bims-camemi Biomed news
on Mitochondrial metabolism in cancer
Issue of 2018‒12‒23
thirty-nine papers selected by
Christian Frezza
University of Cambridge, MRC Cancer Unit

  1. Cancer Metab. 2018 ;6 19
    Niu X, Chen YJ, Crawford PA, Patti GJ.
      Background: Recent in vitro and in vivo work has shown that lactate provides an important source of carbon for metabolic reactions in cancer cell mitochondria. An interesting question is whether lactate is oxidized by lactate dehydrogenase (LDH) in the cytosol and/or in mitochondria. Since metabolic processes in the cytosol and mitochondria are affected by redox balance, the location of LDH may have important regulatory implications in cancer metabolism.Methods: Within most mammalian cells, metabolic processes are physically separated by membrane-bound compartments. Our general understanding of this spatial organization and its role in cellular function, however, suffers from the limited number of techniques to localize enzymatic activities within a cell. Here, we describe an approach to assess metabolic compartmentalization by monitoring the activity of pharmacological inhibitors that cannot be transported into specific cellular compartments.
    Results: Oxamate, which chemically resembles pyruvate, is transported into mitochondria and inhibits LDH activity in purified mitochondria. GSK-2837808A, in contrast, is a competitive inhibitor of NAD, which cannot cross the inner mitochondrial membrane. GSK-2837808A did not inhibit the LDH activity of intact mitochondria, but GSK-2837808A did inhibit LDH activity after the inner mitochondrial membrane was disrupted.
    Conclusions: Our results are consistent with some mitochondrial LDH that is accessible to oxamate, but inaccessible to GSK-2837808A until mitochondria are homogenized. This strategy of using inhibitors with selective access to subcellular compartments, which we refer to as transport-exclusion pharmacology, is broadly applicable to localize other metabolic reactions within cells.
    Keywords:  Lactate; Lactate dehydrogenase; Redox balance; Transport-exclusion pharmacology
  2. Antioxidants (Basel). 2018 Dec 18. pii: E195. [Epub ahead of print]7(12):
    Jhun BS, O-Uchi J, Adaniya SM, Cypress MW, Yoon Y.
      Abnormal mitochondrial morphology, especially fragmented mitochondria, and mitochondrial dysfunction are hallmarks of a variety of human diseases including heart failure (HF). Although emerging evidence suggests a link between mitochondrial fragmentation and cardiac dysfunction, it is still not well described which cardiac signaling pathway regulates mitochondrial morphology and function under pathophysiological conditions such as HF. Mitochondria change their shape and location via the activity of mitochondrial fission and fusion proteins. This mechanism is suggested as an important modulator for mitochondrial and cellular functions including bioenergetics, reactive oxygen species (ROS) generation, spatiotemporal dynamics of Ca2+ signaling, cell growth, and death in the mammalian cell- and tissue-specific manners. Recent reports show that a mitochondrial fission protein, dynamin-like/related protein 1 (DLP1/Drp1), is post-translationally modified via cell signaling pathways, which control its subcellular localization, stability, and activity in cardiomyocytes/heart. In this review, we summarize the possible molecular mechanisms for causing post-translational modifications (PTMs) of DLP1/Drp1 in cardiomyocytes, and further discuss how these PTMs of DLP1/Drp1 mediate abnormal mitochondrial morphology and mitochondrial dysfunction under adrenergic signaling activation that contributes to the development and progression of HF.
    Keywords:  Ca2+/calmodulin-dependent protein kinase II (CaMKII); GTPase; adrenoceptor; apoptosis; calcineurin; mitochondrial permeability transition pore; phosphorylation; protein kinase A (PKA); protein kinase D (PKD)
  3. J Mol Cell Cardiol. 2018 Dec 17. pii: S0022-2828(18)31296-3. [Epub ahead of print]
    Xu P, Damschroder D, Zhang M, Ryall KA, Adler PN, Saucerman JJ, Wessells RJ, Yan Z.
      In yeast, the Atg2-Atg18 complex regulates Atg9 recycling from phagophore assembly site during autophagy; their function in higher eukaryotes remains largely unknown. In a targeted screening in Drosophila melanogaster, we show that Mef2-GAL4-RNAi-mediated knockdown of Atg2, Atg9 or Atg18 in the heart and indirect flight muscles led to shortened healthspan (declined locomotive function) and lifespan. These flies displayed an accelerated age-dependent loss of cardiac function along with cardiac hypertrophy (increased heart tube wall thickness) and structural abnormality (distortion of the lumen surface). Using the Mef2-GAL4-MitoTimer mitochondrial reporter system and transmission electron microscopy, we observed significant elongation of mitochondria and reduced number of lysosome-targeted autophagosomes containing mitochondria in the heart tube but exaggerated mitochondrial fragmentation and reduced mitochondrial density in indirect flight muscles. These findings provide the first direct evidence of the importance of Atg2-Atg18/Atg9 autophagy complex in the maintenance of mitochondrial integrity and, regulation of heart and muscle functions in Drosophila, raising the possibility of augmenting Atg2-Atg18/Atg9 activity in promoting mitochondrial health and, muscle and heart function.
    Keywords:  Autophagy related genes; Cardiac function; Functional aging; Healthspan; Indirect flight muscle; Lifespan; Mitophagy; Negative geotaxis
  4. J Biol Chem. 2018 Dec 18. pii: jbc.RA118.004869. [Epub ahead of print]
    Egnatchik RA, Leamy AK, Sacco SA, Cheah YE, Shiota M, Young JD.
      Hepatocyte lipotoxicity is characterized by aberrant mitochondrial metabolism, which predisposes cells to oxidative stress and apoptosis.  Previously, we reported that translocation of calcium from the ER to mitochondria of palmitate-treated hepatocytes activated anaplerotic flux from glutamine to alpha-ketoglutarate (αKG), which subsequently entered the citric acid cycle (CAC) for oxidation. We hypothesized that increased glutamine anaplerosis fueled elevations in CAC flux and oxidative stress following palmitate treatment. To test this hypothesis, primary rat hepatocytes or immortalized H4IIEC3 rat hepatoma cells were treated with lipotoxic levels of palmitate while modulating anaplerotic pathways leading to αKG. We found that culture media supplemented with glutamine, glutamate, or dimethyl-αKG increased palmitate lipotoxicity compared to media that lacked these anaplerotic substrates. Knockdown of glutamate-oxaloacetate transaminase (GOT) activity significantly reduced the lipotoxic effects of palmitate, while knockdown of glutamate dehydrogenase (Glud1) had no effect on palmitate lipotoxicity. 13C flux analysis of H4IIEC3 cells co-treated with palmitate and the pan-transaminase inhibitor aminooxyacetic acid (AOA) confirmed that reductions in lipotoxic markers were associated with decreases in anaplerosis, CAC flux, and oxygen consumption. Taken together, these results demonstrate that lipotoxic palmitate treatments enhance anaplerosis in cultured rat hepatocytes, causing a shift to aberrant transaminase metabolism that fuels CAC dysregulation and oxidative stress.
    Keywords:  anaplerosis; fatty acid; fatty liver disease; glutamine; hepatocyte; lipotoxicity; metabolic flux analysis; tricarboxylic acid cycle (TCA cycle) (Krebs cycle)
  5. F1000Res. 2018 ;pii: F1000 Faculty Rev-1858. [Epub ahead of print]7
    Pallafacchina G, Zanin S, Rizzuto R.
      In the last few decades, a large body of experimental evidence has highlighted the complex role for mitochondria in eukaryotic cells: they are not only the site of aerobic metabolism (thus providing most of the ATP supply for endergonic processes) but also a crucial checkpoint of cell death processes (both necrosis and apoptosis) and autophagy. For this purpose, mitochondria must receive and decode the wide variety of physiological and pathological stimuli impacting on the cell. The "old" notion that mitochondria possess a sophisticated machinery for accumulating and releasing Ca 2+, the most common and versatile second messenger of eukaryotic cells, is thus no surprise. What may be surprising is that the identification of the molecules involved in mitochondrial Ca 2+ transport occurred only in the last decade for both the influx (the mitochondrial Ca 2+ uniporter, MCU) and the efflux (the sodium calcium exchanger, NCX) pathways. In this review, we will focus on the description of the amazing molecular complexity of the MCU complex, highlighting the numerous functional implications of the tissue-specific expression of the variants of the channel pore components (MCU/MCUb) and of the associated proteins (MICU 1, 2, and 3, EMRE, and MCUR1).
