bims-mibica Biomed News
on Mitochondrial bioenergetics in cancer
Issue of 2020‒05‒03
27 papers selected by
Kelsey Fisher-Wellman
East Carolina University


  1. Cell Signal. 2020 Apr 23. pii: S0898-6568(20)30128-5. [Epub ahead of print] 109651
    Hunter CA, Koc H, Koc EC.
      Src family kinases (SFKs) play a crucial role in the regulation of multiple cellular pathways, including mitochondrial oxidative phosphorylation (OXPHOS). Aberrant activities of one of the most predominant SFKs, c-Src, was identified as a fundamental cause for dysfunctional cell signaling and implicated in cancer development and metastasis, especially in human hepatocellular carcinoma (HCC). Recent work in our laboratory revealed that c-Src is implicated in regulation of mitochondrial energy metabolism in cancer. In this study, we investigated the effect of c-Src expression on mitochondrial energy metabolism by examining changes in the expression and activities of OXPHOS complexes in liver cancer biopsies and cell lines. An increased expression of c-Src was correlated with an impaired expression of nuclear- and mitochondrial-encoded subunits of OXPHOS complexes I and IV, respectively, in metastatic biopsies and cell lines. Additionally, we observed a similar association between high c-Src and reduced OXPHOS complex expression and activity in mouse embryonic fibroblast (MEF) cell lines. Interestingly, the inhibition of c-Src kinase activity with the SFK inhibitor PP2 and c-Src siRNA stimulated the expression of complex I and IV subunits and increased their enzymatic activities in both cancer and normal cells. Evidence provided in this study reveals that c-Src impairs the expression and function of mitochondrial OXPHOS complexes, resulting in a significant defect in mitochondrial energy metabolism, which can be a contributing factor to the development and progression of liver cancer. Furthermore, our findings strongly suggest that SFK inhibitors should be used in the treatment of HCC and other cancers with aberrant c-Src kinase activity to improve mitochondrial energy metabolism.
    Keywords:  Human hepatocellular carcinoma; Liver cancer; Mitochondrial energy metabolism; Oxidative phosphorylation (OXPHOS); Src family kinases (SFK)
    DOI:  https://doi.org/10.1016/j.cellsig.2020.109651
  2. J Physiol. 2020 May 02.
    Glancy B, Kane DA, Kavazis AN, Goodwin ML, Willis WT, Gladden LB.
      Mitochondrial structures were probably observed microscopically in the 1840s, but the idea of oxidative phosphorylation (OXPHOS) within mitochondria did not appear until the 1930s. The foundation for research into energetics arose from Meyerhof's experiments on oxidation of lactate in isolated muscles recovering from electrical contractions in an O2 atmosphere. Today, we know that mitochondria are actually reticula and that the energy released from electron pairs being passed along the electron transport chain from NADH to O2 generates a membrane potential and pH gradient of protons that can enter the molecular machine of ATP Synthase to resynthesize ATP. Lactate stands at the crossroads of glycolytic and oxidative energy metabolism. Based on reported research and our own modeling in silico, we contend that lactate is not directly oxidized in the mitochondrial matrix. Instead, the interim glycolytic products (pyruvate and NADH) are held in cytosolic equilibrium with the products of the lactate dehydrogenase (LDH) reaction and the intermediates of the malate-aspartate and glycerol 3-phosphate shuttles. This equilibrium supplies the glycolytic products to the mitochondrial matrix for OXPHOS. LDH in the mitochondrial matrix is not compatible with the cytoplasmic/matrix redox gradient; its presence would drain matrix reducing power and substantially dissipate the proton motive force. OXPHOS requires O2 as the final electron acceptor, but O2 s upply is sufficient in most situations, including exercise and often acute illness. Recent studies suggest that atmospheric normoxia may constitute a cellular hyperoxia in mitochondrial disease. As research proceeds appropriate oxygenation levels should be carefully considered. Abstract figure legend Credit for the discovery of what would become known as mitochondria is given to Rudolf Albrecht von Kölliker in 1857; these structures were subsequently described in greater detail by Richard Altmann. In 1898, Benda used a derivation of the Greek words for 'thread' and 'granule' to name these structures 'mitochondria'. In 1907, Fletcher and Hopkins reported the disappearance of lactate in the presence of O2 in previously stimulated muscles. Approximately two decades later, Meyerhof's work on O2 consumption and lactate (La- ) resynthesis into glycogen during the recovery of isolated skeletal muscles from prior contractions was an early hint at the intersection of glycolysis and aerobic phosphorylation. Warburg related these phenomena to the metabolic physiology of cancer. Research by both Meyerhof and Emden led to discovery of the glycolytic pathway. In the 1930s, the work of Lundsgaard, Krebs, Kalckar, the Coris, Belitzer and Szent-Gyorgi, and subsequently Lipmann, Ochoa, Bensley & Hoerr and Claude in the 1940s led to establishing the bioenergetics of glycolysis and the TCA cycle and compounds of high phosphoryl transfer potential. The 1950s heralded the age of research using isolated, functioning mitochondria to explore bioenergetics, and featured prominently the work of Lehninger, Estabrook & Saktor, and Chance & Williams. In the 1960s, Peter Mitchell first proposed the chemiosmotic theory of oxidative phosphorylation, for which he was awarded the Nobel Prize. During this same decade, work by Borst clarified the malate-aspartate shuttle, wherein the exchange of anionic aspartate for undissociated glutamate (one negative charge exported from the matrix per exchange) is driven by the membrane potential (ΔΨ). Work by Skulachev in this decade and beyond further clarified mitochondrial bioenergetics and mitochondrial morphology. Boyer elucidated the nature of the ATP synthase, ultimately winning the Nobel Prize for his work. In the 1980s, David Nicholls further clarified mitochondrial bioenergetics, and the work of George Brooks initiated the era of the Cell-to-Cell Lactate Shuttle. Starting in the 1990s, research emerged suggesting that mitochondria are capable of transporting La- across the inner membrane and oxidizing it without the support of the cytosolic-mitochondrial electron shuttles (i.e. the malate-aspartate and glycerol-3-phosphate shuttles). The ultimate combustion of La- obviously takes place in the mitochondria; there is no question about that simple conclusion. However, our view is that La- is not directly oxidized by LDH in the mitochondrial matrix, but rather La- must first be converted to pyruvate (Pyr- ) in the cytosol or intermembrane space. Rationale for this view includes the high activity of the near-equilibrium enzyme LDH, which exceeds glycolytic capacity, the highly oxidized NAD+ /NADH ratio relative to the mitochondrial matrix, and the thermodynamic necessity for an energy-driven accumulation of shuttle species (e.g. ΔΨ-dependent aspartate-glutamate exchanger). Modeling in silico demonstrates that an active LDH in the matrix would render mitochondria nearly incapable of oxidizing Pyr- , a result which is inconsistent with decades of studies from hundreds of laboratories using both isolated mitochondria and permeabilized cells in which the mitochondrial reticulum remains intact. Healthy mitochondria function well, even at low O2 levels such that dysoxia is rare and low O2 is likely a minor factor in the increasing concentrations of La- typical with exercise or even many acute critical care situations.2 This article is protected by copyright. All rights reserved.
