Complex I-complex II ratio strongly differs in various organs of Arabidopsis thaliana.

PubMed

Peters, Katrin; Niessen, Markus; Peterhänsel, Christoph; Späth, Bettina; Hölzle, Angela; Binder, Stefan; Marchfelder, Anita; Braun, Hans-Peter

2012-06-01

In most studies, amounts of protein complexes of the oxidative phosphorylation (OXPHOS) system in different organs or tissues are quantified on the basis of isolated mitochondrial fractions. However, yield of mitochondrial isolations might differ with respect to tissue type due to varying efficiencies of cell disruption during organelle isolation procedures or due to tissue-specific properties of organelles. Here we report an immunological investigation on the ratio of the OXPHOS complexes in different tissues of Arabidopsis thaliana which is based on total protein fractions isolated from five Arabidopsis organs (leaves, stems, flowers, roots and seeds) and from callus. Antibodies were generated against one surface exposed subunit of each of the five OXPHOS complexes and used for systematic immunoblotting experiments. Amounts of all complexes are highest in flowers (likewise with respect to organ fresh weight or total protein content of the flower fraction). Relative amounts of protein complexes in all other fractions were determined with respect to their amounts in flowers. Our investigation reveals high relative amounts of complex I in green organs (leaves and stems) but much lower amounts in non-green organs (roots, callus tissue). In contrast, complex II only is represented by low relative amounts in green organs but by significantly higher amounts in non-green organs, especially in seeds. In fact, the complex I-complex II ratio differs by factor 37 between callus and leaf, indicating drastic differences in electron entry into the respiratory chain in these two fractions. Variation in amounts concerning complexes III, IV and V was less pronounced in different Arabidopsis tissues (quantification of complex V in leaves was not meaningful due to a cross-reaction of the antibody with the chloroplast form of this enzyme). Analyses were complemented by in gel activity measurements for the protein complexes of the OXPHOS system and comparative 2D blue native/SDS PAGE analyses using isolated mitochondria. We suggest that complex I has an especially important role in the context of photosynthesis which might be due to its indirect involvement in photorespiration and its numerous enzymatic side activities in plants.

The mitonuclear compatibility hypothesis of sexual selection

PubMed Central

Hill, Geoffrey E.; Johnson, James D.

2013-01-01

Why females assess ornaments when choosing mates remains a central question in evolutionary biology. We hypothesize that the imperative for a choosing female to find a mate with nuclear oxidative phosphorylation (OXPHOS) genes that are compatible with her mitochondrial OXPHOS genes drives the evolution of ornaments. Indicator traits are proposed to signal the efficiency of OXPHOS function thus enabling females to select mates with nuclear genes that are compatible with maternal mitochondrial genes in the formation of OXPHOS complexes. Species-typical pattern of ornamentation is proposed to serve as a marker of mitochondrial type ensuring that females assess prospective mates with a shared mitochondrial background. The mitonuclear compatibility hypothesis predicts that the production of ornaments will be closely linked to OXPHOS pathways, and that sexual selection for compatible mates will be strongest when genes for nuclear components of OXPHOS complexes are Z-linked. The implications of this hypothesis are that sexual selection may serve as a driver for the evolution of more efficient cellular respiration. PMID:23945683

Multi-omics Reveal Specific Targets of the RNA-Binding Protein Puf3p and Its Orchestration of Mitochondrial Biogenesis.

PubMed

Lapointe, Christopher P; Stefely, Jonathan A; Jochem, Adam; Hutchins, Paul D; Wilson, Gary M; Kwiecien, Nicholas W; Coon, Joshua J; Wickens, Marvin; Pagliarini, David J

2018-01-24

Coenzyme Q (CoQ) is a redox-active lipid required for mitochondrial oxidative phosphorylation (OxPhos). How CoQ biosynthesis is coordinated with the biogenesis of OxPhos protein complexes is unclear. Here, we show that the Saccharomyces cerevisiae RNA-binding protein (RBP) Puf3p regulates CoQ biosynthesis. To establish the mechanism for this regulation, we employed a multi-omic strategy to identify mRNAs that not only bind Puf3p but also are regulated by Puf3p in vivo. The CoQ biosynthesis enzyme Coq5p is a critical Puf3p target: Puf3p regulates the abundance of Coq5p and prevents its detrimental hyperaccumulation, thereby enabling efficient CoQ production. More broadly, Puf3p represses a specific set of proteins involved in mitochondrial protein import, translation, and OxPhos complex assembly (pathways essential to prime mitochondrial biogenesis). Our data reveal a mechanism for post-transcriptionally coordinating CoQ production with OxPhos biogenesis, and they demonstrate the power of multi-omics for defining genuine targets of RBPs. Copyright © 2017 Elsevier Inc. All rights reserved.

Characterization of oxidative phosphorylation enzymes in Euglena gracilis and its white mutant strain W(gm)ZOflL.

PubMed

Krnáčová, Katarína; Rýdlová, Ivana; Vinarčíková, Michaela; Krajčovič, Juraj; Vesteg, Matej; Horváth, Anton

2015-03-12

The enzymes involved in Euglena oxidative phosphorylation (OXPHOS) were characterized in this study. We have demonstrated that Euglena gracilis strain Z and its stable bleached non-photosynthetic mutant strain WgmZOflL both possess fully functional OXPHOS apparatus as well as pathways requiring terminal alternative oxidase(s) and alternative mitochondrial NADH-dehydrogenase(s). Light (or dark) and plastid (non)functionality seem to have little effect on oxygen consumption, the activities of the enzymes involved in OXPHOS and the action of respiration inhibitors in Euglena. This study also demonstrates biochemical properties of complex III (cytochrome c reductase) in Euglena. Copyright © 2015 Federation of European Biochemical Societies. Published by Elsevier B.V. All rights reserved.

Mechanistic insights into selective killing of OXPHOS-dependent cancer cells by arctigenin.

PubMed

Brecht, Karin; Riebel, Virginie; Couttet, Philippe; Paech, Franziska; Wolf, Armin; Chibout, Salah-Dine; Pognan, Francois; Krähenbühl, Stephan; Uteng, Marianne

2017-04-01

Arctigenin has previously been identified as a potential anti-tumor treatment for advanced pancreatic cancer. However, the mechanism of how arctigenin kills cancer cells is not fully understood. In the present work we studied the mechanism of toxicity by arctigenin in the human pancreatic cell line, Panc-1, with special emphasis on the mitochondria. A comparison of Panc-1 cells cultured in glucose versus galactose medium was applied, allowing assessments of effects in glycolytic versus oxidative phosphorylation (OXPHOS)-dependent Panc-1 cells. For control purposes, the mitochondrial toxic response to treatment with arctigenin was compared to the anti-cancer drug, sorafenib, which is a tyrosine kinase inhibitor known for mitochondrial toxic off-target effects (Will et al., 2008). In both Panc-1 OXPHOS-dependent and glycolytic cells, arctigenin dissipated the mitochondrial membrane potential, which was demonstrated to be due to inhibition of the mitochondrial complexes II and IV. However, arctigenin selectively killed only the OXPHOS-dependent Panc-1 cells. This selective killing of OXPHOS-dependent Panc-1 cells was accompanied by generation of ER stress, mitochondrial membrane permeabilization and caspase activation leading to apoptosis and aponecrosis. Copyright © 2016 Elsevier Ltd. All rights reserved.

Anti-cancer analogues ME-143 and ME-344 exert toxicity by directly inhibiting mitochondrial NADH: ubiquinone oxidoreductase (Complex I).

PubMed

Lim, Sze Chern; Carey, Kirstyn T; McKenzie, Matthew

2015-01-01

Isoflavonoids have been shown to inhibit tumor proliferation and metastasis by activating cell death pathways. As such, they have been widely studied as potential therapies for cancer prevention. The second generation synthetic isoflavan analogues ME-143 and ME-344 also exhibit anti-cancer effects, however their specific molecular targets have not been completely defined. To identify these targets, we examined the effects of ME-143 and ME-344 on cellular metabolism and found that they are potent inhibitors of mitochondrial oxidative phosphorylation (OXPHOS) complex I (NADH: ubiquinone oxidoreductase) activity. In isolated HEK293T mitochondria, ME-143 and ME-344 reduced complex I activity to 14.3% and 28.6% of control values respectively. In addition to the inhibition of complex I, ME-344 also significantly inhibited mitochondrial complex III (ubiquinol: ferricytochrome-c oxidoreductase) activity by 10.8%. This inhibition of complex I activity (and to a lesser extent complex III activity) was associated with a reduction in mitochondrial oxygen consumption. In permeabilized HEK293T cells, ME-143 and ME-344 significantly reduced the maximum ADP-stimulated respiration rate to 62.3% and 70.0% of control levels respectively in the presence of complex I-linked substrates. Conversely, complex II-linked respiration was unaffected by either drug. We also observed that the inhibition of complex I-linked respiration caused the dissipation of the mitochondrial membrane potential (ΔΨm). Blue native (BN-PAGE) analysis revealed that prolonged loss of ΔΨm results in the destabilization of the native OXPHOS complexes. In particular, treatment of 143B osteosarcoma, HeLa and HEK293T human embryonic kidney cells with ME-344 for 4 h resulted in reduced steady-state levels of mature complex I. Degradation of the complex I subunit NDUFA9, as well as the complex IV (ferrocytochrome c: oxygen oxidoreductase) subunit COXIV, was also evident. The identification of OXPHOS complex I as a target of ME-143 and ME-344 advances our understanding of how these drugs induce cell death by disrupting mitochondrial metabolism, and will direct future work to maximize the anti-cancer capacity of these and other isoflavone-based compounds.

Microscale oxygraphy reveals OXPHOS impairment in MRC mutant cells

PubMed Central

Invernizzi, F.; D'Amato, I.; Jensen, P.B.; Ravaglia, S.; Zeviani, M.; Tiranti, V.

2012-01-01

Given the complexity of the respiratory chain structure, assembly and regulation, the diagnostic workout for the identification of defects of oxidative phosphorylation (OXPHOS) is a major challenge. Spectrophotometric assays, that measure the activity of individual respiratory complexes in tissue and cell homogenates or isolated mitochondria, are highly specific, but their utilization is limited by the availability of sufficient biological material and intrinsic sensitivity. A further limitation is tissue specificity, which usually determines attenuation, or disappearance, in cultured fibroblasts, of defects detected in muscle or liver. We used numerous fibroblast cell lines derived from patients with OXPHOS deficiencies to set up experimental protocols required for the direct readout of cellular respiration using the Seahorse XF96 apparatus, which measures oxygen consumption rate (OCR) and extra-cellular acidification rate (ECAR) in 96 well plates. Results demonstrate that first level screening based on microscale oxygraphy is more sensitive, cheaper and rapid than spectrophotometry for the biochemical evaluation of cells from patients with suspected mitochondrial disorders. PMID:22310368

Small structural changes on a hydroquinone scaffold determine the complex I inhibition or uncoupling of tumoral oxidative phosphorylation

DOE Office of Scientific and Technical Information (OSTI.GOV)

Urra, Félix A., E-mail: felix.urra@qf.uchile.cl; Córdova-Delgado, Miguel; Lapier, Michel

2016-01-15

Mitochondria participate in several distinctiveness of cancer cell, being a promising target for the design of anti-cancer compounds. Previously, we described that ortho-carbonyl hydroquinone scaffold 14 inhibits the complex I-dependent respiration with selective anti-proliferative effect on mouse mammary adenocarcinoma TA3/Ha cancer cells; however, the structural requirements of this hydroquinone scaffold to affect the oxidative phosphorylation (OXPHOS) of cancer cells have not been studied in detail. Here, we characterize the mitochondrial metabolism of TA3/Ha cancer cells, which exhibit a high oxidative metabolism, and evaluate the effect of small structural changes of the hydroquinone scaffold 14 on the respiration of this cellmore » line. Our results indicate that these structural changes modify the effect on OXPHOS, obtaining compounds with three alternative actions: inhibitors of complex I-dependent respiration, uncoupler of OXPHOS and compounds with both actions. To confirm this, the effect of a bicyclic hydroquinone (9) was evaluated in isolated mitochondria. Hydroquinone 9 increased mitochondrial respiration in state 4o without effects on the ADP-stimulated respiration (state 3{sub ADP}), decreasing the complexes I and II-dependent respiratory control ratio. The effect on mitochondrial respiration was reversed by 6-ketocholestanol addition, indicating that this hydroquinone is a protonophoric uncoupling agent. In intact TA3/Ha cells, hydroquinone 9 caused mitochondrial depolarization, decreasing intracellular ATP and NAD(P)H levels and GSH/GSSG ratio, and slightly increasing the ROS levels. Moreover, it exhibited selective NAD(P)H availability-dependent anti-proliferative effect on cancer cells. Therefore, our results indicate that the ortho-carbonyl hydroquinone scaffold offers the possibility to design compounds with specific actions on OXPHOS of cancer cells. - Highlights: • Small changes on a hydroquinone scaffold modify the action on OXPHOS of cancer cells. • Complex I Inhibitors, uncoupler of OXPHOS and agents with dual action are described. • Cpd. 9 is an uncoupler agent of OXPHOS with selective anti-proliferative effects. • Useful information to design agents with a selective mechanism on OXPHOS is provided.« less

Common Variants within Oxidative Phosphorylation Genes Influence Risk of Ischemic Stroke and Intracerebral Hemorrhage

PubMed Central

Anderson, Christopher D.; Biffi, Alessandro; Nalls, Michael A.; Devan, William J.; Schwab, Kristin; Ayres, Alison M.; Valant, Valerie; Ross, Owen A.; Rost, Natalia S.; Saxena, Richa; Viswanathan, Anand; Worrall, Bradford B.; Brott, Thomas G.; Goldstein, Joshua N.; Brown, Devin; Broderick, Joseph P.; Norrving, Bo; Greenberg, Steven M.; Silliman, Scott L.; Hansen, Björn M.; Tirschwell, David L.; Lindgren, Arne; Slowik, Agnieszka; Schmidt, Reinhold; Selim, Magdy; Roquer, Jaume; Montaner, Joan; Singleton, Andrew B.; Kidwell, Chelsea S.; Woo, Daniel; Furie, Karen L.; Meschia, James F.; Rosand, Jonathan

2013-01-01

Background and Purpose Prior studies demonstrated association between mitochondrial DNA variants and ischemic stroke (IS). We investigated whether variants within a larger set of oxidative phosphorylation (OXPHOS) genes encoded by both autosomal and mitochondrial DNA were associated with risk of IS and, based on our results, extended our investigation to intracerebral hemorrhage (ICH). Methods This association study employed a discovery cohort of 1643 individuals, a validation cohort of 2432 individuals for IS, and an extension cohort of 1476 individuals for ICH. Gene-set enrichment analysis (GSEA) was performed on all structural OXPHOS genes, as well as genes contributing to individual respiratory complexes. Gene-sets passing GSEA were tested by constructing genetic scores using common variants residing within each gene. Associations between each variant and IS that emerged in the discovery cohort were examined in validation and extension cohorts. Results IS was associated with genetic risk scores in OXPHOS as a whole (odds ratio (OR)=1.17, p=0.008) and Complex I (OR=1.06, p=0.050). Among IS subtypes, small vessel (SV) stroke showed association with OXPHOS (OR=1.16, p=0.007), Complex I (OR=1.13, p=0.027) and Complex IV (OR 1.14, p=0.018). To further explore this SV association, we extended our analysis to ICH, revealing association between deep hemispheric ICH and Complex IV (OR=1.08, p=0.008). Conclusions This pathway analysis demonstrates association between common genetic variants within OXPHOS genes and stroke. The associations for SV stroke and deep ICH suggest that genetic variation in OXPHOS influences small vessel pathobiology. Further studies are needed to identify culprit genetic variants and assess their functional consequences. PMID:23362085

Nutrient sensing by the mitochondrial transcription machinery dictates oxidative phosphorylation

PubMed Central

Liu, Lijun; Nam, Minwoo; Fan, Wei; Akie, Thomas E.; Hoaglin, David C.; Gao, Guangping; Keaney, John F.; Cooper, Marcus P.

2014-01-01

Sirtuin 3 (SIRT3), an important regulator of energy metabolism and lipid oxidation, is induced in fasted liver mitochondria and implicated in metabolic syndrome. In fasted liver, SIRT3-mediated increases in substrate flux depend on oxidative phosphorylation (OXPHOS), but precisely how OXPHOS meets the challenge of increased substrate oxidation in fasted liver remains unclear. Here, we show that liver mitochondria in fasting mice adapt to the demand of increased substrate oxidation by increasing their OXPHOS efficiency. In response to cAMP signaling, SIRT3 deacetylated and activated leucine-rich protein 130 (LRP130; official symbol, LRPPRC), promoting a mitochondrial transcriptional program that enhanced hepatic OXPHOS. Using mass spectrometry, we identified SIRT3-regulated lysine residues in LRP130 that generated a lysine-to-arginine (KR) mutant of LRP130 that mimics deacetylated protein. Compared with wild-type LRP130 protein, expression of the KR mutant increased mitochondrial transcription and OXPHOS in vitro. Indeed, even when SIRT3 activity was abolished, activation of mitochondrial transcription and OXPHOS by the KR mutant remained robust, further highlighting the contribution of LRP130 deacetylation to increased OXPHOS in fasted liver. These data establish a link between nutrient sensing and mitochondrial transcription that regulates OXPHOS in fasted liver and may explain how fasted liver adapts to increased substrate oxidation. PMID:24430182

Short Communication: Transplacental Nucleoside Analogue Exposure and Mitochondrial Parameters in HIV-Uninfected Children

PubMed Central

Brogly, Susan B.; DiMauro, Salvatore; Van Dyke, Russell B.; Williams, Paige L.; Naini, Ali; Libutti, Daniel E.; Choi, Julia; Chung, Michelle

2011-01-01

Abstract Transplacental nucleoside analogue exposure can affect infant mitochondrial DNA (mtDNA). We evaluated mitochondria in peripheral blood mononuclear cells of children with and without clinical signs of mitochondrial dysfunction (MD) and antiretroviral (ARV) exposure. We previously identified 20 children with signs of MD (cases) among 1037 HIV-uninfected children born to HIV-infected women. We measured mtDNA copies/cell and oxidative phosphorylation (OXPHOS) NADH dehydrogenase (complex I) and cytochrome c oxidase (complex IV) protein levels and enzyme activities, determined mtDNA haplogroups and deletions in 18 of 20 cases with stored samples and in sex- and age-matched HIV-uninfected children, both ARV exposed and unexposed, (1) within 18 months of birth and (2) at the time of presentation of signs of MD. In specimens drawn within 18 months of birth, mtDNA levels were higher and OXPHOS protein levels and enzyme activities lower in cases than controls. In contrast, at the time of MD presentation, cases and ARV-exposed controls had lower mtDNA levels, 214 and 215 copies/cell, respectively, than ARV-unexposed controls, 254 copies/cell. OXPHOS protein levels and enzyme activities were lower in cases than exposed controls, and higher in cases than unexposed controls, except for complex IV activity, which was higher in cases. Haplotype H was less frequent among cases (6%) than controls (31%). No deletions were found. The long-term significance of these small but potentially important alterations should continue to be studied as these children enter adolescence and adulthood. PMID:21142587

Anti-cancer analogues ME-143 and ME-344 exert toxicity by directly inhibiting mitochondrial NADH: ubiquinone oxidoreductase (Complex I)

PubMed Central

Lim, Sze Chern; Carey, Kirstyn T; McKenzie, Matthew

2015-01-01

Isoflavonoids have been shown to inhibit tumor proliferation and metastasis by activating cell death pathways. As such, they have been widely studied as potential therapies for cancer prevention. The second generation synthetic isoflavan analogues ME-143 and ME-344 also exhibit anti-cancer effects, however their specific molecular targets have not been completely defined. To identify these targets, we examined the effects of ME-143 and ME-344 on cellular metabolism and found that they are potent inhibitors of mitochondrial oxidative phosphorylation (OXPHOS) complex I (NADH: ubiquinone oxidoreductase) activity. In isolated HEK293T mitochondria, ME-143 and ME-344 reduced complex I activity to 14.3% and 28.6% of control values respectively. In addition to the inhibition of complex I, ME-344 also significantly inhibited mitochondrial complex III (ubiquinol: ferricytochrome-c oxidoreductase) activity by 10.8%. This inhibition of complex I activity (and to a lesser extent complex III activity) was associated with a reduction in mitochondrial oxygen consumption. In permeabilized HEK293T cells, ME-143 and ME-344 significantly reduced the maximum ADP-stimulated respiration rate to 62.3% and 70.0% of control levels respectively in the presence of complex I-linked substrates. Conversely, complex II-linked respiration was unaffected by either drug. We also observed that the inhibition of complex I-linked respiration caused the dissipation of the mitochondrial membrane potential (ΔΨm). Blue native (BN-PAGE) analysis revealed that prolonged loss of ΔΨm results in the destabilization of the native OXPHOS complexes. In particular, treatment of 143B osteosarcoma, HeLa and HEK293T human embryonic kidney cells with ME-344 for 4 h resulted in reduced steady-state levels of mature complex I. Degradation of the complex I subunit NDUFA9, as well as the complex IV (ferrocytochrome c: oxygen oxidoreductase) subunit COXIV, was also evident. The identification of OXPHOS complex I as a target of ME-143 and ME-344 advances our understanding of how these drugs induce cell death by disrupting mitochondrial metabolism, and will direct future work to maximize the anti-cancer capacity of these and other isoflavone-based compounds. PMID:25973307

Metabolic Remodeling Precedes Mitochondrial Outer Membrane Permeabilization in Human Glioma Xenograft Cells

PubMed Central

Ponnala, Shivani; Chetty, Chandramu; Veeravalli, Krishna Kumar; Dinh, Dzung H.; Klopfenstein, Jeffrey D.; Rao, Jasti S.

2011-01-01

Glioma cancer cells adapt to changing microenvironment and shift from mitochondrial oxidative phosphorylation to aerobic glycolysis for their metabolic needs irrespective of oxygen availability. In the present study, we show that silencing MMP-9 in combination with uPAR/cathepsin B switch glioma cells glycolytic metabolism to oxidative phosphorylation (OXPHOS) and generate reactive oxygen species (ROS) to predispose glioma cells to mitochondrial outer membrane permeabilization. shRNA for MMP-9 and uPAR (pMU) as well as shRNA for MMP-9 and cathepsin B (pMC) activated complexes of mitochondria involved in OXPHOS and inhibited glycolytic hexokinase expression. The decreased interaction of hexokinase 2 with mitochondria in the treated cells indicated the inhibition of glycolysis activation. Overexpression of Akt reversed the pMU- and pMC-mediated glycolysis to OXPHOS switch. OXPHOS un-coupler oligomycin A altered the expression levels of the Bcl-2 family of proteins; treatment with pMU or pMC reversed this effect and induced mitochondrial outer membrane permeabilization. In addition, our results show changes in mitochondrial pore transition to release cytochrome c due to change in the VDAC-Bcl-XL and BAX-BAK interaction with pMU and pMC treatments. Taken together, our results suggest that pMU and pMC treatments switch glioma cells from glycolytic to OXPHOS pathway through an inhibitory effect on Akt, ROS induction, and an increase of cytosolic cytochrome c accumulation. These results demonstrate the potential of pMU and pMC as therapeutic candidates for treatment of glioma. PMID:22076676

Metabolic remodeling precedes mitochondrial outer membrane permeabilization in human glioma xenograft cells.

PubMed

Ponnala, Shivani; Chetty, Chandramu; Veeravalli, Krishna Kumar; Dinh, Dzung H; Klopfenstein, Jeffrey D; Rao, Jasti S

2012-02-01

Glioma cancer cells adapt to changing microenvironment and shift from mitochondrial oxidative phosphorylation to aerobic glycolysis for their metabolic needs irrespective of oxygen availability. In the present study, we show that silencing MMP-9 in combination with uPAR/cathepsin B switch the glycolytic metabolism of glioma cells to oxidative phosphorylation (OXPHOS) and generate reactive oxygen species (ROS) to predispose glioma cells to mitochondrial outer membrane permeabilization. shRNA for MMP-9 and uPAR (pMU) as well as shRNA for MMP-9 and cathepsin B (pMC) activated complexes of mitochondria involved in OXPHOS and inhibited glycolytic hexokinase expression. The decreased interaction of hexokinase 2 with mitochondria in the treated cells indicated the inhibition of glycolysis activation. Overexpression of Akt reversed the pMU- and pMC-mediated OXPHOS to glycolysis switch. The OXPHOS un-coupler oligomycin A altered the expression levels of the Bcl-2 family of proteins; treatment with pMU or pMC reversed this effect and induced mitochondrial outer membrane permeabilization. In addition, our results show changes in mitochondrial pore transition to release cytochrome c due to changes in the VDAC-Bcl-XL and BAX-BAK interaction with pMU and pMC treatments. Taken together, our results suggest that pMU and pMC treatments switch glioma cells from the glycolytic to the OXPHOS pathway through an inhibitory effect on Akt, ROS induction and an increase of cytosolic cytochrome c accumulation. These results demonstrate the potential of pMU and pMC as therapeutic candidates for the treatment of glioma.

Faster and stronger manifestation of mitochondrial diseases in skeletal muscle than in heart related to cytosolic inorganic phosphate (Pi) accumulation

PubMed Central

2016-01-01

A model of the cell bioenergetic system was used to compare the effect of oxidative phosphorylation (OXPHOS) deficiencies in a broad range of moderate ATP demand in skeletal muscle and heart. Computer simulations revealed that kinetic properties of the system are similar in both cases despite the much higher mitochondria content and “basic” OXPHOS activity in heart than in skeletal muscle, because of a much higher each-step activation (ESA) of OXPHOS in skeletal muscle than in heart. Large OXPHOS deficiencies lead in both tissues to a significant decrease in oxygen consumption (V̇o2) and phosphocreatine (PCr) and increase in cytosolic ADP, Pi, and H+. The main difference between skeletal muscle and heart is a much higher cytosolic Pi concentration in healthy tissue and much higher cytosolic Pi accumulation (level) at low OXPHOS activities in the former, caused by a higher PCr level in healthy tissue (and higher total phosphate pool) and smaller Pi redistribution between cytosol and mitochondria at OXPHOS deficiency. This difference does not depend on ATP demand in a broad range. A much greater Pi increase and PCr decrease during rest-to-moderate work transition in skeletal muscle at OXPHOS deficiencies than at normal OXPHOS activity significantly slows down the V̇o2 on-kinetics. Because high cytosolic Pi concentrations cause fatigue in skeletal muscle and can compromise force generation in skeletal muscle and heart, this system property can contribute to the faster and stronger manifestation of mitochondrial diseases in skeletal muscle than in heart. Shortly, skeletal muscle with large OXPHOS deficiencies becomes fatigued already during low/moderate exercise. PMID:27283913

Faster and stronger manifestation of mitochondrial diseases in skeletal muscle than in heart related to cytosolic inorganic phosphate (Pi) accumulation.

PubMed

Korzeniewski, Bernard

2016-08-01

A model of the cell bioenergetic system was used to compare the effect of oxidative phosphorylation (OXPHOS) deficiencies in a broad range of moderate ATP demand in skeletal muscle and heart. Computer simulations revealed that kinetic properties of the system are similar in both cases despite the much higher mitochondria content and "basic" OXPHOS activity in heart than in skeletal muscle, because of a much higher each-step activation (ESA) of OXPHOS in skeletal muscle than in heart. Large OXPHOS deficiencies lead in both tissues to a significant decrease in oxygen consumption (V̇o2) and phosphocreatine (PCr) and increase in cytosolic ADP, Pi, and H(+) The main difference between skeletal muscle and heart is a much higher cytosolic Pi concentration in healthy tissue and much higher cytosolic Pi accumulation (level) at low OXPHOS activities in the former, caused by a higher PCr level in healthy tissue (and higher total phosphate pool) and smaller Pi redistribution between cytosol and mitochondria at OXPHOS deficiency. This difference does not depend on ATP demand in a broad range. A much greater Pi increase and PCr decrease during rest-to-moderate work transition in skeletal muscle at OXPHOS deficiencies than at normal OXPHOS activity significantly slows down the V̇o2 on-kinetics. Because high cytosolic Pi concentrations cause fatigue in skeletal muscle and can compromise force generation in skeletal muscle and heart, this system property can contribute to the faster and stronger manifestation of mitochondrial diseases in skeletal muscle than in heart. Shortly, skeletal muscle with large OXPHOS deficiencies becomes fatigued already during low/moderate exercise. Copyright © 2016 the American Physiological Society.

An inhibitor of oxidative phosphorylation exploits cancer vulnerability.

PubMed

Molina, Jennifer R; Sun, Yuting; Protopopova, Marina; Gera, Sonal; Bandi, Madhavi; Bristow, Christopher; McAfoos, Timothy; Morlacchi, Pietro; Ackroyd, Jeffrey; Agip, Ahmed-Noor A; Al-Atrash, Gheath; Asara, John; Bardenhagen, Jennifer; Carrillo, Caroline C; Carroll, Christopher; Chang, Edward; Ciurea, Stefan; Cross, Jason B; Czako, Barbara; Deem, Angela; Daver, Naval; de Groot, John Frederick; Dong, Jian-Wen; Feng, Ningping; Gao, Guang; Gay, Jason; Do, Mary Geck; Greer, Jennifer; Giuliani, Virginia; Han, Jing; Han, Lina; Henry, Verlene K; Hirst, Judy; Huang, Sha; Jiang, Yongying; Kang, Zhijun; Khor, Tin; Konoplev, Sergej; Lin, Yu-Hsi; Liu, Gang; Lodi, Alessia; Lofton, Timothy; Ma, Helen; Mahendra, Mikhila; Matre, Polina; Mullinax, Robert; Peoples, Michael; Petrocchi, Alessia; Rodriguez-Canale, Jaime; Serreli, Riccardo; Shi, Thomas; Smith, Melinda; Tabe, Yoko; Theroff, Jay; Tiziani, Stefano; Xu, Quanyun; Zhang, Qi; Muller, Florian; DePinho, Ronald A; Toniatti, Carlo; Draetta, Giulio F; Heffernan, Timothy P; Konopleva, Marina; Jones, Philip; Di Francesco, M Emilia; Marszalek, Joseph R

2018-06-11

Metabolic reprograming is an emerging hallmark of tumor biology and an actively pursued opportunity in discovery of oncology drugs. Extensive efforts have focused on therapeutic targeting of glycolysis, whereas drugging mitochondrial oxidative phosphorylation (OXPHOS) has remained largely unexplored, partly owing to an incomplete understanding of tumor contexts in which OXPHOS is essential. Here, we report the discovery of IACS-010759, a clinical-grade small-molecule inhibitor of complex I of the mitochondrial electron transport chain. Treatment with IACS-010759 robustly inhibited proliferation and induced apoptosis in models of brain cancer and acute myeloid leukemia (AML) reliant on OXPHOS, likely owing to a combination of energy depletion and reduced aspartate production that leads to impaired nucleotide biosynthesis. In models of brain cancer and AML, tumor growth was potently inhibited in vivo following IACS-010759 treatment at well-tolerated doses. IACS-010759 is currently being evaluated in phase 1 clinical trials in relapsed/refractory AML and solid tumors.