    Keywords:  MCU; MICU; Mitochondria; mitochondrial Ca2+ uptake; mitochondrial ion transport
  6. Front Physiol. 2018 ;9 1572
    Newell C, Sabouny R, Hittel DS, Shutt TE, Khan A, Klein MS, Shearer J.
      Mesenchymal stem cells (MSCs) are the most commonly used cells in tissue engineering and regenerative medicine. MSCs can promote host tissue repair through several different mechanisms including donor cell engraftment, release of cell signaling factors, and the transfer of healthy organelles to the host. In the present study, we examine the specific impacts of MSCs on mitochondrial morphology and function in host tissues. Employing in vitro cell culture of inherited mitochondrial disease and an in vivo animal experimental model of low-grade inflammation (high fat feeding), we show human-derived MSCs to alter mitochondrial function. MSC co-culture with skin fibroblasts from mitochondrial disease patients rescued aberrant mitochondrial morphology from a fission state to a more fused appearance indicating an effect of MSC co-culture on host cell mitochondrial network formation. In vivo experiments confirmed mitochondrial abundance and mitochondrial oxygen consumption rates were elevated in host tissues following MSC treatment. Furthermore, microarray profiling identified 226 genes with differential expression in the liver of animals treated with MSC, with cellular signaling, and actin cytoskeleton regulation as key upregulated processes. Collectively, our data indicate that MSC therapy rescues impaired mitochondrial morphology, enhances host metabolic capacity, and induces widespread host gene shifting. These results highlight the potential of MSCs to modulate mitochondria in both inherited and pathological disease states.
    Keywords:  hepatic; high-fat diet; metabolic inflammation; metabolism; mitochondrial regulation
  7. J Cell Biol. 2018 Dec 20. pii: jcb.201807204. [Epub ahead of print]
    Zhou J, Wang X, Wang M, Chang Y, Zhang F, Ban Z, Tang R, Gan Q, Wu S, Guo Y, Zhang Q, Wang F, Zhao L, Jing Y, Qian W, Wang G, Guo W, Yang C.
      Amino acid catabolism is frequently executed in mitochondria; however, it is largely unknown how aberrant amino acid metabolism affects mitochondria. Here we report the requirement for mitochondrial saccharopine degradation in mitochondrial homeostasis and animal development. In Caenorhbditis elegans, mutations in the saccharopine dehydrogenase (SDH) domain of the bi-functional enzyme α-aminoadipic semialdehyde synthase AASS-1 greatly elevate the lysine catabolic intermediate saccharopine, which causes mitochondrial damage by disrupting mitochondrial dynamics, leading to reduced adult animal growth. In mice, failure of mitochondrial saccharopine oxidation causes lethal mitochondrial damage in the liver, leading to postnatal developmental retardation and death. Importantly, genetic inactivation of genes that raise the mitochondrial saccharopine precursors lysine and α-ketoglutarate strongly suppresses SDH mutation-induced saccharopine accumulation and mitochondrial abnormalities in C. elegans Thus, adequate saccharopine catabolism is essential for mitochondrial homeostasis. Our study provides mechanistic and therapeutic insights for understanding and treating hyperlysinemia II (saccharopinuria), an aminoacidopathy with severe developmental defects.
  8. Biotechnol Adv. 2018 Dec 18. pii: S0734-9750(18)30205-2. [Epub ahead of print]
    Shimizu K, Matsuoka Y.
      Overflow metabolism is a common phenomenon observed at higher glycolytic flux in many bacteria, yeast (known as Crabtree effect), and mammalian cells including cancer cells (known as Warburg effect). This phenomenon has recently been characterized as the trade-offs between protein costs and enzyme efficiencies based on coarse-graining approaches. Moreover, it has been recognized that the glycolytic flux increases as the source of energy generation changes from energetically efficient respiration to inefficient respiro-fermentative or fermentative metabolism causing overflow metabolism. It is highly desired to clarify the metabolic regulation mechanisms behind such phenomena. Metabolic fluxes are located on top of the hierarchical regulation systems, and represent the outcome of the integrated response of all levels of cellular regulation systems. In the present article, we discuss about the different levels of regulation systems for the modulation of fluxes depending on the growth rate, growth condition such as oxygen limitation that alters the metabolism towards fermentation, and genetic perturbation affecting the source of energy generation from respiration to respiro-fermentative metabolism in relation to overflow metabolism. The intracellular metabolite of the upper glycolysis such as fructose 1,6-bisphosphate (FBP) plays an important role not only for flux sensing, but also for the regulation of the respiratory activity either directly or indirectly (via transcription factors) at higher growth rate. The glycolytic flux regulation is backed up (enhanced) by unphosphorylated EIIA and HPr of the phosphotransferase system (PTS) components, together with the sugar-phosphate stress regulation, where the transcriptional regulation is further modulated by post-transcriptional regulation via the degradation of mRNA (stability of mRNA) in Escherichia coli. Moreover, the channeling may also play some role in modulating the glycolytic cascade reactions.
    Keywords:  Channeling; Glycolytic flux; Overflow metabolism; Oxygen limitation; Post-transcriptional regulation; Respiration; Respiro-fermentative metabolism; Sugar-phosphate stress; mRNA stability
  9. Arch Biochem Biophys. 2018 Dec 17. pii: S0003-9861(18)30718-5. [Epub ahead of print]
    Seidlmayer LK, Gomez-Garcia MR, Shiba T, Porter GA, Pavlov EV, Bers DM, Dedkova EN.
      We have previously demonstrated that inorganic polyphosphate (polyP) is a potent activator of the mitochondrial permeability transition pore (mPTP) in cardiac myocytes. PolyP depletion protected against Ca2+-induced mPTP opening, however it did not prevent and even exacerbated cell death during ischemia-reperfusion (I/R). The central goal of this study was to investigate potential molecular mechanisms underlying these dichotomous effects of polyP on mitochondrial function. We utilized a Langendorff-perfused heart model of I/R to monitor changes in polyP size and chain length at baseline, 20 min no-flow ischemia, and 15 min reperfusion. Freshly isolated cardiac myocytes and mitochondria from C57BL/6J (WT) and cyclophilin D knock-out (CypD KO) mice were used to measure polyP uptake, mPTP activity, mitochondrial membrane potential, respiration and ATP generation. We found that I/R induced a significant decrease in polyP chain length. We, therefore, tested, the ability of synthetic polyPs with different chain length to accumulate in mitochondria and induce mPTP. Both short and long chain polyPs accumulated in mitochondria in oligomycin-sensitive manner implicating potential involvement of mitochondrial ATP synthase in polyP transport. Notably, only short-chain polyP activated mPTP in WT myocytes, and this effect was prevented by mPTP inhibitor cyclosprorin A and absent in CypD KO myocytes. To the contrary, long-chain polyP suppressed mPTP activation, and enhanced ADP-linked respiration and ATP production. Our data indicate that 1) effect of polyP on cardiac function strongly depends on polymer chain length; and 2) short-chain polyPs (as increased in ischemia-reperfusion) induce mPTP and mitochondrial uncoupling, while long-chain polyPs contribute to energy generation and cell metabolism.
    Keywords:  ATP synthase; Animal models of human disease; Bioenergetics; Inorganic polyphosphate; Ischemia-reperfusion injury; Metabolism; Mitochondrial metabolism; Mitochondrial permeability transition pore
  10. Proteomics. 2018 Dec 17. e1800353
    Sreedhar A, Cassell T, Smith P, Lu D, Nam HW, Lane AN, Zhao Y.