    DOI:  https://doi.org/10.1113/JP278930
  3. Biochim Biophys Acta Bioenerg. 2020 Apr 23. pii: S0005-2728(20)30063-3. [Epub ahead of print] 148213
    Adjobo-Hermans MJW, de Haas R, Willems PHGM, Wojtala A, van Emst-de Vries SE, Wagenaars JA, van den Brand M, Rodenburg RJ, Smeitink JAM, Nijtmans LG, Sazanov LA, Wieckowski MR, Koopman WJH.
      Mutations in NDUFS4, which encodes an accessory subunit of mitochondrial oxidative phosphorylation (OXPHOS) complex I (CI), induce Leigh syndrome (LS). LS is a poorly understood pediatric disorder featuring brain-specific anomalies and early death. To study the LS pathomechanism, we here compared OXPHOS proteomes between various Ndufs4-/- mouse tissues. Ndufs4-/- animals displayed significantly lower CI subunit levels in brain/diaphragm relative to other tissues (liver/heart/kidney/skeletal muscle), whereas other OXPHOS subunit levels were not reduced. Absence of NDUFS4 induced near complete absence of the NDUFA12 accessory subunit, a 50% reduction in other CI subunit levels, and an increase in specific CI assembly factors. Among the latter, NDUFAF2 was most highly increased. Regarding NDUFS4, NDUFA12 and NDUFAF2, identical results were obtained in Ndufs4-/- mouse embryonic fibroblasts (MEFs) and NDUFS4-mutated LS patient cells. Ndufs4-/- MEFs contained active CI in situ but blue-native-PAGE highlighted that NDUFAF2 attached to an inactive CI subcomplex (CI-830) and inactive assemblies of higher MW. In NDUFA12-mutated LS patient cells, NDUFA12 absence did not reduce NDUFS4 levels but triggered NDUFAF2 association to active CI. BN-PAGE revealed no such association in LS patient fibroblasts with mutations in other CI subunit-encoding genes where NDUFAF2 was attached to CI-830 (NDUFS1, NDUFV1 mutation) or not detected (NDUFS7 mutation). Supported by enzymological and CI in silico structural analysis, we conclude that absence of NDUFS4 induces near complete absence of NDUFA12 but not vice versa, and that NDUFAF2 stabilizes active CI in Ndufs4-/- mice and LS patient cells, perhaps in concert with mitochondrial inner membrane lipids.
    Keywords:  Fibroblasts; Leigh syndrome; NADH:ubiquinone oxidoreductase; Proteomics
    DOI:  https://doi.org/10.1016/j.bbabio.2020.148213
  4. Food Funct. 2020 Apr 29.
    Lin X, Wang L, Zhao L, Zhu Z, Chen T, Chen S, Tao Y, Zeng T, Zhong Y, Sun H, Wang Z, Zheng W, Zhang Y, Wu W, Nan K, Chen T.
      Gastric cancer is the fourth most common cancer and the second most frequent cause of cancer death worldwide. Chemotherapy is an important treatment. However, traditional chemotherapy drugs have low bioavailability and targeting ability. Therefore, we developed curcumin-encapsulated micelles for the treatment of gastric cancer and investigated their antitumor efficacy and active mechanism. Gastric cancer cells were treated with different concentrations of curcumin micelles. MTS cell proliferation assays, flow cytometry (FCM), real time cellular analysis (RTCA) and nude mice xenografts were used to evaluate the effects of curcumin micelles on gastric cancer cell growth in vitro and in vivo. Western blotting was performed to analyze the protein levels of the indicated molecules. A Seahorse bioenergetics analyzer was used to investigate alterations in oxygen consumption and the aerobic glycolysis rate. Curcumin micelles significantly inhibited proliferation and colony formation and induced apoptosis in gastric tumor cells compared to the control groups. We further investigated the mechanism of curcumin micelles on gastric tumor cells and demonstrated that curcumin micelles acted on mitochondrial proteins, causing changes in mitochondrial function and affecting mitochondrial bioenergetics. Furthermore, curcumin micelles decreased mitochondrial membrane potential, increased reactive oxygen species (ROS) generation and disrupted redox equilibrium. The nude mouse model verified that curcumin micelle treatment significantly attenuated tumor growth in vivo. Curcumin micelles suppress gastric tumor cell growth in vitro and in vivo. The mechanism may be related to increasing ROS generation, disrupting redox equilibrium and affecting mitochondrial bioenergetics.
