamic consequences of these results and the biological implications are discussed. An integrated model of oxidative phosphorylation that goes beyond the chemiosmotic theory is presented, and a solution to the longstanding fundamental problem of respiratory control is found.
    Keywords:  Adenosine triphosphate (ATP); Coupling of ATP synthesis to oxygen consumption/respiration; Dicarboxylic acids; Energy demand management/economics in biology; Homeostasis; Metabolic engineering; Metabolic regulation and control; Nath's torsional mechanism of energy transduction and ATP synthesis; Nath's two-ion theory of biological energy coupling; Oxidative phosphorylation; P/O ratio; Succinate
    DOI:  https://doi.org/10.1016/j.bpc.2019.106208
  19. Dev Biol. 2019 Jun 21. pii: S0012-1606(19)30078-8. [Epub ahead of print]
    Lim Y, Rubio-Peña K, Sobraske PJ, Molina PA, Brookes PS, Galy V, Nehrke K.
      Paternal mitochondria are eliminated following fertilization by selective autophagy, but the mechanisms that restrict this process to sperm-derived organelles are not well understood. FUNDC1 (FUN14 domain containing1) is a mammalian mitophagy receptor expressed on the mitochondrial outer membrane that contributes to mitochondrial quality control following hypoxic stress. Like FUNDC1, the C. elegans ortholog FNDC-1 is widely expressed in somatic tissues and mediates hypoxic mitophagy. Here, we report that FNDC-1 is strongly expressed in sperm but not oocytes and contributes to paternal mitochondria elimination. Paternal mitochondrial DNA is normally undetectable in wildtype larva, but can be detected in the cross-progeny of fndc-1 mutant males. Moreover, loss of fndc-1 retards the rate of paternal mitochondria degradation, but not that of membranous organelles, a nematode specific membrane compartment whose fusion is required for sperm motility. This is the first example of a ubiquitin-independent mitophagy receptor playing a role in the selective degradation of sperm mitochondria.
    Keywords:  Autophagy; C. elegans; Maternal inheritance; Mitochondria; Mitophagy
    DOI:  https://doi.org/10.1016/j.ydbio.2019.06.016
  20. Mol Cell Biol. 2019 Jun 24. pii: MCB.00153-19. [Epub ahead of print]
    Oleson BJ, Broniowska KA, Yeo CT, Flancher M, Naatz A, Hogg N, Tarakanova VL, Corbett JA.
      In this report, we show that nitric oxide suppresses DNA damage response (DDR) signaling in the pancreatic β-cell line INS 832/13 and rat islets by inhibiting intermediary metabolism. Nitric oxide is known to inhibit complex IV of the electron transport chain and aconitase of the Krebs cycle. Non-β-cells compensate by increasing glycolytic metabolism to maintain ATP levels; however, β-cells lack this metabolic flexibility resulting in the nitric oxide-dependent decrease in ATP and NAD+ Like nitric oxide, mitochondrial toxins inhibit DDR signaling in β-cells by a mechanism that is associated with a decrease in ATP. Non-β-cells compensate for the effects of mitochondrial toxins with an adaptive shift to glycolytic ATP generation that allows for DDR signaling. Forcing non-β-cells to derive ATP via mitochondrial respiration (replacing glucose with galactose in the medium) and glucose deprivation sensitizes these cells to nitric oxide-mediated inhibition of DDR signaling. These findings indicate that metabolic flexibility is necessary to maintain DDR signaling under conditions in which mitochondrial oxidative metabolism is inhibited and support the inhibition of oxidative metabolism (decreased ATP) as one protective mechanism by which nitric oxide attenuates DDR-dependent β-cell apoptosis.
    DOI:  https://doi.org/10.1128/MCB.00153-19
  21. Nat Commun. 2019 Jun 25. 10(1): 2782
    Yang S, Wei J, Cui YH, Park G, Shah P, Deng Y, Aplin AE, Lu Z, Hwang S, He C, He YY.
      Melanoma is one of the most deadly and therapy-resistant cancers. Here we show that N6-methyladenosine (m6A) mRNA demethylation by fat mass and obesity-associated protein (FTO) increases melanoma growth and decreases response to anti-PD-1 blockade immunotherapy. FTO level is increased in human melanoma and enhances melanoma tumorigenesis in mice. FTO is induced by metabolic starvation stress through the autophagy and NF-κB pathway. Knockdown of FTO increases m6A methylation in the critical protumorigenic melanoma cell-intrinsic genes including PD-1 (PDCD1), CXCR4, and SOX10, leading to increased RNA decay through the m6A reader YTHDF2. Knockdown of FTO sensitizes melanoma cells to interferon gamma (IFNγ) and sensitizes melanoma to anti-PD-1 treatment in mice, depending on adaptive immunity. Our findings demonstrate a crucial role of FTO as an m6A demethylase in promoting melanoma tumorigenesis and anti-PD-1 resistance, and suggest that the combination of FTO inhibition with anti-PD-1 blockade may reduce the resistance to immunotherapy in melanoma.