      Uncoupling protein 2 (UCP2) is often upregulated in cancer cells. The UCP2 upregulation is positively correlated with enhanced proliferation, tumorigenesis, and metabolic alterations, thus suggesting that UCP2 upregulation could play a key role in sensing metabolic changes to promote tumorigenesis. To determine the global metabolic impact of UCP2 upregulation, we used 13 C6 glucose as a source molecule to 'trace' the metabolic fate of carbon atoms derived from glucose. UCP2 overexpression in skin epidermal cells enhanced the incorporation of 13 C-label to pyruvate, TCA cycle intermediates, nucleotides, and amino acids, suggesting that UCP2 upregulation reprograms cellular metabolism towards macromolecule synthesis. To the best of our knowledge, this is the first study to bring to light the overall metabolic differences caused by UCP2 upregulation. This article is protected by copyright. All rights reserved.
    Keywords:  Uncoupling protein 2; bioenergetics; metabolite profiling; metabolomics; mitochondrial metabolism; tumorigenesis
  11. Cell Rep. 2018 Dec 18. pii: S2211-1247(18)31875-8. [Epub ahead of print]25(12): 3465-3475.e4
    Kostic M, Katoshevski T, Sekler I.
      Calcium is a key regulator of mitochondrial function under both normal and pathological conditions. The mechanisms linking metabolic activity to mitochondrial Ca2+ signaling remain elusive, however. Here, by monitoring mitochondrial Ca2+ transients while manipulating mitochondrial membrane potential (ΔΨm), we found that mild fluctuations in ΔΨm, which do not affect Ca2+ influx, are sufficient to strongly regulate NCLX, the major efflux pathway of Ca2+ from the mitochondria. Phosphorylation of NCLX or expression of phosphomimicking mutant (S258D) rescued NCLX activity from ΔΨm-driven allosteric inhibition. By screening ΔΨm sensitivity of NCLX mutants, we also identified amino acid residues that, through functional interaction with Ser258, control NCLX regulation. Finally, we find that glucose-driven ΔΨm changes in pancreatic β-cells control mitochondrial Ca2+ signaling primarily via NCLX regulation. Our results identify a feedback control between metabolic activity and mitochondrial Ca2+ signaling and the "safety valve" NCLX phosphorylation that can rescue Ca2+ efflux in depolarized mitochondria.
    Keywords:  NCLX; mitochondrial Ca(2+) signaling; mitochondrial membrane potential
  12. J Physiol Sci. 2018 Dec 20.
    Mon EE, Wei FY, Ahmad RNR, Yamamoto T, Moroishi T, Tomizawa K.
      Mitochondrial iron is indispensable for heme biosynthesis and iron-sulfur cluster assembly. Several mitochondrial transmembrane proteins have been implicated to function in the biosynthesis of heme and iron-sulfur clusters by transporting reaction intermediates. However, several mitochondrial proteins related to iron metabolism remain uncharacterized. Here, we show that human sideroflexin 2 (SFXN2), a member of the SFXN protein family, is involved in mitochondrial iron metabolism. SFXN2 is an evolutionarily conserved protein that localized to mitochondria via its transmembrane domain. SFXN2-knockout (KO) cells had an increased mitochondrial iron content, which was associated with decreases in the heme content and heme-dependent enzyme activities. By contrast, the activities of iron-sulfur cluster-dependent enzymes were unchanged in SFXN2-KO cells. Moreover, abnormal iron metabolism impaired mitochondrial respiration in SFXN2-KO cells and accelerated iron-mediated death of these cells. Our findings demonstrate that SFXN2 functions in mitochondrial iron metabolism by regulating heme biosynthesis.
    Keywords:  Heme; Iron; Mitochondria; OXPHOS; Respiration
  13. Cancer Sci. 2018 Dec 18.
    Wang J, Konishi T.
      It has been reported that DNA double-strand breaks (DSB) can be induced by cytoplasm irradiation, and that both reactive free radicals and mitochondria are involved in DSB formation. However, the cellular antioxidative responses that are stimulated and the biological consequences of cytoplasmic irradiation remain unknown. Using the Single Particle Irradiation System to Cells proton microbeam facility at the National Institute of Radiological Sciences (Japan), the response of nuclear factor (erythroid-derived 2)-like 2 (NRF2) antioxidative signaling to cytoplasmic irradiation was studied in normal human lung fibroblast WI-38 cells. Cytoplasmic irradiation stimulated the localization of NRF2 to the nucleus and the expression of its target protein, heme oxygenase 1. Activation of NRF2 by tert-Butylhydroquinone mitigated the levels of DSB induced by cytoplasmic irradiation. Mitochondrial fragmentation was also promoted by cytoplasmic irradiation, and treatment with the mitochondrial fragmentation inhibitor Mdivi-1 suppressed cytoplasmic irradiation-induced NRF2 activation and aggravated DSB formation. Furthermore, p53 contributed to the induction of mitochondrial fragmentation and activation of NRF2, although the expression of p53 was significantly downregulated by cytoplasmic irradiation. Finally, mitochondrial superoxide (MitoSOX) production was enhanced under cytoplasmic irradiation, and the usage of the MitoSOX scavenger mito-tempol indicated that MitoSOX elicited alterations in p53 expression, mitochondrial dynamics, and NRF2 activation. Overall, NRF2 antioxidative response is suggested to play a key role against genomic DNA damage under cytoplasmic irradiation. Additionally, the upstream regulators of NRF2 provide new clues on cytoplasmic irradiation-induced biological processes and prevention of radiation risks. This article is protected by copyright. All rights reserved.
    Keywords:  Double-Strand DNA Breaks; NFE2L2 protein; cytoplasm; mitochondria; radiation biology
  14. Mol Metab. 2018 Dec 05. pii: S2212-8778(18)31034-2. [Epub ahead of print]
    Yi M, Ban Y, Tan Y, Xiong W, Li G, Xiang B.
      BACKGROUND: Cancer cells favor the use of less efficient glycolysis rather than mitochondrial oxidative phosphorylation to metabolize glucose, even in oxygen-rich conditions, a distinct metabolic alteration named the Warburg effect or aerobic glycolysis. In adult cells, bifunctional 6-phosphofructo-2-kinase/fructose-2, 6-bisphosphatase (PFKFB) family members are responsible for controlling the steady-state cytoplasmic levels of fructose-2,6-bisphosphate, which allosterically activates 6-phosphofructo-1-kinase, the key enzyme catalyzing the rate-limiting reaction of glycolysis. PFKFB3 and PFKFB4 are the two main isoenzymes overexpressed in various human cancers.SCOPE OF REVIEW: In this review, we summarize recent findings on the glycolytic and extraglycolytic roles of PFKFB3 and PFKFB4 in cancer progression and discuss potential therapies for targeting of PFKFB3 and PFKFB4.
    MAJOR CONCLUSIONS: PFKFB3 has the highest kinase activity to shunt glucose toward glycolysis, whereas PFKFB4 has more FBPase-2 activity, redirecting glucose toward the pentose phosphate pathway, providing reducing power for lipid biosynthesis and scavenging reactive oxygen species. Co-expression of PFKFB3 and PFKFB4 provides sufficient glucose metabolism to satisfy the bioenergetics demand and redox homeostasis requirements of cancer cells. Various reversible post-translational modifications of PFKFB3 enable cancer cells to flexibly adapt glucose metabolism in response to diverse stress conditions. In addition to playing important roles in tumor cell glucose metabolism, PFKFB3 and PFKFB4 are widely involved in multiple biological processes, such as cell cycle regulation, autophagy, and transcriptional regulation in a non-glycolysis-dependent manner.
    Keywords:  6-Phosphofructo-2-kinase/fructose-2,6-bisphosphatase; Aerobic glycolysis; Metabolic reprogramming; Pentose phosphate pathway; Warburg effect
  15. Curr Opin Physiol. 2018 Jun;3 94-100
    Moore AS, Holzbaur ELF.