    DOI:  https://doi.org/10.1039/d0fo00260g
  5. Methods Protoc. 2020 Apr 27. pii: E32. [Epub ahead of print]3(2):
    Sim JJ, Jeong KY.
      In this protocol, we introduced a method of measuring mitochondrial dysfunction to confirm the epithelial-mesenchymal transition (EMT) in pancreatic cancer cells under a hypoxic environment. There are many expertized and complicated methods to verify EMT. However, our methods have indicated that EMT can be identified by examining changes in reactive oxygen species (ROS) generation and membrane potential in mitochondria. To demonstrate whether the changes in the indicators of mitochondrial dysfunction are correlative to EMT, cell morphology, and expression of E-cadherin and N-cadherin were additionally observed. The results verified that a decrease in membrane potential and an increase in ROS in mitochondria were associated with EMT of pancreatic cancer cells. This protocol would be useful as a basis for providing an additional indicator for changes in the tumor microenvironment of pancreatic cancer cells relating to EMT under a hypoxic environment.
    Keywords:  ROS; epithelial–mesenchymal transition; hypoxia; membrane potential; mitochondria; pancreatic cancer cell
    DOI:  https://doi.org/10.3390/mps3020032
  6. J Am Chem Soc. 2020 Apr 29.
    Gupta C, Khaniya U, Chan CK, Dehez F, Shekhar M, Gunner MR, Sazanov L, Chipot C, Singharoy A.
      The mitochondrial respiratory chain, formed by five protein complexes, utilizes energy from catabolic processes to synthesize ATP. Complex I, the first and the largest protein complex of the chain, harvests electrons from NADH to reduce quinone, while pumping protons across the mitochondrial membrane. Detailed knowledge of the working principle of such coupled charge-transfer processes remains, however, fragmentary due to bottlenecks in understanding redox-driven conformational transitions and their interplay with the hydrated proton pathways. Complex I from Thermus thermophilus encases 16 subunits with nine iron-sulfur clusters, reduced by electrons from NADH. Here, employing the latest crystal structure of T. thermophilus complex I, we have used microsecond-scale molecular dynamics simulations to study the chemo-mechanical coupling between redox changes of the iron-sulfur clusters and conformational transitions across complex I. First, we identify the redox switches within complex I, which allosterically couple the dynamics of the quinone binding pocket to the site of NADH reduction. Second, our free-energy calculations reveal that the affinity of the quinone, specifically menaquinone, for the binding-site is higher than that of its reduced, menaquinol form - a design essential for menaquinol release. Remarkably, the barriers to diffusive menaquinone dynamics are lesser than that of the more ubiquitous ubiquinone, and the naphthoquinone headgroup of the former furnishes stronger binding interactions with the pocket, favoring menaquinone for charge transport in T. thermophilus. Our computations are consistent with experimentally validated mutations and hierarchize the key residues into three function-al classes, identifying new mutation targets. Third, long-range hydrogen-bond networks connecting the quinone-binding site to the transmembrane subunits are found to be responsible for proton pumping. Put together, the simulations reveal the molecular design principles linking redox reactions to quinone turnover to proton translocation in complex I.
    DOI:  https://doi.org/10.1021/jacs.9b13450
  7. Aging (Albany NY). 2020 Apr 28. 12
    Liu Y, Bai H, Guo F, Thai PN, Luo X, Zhang P, Yang C, Feng X, Zhu D, Guo J, Liang P, Xu Z, Yang H, Lu X.
      Human pluripotent stem cell-derived cardiomyocytes (hPSC-CMs) have great potential in biomedical applications. However, the immature state of cardiomyocytes obtained using existing protocols limits the application of hPSC-CMs. Unlike adult cardiac myocytes, hPSC-CMs generate ATP through an immature metabolic pathway-aerobic glycolysis, instead of mitochondrial oxidative phosphorylation (OXPHOS). Hence, metabolic switching is critical for functional maturation in hPSC-CMs. Peroxisome proliferator-activated receptor gamma coactivator 1α (PGC-1α) is a key regulator of mitochondrial biogenesis and metabolism, which may help promote cardiac maturation during development. In this study, we investigated the effects of PGC-1α and its activator ZLN005 on the maturation of human embryonic stem cell-derived cardiomyocyte (hESC-CM). hESC-CMs were generated using a chemically defined differentiation protocol and supplemented with either ZLN005 or DMSO (control) on differentiating days 10 to 12. Biological assays were then performed around day 30. ZLN005 treatment upregulated the expressions of PGC-1α and mitochondrial function-related genes in hESC-CMs and induced more mature energy metabolism compared with the control group. In addition, ZLN005 treatment increased cell sarcomere length, improved cell calcium handling, and enhanced intercellular connectivity. These findings support an effective approach to promote hESC-CM maturation, which is critical for the application of hESC-CM in disease modeling, drug screening, and engineering cardiac tissue.
    Keywords:  ZLN005; cardiomyocyte maturation; embryonic stem cells; metabolism; peroxisome proliferator-activated receptor gamma coactivator 1α
    DOI:  https://doi.org/10.18632/aging.103088
  8. Mol Biol Cell. 2020 Apr 29. mbcE19070413
    Black M, Arumugam P, Shukla S, Pradhan A, Ustiyan V, Milewski D, Kalinichenko VV, Kalin TV.