Complex I Disorders: Causes, Mechanisms, and Development of Treatment Strategies at the Cellular Level

ERIC Educational Resources Information Center

Valsecchi, Federica; Koopman, Werner J. H.; Manjeri, Ganesh R.; Rodenburg, Richard J.; Smeitink, Jan A. M.; Willems, Peter H. G. M.

2010-01-01

Mitochondrial oxidative phosphorylation (OXPHOS) represents the final step in the conversion of nutrients into cellular energy. Genetic defects in the OXPHOS system have an incidence between 1:5,000 and 1:10,000 live births. Inherited isolated deficiency of the first complex (CI) of this system, a multisubunit assembly of 45 different proteins,…

Natural Product Screening Reveals Naphthoquinone Complex I Bypass Factors

PubMed Central

Mevers, Emily; Higgins, Kathleen W.; Fomina, Yevgenia; Zhang, Jianming; Mandinova, Anna; Newman, David; Shaw, Stanley Y.; Clardy, Jon; Mootha, Vamsi K.

2016-01-01

Deficiency of mitochondrial complex I is encountered in both rare and common diseases, but we have limited therapeutic options to treat this lesion to the oxidative phosphorylation system (OXPHOS). Idebenone and menadione are redox-active molecules capable of rescuing OXPHOS activity by engaging complex I-independent pathways of entry, often referred to as “complex I bypass.” In the present study, we created a cellular model of complex I deficiency by using CRISPR genome editing to knock out Ndufa9 in mouse myoblasts, and utilized this cell line to develop a high-throughput screening platform for novel complex I bypass factors. We screened a library of ~40,000 natural product extracts and performed bioassay-guided fractionation on a subset of the top scoring hits. We isolated four plant-derived 1,4-naphthoquinone complex I bypass factors with structural similarity to menadione: chimaphilin and 3-chloro-chimaphilin from Chimaphila umbellata and dehydro-α-lapachone and dehydroiso-α-lapachone from Stereospermum euphoroides. We also tested a small number of structurally related naphthoquinones from commercial sources and identified two additional compounds with complex I bypass activity: 2-methoxy-1,4-naphthoquinone and 2-methoxy-3-methyl-1,4,-naphthoquinone. The six novel complex I bypass factors reported here expand this class of molecules and will be useful as tool compounds for investigating complex I disease biology. PMID:27622560

Nutrient deprivation-related OXPHOS/glycolysis interconversion via HIF-1α/C-MYC pathway in U251 cells.

PubMed

Liu, Zhongjian; Sun, Yang; Tan, Shirui; Liu, Liang; Hu, Suqiong; Huo, Hongyu; Li, Meizhang; Cui, Qinghua; Yu, Min

2016-05-01

Although the Warburg effect is a dominant metabolic phenotype observed in cancers, the metabolic changes and adaptation occurring in tumors have been demonstrated to extend beyond the Warburg effect and thus considered a secondary effect to the transformation process of carcinogenesis, including nutritional deficiencies. However, the role of nutritional deficiencies in this metabolic reprogramming (e. g., oxidative phosphorylation (OXPHOS)/glycolysis interconversion) is not completely known yet. Here, we showed that under regular culture condition, the proliferation of U251 cells, but not other tumor cell lines, preferentially performed the Warburg effect and was remarkably inhibited by oxamic acid which can inhibit the activity of lactate dehydrogenase (LDH); whereas under serum starvation, glycolysis was depressed, tricarboxylic acid cycle (TCA) was enhanced, and the activity of OXPHOS was reinforced to maintain cellular ATP content in a high level, but interestingly, we observed a decreased expression of reactive oxygen species (ROS). Moreover, the upregulated activity of mitochondrial complex I was confirmed by Western blots and showed that the mitochondrial-related protein, NDUFA9, NDUFB8, ND1, and VDAC1 were remarkably increased after serum starved. Mechanistically, nutritional deficiencies could reduce hypoxia-inducible factor α (HIF-1α) protein expression to increase C-MYC protein level, which in turn increased nuclear respiratory factor 1 (NRF1) and mitochondrial transcription factor A (TFAM) transcription to enhance the activity of OXPHOS, suggesting that metabolic reprogramming by the changes of microenvironment during the carcinogenesis can provide some novel therapeutic clues to traditional cancer treatments.

Inborn oxidative phosphorylation defect as risk factor for propofol infusion syndrome.

PubMed

Vanlander, A V; Jorens, P G; Smet, J; De Paepe, B; Verbrugghe, W; Van den Eynden, G G; Meire, F; Pauwels, P; Van der Aa, N; Seneca, S; Lissens, W; Okun, J G; Van Coster, R

2012-04-01

Propofol is an anesthetic agent widely used for induction and maintenance of anesthesia, and sedation in children. Although generally considered as reliable and safe, administration of propofol can occasionally induce a potentially fatal complication known as propofol infusion syndrome (PRIS). Mitochondrial dysfunction has been implicated in the pathogenesis of PRIS. We report on an adult patient with Leber hereditary optic neuropathy (LHON) who developed PRIS. He was a carrier of the m.3460G>A mutation, one of the major three pathogenic point mutations associated with LHON. The propositus was blind and underwent propofol sedation after severe head injury. Five days after start of propofol infusion, the patient died. The activity of complex I of the oxidative phosphorylation (OXPHOS) system was severely deficient in skeletal muscle. Our observation indicates that fulminate PRIS can occur in an adult patient with an inborn OXPHOS defect and corroborates the hypothesis that PRIS is caused by inhibition of the OXPHOS system. © 2012 The Authors. Acta Anaesthesiologica Scandinavica © 2012 The Acta Anaesthesiologica Scandinavica Foundation.

Glycoprotein Ib activation by thrombin stimulates the energy metabolism in human platelets

PubMed Central

Corona de la Peña, Norma; Gutiérrez-Aguilar, Manuel; Hernández-Reséndiz, Ileana; Marín-Hernández, Álvaro

2017-01-01

Thrombin-induced platelet activation requires substantial amounts of ATP. However, the specific contribution of each ATP-generating pathway i.e., oxidative phosphorylation (OxPhos) versus glycolysis and the biochemical mechanisms involved in the thrombin-induced activation of energy metabolism remain unclear. Here we report an integral analysis on the role of both energy pathways in human platelets activated by several agonists, and the signal transducing mechanisms associated with such activation. We found that thrombin, Trap-6, arachidonic acid, collagen, A23187, epinephrine and ADP significantly increased glycolytic flux (3–38 times vs. non-activated platelets) whereas ristocetin was ineffective. OxPhos (33 times) and mitochondrial transmembrane potential (88%) were increased only by thrombin. OxPhos was the main source of ATP in thrombin-activated platelets, whereas in platelets activated by any of the other agonists, glycolysis was the principal ATP supplier. In order to establish the biochemical mechanisms involved in the thrombin-induced OxPhos activation in platelets, several signaling pathways associated with mitochondrial activation were analyzed. Wortmannin and LY294002 (PI3K/Akt pathway inhibitors), ristocetin and heparin (GPIb inhibitors) as well as resveratrol, ATP (calcium-release inhibitors) and PP1 (Tyr-phosphorylation inhibitor) prevented the thrombin-induced platelet activation. These results suggest that thrombin activates OxPhos and glycolysis through GPIb-dependent signaling involving PI3K and Akt activation, calcium mobilization and protein phosphorylation. PMID:28817667

Transcriptomic and proteomic landscape of mitochondrial dysfunction reveals secondary coenzyme Q deficiency in mammals

PubMed Central

Atanassov, Ilian; Kuznetsova, Irina; Hinze, Yvonne; Mourier, Arnaud; Filipovska, Aleksandra

2017-01-01

Dysfunction of the oxidative phosphorylation (OXPHOS) system is a major cause of human disease and the cellular consequences are highly complex. Here, we present comparative analyses of mitochondrial proteomes, cellular transcriptomes and targeted metabolomics of five knockout mouse strains deficient in essential factors required for mitochondrial DNA gene expression, leading to OXPHOS dysfunction. Moreover, we describe sequential protein changes during post-natal development and progressive OXPHOS dysfunction in time course analyses in control mice and a middle lifespan knockout, respectively. Very unexpectedly, we identify a new response pathway to OXPHOS dysfunction in which the intra-mitochondrial synthesis of coenzyme Q (ubiquinone, Q) and Q levels are profoundly decreased, pointing towards novel possibilities for therapy. Our extensive omics analyses provide a high-quality resource of altered gene expression patterns under severe OXPHOS deficiency comparing several mouse models, that will deepen our understanding, open avenues for research and provide an important reference for diagnosis and treatment. PMID:29132502

Oxidative Phosphorylation System in Gastric Carcinomas and Gastritis.

PubMed

Feichtinger, René G; Neureiter, Daniel; Skaria, Tom; Wessler, Silja; Cover, Timothy L; Mayr, Johannes A; Zimmermann, Franz A; Posselt, Gernot; Sperl, Wolfgang; Kofler, Barbara

2017-01-01

Switching of cellular energy production from oxidative phosphorylation (OXPHOS) by mitochondria to aerobic glycolysis occurs in many types of tumors. However, the significance of this switching for the development of gastric carcinoma and what connection it may have to Helicobacter pylori infection of the gut, a primary cause of gastric cancer, are poorly understood. Therefore, we investigated the expression of OXPHOS complexes in two types of human gastric carcinomas ("intestinal" and "diffuse"), bacterial gastritis with and without metaplasia, and chemically induced gastritis by using immunohistochemistry. Furthermore, we analyzed the effect of HP infection on several key mitochondrial proteins. Complex I expression was significantly reduced in intestinal type (but not diffuse) gastric carcinomas compared to adjacent control tissue, and the reduction was independent of HP infection. Significantly, higher complex I and complex II expression was present in large tumors. Furthermore, higher complex II and complex III protein levels were also obvious in grade 3 versus grade 2. No differences of OXPHOS complexes and markers of mitochondrial biogenesis were found between bacterially caused and chemically induced gastritis. Thus, intestinal gastric carcinomas, but not precancerous stages, are frequently characterized by loss of complex I, and this pathophysiology occurs independently of HP infection.

Oxidative Phosphorylation System in Gastric Carcinomas and Gastritis

PubMed Central

Skaria, Tom; Wessler, Silja; Cover, Timothy L.; Posselt, Gernot; Sperl, Wolfgang; Kofler, Barbara

2017-01-01

Switching of cellular energy production from oxidative phosphorylation (OXPHOS) by mitochondria to aerobic glycolysis occurs in many types of tumors. However, the significance of this switching for the development of gastric carcinoma and what connection it may have to Helicobacter pylori infection of the gut, a primary cause of gastric cancer, are poorly understood. Therefore, we investigated the expression of OXPHOS complexes in two types of human gastric carcinomas (“intestinal” and “diffuse”), bacterial gastritis with and without metaplasia, and chemically induced gastritis by using immunohistochemistry. Furthermore, we analyzed the effect of HP infection on several key mitochondrial proteins. Complex I expression was significantly reduced in intestinal type (but not diffuse) gastric carcinomas compared to adjacent control tissue, and the reduction was independent of HP infection. Significantly, higher complex I and complex II expression was present in large tumors. Furthermore, higher complex II and complex III protein levels were also obvious in grade 3 versus grade 2. No differences of OXPHOS complexes and markers of mitochondrial biogenesis were found between bacterially caused and chemically induced gastritis. Thus, intestinal gastric carcinomas, but not precancerous stages, are frequently characterized by loss of complex I, and this pathophysiology occurs independently of HP infection. PMID:28744336

Neonatal liver failure and Leigh syndrome possibly due to CoQ-responsive OXPHOS deficiency.

PubMed

Leshinsky-Silver, E; Levine, A; Nissenkorn, A; Barash, V; Perach, M; Buzhaker, E; Shahmurov, M; Polak-Charcon, S; Lev, D; Lerman-Sagie, T

2003-08-01

CoQ transfers electrons from complexes I and II of the mitochondrial respiratory chain to complex III. There are very few reports on human CoQ deficiency. The clinical presentation is usually characterized by: epilepsy, muscle weakness, ataxia, cerebellar atrophy, migraine, myogloblinuria and developmental delay. We describe a patient who presented with neonatal liver and pancreatic insufficiency, tyrosinemia and hyperammonemia and later developed sensorineural hearing loss and Leigh syndrome. Liver biopsy revealed markedly reduced complex I+III and II+III. Addition of CoQ to the liver homogenate restored the activities, suggesting CoQ depletion. Histological staining showed prominent bridging; septal fibrosis and widening of portal spaces with prominent mixed inflammatory infiltrate, associated with interface hepatitis, bile duct proliferation with numerous bile plugs. Electron microscopy revealed a large number of mitochondria, which were altered in shape and size, widened and disordered intercristal spaces. This may be the first case of Leigh syndrome with liver and pancreas insufficiency, possibly caused by CoQ responsive oxphos deficiency.

Physical Fitness and Mitochondrial Respiratory Capacity in Horse Skeletal Muscle

PubMed Central

Lemieux, Hélène; Mouithys-Mickalad, Ange; Serteyn, Didier

2012-01-01

Background Within the animal kingdom, horses are among the most powerful aerobic athletic mammals. Determination of muscle respiratory capacity and control improves our knowledge of mitochondrial physiology in horses and high aerobic performance in general. Methodology/Principal Findings We applied high-resolution respirometry and multiple substrate-uncoupler-inhibitor titration protocols to study mitochondrial physiology in small (1.0–2.5 mg) permeabilized muscle fibres sampled from triceps brachii of healthy horses. Oxidative phosphorylation (OXPHOS) capacity (pmol O2•s−1•mg−1 wet weight) with combined Complex I and II (CI+II) substrate supply (malate+glutamate+succinate) increased from 77±18 in overweight horses to 103±18, 122±15, and 129±12 in untrained, trained and competitive horses (N = 3, 8, 16, and 5, respectively). Similar to human muscle mitochondria, equine OXPHOS capacity was limited by the phosphorylation system to 0.85±0.10 (N = 32) of electron transfer capacity, independent of fitness level. In 15 trained horses, OXPHOS capacity increased from 119±12 to 134±37 when pyruvate was included in the CI+II substrate cocktail. Relative to this maximum OXPHOS capacity, Complex I (CI)-linked OXPHOS capacities were only 50% with glutamate+malate, 64% with pyruvate+malate, and 68% with pyruvate+malate+glutamate, and ∼78% with CII-linked succinate+rotenone. OXPHOS capacity with glutamate+malate increased with fitness relative to CI+II-supported ETS capacity from a flux control ratio of 0.38 to 0.40, 0.41 and 0.46 in overweight to competitive horses, whereas the CII/CI+II substrate control ratio remained constant at 0.70. Therefore, the apparent deficit of the CI- over CII-linked pathway capacity was reduced with physical fitness. Conclusions/Significance The scope of mitochondrial density-dependent OXPHOS capacity and the density-independent (qualitative) increase of CI-linked respiratory capacity with increased fitness open up new perspectives of integrative and comparative mitochondrial respiratory physiology. PMID:22529950

Lack of FTSH4 Protease Affects Protein Carbonylation, Mitochondrial Morphology, and Phospholipid Content in Mitochondria of Arabidopsis: New Insights into a Complex Interplay.

PubMed

Smakowska, Elwira; Skibior-Blaszczyk, Renata; Czarna, Malgorzata; Kolodziejczak, Marta; Kwasniak-Owczarek, Malgorzata; Parys, Katarzyna; Funk, Christiane; Janska, Hanna

2016-08-01

FTSH4 is one of the inner membrane-embedded ATP-dependent metalloproteases in mitochondria of Arabidopsis (Arabidopsis thaliana). In mutants impaired to express FTSH4, carbonylated proteins accumulated and leaf morphology was altered when grown under a short-day photoperiod, at 22°C, and a long-day photoperiod, at 30°C. To provide better insight into the function of FTSH4, we compared the mitochondrial proteomes and oxyproteomes of two ftsh4 mutants and wild-type plants grown under conditions inducing the phenotypic alterations. Numerous proteins from various submitochondrial compartments were observed to be carbonylated in the ftsh4 mutants, indicating a widespread oxidative stress. One of the reasons for the accumulation of carbonylated proteins in ftsh4 was the limited ATP-dependent proteolytic capacity of ftsh4 mitochondria, arising from insufficient ATP amount, probably as a result of an impaired oxidative phosphorylation (OXPHOS), especially complex V. In ftsh4, we further observed giant, spherical mitochondria coexisting among normal ones. Both effects, the increased number of abnormal mitochondria and the decreased stability/activity of the OXPHOS complexes, were probably caused by the lower amount of the mitochondrial membrane phospholipid cardiolipin. We postulate that the reduced cardiolipin content in ftsh4 mitochondria leads to perturbations within the OXPHOS complexes, generating more reactive oxygen species and less ATP, and to the deregulation of mitochondrial dynamics, causing in consequence the accumulation of oxidative damage. © 2016 American Society of Plant Biologists. All Rights Reserved.

Mitochondrial function in skeletal muscle of patients with protracted critical illness and ICU-acquired weakness.

PubMed

Jiroutková, Kateřina; Krajčová, Adéla; Ziak, Jakub; Fric, Michal; Waldauf, Petr; Džupa, Valér; Gojda, Jan; Němcova-Fürstová, Vlasta; Kovář, Jan; Elkalaf, Moustafa; Trnka, Jan; Duška, František

2015-12-24

Mitochondrial damage occurs in the acute phase of critical illness, followed by activation of mitochondrial biogenesis in survivors. It has been hypothesized that bioenergetics failure of skeletal muscle may contribute to the development of ICU-acquired weakness. The aim of the present study was to determine whether mitochondrial dysfunction persists until protracted phase of critical illness. In this single-centre controlled-cohort ex vivo proof-of-concept pilot study, we obtained vastus lateralis biopsies from ventilated patients with ICU-acquired weakness (n = 8) and from age and sex-matched metabolically healthy controls (n = 8). Mitochondrial functional indices were measured in cytosolic context by high-resolution respirometry in tissue homogenates, activities of respiratory complexes by spectrophotometry and individual functional capacities were correlated with concentrations of electron transport chain key subunits from respiratory complexes II, III, IV and V measured by western blot. The ability of aerobic ATP synthesis (OXPHOS) was reduced to ~54% in ICU patients (p<0.01), in correlation with the depletion of complexes III (~38% of control, p = 0.02) and IV (~26% of controls, p<0.01) and without signs of mitochondrial uncoupling. When mitochondrial functional indices were adjusted to citrate synthase activity, OXPHOS and the activity of complexes I and IV were not different, whilst the activities of complexes II and III were increased in ICU patients 3-fold (p<0.01) respectively 2-fold (p<0.01). Compared to healthy controls, in ICU patients we have demonstrated a ~50% reduction of the ability of skeletal muscle to synthetize ATP in mitochondria. We found a depletion of complex III and IV concentrations and relative increases in functional capacities of complex II and glycerol-3-phosphate dehydrogenase/complex III.

Human primitive brain displays negative mitochondrial-nuclear expression correlation of respiratory genes.

PubMed

Barshad, Gilad; Blumberg, Amit; Cohen, Tal; Mishmar, Dan

2018-06-14

Oxidative phosphorylation (OXPHOS), a fundamental energy source in all human tissues, requires interactions between mitochondrial (mtDNA)- and nuclear (nDNA)-encoded protein subunits. Although such interactions are fundamental to OXPHOS, bi-genomic coregulation is poorly understood. To address this question, we analyzed ∼8500 RNA-seq experiments from 48 human body sites. Despite well-known variation in mitochondrial activity, quantity, and morphology, we found overall positive mtDNA-nDNA OXPHOS genes' co-expression across human tissues. Nevertheless, negative mtDNA-nDNA gene expression correlation was identified in the hypothalamus, basal ganglia, and amygdala (subcortical brain regions, collectively termed the "primitive" brain). Single-cell RNA-seq analysis of mouse and human brains revealed that this phenomenon is evolutionarily conserved, and both are influenced by brain cell types (involving excitatory/inhibitory neurons and nonneuronal cells) and by their spatial brain location. As the "primitive" brain is highly oxidative, we hypothesized that such negative mtDNA-nDNA co-expression likely controls for the high mtDNA transcript levels, which enforce tight OXPHOS regulation, rather than rewiring toward glycolysis. Accordingly, we found "primitive" brain-specific up-regulation of lactate dehydrogenase B ( LDHB ), which associates with high OXPHOS activity, at the expense of LDHA , which promotes glycolysis. Analyses of co-expression, DNase-seq, and ChIP-seq experiments revealed candidate RNA-binding proteins and CEBPB as the best regulatory candidates to explain these phenomena. Finally, cross-tissue expression analysis unearthed tissue-dependent splice variants and OXPHOS subunit paralogs and allowed revising the list of canonical OXPHOS transcripts. Taken together, our analysis provides a comprehensive view of mito-nuclear gene co-expression across human tissues and provides overall insights into the bi-genomic regulation of mitochondrial activities. © 2018 Barshad et al.; Published by Cold Spring Harbor Laboratory Press.

Oxidative stress in myelin sheath: The other face of the extramitochondrial oxidative phosphorylation ability.

PubMed

Ravera, S; Bartolucci, M; Cuccarolo, P; Litamè, E; Illarcio, M; Calzia, D; Degan, P; Morelli, A; Panfoli, I

2015-01-01

Oxidative phosphorylation (OXPHOS) is not only the main source of ATP for the cell, but also a major source of reactive oxygen species (ROS), which lead to oxidative stress. At present, mitochondria are considered the organelles responsible for the OXPHOS, but in the last years we have demonstrated that it can also occur outside the mitochondrion. Myelin sheath is able to conduct an aerobic metabolism, producing ATP that we have hypothesized is transferred to the axon, to support its energetic demand. In this work, spectrophotometric, cytofluorimetric, and luminometric analyses were employed to investigate the oxidative stress production in isolated myelin, as far as its respiratory activity is concerned. We have evaluated the levels of malondialdehyde (MDA) and 4-hydroxynonenal (4-HNE), markers of lipid peroxidation, as well as of hydrogen peroxide (H2O2), marker of ROS production. To assess the presence of endogenous antioxidant systems, superoxide dismutase, catalase, and glutathione peroxidase activities were assayed. The effect of certain uncoupling or antioxidant molecules on oxidative stress in myelin was also investigated. We report that isolated myelin produces high levels of MDA, 4-HNE, and H2O2, likely through the pathway composed by Complex I-III-IV, but it also contains active superoxide dismutase, catalase, and glutathione peroxidase, as antioxidant defense. Uncoupling compounds or Complex I inhibitors increase oxidative stress, while antioxidant compounds limit ROS generation. Data may shed new light on the role of myelin sheath in physiology and pathology. In particular, it can be presumed that the axonal degeneration associated with myelin loss in demyelinating diseases is related to oxidative stress caused by impaired OXPHOS.

Distinct intracellular sAC-cAMP domains regulate ER Ca2+ signaling and OXPHOS function.

PubMed

Valsecchi, Federica; Konrad, Csaba; D'Aurelio, Marilena; Ramos-Espiritu, Lavoisier S; Stepanova, Anna; Burstein, Suzanne R; Galkin, Alexander; Magranè, Jordi; Starkov, Anatoly; Buck, Jochen; Levin, Lonny R; Manfredi, Giovanni

2017-11-01

cAMP regulates a wide variety of physiological functions in mammals. This single second messenger can regulate multiple, seemingly disparate functions within independently regulated cell compartments. We have previously identified one such compartment inside the matrix of the mitochondria, where soluble adenylyl cyclase (sAC) regulates oxidative phosphorylation (OXPHOS). We now show that sAC knockout fibroblasts have a defect in OXPHOS activity and attempt to compensate for this defect by increasing OXPHOS proteins. Importantly, sAC knockout cells also exhibit decreased probability of endoplasmic reticulum (ER) Ca 2+ release associated with diminished phosphorylation of the inositol 3-phosphate receptor. Restoring sAC expression exclusively in the mitochondrial matrix rescues OXPHOS activity and reduces mitochondrial biogenesis, indicating that these phenotypes are regulated by intramitochondrial sAC. In contrast, Ca 2+ release from the ER is only rescued when sAC expression is restored throughout the cell. Thus, we show that functionally distinct, sAC-defined, intracellular cAMP signaling domains regulate metabolism and Ca 2+ signaling. © 2017. Published by The Company of Biologists Ltd.

OXPHOS-Mediated Induction of NAD+ Promotes Complete Oxidation of Fatty Acids and Interdicts Non-Alcoholic Fatty Liver Disease.

PubMed

Akie, Thomas E; Liu, Lijun; Nam, Minwoo; Lei, Shi; Cooper, Marcus P

2015-01-01

OXPHOS is believed to play an important role in non-alcoholic fatty liver disease (NAFLD), however, precise mechanisms whereby OXPHOS influences lipid homeostasis are incompletely understood. We previously reported that ectopic expression of LRPPRC, a protein that increases cristae density and OXPHOS, promoted fatty acid oxidation in cultured primary hepatocytes. To determine the biological significance of that observation and define underlying mechanisms, we have ectopically expressed LRPPRC in mouse liver in the setting of NAFLD. Interestingly, ectopic expression of LRPPRC in mouse liver completely interdicted NAFLD, including inflammation. Consistent with mitigation of NAFLD, two markers of hepatic insulin resistance--ROS and PKCε activity--were both modestly reduced. As reported by others, improvement of NAFLD was associated with improved whole-body insulin sensitivity. Regarding hepatic lipid homeostasis, the ratio of NAD+ to NADH was dramatically increased in mouse liver replete with LRPPRC. Pharmacological activators and inhibitors of the cellular respiration respectively increased and decreased the [NAD+]/[NADH] ratio, indicating respiration-mediated control of the [NAD+]/[NADH] ratio. Supporting a prominent role for NAD+, increasing the concentration of NAD+ stimulated complete oxidation of fatty acids. Importantly, NAD+ rescued impaired fatty acid oxidation in hepatocytes deficient for either OXPHOS or SIRT3. These data are consistent with a model whereby augmented hepatic OXPHOS increases NAD+, which in turn promotes complete oxidation of fatty acids and protects against NAFLD.

C1orf163/RESA1 is a novel mitochondrial intermembrane space protein connected to respiratory chain assembly.

PubMed

Kozjak-Pavlovic, Vera; Prell, Florian; Thiede, Bernd; Götz, Monika; Wosiek, Dominik; Ott, Christine; Rudel, Thomas

2014-02-20

Oxidative phosphorylation (OXPHOS) in mitochondria takes place at the inner membrane, which folds into numerous cristae. The stability of cristae depends, among other things, on the mitochondrial intermembrane space bridging complex. Its components include inner mitochondrial membrane protein mitofilin and outer membrane protein Sam50. We identified a conserved, uncharacterized protein, C1orf163 [SEL1 repeat containing 1 protein (SELRC1)], as one of the proteins significantly reduced after the knockdown of Sam50 and mitofilin. We show that C1orf163 is a mitochondrial soluble intermembrane space protein. Sam50 depletion affects moderately the import and assembly of C1orf163 into two protein complexes of approximately 60kDa and 150kDa. We observe that the knockdown of C1orf163 leads to reduction of levels of proteins belonging to the OXPHOS complexes. The activity of complexes I and IV is reduced in C1orf163-depleted cells, and we observe the strongest defects in the assembly of complex IV. Therefore, we propose C1orf163 to be a novel factor important for the assembly of respiratory chain complexes in human mitochondria and suggest to name it RESA1 (for RESpiratory chain Assembly 1). Copyright © 2013 Elsevier Ltd. All rights reserved.

Pleiotropic Effects of Biguanides on Mitochondrial Reactive Oxygen Species Production.

PubMed

Pecinova, Alena; Drahota, Zdenek; Kovalcikova, Jana; Kovarova, Nikola; Pecina, Petr; Alan, Lukas; Zima, Michal; Houstek, Josef; Mracek, Tomas

2017-01-01

Metformin is widely prescribed as a first-choice antihyperglycemic drug for treatment of type 2 diabetes mellitus, and recent epidemiological studies showed its utility also in cancer therapy. Although it is in use since the 1970s, its molecular target, either for antihyperglycemic or antineoplastic action, remains elusive. However, the body of the research on metformin effect oscillates around mitochondrial metabolism, including the function of oxidative phosphorylation (OXPHOS) apparatus. In this study, we focused on direct inhibitory mechanism of biguanides (metformin and phenformin) on OXPHOS complexes and its functional impact, using the model of isolated brown adipose tissue mitochondria. We demonstrate that biguanides nonspecifically target the activities of all respiratory chain dehydrogenases (mitochondrial NADH, succinate, and glycerophosphate dehydrogenases), but only at very high concentrations (10 -2 -10 -1  M) that highly exceed cellular concentrations observed during the treatment. In addition, these concentrations of biguanides also trigger burst of reactive oxygen species production which, in combination with pleiotropic OXPHOS inhibition, can be toxic for the organism. We conclude that the beneficial effect of biguanides should probably be associated with subtler mechanism, different from the generalized inhibition of the respiratory chain.

Comprehensive mathematical model of oxidative phosphorylation valid for physiological and pathological conditions.

PubMed

Heiske, Margit; Letellier, Thierry; Klipp, Edda

2017-09-01

We developed a mathematical model of oxidative phosphorylation (OXPHOS) that allows for a precise description of mitochondrial function with respect to the respiratory flux and the ATP production. The model reproduced flux-force relationships under various experimental conditions (state 3 and 4, uncoupling, and shortage of respiratory substrate) as well as time courses, exhibiting correct P/O ratios. The model was able to reproduce experimental threshold curves for perturbations of the respiratory chain complexes, the F 1 F 0 -ATP synthase, the ADP/ATP carrier, the phosphate/OH carrier, and the proton leak. Thus, the model is well suited to study complex interactions within the OXPHOS system, especially with respect to physiological adaptations or pathological modifications, influencing substrate and product affinities or maximal catalytic rates. Moreover, it could be a useful tool to study the role of OXPHOS and its capacity to compensate or enhance physiopathologies of the mitochondrial and cellular energy metabolism. © 2017 Federation of European Biochemical Societies.

Myopathology of Adult and Paediatric Mitochondrial Diseases

PubMed Central

Phadke, Rahul

2017-01-01

Mitochondria are dynamic organelles ubiquitously present in nucleated eukaryotic cells, subserving multiple metabolic functions, including cellular ATP generation by oxidative phosphorylation (OXPHOS). The OXPHOS machinery comprises five transmembrane respiratory chain enzyme complexes (RC). Defective OXPHOS gives rise to mitochondrial diseases (mtD). The incredible phenotypic and genetic diversity of mtD can be attributed at least in part to the RC dual genetic control (nuclear DNA (nDNA) and mitochondrial DNA (mtDNA)) and the complex interaction between the two genomes. Despite the increasing use of next-generation-sequencing (NGS) and various omics platforms in unravelling novel mtD genes and pathomechanisms, current clinical practice for investigating mtD essentially involves a multipronged approach including clinical assessment, metabolic screening, imaging, pathological, biochemical and functional testing to guide molecular genetic analysis. This review addresses the broad muscle pathology landscape including genotype–phenotype correlations in adult and paediatric mtD, the role of immunodiagnostics in understanding some of the pathomechanisms underpinning the canonical features of mtD, and recent diagnostic advances in the field. PMID:28677615

Palmitoleic acid (16:1n7) increases oxygen consumption, fatty acid oxidation and ATP content in white adipocytes.