      Mitochondria are dynamic organelles that can form complex networks in the cell. These networks can be rapidly remodeled in response to environmental changes or to support cellular needs. Mitochondrial dynamics are dependent on interactions with the cellular cytoskeleton - both microtubules and actin filaments. Mitochondrial-cytoskeletal interactions have a well-established role in mitochondrial motility. Recent progress indicates that these interactions also regulate the balance of mitochondrial fission/fusion, as well as mitochondria turnover and mitochondrial inheritance during cell division. We review these advances, and how this work has deepened our understanding of mitochondrial dynamics in the cell.
  16. Elife. 2018 Dec 18. pii: e40986. [Epub ahead of print]7
    Agrawal A, Pekkurnaz G, Koslover EF.
      Eukaryotic cells modulate their metabolism by organizing metabolic components in response to varying nutrient availability and energy demands. In rat axons, mitochondria respond to glucose levels by halting active transport in high glucose regions. We employ quantitative modeling to explore physical limits on spatial organization of mitochondria and localized metabolic enhancement through regulated stopping of processive motion. We delineate the role of key parameters, including cellular glucose uptake and consumption rates, that are expected to modulate mitochondrial distribution and metabolic response in spatially varying glucose conditions. Our estimates indicate that physiological brain glucose levels fall within the limited range necessary for metabolic enhancement. Hence mitochondrial localization is shown to be a plausible regulatory mechanism for neuronal metabolic flexibility in the presence of spatially heterogeneous glucose, as may occur in long processes of projection neurons. These findings provide a framework for the control of cellular bioenergetics through organelle trafficking.
    Keywords:  human; physics of living systems; rat
  17. Front Genet. 2018 ;9 593
    Towarnicki SG, Ballard JWO.
      Mitochondrial DNA (mtDNA) and the dietary macronutrient ratio are known to influence a wide range of phenotypic traits including longevity, fitness and energy production. Commonly mtDNA mutations are posited to be selectively neutral or reduce fitness and, to date, no selectively advantageous mtDNA mutations have been experimentally demonstrated in adult female Drosophila. Here we propose that a ND V161L mutation interacted with diets differing in their macronutrient ratios to influence organismal physiology and mitochondrial traits, but further studies are required to definitively show no linked mtDNA mutations are functionally significant. We utilized two mtDNA types (mitotypes) fed either a 1:2 Protein: Carbohydrate (P:C) or 1:16 P:C diet. When fed the former diet, Dahomey females harboring the V161L mitotype lived longer than those with the Alstonville mitotype and had higher climbing, basal reactive oxygen species (ROS) and elevated glutathione S-transferase E1 expression. The short lived Alstonville females ate more, had higher walking speed and elevated mitochondrial functions as suggested by respiratory control ratio (RCR), mtDNA copy number and expression of mitochondrial transcription termination factor 3. In contrast, Dahomey females fed 1:16 P:C were shorter lived, had higher fecundity, walking speed and mitochondrial functions. They had reduced climbing. This result suggests that mtDNA cannot be assumed to be a strictly neutral evolutionary marker when the dietary macronutrient ratio of a species varies over time and space and supports the hypothesis that mtDNA diversity may reflect the amount of time since the last selective sweep rather than strictly demographic processes.
    Keywords:  Drosophila; OXPHOS = oxidative phosphorylation; diet; mitochondria – DNA; mitotype
  18. Nat Commun. 2018 Dec 21. 9(1): 5442
    Reid MA, Allen AE, Liu S, Liberti MV, Liu P, Liu X, Dai Z, Gao X, Wang Q, Liu Y, Lai L, Locasale JW.
      Phosphoglycerate dehydrogenase (PHGDH) catalyzes the committed step in de novo serine biosynthesis. Paradoxically, PHGDH and serine synthesis are required in the presence of abundant environmental serine even when serine uptake exceeds the requirements for nucleotide synthesis. Here, we establish a mechanism for how PHGDH maintains nucleotide metabolism. We show that inhibition of PHGDH induces alterations in nucleotide metabolism independent of serine utilization. These changes are not attributable to defects in serine-derived nucleotide synthesis and redox maintenance, another key aspect of serine metabolism, but result from disruption of mass balance within central carbon metabolism. Mechanistically, this leads to simultaneous alterations in both the pentose phosphate pathway and the tri-carboxylic acid cycle, as we demonstrate based on a quantitative model. These findings define a mechanism whereby disruption of one metabolic pathway induces toxicity by simultaneously affecting the activity of multiple related pathways.
  19. BMC Biol. 2018 Dec 18. 16(1): 147
    Wu Z, Senchuk MM, Dues DJ, Johnson BK, Cooper JF, Lew L, Machiela E, Schaar CE, DeJonge H, Blackwell TK, Van Raamsdonk JM.
      BACKGROUND: The mitochondrial unfolded protein response (mitoUPR) is a stress response pathway activated by disruption of proteostasis in the mitochondria. This pathway has been proposed to influence lifespan, with studies suggesting that mitoUPR activation has complex effects on longevity.RESULTS: Here, we examined the contribution of the mitoUPR to the survival and lifespan of three long-lived mitochondrial mutants in Caenorhabditis elegans by modulating the levels of ATFS-1, the central transcription factor that mediates the mitoUPR. We found that clk-1, isp-1, and nuo-6 worms all exhibit an ATFS-1-dependent activation of the mitoUPR. While loss of atfs-1 during adulthood does not affect lifespan in any of these strains, absence of atfs-1 during development prevents clk-1 and isp-1 worms from reaching adulthood and reduces the lifespan of nuo-6 mutants. Examining the mechanism by which deletion of atfs-1 reverts nuo-6 lifespan to wild-type, we find that many of the transcriptional changes present in nuo-6 worms are mediated by ATFS-1. Genes exhibiting an ATFS-1-dependent upregulation in nuo-6 worms are enriched for transcripts that function in stress response and metabolism. Consistent, with this finding, loss of atfs-1 abolishes the enhanced stress resistance observed in nuo-6 mutants and prevents upregulation of multiple stress response pathways including the HIF-1-mediated hypoxia response, SKN-1-mediated oxidative stress response and DAF-16-mediated stress response.
    CONCLUSIONS: Our results suggest that in the long-lived mitochondrial mutant nuo-6 activation of the mitoUPR causes atfs-1-dependent changes in the expression of genes involved in stress response and metabolism, which contributes to the extended longevity observed in this mutant. This work demonstrates that the mitoUPR can modulate multiple stress response pathways and suggests that it is crucial for the development and lifespan of long-lived mitochondrial mutants.
    Keywords:  ATFS-1; Aging; C. elegans; Genetics; Lifespan; Mitochondria; Mitochondrial unfolded protein response; clk-1; isp-1; nuo-6
  20. Cytometry A. 2018 Dec 21.
    Clutton G, Mollan K, Hudgens M, Goonetilleke N.
      MitoTracker ® dyes are fluorescent compounds that allow cellular mitochondrial content to be measured semi-quantitatively by flow cytometry and have been used extensively in immunology publications. However, the parameters commonly reported, mean or median fluorescence intensity and percentage of cells that are MitoTracker® "high", can be influenced by variability in cytometer setup, dye stability, and operator subjectivity, making it difficult to compare data between experiments. Here, we describe a method to identify MitoTracker® "high" populations in an objective manner. When analyzing data, we first removed outliers using a pre-specified threshold, determined the fluorescence intensity of the brightest and dimmest events to obtain the fluorescence range and then gated cells within the top 90% of this range. This strategy substantially reduced variability between technical replicates and produced consistent results when data were analyzed by different operators. Consistent with previous reports and other analysis strategies, this analysis method demonstrated that within an individual, CD4+ T cells exhibit significantly higher mitochondrial mass than CD8+ T cells. Objective gating increases the reliability and utility of data generated using MitoTracker® dyes. © 2018 The Authors. Cytometry Part A published by Wiley Periodicals, Inc. on behalf of International Society for Advancement of Cytometry.
    Keywords:  T-lymphocytes; flow cytometry; immunologic techniques; metabolism; mitochondria
  21. BMC Mol Biol. 2018 Dec 19. 19(1): 12
    Harman A, Barth C.