    DOI:  https://doi.org/10.1038/s41467-019-10669-0
  22. Cell Rep. 2019 Jun 25. pii: S2211-1247(19)30731-4. [Epub ahead of print]27(13): 3939-3955.e6
    Yucel N, Wang YX, Mai T, Porpiglia E, Lund PJ, Markov G, Garcia BA, Bendall SC, Angelo M, Blau HM.
      The impact of glucose metabolism on muscle regeneration remains unresolved. We identify glucose metabolism as a crucial driver of histone acetylation and myogenic cell fate. We use single-cell mass cytometry (CyTOF) and flow cytometry to characterize the histone acetylation and metabolic states of quiescent, activated, and differentiating muscle stem cells (MuSCs). We find glucose is dispensable for mitochondrial respiration in proliferating MuSCs, so that glucose becomes available for maintaining high histone acetylation via acetyl-CoA. Conversely, quiescent and differentiating MuSCs increase glucose utilization for respiration and have consequently reduced acetylation. Pyruvate dehydrogenase (PDH) activity serves as a rheostat for histone acetylation and must be controlled for muscle regeneration. Increased PDH activity in proliferation increases histone acetylation and chromatin accessibility at genes that must be silenced for differentiation to proceed, and thus promotes self-renewal. These results highlight metabolism as a determinant of MuSC histone acetylation, fate, and function during muscle regeneration.
    DOI:  https://doi.org/10.1016/j.celrep.2019.05.092
  23. Sci Rep. 2019 Jun 26. 9(1): 9299
    Zwarts I, van Zutphen T, Kruit JK, Liu W, Oosterveer MH, Verkade HJ, Uhlenhaut NH, Jonker JW.
      Fructose has become a major constituent of our modern diet and is implicated as an underlying cause in the development of metabolic diseases. The fructose transporter GLUT5 (SLC2A5) is required for intestinal fructose absorption. GLUT5 expression is induced in the intestine and skeletal muscle of type 2 diabetes (T2D) patients and in certain cancers that are dependent on fructose metabolism, indicating that modulation of GLUT5 levels could have potential in the treatment of these diseases. Using an unbiased screen for transcriptional control of the human GLUT5 promoter we identified a strong and specific regulation by liver X receptor α (LXRα, NR1H3). Using promoter truncations and site-directed mutagenesis we identified a functional LXR response element (LXRE) in the human GLUT5 promoter, located at -385 bp relative to the transcriptional start site (TSS). Finally, mice treated with LXR agonist T0901317 showed an increase in Glut5 mRNA and protein levels in duodenum and adipose tissue, underscoring the in vivo relevance of its regulation by LXR. Together, our findings show that LXRα regulates GLUT5 in mice and humans. As a ligand-activated transcription factor, LXRα might provide novel pharmacologic strategies for the selective modulation of GLUT5 activity in the treatment of metabolic disease as well as cancer.
    DOI:  https://doi.org/10.1038/s41598-019-45803-x
  24. J Mol Biol. 2019 Jun 20. pii: S0022-2836(19)30400-0. [Epub ahead of print]
    Dukatz M, Requena CE, Emperle M, Hajkova P, Sarkies P, Jeltsch A.
      Recently, it has been discovered that different DNA-(cytosine C5)-methyltransferases including DNMT3A generate low levels of 3mC (Rosic et al., Nat. Genet. 2018, 50, 452-459). This reaction resulted in the co-evolution of DNMTs and ALKB2 DNA repair enzymes, but its mechanism remained elusive. Here, we investigated the catalytic mechanism of DNMT3A for cytosine N3 methylation. We generated several DNMT3A variants with mutated catalytic residues and measured their activities in 5mC and 3mC generation by liquid chromatography linked to tandem mass spectrometry. Our data suggest that the methylation of N3 instead of C5 is caused by an inverted binding of the flipped cytosine target base into the active site pocket of the DNA methyltransferase, which is partially compatible with the arrangement of catalytic amino acid residues. Given that all DNA-(cytosine C5)-methyltransferases have a common catalytic mechanism it is likely that other enzymes of this class generate 3mC following the same mechanism.
    Keywords:  DNA methylation; DNA methyltransferase; Enzyme catalysis; Enzyme mechanism
    DOI:  https://doi.org/10.1016/j.jmb.2019.06.015
  25. Autophagy. 2019 Jun 25.
    Di Rienzo M, Piacentini M, Fimia GM.