PubMed

Cruz, Maysa M; Lopes, Andressa B; Crisma, Amanda R; de Sá, Roberta C C; Kuwabara, Wilson M T; Curi, Rui; de Andrade, Paula B M; Alonso-Vale, Maria I C

2018-03-20

We have recently demonstrated that palmitoleic acid (16:1n7) increases lipolysis, glucose uptake and glucose utilization for energy production in white adipose cells. In the present study, we tested the hypothesis that palmitoleic acid modulates bioenergetic activity in white adipocytes. For this, 3 T3-L1 pre-adipocytes were differentiated into mature adipocytes in the presence (or absence) of palmitic (16:0) or palmitoleic (16:1n7) acid at 100 or 200 μM. The following parameters were evaluated: lipolysis, lipogenesis, fatty acid (FA) oxidation, ATP content, oxygen consumption, mitochondrial mass, citrate synthase activity and protein content of mitochondrial oxidative phosphorylation (OXPHOS) complexes. Treatment with 16:1n7 during 9 days raised basal and isoproterenol-stimulated lipolysis, FA incorporation into triacylglycerol (TAG), FA oxidation, oxygen consumption, protein expression of subunits representing OXPHOS complex II, III, and V and intracellular ATP content. These effects were not observed in adipocytes treated with 16:0. Palmitoleic acid, by concerted action on lipolysis, FA esterification, mitochondrial FA oxidation, oxygen consumption and ATP content, does enhance white adipocyte energy expenditure and may act as local hormone.

Small structural changes on a hydroquinone scaffold determine the complex I inhibition or uncoupling of tumoral oxidative phosphorylation.

PubMed

Urra, Félix A; Córdova-Delgado, Miguel; Lapier, Michel; Orellana-Manzano, Andrea; Acevedo-Arévalo, Luis; Pessoa-Mahana, Hernán; González-Vivanco, Jaime M; Martínez-Cifuentes, Maximiliano; Ramírez-Rodríguez, Oney; Millas-Vargas, Juan Pablo; Weiss-López, Boris; Pavani, Mario; Ferreira, Jorge; Araya-Maturana, Ramiro

2016-01-15

Mitochondria participate in several distinctiveness of cancer cell, being a promising target for the design of anti-cancer compounds. Previously, we described that ortho-carbonyl hydroquinone scaffold 14 inhibits the complex I-dependent respiration with selective anti-proliferative effect on mouse mammary adenocarcinoma TA3/Ha cancer cells; however, the structural requirements of this hydroquinone scaffold to affect the oxidative phosphorylation (OXPHOS) of cancer cells have not been studied in detail. Here, we characterize the mitochondrial metabolism of TA3/Ha cancer cells, which exhibit a high oxidative metabolism, and evaluate the effect of small structural changes of the hydroquinone scaffold 14 on the respiration of this cell line. Our results indicate that these structural changes modify the effect on OXPHOS, obtaining compounds with three alternative actions: inhibitors of complex I-dependent respiration, uncoupler of OXPHOS and compounds with both actions. To confirm this, the effect of a bicyclic hydroquinone (9) was evaluated in isolated mitochondria. Hydroquinone 9 increased mitochondrial respiration in state 4o without effects on the ADP-stimulated respiration (state 3ADP), decreasing the complexes I and II-dependent respiratory control ratio. The effect on mitochondrial respiration was reversed by 6-ketocholestanol addition, indicating that this hydroquinone is a protonophoric uncoupling agent. In intact TA3/Ha cells, hydroquinone 9 caused mitochondrial depolarization, decreasing intracellular ATP and NAD(P)H levels and GSH/GSSG ratio, and slightly increasing the ROS levels. Moreover, it exhibited selective NAD(P)H availability-dependent anti-proliferative effect on cancer cells. Therefore, our results indicate that the ortho-carbonyl hydroquinone scaffold offers the possibility to design compounds with specific actions on OXPHOS of cancer cells.

Assessment of mitochondrial functions in Daphnia pulex clones using high-resolution respirometry.

PubMed

Kake-Guena, Sandrine A; Touisse, Kamal; Vergilino, Roland; Dufresne, France; Blier, Pierre U; Lemieux, Hélène

2015-06-01

The objectives of our study were to adapt a method to measure mitochondrial function in intact mitochondria from the small crustacean Daphnia pulex and to validate if this method was sensitive enough to characterize mitochondrial metabolism in clones of the pulex complex differing in ploidy levels, mitochondrial DNA haplotypes, and geographic origins. Daphnia clones belonging to the Daphnia pulex complex represent a powerful model to delineate the link between mitochondrial DNA evolution and mitochondrial phenotypes, as single genotypes with divergent mtDNA can be grown under various experimental conditions. Our study included two diploid clones from temperate environments and two triploid clones from subarctic environments. The whole animal permeabilization and measurement of respiration with high-resolution respirometry enabled the measurement of the functional capacity of specific mitochondrial complexes in four clones. When expressing the activity as ratios, our method detected significant interclonal variations. In the triploid subarctic clone from Kuujjurapik, a higher proportion of the maximal physiological oxidative phosphorylation (OXPHOS) capacity of mitochondria was supported by complex II, and a lower proportion by complex I. The triploid subarctic clone from Churchill (Manitoba) showed the lowest proportion of the maximal OXPHOS supported by complex II. Additional studies are required to determine if these differences in mitochondrial functions are related to differences in mitochondrial haplotypes or ploidy level and if they might be associated with fitness divergences and therefore selective value. © 2015 Wiley Periodicals, Inc.

Absence of Complex I Implicates Rearrangement of the Respiratory Chain in European Mistletoe.

PubMed

Senkler, Jennifer; Rugen, Nils; Eubel, Holger; Hegermann, Jan; Braun, Hans-Peter

2018-05-21

The mitochondrial oxidative phosphorylation (OXPHOS) system, which is based on the presence of five protein complexes, is in the very center of cellular ATP production. Complexes I to IV are components of the respiratory electron transport chain that drives proton translocation across the inner mitochondrial membrane. The resulting proton gradient is used by complex V (the ATP synthase complex) for the phosphorylation of ADP. Occurrence of complexes I to V is highly conserved in eukaryotes, with exceptions being restricted to unicellular parasites that take up energy-rich compounds from their hosts. Here we present biochemical evidence that the European mistletoe (Viscum album), an obligate semi-parasite living on branches of trees, has a highly unusual OXPHOS system. V. album mitochondria completely lack complex I and have greatly reduced amounts of complexes II and V. At the same time, the complexes III and IV form remarkably stable respiratory supercomplexes. Furthermore, complexome profiling revealed the presence of 150 kDa complexes that include type II NAD(P)H dehydrogenases and an alternative oxidase. Although the absence of complex I genes in mitochondrial genomes of mistletoe species has recently been reported, this is the first biochemical proof that these genes have not been transferred to the nuclear genome and that this respiratory complex indeed is not assembled. As a consequence, the whole respiratory chain is remodeled. Our results demonstrate that, in the context of parasitism, multicellular life can cope with lack of one of the OXPHOS complexes and give new insights into the life strategy of mistletoe species. Copyright © 2018 Elsevier Ltd. All rights reserved.

Metabolic determinants of cancer cell sensitivity to glucose limitation and biguanides

NASA Astrophysics Data System (ADS)

Birsoy, Kıvanç; Possemato, Richard; Lorbeer, Franziska K.; Bayraktar, Erol C.; Thiru, Prathapan; Yucel, Burcu; Wang, Tim; Chen, Walter W.; Clish, Clary B.; Sabatini, David M.

2014-04-01

As the concentrations of highly consumed nutrients, particularly glucose, are generally lower in tumours than in normal tissues, cancer cells must adapt their metabolism to the tumour microenvironment. A better understanding of these adaptations might reveal cancer cell liabilities that can be exploited for therapeutic benefit. Here we developed a continuous-flow culture apparatus (Nutrostat) for maintaining proliferating cells in low-nutrient media for long periods of time, and used it to undertake competitive proliferation assays on a pooled collection of barcoded cancer cell lines cultured in low-glucose conditions. Sensitivity to low glucose varies amongst cell lines, and an RNA interference (RNAi) screen pinpointed mitochondrial oxidative phosphorylation (OXPHOS) as the major pathway required for optimal proliferation in low glucose. We found that cell lines most sensitive to low glucose are defective in the OXPHOS upregulation that is normally caused by glucose limitation as a result of either mitochondrial DNA (mtDNA) mutations in complex I genes or impaired glucose utilization. These defects predict sensitivity to biguanides, antidiabetic drugs that inhibit OXPHOS, when cancer cells are grown in low glucose or as tumour xenografts. Notably, the biguanide sensitivity of cancer cells with mtDNA mutations was reversed by ectopic expression of yeast NDI1, a ubiquinone oxidoreductase that allows bypass of complex I function. Thus, we conclude that mtDNA mutations and impaired glucose utilization are potential biomarkers for identifying tumours with increased sensitivity to OXPHOS inhibitors.

Effects of alpha-melanocyte-stimulating hormone on mitochondrial energy metabolism in rats of different age-groups.

PubMed

Feichtinger, René G; Pétervári, Erika; Zopf, Michaela; Vidali, Silvia; Aminzadeh-Gohari, Sepideh; Mayr, Johannes A; Kofler, Barbara; Balaskó, Márta

2017-08-01

Hypothalamic alpha-melanocyte-stimulating hormone (α-MSH) is a key catabolic mediator of energy homeostasis. Its anorexigenic and hypermetabolic effects show characteristic age-related alterations that may be part of the mechanism of middle-aged obesity and geriatric anorexia/cachexia seen in humans and other mammals. We aimed to investigate the role of α-MSH in mitochondrial energy metabolism during the course of aging in a rodent model. To determine the role of α-MSH in mitochondrial energy metabolism in muscle, we administered intracerebroventricular (ICV) infusions of α-MSH for 7-days to different age-groups of male Wistar rats. The activities of oxidative phosphorylation complexes I to V and citrate synthase were determined and compared to those of age-matched controls. We also quantified mitochondrial DNA (mtDNA) copy number and measured the expression of the master regulators of mitochondrial biogenesis, peroxisome proliferator-activated receptor gamma coactivator 1-alpha (PGC-1α) and peroxisome proliferator-activated receptor gamma (PPARγ). The peptide reduced weight gain in juvenile rats to one fifth of that of controls and increased the weight loss in older animals by about five fold. Mitochondrial DNA copy number inversely correlated with changes in body weight in controls, but not in α-MSH-treated animals. The strong increase in body weight in young rats was associated with a low mtDNA copy number and high PPARγ mRNA levels in controls. Expression of PGC-1α and PPARγ declined with age, whereas OXPHOS and citrate synthase enzyme activities were unchanged. In contrast, α-MSH treatment suppressed OXPHOS enzyme and citrate synthase activity. In conclusion, our results showed age-related differences in the metabolic effects of α-MSH. In addition, administration of α-MSH suppressed citrate synthase and OXPHOS activities independent of age. These findings suggest that α-MSH exposure may inhibit mitochondrial biogenesis. Copyright © 2016 Elsevier Ltd. All rights reserved.

Lactic acidosis switches cancer cells from aerobic glycolysis back to dominant oxidative phosphorylation.

PubMed

Wu, Hao; Ying, Minfeng; Hu, Xun

2016-06-28

While transformation of normal cells to cancer cells is accompanied with a switch from oxidative phosphorylation (OXPHOS) to aerobic glycolysis, it is interesting to ask if cancer cells can revert from Warburg effect to OXPHOS. Our previous works suggested that cancer cells reverted to OXPHOS, when they were exposed to lactic acidosis, a common factor in tumor environment. However, the conclusion cannot be drawn unless ATP output from glycolysis and OXPHOS is quantitatively determined. Here we quantitatively measured ATP generation from glycolysis and OXPHOS in 9 randomly selected cancer cell lines. Without lactic acidosis, glycolysis and OXPHOS generated 23.7% - 52.2 % and 47.8% - 76.3% of total ATP, respectively; with lactic acidosis (20 mM lactate with pH 6.7), glycolysis and OXPHOS provided 5.7% - 13.4% and 86.6% - 94.3% of total ATP. We concluded that cancer cells under lactic acidosis reverted from Warburg effect to OXPHOS phenotype.

Voluntary exercise promotes beneficial anti-aging mechanisms in SAMP8 female brain.

PubMed

Bayod, Sergi; Guzmán-Brambila, Carolina; Sanchez-Roige, Sandra; Lalanza, Jaume F; Kaliman, Perla; Ortuño-Sahagun, Daniel; Escorihuela, Rosa M; Pallàs, Mercè

2015-02-01

Regular physical exercise mediates health and longevity promotion involving Sirtuin 1 (SIRT1)-regulated pathways. The anti-aging activity of SIRT1 is achieved, at least in part, by means of fine-tuning the adenosine monophosphate (AMP)-activated protein kinase (AMPK) pathway by preventing the transition of an originally pro-survival program into a pro-aging mechanism. Additionally, SIRT1 promotes mitochondrial function and reduces the production of reactive oxygen species (ROS) through regulating peroxisome proliferator-activated receptor γ coactivator 1α (PGC-1α), the master controller of mitochondrial biogenesis. Here, by using senescence-accelerated mice prone 8 (SAMP8) as a model for aging, we determined the effect of wheel-running as a paradigm for long-term voluntary exercise on SIRT1-AMPK pathway and mitochondrial functionality measured by oxidative phosphorylation (OXPHOS) complex content in the hippocampus and cortex. We found differential activation of SIRT1 in both tissues and hippocampal-specific activation of AMPK. These findings correlated well with significant changes in OXPHOS in the hippocampal, but not in the cerebral cortex, area. Collectively, the results revealed greater benefits of the exercise in the wheel-running intervention in a murine model of senescence, which was directly related with mitochondrial function and which was mediated through the modulation of SIRT1 and AMPK pathways.

Self-renewal of CD133(hi) cells by IL6/Notch3 signalling regulates endocrine resistance in metastatic breast cancer.

PubMed

Sansone, Pasquale; Ceccarelli, Claudio; Berishaj, Marjan; Chang, Qing; Rajasekhar, Vinagolu K; Perna, Fabiana; Bowman, Robert L; Vidone, Michele; Daly, Laura; Nnoli, Jennifer; Santini, Donatella; Taffurelli, Mario; Shih, Natalie N C; Feldman, Michael; Mao, Jun J; Colameco, Christopher; Chen, Jinbo; DeMichele, Angela; Fabbri, Nicola; Healey, John H; Cricca, Monica; Gasparre, Giuseppe; Lyden, David; Bonafé, Massimiliano; Bromberg, Jacqueline

2016-02-09

The mechanisms of metastatic progression from hormonal therapy (HT) are largely unknown in luminal breast cancer. Here we demonstrate the enrichment of CD133(hi)/ER(lo) cancer cells in clinical specimens following neoadjuvant endocrine therapy and in HT refractory metastatic disease. We develop experimental models of metastatic luminal breast cancer and demonstrate that HT can promote the generation of HT-resistant, self-renewing CD133(hi)/ER(lo)/IL6(hi) cancer stem cells (CSCs). HT initially abrogates oxidative phosphorylation (OXPHOS) generating self-renewal-deficient cancer cells, CD133(hi)/ER(lo)/OXPHOS(lo). These cells exit metabolic dormancy via an IL6-driven feed-forward ER(lo)-IL6(hi)-Notch(hi) loop, activating OXPHOS, in the absence of ER activity. The inhibition of IL6R/IL6-Notch pathways switches the self-renewal of CD133(hi) CSCs, from an IL6/Notch-dependent one to an ER-dependent one, through the re-expression of ER. Thus, HT induces an OXPHOS metabolic editing of luminal breast cancers, paradoxically establishing HT-driven self-renewal of dormant CD133(hi)/ER(lo) cells mediating metastatic progression, which is sensitive to dual targeted therapy.

Lactic acidosis switches cancer cells from aerobic glycolysis back to dominant oxidative phosphorylation

PubMed Central

Wu, Hao; Ying, Minfeng; Hu, Xun

2016-01-01

While transformation of normal cells to cancer cells is accompanied with a switch from oxidative phosphorylation (OXPHOS) to aerobic glycolysis, it is interesting to ask if cancer cells can revert from Warburg effect to OXPHOS. Our previous works suggested that cancer cells reverted to OXPHOS, when they were exposed to lactic acidosis, a common factor in tumor environment. However, the conclusion cannot be drawn unless ATP output from glycolysis and OXPHOS is quantitatively determined. Here we quantitatively measured ATP generation from glycolysis and OXPHOS in 9 randomly selected cancer cell lines. Without lactic acidosis, glycolysis and OXPHOS generated 23.7% − 52.2 % and 47.8% − 76.3% of total ATP, respectively; with lactic acidosis (20 mM lactate with pH 6.7), glycolysis and OXPHOS provided 5.7% − 13.4% and 86.6% − 94.3% of total ATP. We concluded that cancer cells under lactic acidosis reverted from Warburg effect to OXPHOS phenotype. PMID:27259254

Upregulation of glycolysis and oxidative phosphorylation in benzo[β]pyrene and arsenic-induced rat lung epithelial transformed cells

PubMed Central

Li, Guanwu; Tsao, Sai-Wah; Chiu, Jen-Fu

2016-01-01

Arsenic and benzo[β]pyrene (B[a]P) are common contaminants in developing countries. Many studies have investigated the consequences of arsenic and/or B[a]P-induced cellular transformation, including altered metabolism. In the present study, we show that, in addition to elevated glycolysis, B[a]P/arsenic-induced transformation also stimulates oxidative phosphorylation (OXPHOS). Proteomic data and immunoblot studies demonstrated that enzymatic activities, involved in both glycolysis and OXPHOS, are upregulated in the primary transformed rat lung epithelial cell (TLEC) culture, as well as in subcloned TLEC cell lines (TMCs), indicating that OXPHOS was active and still contributed to energy production. LEC expression, of the glycolytic enzyme phosphoglycerate mutase (PGAM) and the TCA cycle enzyme alpha-ketoglutarate dehydrogenase (OGDH), revealed an alternating cyclic pattern of glycolysis and OXPHOS during cell transformation. We also found that the expression levels of hypoxia-inducible factor-1β were consistent with the pattern of glycolysis during the course of transformation. Low doses of an ATP synthase inhibitor depleted endogenous ATP levels to a greater extent in TLECs, compared to parental LECs, indicating greater sensitivity of B[a]P/arsenic-transformed cells to ATP depletion. However, TLEC cells exhibited better survival under hypoxia, possibly due to further induction of anaerobic glycolysis. Collectively, our data indicate that B[a]P/arsenic-transformed cells can maintain energy production through upregulation of both glycolysis and OXPHOS. Selective inhibition of metabolic pathways may serve as a therapeutic option for cancer therapy. PMID:27276679

Mechanisms responsible for the acceleration of pulmonary V̇O2 on-kinetics in humans after prolonged endurance training.

PubMed

Zoladz, Jerzy A; Grassi, Bruno; Majerczak, Joanna; Szkutnik, Zbigniew; Korostyński, Michal; Grandys, Marcin; Jarmuszkiewicz, Wieslawa; Korzeniewski, Bernard

2014-11-01

The effect of prolonged endurance training on the pulmonary V̇O2 on- and off-kinetics in humans, in relation to muscle mitochondria biogenesis, is investigated. Eleven untrained physically active men (means±SD: age 22.4±1.5 years, V̇O2peak 3,187±479 ml/min) performed endurance cycling training (4 sessions per week) lasting 20 wk. Training shortened τp of the pulmonary V̇O2 on-kinetics during moderate-intensity cycling by ∼19% from 28.3±5.2 to 23.0±4.0 s (P=0.005). τp of the pulmonary V̇O2 off-kinetics decreased by ∼11% from 33.7±7.2 to 30.0±6.6 (P=0.02). Training increased (in vastus lateralis muscle) mitochondrial DNA copy number in relation to nuclear DNA (mtDNA/nDNA) (+53%) (P=0.014), maximal citrate synthase (CS) activity (+38%), and CS protein content (+38%) (P=0.004), whereas maximal cytochrome c oxidase (COX) activity after training tended to be only slightly (+5%) elevated (P=0.08). By applying to the experimental data, our computer model of oxidative phosphorylation (OXPHOS) and using metabolic control analysis, we argue that COX activity is a much better measure of OXPHOS intensity than CS activity. According to the model, in the present study a training-induced increase in OXPHOS activity accounted for about 0-10% of the decrease in τp of muscle and pulmonary V̇O2 for the on-transient, whereas the remaining 90-100% is caused by an increase in each-step parallel activation of OXPHOS. Copyright © 2014 the American Physiological Society.

Clinical differences in patients with mitochondriocytopathies due to nuclear versus mitochondrial DNA mutations.

PubMed

Rubio-Gozalbo, M E; Dijkman, K P; van den Heuvel, L P; Sengers, R C; Wendel, U; Smeitink, J A

2000-01-01

Defects in oxidative phosphorylation (OXPHOS) are genetically unique because the different components involved in this process, respiratory chain enzyme complexes (I, III, and IV) and complex V, are encoded by nuclear and mitochondrial genome. The objective of the study was to assess whether there are clinical differences in patients suffering from OXPHOS defects caused by nuclear or mitochondrial DNA (mtDNA) mutations. We studied 16 families with > or = two siblings with a genetically established OXPHOS deficiency, four due to a nuclear gene mutation and 12 due to a mtDNA mutation. Siblings with a nuclear gene mutation showed very similar clinical pictures that became manifest in the first years (ranging from first months to early childhood). There was a severe progressive course. Seven of the eight children died in their first decade. Conversely, siblings with a mtDNA mutation had clinical pictures that varied from almost alike to very distinct. They became symptomatic at an older age (ranging from childhood to adulthood), with the exception of defects associated with Leigh or Leigh-like phenotype. The clinical course was more gradual and relatively less severe; four of the 26 patients died, one in his second year, another in her second decade and two in their sixth decade. There are differences in age at onset, severity of clinical course, outcome, and intrafamilial variability in patients affected of an OXPHOS defect due to nuclear or mtDNA mutations. Patients with nuclear mutations become symptomatic at a young age, and have a severe clinical course. Patients with mtDNA mutations show a wider clinical spectrum of age at onset and severity. These differences may be of importance regarding the choice of which genome to study in affected patients as well as with respect to genetic counseling. Copyright 2000 Wiley-Liss, Inc.

Effects of β-hydroxy-β-methylbutyrate on skeletal muscle mitochondrial content and dynamics, and lipids after 10 days of bed rest in older adults.

PubMed

Standley, Robert A; Distefano, Giovanna; Pereira, Suzette L; Tian, Min; Kelly, Owen J; Coen, Paul M; Deutz, Nicolaas E P; Wolfe, Robert R; Goodpaster, Bret H

2017-11-01

Loss of muscle mass during periods of disuse likely has negative health consequences for older adults. We have previously shown that β-hydroxy-β-methylbutyrate (HMB) supplementation during 10 days of strict bed rest (BR) attenuates the loss of lean mass in older adults. To elucidate potential molecular mechanisms of HMB effects on muscle during BR and resistance training rehabilitation (RT), we examined mediators of skeletal muscle mitochondrial dynamics, autophagy and atrophy, and intramyocellular lipids. Nineteen older adults (60-76 yr) completed 10 days BR followed by 8-wk RT rehabilitation. Subjects were randomized to either HMB (3 g/day HMB; n = 11) or control (CON; n = 8) groups. Skeletal muscle cross-sectional area (CSA) was determined by histology from percutaneous vastus lateralis biopsies. We measured protein markers of mitochondrial content [oxidative phosphorylation (OXPHOS)], fusion and fission (MFN2, OPA1, FIS1, and DRP1), autophagy (Beclin1, LC3B, and BNIP3), and atrophy [poly-ubiquinated proteins (poly-ub)] by Western blot. Fatty acid composition of several lipid classes in skeletal muscle was measured by infusion-MS analysis. Poly-ub proteins and OXPHOS complex I increased in both groups following BR ( P < 0.05, main effect for time), and muscle triglyceride content tended to increase following BR in the HMB group ( P = 0.055). RT rehabilitation increased OXPHOS complex II protein ( P < 0.05), and total OXPHOS content tended ( P = 0.0504) to be higher in HMB group. In addition, higher levels of DRP1 and MFN2 were maintained in the HMB group after RT ( P < 0.05). BNIP3 and poly-ub proteins were significantly reduced following rehabilitation in both groups ( P < 0.05). Collectively, these data suggest that HMB influences mitochondrial dynamics and lipid metabolism during disuse atrophy and rehabilitation. NEW & NOTEWORTHY Mitochondrial content and dynamics remained unchanged over 10 days of BR in older adults. HMB stimulated intramuscular lipid storage as triacylglycerol following 10 days of bed rest (BR) and maintained higher mitochondrial OXPHOS content and dynamics during the 8-wk resistance exercise rehabilitation program. Copyright © 2017 the American Physiological Society.

Cold and Heat Stress Diversely Alter Both Cauliflower Respiration and Distinct Mitochondrial Proteins Including OXPHOS Components and Matrix Enzymes

PubMed Central

Rurek, Michał; Czołpińska, Magdalena; Pawłowski, Tomasz Andrzej; Krzesiński, Włodzimierz; Spiżewski, Tomasz

2018-01-01

Complex proteomic and physiological approaches for studying cold and heat stress responses in plant mitochondria are still limited. Variations in the mitochondrial proteome of cauliflower (Brassica oleracea var. botrytis) curds after cold and heat and after stress recovery were assayed by two-dimensional polyacrylamide gel electrophoresis (2D PAGE) in relation to mRNA abundance and respiratory parameters. Quantitative analysis of the mitochondrial proteome revealed numerous stress-affected protein spots. In cold, major downregulations in the level of photorespiratory enzymes, porine isoforms, oxidative phosphorylation (OXPHOS) and some low-abundant proteins were observed. In contrast, carbohydrate metabolism enzymes, heat-shock proteins, translation, protein import, and OXPHOS components were involved in heat response and recovery. Several transcriptomic and metabolic regulation mechanisms are also suggested. Cauliflower plants appeared less susceptible to heat; closed stomata in heat stress resulted in moderate photosynthetic, but only minor respiratory impairments, however, photosystem II performance was unaffected. Decreased photorespiration corresponded with proteomic alterations in cold. Our results show that cold and heat stress not only operate in diverse modes (exemplified by cold-specific accumulation of some heat shock proteins), but exert some associations at molecular and physiological levels. This implies a more complex model of action of investigated stresses on plant mitochondria. PMID:29547512

Modulation of oxidative phosphorylation (OXPHOS) by radiation- induced biophotons.

PubMed

Le, Michelle; McNeill, Fiona E; Seymour, Colin B; Rusin, Andrej; Diamond, Kevin; Rainbow, Andrew J; Murphy, James; Mothersill, Carmel E

2018-05-01

Radiation-induced biophotons are an electromagnetic form of bystander signalling. In human cells, biophoton signalling is capable of eliciting effects in non-irradiated bystander cells. However, the mechanisms by which the biophotons interact and act upon the bystander cells are not clearly understood. Mitochondrial energy production and ROS are known to be involved but the precise interactions are not known. To address this question, we have investigated the effect of biophoton emission upon the function of the complexes of oxidative phosphorylation (OXPHOS). The exposure of bystander HCT116 p53 +/+ cells to biophoton signals emitted from β-irradiated HCT116 p53 +/+ cells induced significant modifications in the activity of Complex I (NADH dehydrogenase or NADH:ubiquinone oxidoreductase) such that the activity was severely diminished compared to non-irradiated controls. The enzymatic assay showed that the efficiency of NADH oxidation to NAD+ was severely compromised. It is suspected that this impairment may be linked to the photoabsorption of biophotons in the blue wavelength range (492-455 nm). The photobiomodulation to Complex I was suspected to contribute greatly to the inefficiency of ATP synthase function since it resulted in a lower quantity of H + ions to be available for use in the process of chemiosmosis. Other reactions of the ETC were not significantly impacted. Overall, these results provide evidence for a link between biophoton emission and biomodulation of the mitochondrial ATP synthesis process. However, there are many aspects of biological modulation by radiation-induced biophotons which will require further elucidation. Copyright © 2018 Elsevier Inc. All rights reserved.

Changes in mitochondrial function and mitochondria associated protein expression in response to 2-weeks of high intensity interval training

PubMed Central

Vincent, Grace; Lamon, Séverine; Gant, Nicholas; Vincent, Peter J.; MacDonald, Julia R.; Markworth, James F.; Edge, Johann A.; Hickey, Anthony J. R.

2015-01-01

Purpose: High-intensity short-duration interval training (HIT) stimulates functional and metabolic adaptation in skeletal muscle, but the influence of HIT on mitochondrial function remains poorly studied in humans. Mitochondrial metabolism as well as mitochondrial-associated protein expression were tested in untrained participants performing HIT over a 2-week period. Methods: Eight males performed a single-leg cycling protocol (12 × 1 min intervals at 120% peak power output, 90 s recovery, 4 days/week). Muscle biopsies (vastus lateralis) were taken pre- and post-HIT. Mitochondrial respiration in permeabilized fibers, citrate synthase (CS) activity and protein expression of peroxisome proliferator-activated receptor gamma coactivator (PGC-1α) and respiratory complex components were measured. Results: HIT training improved peak power and time to fatigue. Increases in absolute oxidative phosphorylation (OXPHOS) capacities and CS activity were observed, but not in the ratio of CCO to the electron transport system (CCO/ETS), the respiratory control ratios (RCR-1 and RCR-2) or mitochondrial-associated protein expression. Specific increases in OXPHOS flux were not apparent after normalization to CS, indicating that gross changes mainly resulted from increased mitochondrial mass. Conclusion: Over only 2 weeks HIT significantly increased mitochondrial function in skeletal muscle independently of detectable changes in mitochondrial-associated and mitogenic protein expression. PMID:25759671

OXPHOS-Dependent Cells Identify Environmental Disruptors of Mitochondrial Function

EPA Science Inventory

Mitochondrial dysfunction is associated with numerous chronic diseases including metabolic syndrome. Environmental chemicals can impair mitochondrial function through numerous mechanisms such as membrane disruption, complex inhibition and electron transport chain uncoupling. Curr...

OXPHOS Defects Due to mtDNA Mutations: Glutamine to the Rescue!

PubMed

Chinopoulos, Christos

2018-06-05

Mutations in mtDNA associated with OXPHOS defects preclude energy harnessing by OXPHOS. The work of Chen et al. (2018) is previewed, reporting flux pathways of glutamine catabolism in mtDNA mutant cells yielding high-energy phosphates through substrate-level phosphorylation and the influence exerted by the severity of OXPHOS impairment. Copyright © 2018 Elsevier Inc. All rights reserved.