      BACKGROUND: DNA replication requires contributions from various proteins, such as DNA helicases; in mitochondria Twinkle is important for maintaining and replicating mitochondrial DNA. Twinkle helicases are predicted to also possess primase activity, as has been shown in plants; however this activity appears to have been lost in metazoans. Given this, the study of Twinkle in other organisms is required to better understand the evolution of this family and the roles it performs within mitochondria.RESULTS: Here we describe the characterization of a Twinkle homologue, Twm1, in the amoeba Dictyostelium discoideum, a model organism for mitochondrial genetics and disease. We show that Twm1 is important for mitochondrial function as it maintains mitochondrial DNA copy number in vivo. Twm1 is a helicase which unwinds DNA resembling open forks, although it can act upon substrates with a single 3' overhang, albeit less efficiently. Furthermore, unlike human Twinkle, Twm1 has primase activity in vitro. Finally, using a novel in bacterio approach, we demonstrated that Twm1 promotes DNA replication.
    CONCLUSIONS: We conclude that Twm1 is a replicative mitochondrial DNA helicase which is capable of priming DNA for replication. Our results also suggest that non-metazoan Twinkle could function in the initiation of mitochondrial DNA replication. While further work is required, this study has illuminated several alternative processes of mitochondrial DNA maintenance which might also be performed by the Twinkle family of helicases.
    Keywords:  DNA helicase; DNA primase; Dictyostelium discoideum; Mitochondrial DNA replication; Twinkle
  22. Cell Rep. 2018 Dec 18. pii: S2211-1247(18)31871-0. [Epub ahead of print]25(12): 3315-3328.e6
    Agnew T, Goldsworthy M, Aguilar C, Morgan A, Simon M, Hilton H, Esapa C, Wu Y, Cater H, Bentley L, Scudamore C, Poulton J, Morten KJ, Thompson K, He L, Brown SDM, Taylor RW, Bowl MR, Cox RD.
      Mutations in genes essential for mitochondrial function have pleiotropic effects. The mechanisms underlying these traits yield insights into metabolic homeostasis and potential therapies. Here we report the characterization of a mouse model harboring a mutation in the tryptophanyl-tRNA synthetase 2 (Wars2) gene, encoding the mitochondrial-localized WARS2 protein. This hypomorphic allele causes progressive tissue-specific pathologies, including hearing loss, reduced adiposity, adipose tissue dysfunction, and hypertrophic cardiomyopathy. We demonstrate the tissue heterogeneity arises as a result of variable activation of the integrated stress response (ISR) pathway and the ability of certain tissues to respond to impaired mitochondrial translation. Many of the systemic metabolic effects are likely mediated through elevated fibroblast growth factor 21 (FGF21) following activation of the ISR in certain tissues. These findings demonstrate the potential pleiotropy associated with Wars2 mutations in patients.
    Keywords:  ISR; WARS2; adiposity; deafness; hypertrophic cardiomyopathy; mitochondrial dysfunction; pleiotropic
  23. PLoS Biol. 2018 Dec 20. 16(12): e2006265
    Bardai FH, Ordonez DG, Bailey RM, Hamm M, Lewis J, Feany MB.
      Mutations in leucine-rich repeat kinase 2 (LRRK2) are the most common cause of familial Parkinson disease. Genetics and neuropathology link Parkinson disease with the microtubule-binding protein tau, but the mechanism of action of LRRK2 mutations and the molecular connection between tau and Parkinson disease are unclear. Here, we investigate the interaction of LRRK and tau in Drosophila and mouse models of tauopathy. We find that either increasing or decreasing the level of fly Lrrk enhances tau neurotoxicity, which is further exacerbated by expressing Lrrk with dominantly acting Parkinson disease-associated mutations. At the cellular level, altering Lrrk expression promotes tau neurotoxicity via excess stabilization of filamentous actin (F-actin) and subsequent mislocalization of the critical mitochondrial fission protein dynamin-1-like protein (Drp1). Biochemically, monomeric LRRK2 exhibits actin-severing activity, which is reduced as increasing concentrations of wild-type LRRK2, or expression of mutant forms of LRRK2 promote oligomerization of the protein. Overall, our findings provide a potential mechanistic basis for a dominant negative mechanism in LRRK2-mediated Parkinson disease, suggest a common molecular pathway with other familial forms of Parkinson disease linked to abnormalities of mitochondrial dynamics and quality control, and raise the possibility of new therapeutic approaches to Parkinson disease and related disorders.
  24. EMBO J. 2018 Dec 20. pii: e99916. [Epub ahead of print]
    Bernardini JP, Brouwer JM, Tan IK, Sandow JJ, Huang S, Stafford CA, Bankovacki A, Riffkin CD, Wardak AZ, Czabotar PE, Lazarou M, Dewson G.
      The E3 ubiquitin ligase Parkin is a key effector of the removal of damaged mitochondria by mitophagy. Parkin determines cell fate in response to mitochondrial damage, with its loss promoting early onset Parkinson's disease and potentially also cancer progression. Controlling a cell's apoptotic response is essential to co-ordinate the removal of damaged mitochondria. We report that following mitochondrial damage-induced mitophagy, Parkin directly ubiquitinates the apoptotic effector protein BAK at a conserved lysine in its hydrophobic groove, a region that is crucial for BAK activation by BH3-only proteins and its homo-dimerisation during apoptosis. Ubiquitination inhibited BAK activity by impairing its activation and the formation of lethal BAK oligomers. Parkin also suppresses BAX-mediated apoptosis, but in the absence of BAX ubiquitination suggesting an indirect mechanism. In addition, we find that BAK-dependent mitochondrial outer membrane permeabilisation during apoptosis promotes PINK1-dependent Parkin activation. Hence, we propose that Parkin directly inhibits BAK to suppress errant apoptosis, thereby allowing the effective clearance of damaged mitochondria, but also promotes clearance of apoptotic mitochondria to limit their potential pro-inflammatory effect.
    Keywords:   BAK ; BAX ; Parkin; apoptosis; mitophagy
  25. Proc Natl Acad Sci U S A. 2018 Dec 17. pii: 201808950. [Epub ahead of print]
    Sun C, Li T, Song X, Huang L, Zang Q, Xu J, Bi N, Jiao G, Hao Y, Chen Y, Zhang R, Luo Z, Li X, Wang L, Wang Z, Song Y, He J, Abliz Z.
      Characterization of tumor metabolism with spatial information contributes to our understanding of complex cancer metabolic reprogramming, facilitating the discovery of potential metabolic vulnerabilities that might be targeted for tumor therapy. However, given the metabolic variability and flexibility of tumors, it is still challenging to characterize global metabolic alterations in heterogeneous cancer. Here, we propose a spatially resolved metabolomics approach to discover tumor-associated metabolites and metabolic enzymes directly in their native state. A variety of metabolites localized in different metabolic pathways were mapped by airflow-assisted desorption electrospray ionization mass spectrometry imaging (AFADESI-MSI) in tissues from 256 esophageal cancer patients. In combination with in situ metabolomics analysis, this method provided clues into tumor-associated metabolic pathways, including proline biosynthesis, glutamine metabolism, uridine metabolism, histidine metabolism, fatty acid biosynthesis, and polyamine biosynthesis. Six abnormally expressed metabolic enzymes that are closely associated with the altered metabolic pathways were further discovered in esophageal squamous cell carcinoma (ESCC). Notably, pyrroline-5-carboxylate reductase 2 (PYCR2) and uridine phosphorylase 1 (UPase1) were found to be altered in ESCC. The spatially resolved metabolomics reveal what occurs in cancer at the molecular level, from metabolites to enzymes, and thus provide insights into the understanding of cancer metabolic reprogramming.
    Keywords:  airflow-assisted ionization; esophageal cancer; mass spectrometry imaging; metabolic alterations; metabolomics
  26. Am J Physiol Endocrinol Metab. 2018 Dec 21.
    Ngo DT, Sverdlov AL, Karki S, Macartney-Coxson D, Stubbs RS, Farb MG, Carmine B, Hess DT, Colucci WS, Gokce N.