      Forkhead box M1 (FOXM1), a nuclear transcription factor which activates cell cycle regulatory genes, is highly expressed in a majority of human cancers. The function of FOXM1 independent of nuclear transcription is unknown. In the present study, we found the FOXM1 protein inside of the mitochondria. Using site-directed mutagenesis, we generated FOXM1 mutant proteins that localized to distinct cellular compartments, uncoupling the nuclear and mitochondrial functions of FOXM1. Directing FOXM1 into the mitochondria decreased mitochondrial mass, membrane potential, respiration and electron transport chain (ETC) activity. In mitochondria, the FOXM1 directly bound to and increased the pentatricopeptide repeat domain 1 (PTCD1) protein, a mitochondrial leucine-specific tRNA binding protein that inhibits leucine-rich ETC complexes. Mitochondrial FOXM1 did not change cellular proliferation. Thus, FOXM1 translocates into mitochondria and inhibits mitochondrial respiration by increasing PTCD1. We identify a new paradigm that FOXM1 regulates mitochondrial homeostasis in a process independent of nuclear transcription.
    DOI:  https://doi.org/10.1091/mbc.E19-07-0413
  9. Sci Rep. 2020 Apr 28. 10(1): 7110
    Chicherin IV, Baleva MV, Levitskii SA, Dashinimaev EB, Krasheninnikov IA, Kamenski P.
      The initiation of protein synthesis in bacteria is ruled by three canonical factors: IF1, IF2, and IF3. This system persists in human mitochondria; however, it functions in a rather different way due to specialization and adaptation to the organellar micro-environment. We focused on human mitochondrial IF3, which was earlier studied in vitro, but no knock-out cellular models have been published up to date. In this work, we generated human HeLa cell lines deficient in the MTIF3 gene and analyzed their mitochondrial function. Despite the lack of IF3mt in these cells, they preserved functional mitochondria capable of oxygen consumption and protein synthesis; however, the translation of ATP6 mRNA was selectively decreased which compromised the assembly of ATP synthase. Together with the analogous results obtained earlier for baker's yeast mitochondrial IF3, our findings point to a functional divergence of mitochondrial initiation factors from their bacterial ancestors.
    DOI:  https://doi.org/10.1038/s41598-020-64139-5
  10. Proc Natl Acad Sci U S A. 2020 May 01. pii: 202001387. [Epub ahead of print]
    Jedrychowski MP, Lu GZ, Szpyt J, Mariotti M, Garrity R, Paulo JA, Schweppe DK, Laznik-Bogoslavski D, Kazak L, Murphy MP, Gladyshev VN, Gygi SP, Chouchani ET, Spiegelman BM.
      Oxidation of cysteine thiols by physiological reactive oxygen species (ROS) initiates thermogenesis in brown and beige adipose tissues. Cellular selenocysteines, where sulfur is replaced with selenium, exhibit enhanced reactivity with ROS. Despite their critical roles in physiology, methods for broad and direct detection of proteogenic selenocysteines are limited. Here we developed a mass spectrometric method to interrogate incorporation of selenium into proteins. Unexpectedly, this approach revealed facultative incorporation of selenium as selenocysteine or selenomethionine into proteins that lack canonical encoding for selenocysteine. Selenium was selectively incorporated into regulatory sites on key metabolic proteins, including as selenocysteine-replacing cysteine at position 253 in uncoupling protein 1 (UCP1). This facultative utilization of selenium was initiated by increasing cellular levels of organic, but not inorganic, forms of selenium. Remarkably, dietary selenium supplementation elevated facultative incorporation into UCP1, elevated energy expenditure through thermogenic adipose tissue, and protected against obesity. Together, these findings reveal the existence of facultative protein selenation, which correlates with impacts on thermogenic adipocyte function and presumably other biological processes as well.
    Keywords:  ROS; brown adipose tissue; cysteine; selenocysteine
    DOI:  https://doi.org/10.1073/pnas.2001387117
  11. Redox Biol. 2020 Apr 20. pii: S2213-2317(20)30328-1. [Epub ahead of print]34 101539
    Mota-Martorell N, Jove M, Pradas I, Sanchez I, Gómez J, Naudi A, Barja G, Pamplona R.
      Mitochondrial reactive oxygen species (ROS) production, specifically at complex I (Cx I), has been widely suggested to be one of the determinants of species longevity. The present study follows a comparative approach to analyse complex I in heart tissue from 8 mammalian species with a longevity ranging from 3.5 to 46 years. Gene expression and protein content of selected Cx I subunits were analysed using droplet digital PCR (ddPCR) and western blot, respectively. Our results demonstrate: 1) the existence of species-specific differences in gene expression and protein content of Cx I in relation to longevity; 2) the achievement of a longevity phenotype is associated with low protein abundance of subunits NDUFV2 and NDUFS4 from the matrix hydrophilic domain of Cx I; and 3) long-lived mammals show also lower levels of VDAC (voltage-dependent anion channel) amount. These differences could be associated with the lower mitochondrial ROS production and slower aging rate of long-lived animals and, unexpectedly, with a low content of the mitochondrial permeability transition pore in these species.
    Keywords:  Complex I; Droplet digital PCR; Longevity; Mammals; Mitochondria; NDUFS4 subunit; NDUFV2 subunit; VDAC; Western blot
    DOI:  https://doi.org/10.1016/j.redox.2020.101539
  12. Diabetologia. 2020 Apr 29.
    Georgiadou E, Haythorne E, Dickerson MT, Lopez-Noriega L, Pullen TJ, da Silva Xavier G, Davis SPX, Martinez-Sanchez A, Semplici F, Rizzuto R, McGinty JA, French PM, Cane MC, Jacobson DA, Leclerc I, Rutter GA.
      AIMS/HYPOTHESIS: Mitochondrial oxidative metabolism is central to glucose-stimulated insulin secretion (GSIS). Whether Ca2+ uptake into pancreatic beta cell mitochondria potentiates or antagonises this process is still a matter of debate. Although the mitochondrial Ca2+ importer (MCU) complex is thought to represent the main route for Ca2+ transport across the inner mitochondrial membrane, its role in beta cells has not previously been examined in vivo.METHODS: Here, we inactivated the pore-forming subunit of the MCU, encoded by Mcu, selectively in mouse beta cells using Ins1Cre-mediated recombination. Whole or dissociated pancreatic islets were isolated and used for live beta cell fluorescence imaging of cytosolic or mitochondrial Ca2+ concentration and ATP production in response to increasing glucose concentrations. Electrophysiological recordings were also performed on whole islets. Serum and blood samples were collected to examine oral and i.p. glucose tolerance.