Pyruvate dehydrogenase expression is negatively associated with cell stemness and worse clinical outcome in prostate cancers

PubMed Central

Zhong, Yali; Li, Xiaoli; Ji, Yasai; Li, Xiaoran; Li, Yaqing; Yu, Dandan; Yuan, Yuan; Liu, Jian; Li, Huixiang; Zhang, Mingzhi; Ji, Zhenyu; Fan, Dandan; Wen, Jianguo; Goscinski, Mariusz Adam; Yuan, Long; Hao, Bin; Nesland, Jahn M; Suo, Zhenhe

2017-01-01

Cells generate adenosine-5′-triphosphate (ATP), the major currency for energy-consuming reactions, through mitochondrial oxidative phosphorylation (OXPHOS) and glycolysis. One of the remarkable features of cancer cells is aerobic glycolysis, also known as the “Warburg Effect”, in which cancer cells rely preferentially on glycolysis instead of mitochondrial OXPHOS as the main energy source even in the presence of high oxygen tension. One of the main players in controlling OXPHOS is the mitochondrial gatekeeperpyruvate dehydrogenase complex (PDHc) and its major subunit is E1α (PDHA1). To further analyze the function of PDHA1 in cancer cells, it was knock out (KO) in the human prostate cancer cell line LnCap and a stable KO cell line was established. We demonstrated that PDHA1 gene KO significantly decreased mitochondrial OXPHOS and promoted anaerobic glycolysis, accompanied with higher stemness phenotype including resistance to chemotherapy, enhanced migration ability and increased expression of cancer stem cell markers. We also examined PDHA1 protein expression in prostate cancer tissues by immunohistochemistry and observed that reduced PDHA1 protein expression in clinical prostate carcinomas was significantly correlated with poor prognosis. Collectively, our results show that negative PDHA1 gene expressionis associated with significantly higher cell stemness in prostate cancer cells and reduced protein expression of this gene is associated with shorter clinical outcome in prostate cancers. PMID:28076853

Tumor cell death induced by the inhibition of mitochondrial electron transport: The effect of 3-hydroxybakuchiol

DOE Office of Scientific and Technical Information (OSTI.GOV)

Jaña, Fabián; Faini, Francesca; Lapier, Michel

Changes in mitochondrial ATP synthesis can affect the function of tumor cells due to the dependence of the first step of glycolysis on mitochondrial ATP. The oxidative phosphorylation (OXPHOS) system is responsible for the synthesis of approximately 90% of the ATP in normal cells and up to 50% in most glycolytic cancers; therefore, inhibition of the electron transport chain (ETC) emerges as an attractive therapeutic target. We studied the effect of a lipophilic isoprenylated catechol, 3-hydroxybakuchiol (3-OHbk), a putative ETC inhibitor isolated from Psoralea glandulosa. 3-OHbk exerted cytotoxic and anti-proliferative effects on the TA3/Ha mouse mammary adenocarcinoma cell line andmore » induced a decrease in the mitochondrial transmembrane potential, the activation of caspase-3, the opening of the mitochondrial permeability transport pore (MPTP) and nuclear DNA fragmentation. Additionally, 3-OHbk inhibited oxygen consumption, an effect that was completely reversed by succinate (an electron donor for Complex II) and duroquinol (electron donor for Complex III), suggesting that 3-OHbk disrupted the electron flow at the level of Complex I. The inhibition of OXPHOS did not increase the level of reactive oxygen species (ROS) but caused a large decrease in the intracellular ATP level. ETC inhibitors have been shown to induce cell death through necrosis and apoptosis by increasing ROS generation. Nevertheless, we demonstrated that 3-OHbk inhibited the ETC and induced apoptosis through an interaction with Complex I. By delivering electrons directly to Complex III with duroquinol, cell death was almost completely abrogated. These results suggest that 3-OHbk has antitumor activity resulting from interactions with the ETC, a system that is already deficient in cancer cells. - Highlights: • We studied the anticancer activity of a natural compound, 3-OHbk, on TA3/Ha cells. • 3-OHbk inhibited mitochondrial electron flow by interacting with Complex I. • Complex I inhibition did not induce ROS generation. • 3-OHbk induced apoptosis in tumor cells with no effect on mammary epithelial cells. • Mitochondrial bioenergetics is implicated in anticancer action of 3-OHbk.« less

Clarifying the supercomplex: the higher-order organization of the mitochondrial electron transport chain.

PubMed

Letts, James A; Sazanov, Leonid A

2017-10-05

The oxidative phosphorylation electron transport chain (OXPHOS-ETC) of the inner mitochondrial membrane is composed of five large protein complexes, named CI-CV. These complexes convert energy from the food we eat into ATP, a small molecule used to power a multitude of essential reactions throughout the cell. OXPHOS-ETC complexes are organized into supercomplexes (SCs) of defined stoichiometry: CI forms a supercomplex with CIII 2 and CIV (SC I+III 2 +IV, known as the respirasome), as well as with CIII 2 alone (SC I+III 2 ). CIII 2 forms a supercomplex with CIV (SC III 2 +IV) and CV forms dimers (CV 2 ). Recent cryo-EM studies have revealed the structures of SC I+III 2 +IV and SC I+III 2 . Furthermore, recent work has shed light on the assembly and function of the SCs. Here we review and compare these recent studies and discuss how they have advanced our understanding of mitochondrial electron transport.

Crif1 Deficiency Reduces Adipose OXPHOS Capacity and Triggers Inflammation and Insulin Resistance in Mice

PubMed Central

Ryu, Min Jeong; Kim, Soung Jung; Kim, Yong Kyung; Choi, Min Jeong; Tadi, Surendar; Lee, Min Hee; Lee, Seong Eun; Chung, Hyo Kyun; Jung, Saet Byel; Kim, Hyun-Jin; Jo, Young Suk; Kim, Koon Soon; Lee, Sang-Hee; Kim, Jin Man; Kweon, Gi Ryang; Park, Ki Cheol; Lee, Jung Uee; Kong, Young Yun; Lee, Chul-Ho; Chung, Jongkyeong; Shong, Minho

2013-01-01

Impaired mitochondrial oxidative phosphorylation (OXPHOS) has been proposed as an etiological mechanism underlying insulin resistance. However, the initiating organ of OXPHOS dysfunction during the development of systemic insulin resistance has yet to be identified. To determine whether adipose OXPHOS deficiency plays an etiological role in systemic insulin resistance, the metabolic phenotype of mice with OXPHOS–deficient adipose tissue was examined. Crif1 is a protein required for the intramitochondrial production of mtDNA–encoded OXPHOS subunits; therefore, Crif1 haploinsufficient deficiency in mice results in a mild, but specific, failure of OXPHOS capacity in vivo. Although adipose-specific Crif1-haploinsufficient mice showed normal growth and development, they became insulin-resistant. Crif1-silenced adipocytes showed higher expression of chemokines, the expression of which is dependent upon stress kinases and antioxidant. Accordingly, examination of adipose tissue from Crif1-haploinsufficient mice revealed increased secretion of MCP1 and TNFα, as well as marked infiltration by macrophages. These findings indicate that the OXPHOS status of adipose tissue determines its metabolic and inflammatory responses, and may cause systemic inflammation and insulin resistance. PMID:23516375

Extramitochondrial energy production in platelets.

PubMed

Ravera, Silvia; Signorello, Maria Grazia; Bartolucci, Martina; Ferrando, Sara; Manni, Lucia; Caicci, Federico; Calzia, Daniela; Panfoli, Isabella; Morelli, Alessandro; Leoncini, Giuliana

2018-05-01

Energy demand in human platelets is very high, to carry out their functions. As for most human cells, the aerobic metabolism represents the primary energy source in platelets, even though mitochondria are negligibly represented. Following the hypothesis that other structures could be involved in chemical energy production, in this work, we have investigated the functional expression of an extramitochondrial aerobic metabolism in platelets. Oximetric and luminometric analyses showed that platelets consume large amounts of oxygen and produce ATP in the presence of common respiring substrates, such as pyruvate + malate or succinate, although morphological electron microscopy analysis showed that these contain few mitochondria. However, evaluation of the anaerobic glycolytic metabolism showed that only 13% of consumed glucose was converted to lactate. Interestingly, the highest OXPHOS activity was observed in the presence of NADH, not a readily permeant respiring substrate for mitochondria. Also, oxygen consumption and ATP synthesis fuelled by NADH were not affected by atractyloside, an inhibitor of the adenine nucleotide translocase, suggesting that these processes may not be ascribed to mitochondria. Functional data were confirmed by immunofluorescence microscopy and Western blot analyses, showing a consistent expression of the β subunit of F 1 F o -ATP synthase and COXII, a subunit of Complex IV, but a low signal of translocase of the inner mitochondrial membrane (a protein not involved in OXPHOS metabolism). Interestingly, the NADH-stimulated oxygen consumption and ATP synthesis increased in the presence of the physiological platelets agonists, thrombin or collagen. Data suggest that in platelets, aerobic energy production is mainly driven by an extramitochondrial OXPHOS machinery, originated inside the megakaryocyte, and that this metabolism plays a pivotal role in platelet activation. This work represents a further example of the existence of an extramitochondrial aerobic metabolism, which can contribute to the cellular energy balance. © 2018 Société Française des Microscopies and Société de Biologie Cellulaire de France. Published by John Wiley & Sons Ltd.

The antioxidant uncoupling protein 2 stimulates hnRNPA2/B1, GLUT1 and PKM2 expression and sensitizes pancreas cancer cells to glycolysis inhibition.

PubMed

Brandi, Jessica; Cecconi, Daniela; Cordani, Marco; Torrens-Mas, Margalida; Pacchiana, Raffaella; Dalla Pozza, Elisa; Butera, Giovanna; Manfredi, Marcello; Marengo, Emilio; Oliver, Jordi; Roca, Pilar; Dando, Ilaria; Donadelli, Massimo

2016-12-01

Several evidence indicate that metabolic alterations play a pivotal role in cancer development. Here, we report that the mitochondrial uncoupling protein 2 (UCP2) sustains the metabolic shift from mitochondrial oxidative phosphorylation (mtOXPHOS) to glycolysis in pancreas cancer cells. Indeed, we show that UCP2 sensitizes pancreas cancer cells to the treatment with the glycolytic inhibitor 2-deoxy-D-glucose. Through a bidimensional electrophoresis analysis, we identify 19 protein species differentially expressed after treatment with the UCP2 inhibitor genipin and, by bioinformatic analyses, we show that these proteins are mainly involved in metabolic processes. In particular, we demonstrate that the antioxidant UCP2 induces the expression of hnRNPA2/B1, which is involved in the regulation of both GLUT1 and PKM2 mRNAs, and of lactate dehydrogenase (LDH) increasing the secretion of L-lactic acid. We further demonstrate that the radical scavenger N-acetyl-L-cysteine reverts hnRNPA2/B1 and PKM2 inhibition by genipin indicating a role for reactive oxygen species in the metabolic reprogramming of cancer cells mediated by UCP2. We also observe an UCP2-dependent decrease in mtOXPHOS complex I (NADH dehydrogenase), complex IV (cytochrome c oxidase), complex V (ATPase) and in mitochondrial oxygen consumption, suggesting a role for UCP2 in the counteraction of pancreatic cancer cellular respiration. All these results reveal novel mechanisms through which UCP2 promotes cancer cell proliferation with the concomitant metabolic shift from mtOXPHOS to the glycolytic pathway. Copyright © 2016 Elsevier Inc. All rights reserved.

PDE 5 inhibitor improves insulin sensitivity by enhancing mitochondrial function in adipocytes.

PubMed

Yu, Hea Min; Chung, Hyo Kyun; Kim, Koon Soon; Lee, Jae Min; Hong, Jun Hwa; Park, Kang Seo

2017-11-04

Adipocytes are involved in many metabolic disorders. It was recently reported that phosphodiesterase type 5 (PDE5) is expressed in human adipose tissue. In addition, PDE5 inhibitors have been shown to improve insulin sensitivity in humans. However, the mechanism underlying the role of PDE5 inhibitors as an insulin sensitizer remains largely unknown. The present study was undertaken to investigate the role of the PDE5 inhibitor udenafil in insulin signaling in adipocytes and whether this is mediated through the regulation of mitochondrial function. To study the mechanism underlying the insulin sensitizing action of PDE5 inhibitors, we evaluated quantitative changes in protein or mRNA levels of mitochondrial oxidative phosphorylation (OxPhos) complex, oxygen consumption rate (OCR), and fatty acid oxidation with varying udenafil concentrations in 3T3-L1 cells. Our cell study suggested that udenafil enhanced the insulin signaling pathway in 3T3-L1 cells. Following udenafil treatment, basal mitochondrial OCR, maximal OxPhos capacity, and OxPhos gene expression significantly increased. Finally, we examined whether udenafil can affect the fatty acid oxidation process. Treatment of 3T3-L1 cells with udenafil (10 and 20 μM) significantly increased fatty acid oxidation rate in a dose-dependent manner. In addition, the expression of peroxisome proliferator-activated receptor gamma coactivator 1-alpha (PGC-1α) significantly increased. We demonstrated that the PDE5 inhibitor udenafil enhances insulin sensitivity by improving mitochondrial function in 3T3-L1 cells. This might be the mechanism underlying the PDE5 inhibitor-enhanced insulin signaling in adipocytes. This also suggests that udenafil may provide benefit in the treatment of type 2 diabetes and other related cardiovascular diseases. Copyright © 2017 Elsevier Inc. All rights reserved.

GILZ overexpression attenuates endoplasmic reticulum stress-mediated cell death via the activation of mitochondrial oxidative phosphorylation

DOE Office of Scientific and Technical Information (OSTI.GOV)

André, Fanny; Corazao-Rozas, Paola; Idziorek, Thierry

The Glucocorticoïd-induced leucine zipper (GILZ) protein has profound anti-inflammatory activities in haematopoietic cells. GILZ regulates numerous signal transduction pathways involved in proliferation and survival of normal and neoplastic cells. Here, we have demonstrated the potential of GILZ in alleviating apoptosis induced by ER stress inducers. Whereas the glucocorticoid, dexamethasone, protects from tunicamycin-induced cell death, silencing endogeneous GILZ in dexamethasone-treated cancer cells alter the capacity of glucocorticoids to protect from tunicamycin-mediated apoptosis. Under ER stress conditions, overexpression of GILZ significantly reduced activation of mitochondrial pathway of apoptosis by maintaining Bcl-xl level. GILZ protein affects the UPR signaling shifting the balance towardsmore » pro-survival signals as judged by down-regulation of CHOP, ATF4, XBP1s mRNA and increase in GRP78 protein level. Interestingly, GILZ sustains high mitochondrial OXPHOS during ER stress and cytoprotection mediated by GILZ is abolished in cells depleted of mitochondrial DNA, which are OXPHOS-deficient. These findings reveal a new role of GILZ, which acts as a cytoprotector against ER stress through a pathway involving mitochondrial OXPHOS. - Highlights: • GILZ attenuates apoptotic cell death induced by ER stress conditions. • GILZ promotes pro-survival signaling of the UPR. • GILZ overexpression sustains high mitochondrial activity under ER stress. • Mitochondrial OXPHOX is required for GILZ protective effects against ER stress-mediated apoptosis.« less

Exercise in claudicants increase or decrease walking ability and the response relates to mitochondrial function.

PubMed

van Schaardenburgh, Michel; Wohlwend, Martin; Rognmo, Øivind; Mattsson, Erney J R

2017-06-07

Exercise of patients with intermittent claudication improves walking performance. Exercise does not usually increase blood flow, but seems to increase muscle mitochondrial enzyme activities. Although exercise is beneficial in most patients, it might be harmful in some. The mitochondrial response to exercise might therefore differ between patients. Our hypothesis was that changes in walking performance relate to changes in mitochondrial function after 8 weeks of exercise. At a subgroup level, negative responders decrease and positive responders increase mitochondrial capacity. Two types of exercise were studied, calf raising and walking (n = 28). We wanted to see whether there were negative and positive responders, independent of type of exercise. Measurements of walking performance, peripheral hemodynamics, mitochondrial respiration and content (citrate synthase activity) were obtained on each patient before and after the intervention period. Multiple linear regression was used to test whether changes in peak walking time relate to mitochondrial function. Subgroups of negative (n = 8) and positive responders (n = 8) were defined as those that either decreased or increased peak walking time following exercise. Paired t test and analysis of covariance was used to test changes within and between subgroups. Changes in peak walking time were related to changes in mitochondrial respiration supported by electron transferring flavoprotein (ETF + CI) P (p = 0.004), complex I (CI + ETF) P (p = 0.003), complex I + complex II (CI + CII + ETF) P (p = 0.037) and OXPHOS coupling efficiency (p = 0.046) in the whole group. Negative responders had more advanced peripheral arterial disease. Mitochondrial respiration supported by electron transferring flavoprotein (ETF + CI) P (p = 0.0013), complex I (CI + ETF) P (p = 0.0005), complex I + complex II (CI + CII + ETF) P (p = 0.011) and electron transfer system capacity (CI + CII + ETF) E (p = 0.021) and OXPHOS coupling efficiency decreased in negative responders (p = 0.0007) after exercise. Positive responders increased citrate synthase activity (p = 0.010). Changes in walking performance seem to relate to changes in mitochondrial function after exercise. Negative responders have more advanced peripheral arterial disease and decrease, while positive responders increase mitochondrial capacity. Trial registration ClinicalTrials.gov ID: NCT023110256.

Mitochondrial proteomic profile of complex IV deficiency fibroblasts: rearrangement of oxidative phosphorylation complex/supercomplex and other metabolic pathways.

PubMed

Salvador-Severo, Karina; Gómez-Caudillo, Leopoldo; Quezada, Héctor; García-Trejo, José de Jesús; Cárdenas-Conejo, Alan; Vázquez-Memije, Martha Elisa; Minauro-Sanmiguel, Fernando

Mitochondriopathies are multisystem diseases affecting the oxidative phosphorylation (OXPHOS) system. Skin fibroblasts are a good model for the study of these diseases. Fibroblasts with a complex IV mitochondriopathy were used to determine the molecular mechanism and the main affected functions in this disease. Skin fibroblast were grown to assure disease phenotype. Mitochondria were isolated from these cells and their proteome extracted for protein identification. Identified proteins were validated with the MitoMiner database. Disease phenotype was corroborated on skin fibroblasts, which presented a complex IV defect. The mitochondrial proteome of these cells showed that the most affected proteins belonged to the OXPHOS system, mainly to the complexes that form supercomplexes or respirosomes (I, III, IV, and V). Defects in complex IV seemed to be due to assembly issues, which might prevent supercomplexes formation and efficient substrate channeling. It was also found that this mitochondriopathy affects other processes that are related to DNA genetic information flow (replication, transcription, and translation) as well as beta oxidation and tricarboxylic acid cycle. These data, as a whole, could be used for the better stratification of these diseases, as well as to optimize management and treatment options. Copyright © 2017 Hospital Infantil de México Federico Gómez. Publicado por Masson Doyma México S.A. All rights reserved.

Combined defects in oxidative phosphorylation and fatty acid β-oxidation in mitochondrial disease

PubMed Central

Nsiah-Sefaa, Abena; McKenzie, Matthew

2016-01-01

Mitochondria provide the main source of energy to eukaryotic cells, oxidizing fats and sugars to generate ATP. Mitochondrial fatty acid β-oxidation (FAO) and oxidative phosphorylation (OXPHOS) are two metabolic pathways which are central to this process. Defects in these pathways can result in diseases of the brain, skeletal muscle, heart and liver, affecting approximately 1 in 5000 live births. There are no effective therapies for these disorders, with quality of life severely reduced for most patients. The pathology underlying many aspects of these diseases is not well understood; for example, it is not clear why some patients with primary FAO deficiencies exhibit secondary OXPHOS defects. However, recent findings suggest that physical interactions exist between FAO and OXPHOS proteins, and that these interactions are critical for both FAO and OXPHOS function. Here, we review our current understanding of the interactions between FAO and OXPHOS proteins and how defects in these two metabolic pathways contribute to mitochondrial disease pathogenesis. PMID:26839416

Mitochondrial-nuclear crosstalk, haplotype and copy number variation distinct in muscle fiber type, mitochondrial respiratory and metabolic enzyme activities.

PubMed

Liu, Xuan; Trakooljul, Nares; Hadlich, Frieder; Murani, Eduard; Wimmers, Klaus; Ponsuksili, Siriluck

2017-10-25

Genes expressed in mitochondria work in concert with those expressed in the nucleus to mediate oxidative phosphorylation (OXPHOS), a process that is relevant for muscle metabolism and meat quality. Mitochondrial genome activity can be efficiently studied and compared in Duroc and Pietrain pigs, which harbor different mitochondrial haplotypes and distinct muscle fiber types, mitochondrial respiratory activities, and fat content. Pietrain pigs homozygous-positive for malignant hyperthermia susceptibility (PiPP) carried only haplotype 8 and showed the lowest absolute mtDNA copy number accompanied by a decrease transcript abundance of mitochondrial-encoded subunits ND1, ND6, and ATP6 and nuclear-encoded subunits NDUFA11 and NDUFB8. In contrast, we found that haplotype 4 of Duroc pigs had significantly higher mitochondrial DNA (mtDNA) copy numbers and an increase transcript abundance of mitochondrial-encoded subunits ND1, ND6, and ATP6. These results suggest that the variation in mitochondrial and nuclear genetic background among these animals has an effect on mitochondrial content and OXPHOS system subunit expression. We observed the co-expression pattern of mitochondrial and nuclear encoded OXPHOS subunits suggesting that the mitochondrial-nuclear crosstalk functionally involves in muscle metabolism. The findings provide valuable information for understanding muscle biology processes and energy metabolism, and may direct use for breeding strategies to improve meat quality and animal health.

Influence of endurance training on skeletal muscle mitophagy regulatory proteins in type 2 diabetic men.

PubMed

Brinkmann, Christian; Przyklenk, Axel; Metten, Alexander; Schiffer, Thorsten; Bloch, Wilhelm; Brixius, Klara; Gehlert, Sebastian

2017-11-01

Mitophagy is a form of autophagy for the elimination of mitochondria. Mitochondrial content and function are reduced in the skeletal muscle of patients with type 2 diabetes mellitus (T2DM). Physical training has been shown to restore mitochondrial capacity in T2DM patients, but the role of mitophagy has not been examined in this context. This study analyzes the impact of a 3-month endurance training on important skeletal muscle mitophagy regulatory proteins and oxidative phosphorylation (OXPHOS) complexes in T2DM patients. Muscle biopsies were obtained from eight overweight/obese T2DM men (61±10 years) at T1 (6 weeks pre-training), T2 (1 week pre-training), and T3 (3 to 4 days post-training). Protein contents were determined by Western blotting. The training increased mitochondrial complex II significantly (T2-T3: +29%, p = 0.037). The protein contents of mitophagy regulatory proteins (phosphorylated form of forkhead box O3A (pFOXO3A), mitochondrial E3 ubiquitin protein ligase-1 (MUL1), Bcl-2/adenovirus E1B 19-kD interacting protein-3 (BNIP3), microtubule-associated protein 1 light chain-3B (the ratio LC3B-II/LC3B-I was determined)) did not differ significantly between T1, T2, and T3. The results imply that training-induced changes in OXPHOS subunits (significant increase in complex II) are not accompanied by changes in mitophagy regulatory proteins in T2DM men. Future studies should elucidate whether acute exercise might affect mitophagic processes in T2DM patients (and whether a transient regulation of mitophagy regulatory proteins is evident) to fully clarify the role of physical activity and mitophagy for mitochondrial health in this particular patient group.

Toxoplasma gondii Infection Is Associated with Mitochondrial Dysfunction in-Vitro

PubMed Central

Syn, Genevieve; Anderson, Denise; Blackwell, Jenefer M.; Jamieson, Sarra E.

2017-01-01

Upon invasion of host cells, the ubiquitous pathogen Toxoplasma gondii manipulates several host processes, including re-organization of host organelles, to create a replicative niche. Host mitochondrial association to T. gondii parasitophorous vacuoles is rapid and has roles in modulating host immune responses. Here gene expression profiling of T. gondii infected cells reveals enrichment of genes involved in oxidative phosphorylation (OXPHOS) and mitochondrial dysfunction 6 h post-infection. We identified 11 hub genes (HIF-1α, CASP8, FN1, POU5F1, CD44, ISG15, HNRNPA1, MDM2, RPL35, VHL, and NUPR1) and 10 predicted upstream regulators, including 4 endogenous regulators RICTOR, KDM5A, RB1, and D-glucose. We characterized a number of mitochondrial parameters in T. gondii infected human foreskin fibroblast cells over a 36 h time-course. In addition to the usual rapid recruitment and apparent enlargement of mitochondria around the parasitophorous vacuole we observed fragmented host mitochondria in infected cells, not linked to cellular apoptosis, from 24 h post-infection. An increase in mitochondrial superoxide levels in T. gondii infected cells was observed that required active parasite invasion and peaked at 30 h post-infection. Measurement of OXPHOS proteins showed decreased expression of Complex IV in infected cells at 24 h post-infection, followed by decreased expression of Complexes I and II at 36 h post-infection. No change occurred in Complex V. No difference in host mitochondrial membrane potential between infected and mock-infected cells was observed at any time. Our results show perturbation of host mitochondrial function following T. gondii infection that likely impacts on pathogenesis of disease. PMID:29312892

Defining the action spectrum of potential PGC-1α activators on a mitochondrial and cellular level in vivo.

PubMed

Hofer, Annette; Noe, Natalie; Tischner, Christin; Kladt, Nikolay; Lellek, Veronika; Schauß, Astrid; Wenz, Tina

2014-05-01

Previous studies have demonstrated a therapeutic benefit of pharmaceutical PGC-1α activation in cellular and murine model of disorders linked to mitochondrial dysfunction. While in some cases, this effect seems to be clearly associated with boosting of mitochondrial function, additional alterations as well as tissue- and cell-type-specific effects might play an important role. We initiated a comprehensive analysis of the effects of potential PGC-1α-activating drugs and pharmaceutically targeted the PPAR (bezafibrate, rosiglitazone), AMPK (AICAR, metformin) and Sirt1 (resveratrol) pathways in HeLa cells, neuronal cells and PGC-1α-deficient MEFs to get insight into cell type specificity and PGC-1α dependence of their working action. We used bezafibrate as a model drug to assess the effect on a tissue-specific level in a murine model. Not all analyzed drugs activate the PGC pathway or alter mitochondrial protein levels. However, they all affect supramolecular assembly of OXPHOS complexes and OXPHOS protein stability. In addition, a clear drug- and cell-type-specific influence on several cellular stress pathways as well as on post-translational modifications could be demonstrated, which might be relevant to fully understand the action of the analyzed drugs in the disease state. Importantly, the effect on the activation of mitochondrial biogenesis and stress response program upon drug treatment is PGC-1α dependent in MEFs demonstrating not only the pleiotropic effects of this molecule but points also to the working mechanism of the analyzed drugs. The definition of the action spectrum of the different drugs forms the basis for a defect-specific compensation strategy and a future personalized therapeutic approach.

The First Chameleon Transcriptome: Comparative Genomic Analysis of the OXPHOS System Reveals Loss of COX8 in Iguanian Lizards

PubMed Central

Bar-Yaacov, Dan; Bouskila, Amos; Mishmar, Dan

2013-01-01

Recently, we found dramatic mitochondrial DNA divergence of Israeli Chamaeleo chamaeleon populations into two geographically distinct groups. We aimed to examine whether the same pattern of divergence could be found in nuclear genes. However, no genomic resource is available for any chameleon species. Here we present the first chameleon transcriptome, obtained using deep sequencing (SOLiD). Our analysis identified 164,000 sequence contigs of which 19,000 yielded unique BlastX hits. To test the efficacy of our sequencing effort, we examined whether the chameleon and other available reptilian transcriptomes harbored complete sets of genes comprising known biochemical pathways, focusing on the nDNA-encoded oxidative phosphorylation (OXPHOS) genes as a model. As a reference for the screen, we used the human 86 (including isoforms) known structural nDNA-encoded OXPHOS subunits. Analysis of 34 publicly available vertebrate transcriptomes revealed orthologs for most human OXPHOS genes. However, OXPHOS subunit COX8 (Cytochrome C oxidase subunit 8), including all its known isoforms, was consistently absent in transcriptomes of iguanian lizards, implying loss of this subunit during the radiation of this suborder. The lack of COX8 in the suborder Iguania is intriguing, since it is important for cellular respiration and ATP production. Our sequencing effort added a new resource for comparative genomic studies, and shed new light on the evolutionary dynamics of the OXPHOS system. PMID:24009133

The first Chameleon transcriptome: comparative genomic analysis of the OXPHOS system reveals loss of COX8 in Iguanian lizards.

PubMed

Bar-Yaacov, Dan; Bouskila, Amos; Mishmar, Dan

2013-01-01

Recently, we found dramatic mitochondrial DNA divergence of Israeli Chamaeleo chamaeleon populations into two geographically distinct groups. We aimed to examine whether the same pattern of divergence could be found in nuclear genes. However, no genomic resource is available for any chameleon species. Here we present the first chameleon transcriptome, obtained using deep sequencing (SOLiD). Our analysis identified 164,000 sequence contigs of which 19,000 yielded unique BlastX hits. To test the efficacy of our sequencing effort, we examined whether the chameleon and other available reptilian transcriptomes harbored complete sets of genes comprising known biochemical pathways, focusing on the nDNA-encoded oxidative phosphorylation (OXPHOS) genes as a model. As a reference for the screen, we used the human 86 (including isoforms) known structural nDNA-encoded OXPHOS subunits. Analysis of 34 publicly available vertebrate transcriptomes revealed orthologs for most human OXPHOS genes. However, OXPHOS subunit COX8 (Cytochrome C oxidase subunit 8), including all its known isoforms, was consistently absent in transcriptomes of iguanian lizards, implying loss of this subunit during the radiation of this suborder. The lack of COX8 in the suborder Iguania is intriguing, since it is important for cellular respiration and ATP production. Our sequencing effort added a new resource for comparative genomic studies, and shed new light on the evolutionary dynamics of the OXPHOS system.

Mitochondrial genes are altered in blood early in Alzheimer's disease.