      AIM: Obesity, particularly visceral adiposity, has been linked to mitochondrial dysfunction and increased oxidative stress, which have been suggested as mechanisms of insulin resistance. The mechanism(s) behind this remain incompletely understood. In this study, we hypothesized that mitochondrial complex II dysfunction plays a role in impaired insulin sensitivity in visceral adipose tissue of patients with obesity.METHODS AND RESULTS: We obtained subcutaneous and visceral adipose tissue biopsies from 43 patients with obesity (BMI ≥ 30 kg/m2) during planned bariatric surgery. Compared to subcutaneous, visceral adipose tissue exhibited decreased complex II activity, which was restored with the reducing agent dithiothreitol (5mM) (p<0.01). A biotin switch assay identified that cysteine OPTM in complex II subunit A (SDHA) were increased in visceral vs. subcutaneous fat (p<0.05). Insulin treatment (100nM) stimulated complex II activity in subcutaneous fat (p<0.05). In contrast, insulin treatment of visceral fat led to a decrease in complex II activity (p<0.01), which was restored with the addition of the mitochondria-specific oxidant scavenger, mito-TEMPO (10µM). In a cohort of 10 patients with severe obesity, weight loss decreased OPTM and restored complex II activity, exclusively in the visceral depot.
    CONCLUSIONS: Mitochondrial complex II may be an unrecognized and novel mediator of insulin resistance associated with visceral adiposity. The activity of complex II is improved by weight loss which may contribute to metabolic improvements associated with bariatric surgery.
    Keywords:  Obesity; adipose tissue; mitochondrial complex II; mitochondrial function; succinate dehydrogenase
  27. Am J Hypertens. 2018 Dec 18.
    Zhang YL, Zhang R, Shen YF, Huang KY, He YY, Zhao JH, Jing ZC.
      Background: The shift of metabolism from mitochondrial oxidative phosphorylation to glycolysis and mitochondria binding partner of hexokinase are features common to cancer. These have been seen in pulmonary hypertension (PH) as well. An inhibitor of hexokinase 2 (HK 2), the small molecule 3-bromopyruvate (3-BrPA) is incredibly powerful and swift acting anticancer agent. However, whether it could be of potential benefit to PH was still unknown.METHODS: Sprague-Dawley rats with monocrotaline (MCT)-induced PH were administered two different oral doses of 3-BrPA (15mg/kg/d and 30mg/kg/d, respectively) for 14 days. Hemodynamic parameters were acquired by right heart catheterization. Histopathology, immunohistochemistry, transmission electron microscopy, flow cytometry, and assessments of relative protein expressions were done.
    RESULTS: Compared with MCT-treated group, 3-BrPA decreased mean pulmonary arterial pressure and pulmonary vascular resistance, and increased cardiac output. The 3-BrPA significantly suppressed proliferation in addition to enhancing apoptosis of pulmonary artery smooth muscle cells, attenuating small pulmonary artery remodeling and right ventricular hypertrophy. Treatment with 3-BrPA markedly reduced the mitochondrial membrane potential and restored mitochondrial structure. Furthermore, 3-BrPA significantly inhibited HK 2 expression but not HK 1. The expression of both pyruvate dehydrogenase kinase and lactate dehydrogenase was decreased, while pyruvate dehydrogenase and cytosolic Cytochrome c expression was up-regulated with 3-BrPA administration.
    CONCLUSIONS: This study demonstrates the reversal of PH by 3-BrPA is related to alteration in glycolysis and improved mitochondria function, indicating the "metabolic targeting" as a rational therapeutic strategy for PH.
  28. PLoS One. 2018 ;13(12): e0202784
    Real-Hohn A, Navegantes C, Ramos K, Ramos-Filho D, Cahuê F, Galina A, Salerno VP.
      Visceral lipid accumulation, organ hypertrophy and a reduction in skeletal muscle strength are all signs associated with the severity of obesity-related disease. Intermittent fasting (IF) and high-intensity intermittent exercise (HIIE) are natural strategies that, individually, can prevent and help treat obesity along with metabolic syndrome and its associated diseases. However, the combinatorial effect of IF and HIIE on energetic metabolism is currently not well understood. We hypothesized that their combination could have a potential for more than strictly additive benefits. Here, we show that two months of every-other-day intermittent fasting regimen combined with a high-intensity intermittent exercise protocol (IF/HIIE) produced a synergistic effect, enhancing physical endurance (vs. control, HIIE and IF) and optimizing metabolic pathways of energy production in male Wistar rats. The IF/HIIE group presented enhanced glucose tolerance (vs. control, HIIE and IF), lower levels of plasma insulin (vs. control and HIIE), and a global activation of low Km hexokinases in liver (vs. control, HIIE and IF), heart (vs. control and HIIE) and skeletal muscle (vs. control, HIIE and IF). The IF/HIIE synergism, rather than a simply additive effect, is evidenced by increase in muscle mass and cross-section area, activation of the FoF1 ATP synthase, and the gain of characteristics suggestive of augmented mitochondrial mass and efficiency observed in this group. Finally, important reductions in plasma oxidative stress markers were present preferentially in IF/HIIE group. These findings provide new insights for the implementation of non-pharmaceutical strategies to prevent/treat metabolic syndrome and associated diseases.
  29. PLoS One. 2018 ;13(12): e0209489
    Jackson M, Serada N, Sheehan M, Srinivasan S, Mason N, Guha M, Avadhani N.
      Osteosarcoma (OSA) is an aggressive mesenchymal tumor of the bone that affects children and occurs spontaneously in dogs. Human and canine OSA share similar clinical, biological and genetic features, which make dogs an excellent comparative model to investigate the etiology and pathogenesis of OSA. Mitochondrial (mt) defects have been reported in many different cancers including OSA, although it is not known whether these defects contribute to OSA progression and metastasis. Taking a comparative approach using canine OSA cell lines and tumor tissues we investigated the effects of mtDNA content and dysfunction on OSA biology. OSA tumor tissues had low mtDNA contents compared to the matched non-tumor tissues. We observed mitochondrial heterogeneity among the OSA cell lines and the most invasive cells expressing increased levels of OSA metastasis genes contained the highest amount of mitochondrial defects (reduced mtDNA copies, mt respiration, and expression of electron transport chain proteins). While mitochondria maintain a filamentous network in healthy cells, the mitochondrial morphology in OSA cells were mostly "donut shaped", typical of "stressed" mitochondria. Moreover the expression levels of mitochondrial retrograde signaling proteins Akt1, IGF1R, hnRNPA2 and NFkB correlated with the invasiveness of the OSA cells. Furthermore, we demonstrate the causal role of mitochondrial defects in inducing the invasive phenotype by Ethidium Bromide induced-mtDNA depletion in OSA cells. Our data suggest that defects in mitochondrial genome and function are prevalent in OSA and that lower mtDNA content is associated with higher tumor cell invasiveness. We propose that mt defects in OSA might serve as a prognostic biomarker and a target for therapeutic intervention in OSA patients.
  30. Proc Natl Acad Sci U S A. 2018 Dec 21. pii: 201811938. [Epub ahead of print]
    Spinazzi M, Radaelli E, Horré K, Arranz AM, Gounko NV, Agostinis P, Maia TM, Impens F, Morais VA, Lopez-Lluch G, Serneels L, Navas P, De Strooper B.
      The mitochondrial intramembrane rhomboid protease PARL has been implicated in diverse functions in vitro, but its physiological role in vivo remains unclear. Here we show that Parl ablation in mouse causes a necrotizing encephalomyelopathy similar to Leigh syndrome, a mitochondrial disease characterized by disrupted energy production. Mice with conditional PARL deficiency in the nervous system, but not in muscle, develop a similar phenotype as germline Parl KOs, demonstrating the vital role of PARL in neurological homeostasis. Genetic modification of two major PARL substrates, PINK1 and PGAM5, do not modify this severe neurological phenotype. Parl -/- brain mitochondria are affected by progressive ultrastructural changes and by defects in Complex III (CIII) activity, coenzyme Q (CoQ) biosynthesis, and mitochondrial calcium metabolism. PARL is necessary for the stable expression of TTC19, which is required for CIII activity, and of COQ4, which is essential in CoQ biosynthesis. Thus, PARL plays a previously overlooked constitutive role in the maintenance of the respiratory chain in the nervous system, and its deficiency causes progressive mitochondrial dysfunction and structural abnormalities leading to neuronal necrosis and Leigh-like syndrome.