    RESULTS: Glucose-stimulated mitochondrial Ca2+ accumulation (p< 0.05), ATP production (p< 0.05) and insulin secretion (p< 0.01) were strongly inhibited in beta cell-specific Mcu-null (βMcu-KO) animals, in vitro, as compared with wild-type (WT) mice. Interestingly, cytosolic Ca2+ concentrations increased (p< 0.001), whereas mitochondrial membrane depolarisation improved in βMcu-KO animals. βMcu-KO mice displayed impaired in vivo insulin secretion at 5 min (p< 0.001) but not 15 min post-i.p. injection of glucose, whilst the opposite phenomenon was observed following an oral gavage at 5 min. Unexpectedly, glucose tolerance was improved (p< 0.05) in young βMcu-KO (<12 weeks), but not in older animals vs WT mice.
    CONCLUSIONS/INTERPRETATION: MCU is crucial for mitochondrial Ca2+ uptake in pancreatic beta cells and is required for normal GSIS. The apparent compensatory mechanisms that maintain glucose tolerance in βMcu-KO mice remain to be established.
    Keywords:  Calcium; Glucose homeostasis; Insulin secretion; Mitochondria; Mitochondrial Ca2+ uniporter (MCU); Pancreatic beta cells; Type 2 diabetes
    DOI:  https://doi.org/10.1007/s00125-020-05148-x
  13. Proc Natl Acad Sci U S A. 2020 Apr 27. pii: 201919250. [Epub ahead of print]
    Nilsson A, Haanstra JR, Engqvist M, Gerding A, Bakker BM, Klingmüller U, Teusink B, Nielsen J.
      Many cancer cells consume glutamine at high rates; counterintuitively, they simultaneously excrete glutamate, the first intermediate in glutamine metabolism. Glutamine consumption has been linked to replenishment of tricarboxylic acid cycle (TCA) intermediates and synthesis of adenosine triphosphate (ATP), but the reason for glutamate excretion is unclear. Here, we dynamically profile the uptake and excretion fluxes of a liver cancer cell line (HepG2) and use genome-scale metabolic modeling for in-depth analysis. We find that up to 30% of the glutamine is metabolized in the cytosol, primarily for nucleotide synthesis, producing cytosolic glutamate. We hypothesize that excreting glutamate helps the cell to increase the nucleotide synthesis rate to sustain growth. Indeed, we show experimentally that partial inhibition of glutamate excretion reduces cell growth. Our integrative approach thus links glutamine addiction to glutamate excretion in cancer and points toward potential drug targets.
    Keywords:  flux-balance analysis; genome-scale modeling; metabolic engineering; systems biology
    DOI:  https://doi.org/10.1073/pnas.1919250117
  14. J Mol Cell Cardiol. 2020 Apr 27. pii: S0022-2828(20)30119-X. [Epub ahead of print]
    Garbincius JF, Luongo TS, Elrod JW.
      Since the initial identification of the mitochondrial calcium uniporter (MCU) in 2011, several studies employing genetic models have attempted to decipher the role of mitochondrial calcium uptake in cardiac physiology. Confounding results in various mutant mouse models have led to an ongoing debate regarding the function of MCU in the heart. In this review, we evaluate and discuss the totality of evidence for mitochondrial calcium uptake in the cardiac stress response. We highlight recent reports that implicate MCU in the control of homeostatic cardiac metabolism and function. This review concludes with a discussion of current gaps in knowledge and remaining experiments to define how MCU contributes to contractile function, cell death, metabolic regulation, and heart failure progression.
    Keywords:  Calcium; Cardiac function; Energetics; Ischemia reperfusion; MCU; MICU1; Mitochondria; NCLX; Permeability transition
    DOI:  https://doi.org/10.1016/j.yjmcc.2020.04.029
  15. Trends Cancer. 2020 May;pii: S2405-8033(20)30073-X. [Epub ahead of print]6(5): 359-361
    Quon E, Hart ML, Sullivan LB.
      Lactate dehydrogenase (LDH) accounts for the fermentative component of aerobic glycolysis, a near ubiquitous metabolic alteration in cancer. Recently, Oshima et al. developed a bioavailable LDH inhibitor that decreases tumor growth in mice and functions synergistically with mitochondrial respiration inhibitors. These findings suggest a cooperative mechanism of action that targets redox homeostasis.
    DOI:  https://doi.org/10.1016/j.trecan.2020.02.012
  16. PLoS One. 2020 ;15(4): e0231761
    Gangcuangco LMA, Mitchell BI, Siriwardhana C, Kohorn LB, Chew GM, Bowler S, Kallianpur KJ, Chow DC, Ndhlovu LC, Gerschenson M, Shikuma CM.
      BACKGROUND: Cellular immunometabolism among people living with HIV (PLWH) on antiretroviral therapy (ART) remains under investigated. We assessed the relationships between mitochondrial oxidative phosphorylation (OXPHOS) in peripheral blood mononuclear cells (PBMCs) and blood parameters associated with HIV immune dysregulation.METHODS: PLWH ≥40 years old and on stable ART ≥3 months were enrolled (N = 149). OXPHOS complex I (CI, NADH dehydrogenase) and complex IV (CIV, cytochrome c oxidase) protein levels in PBMCs were quantified using immunoassays. Monocyte subsets and markers of T-cell activation, senescence, and exhaustion were measured on PBMC by flow cytometry. Plasma inflammatory mediators were quantified using a multiplex assay. HIV-uninfected group (N = 44) of similar age, gender, and ethnicity had available OXPHOS levels.