PubMed

Lunnon, Katie; Keohane, Aoife; Pidsley, Ruth; Newhouse, Stephen; Riddoch-Contreras, Joanna; Thubron, Elisabeth B; Devall, Matthew; Soininen, Hikka; Kłoszewska, Iwona; Mecocci, Patrizia; Tsolaki, Magda; Vellas, Bruno; Schalkwyk, Leonard; Dobson, Richard; Malik, Afshan N; Powell, John; Lovestone, Simon; Hodges, Angela

2017-05-01

Although mitochondrial dysfunction is a consistent feature of Alzheimer's disease in the brain and blood, the molecular mechanisms behind these phenomena are unknown. Here we have replicated our previous findings demonstrating reduced expression of nuclear-encoded oxidative phosphorylation (OXPHOS) subunits and subunits required for the translation of mitochondrial-encoded OXPHOS genes in blood from people with Alzheimer's disease and mild cognitive impairment. Interestingly this was accompanied by increased expression of some mitochondrial-encoded OXPHOS genes, namely those residing closest to the transcription start site of the polycistronic heavy chain mitochondrial transcript (MT-ND1, MT-ND2, MT-ATP6, MT-CO1, MT-CO2, MT-C03) and MT-ND6 transcribed from the light chain. Further we show that mitochondrial DNA copy number was unchanged suggesting no change in steady-state numbers of mitochondria. We suggest that an imbalance in nuclear and mitochondrial genome-encoded OXPHOS transcripts may drive a negative feedback loop reducing mitochondrial translation and compromising OXPHOS efficiency, which is likely to generate damaging reactive oxygen species. Copyright © 2017 The Author(s). Published by Elsevier Inc. All rights reserved.

Contribution of proton leak to oxygen consumption in skeletal muscle during intense exercise is very low despite large contribution at rest

PubMed Central

2017-01-01

A computer model was used to simulate the dependence of protonmotive force (Δp), proton leak and phenomenological (involving proton leak) ATP/O2 ratio on work intensity in skeletal muscle. Δp, NADH and proton leak decreased with work intensity. The contribution of proton leak to oxygen consumption (V˙O2) decreased from about 60% at rest to about 3 and 1% at moderate and heavy/severe exercise, respectively, while the ATP/O2 ratio increased from 2.1 to 5.5 and 5.7. A two-fold increase in proton leak activity or its decrease to zero decreased/increased the ATP/O2 ratio by only about 3 and 1% during moderate and heavy/severe exercise, respectively. The low contribution of proton leak to V˙O2 in intensively working skeletal muscle was mostly caused by a huge increase in ATP usage intensity during rest-to-work transition, while OXPHOS, and thus oxidative ATP supply and V˙O2 related to it, was mostly stimulated by high each-step activation (ESA) of OXPHOS complexes. The contribution of proton leak to V˙O2 and ATP/O2 ratio in isolated mitochondria should not be directly extrapolated to working muscle, as mitochondria lack ESA, at least in the absence of Ca2+, and therefore V˙O2 cannot be elevated as much as in intact muscle. PMID:29045413

Contribution of proton leak to oxygen consumption in skeletal muscle during intense exercise is very low despite large contribution at rest.

PubMed

Korzeniewski, Bernard

2017-01-01

A computer model was used to simulate the dependence of protonmotive force (Δp), proton leak and phenomenological (involving proton leak) ATP/O2 ratio on work intensity in skeletal muscle. Δp, NADH and proton leak decreased with work intensity. The contribution of proton leak to oxygen consumption ([Formula: see text]) decreased from about 60% at rest to about 3 and 1% at moderate and heavy/severe exercise, respectively, while the ATP/O2 ratio increased from 2.1 to 5.5 and 5.7. A two-fold increase in proton leak activity or its decrease to zero decreased/increased the ATP/O2 ratio by only about 3 and 1% during moderate and heavy/severe exercise, respectively. The low contribution of proton leak to [Formula: see text] in intensively working skeletal muscle was mostly caused by a huge increase in ATP usage intensity during rest-to-work transition, while OXPHOS, and thus oxidative ATP supply and [Formula: see text] related to it, was mostly stimulated by high each-step activation (ESA) of OXPHOS complexes. The contribution of proton leak to [Formula: see text] and ATP/O2 ratio in isolated mitochondria should not be directly extrapolated to working muscle, as mitochondria lack ESA, at least in the absence of Ca2+, and therefore [Formula: see text] cannot be elevated as much as in intact muscle.

Combined targeting of PDK1 and EGFR triggers regression of glioblastoma by reversing the Warburg effect.

PubMed

Velpula, Kiran Kumar; Bhasin, Arnima; Asuthkar, Swapna; Tsung, Andrew J

2013-12-15

Glioblastoma multiforme is the most aggressive primary brain tumor in adults. Overexpression of the EGF receptor (EGFR) is recognized as a widespread oncogenic signature in glioblastoma multiforme, but the complexity of its contributions is not fully understood, nor the most effective ways to leverage anti-EGFR therapy in this setting. Hypoxia is known to drive the aggressive character of glioblastoma multiforme by promoting aerobic glycolysis rather than pyruvate oxidation carried out in mitochondria (OXPHOS), a phenomenon termed the Warburg effect, which is a general feature of oncogenesis. In this study, we report that hypoxia drives expression of the pyruvate dehydrogenase kinase (PDK1) and EGFR along with the hypoxia-inducing factor (HIF)-1α in human glioblastoma multiforme cells. PDK1 is a HIF-1-regulated gene and our findings indicated that hypoxia-induced PDK1 expression may promote EGFR activation, initiating a feed-forward loop that can sustain malignant progression. RNAi-mediated attenuation of PDK1 and EGFR lowered PDK1-EGFR activation and decreased HIF-1α expression, shifting the Warburg phenotype to OXPHOS and inhibiting glioblastoma multiforme growth and proliferation. In clinical specimens of glioblastoma multiforme, we found that immunohistochemical expression of PDK1, EGFR, and HIF-1α were elevated in glioblastoma multiforme specimens when compared with normal brain tissues. Collectively, our studies establish PDK1 as a key driver and candidate therapeutic target in glioblastoma multiforme. ©2013 AACR.

c-Myc and AMPK Control Cellular Energy Levels by Cooperatively Regulating Mitochondrial Structure and Function

PubMed Central

Edmunds, Lia R.; Sharma, Lokendra; Wang, Huabo; Kang, Audry; d’Souza, Sonia; Lu, Jie; McLaughlin, Michael; Dolezal, James M.; Gao, Xiaoli; Weintraub, Susan T.; Ding, Ying; Zeng, Xuemei; Yates, Nathan; Prochownik, Edward V.

2015-01-01

The c-Myc (Myc) oncoprotein and AMP-activated protein kinase (AMPK) regulate glycolysis and oxidative phosphorylation (Oxphos) although often for different purposes. Because Myc over-expression depletes ATP with the resultant activation of AMPK, we explored the potential co-dependency of and cross-talk between these proteins by comparing the consequences of acute Myc induction in ampk+/+ (WT) and ampk-/- (KO) murine embryo fibroblasts (MEFs). KO MEFs showed a higher basal rate of glycolysis than WT MEFs and an appropriate increase in response to activation of a Myc-estrogen receptor (MycER) fusion protein. However, KO MEFs had a diminished ability to increase Oxphos, mitochondrial mass and reactive oxygen species in response to MycER activation. Other differences between WT and KO MEFs, either in the basal state or following MycER induction, included abnormalities in electron transport chain function, levels of TCA cycle-related oxidoreductases and cytoplasmic and mitochondrial redox states. Transcriptional profiling of pathways pertinent to glycolysis, Oxphos and mitochondrial structure and function also uncovered significant differences between WT and KO MEFs and their response to MycER activation. Finally, an unbiased mass-spectrometry (MS)-based survey capable of quantifying ~40% of all mitochondrial proteins, showed about 15% of them to be AMPK- and/or Myc-dependent in their steady state. Significant differences in the activities of the rate-limiting enzymes pyruvate kinase and pyruvate dehydrogenase, which dictate pyruvate and acetyl coenzyme A abundance, were also differentially responsive to Myc and AMPK and could account for some of the differences in basal metabolite levels that were also detected by MS. Thus, Myc and AMPK are highly co-dependent and appear to engage in significant cross-talk across numerous pathways which support metabolic and ATP-generating functions. PMID:26230505

Balancing glycolysis and mitochondrial OXPHOS: lessons from the hematopoietic system and exercising muscles.

PubMed

Haran, Michal; Gross, Atan

2014-11-01

Living organisms require a constant supply of safe and efficient energy to maintain homeostasis and to allow locomotion of single cells, tissues and the entire organism. The source of energy can be glycolysis, a simple series of enzymatic reactions in the cytosol, or a much more complex process in the mitochondria, oxidative phosphorylation (OXPHOS). In this review we will examine how does the organism balance its source of energy in two seemingly distinct and unrelated processes: hematopoiesis and exercise. In both processes we will show the importance of the metabolic program and its regulation. We will also discuss the importance of oxygen availability not as a sole determinant, but in the context of the nutrient and cellular state, and address the emerging role of lactate as an energy source and signaling molecule in health and disease. Copyright © 2014 Elsevier B.V. and Mitochondria Research Society. All rights reserved.

Autocrine IL-10 functions as a rheostat for M1 macrophage glycolytic commitment by tuning nitric oxide production.

PubMed

Baseler, Walter A; Davies, Luke C; Quigley, Laura; Ridnour, Lisa A; Weiss, Jonathan M; Hussain, S Perwez; Wink, David A; McVicar, Daniel W

2016-12-01

Inflammatory maturation of M1 macrophages by proinflammatory stimuli such as toll like receptor ligands results in profound metabolic reprogramming resulting in commitment to aerobic glycolysis as evidenced by repression of mitochondrial oxidative phosphorylation (OXPHOS) and enhanced glucose utilization. In contrast, "alternatively activated" macrophages adopt a metabolic program dominated by fatty acid-fueled OXPHOS. Despite the known importance of these developmental stages on the qualitative aspects of an inflammatory response, relatively little is know regarding the regulation of these metabolic adjustments. Here we provide evidence that the immunosuppressive cytokine IL-10 defines a metabolic regulatory loop. Our data show for the first time that lipopolysaccharide (LPS)-induced glycolytic flux controls IL-10-production via regulation of mammalian target of rapamycin (mTOR) and that autocrine IL-10 in turn regulates macrophage nitric oxide (NO) production. Genetic and pharmacological manipulation of IL-10 and nitric oxide (NO) establish that metabolically regulated autocrine IL-10 controls glycolytic commitment by limiting NO-mediated suppression of OXPHOS. Together these data support a model where autocine IL-10 production is controlled by glycolytic flux in turn regulating glycolytic commitment by preserving OXPHOS via suppression of NO. We propose that this IL-10-driven metabolic rheostat maintains metabolic equilibrium during M1 macrophage differentiation and that perturbation of this regulatory loop, either directly by exogenous cellular sources of IL-10 or indirectly via limitations in glucose availability, skews the cellular metabolic program altering the balance between inflammatory and immunosuppressive phenotypes. Copyright © 2016. Published by Elsevier B.V.

MITOCHONDRIAL DISEASES PART II: MOUSE MODELS OF OXPHOS DEFICIENCIES CAUSED BY DEFECTS IN REGULATORY FACTORS AND OTHER COMPONENTS REQUIRED FOR MITOCHONDRIAL FUNCTION

PubMed Central

Iommarini, Luisa; Peralta, Susana; Torraco, Alessandra; Diaz, Francisca

2015-01-01

Mitochondrial disorders are defined as defects that affect the oxidative phosphorylation system (OXPHOS). They are characterized by a heterogeneous array of clinical presentations due in part to a wide variety of factors required for proper function of the components of the OXPHOS system. There is no cure for these disorders owing our poor knowledge of the pathogenic mechanisms of disease. To understand the mechanisms of human disease numerous mouse models have been developed in recent years. Here we summarize the features of several mouse models of mitochondrial diseases directly related to those factors affecting mtDNA maintenance, replication, transcription, translation as well to other proteins that are involved in mitochondrial dynamics and quality control which affect mitochondrial OXPHOS function without been intrinsic components of the system. We discuss how these models have contributed to our understanding of mitochondrial diseases and their pathogenic mechanisms. PMID:25640959

Beyond AICA Riboside: In Search of New Specific AMP-activated Protein Kinase Activators

PubMed Central

Guigas, Bruno; Sakamoto, Kei; Taleux, Nellie; Reyna, Sara M.; Musi, Nicolas; Viollet, Benoit; Hue, Louis

2010-01-01

Summary 5-Aminoimidazole-4-carboxamide-1-β-d-ribofuranoside (AICA riboside) has been extensively used in vitro and in vivo to activate the AMP-activated protein kinase (AMPK), a metabolic sensor involved in both cellular and whole body energy homeostasis. However, it has been recently highlighted that AICA riboside also exerts AMPK-independent effects, mainly on AMP-regulated enzymes and mitochondrial oxidative phosphorylation (OXPHOS), leading to the conclusion that new compounds with reduced off target effects are needed to specifically activate AMPK. Here, we review recent findings on newly discovered AMPK activators, notably on A-769662, a nonnucleoside compound from the thienopyridone family. We also report that A-769662 is able to activate AMPK and stimulate glucose uptake in both L6 cells and primary myotubes derived from human satellite cells. In addition, A-769662 increases AMPK activity and phosphorylation of its main downstream targets in primary cultured rat hepatocytes but, by contrast with AICA riboside, does neither affect mitochondrial OXPHOS nor change cellular AMP:ATP ratio. We conclude that A-769662 could be one of the new promising chemical agents to activate AMPK with limited AMPK-independent side effects. PMID:18798311

Assessment of mitochondrial dysfunction-related, drug-induced hepatotoxicity in primary rat hepatocytes

DOE Office of Scientific and Technical Information (OSTI.GOV)

Liu, Cong; Sekine, Shuichi, E-mail: ssekine@facult

Evidence that mitochondrial dysfunction plays a central role in drug-induced liver injury is rapidly accumulating. In contrast to physiological conditions, in which almost all adenosine triphosphate (ATP) in hepatocytes is generated in mitochondria via aerobic respiration, the high glucose content and limited oxygen supply of conventional culture systems force primary hepatocytes to generate most ATP via cytosolic glycolysis. Thus, such anaerobically poised cells are resistant to xenobiotics that impair mitochondrial function, and are not suitable to identify drugs with mitochondrial liabilities. In this study, primary rat hepatocytes were cultured in galactose-based medium, instead of the conventional glucose-based medium, and inmore » hyperoxia to improve the reliance of energy generation on aerobic respiration. Activation of mitochondria was verified by diminished cellular lactate release and increased oxygen consumption. These conditions improved sensitivity to the mitochondrial complex I inhibitor rotenone. Since oxidative stress is also a general cause of mitochondrial impairment, cells were exposed to test compounds in the presence of transferrin to increase the generation of reactive oxygen species via increased uptake of iron. Finally, 14 compounds with reported mitochondrial liabilities were tested to validate this new drug-induced mitochondrial toxicity assay. Overall, the culture of primary rat hepatocytes in galactose, hyperoxia and transferrin is a useful model for the identification of mitochondrial dysfunction-related drug-induced hepatotoxicity. - Highlights: • Drug-induced mitochondrial toxicity was evaluated using primary rat hepatocytes. • Galactose and hyperoxia could activate OXPHOS in primary rat hepatocytes. • Cells with enhanced OXPHOS exhibit improved sensitivity to mitochondrial toxins. • Transferrin potentiate mitochondrial toxicity via increased ROS production.« less

Melatonin-induced increase of lipid droplets accumulation and in vitro maturation in porcine oocytes is mediated by mitochondrial quiescence.

PubMed

He, Bin; Yin, Chao; Gong, Yabin; Liu, Jie; Guo, Huiduo; Zhao, Ruqian

2018-01-01

Melatonin, the major pineal secretory product, has a significant impact on the female reproductive system. Recently, the beneficial effects of melatonin on mammalian oocyte maturation and embryonic development have drawn increased attention. However, the exact underlying mechanisms remain to be fully elucidated. This study demonstrates that supplementing melatonin to in vitro maturation (IVM) medium enhances IVM rate, lipid droplets (LDs) accumulation as well as triglyceride content in porcine oocytes. Decrease of mitochondrial membrane potential, mitochondrial respiratory chain complex IV activity as well as mitochondrial reactive oxygen species (mROS) content indicated that melatonin induced a decrease of mitochondrial activity. The copy number of mitochondrial DNA (mtDNA) which encodes essential subunits of oxidative phosphorylation (OXPHOS), was not affected by melatonin. However, the expression of mtDNA-encoded genes was significantly down-regulated after melatonin treatment. The DNA methyltransferase DNMT1, which regulates methylation and expression of mtDNA, was increased and translocated into the mitochondria in melatonin-treated oocytes. The inhibitory effect of melatonin on the expression of mtDNA was significantly prevented by simultaneous addition of DNMT1 inhibitor, which suggests that melatonin regulates the transcription of mtDNA through up-regulation of DNMT1 and mtDNA methylation. Increase of triglyceride contents after inhibition of OXPHOS indicated that mitochondrial quiescence is crucial for LDs accumulation in oocytes. Taken together, our results suggest that melatonin-induced reduction in mROS production and increase in IVM, and LDs accumulation in porcine oocytes is mediated by mitochondrial quiescence. © 2017 Wiley Periodicals, Inc.

Focal adhesion kinase-promoted tumor glucose metabolism is associated with a shift of mitochondrial respiration to glycolysis.

PubMed

Zhang, J; Gao, Q; Zhou, Y; Dier, U; Hempel, N; Hochwald, S N

2016-04-14

Cancer cells often gains a growth advantage by taking up glucose at a high rate and undergoing aerobic glycolysis through intrinsic cellular factors that reprogram glucose metabolism. Focal adhesion kinase (FAK), a key transmitter of growth factor and anchorage stimulation, is aberrantly overexpressed or activated in most solid tumors, including pancreatic ductal adenocarcinomas (PDACs). We determined whether FAK can act as an intrinsic driver to promote aerobic glycolysis and tumorigenesis. FAK inhibition decreases and overexpression increases intracellular glucose levels during unfavorable conditions, including growth factor deficiency and cell detachment. Amplex glucose assay, fluorescence and carbon-13 tracing studies demonstrate that FAK promotes glucose consumption and glucose-to-lactate conversion. Extracellular flux analysis indicates that FAK enhances glycolysis and decreases mitochondrial respiration. FAK increases key glycolytic proteins, including enolase, pyruvate kinase M2 (PKM2), lactate dehydrogenase and monocarboxylate transporter. Furthermore, active/tyrosine-phosphorylated FAK directly binds to PKM2 and promotes PKM2-mediated glycolysis. On the other hand, FAK-decreased levels of mitochondrial complex I can result in reduced oxidative phosphorylation (OXPHOS). Attenuation of FAK-enhanced glycolysis re-sensitizes cancer cells to growth factor withdrawal, decreases cell viability and reduces growth of tumor xenografts. These observations, for the first time, establish a vital role of FAK in cancer glucose metabolism through alterations in the OXPHOS-to-glycolysis balance. Broadly targeting the common phenotype of aerobic glycolysis and more specifically FAK-reprogrammed glucose metabolism will disrupt the bioenergetic and biosynthetic supply for uncontrolled growth of tumors, particularly glycolytic PDAC.

Disruption of oxidative phosphorylation (OXPHOS) by hydroxylated polybrominated diphenyl ethers (OH-PBDEs) present in the marine environment.

PubMed

Legradi, Jessica; Dahlberg, Anna-Karin; Cenijn, Peter; Marsh, Göran; Asplund, Lillemor; Bergman, Åke; Legler, Juliette

2014-12-16

Hydroxylated polybrominated diphenyl ethers (OH-PBDEs) are of growing concern, as they have been detected in both humans and wildlife and have been shown to be toxic. Recent studies have indicated that OH-PBDEs can be more toxic than PBDEs, partly due to their ability to disrupt oxidative phosphorylation (OXPHOS), an essential process in energy metabolism. In this study, we determined the OXPHOS disruption potential of 18 OH-PBDE congeners reported in marine wildlife using two in vitro bioassays, namely the classic rat mitochondrial respiration assay, and a mitochondrial membrane potential assay using zebrafish PAC2 cells. Single OH-PBDE congeners as well as mixtures were tested to study potential additive or synergistic effects. An environmental mixture composed of seven OH-PBDE congeners mimicking the concentrations reported in Baltic blue mussels were also studied. We report that all OH-PBDEs tested were able to disrupt OXPHOS via either protonophoric uncoupling and/or inhibition of the electron transport chain. Additionally we show that OH-PBDEs tested in combinations as found in the environment have the potential to disrupt OXPHOS. Importantly, mixtures of OH-PBDEs may show very strong synergistic effects, stressing the importance of further research on the in vivo impacts of these compounds in the environment.

Biallelic C1QBP Mutations Cause Severe Neonatal-, Childhood-, or Later-Onset Cardiomyopathy Associated with Combined Respiratory-Chain Deficiencies.

PubMed

Feichtinger, René G; Oláhová, Monika; Kishita, Yoshihito; Garone, Caterina; Kremer, Laura S; Yagi, Mikako; Uchiumi, Takeshi; Jourdain, Alexis A; Thompson, Kyle; D'Souza, Aaron R; Kopajtich, Robert; Alston, Charlotte L; Koch, Johannes; Sperl, Wolfgang; Mastantuono, Elisa; Strom, Tim M; Wortmann, Saskia B; Meitinger, Thomas; Pierre, Germaine; Chinnery, Patrick F; Chrzanowska-Lightowlers, Zofia M; Lightowlers, Robert N; DiMauro, Salvatore; Calvo, Sarah E; Mootha, Vamsi K; Moggio, Maurizio; Sciacco, Monica; Comi, Giacomo P; Ronchi, Dario; Murayama, Kei; Ohtake, Akira; Rebelo-Guiomar, Pedro; Kohda, Masakazu; Kang, Dongchon; Mayr, Johannes A; Taylor, Robert W; Okazaki, Yasushi; Minczuk, Michal; Prokisch, Holger

2017-10-05

Complement component 1 Q subcomponent-binding protein (C1QBP; also known as p32) is a multi-compartmental protein whose precise function remains unknown. It is an evolutionary conserved multifunctional protein localized primarily in the mitochondrial matrix and has roles in inflammation and infection processes, mitochondrial ribosome biogenesis, and regulation of apoptosis and nuclear transcription. It has an N-terminal mitochondrial targeting peptide that is proteolytically processed after import into the mitochondrial matrix, where it forms a homotrimeric complex organized in a doughnut-shaped structure. Although C1QBP has been reported to exert pleiotropic effects on many cellular processes, we report here four individuals from unrelated families where biallelic mutations in C1QBP cause a defect in mitochondrial energy metabolism. Infants presented with cardiomyopathy accompanied by multisystemic involvement (liver, kidney, and brain), and children and adults presented with myopathy and progressive external ophthalmoplegia. Multiple mitochondrial respiratory-chain defects, associated with the accumulation of multiple deletions of mitochondrial DNA in the later-onset myopathic cases, were identified in all affected individuals. Steady-state C1QBP levels were decreased in all individuals' samples, leading to combined respiratory-chain enzyme deficiency of complexes I, III, and IV. C1qbp -/- mouse embryonic fibroblasts (MEFs) resembled the human disease phenotype by showing multiple defects in oxidative phosphorylation (OXPHOS). Complementation with wild-type, but not mutagenized, C1qbp restored OXPHOS protein levels and mitochondrial enzyme activities in C1qbp -/- MEFs. C1QBP deficiency represents an important mitochondrial disorder associated with a clinical spectrum ranging from infantile lactic acidosis to childhood (cardio)myopathy and late-onset progressive external ophthalmoplegia. Copyright © 2017 The Authors. Published by Elsevier Inc. All rights reserved.

Dysfunctional oxidative phosphorylation makes malignant melanoma cells addicted to glycolysis driven by the (V600E)BRAF oncogene.

PubMed

Hall, Arnaldur; Meyle, Kathrine Damm; Lange, Marina Krarup; Klima, Martin; Sanderhoff, May; Dahl, Christina; Abildgaard, Cecilie; Thorup, Katrine; Moghimi, Seyed Moein; Jensen, Per Bo; Bartek, Jiri; Guldberg, Per; Christensen, Claus

2013-04-01

Oncogene addiction describes how cancer cells exhibit dependence on single oncogenes to escape apoptosis and senescence. While oncogene addiction constitutes the basis for new cancer treatment strategies targeting individual kinases and pathways activated by oncogenic mutations, the biochemical basis for this addiction is largely unknown. Here we provide evidence for a metabolic rationale behind the addiction to (V600E)BRAF in two malignant melanoma cell lines. Both cell lines display a striking addiction to glycolysis due to underlying dysfunction of oxidative phosphorylation (OXPHOS). Notably, even minor reductions in glycolytic activity lead to increased OXPHOS activity (reversed Warburg effect), however the mitochondria are unable to sustain ATP production. We show that (V600E)BRAF upholds the activity of glycolysis and therefore the addiction to glycolysis de facto becomes an addiction to (V600E)BRAF. Finally, the senescence response associated with inhibition of (V600E)BRAF is rescued by overexpression of glyceraldehyde-3-phosphate dehydrogenase (GAPDH), providing direct evidence that oncogene addiction rests on a metabolic foundation.

Mitochondrial free fatty acid β-oxidation supports oxidative phosphorylation and proliferation in cancer cells.

PubMed

Rodríguez-Enríquez, Sara; Hernández-Esquivel, Luz; Marín-Hernández, Alvaro; El Hafidi, Mohammed; Gallardo-Pérez, Juan Carlos; Hernández-Reséndiz, Ileana; Rodríguez-Zavala, José S; Pacheco-Velázquez, Silvia C; Moreno-Sánchez, Rafael

2015-08-01

Oxidative phosphorylation (OxPhos) is functional and sustains tumor proliferation in several cancer cell types. To establish whether mitochondrial β-oxidation of free fatty acids (FFAs) contributes to cancer OxPhos functioning, its protein contents and enzyme activities, as well as respiratory rates and electrical membrane potential (ΔΨm) driven by FFA oxidation were assessed in rat AS-30D hepatoma and liver (RLM) mitochondria. Higher protein contents (1.4-3 times) of β-oxidation (CPT1, SCAD) as well as proteins and enzyme activities (1.7-13-times) of Krebs cycle (KC: ICD, 2OGDH, PDH, ME, GA), and respiratory chain (RC: COX) were determined in hepatoma mitochondria vs. RLM. Although increased cholesterol content (9-times vs. RLM) was determined in the hepatoma mitochondrial membranes, FFAs and other NAD-linked substrates were oxidized faster (1.6-6.6 times) by hepatoma mitochondria than RLM, maintaining similar ΔΨm values. The contents of β-oxidation, KC and RC enzymes were also assessed in cells. The mitochondrial enzyme levels in human cervix cancer HeLa and AS-30D cells were higher than those observed in rat hepatocytes whereas in human breast cancer biopsies, CPT1 and SCAD contents were lower than in human breast normal tissue. The presence of CPT1 and SCAD in AS-30D mitochondria and HeLa cells correlated with an active FFA utilization in HeLa cells. Furthermore, the β-oxidation inhibitor perhexiline blocked FFA utilization, OxPhos and proliferation in HeLa and other cancer cells. In conclusion, functional mitochondria supported by FFA β-oxidation are essential for the accelerated cancer cell proliferation and hence anti-β-oxidation therapeutics appears as an alternative promising approach to deter malignant tumor growth. Copyright © 2015 Elsevier Ltd. All rights reserved.

Amino acid-dependent cMyc expression is essential for NK cell metabolic and functional responses in mice.

PubMed

Loftus, Róisín M; Assmann, Nadine; Kedia-Mehta, Nidhi; O'Brien, Katie L; Garcia, Arianne; Gillespie, Conor; Hukelmann, Jens L; Oefner, Peter J; Lamond, Angus I; Gardiner, Clair M; Dettmer, Katja; Cantrell, Doreen A; Sinclair, Linda V; Finlay, David K

2018-06-14

Natural killer (NK) cells are lymphocytes with important anti-tumour functions. Cytokine activation of NK cell glycolysis and oxidative phosphorylation (OXPHOS) are essential for robust NK cell responses. However, the mechanisms leading to this metabolic phenotype are unclear. Here we show that the transcription factor cMyc is essential for IL-2/IL-12-induced metabolic and functional responses in mice. cMyc protein levels are acutely regulated by amino acids; cMyc protein is lost rapidly when glutamine is withdrawn or when system L-amino acid transport is blocked. We identify SLC7A5 as the predominant system L-amino acid transporter in activated NK cells. Unlike other lymphocyte subsets, glutaminolysis and the tricarboxylic acid cycle do not sustain OXPHOS in activated NK cells. Glutamine withdrawal, but not the inhibition of glutaminolysis, results in the loss of cMyc protein, reduced cell growth and impaired NK cell responses. These data identify an essential role for amino acid-controlled cMyc for NK cell metabolism and function.

Doubling diet fat on sugar ratio in children with mitochondrial OXPHOS disorders: Effects of a randomized trial on resting energy expenditure, diet induced thermogenesis and body composition.

PubMed

Béghin, Laurent; Coopman, Stéphanie; Schiff, Manuel; Vamecq, Joseph; Mention-Mulliez, Karine; Hankard, Régis; Cuisset, Jean-Marie; Ogier, Hélène; Gottrand, Frédéric; Dobbelaere, Dries

2016-12-01

Mitochondrial OXPHOS disorders (MODs) affect one or several complexes of respiratory chain oxidative phosphorylation. An increased fat/low-carbohydrate ratio of the diet was recommended for treating MODs without, however, evaluating its potential benefits through changes in the respective contributions of cell pathways (glycolysis, fatty acid oxidation) initiating energy production. Therefore, the objective of the present work was to compare Resting Energy Expenditure (REE) under basal diet (BD) and challenging diet (CD) in which fat on sugar content ratio was doubled. Diet-induced thermogenesis (DIT) and body compositions were also compared. Energetic vs regulatory aspects of increasing fat contribution to total nutritional energy input were essentially addressed through measures primarily aiming at modifying total fat amounts and not the types of fats in designed diets. In this randomized cross-over study, BD contained 10% proteins/30% lipids/60% carbohydrates (fat on sugar ratio = 0.5) and was the imposed diet at baseline. CD contained 10% proteins/45% lipids/45% carbohydrates (fat on sugar ratio = 1). Main and second evaluation criteria measured by indirect calorimetry (QUARK RMR ® , Cosmed, Pavona; Italy) were REE and DIT, respectively. Thirty four MOD patients were included; 22 (mean age 13.2 ± 4.7 years, 50% female; BMI 16.9 ± 4.2 kg/m 2 ) were evaluated for REE, and 12 (mean age 13.8 ± 4.8 years, 60% female; BMI 17.4 ± 4.6 kg/m 2 ) also for DIT. OXPHOS complex deficiency repartition in 22 analysed patients was 55% for complex I, 9% for complex III, 27% for complex IV and 9% for other proteins. Neither carry-over nor period effects were detected (p = 0.878; ANOVA for repeated measures). REE was similar between BD vs CD (1148.8 ± 301.7 vs 1156.1 ± 278.8 kcal/day; p = 0.942) as well as DIT (peak DIT 260 vs 265 kcal/day; p = 0.842) and body composition (21.9 ± 13.0 vs 21.6 ± 13.3% of fat mass; p = 0.810). Doubling diet fat on sugar ratio does not appear to improve, per se, energetic status and body composition of patients with MODs. Copyright © 2016 Elsevier Ltd and European Society for Clinical Nutrition and Metabolism. All rights reserved.

m-AAA Complexes Are Not Crucial for the Survival of Arabidopsis Under Optimal Growth Conditions Despite Their Importance for Mitochondrial Translation.