    Keywords:  Leigh syndrome; mitochondria; neurodegeneration; respiratory chain; rhomboid protease
  31. Front Oncol. 2018 ;8 575
    Hossain F, Sorrentino C, Ucar DA, Peng Y, Matossian M, Wyczechowska D, Crabtree J, Zabaleta J, Morello S, Del Valle L, Burow M, Collins-Burow B, Pannuti A, Minter LM, Golde TE, Osborne BA, Miele L.
      Triple negative breast cancer (TNBC) patients have high risk of recurrence and metastasis, and current treatment options remain limited. Cancer stem-like cells (CSCs) have been linked to cancer initiation, progression and chemotherapy resistance. Notch signaling is a key pathway regulating TNBC CSC survival. Treatment of TNBC with PI3K or mTORC1/2 inhibitors results in drug-resistant, Notch-dependent CSC. However, downstream mechanisms and potentially druggable Notch effectors in TNBC CSCs are largely unknown. We studied the role of the AKT pathway and mitochondrial metabolism downstream of Notch signaling in TNBC CSC from cell lines representative of different TNBC molecular subtypes as well as a novel patient-derived model. We demonstrate that exposure of TNBC cells to recombinant Notch ligand Jagged1 leads to rapid AKT phosphorylation in a Notch1-dependent but RBP-Jκ independent fashion. This requires mTOR and IKKα. Jagged1 also stimulates mitochondrial respiration and fermentation in an AKT- and IKK-dependent fashion. Notch1 co-localizes with mitochondria in TNBC cells. Pharmacological inhibition of Notch cleavage by gamma secretase inhibitor PF-03084014 in combination with AKT inhibitor MK-2206 or IKK-targeted NF-κB inhibitor Bay11-7082 blocks secondary mammosphere formation from sorted CD90hi or CD44+CD24low (CSCs) cells. A TNBC patient-derived model gave comparable results. Besides mitochondrial oxidative metabolism, Jagged1 also triggers nuclear, NF-κB-dependent transcription of anti-apoptotic gene cIAP-2. This requires recruitment of Notch1, IKKα and NF-κB to the cIAP-2 promoter. Our observations support a model where Jagged1 triggers IKKα-dependent, mitochondrial and nuclear Notch1 signals that stimulate AKT phosphorylation, oxidative metabolism and transcription of survival genes in PTEN wild-type TNBC cells. These data suggest that combination treatments targeting the intersection of the Notch, AKT and NF-κB pathways have potential therapeutic applications against CSCs in TNBC cases with Notch1 and wild-type PTEN expression.
    Keywords:  NF-κB; cancer stem-like cells (CSCs); jagged; mitochondria; notch; triple-negative breast cancer
  32. J Physiol. 2018 Nov 22.
    Tarpey MD, Valencia AP, Jackson KC, Amorese AJ, Balestrieri NP, Renegar RH, Pratt SJP, Ryan TE, McClung JM, Lovering RM, Spangenburg EE.
      KEY POINTS: Breast cancer 1 early onset gene codes for the DNA repair enzyme, breast cancer type 1 susceptibility protein (BRCA1). The gene is prone to mutations that cause a loss of protein function. BRCA1/Brca1 has recently been found to regulate several cellular pathways beyond DNA repair and is expressed in skeletal muscle. Skeletal muscle specific knockout of Brca1 in mice caused a loss of muscle quality, identifiable by reductions in muscle force production and mitochondrial respiratory capacity. Loss of muscle quality was associated with a shift in muscle phenotype and an accumulation of mitochondrial DNA mutations. These results demonstrate that BRCA1 is necessary for skeletal muscle function and that increased mitochondrial DNA mutations may represent a potential underlying mechanism.ABSTRACT: Recent evidence suggests that the breast cancer 1 early onset gene (BRCA1) influences numerous peripheral tissues, including skeletal muscle. The present study aimed to determine whether induced-loss of the breast cancer type 1 susceptibility protein (Brca1) alters skeletal muscle function. We induced genetic ablation of exon 11 in the Brca1 gene specifically in the skeletal muscle of adult mice to generate skeletal muscle-specific Brca1 homozygote knockout (Brca1KOsmi ) mice. Brca1KOsmi exhibited kyphosis and decreased maximal isometric force in limb muscles compared to age-matched wild-type mice. Brca1KOsmi skeletal muscle shifted toward an oxidative muscle fibre type and, in parallel, increased myofibre size and reduced capillary numbers. Unexpectedly, myofibre bundle mitochondrial respiration was reduced, whereas contraction-induced lactate production was elevated in Brca1KOsmi muscle. Brca1KOsmi mice accumulated mitochondrial DNA mutations and exhibited an altered mitochondrial morphology characterized by distorted and enlarged mitochondria, and these were more susceptible to swelling. In summary, skeletal muscle-specific loss of Brca1 leads to a myopathy and mitochondriopathy characterized by reductions in skeletal muscle quality and a consequent kyphosis. Given the substantial impact of BRCA1 mutations on cancer development risk in humans, a parallel loss of BRCA1 function in patient skeletal muscle cells would potentially result in implications for human health.
    Keywords:  BRCA1; Mitochondria; Muscle physiology; Skeletal muscle; kyphosis; mtDNA damage
  33. Cell Cycle. 2018 Dec 17.
    Drake JC, Laker RC, Wilson RJ, Zhang M, Yan Z.
      Maintenance of mitochondrial quality is essential for skeletal muscle function and overall health. Exercise training elicits profound adaptations to mitochondria to improve mitochondrial quality in skeletal muscle. We have recently demonstrated that acute exercise promotes removal of damaged/dysfunctional mitochondria via mitophagy in skeletal muscle during recovery through the Ampk-Ulk1 signaling cascade. In this Extra View, we explore whether Pink1 is stabilized on mitochondria following exercise as the signal for mitophagy. We observed no discernable presence of Pink1 in isolated mitochondria from skeletal muscle at any time point following acute exercise, in contrast to clear evidence of stabilization of Pink1 on mitochondria in HeLa cells following treatment with the uncoupler carbonyl cyanide m-chlorophenyl hydrazone (CCCP). Taken together, we conclude that Pink1 is not involved in exercise-induced mitophagy in skeletal muscle.
  34. Metabolites. 2018 Dec 15. pii: E95. [Epub ahead of print]8(4):
    You R, Dai J, Zhang P, Barding GA, Raftery D.
      Cellular senescence displays a heterogeneous set of phenotypes linked to tumor suppression; however, after drug treatment, senescence may also be involved in stable or recurrent cancer. Metabolic changes during senescence can provide detailed information on cellular status and may also have implications for the development of effective treatment strategies. The metabolic response to Adriamycin (ADR) treatment, which causes senescence as well as cell death, was obtained with the aid of metabolic profiling and isotope tracing in two human breast cancer cell lines, MCF7 and MDA-MB-231. After 5 days of ADR treatment, more than 60% of remaining, intact cells entered into a senescent state, characterized by enlarged and flattened morphology and positive blue staining using SA-β-gal. Metabolic trajectory analysis showed that the two cell lines' responses were significantly different and were divided into two distinct stages. The metabolic shift from the first stage to the second was reflected by a partial recovery of the TCA cycle, as well as amino acid and lipid metabolisms. Isotope tracing analysis indicated that the higher level of glutamine metabolism helped maintain senescence. The results suggest that the dynamic changes during senescence indicate a multi-step process involving important metabolic pathways which might allow breast cancer cells to adapt to persistent ADR treatment, while the higher level of anapleurosis may be important for maintaining the senescent state. Ultimately, a better understanding of metabolic changes during senescence might provide targets for cancer therapy and tumor eradication.