    RESULTS: PLWH had a median age of 51 years. Majority were male (88.6%), Caucasian (57.7%), and with undetectable plasma HIV RNA <50 copies/mL (84.6%). Median CI level was lower in PLWH compared with the HIV-seronegative group (65.5 vs 155.0 optical density/μg protein x 103, p <0.0001). There was no significant difference in median CIV levels. Lower OXPHOS levels correlated with lower CD4% and CD4/CD8 ratio. On multivariable linear regression adjusted for age, current use of zidovudine/didanosine, and HIV RNA (detectable versus undetectable), lower OXPHOS levels were significantly associated with higher MPO, SAA, SAP, and sVCAM, and higher frequencies of intermediate (CD14++CD16+) monocytes and TIGIT+TIM3+ CD4 T-cell (p<0.01).
    CONCLUSION: CI PBMC protein levels were decreased in PLWH on ART. Decreased OXPHOS correlated with disease severity and inflammation. Further studies on the relationship between immunometabolism and immune dysregulation in HIV are warranted.
    DOI:  https://doi.org/10.1371/journal.pone.0231761
  17. Proc Biol Sci. 2020 May 13. 287(1926): 20192713
    Gyllenhammer LE, Entringer S, Buss C, Wadhwa PD.
      Research on mechanisms underlying the phenomenon of developmental programming of health and disease has focused primarily on processes that are specific to cell types, organs and phenotypes of interest. However, the observation that exposure to suboptimal or adverse developmental conditions concomitantly influences a broad range of phenotypes suggests that these exposures may additionally exert effects through cellular mechanisms that are common, or shared, across these different cell and tissue types. It is in this context that we focus on cellular bioenergetics and propose that mitochondria, bioenergetic and signalling organelles, may represent a key cellular target underlying developmental programming. In this review, we discuss empirical findings in animals and humans that suggest that key structural and functional features of mitochondrial biology exhibit developmental plasticity, and are influenced by the same physiological pathways that are implicated in susceptibility for complex, common age-related disorders, and that these targets of mitochondrial developmental programming exhibit long-term temporal stability. We conclude by articulating current knowledge gaps and propose future research directions to bridge these gaps.
    Keywords:  bioenergetics; developmental programming; fetal programming; maternal–fetal–placental biology; mitochondria
    DOI:  https://doi.org/10.1098/rspb.2019.2713
  18. Cancer Discov. 2020 Apr 27. pii: CD-19-0959. [Epub ahead of print]
    Parker SJ, Amendola CR, Hollinshead KER, Yu Q, Yamamoto K, Encarnacion-Rosado J, Rose RE, LaRue MM, Sohn ASW, Biancur DE, Paulo JA, Gygi SP, Jones DR, Wang H, Philips MR, Bar-Sagi D, Mancias JD, Kimmelman AC.
      Pancreatic ductal adenocarcinoma (PDAC) evolves a complex microenvironment comprised of multiple cell types, including pancreatic stellate cells (PSCs). Previous studies have demonstrated that stromal supply of alanine, lipids, and nucleotides supports the metabolism, growth, and therapeutic resistance of PDAC. Here we demonstrate that alanine crosstalk between PSCs and PDAC is orchestrated by the utilization of specific transporters. PSCs utilize SLC1A4 and other transporter(s) to rapidly exchange and maintain environmental alanine concentrations. Moreover, PDAC cells upregulate SLC38A2 to supply their increased alanine demand. Cells lacking SLC38A2 fail to concentrate intracellular alanine and undergo a profound metabolic crisis resulting in markedly impaired tumor growth. Our results demonstrate that stromal-cancer metabolic niches can form through differential transporter expression, creating unique therapeutic opportunities to target metabolic demands of cancer.
    DOI:  https://doi.org/10.1158/2159-8290.CD-19-0959
  19. Antioxid Redox Signal. 2020 Apr 25.
    Novera W, Lee ZW, Nin DS, Dai MZ, Binte Idres S, Wu H, Damen JMA, Tuan ZT, Sim AYL, Long YC, Wu W, Huang RY, Deng LW.
      AIMS: Current treatment options for ovarian clear cell carcinoma (OCCC) are limited to combination of platinum-based and other cytotoxic agents to which patients respond poorly due to intrinsic chemo-resistance. There is therefore an urgent need to develop alternative therapeutic strategies for OCCC.RESULTS: Cysteine deprivation suppresses OCCC growth in vitro and in vivo with no apparent toxicity. Modes of cell death induced by cysteine deprivation in OCCC is determined by their innate metabolic profiles. Cysteine-deprived glycolytic OCCC is abolished primarily by oxidative stress-dependent necrosis and ferroptosis that can otherwise be prevented by pre-treatment with anti-oxidative agents. Meanwhile, OCCC that relies on mitochondria respiration for its bioenergetics, is suppressed through apoptosis that can otherwise be averted by pre-treatment with cysteine precursor alone, but not with anti-oxidative agents. Cysteine deprivation induces apoptosis in respiring OCCC by limiting iron-sulfur (Fe-S) cluster synthesis in the mitochondria, without which electron transport chain may be disrupted. Respiring OCCC responds to Fe-S cluster deficit by increasing iron influx into the mitochondria that leads to iron-overload, mitochondria damage and eventual cell death. Innovation/Conclusion: This study demonstrates the importance of cysteine availability in OCCC that is for its anti-oxidative property and its less appreciated role in mitochondria respiration. Regardless of OCCC metabolic profiles, cysteine deprivation abolishes both glycolytic and respiring OCCC growth in vitro and in vivo. This study highlights the therapeutic potential of cysteine deprivation for OCCC.
    DOI:  https://doi.org/10.1089/ars.2019.7850
  20. Biochim Biophys Acta Mol Basis Dis. 2020 Apr 22. pii: S0925-4439(20)30153-8. [Epub ahead of print] 165808
    Cho H, Cho YY, Shim MS, Lee JY, Lee HS, Kang HC.