PubMed

Kolodziejczak, Marta; Skibior-Blaszczyk, Renata; Janska, Hanna

2018-05-01

For optimal mitochondrial activity, the mitochondrial proteome must be properly maintained or altered in response to developmental and environmental stimuli. Based on studies of yeast and humans, one of the key players in this control are m-AAA proteases, mitochondrial inner membrane-bound ATP-dependent metalloenzymes. This study focuses on the importance of m-AAA proteases in plant mitochondria, providing their first experimentally proven physiological substrate. We found that the Arabidopsis m- AAA complexes composed of AtFTSH3 and/or AtFTSH10 are involved in the proteolytic maturation of ribosomal subunit L32. Consequently, in the double Arabidopsis ftsh3/10 mutant, mitoribosome biogenesis, mitochondrial translation and functionality of OXPHOS (oxidative phosphorylation) complexes are impaired. However, in contrast to their mammalian or yeast counterparts, plant m-AAA complexes are not critical for the survival of Arabidopsis under optimal conditions; ftsh3/10 plants are only slightly smaller in size at the early developmental stage compared with plants containing m-AAA complexes. Our data suggest that a lack of significant visible morphological alterations under optimal growth conditions involves mechanisms which rely on existing functional redundancy and induced functional compensation in Arabidopsis mitochondria.

Thymidine kinase 2 deficiency-induced mitochondrial DNA depletion causes abnormal development of adipose tissues and adipokine levels in mice.

PubMed

Villarroya, Joan; Dorado, Beatriz; Vilà, Maya R; Garcia-Arumí, Elena; Domingo, Pere; Giralt, Marta; Hirano, Michio; Villarroya, Francesc

2011-01-01

Mammal adipose tissues require mitochondrial activity for proper development and differentiation. The components of the mitochondrial respiratory chain/oxidative phosphorylation system (OXPHOS) are encoded by both mitochondrial and nuclear genomes. The maintenance of mitochondrial DNA (mtDNA) is a key element for a functional mitochondrial oxidative activity in mammalian cells. To ascertain the role of mtDNA levels in adipose tissue, we have analyzed the alterations in white (WAT) and brown (BAT) adipose tissues in thymidine kinase 2 (Tk2) H126N knockin mice, a model of TK2 deficiency-induced mtDNA depletion. We observed respectively severe and moderate mtDNA depletion in TK2-deficient BAT and WAT, showing both tissues moderate hypotrophy and reduced fat accumulation. Electron microscopy revealed altered mitochondrial morphology in brown but not in white adipocytes from TK2-deficient mice. Although significant reduction in mtDNA-encoded transcripts was observed both in WAT and BAT, protein levels from distinct OXPHOS complexes were significantly reduced only in TK2-deficient BAT. Accordingly, the activity of cytochrome c oxidase was significantly lowered only in BAT from TK2-deficient mice. The analysis of transcripts encoding up to fourteen components of specific adipose tissue functions revealed that, in both TK2-deficient WAT and BAT, there was a consistent reduction of thermogenesis related gene expression and a severe reduction in leptin mRNA. Reduced levels of resistin mRNA were found in BAT from TK2-deficient mice. Analysis of serum indicated a dramatic reduction in circulating levels of leptin and resistin. In summary, our present study establishes that mtDNA depletion leads to a moderate impairment in mitochondrial respiratory function, especially in BAT, causes substantial alterations in WAT and BAT development, and has a profound impact in the endocrine properties of adipose tissues. © 2011 Villarroya et al.

Thymidine Kinase 2 Deficiency-Induced Mitochondrial DNA Depletion Causes Abnormal Development of Adipose Tissues and Adipokine Levels in Mice

PubMed Central

Villarroya, Joan; Dorado, Beatriz; Vilà, Maya R.; Garcia-Arumí, Elena; Domingo, Pere; Giralt, Marta; Hirano, Michio; Villarroya, Francesc

2011-01-01

Mammal adipose tissues require mitochondrial activity for proper development and differentiation. The components of the mitochondrial respiratory chain/oxidative phosphorylation system (OXPHOS) are encoded by both mitochondrial and nuclear genomes. The maintenance of mitochondrial DNA (mtDNA) is a key element for a functional mitochondrial oxidative activity in mammalian cells. To ascertain the role of mtDNA levels in adipose tissue, we have analyzed the alterations in white (WAT) and brown (BAT) adipose tissues in thymidine kinase 2 (Tk2) H126N knockin mice, a model of TK2 deficiency-induced mtDNA depletion. We observed respectively severe and moderate mtDNA depletion in TK2-deficient BAT and WAT, showing both tissues moderate hypotrophy and reduced fat accumulation. Electron microscopy revealed altered mitochondrial morphology in brown but not in white adipocytes from TK2-deficient mice. Although significant reduction in mtDNA-encoded transcripts was observed both in WAT and BAT, protein levels from distinct OXPHOS complexes were significantly reduced only in TK2-deficient BAT. Accordingly, the activity of cytochrome c oxidase was significantly lowered only in BAT from TK2-deficient mice. The analysis of transcripts encoding up to fourteen components of specific adipose tissue functions revealed that, in both TK2-deficient WAT and BAT, there was a consistent reduction of thermogenesis related gene expression and a severe reduction in leptin mRNA. Reduced levels of resistin mRNA were found in BAT from TK2-deficient mice. Analysis of serum indicated a dramatic reduction in circulating levels of leptin and resistin. In summary, our present study establishes that mtDNA depletion leads to a moderate impairment in mitochondrial respiratory function, especially in BAT, causes substantial alterations in WAT and BAT development, and has a profound impact in the endocrine properties of adipose tissues. PMID:22216345

Heterogeneity in Cancer Metabolism: New Concepts in an Old Field

PubMed Central

Gentric, Géraldine; Mieulet, Virginie

2017-01-01

Abstract Significance: In the last years, metabolic reprogramming, fluctuations in bioenergetic fuels, and modulation of oxidative stress became new key hallmarks of tumor development. In cancer, elevated glucose uptake and high glycolytic rate, as a source of adenosine triphosphate, constitute a growth advantage for tumors. This represents the universally known Warburg effect, which gave rise to one major clinical application for detecting cancer cells using glucose analogs: the positron emission tomography scan imaging. Recent Advances: Glucose utilization and carbon sources in tumors are much more heterogeneous than initially thought. Indeed, new studies emerged and revealed a dual capacity of tumor cells for glycolytic and oxidative phosphorylation (OXPHOS) metabolism. OXPHOS metabolism, which relies predominantly on mitochondrial respiration, exhibits fine-tuned regulation of respiratory chain complexes and enhanced antioxidant response or detoxification capacity. Critical Issues: OXPHOS-dependent cancer cells use alternative oxidizable substrates, such as glutamine and fatty acids. The diversity of carbon substrates fueling neoplastic cells is indicative of metabolic heterogeneity, even within tumors sharing the same clinical diagnosis. Metabolic switch supports cancer cell stemness and their bioenergy-consuming functions, such as proliferation, survival, migration, and invasion. Moreover, reactive oxygen species-induced mitochondrial metabolism and nutrient availability are important for interaction with tumor microenvironment components. Carcinoma-associated fibroblasts and immune cells participate in the metabolic interplay with neoplastic cells. They collectively adapt in a dynamic manner to the metabolic needs of cancer cells, thus participating in tumorigenesis and resistance to treatments. Future Directions: Characterizing the reciprocal metabolic interplay between stromal, immune, and neoplastic cells will provide a better understanding of treatment resistance. Antioxid. Redox Signal. 26, 462–485. PMID:27228792

Repurposing atovaquone: Targeting mitochondrial complex III and OXPHOS to eradicate cancer stem cells

PubMed Central

Fiorillo, Marco; Lamb, Rebecca; Tanowitz, Herbert B.; Mutti, Luciano; Krstic-Demonacos, Marija; Cappello, Anna Rita; Martinez-Outschoorn, Ubaldo E.; Sotgia, Federica; Lisanti, Michael P.

2016-01-01

Atovaquone is an FDA-approved anti-malarial drug, which first became clinically available in the year 2000. Currently, its main usage is for the treatment of pneumocystis pneumonia (PCP) and/or toxoplasmosis in immune-compromised patients. Atovaquone is a hydroxy-1,4-naphthoquinone analogue of ubiquinone, also known as Co-enzyme Q10 (CoQ10). It is a well-tolerated drug that does not cause myelo-suppression. Mechanistically, it is thought to act as a potent and selective OXPHOS inhibitor, by targeting the CoQ10-dependence of mitochondrial complex III. Here, we show for the first time that atovaquone also has anti-cancer activity, directed against Cancer Stem-like Cells (CSCs). More specifically, we demonstrate that atovaquone treatment of MCF7 breast cancer cells inhibits oxygen-consumption and metabolically induces aerobic glycolysis (the Warburg effect), as well as oxidative stress. Remarkably, atovaquone potently inhibits the propagation of MCF7-derived CSCs, with an IC-50 of 1 μM, as measured using the mammosphere assay. Atovaquone also maintains this selectivity and potency in mixed populations of CSCs and non-CSCs. Importantly, these results indicate that glycolysis itself is not sufficient to maintain the proliferation of CSCs, which is instead strictly dependent on mitochondrial function. In addition to targeting the proliferation of CSCs, atovaquone also induces apoptosis in both CD44+/CD24low/− CSC and ALDH+ CSC populations, during exposure to anchorage-independent conditions for 12 hours. However, it has no effect on oxygen consumption in normal human fibroblasts and, in this cellular context, behaves as an anti-inflammatory, consistent with the fact that it is well-tolerated in patients treated for infections. Future studies in xenograft models and human clinical trials may be warranted, as the IC-50 of atovaquone's action on CSCs (1 μM) is >50 times less than its average serum concentration in humans. PMID:27136895

Mitochondrial transfer from Wharton's jelly-derived mesenchymal stem cells to mitochondria-defective cells recaptures impaired mitochondrial function.

PubMed

Lin, Hung-Yu; Liou, Chia-Wei; Chen, Shang-Der; Hsu, Te-Yao; Chuang, Jiin-Haur; Wang, Pei-Wen; Huang, Sheng-Teng; Tiao, Mao-Meng; Chen, Jin-Bor; Lin, Tsu-Kung; Chuang, Yao-Chung

2015-05-01

Adult mesenchymal stem cell (MSC)-conducted mitochondrial transfer has been recently shown to rescue cellular bioenergetics and prevent cell death caused by mitochondrial dysfunction. Wharton's jelly-derived MSCs (WJMSCs) harvested from postpartum umbilical cords are an accessible and abundant source of stem cells. This study aimed to determine the capability of WJMSCs to transfer their own mitochondria and rescue impaired oxidative phosphorylation (OXPHOS) and bioenergetics caused by mitochondrial DNA defects. To do this, WJMSCs were co-cultured with mitochondrial DNA (mtDNA)-depleted ρ(0) cells and the recapture of mitochondrial function was evaluated. WJMSCs were shown to be capable of transferring their own mitochondria into ρ(0) cells and underwent interorganellar mixture within these cells. Permissive culture media (BrdU-containing and pyruvate- and uridine-free) sieved out a survival cell population from the co-cultured WJMSCs (BrdU-sensitive) and ρ(0) cells (pyruvate/uridine-free). The survival cells had mtDNA identical to that of WJMSCs, whereas they expressed cellular markers identical to that of ρ(0) cells. Importantly, these ρ(0)-plus -WJMSC-mtDNA (ρ(+W)) cells recovered the expression of mtDNA-encoded proteins and exhibited functional oxygen consumption and respiratory control, as well as the activity of electron transport chain (ETC) complexes I, II, III and IV. In addition, ETC complex V-inhibitor-sensitive ATP production and metabolic shifting were also recovered. Furthermore, cellular behaviors including attachment-free proliferation, aerobic viability and OXPHOS-reliant cellular motility were also regained after mitochondrial transfer by WJMSCs. The therapeutic effect of WJMSCs-derived mitochondrial transfer was able to stably sustain for at least 45 passages. In conclusion, this study suggests that WJMSCs may serve as a potential therapeutic strategy for diseases linked to mitochondrial dysfunction through the donation of healthy mitochondria to cells with genetic mitochondrial defects. Copyright © 2015 Elsevier B.V. All rights reserved.

Lipopolysaccharide-induced mitochondrial dysfunction in boar sperm is mediated by activation of oxidative phosphorylation.

PubMed

He, Bin; Guo, Huiduo; Gong, Yabin; Zhao, Ruqian

2017-01-01

Lipopolysaccharide (LPS) has been reported to exert detrimental effects on boar sperm viability. In the present study, LPS was detected in boar semen samples at an average level of 0.62 ± 0.14 μg/mL. We treated boar sperm with 1 μg/mL LPS for 6 hours and examined alterations in sperm motility and apoptosis, together with mitochondrial functionality and mitochondrial reactive oxygen species generation. The expression and the location of toll-like receptor 4 (TLR4) and mitochondrial transcription factor A (TFAM) were determined to reveal possible mechanisms. LPS-treated sperm showed significant reduction in motility (P < 0.05) and viability (P < 0.05). LPS induced sperm mitochondrial damage via oxidative stress which is indicated by marked ultrastructural changes in the mitochondria including swelling, disorientation and vacuole, a decrease of mitochondrial membrane potential (ΔΨm; P < 0.05), as well as an increase of malondialdehyde levels (P < 0.01). Moreover, the production of mitochondrial reactive oxygen species through oxidative phosphorylation (OXPHOS) was significantly (P < 0.05) increased, which leads to oxidative stress. The copy number of mitochondrial DNA was significantly (P < 0.05) higher in LPS-treated sperm. Moreover, cytochrome c oxidase subunit IV (COXIV), an important subunit in mitochondrial electron transport chain and OXPHOS, was significantly (P < 0.05) upregulated after LPS treatment. TFAM, the key transcription factor that activates mitochondrial DNA replication and transcription, was translocated from the head to the midpiece of sperm where mitochondria are distributed in LPS-treated sperm. Taken together, these results indicate that LPS-induced decrease of motility and viability in boar sperm is mediated by abnormal activation of OXPHOS and mitochondrial membrane lipid peroxidation. These findings may provide new insights in understanding the mechanisms underlying the bacterial infection-induced sperm damage. Copyright © 2016 Elsevier Inc. All rights reserved.

The nuclear OXPHOS genes in insecta: a common evolutionary origin, a common cis-regulatory motif, a common destiny for gene duplicates

PubMed Central

Porcelli, Damiano; Barsanti, Paolo; Pesole, Graziano; Caggese, Corrado

2007-01-01

Background When orthologous sequences from species distributed throughout an optimal range of divergence times are available, comparative genomics is a powerful tool to address problems such as the identification of the forces that shape gene structure during evolution, although the functional constraints involved may vary in different genes and lineages. Results We identified and annotated in the MitoComp2 dataset the orthologs of 68 nuclear genes controlling oxidative phosphorylation in 11 Drosophilidae species and in five non-Drosophilidae insects, and compared them with each other and with their counterparts in three vertebrates (Fugu rubripes, Danio rerio and Homo sapiens) and in the cnidarian Nematostella vectensis, taking into account conservation of gene structure and regulatory motifs, and preservation of gene paralogs in the genome. Comparative analysis indicates that the ancestral insect OXPHOS genes were intron rich and that extensive intron loss and lineage-specific intron gain occurred during evolution. Comparison with vertebrates and cnidarians also shows that many OXPHOS gene introns predate the cnidarian/Bilateria evolutionary split. The nuclear respiratory gene element (NRG) has played a key role in the evolution of the insect OXPHOS genes; it is constantly conserved in the OXPHOS orthologs of all the insect species examined, while their duplicates either completely lack the element or possess only relics of the motif. Conclusion Our observations reinforce the notion that the common ancestor of most animal phyla had intron-rich gene, and suggest that changes in the pattern of expression of the gene facilitate the fixation of duplications in the genome and the development of novel genetic functions. PMID:18315839

Light might directly affect retinal ganglion cell mitochondria to potentially influence function.

PubMed

del Olmo-Aguado, Susana; Manso, Alberto G; Osborne, Neville N

2012-01-01

Visible light (360-760 nm) entering the eye impinges on the many ganglion cell mitochondria in the non-myelinated part of their axons. The same light also disrupts isolated mitochondrial function in vitro and kills cells in culture with the blue light component being particularly lethal whereas red light has little effect. Significantly, a defined light insult only affects the survival of fibroblasts in vitro that contain functional mitochondria supporting the view that mitochondrial photosensitizers are influenced by light. Moreover, a blue light insult to cells in culture causes a change in mitochondrial structure and membrane potential and results in a release of cytochrome c. Blue light also causes an alteration in mitochondria located components of the OXPHOS (oxidative phosphorylation system). Complexes III and IV as well as complex V are significantly upregulated whereas complexes I and II are slightly but significantly up- and downregulated, respectively. Also, blue light causes Dexras1 and reactive oxygen species to be upregulated and for mitochondrial located apoptosis-inducing factor to be activated. A blue light detrimental insult to cells in culture does not involve the activation of caspases but is known to be attenuated by necrostatin-1, typical of a death mechanism named necroptosis. © 2012 Wiley Periodicals, Inc. Photochemistry and Photobiology © 2012 The American Society of Photobiology.

Alterations in mitochondrial electron transport system activity in response to warm acclimation, hypoxia-reoxygenation and copper in rainbow trout, Oncorhynchus mykiss.

PubMed

Sappal, Ravinder; MacDougald, Michelle; Fast, Mark; Stevens, Don; Kibenge, Fred; Siah, Ahmed; Kamunde, Collins

2015-08-01

Fish expend significant amounts of energy to handle the numerous potentially stressful biotic and abiotic factors that they commonly encounter in aquatic environments. This universal requirement for energy singularizes mitochondria, the primary cellular energy transformers, as fundamental drivers of responses to environmental change. Our study probed the interacting effects of thermal stress, hypoxia-reoxygenation (HRO) and copper (Cu) exposure in rainbow trout to test the prediction that they act jointly to impair mitochondrial function. Rainbow trout were acclimated to 11 (controls) or 20°C for 2 months. Liver mitochondria were then isolated and their responses in vitro to Cu (0-20μM) without and with HRO were assessed. Sequential inhibition and activation of mitochondrial electron transport system (ETS) enzyme complexes permitted the measurement of respiratory activities supported by complex I-IV (CI-IV) in one run. The results showed that warm acclimation reduced fish and liver weights but increased mitochondrial protein indicating impairment of energy metabolism, increased synthesis of defense proteins and/or reduced liver water content. Whereas acute rise (11→20°C) in temperature increased mitochondrial oxidation rates supported by CI-IV, warm acclimation reduced the maximal (state 3) and increased the basal (state 4) respiration leading to global uncoupling of oxidative phosphorylation (OXPHOS). HRO profoundly inhibited both maximal and basal respiration rates supported by CI-IV, reduced RCR for all except CII and lowered CI:CII respiration ratio, an indication of decreased OXPHOS efficiency. The effects of Cu were less pronounced but more variable and included inhibition of CII-IV maximal respiration rates and stimulation of both CI and CIII basal respiration rates. Surprisingly, only CII and CIII indices exhibited significant 3-way interactions whereas 2-way interactions of acclimation either with Cu or HRO were portrayed mostly by CIV, and those of HRO and Cu were most common in CI and II respiratory indices. Our study suggests that warm acclimation blunts sensitivity of the ETS to temperature rise and that HRO and warm acclimation impose mitochondrial changes that sensitize the ETS to Cu. Overall, our study highlights the significance of the ETS in mitochondrial bioenergetic dysfunction caused by thermal stress, HRO and Cu exposure. Copyright © 2015 Elsevier B.V. All rights reserved.

An animal model to study human muscular diseases involving mitochondrial oxidative phosphorylation.

PubMed

Lemieux, Hélène; Warren, Blair E

2012-08-01

Mitochondria are producing most of the energy needed for many cellular functions by a process named oxidative phosphorylation (OXPHOS). It is now well recognized that mitochondrial dysfunctions are involved in several pathologies or degenerative processes, including cardiovascular diseases, diabetes, and aging. Animal models are currently used to try to understand the role of mitochondria in human diseases but a major problem is that mitochondria from different species and tissues are variable in terms of regulation. Analysis of mitochondrial function in three species of planarian flatworms (Tricladia, Platyhelminthes) shows that they share a very rare characteristic with human mitochondria: a strong control of oxidative phosphorylation by the phosphorylation system. The ratio of coupled OXPHOS over maximal electron transport capacity after uncoupling (electron transport system; ETS) well below 1.0 indicates that the phosphorylation system is limiting the rate of OXPHOS. The OXPHOS/ETS ratios are 0.62 ± 0.06 in Dugesia tigrina, 0.63 ± 0.05 in D. dorotocephala and 0.62 ± 0.05 in Procotyla fluviatilis, comparable to the value measured in human muscles. To our knowledge, no other animal model displays this peculiarity. This new model offers a venue in which to test the phosphorylation system as a potential therapeutic control point within humans.

Permeabilized Rat Cardiomyocyte Response Demonstrates Intracellular Origin of Diffusion Obstacles

PubMed Central

Jepihhina, Natalja; Beraud, Nathalie; Sepp, Mervi; Birkedal, Rikke; Vendelin, Marko

2011-01-01

Intracellular diffusion restrictions for ADP and other molecules have been predicted earlier based on experiments on permeabilized fibers or cardiomyocytes. However, it is possible that the effective diffusion distance is larger than the cell dimensions due to clumping of cells and incomplete separation of cells in fiber preparations. The aim of this work was to check whether diffusion restrictions exist inside rat cardiomyocytes or are caused by large effective diffusion distance. For that, we determined the response of oxidative phosphorylation (OxPhos) to exogenous ADP and ATP stimulation in permeabilized rat cardiomyocytes using fluorescence microscopy. The state of OxPhos was monitored via NADH and flavoprotein autofluorescence. By varying the ADP or ATP concentration in flow chamber, we determined that OxPhos has a low affinity in cardiomyocytes. The experiments were repeated in a fluorometer on cardiomyocyte suspensions leading to similar autofluorescence changes induced by ADP as recorded under the microscope. ATP stimulated OxPhos more in a fluorometer than under the microscope, which was attributed to accumulation of ADP in fluorometer chamber. By calculating the flow profile around the cell in the microscope chamber and comparing model solutions to measured data, we demonstrate that intracellular structures impose significant diffusion obstacles in rat cardiomyocytes. PMID:22067148

Defects of mtDNA Replication Impaired Mitochondrial Biogenesis During Trypanosoma cruzi Infection in Human Cardiomyocytes and Chagasic Patients: The Role of Nrf1/2 and Antioxidant Response

PubMed Central

Wan, Xianxiu; Gupta, Shivali; Zago, Maria P.; Davidson, Mercy M.; Dousset, Pierre; Amoroso, Alejandro; Garg, Nisha Jain

2012-01-01

Background Mitochondrial dysfunction is a key determinant in chagasic cardiomyopathy development in mice; however, its relevance in human Chagas disease is not known. We determined if defects in mitochondrial biogenesis and dysregulation of peroxisome proliferator-activated receptor gamma (PPARγ) coactivator-1 (PGC-1)–regulated transcriptional pathways constitute a mechanism or mechanisms underlying mitochondrial oxidative-phosphorylation (OXPHOS) deficiency in human Chagas disease. Methods and Results We utilized human cardiomyocytes and left-ventricular tissue from chagasic and other cardiomyopathy patients and healthy donors (n>6/group). We noted no change in citrate synthase activity, yet mRNA and/or protein levels of subunits of the respiratory complexes were significantly decreased in Trypanosoma cruzi–infected cardiomyocytes (0 to 24 hours) and chagasic hearts. We observed increased mRNA and decreased nuclear localization of PGC-1-coactivated transcription factors, yet the expression of genes for PPARγ-regulated fatty acid oxidation and nuclear respiratory factor (NRF1/2)–regulated mtDNA replication and transcription machinery was enhanced in infected cardiomyocytes and chagasic hearts. The D-loop formation was normal or higher, but mtDNA replication and mtDNA content were decreased by 83% and 40% to 65%, respectively. Subsequently, we noted that reactive oxygen species (ROS), oxidative stress, and mtDNA oxidation were significantly increased, yet NRF1/2-regulated antioxidant gene expression remained compromised in infected cardiomyocytes and chagasic hearts. Conclusions The replication of mtDNA was severely compromised, resulting in a significant loss of mtDNA and expression of OXPHOS genes in T cruzi–infected cardiomyocytes and chagasic hearts. Our data suggest increased ROS generation and selective functional incapacity of NRF2-mediated antioxidant gene expression played a role in the defects in mtDNA replication and unfitness of mtDNA for replication and gene expression in Chagas disease. PMID:23316324

The 12-day thermoregulatory metamorphosis of Red-winged Blackbirds (Agelaius phoeniceus).

PubMed

Sirsat, Sarah K Goy; Sirsat, Tushar S; Crossley, Janna L; Sotherland, Paul R; Dzialowski, Edward M

2016-07-01

We examined development of endothermy in altricial Red-winged Blackbirds (Agelaius phoeniceus) by measuring oxygen consumption [Formula: see text], body temperature and ventilation at ambient temperatures from 35 to 15 °C. Mitochondrial respiration of permeabilized skeletal muscle was also measured from breast (pectoralis) and thigh (femorotibialis) muscles. Animals were studied from the first day of hatching through fledging (12 days post-hatch, dph). Nestling whole-body metabolic rate began to show an endothermic response to cold temperature midway between hatching and fledging. Nestlings less than 5 dph were unable to maintain elevated [Formula: see text] and body temperature when exposed to gradually decreasing temperature, whereas 7 dph nestlings maintained [Formula: see text] until ~25 °C, after which [Formula: see text] decreased. From 10 dph to fledging, animals maintained elevated [Formula: see text] and body temperature when exposed to gradual cooling; full endothermic capacity was achieved. Ventilation followed a similar developmental trend to that of [Formula: see text], with increases in 10 dph fledglings occurring in tidal volume rather than ventilation frequency. LEAK respiration and oxidative phosphorylation (OXPHOS) through complex I of breast muscle mitochondria increased significantly after 3 dph. Expression of avUCP and PCG-1α mRNA increased significantly at 3 dph and remained elevated in both skeletal muscle types. Increased metabolic capacity at the cellular level occurred prior to that of the whole animal. This change in whole animal metabolic capacity increased steadily upon hatching as evidenced by the shift of metabolic rate from an ectothermic to endothermic phenotype and the increase of mitochondrial OXPHOS activity of the shivering muscles of this altricial avian species.

The m-AAA Protease Associated with Neurodegeneration Limits MCU Activity in Mitochondria.

PubMed

König, Tim; Tröder, Simon E; Bakka, Kavya; Korwitz, Anne; Richter-Dennerlein, Ricarda; Lampe, Philipp A; Patron, Maria; Mühlmeister, Mareike; Guerrero-Castillo, Sergio; Brandt, Ulrich; Decker, Thorsten; Lauria, Ines; Paggio, Angela; Rizzuto, Rosario; Rugarli, Elena I; De Stefani, Diego; Langer, Thomas

2016-10-06

Mutations in subunits of mitochondrial m-AAA proteases in the inner membrane cause neurodegeneration in spinocerebellar ataxia (SCA28) and hereditary spastic paraplegia (HSP7). m-AAA proteases preserve mitochondrial proteostasis, mitochondrial morphology, and efficient OXPHOS activity, but the cause for neuronal loss in disease is unknown. We have determined the neuronal interactome of m-AAA proteases in mice and identified a complex with C2ORF47 (termed MAIP1), which counteracts cell death by regulating the assembly of the mitochondrial Ca 2+ uniporter MCU. While MAIP1 assists biogenesis of the MCU subunit EMRE, the m-AAA protease degrades non-assembled EMRE and ensures efficient assembly of gatekeeper subunits with MCU. Loss of the m-AAA protease results in accumulation of constitutively active MCU-EMRE channels lacking gatekeeper subunits in neuronal mitochondria and facilitates mitochondrial Ca 2+ overload, mitochondrial permeability transition pore opening, and neuronal death. Together, our results explain neuronal loss in m-AAA protease deficiency by deregulated mitochondrial Ca 2+ homeostasis. Copyright © 2016 Elsevier Inc. All rights reserved.

FOXO1 is a TXN- and p300-dependent sensor and effector of oxidative stress in diffuse large B-cell lymphomas characterized by increased oxidative metabolism.

PubMed

Sewastianik, T; Szydlowski, M; Jablonska, E; Bialopiotrowicz, E; Kiliszek, P; Gorniak, P; Polak, A; Prochorec-Sobieszek, M; Szumera-Cieckiewicz, A; Kaminski, T S; Markowicz, S; Nowak, E; Grygorowicz, M A; Warzocha, K; Juszczynski, P

2016-11-17

Molecular profiling has led to identification of subtypes of diffuse large B-cell lymphomas (DLBCLs) differing in terms of oncogenic signaling and metabolic programs. The OxPhos-DLBCL subtype is characterized by enhanced mitochondrial oxidative phosphorylation. As increased oxidative metabolism leads to overproduction of potentially toxic reactive oxygen species (ROS), we sought to identify mechanisms responsible for adaptation of OxPhos cells to these conditions. Herein, we describe a mechanism involving the FOXO1-TXN-p300 redox-dependent circuit protecting OxPhos-DLBCL cells from ROS toxicity. We identify a BCL6-dependent transcriptional mechanism leading to relative TXN overexpression in OxPhos cells. We found that OxPhos cells lacking TXN were uniformly more sensitive to ROS and doxorubicin than control cells. Consistent with this, the overall survival of patients with high TXN mRNA expression, treated with doxorubicin-containing regimens, is significantly shorter than of those with low TXN mRNA expression. TXN overexpression curtails p300-mediated FOXO1 acetylation and its nuclear translocation in response to oxidative stress, thus attenuating FOXO1 transcriptional activity toward genes involved in apoptosis and cell cycle inhibition. We also demonstrate that FOXO1 knockdown in cells with silenced TXN expression markedly reduces ROS-induced apoptosis, indicating that FOXO1 is the major sensor and effector of oxidative stress in OxPhos-DLBCLs. These data highlight dynamic, context-dependent modulation of FOXO1 tumor-suppressor functions via acetylation and reveal potentially targetable vulnerabilities in these DLBCLs.