    Keywords:  MDA-MB-231; gas chromatography–mass spectrometry (GC–MS); isotope tracing analysis; metabolomics; senescence MCF7
  35. Cancer Res. 2018 Dec 20. pii: canres.1062.2018. [Epub ahead of print]
    Erez A, Silberman A, Goldman O, Boukobza Assayag O, Jacob A, Limanovich S, Adler L, Lee JS, Keshet R, Sarver A, Frug J, Stettner N, Galai S, Persi E, Bahar Halpern K, Zaltsman-Amir Y, Pode-Shakked B, Eilam R, Anikster Y, Nagamani SC, Ulitsky I, Ruppin E.
      Downregulation of the urea cycle enzyme argininosuccinate synthase (ASS1) by either promoter methylation or by HIF1α is associated with increased metastasis and poor prognosis in multiple cancers. We have previously shown that in normoxic conditions, ASS1 downregulation facilitates cancer cell proliferation by increasing aspartate availability for pyrimidine synthesis by the enzyme complex CAD. Here we report that in hypoxia, ASS1 expression in cancerous cells is downregulated further by Hif1α-mediated induction of miR224-5p, making the cells more invasive and dependent on upstream substrates of ASS1 for survival. ASS1 was downregulated under acidic conditions, and ASS1-depleted cancer cells maintained a higher intracellular pH (pHi), depended less on extracellular glutamine, and displayed higher glutathione levels. Depletion of substrates of urea cycle enzymes in ASS1-deficient cancers decreased cancer cell survival. Thus, ASS1 levels in cancer are differentially regulated in various environmental conditions to metabolically benefit cancer progression. Understanding these alterations may help uncover specific context-dependent cancer vulnerabilities that may be targeted for therapeutic purposes.
  36. Histopathology. 2019 Jan;74(1): 31-59
    Trpkov K, Hes O.
      Renal tumours include a heterogeneous and diverse spectrum of neoplasms. Recent advances in this field have significantly improved our understanding of the morphological, immunohistochemical, molecular, epidemiological and clinical characteristics of renal tumours, which led to the new Vancouver classification of renal neoplasia and the new World Health Organization (WHO) classification of renal cell tumours. This review aims to summarise the new information and evidence on several new and emerging/provisional renal entities, which were mostly generated after the recent classification of renal neoplasia. We include in this review the following new and emerging/provisional renal entities: succinate dehydrogenase-deficient renal cell carcinoma, thyroid-like follicular carcinoma of the kidney, anaplastic lymphoma kinase rearrangement-associated renal cell carcinoma, renal cell carcinomas with prominent smooth muscle stroma, fumarate hydratase-deficient renal cell carcinoma, biphasic squamoid papillary renal cell carcinoma, eosinophilic solid and cystic renal cell carcinoma, atrophic kidney-like renal cell carcinoma, clear cell renal cell carcinoma with giant cells and emperipolesis, Warthin-like papillary renal cell carcinoma, and low-grade oncocytic renal tumour (CD117-negative; cytokeratin 7-positive). Some of these entities, such as succinate dehydrogenase-deficient renal cell carcinoma, have already been recognised as new entities in the WHO classification, and some have been recognised as provisional/emerging entities. However, we include in this review several additional entities that, on the basis of the published evidence, also warrant this designation. We hope that this review will ease the navigation through this complex and evolving field, and will inform and stimulate new studies and discussions.
    Keywords:   WHO ; emerging entity; kidney; new entity; renal cell carcinoma
  37. Biochemistry. 2018 Dec 20.
    Saleh L, Parker MJ, Weigele PR.
      A tight link exists between patterns of DNA methylation at carbon 5 of 2-deoxycytosine and differential gene expression in mammalian tissues. Indeed, aberrant DNA methylation results in various human diseases, including neurologic and immune disorders, and contributes to the initiation and progression of various cancers. Proper DNA methylation depends on the fidelity and control of the underlying mechanisms that write, maintain, and erase these epigenetic marks. In this perspective, we focus on key players in active demethylation: the ten-eleven translocation enzymes or TETs. These enzymes belong to the Fe2+/-ketoglutarate-dependent dioxygenase superfamily and iteratively oxidize 5-methylcytosine in DNA to produce 5-hydroxymethylcytosine, 5-formylcytosine, and 5-carboxycytosine. The latter three bases may convey additional layers of epigenetic information in addition to being intermediates in active demethylation. Despite the intense interest in understanding the physiological roles TETs play in active demethylation and cell regulation, less has been done, in comparison, to illuminate details of the chemistry and factors involved in regulating the three-step oxidation mechanism. Herein, we examine what is known about the biochemical features of TETs and explore questions whose answers will lead to a more detailed understanding of the in vivo modus operandi of these enzymes. We also summarize the membership and evolutionary history of the TET/JBP family and highlight the prokaryotic homologs as a reservoir of potentially diverse functionalities awaiting discovery. Finally, we spotlight sequencing methods that utilize TET for mapping 5mC and its oxidation products in genomic DNA and comment on possible improvements to these approaches.
  38. J Cell Biol. 2018 Dec 21. pii: jcb.201807154. [Epub ahead of print]
    Locke MN, Thorner J.
      Target of rapamycin complex-2 (TORC2), a conserved protein kinase complex, is an indispensable regulator of plasma membrane homeostasis. In budding yeast (Saccharomyces cerevisiae), the essential downstream effector of TORC2 is protein kinase Ypk1 and its paralog Ypk2. Muk1, a Rab5-specific guanine nucleotide exchange factor (GEF), was identified in our prior global screen for candidate Ypk1 targets. We confirm here that Muk1 is a substrate of Ypk1 and demonstrate that Ypk1-mediated phosphorylation stimulates Muk1 function in vivo. Strikingly, yeast lacking its two Rab5 GEFs (Muk1 and Vps9) or its three Rab5 paralogs (Vps21/Ypt51, Ypt52, and Ypt53) or overexpressing Msb3, a Rab5-directed GTPase-activating protein, all exhibit pronounced reduction in TORC2-mediated phosphorylation and activation of Ypk1. Vps21 coimmunoprecipitates with TORC2, and immuno-enriched TORC2 is less active in vitro in the absence of Rab5 GTPases. Thus, TORC2-dependent and Ypk1-mediated activation of Muk1 provides a control circuit for positive (self-reinforcing) up-regulation to sustain TORC2-Ypk1 signaling.
  39. Traffic. 2018 Dec 20.
    Ganesan S, Tavassoli M, Sosa Ponce ML, Shabits B, Mahadeo M, Prenner EJ, Terebiznik MR, Zaremberg V.
      Diacylglycerol (DAG) is a key signaling lipid and intermediate in lipid metabolism. Our knowledge of DAG distribution and dynamics in cell membranes is limited. Using live-cell fluorescence microscopy we investigated the localization of yeast cytosolic-facing pools of DAG in response to conditions where lipid homeostasis and DAG levels were known to be altered. Two main pools were monitored over time using DAG sensors. One pool was associated with vacuolar membranes and the other localized to sites of polarized growth. Dynamic changes in DAG distribution were observed during resumption of growth from stationary phase, when DAG is used to support phospholipid synthesis for membrane proliferation. Vacuolar membranes experienced constant morphological changes displaying DAG enriched microdomains co-existing with liquid-disordered areas demarcated by Vph1. Formation of these domains was dependent on TAG lipolysis. DAG domains and puncta were closely connected to lipid droplets. Lack of conversion of DAG to phosphatidate in growth conditions dependent on TAG mobilization, led to the accumulation of DAG in a vacuolar associated compartment, impacting the polarized distribution of DAG at budding sites. DAG polarization was also regulated by phosphatidylserine synthesis/traffic and sphingolipid synthesis in the Golgi. This article is protected by copyright. All rights reserved.
    Keywords:  Diacylglycerol; Saccharomyces cerevisiae; glycerolipid; metabolism; microdomain; phosphatidic acid; phosphatidylserine; polarized growth; sphingolipid; vacuole