      Mitochondria are considered one of the most important subcellular organelles for targeting and delivering drugs because mitochondria are the main location for various cellular functions and energy (i.e., ATP) production, and mitochondrial dysfunctions and malfunctions cause diverse diseases such as neurodegenerative disorders, cardiovascular disorders, metabolic disorders, and cancers. In particular, unique mitochondrial characteristics (e.g., negatively polarized membrane potential, alkaline pH, high reactive oxygen species (ROS) level, high glutathione level, high temperature, and paradoxical mitochondrial dynamics) in pathological cancers have been used as targets, signals, triggers, or driving forces for specific sensing/diagnosing/imaging of characteristic changes in mitochondria, targeted drug delivery on mitochondria, targeted drug delivery/accumulation into mitochondria, or stimuli-triggered drug release in mitochondria. In this review, we describe the distinctive structures, functions, and physiological properties of cancer mitochondria and discuss recent technologies of mitochondria-specific "key characteristic" sensing systems, mitochondria-targeted "drug delivery" systems, and mitochondrial stimuli-specific "drug release" systems as well as their strengths and weaknesses.
    Keywords:  Cancer targeting; Drug targeting; Mitochondria; Mitochondria targeting; Mitochondria-targeted drug delivery; Organelle targeting
    DOI:  https://doi.org/10.1016/j.bbadis.2020.165808
  21. iScience. 2020 Apr 11. pii: S2589-0042(20)30233-9. [Epub ahead of print]23(5): 101048
    Shiba-Fukushima K, Inoshita T, Sano O, Iwata H, Ishikawa KI, Okano H, Akamatsu W, Imai Y, Hattori N.
      Early-onset Parkinson's disease-associated PINK1-Parkin signaling maintains mitochondrial health. Therapeutic approaches for enhancing PINK1-Parkin signaling present a potential strategy for treating various diseases caused by mitochondrial dysfunction. We report two chemical enhancers of PINK1-Parkin signaling, identified using a robust cell-based high-throughput screening system. These small molecules, T0466 and T0467, activate Parkin mitochondrial translocation in dopaminergic neurons and myoblasts at low doses that do not induce mitochondrial accumulation of PINK1. Moreover, both compounds reduce unfolded mitochondrial protein levels, presumably through enhanced PINK1-Parkin signaling. These molecules also mitigate the locomotion defect, reduced ATP production, and disturbed mitochondrial Ca2+ response in the muscles along with the mitochondrial aggregation in dopaminergic neurons through reduced PINK1 activity in Drosophila. Our results suggested that T0466 and T0467 may hold promise as therapeutic reagents in Parkinson's disease and related disorders.
    Keywords:  Biological Sciences; Cell Biology; Neuroscience
    DOI:  https://doi.org/10.1016/j.isci.2020.101048
  22. Science. 2020 May 01. pii: eaat3987. [Epub ahead of print]368(6490):
    Knudsen NH, Stanya KJ, Hyde AL, Chalom MM, Alexander RK, Liou YH, Starost KA, Gangl MR, Jacobi D, Liu S, Sopariwala DH, Fonseca-Pereira D, Li J, Hu FB, Garrett WS, Narkar VA, Ortlund EA, Kim JH, Paton CM, Cooper JA, Lee CH.
      Repeated bouts of exercise condition muscle mitochondria to meet increased energy demand-an adaptive response associated with improved metabolic fitness. We found that the type 2 cytokine interleukin-13 (IL-13) is induced in exercising muscle, where it orchestrates metabolic reprogramming that preserves glycogen in favor of fatty acid oxidation and mitochondrial respiration. Exercise training-mediated mitochondrial biogenesis, running endurance, and beneficial glycemic effects were lost in Il13-/- mice. By contrast, enhanced muscle IL-13 signaling was sufficient to increase running distance, glucose tolerance, and mitochondrial activity similar to the effects of exercise training. In muscle, IL-13 acts through both its receptor IL-13Rα1 and the transcription factor Stat3. The genetic ablation of either of these downstream effectors reduced running capacity in mice. Thus, coordinated immunological and physiological responses mediate exercise-elicited metabolic adaptations that maximize muscle fuel economy.
    DOI:  https://doi.org/10.1126/science.aat3987
  23. Cell Metab. 2020 Apr 23. pii: S1550-4131(20)30183-2. [Epub ahead of print]
    Kulkarni AS, Gubbi S, Barzilai N.
      Biological aging involves an interplay of conserved and targetable molecular mechanisms, summarized as the hallmarks of aging. Metformin, a biguanide that combats age-related disorders and improves health span, is the first drug to be tested for its age-targeting effects in the large clinical trial-TAME (targeting aging by metformin). This review focuses on metformin's mechanisms in attenuating hallmarks of aging and their interconnectivity, by improving nutrient sensing, enhancing autophagy and intercellular communication, protecting against macromolecular damage, delaying stem cell aging, modulating mitochondrial function, regulating transcription, and lowering telomere attrition and senescence. These characteristics make metformin an attractive gerotherapeutic to translate to human trials.
    Keywords:  TAME; aging; aging hallmarks; health span; longevity; metabolism; metformin
    DOI:  https://doi.org/10.1016/j.cmet.2020.04.001
  24. Cell Rep. 2020 Apr 28. pii: S2211-1247(20)30514-3. [Epub ahead of print]31(4): 107565
    Gebert N, Cheng CW, Kirkpatrick JM, Di Fraia D, Yun J, Schädel P, Pace S, Garside GB, Werz O, Rudolph KL, Jasper H, Yilmaz ÖH, Ori A.