      The Ser/Thr protein kinase ULK1 is an upstream macroautophagy/autophagy regulator that is rapidly activated to ensure a proper adaptive response to stress conditions. Signaling pathways modulating ULK1 activity have been extensively characterized in response to nutrient/energy shortage, which mainly act by mediating ULK1 post-translational modifications, such as phosphorylation, acetylation and ubiquitination. Less characterized is how tissue-specific stress signals are able to activate ULK1 to induce autophagy. Our recent study has uncovered the E3 ubiquitin ligase TRIM32 as a novel ULK1 activator that regulates autophagy in muscle cells upon atrophy induction. TRIM32 is conveyed to ULK1 by the autophagy cofactor AMBRA1 to stimulate its kinase activity through unanchored K63-linked polyubiquitin chains. Notably, mutations in TRIM32 responsible for limb-girdle muscular dystrophy 2H disrupt its ability to bind ULK1 and to induce autophagy in muscle cells, resulting in a dysregulated activation of the atrophic process. In conclusion, we have identified a novel molecular mechanism by which autophagy is regulated in muscles, whose alteration is associated with the development of muscular dystrophy.
    Keywords:  AMBRA1; TRIM32; ULK1; muscle atrophy; muscular dystrophy; unanchored polyubiquitin
    DOI:  https://doi.org/10.1080/15548627.2019.1635385
  26. Tomography. 2019 Jun;5(2): 239-247
    Laustsen C, Lipsø K, Østergaard JA, Nielsen PM, Bertelsen LB, Flyvbjerg A, Pedersen M, Palm F, Ardenkjær-Larsen JH.
      Intrarenal hypoxia develops within a few days after the onset of insulinopenic diabetes in an experimental animal model (ie, a model of type-1 diabetes). Although diabetes-induced hypoxia results in increased renal lactate formation, mitochondrial function is well maintained, a condition commonly referred to as pseudohypoxia. However, the metabolic effects of significantly elevated lactate levels remain unclear. We therefore investigated in diabetic animals the response to acute intrarenal hypoxia in the presence of high renal lactate formation to delineate mechanistic pathways and compare these findings to healthy control animals. Hyperpolarized 13C-MRI and blood oxygenation level-dependent 1H-MRI was used to investigate the renal metabolism of [1-13C]pyruvate and oxygenation following acutely altered oxygen content in the breathing gas in a streptozotocin rat model of type-1 diabetes with and without insulin treatment and compared with healthy control rats. The lactate signal in the diabetic kidney was reduced by 12%-16% during hypoxia in diabetic rats irrespective of insulin supplementation. In contrast, healthy controls displayed the well-known Pasteur effect manifested as a 10% increased lactate signal following reduction of oxygen in the inspired air. Reduced expression of the monocarboxyl transporter-4 may account for altered response to hypoxia in diabetes with a high intrarenal pyruvate-to-lactate conversion. Reduced intrarenal lactate formation in response to hypoxia in diabetes shows the existence of a different metabolic phenotype, which is independent of insulin, as insulin supplementation was unable to affect the pyruvate-to-lactate conversion in the diabetic kidney.
    Keywords:  MRI; hyperpolarization; kidney; renal metabolism; type 1-diabetes
    DOI:  https://doi.org/10.18383/j.tom.2019.00003
  27. Nat Commun. 2019 06 24. 10(1): 2767
    Agudelo LZ, Ferreira DMS, Dadvar S, Cervenka I, Ketscher L, Izadi M, Zhengye L, Furrer R, Handschin C, Venckunas T, Brazaitis M, Kamandulis S, Lanner JT, Ruas JL.
      The coactivator PGC-1α1 is activated by exercise training in skeletal muscle and promotes fatigue-resistance. In exercised muscle, PGC-1α1 enhances the expression of kynurenine aminotransferases (Kats), which convert kynurenine into kynurenic acid. This reduces kynurenine-associated neurotoxicity and generates glutamate as a byproduct. Here, we show that PGC-1α1 elevates aspartate and glutamate levels and increases the expression of glycolysis and malate-aspartate shuttle (MAS) genes. These interconnected processes improve energy utilization and transfer fuel-derived electrons to mitochondrial respiration. This PGC-1α1-dependent mechanism allows trained muscle to use kynurenine metabolism to increase the bioenergetic efficiency of glucose oxidation. Kat inhibition with carbidopa impairs aspartate biosynthesis, mitochondrial respiration, and reduces exercise performance and muscle force in mice. Our findings show that PGC-1α1 activates the MAS in skeletal muscle, supported by kynurenine catabolism, as part of the adaptations to endurance exercise. This crosstalk between kynurenine metabolism and the MAS may have important physiological and clinical implications.
    DOI:  https://doi.org/10.1038/s41467-019-10712-0