MHC-I modulation due to changes in tumor cell metabolism regulates tumor sensitivity to CTL and NK cells

PubMed Central

Catalán, Elena; Charni, Seyma; Jaime, Paula; Aguiló, Juan Ignacio; Enríquez, José Antonio; Naval, Javier; Pardo, Julián; Villalba, Martín; Anel, Alberto

2015-01-01

Tumor cells have a tendency to use glucose fermentation to obtain energy instead of mitochondrial oxidative phosphorylation (OXPHOS). We demonstrated that this phenotype correlated with loss of ERK5 expression and with reduced MHC class I expression. Consequently, tumor cells could evade cytotoxic T lymphocyte (CTL)-mediated immune surveillance, but also increase their sensitivity to natural killer (NK) cells. These outcomes were evaluated using two cellular models: leukemic EL4 cells and L929 transformed fibroblasts and their derived ρ° cell lines, which lack mitochondrial DNA. We have also used a L929 cell sub-line that spontaneously lost matrix attachment (L929dt), reminiscent of metastasis generation, that also downregulated MHC-I and ERK5 expression. MHC-I expression is lower in ρ° cells than in the parental cell lines, but they were equally sensitive to CTL. On the contrary, ρ° cells were more sensitive to activated NK cells than parental cells. On the other hand, L929dt cells were resistant to CTL and NK cells, showed reduced viability when forced to perform OXPHOS, and surviving cells increased MHC-I expression and became sensitive to CTL. The present results suggest that when the reduction in MHC-I levels in tumor cells due to glycolytic metabolism is partial, the increase in sensitivity to NK cells seems to predominate. However, when tumor cells completely lose MHC-I expression, the combination of treatments that increase OXPHOS with CTL-mediated immunotherapy could be a promising therapeutic approach. PMID:25949869

MHC-I modulation due to changes in tumor cell metabolism regulates tumor sensitivity to CTL and NK cells.

PubMed

Catalán, Elena; Charni, Seyma; Jaime, Paula; Aguiló, Juan Ignacio; Enríquez, José Antonio; Naval, Javier; Pardo, Julián; Villalba, Martín; Anel, Alberto

2015-01-01

Tumor cells have a tendency to use glucose fermentation to obtain energy instead of mitochondrial oxidative phosphorylation (OXPHOS). We demonstrated that this phenotype correlated with loss of ERK5 expression and with reduced MHC class I expression. Consequently, tumor cells could evade cytotoxic T lymphocyte (CTL)-mediated immune surveillance, but also increase their sensitivity to natural killer (NK) cells. These outcomes were evaluated using two cellular models: leukemic EL4 cells and L929 transformed fibroblasts and their derived ρ° cell lines, which lack mitochondrial DNA. We have also used a L929 cell sub-line that spontaneously lost matrix attachment (L929dt), reminiscent of metastasis generation, that also downregulated MHC-I and ERK5 expression. MHC-I expression is lower in ρ° cells than in the parental cell lines, but they were equally sensitive to CTL. On the contrary, ρ° cells were more sensitive to activated NK cells than parental cells. On the other hand, L929dt cells were resistant to CTL and NK cells, showed reduced viability when forced to perform OXPHOS, and surviving cells increased MHC-I expression and became sensitive to CTL. The present results suggest that when the reduction in MHC-I levels in tumor cells due to glycolytic metabolism is partial, the increase in sensitivity to NK cells seems to predominate. However, when tumor cells completely lose MHC-I expression, the combination of treatments that increase OXPHOS with CTL-mediated immunotherapy could be a promising therapeutic approach.

Burn after feeding. An old uncoupler of oxidative phosphorylation is redesigned for the treatment of nonalcoholic fatty liver disease.

PubMed

Fromenty, B

2014-10-01

Uncoupling of oxidative phosphorylation (OXPHOS) in brown adipose tissue can be used by hibernating animals to produce heat at the expense of their fat mass. In a recent work, Dr Shulman et al. generated a liver-targeted derivative of the prototypical OXPHOS uncoupler 2,4-dinitrophenol that alleviated steatosis, hypertriglyceridemia and insulin resistance in several models of nonalcoholic fatty liver disease and type 2 diabetes. Copyright © 2014 Elsevier Masson SAS. All rights reserved.

Mitochondrial acclimation potential to ocean acidification and warming of Polar cod (Boreogadus saida) and Atlantic cod (Gadus morhua).

PubMed

Leo, Elettra; Kunz, Kristina L; Schmidt, Matthias; Storch, Daniela; Pörtner, Hans-O; Mark, Felix C

2017-01-01

Ocean acidification and warming are happening fast in the Arctic but little is known about the effects of ocean acidification and warming on the physiological performance and survival of Arctic fish. In this study we investigated the metabolic background of performance through analyses of cardiac mitochondrial function in response to control and elevated water temperatures and P CO 2 of two gadoid fish species, Polar cod ( Boreogadus saida ), an endemic Arctic species, and Atlantic cod ( Gadus morhua ), which is a temperate to cold eurytherm and currently expanding into Arctic waters in the wake of ocean warming. We studied their responses to the above-mentioned drivers and their acclimation potential through analysing the cardiac mitochondrial function in permeabilised cardiac muscle fibres after 4 months of incubation at different temperatures (Polar cod: 0, 3, 6, 8 °C and Atlantic cod: 3, 8, 12, 16 °C), combined with exposure to present (400μatm) and year 2100 (1170μatm) levels of CO 2 . OXPHOS, proton leak and ATP production efficiency in Polar cod were similar in the groups acclimated at 400μatm and 1170μatm of CO 2 , while incubation at 8 °C evoked increased proton leak resulting in decreased ATP production efficiency and decreased Complex IV capacity. In contrast, OXPHOS of Atlantic cod increased with temperature without compromising the ATP production efficiency, whereas the combination of high temperature and high P CO 2 depressed OXPHOS and ATP production efficiency. Polar cod mitochondrial efficiency decreased at 8 °C while Atlantic cod mitochondria were more resilient to elevated temperature; however, this resilience was constrained by high P CO 2 . In line with its lower habitat temperature and higher degree of stenothermy, Polar cod has a lower acclimation potential to warming than Atlantic cod.

Markers of Skeletal Muscle Mitochondrial Function and Lipid Accumulation Are Moderately Associated with the Homeostasis Model Assessment Index of Insulin Resistance in Obese Men

PubMed Central

Samjoo, Imtiaz A.; Safdar, Adeel; Hamadeh, Mazen J.; Glover, Alexander W.; Mocellin, Nicholas J.; Santana, Jose; Little, Jonathan P.; Steinberg, Gregory R.; Raha, Sandeep; Tarnopolsky, Mark A.

2013-01-01

Lower skeletal muscle mitochondrial oxidative phosphorylation capacity (OXPHOS) and intramyocellular lipid (IMCL) accumulation have been implicated in the etiology of insulin resistance (IR) in obesity. The purpose of this study was to examine the impact of endurance exercise on biochemical and morphological measures of IMCL and mitochondrial content, and their relationship to IR in obese individuals. We examined mitochondrial content (subunit protein abundance and maximal activity of electron transport chain enzymes), IMCL/mitochondrial morphology in both subsarcolemmal (SS) and intermyofibrillar (IMF) regions by transmission electron microscopy, and intracellular lipid metabolites (diacylglycerol and ceramide) in vastus lateralis biopsies, as well as, the homeostasis model assessment index of IR (HOMA-IR) prior to and following twelve weeks of an endurance exercise regimen in healthy age- and physical activity-matched lean and obese men. Obese men did not show evidence of mitochondrial OXPHOS dysfunction, disproportionate IMCL content in sub-cellular regions, or diacylglycerol/ceramide accretion despite marked IR vs. lean controls. Endurance exercise increased OXPHOS and mitochondrial size and density, but not number of individual mitochondrial fragments, with moderate improvements in HOMA-IR. Exercise reduced SS IMCL content (size, number and density), increased IMF IMCL content, while increasing IMCL/mitochondrial juxtaposition in both regions. HOMA-IR was inversely associated with SS (r = −0.34; P = 0.051) and IMF mitochondrial density (r = −0.29; P = 0.096), IMF IMCL/mitochondrial juxtaposition (r = −0.30; P = 0.086), and COXII (r = −0.32; P = 0.095) and COXIV protein abundance (r = −0.35; P = 0.052); while positively associated with SS IMCL size (r = 0.28; P = 0.119) and SS IMCL density (r = 0.25; P = 0.152). Our findings suggest that once physical activity and cardiorespiratory fitness have been controlled for, skeletal muscle mitochondrial and IMCL profile in obesity may only partially contribute to the development of IR. PMID:23776659

The Elusive Nature of Adaptive Mitochondrial DNA Evolution of an Arctic Lineage Prone to Frequent Introgression

PubMed Central

Melo-Ferreira, José; Vilela, Joana; Fonseca, Miguel M.; da Fonseca, Rute R.; Boursot, Pierre; Alves, Paulo C.

2014-01-01

Mitochondria play a fundamental role in cellular metabolism, being responsible for most of the energy production of the cell in the oxidative phosphorylation (OXPHOS) pathway. Mitochondrial DNA (mtDNA) encodes for key components of this process, but its direct role in adaptation remains far from understood. Hares (Lepus spp.) are privileged models to study the impact of natural selection on mitogenomic evolution because 1) species are adapted to contrasting environments, including arctic, with different metabolic pressures, and 2) mtDNA introgression from arctic into temperate species is widespread. Here, we analyzed the sequences of 11 complete mitogenomes (ten newly obtained) of hares of temperate and arctic origins (including two of arctic origin introgressed into temperate species). The analysis of patterns of codon substitutions along the reconstructed phylogeny showed evidence for positive selection in several codons in genes of the OXPHOS complexes, most notably affecting the arctic lineage. However, using theoretical models, no predictable effect of these differences was found on the structure and physicochemical properties of the encoded proteins, suggesting that the focus of selection may lie on complex interactions with nuclear encoded peptides. Also, a cloverleaf structure was detected in the control region only from the arctic mtDNA lineage, which may influence mtDNA replication and transcription. These results suggest that adaptation impacted the evolution of hare mtDNA and may have influenced the occurrence and consequences of the many reported cases of massive mtDNA introgression. However, the origin of adaptation remains elusive. PMID:24696399

The emerging role of ASC in dendritic cell metabolism during Chlamydia infection

PubMed Central

McKeithen, Danielle N.; Ryans, Khamia; Mu, Jing; Xie, Zhonglin; Simoneaux, Tankya; Blas-machado, Uriel; Eko, Francis O.; Black, Carolyn M.; Igietseme, Joseph U.; He, Qing

2017-01-01

Chlamydia trachomatis is a bacterial agent that causes sexually transmitted infections worldwide. The regulatory functions of dendritic cells (DCs) play a major role in protective immunity against Chlamydia infections. Here, we investigated the role of ASC in DCs metabolism and the regulation of DCs activation and function during Chlamydia infection. Following Chlamydia stimulation, maturation and antigen presenting functions were impaired in ASC-/- DCs compared to wild type (WT) DCs, in addition, ASC deficiency induced a tolerant phenotype in Chlamydia stimulated DCs. Using real-time extracellular flux analysis, we showed that activation in Chlamydia stimulated WT DCs is associated with a metabolic change in which mitochondrial oxidative phosphorylation (OXPHOS) is inhibited and the cells become committed to utilizing glucose through aerobic glycolysis for differentiation and antigen presenting functions. However, in ASC-/- DCs Chlamydia-induced metabolic change was prevented and there was a significant effect on mitochondrial morphology. The mitochondria of Chlamydia stimulated ASC-/- DCs had disrupted cristae compared to the normal narrow pleomorphic cristae found in stimulated WT DCs. In conclusion, our results suggest that Chlamydia-mediated activation of DCs is associated with a metabolic transition in which OXPHOS is inhibited, thereby dedicating the DCs to aerobic glycolysis, while ASC deficiency disrupts DCs function by inhibiting the reprogramming of DCs metabolism within the mitochondria, from glycolysis to electron transport chain. PMID:29216217

The paradoxical relationship between stallion fertility and oxidative stress.

PubMed

Gibb, Zamira; Lambourne, Sarah R; Aitken, Robert J

2014-09-01

The relationship between stallion fertility and oxidative stress remains poorly understood. The purpose of this study was to identify criteria for thoroughbred fertility assessment by performing a logistical regression analysis using "dismount" sperm parameters as predictors and weekly per-cycle conception rate as the dependent variable. Paradoxically, positive relationships between fertility and oxidative stress were revealed, such that samples that produced pregnancies exhibited higher rates of 8-hydroxy-2'-deoxyguanosine release (1490.2% vs. 705.5 pg/ml/24 h) and lower vitality (60.5% vs. 69.6%) and acrosome integrity (40.2% vs. 50.1%) than those that did not. We hypothesized that the most fertile spermatozoa exhibited the highest levels of oxidative phosphorylation (OXPHOS), with oxidative stress simply being a by-product of intense mitochondrial activity. Accordingly, an experiment to investigate the relationship between oxidative stress and motility was conducted and revealed positive correlations between mitochondrial ROS and total motility (R² = 0.90), rapid motility (R² = 0.89), average path velocity (VAP; R² = 0.59), and curvilinear velocity (VCL; R² = 0.66). Similarly, lipid peroxidation was positively correlated with total motility (R² = 0.46), rapid motility (R² = 0.51), average path velocity (R² = 0.62), and VCL (R² = 0.56), supporting the aforementioned hypothesis. The relative importance of OXPHOS in supporting the motility of equine spermatozoa was contrasted with human spermatozoa, which primarily utilize glycolysis. In this study, mitochondrial inhibition significantly reduced the velocity (P < 0.01) and ATP (P < 0.05) content of equine, but not human, spermatozoa, emphasizing the former's relative dependence on OXPHOS. The equine is the first mammal in which such a positive relationship between oxidative stress and functionality has been observed, with implications for the management of stallion fertility in vitro and in vivo. © 2014 by the Society for the Study of Reproduction, Inc.

Different effects of guanine nucleotides (GDP and GTP) on protein-mediated mitochondrial proton leak.

PubMed

Woyda-Ploszczyca, Andrzej M; Jarmuszkiewicz, Wieslawa

2014-01-01

In this study, we compared the influence of GDP and GTP on isolated mitochondria respiring under conditions favoring oxidative phosphorylation (OXPHOS) and under conditions excluding this process, i.e., in the presence of carboxyatractyloside, an adenine nucleotide translocase inhibitor, and/or oligomycin, an FOF1-ATP synthase inhibitor. Using mitochondria isolated from rat kidney and human endothelial cells, we found that the action of GDP and GTP can differ diametrically depending on the conditions. Namely, under conditions favoring OXPHOS, both in the absence and presence of linoleic acid, an activator of uncoupling proteins (UCPs), the addition of 1 mM GDP resulted in the state 4 (non-phosphorylating respiration)-state 3 (phosphorylating respiration) transition, which is characteristic of ADP oxidative phosphorylation. In contrast, the addition of 1 mM GTP resulted in a decrease in the respiratory rate and an increase in the membrane potential, which is characteristic of UCP inhibition. The stimulatory effect of GDP, but not GTP, was also observed in inside-out submitochondrial particles prepared from rat kidney mitochondria. However, the effects of GDP and GTP were more similar in the presence of OXPHOS inhibitors. The importance of these observations in connection with the action of UCPs, adenine nucleotide translocase (or other carboxyatractyloside-sensitive carriers), carboxyatractyloside- and purine nucleotide-insensitive carriers, as well as nucleoside-diphosphate kinase (NDPK) are considered. Because the measurements favoring oxidative phosphorylation better reflect in vivo conditions, our study strongly supports the idea that GDP cannot be considered a significant physiological inhibitor of UCP. Moreover, it appears that, under native conditions, GTP functions as a more efficient UCP inhibitor than GDP and ATP.

Different Effects of Guanine Nucleotides (GDP and GTP) on Protein-Mediated Mitochondrial Proton Leak

PubMed Central

Woyda-Ploszczyca, Andrzej M.; Jarmuszkiewicz, Wieslawa

2014-01-01

In this study, we compared the influence of GDP and GTP on isolated mitochondria respiring under conditions favoring oxidative phosphorylation (OXPHOS) and under conditions excluding this process, i.e., in the presence of carboxyatractyloside, an adenine nucleotide translocase inhibitor, and/or oligomycin, an FOF1-ATP synthase inhibitor. Using mitochondria isolated from rat kidney and human endothelial cells, we found that the action of GDP and GTP can differ diametrically depending on the conditions. Namely, under conditions favoring OXPHOS, both in the absence and presence of linoleic acid, an activator of uncoupling proteins (UCPs), the addition of 1 mM GDP resulted in the state 4 (non-phosphorylating respiration)-state 3 (phosphorylating respiration) transition, which is characteristic of ADP oxidative phosphorylation. In contrast, the addition of 1 mM GTP resulted in a decrease in the respiratory rate and an increase in the membrane potential, which is characteristic of UCP inhibition. The stimulatory effect of GDP, but not GTP, was also observed in inside-out submitochondrial particles prepared from rat kidney mitochondria. However, the effects of GDP and GTP were more similar in the presence of OXPHOS inhibitors. The importance of these observations in connection with the action of UCPs, adenine nucleotide translocase (or other carboxyatractyloside-sensitive carriers), carboxyatractyloside- and purine nucleotide-insensitive carriers, as well as nucleoside-diphosphate kinase (NDPK) are considered. Because the measurements favoring oxidative phosphorylation better reflect in vivo conditions, our study strongly supports the idea that GDP cannot be considered a significant physiological inhibitor of UCP. Moreover, it appears that, under native conditions, GTP functions as a more efficient UCP inhibitor than GDP and ATP. PMID:24904988

Absence of Complex I Is Associated with Diminished Respiratory Chain Function in European Mistletoe.

PubMed

Maclean, Andrew E; Hertle, Alexander P; Ligas, Joanna; Bock, Ralph; Balk, Janneke; Meyer, Etienne H

2018-05-21

Parasitism is a life history strategy found across all domains of life whereby nutrition is obtained from a host. It is often associated with reductive evolution of the genome, including loss of genes from the organellar genomes [1, 2]. In some unicellular parasites, the mitochondrial genome (mitogenome) has been lost entirely, with far-reaching consequences for the physiology of the organism [3, 4]. Recently, mitogenome sequences of several species of the hemiparasitic plant mistletoe (Viscum sp.) have been reported [5, 6], revealing a striking loss of genes not seen in any other multicellular eukaryotes. In particular, the nad genes encoding subunits of respiratory complex I are all absent and other protein-coding genes are also lost or highly diverged in sequence, raising the question what remains of the respiratory complexes and mitochondrial functions. Here we show that oxidative phosphorylation (OXPHOS) in European mistletoe, Viscum album, is highly diminished. Complex I activity and protein subunits of complex I could not be detected. The levels of complex IV and ATP synthase were at least 5-fold lower than in the non-parasitic model plant Arabidopsis thaliana, whereas alternative dehydrogenases and oxidases were higher in abundance. Carbon flux analysis indicates that cytosolic reactions including glycolysis are greater contributors to ATP synthesis than the mitochondrial tricarboxylic acid (TCA) cycle. Our results describe the extreme adjustments in mitochondrial functions of the first reported multicellular eukaryote without complex I. Copyright © 2018 Elsevier Ltd. All rights reserved.

Leucine reduces reactive oxygen species levels via an energy metabolism switch by activation of the mTOR-HIF-1α pathway in porcine intestinal epithelial cells.

PubMed

Hu, Jun; Nie, Yangfan; Chen, Shifeng; Xie, Chunlin; Fan, Qiwen; Wang, Zhichang; Long, Baisheng; Yan, Guokai; Zhong, Qing; Yan, Xianghua

2017-08-01

Leucine serves not only as a substrate for protein synthesis, but also as a signal molecule involved in protein metabolism. However, whether the levels of cellular reactive oxygen species (ROS), which have damaging effects on cellular DNA, proteins, and lipids, are regulated by leucine is still unclear. Here, we report that leucine supplementation reduces ROS levels in intestinal epithelial cells of weaned piglets. A proteomics analysis revealed that leucine supplementation induces an energy metabolism switch from oxidative phosphorylation (OXPHOS) towards glycolysis. The leucine-induced ROS reduction and the energy metabolism switch were further validated in cultured cells. Mechanistically, our data revealed that leucine-induced ROS reduction actually depends on the energy metabolism switch from OXPHOS towards glycolysis through the mechanistic target of rapamycin (mTOR)- hypoxia-inducible factor-1alpha (HIF-1α) pathway. These findings reveal a vital regulatory role of leucine as the signal molecule involved in an energy metabolism switch in mammals. Copyright © 2017 Elsevier Ltd. All rights reserved.

Human REV3 DNA Polymerase Zeta Localizes to Mitochondria and Protects the Mitochondrial Genome.

PubMed

Singh, Bhupendra; Li, Xiurong; Owens, Kjerstin M; Vanniarajan, Ayyasamy; Liang, Ping; Singh, Keshav K

2015-01-01

To date, mitochondrial DNA polymerase γ (POLG) is the only polymerase known to be present in mammalian mitochondria. A dogma in the mitochondria field is that there is no other polymerase present in the mitochondria of mammalian cells. Here we demonstrate localization of REV3 DNA polymerase in the mammalian mitochondria. We demonstrate localization of REV3 in the mitochondria of mammalian tissue as well as cell lines. REV3 associates with POLG and mitochondrial DNA and protects the mitochondrial genome from DNA damage. Inactivation of Rev3 leads to reduced mitochondrial membrane potential, reduced OXPHOS activity, and increased glucose consumption. Conversely, inhibition of the OXPHOS increases expression of Rev3. Rev3 expression is increased in human primary breast tumors and breast cancer cell lines. Inactivation of Rev3 decreases cell migration and invasion, and localization of Rev3 in mitochondria increases survival and the invasive potential of cancer cells. Taken together, we demonstrate that REV3 functions in mammalian mitochondria and that mitochondrial REV3 is associated with the tumorigenic potential of cells.

The cellular and compartmental profile of mouse retinal glycolysis, tricarboxylic acid cycle, oxidative phosphorylation, and ~P transferring kinases

PubMed Central

Rueda, Elda M.; Johnson, Jerry E.; Giddabasappa, Anand; Swaroop, Anand; Brooks, Matthew J.; Sigel, Irena; Chaney, Shawnta Y.

2016-01-01

Purpose The homeostatic regulation of cellular ATP is achieved by the coordinated activity of ATP utilization, synthesis, and buffering. Glucose is the major substrate for ATP synthesis through glycolysis and oxidative phosphorylation (OXPHOS), whereas intermediary metabolism through the tricarboxylic acid (TCA) cycle utilizes non-glucose-derived monocarboxylates, amino acids, and alpha ketoacids to support mitochondrial ATP and GTP synthesis. Cellular ATP is buffered by specialized equilibrium-driven high-energy phosphate (~P) transferring kinases. Our goals were twofold: 1) to characterize the gene expression, protein expression, and activity of key synthesizing and regulating enzymes of energy metabolism in the whole mouse retina, retinal compartments, and/or cells and 2) to provide an integrative analysis of the results related to function. Methods mRNA expression data of energy-related genes were extracted from our whole retinal Affymetrix microarray data. Fixed-frozen retinas from adult C57BL/6N mice were used for immunohistochemistry, laser scanning confocal microscopy, and enzymatic histochemistry. The immunoreactivity levels of well-characterized antibodies, for all major retinal cells and their compartments, were obtained using our established semiquantitative confocal and imaging techniques. Quantitative cytochrome oxidase (COX) and lactate dehydrogenase (LDH) activity was determined histochemically. Results The Affymetrix data revealed varied gene expression patterns of the ATP synthesizing and regulating enzymes found in the muscle, liver, and brain. Confocal studies showed differential cellular and compartmental distribution of isozymes involved in glucose, glutamate, glutamine, lactate, and creatine metabolism. The pattern and intensity of the antibodies and of the COX and LDH activity showed the high capacity of photoreceptors for aerobic glycolysis and OXPHOS. Competition assays with pyruvate revealed that LDH-5 was localized in the photoreceptor inner segments. The combined results indicate that glycolysis is regulated by the compartmental expression of hexokinase 2, pyruvate kinase M1, and pyruvate kinase M2 in photoreceptors, whereas the inner retinal neurons exhibit a lower capacity for glycolysis and aerobic glycolysis. Expression of nucleoside diphosphate kinase, mitochondria-associated adenylate kinase, and several mitochondria-associated creatine kinase isozymes was highest in the outer retina, whereas expression of cytosolic adenylate kinase and brain creatine kinase was higher in the cones, horizontal cells, and amacrine cells indicating the diversity of ATP-buffering strategies among retinal neurons. Based on the antibody intensities and the COX and LDH activity, Müller glial cells (MGCs) had the lowest capacity for glycolysis, aerobic glycolysis, and OXPHOS. However, they showed high expression of glutamate dehydrogenase, alpha-ketoglutarate dehydrogenase, succinate thiokinase, GABA transaminase, and ~P transferring kinases. This suggests that MGCs utilize TCA cycle anaplerosis and cataplerosis to generate GTP and ~P transferring kinases to produce ATP that supports MGC energy requirements. Conclusions Our comprehensive and integrated results reveal that the adult mouse retina expresses numerous isoforms of ATP synthesizing, regulating, and buffering genes; expresses differential cellular and compartmental levels of glycolytic, OXPHOS, TCA cycle, and ~P transferring kinase proteins; and exhibits differential layer-by-layer LDH and COX activity. New insights into cell-specific and compartmental ATP and GTP production, as well as utilization and buffering strategies and their relationship with known retinal and cellular functions, are discussed. Developing therapeutic strategies for neuroprotection and treating retinal deficits and degeneration in a cell-specific manner will require such knowledge. This work provides a platform for future research directed at identifying the molecular targets and proteins that regulate these processes. PMID:27499608

The cellular and compartmental profile of mouse retinal glycolysis, tricarboxylic acid cycle, oxidative phosphorylation, and ~P transferring kinases.

PubMed

Rueda, Elda M; Johnson, Jerry E; Giddabasappa, Anand; Swaroop, Anand; Brooks, Matthew J; Sigel, Irena; Chaney, Shawnta Y; Fox, Donald A

2016-01-01

The homeostatic regulation of cellular ATP is achieved by the coordinated activity of ATP utilization, synthesis, and buffering. Glucose is the major substrate for ATP synthesis through glycolysis and oxidative phosphorylation (OXPHOS), whereas intermediary metabolism through the tricarboxylic acid (TCA) cycle utilizes non-glucose-derived monocarboxylates, amino acids, and alpha ketoacids to support mitochondrial ATP and GTP synthesis. Cellular ATP is buffered by specialized equilibrium-driven high-energy phosphate (~P) transferring kinases. Our goals were twofold: 1) to characterize the gene expression, protein expression, and activity of key synthesizing and regulating enzymes of energy metabolism in the whole mouse retina, retinal compartments, and/or cells and 2) to provide an integrative analysis of the results related to function. mRNA expression data of energy-related genes were extracted from our whole retinal Affymetrix microarray data. Fixed-frozen retinas from adult C57BL/6N mice were used for immunohistochemistry, laser scanning confocal microscopy, and enzymatic histochemistry. The immunoreactivity levels of well-characterized antibodies, for all major retinal cells and their compartments, were obtained using our established semiquantitative confocal and imaging techniques. Quantitative cytochrome oxidase (COX) and lactate dehydrogenase (LDH) activity was determined histochemically. The Affymetrix data revealed varied gene expression patterns of the ATP synthesizing and regulating enzymes found in the muscle, liver, and brain. Confocal studies showed differential cellular and compartmental distribution of isozymes involved in glucose, glutamate, glutamine, lactate, and creatine metabolism. The pattern and intensity of the antibodies and of the COX and LDH activity showed the high capacity of photoreceptors for aerobic glycolysis and OXPHOS. Competition assays with pyruvate revealed that LDH-5 was localized in the photoreceptor inner segments. The combined results indicate that glycolysis is regulated by the compartmental expression of hexokinase 2, pyruvate kinase M1, and pyruvate kinase M2 in photoreceptors, whereas the inner retinal neurons exhibit a lower capacity for glycolysis and aerobic glycolysis. Expression of nucleoside diphosphate kinase, mitochondria-associated adenylate kinase, and several mitochondria-associated creatine kinase isozymes was highest in the outer retina, whereas expression of cytosolic adenylate kinase and brain creatine kinase was higher in the cones, horizontal cells, and amacrine cells indicating the diversity of ATP-buffering strategies among retinal neurons. Based on the antibody intensities and the COX and LDH activity, Müller glial cells (MGCs) had the lowest capacity for glycolysis, aerobic glycolysis, and OXPHOS. However, they showed high expression of glutamate dehydrogenase, alpha-ketoglutarate dehydrogenase, succinate thiokinase, GABA transaminase, and ~P transferring kinases. This suggests that MGCs utilize TCA cycle anaplerosis and cataplerosis to generate GTP and ~P transferring kinases to produce ATP that supports MGC energy requirements. Our comprehensive and integrated results reveal that the adult mouse retina expresses numerous isoforms of ATP synthesizing, regulating, and buffering genes; expresses differential cellular and compartmental levels of glycolytic, OXPHOS, TCA cycle, and ~P transferring kinase proteins; and exhibits differential layer-by-layer LDH and COX activity. New insights into cell-specific and compartmental ATP and GTP production, as well as utilization and buffering strategies and their relationship with known retinal and cellular functions, are discussed. Developing therapeutic strategies for neuroprotection and treating retinal deficits and degeneration in a cell-specific manner will require such knowledge. This work provides a platform for future research directed at identifying the molecular targets and proteins that regulate these processes.

Mitochondrial Aging Defects Emerge in Directly Reprogrammed Human Neurons due to Their Metabolic Profile.

PubMed

Kim, Yongsung; Zheng, Xinde; Ansari, Zoya; Bunnell, Mark C; Herdy, Joseph R; Traxler, Larissa; Lee, Hyungjun; Paquola, Apua C M; Blithikioti, Chrysanthi; Ku, Manching; Schlachetzki, Johannes C M; Winkler, Jürgen; Edenhofer, Frank; Glass, Christopher K; Paucar, Andres A; Jaeger, Baptiste N; Pham, Son; Boyer, Leah; Campbell, Benjamin C; Hunter, Tony; Mertens, Jerome; Gage, Fred H

2018-05-29

Mitochondria are a major target for aging and are instrumental in the age-dependent deterioration of the human brain, but studying mitochondria in aging human neurons has been challenging. Direct fibroblast-to-induced neuron (iN) conversion yields functional neurons that retain important signs of aging, in contrast to iPSC differentiation. Here, we analyzed mitochondrial features in iNs from individuals of different ages. iNs from old donors display decreased oxidative phosphorylation (OXPHOS)-related gene expression, impaired axonal mitochondrial morphologies, lower mitochondrial membrane potentials, reduced energy production, and increased oxidized proteins levels. In contrast, the fibroblasts from which iNs were generated show only mild age-dependent changes, consistent with a metabolic shift from glycolysis-dependent fibroblasts to OXPHOS-dependent iNs. Indeed, OXPHOS-induced old fibroblasts show increased mitochondrial aging features similar to iNs. Our data indicate that iNs are a valuable tool for studying mitochondrial aging and support a bioenergetic explanation for the high susceptibility of the brain to aging. Copyright © 2018 The Author(s). Published by Elsevier Inc. All rights reserved.

Selective downregulation of mitochondrial electron transport chain activity and increased oxidative stress in human atrial fibrillation.