      The small intestine is responsible for nutrient absorption and one of the most important interfaces between the environment and the body. During aging, changes of the epithelium lead to food malabsorption and reduced barrier function, thus increasing disease risk. The drivers of these alterations remain poorly understood. Here, we compare the proteomes of intestinal crypts from mice across different anatomical regions and ages. We find that aging alters epithelial immunity, metabolism, and cell proliferation and is accompanied by region-dependent skewing in the cellular composition of the epithelium. Of note, short-term dietary restriction followed by refeeding partially restores the epithelium by promoting stem cell differentiation toward the secretory lineage. We identify Hmgcs2 (3-hydroxy-3-methylglutaryl-coenzyme A [CoA] synthetase 2), the rate-limiting enzyme for ketogenesis, as a modulator of stem cell differentiation that responds to dietary changes, and we provide an atlas of region- and age-dependent proteome changes of the small intestine.
    Keywords:  aging; dietary restriction; hmgcs2; intestine; ketone bodies; proteomics; stem cells
    DOI:  https://doi.org/10.1016/j.celrep.2020.107565
  25. Signal Transduct Target Ther. 2020 May 01. 5(1): 52
    Chen X, Wang C, Jiang Y, Wang Q, Tao Y, Zhang H, Zhao Y, Hu Y, Li C, Ye D, Liu D, Jiang W, Chin EY, Chen S, Liu Y, Wang M, Liu S, Zhang X.
      Wnt/β-catenin signaling plays a critical role in colorectal cancer (CRC) tumorigenesis and the homeostasis of colorectal cancer stem cells (CSCs), but its molecular mechanism remains unclear. B-cell lymphoma 3 (Bcl-3), a member of the IκB family, is overexpressed in CRC and promotes tumorigenicity. Here, we report a novel function of Bcl-3 in maintaining colorectal CSC homeostasis by activating Wnt/β-catenin signaling. Silencing Bcl-3 suppresses the self-renewal capacity of colorectal CSCs and sensitizes CRC cells to chemotherapeutic drugs through a decrease in Wnt/β-catenin signaling. Moreover, our data show that Bcl-3 is a crucial component of Wnt/β-catenin signaling and is essential for β-catenin transcriptional activity in CRC cells. Interestingly, Wnt3a increases the level and nuclear translocation of Bcl-3, which binds directly to β-catenin and enhances the acetylation of β-catenin at lysine 49 (Ac-K49-β-catenin) and transcriptional activity. Bcl-3 depletion decreases the Ac-K49-β-catenin level by increasing the level of histone deacetylase 1 to remove acetyl groups from β-catenin, thus interrupting Wnt/β-catenin activity. In CRC clinical specimens, Bcl-3 expression negatively correlates with the overall survival of CRC patients. A significantly positive correlation was found between the expression of Bcl-3 and Ac-K49-β-catenin. Collectively, our data reveal that Bcl-3 plays a crucial role in CRC chemoresistance and colorectal CSC maintenance via its modulation of the Ac-K49-β-catenin, which serves as a promising therapeutic target for CRC.
    DOI:  https://doi.org/10.1038/s41392-020-0138-6
  26. Int J Mol Sci. 2020 Apr 28. pii: E3096. [Epub ahead of print]21(9):
    Buttacavoli M, Albanese NN, Roz E, Pucci-Minafra I, Feo S, Cancemi P.
      Colon cancer is an aggressive tumor form with a poor prognosis. This study reports a comparative proteomic analysis performed by using two-dimensional differential in-gel electrophoresis (2D-DIGE) between 26 pooled colon cancer surgical tissues and adjacent non-tumoral tissues, to identify potential target proteins correlated with carcinogenesis. The DAVID functional classification tool revealed that most of the differentially regulated proteins, acting both intracellularly and extracellularly, concur across multiple cancer steps. The identified protein classes include proteins involved in cell proliferation, apoptosis, metabolic pathways, oxidative stress, cell motility, Ras signal transduction, and cytoskeleton. Interestingly, networks and pathways analysis showed that the identified proteins could be biologically inter-connected to the tumor-host microenvironment, including innate immune response, platelet and neutrophil degranulation, and hemostasis. Finally, transgelin (TAGL), here identified for the first time with four different protein species, collectively down-regulated in colon cancer tissues, emerged as a top-ranked biomarker for colorectal cancer (CRC). In conclusion, our findings revealed a different proteomic profiling in colon cancer tissues characterized by the deregulation of specific pathways involved in hallmarks of cancer. All of these proteins may represent promising novel colon cancer biomarkers and potential therapeutic targets, if validated in larger cohorts of patients.
    Keywords:  TAGL; colon cancer; pathway analysis; proteomic profiling; transgelin
    DOI:  https://doi.org/10.3390/ijms21093096
  27. Elife. 2020 May 01. pii: e53560. [Epub ahead of print]9
    Corrigan JK, Ramachandran D, He Y, Palmer CJ, Jurczak MJ, Chen R, Li B, Friedline RH, Kim JK, Ramsey JJ, Lantier L, McGuinness OP, , Banks A.
      Maintaining a healthy body weight requires an exquisite balance between energy intake and energy expenditure. To understand the genetic and environmental factors that contribute to the regulation of body weight, an important first step is to establish the normal range of metabolic values and primary sources contributing to variability. Energy metabolism is measured by powerful and sensitive indirect calorimetry devices. Analysis of nearly 10,000 wild-type mice from two large-scale experiments revealed that the largest variation in energy expenditure is due to body composition, ambient temperature, and institutional site of experimentation. We also analyze variation in 2,329 knockout strains and establish a reference for the magnitude of metabolic changes. Based on these findings, we provide suggestions for how best to design and conduct energy balance experiments in rodents. These recommendations will move us closer to the goal of a centralized physiological repository to foster transparency, rigor and reproducibility in metabolic physiology experimentation.
    Keywords:  human biology; medicine; mouse; neuroscience
    DOI:  https://doi.org/10.7554/eLife.53560