PubMed

Emelyanova, Larisa; Ashary, Zain; Cosic, Milanka; Negmadjanov, Ulugbek; Ross, Gracious; Rizvi, Farhan; Olet, Susan; Kress, David; Sra, Jasbir; Tajik, A Jamil; Holmuhamedov, Ekhson L; Shi, Yang; Jahangir, Arshad

2016-07-01

Mitochondria are critical for maintaining normal cardiac function, and a deficit in mitochondrial energetics can lead to the development of the substrate that promotes atrial fibrillation (AF) and its progression. However, the link between mitochondrial dysfunction and AF in humans is still not fully defined. The aim of this study was to elucidate differences in the functional activity of mitochondrial oxidative phosphorylation (OXPHOS) complexes and oxidative stress in right atrial tissue from patients without (non-AF) and with AF (AF) who were undergoing open-heart surgery and were not significantly different for age, sex, major comorbidities, and medications. The overall functional activity of the electron transport chain (ETC), NADH:O2 oxidoreductase activity, was reduced by 30% in atrial tissue from AF compared with non-AF patients. This was predominantly due to a selective reduction in complex I (0.06 ± 0.007 vs. 0.09 ± 0.006 nmol·min(-1)·citrate synthase activity(-1), P = 0.02) and II (0.11 ± 0.012 vs. 0.16 ± 0.012 nmol·min(-1)·citrate synthase activity(-1), P = 0.003) functional activity in AF patients. Conversely, complex V activity was significantly increased in AF patients (0.21 ± 0.027 vs. 0.12 ± 0.01 nmol·min(-1)·citrate synthase activity(-1), P = 0.005). In addition, AF patients exhibited a higher oxidative stress with increased production of mitochondrial superoxide (73 ± 17 vs. 11 ± 2 arbitrary units, P = 0.03) and 4-hydroxynonenal level (77.64 ± 30.2 vs. 9.83 ± 2.83 ng·mg(-1) protein, P = 0.048). Our findings suggest that AF is associated with selective downregulation of ETC activity and increased oxidative stress that can contribute to the progression of the substrate for AF. Copyright © 2016 the American Physiological Society.

Uncoupling of oxidative phosphorylation and ATP synthase reversal within the hyperthermic heart

PubMed Central

Power, Amelia; Pearson, Nicholas; Pham, Toan; Cheung, Carlos; Phillips, Anthony; Hickey, Anthony

2014-01-01

Abstract Heart failure is a common cause of death with hyperthermia, and the exact cause of hyperthermic heart failure appears elusive. We hypothesize that the energy supply (ATP) of the heart may become impaired due to increased inner‐mitochondrial membrane permeability and inefficient oxidative phosphorylation (OXPHOS). Therefore, we assessed isolated working heart and mitochondrial function. Ex vivo working rat hearts were perfused between 37 and 43.5°C and showed break points in all functional parameters at ~40.5°C. Mitochondrial high‐resolution respirometry coupled to fluorometry was employed to determine the effects of hyperthermia on OXPHOS and mitochondrial membrane potential (ΔΨ) in vitro using a comprehensive metabolic substrate complement with isolated mitochondria. Relative to 37 and 40°C, 43°C elevated Leak O2 flux and depressed OXPHOS O2 flux and ∆Ψ. Measurement of steady‐state ATP production from mitochondria revealed decreased ATP synthesis capacity, and a negative steady‐state P:O ratio at 43°C. This approach offers a more powerful analysis of the effects of temperature on OXPHOS that cannot be measured using simple measures such as the traditional respiratory control ratio (RCR) or P:O ratio, which, respectively, can only approach 1 or 0 with inner‐membrane failure. At 40°C there was only a slight enhancement of the Leak O2 flux and this did not significantly affect ATP production rate. Therefore, during mild hyperthermia (40°C) there is no enhancement of ATP supply by mitochondria, to accompany increasing cardiac energy demands, while between this and critical hyperthermia (43°C), mitochondria become net consumers of ATP. This consumption may contribute to cardiac failure or permanent damage during severe hyperthermia. PMID:25263202

Pioglitazone Enhances Mitochondrial Biogenesis and Ribosomal Protein Biosynthesis in Skeletal Muscle in Polycystic Ovary Syndrome

PubMed Central

Skov, Vibe; Glintborg, Dorte; Knudsen, Steen; Tan, Qihua; Jensen, Thomas; Kruse, Torben A.; Beck-Nielsen, Henning; Højlund, Kurt

2008-01-01

Insulin resistance is a common metabolic abnormality in women with PCOS and leads to an elevated risk of type 2 diabetes. Studies have shown that thiazolidinediones (TZDs) improve metabolic disturbances in PCOS patients. We hypothesized that the effect of TZDs in PCOS is, in part, mediated by changes in the transcriptional profile of muscle favoring insulin sensitivity. Using Affymetrix microarrays, we examined the effect of pioglitazone (30 mg/day for 16 weeks) on gene expression in skeletal muscle of 10 obese women with PCOS metabolically characterized by a euglycemic-hyperinsulinemic clamp. Moreover, we explored gene expression changes between these PCOS patients before treatment and 13 healthy women. Treatment with pioglitazone improved insulin-stimulated glucose metabolism and plasma adiponectin, and reduced fasting serum insulin (all P<0.05). Global pathway analysis using Gene Map Annotator and Pathway Profiler (GenMAPP 2.1) and Gene Set Enrichment Analysis (GSEA 2.0.1) revealed a significant upregulation of genes representing mitochondrial oxidative phosphorylation (OXPHOS), ribosomal proteins, mRNA processing reactome, translation factors, and proteasome degradation in PCOS after pioglitazone therapy. Quantitative real-time PCR suggested that upregulation of OXPHOS genes was mediated by an increase in PGC-1α expression (P<0.05). Pretreatment expression of genes representing OXPHOS and ribosomal proteins was down-regulated in PCOS patients compared to healthy women. These data indicate that pioglitazone therapy restores insulin sensitivity, in part, by a coordinated upregulation of genes involved in mitochondrial OXPHOS and ribosomal protein biosynthesis in muscle in PCOS. These transcriptional effects of pioglitazone may contribute to prevent the onset of type 2 diabetes in these women. PMID:18560589

Energy metabolism analysis reveals the mechanism of inhibition of breast cancer cell metastasis by PEG-modified graphene oxide nanosheets.

PubMed

Zhou, Teng; Zhang, Bo; Wei, Peng; Du, Yipeng; Zhou, Hejiang; Yu, Meifang; Yan, Liang; Zhang, Wendi; Nie, Guangjun; Chen, Chunying; Tu, Yaping; Wei, Taotao

2014-12-01

Recent advances in nanomedicine provide promising alternatives for cancer treatment that may improve the survival of patients with metastatic disease. The goal of the present study was to evaluate graphene oxide (GO) as a potential anti-metastatic agent. For this purpose, GO was modified with polyethylene glycol (PEG) to form PEG-modified GO (PEG-GO), which improves its aqueous stability and biocompatibility. We show here that PEG-GO exhibited no apparent effects on the viability of breast cancer cells (MDA-MB-231, MDA-MB-436, and SK-BR-3) or non-cancerous cells (MCF-10A), but inhibited cancer cell migration in vitro and in vivo. Analysis of cellular energy metabolism revealed that PEG-GO significantly impaired mitochondrial oxidative phosphorylation (OXPHOS) in breast cancer cells; however, PEG-GO showed no effect on OXPHOS in non-cancerous cells. To explore the underlying mechanisms, a SILAC (Stable Isotope Labeling by Amino acids in Cell culture) labeling strategy was used to quantify protein expression in PEG-GO-exposed breast cancer versus non-cancerous cells. The results indicated that PEG-GO selectively down-regulated PGC-1α in breast cancer cells and thus modified the expression of diverse energy generation-related proteins, which accounts for the inhibition of OXPHOS. The inhibition of OXPHOS by PEG-GO significantly reduced ATP production and impaired assembly of the F-actin cytoskeleton in breast cancer cells, which is required for the migratory and invasive phenotype of cancer cells. Taken together, these effects of PEG-GO on cancer cell metastasis may allow the development of a new approach to treat metastatic breast cancer. Copyright © 2014 Elsevier Ltd. All rights reserved.

Mutations in the Caenorhabditis elegans orthologs of human genes required for mitochondrial tRNA modification cause similar electron transport chain defects but different nuclear responses.

PubMed

Navarro-González, Carmen; Moukadiri, Ismaïl; Villarroya, Magda; López-Pascual, Ernesto; Tuck, Simon; Armengod, M-Eugenia

2017-07-01

Several oxidative phosphorylation (OXPHOS) diseases are caused by defects in the post-transcriptional modification of mitochondrial tRNAs (mt-tRNAs). Mutations in MTO1 or GTPBP3 impair the modification of the wobble uridine at position 5 of the pyrimidine ring and cause heart failure. Mutations in TRMU affect modification at position 2 and cause liver disease. Presently, the molecular basis of the diseases and why mutations in the different genes lead to such different clinical symptoms is poorly understood. Here we use Caenorhabditis elegans as a model organism to investigate how defects in the TRMU, GTPBP3 and MTO1 orthologues (designated as mttu-1, mtcu-1, and mtcu-2, respectively) exert their effects. We found that whereas the inactivation of each C. elegans gene is associated with a mild OXPHOS dysfunction, mutations in mtcu-1 or mtcu-2 cause changes in the expression of metabolic and mitochondrial stress response genes that are quite different from those caused by mttu-1 mutations. Our data suggest that retrograde signaling promotes defect-specific metabolic reprogramming, which is able to rescue the OXPHOS dysfunction in the single mutants by stimulating the oxidative tricarboxylic acid cycle flux through complex II. This adaptive response, however, appears to be associated with a biological cost since the single mutant worms exhibit thermosensitivity and decreased fertility and, in the case of mttu-1, longer reproductive cycle. Notably, mttu-1 worms also exhibit increased lifespan. We further show that mtcu-1; mttu-1 and mtcu-2; mttu-1 double mutants display severe growth defects and sterility. The animal models presented here support the idea that the pathological states in humans may initially develop not as a direct consequence of a bioenergetic defect, but from the cell's maladaptive response to the hypomodification status of mt-tRNAs. Our work highlights the important association of the defect-specific metabolic rewiring with the pathological phenotype, which must be taken into consideration in exploring specific therapeutic interventions.

Application of mitochondrial pyruvate carrier blocker UK5099 creates metabolic reprogram and greater stem-like properties in LnCap prostate cancer cells in vitro.

PubMed

Zhong, Yali; Li, Xiaoran; Yu, Dandan; Li, Xiaoli; Li, Yaqing; Long, Yuan; Yuan, Yuan; Ji, Zhenyu; Zhang, Mingzhi; Wen, Jian-Guo; Nesland, Jahn M; Suo, Zhenhe

2015-11-10

Aerobic glycolysis is one of the important hallmarks of cancer cells and eukaryotic cells. In this study, we have investigated the relationship between blocking mitochondrial pyruvate carrier (MPC) with UK5099 and the metabolic alteration as well as stemness phenotype of prostatic cancer cells. It was found that blocking pyruvate transportation into mitochondrial attenuated mitochondrial oxidative phosphorylation (OXPHOS) and increased glycolysis. The UK5099 treated cells showed significantly higher proportion of side population (SP) fraction and expressed higher levels of stemness markers Oct3/4 and Nanog. Chemosensitivity examinations revealed that the UK5099 treated cells became more resistant to chemotherapy compared to the non-treated cells. These results demonstrate probably an intimate connection between metabolic reprogram and stem-like phenotype of LnCap cells in vitro. We propose that MPC blocker (UK5099) application may be an ideal model for Warburg effect studies, since it attenuates mitochondrial OXPHOS and increases aerobic glycolysis, a phenomenon typically reflected in the Warburg effect. We conclude that impaired mitochondrial OXPHOS and upregulated glycolysis are related with stem-like phenotype shift in prostatic cancer cells.

Application of mitochondrial pyruvate carrier blocker UK5099 creates metabolic reprogram and greater stem-like properties in LnCap prostate cancer cells in vitro

PubMed Central

Zhong, Yali; Li, Xiaoran; Yu, Dandan; Li, Xiaoli; Li, Yaqing; Long, Yuan; Yuan, Yuan; Ji, Zhenyu; Zhang, Mingzhi; Wen, Jian-Guo; Nesland, Jahn M.; Suo, Zhenhe

2015-01-01

Aerobic glycolysis is one of the important hallmarks of cancer cells and eukaryotic cells. In this study, we have investigated the relationship between blocking mitochondrial pyruvate carrier (MPC) with UK5099 and the metabolic alteration as well as stemness phenotype of prostatic cancer cells. It was found that blocking pyruvate transportation into mitochondrial attenuated mitochondrial oxidative phosphorylation (OXPHOS) and increased glycolysis. The UK5099 treated cells showed significantly higher proportion of side population (SP) fraction and expressed higher levels of stemness markers Oct3/4 and Nanog. Chemosensitivity examinations revealed that the UK5099 treated cells became more resistant to chemotherapy compared to the non-treated cells. These results demonstrate probably an intimate connection between metabolic reprogram and stem-like phenotype of LnCap cells in vitro. We propose that MPC blocker (UK5099) application may be an ideal model for Warburg effect studies, since it attenuates mitochondrial OXPHOS and increases aerobic glycolysis, a phenomenon typically reflected in the Warburg effect. We conclude that impaired mitochondrial OXPHOS and upregulated glycolysis are related with stem-like phenotype shift in prostatic cancer cells. PMID:26413751

Inhibition of AMPK and Krebs cycle gene expression drives metabolic remodeling of Pten-deficient preneoplastic thyroid cells.

PubMed

Antico Arciuch, Valeria G; Russo, Marika A; Kang, Kristy S; Di Cristofano, Antonio

2013-09-01

Rapidly proliferating and neoplastically transformed cells generate the energy required to support rapid cell division by increasing glycolysis and decreasing flux through the oxidative phosphorylation (OXPHOS) pathway, usually without alterations in mitochondrial function. In contrast, little is known of the metabolic alterations, if any, which occur in cells harboring mutations that prime their neoplastic transformation. To address this question, we used a Pten-deficient mouse model to examine thyroid cells where a mild hyperplasia progresses slowly to follicular thyroid carcinoma. Using this model, we report that constitutive phosphoinositide 3-kinase (PI3K) activation caused by PTEN deficiency in nontransformed thyrocytes results in a global downregulation of Krebs cycle and OXPHOS gene expression, defective mitochondria, reduced respiration, and an enhancement in compensatory glycolysis. We found that this process does not involve any of the pathways classically associated with the Warburg effect. Moreover, this process was independent of proliferation but contributed directly to thyroid hyperplasia. Our findings define a novel metabolic switch to glycolysis driven by PI3K-dependent AMPK inactivation with a consequent repression in the expression of key metabolic transcription regulators. ©2013 AACR.

Green Tea Polyphenols Stimulate Mitochondrial Biogenesis and Improve Renal Function after Chronic Cyclosporin A Treatment in Rats

PubMed Central

Rehman, Hasibur; Krishnasamy, Yasodha; Haque, Khujista; Lemasters, John J.; Schnellmann, Rick G.; Zhong, Zhi

2013-01-01

Our previous studies showed that an extract from Camellia sinenesis (green tea), which contains several polyphenols, attenuates nephrotoxicity caused by cyclosporine A (CsA). Since polyphenols are stimulators of mitochondrial biogenesis (MB), this study investigated whether stimulation of MB plays a role in green tea polyphenol protection against CsA renal toxicity. Rats were fed a powdered diet containing green tea polyphenolic extract (0.1%) starting 3 days prior to CsA treatment (25 mg/kg, i.g. daily for 3 weeks). CsA alone decreased renal nuclear DNA-encoded oxidative phosphorylation (OXPHOS) protein ATP synthase-β (AS-β) by 42%, mitochondrial DNA (mtDNA)-encoded OXPHOS protein NADH dehydrogenase-3 (ND3) by 87% and their associated mRNAs. Mitochondrial DNA copy number was also decreased by 78% by CsA. Immunohistochemical analysis showed decreased cytochrome c oxidase subunit IV (COX-IV), an OXPHOS protein, in tubular cells. Peroxisome proliferator-activated receptor-γ coactivator (PGC)-1α, the master regulator of MB, and mitochondrial transcription factor-A (Tfam), the transcription factor that regulates mtDNA replication and transcription, were 42% and 90% lower, respectively, in the kidneys of CsA-treated than in untreated rats. These results indicate suppression of MB by chronic CsA treatment. Green tea polyphenols alone and following CsA increased AS-β, ND3, COX-IV, mtDNA copy number, PGC-1α mRNA and protein, decreased acetylated PGC-1α, and increased Tfam mRNA and protein. In association with suppressed MB, CsA increased serum creatinine, caused loss of brush border and dilatation of proximal tubules, tubular atrophy, vacuolization, apoptosis, calcification, and increased neutrophil gelatinase-associated lipocalin expression, leukocyte infiltration, and renal fibrosis. Green tea polyphenols markedly attenuated CsA-induced renal injury and improved renal function. Together, these results demonstrate that green tea polyphenols attenuate CsA-induced kidney injury, at least in part, through the stimulation of MB. PMID:23755172

SCO2 induces p53-mediated apoptosis by Thr845 phosphorylation of ASK-1 and dissociation of the ASK-1-Trx complex.

PubMed

Madan, Esha; Gogna, Rajan; Kuppusamy, Periannan; Bhatt, Madan; Mahdi, Abbas Ali; Pati, Uttam

2013-04-01

p53 prevents cancer via cell cycle arrest, apoptosis, and the maintenance of genome stability. p53 also regulates energy-generating metabolic pathways such as oxidative phosphorylation (OXPHOS) and glycolysis via transcriptional regulation of SCO2 and TIGAR. SCO2, a cytochrome c oxidase assembly factor, is a metallochaperone which is involved in the biogenesis of cytochrome c oxidase subunit II. Here we have shown that SCO2 functions as an apoptotic protein in tumor xenografts, thus providing an alternative pathway for p53-mediated apoptosis. SCO2 increases the generation of reactive oxygen species (ROS) and induces dissociation of the protein complex between apoptosis signal-regulating kinase 1 (ASK-1) (mitogen-activated protein kinase kinase kinase [MAPKKK]) and its cellular inhibitor, the redox-active protein thioredoxin (Trx). Furthermore, SCO2 induces phosphorylation of ASK-1 at the Thr(845) residue, resulting in the activation of the ASK-1 kinase pathway. The phosphorylation of ASK-1 induces the activation of mitogen-activated protein kinase kinases 4 and 7 (MAP2K4/7) and MAP2K3/6, which switches the c-Jun N-terminal protein kinase (JNK)/p38-dependent apoptotic cascades in cancer cells. Exogenous addition of the SCO2 gene to hypoxic cancer cells and hypoxic tumors induces apoptosis and causes significant regression of tumor xenografts. We have thus discovered a novel apoptotic function of SCO2, which activates the ASK-1 kinase pathway in switching "on" an alternate mode of p53-mediated apoptosis. We propose that SCO2 might possess a novel tumor suppressor function via the ROS-ASK-1 kinase pathway and thus could be an important candidate for anticancer gene therapy.

Inverse relationship between exercise economy and oxidative capacity in muscle.

PubMed

Hunter, Gary R; Bamman, Marcas M; Larson-Meyer, D Enette; Joanisse, Denis R; McCarthy, John P; Blaudeau, Tamilane E; Newcomer, Bradley R

2005-08-01

An inverse relationship has been shown between running and cycling exercise economy and maximum oxygen uptake (VO2max). The purposes were: 1) determine the relationship between walking economy and VO2max; and 2) determine the relationship between muscle metabolic economy and muscle oxidative capacity and fiber type. Subjects were 77 premenopausal normal weight women. Walking economy (1/VO2max) was measured at 3 mph and VO2max during graded treadmill test. Muscle oxidative phosphorylation rate (OxPhos), and muscle metabolic economy (force/ATP) were measured in calf muscle using 31P MRS during isometric plantar flexion at 70 and 100% of maximum force, (HI) and (MI) respectively. Muscle fiber type and citrate synthase activity were determined in the lateral gastrocnemius. Significant inverse relationships (r from -0.28 to -0.74) were observed between oxidative metabolism measures and exercise economy (walking and muscle). Type IIa fiber distribution was inversely related to all measures of exercise economy (r from -0.51 to -0.64) and citrate synthase activity was inversely related to muscle metabolic economy at MI (r = -0.56). In addition, Type IIa fiber distribution and citrate synthase activity were positively related to VO2max and muscle OxPhos at HI and MI (r from 0.49 to 0.70). Type I fiber distribution was not related to any measure of exercise economy or oxidative capacity. Our results support the concept that exercise economy and oxidative capacity are inversely related. We have demonstrated this inverse relationship in women both by indirect calorimetry during walking and in muscle tissue by 31P MRS.

Effects of argan oil on the mitochondrial function, antioxidant system and the activity of NADPH- generating enzymes in acrylamide treated rat brain.

PubMed

Aydın, Birsen

2017-03-01

Argan oil (AO) is rich in minor compounds such as polyphenols and tocopherols which are powerful antioxidants. Acrylamide (ACR) has been classified as a neurotoxic agent in animals and humans. Mitochondrial oxidative stress and dysfunction is one of the most probable molecular mechanisms of neurodegenerative diseases. Female Sprague Dawley rats were exposed to ACR (50mg/kg i.p. three times a week), AO (6ml/kg,o.p, per day) or together for 30days. The activities of cytosolic enzymes such as xanthine oxidase (XO), glucose 6-phosphate dehydrogenase (G6PDH), glutathione-S-transferase (GST), mitochondrial oxidative stress, oxidative phosphorylation (OXPHOS) and tricarboxylic acid cycle (TCA) enzymes, mitochondrial metabolic function, adenosine triphosphate (ATP) level and acetylcholinesterase (AChE) activity were assessed in rat brain. Cytosolic and mitochondrial antioxidant enzymes were significantly diminished in the brains of rats treated with ACR compared to those in control. Besides, ACR treatment resulted in a significant reduction in brain ATP level, mitochondrial metabolic function, OXPHOS and TCA enzymes. Administration of AO restored both the cytosolic and mitochondrial oxidative stress by normalizing nicotinamide adenine dinucleotide phosphate (NADPH) generating enzymes. In addition, improved mitochondrial function primarily enhancing nicotinamide adenine dinucleotide (NADH) generated enzymes activities and ATP level in the mitochondria. The reason for AO's obvious beneficial effects in this study may be due to synergistic effects of its different bioactive compounds which is especially effective on mitochondria. Modulation of the brain mitochondrial functions and antioxidant systems by AO may lead to the development of new mitochondria-targeted antioxidants in the future. Copyright © 2016 Elsevier Masson SAS. All rights reserved.

Uncoupling of oxidative phosphorylation and ATP synthase reversal within the hyperthermic heart.

PubMed

Power, Amelia; Pearson, Nicholas; Pham, Toan; Cheung, Carlos; Phillips, Anthony; Hickey, Anthony

2014-09-01

Heart failure is a common cause of death with hyperthermia, and the exact cause of hyperthermic heart failure appears elusive. We hypothesize that the energy supply (ATP) of the heart may become impaired due to increased inner-mitochondrial membrane permeability and inefficient oxidative phosphorylation (OXPHOS). Therefore, we assessed isolated working heart and mitochondrial function. Ex vivo working rat hearts were perfused between 37 and 43.5°C and showed break points in all functional parameters at ~40.5°C. Mitochondrial high-resolution respirometry coupled to fluorometry was employed to determine the effects of hyperthermia on OXPHOS and mitochondrial membrane potential (ΔΨ) in vitro using a comprehensive metabolic substrate complement with isolated mitochondria. Relative to 37 and 40°C, 43°C elevated Leak O2 flux and depressed OXPHOS O2 flux and ∆Ψ. Measurement of steady-state ATP production from mitochondria revealed decreased ATP synthesis capacity, and a negative steady-state P:O ratio at 43°C. This approach offers a more powerful analysis of the effects of temperature on OXPHOS that cannot be measured using simple measures such as the traditional respiratory control ratio (RCR) or P:O ratio, which, respectively, can only approach 1 or 0 with inner-membrane failure. At 40°C there was only a slight enhancement of the Leak O2 flux and this did not significantly affect ATP production rate. Therefore, during mild hyperthermia (40°C) there is no enhancement of ATP supply by mitochondria, to accompany increasing cardiac energy demands, while between this and critical hyperthermia (43°C), mitochondria become net consumers of ATP. This consumption may contribute to cardiac failure or permanent damage during severe hyperthermia. © 2014 The Authors. Physiological Reports published by Wiley Periodicals, Inc. on behalf of the American Physiological Society and The Physiological Society.

Biallelic Mutations in MRPS34 Lead to Instability of the Small Mitoribosomal Subunit and Leigh Syndrome.

PubMed

Lake, Nicole J; Webb, Bryn D; Stroud, David A; Richman, Tara R; Ruzzenente, Benedetta; Compton, Alison G; Mountford, Hayley S; Pulman, Juliette; Zangarelli, Coralie; Rio, Marlene; Boddaert, Nathalie; Assouline, Zahra; Sherpa, Mingma D; Schadt, Eric E; Houten, Sander M; Byrnes, James; McCormick, Elizabeth M; Zolkipli-Cunningham, Zarazuela; Haude, Katrina; Zhang, Zhancheng; Retterer, Kyle; Bai, Renkui; Calvo, Sarah E; Mootha, Vamsi K; Christodoulou, John; Rötig, Agnes; Filipovska, Aleksandra; Cristian, Ingrid; Falk, Marni J; Metodiev, Metodi D; Thorburn, David R

2017-08-03

The synthesis of all 13 mitochondrial DNA (mtDNA)-encoded protein subunits of the human oxidative phosphorylation (OXPHOS) system is carried out by mitochondrial ribosomes (mitoribosomes). Defects in the stability of mitoribosomal proteins or mitoribosome assembly impair mitochondrial protein translation, causing combined OXPHOS enzyme deficiency and clinical disease. Here we report four autosomal-recessive pathogenic mutations in the gene encoding the small mitoribosomal subunit protein, MRPS34, in six subjects from four unrelated families with Leigh syndrome and combined OXPHOS defects. Whole-exome sequencing was used to independently identify all variants. Two splice-site mutations were identified, including homozygous c.321+1G>T in a subject of Italian ancestry and homozygous c.322-10G>A in affected sibling pairs from two unrelated families of Puerto Rican descent. In addition, compound heterozygous MRPS34 mutations were identified in a proband of French ancestry; a missense (c.37G>A [p.Glu13Lys]) and a nonsense (c.94C>T [p.Gln32 ∗ ]) variant. We demonstrated that these mutations reduce MRPS34 protein levels and the synthesis of OXPHOS subunits encoded by mtDNA. Examination of the mitoribosome profile and quantitative proteomics showed that the mitochondrial translation defect was caused by destabilization of the small mitoribosomal subunit and impaired monosome assembly. Lentiviral-mediated expression of wild-type MRPS34 rescued the defect in mitochondrial translation observed in skin fibroblasts from affected subjects, confirming the pathogenicity of MRPS34 mutations. Our data establish that MRPS34 is required for normal function of the mitoribosome in humans and furthermore demonstrate the power of quantitative proteomic analysis to identify signatures of defects in specific cellular pathways in fibroblasts from subjects with inherited disease. Copyright © 2017 American Society of Human Genetics. Published by Elsevier Inc. All rights reserved.

Renal oncocytoma characterized by the defective complex I of the respiratory chain boosts the synthesis of the ROS scavenger glutathione.

PubMed

Kürschner, Gerrit; Zhang, Qingzhou; Clima, Rosanna; Xiao, Yi; Busch, Jonas Felix; Kilic, Ergin; Jung, Klaus; Berndt, Nikolaus; Bulik, Sascha; Holzhütter, Hermann-Georg; Gasparre, Giuseppe; Attimonelli, Marcella; Babu, Mohan; Meierhofer, David

2017-12-01

Renal oncocytomas are rare benign tumors of the kidney and characterized by a deficient complex I (CI) enzyme activity of the oxidative phosphorylation (OXPHOS) system caused by mitochondrial DNA (mtDNA) mutations. Yet, little is known about the underlying molecular mechanisms and alterations of metabolic pathways in this tumor. We compared renal oncocytomas with adjacent matched normal kidney tissues on a global scale by multi-omics approaches, including whole exome sequencing (WES), proteomics, metabolomics, and metabolic pathway simulation. The abundance of proteins localized to mitochondria increased more than 2-fold, the only exception was a strong decrease in the abundance for CI subunits that revealed several pathogenic heteroplasmic mtDNA mutations by WES. We also observed renal oncocytomas to dysregulate main metabolic pathways, shunting away from gluconeogenesis and lipid metabolism. Nevertheless, the abundance of energy carrier molecules such as NAD + , NADH, NADP, ATP, and ADP were significantly higher in renal oncocytomas. Finally, a substantial 5000-fold increase of the reactive oxygen species scavenger glutathione can be regarded as a new hallmark of renal oncocytoma. Our findings demonstrate that renal oncocytomas undergo a metabolic switch to eliminate ATP consuming processes to ensure a sufficient energy supply for the tumor.

Renal oncocytoma characterized by the defective complex I of the respiratory chain boosts the synthesis of the ROS scavenger glutathione

PubMed Central

Clima, Rosanna; Xiao, Yi; Busch, Jonas Felix; Kilic, Ergin; Jung, Klaus; Berndt, Nikolaus; Bulik, Sascha; Holzhütter, Hermann-Georg; Gasparre, Giuseppe; Attimonelli, Marcella; Babu, Mohan; Meierhofer, David

2017-01-01

Renal oncocytomas are rare benign tumors of the kidney and characterized by a deficient complex I (CI) enzyme activity of the oxidative phosphorylation (OXPHOS) system caused by mitochondrial DNA (mtDNA) mutations. Yet, little is known about the underlying molecular mechanisms and alterations of metabolic pathways in this tumor. We compared renal oncocytomas with adjacent matched normal kidney tissues on a global scale by multi-omics approaches, including whole exome sequencing (WES), proteomics, metabolomics, and metabolic pathway simulation. The abundance of proteins localized to mitochondria increased more than 2-fold, the only exception was a strong decrease in the abundance for CI subunits that revealed several pathogenic heteroplasmic mtDNA mutations by WES. We also observed renal oncocytomas to dysregulate main metabolic pathways, shunting away from gluconeogenesis and lipid metabolism. Nevertheless, the abundance of energy carrier molecules such as NAD+, NADH, NADP, ATP, and ADP were significantly higher in renal oncocytomas. Finally, a substantial 5000-fold increase of the reactive oxygen species scavenger glutathione can be regarded as a new hallmark of renal oncocytoma. Our findings demonstrate that renal oncocytomas undergo a metabolic switch to eliminate ATP consuming processes to ensure a sufficient energy supply for the tumor. PMID:29285300

Polyethylenimine architecture-dependent metabolic imprints and perturbation of cellular redox homeostasis.

PubMed

Hall, Arnaldur; Parhamifar, Ladan; Lange, Marina Krarup; Meyle, Kathrine Damm; Sanderhoff, May; Andersen, Helene; Roursgaard, Martin; Larsen, Anna Karina; Jensen, Per Bo; Christensen, Claus; Bartek, Jiri; Moghimi, Seyed Moein

2015-03-01

Polyethylenimines (PEIs) are among the most efficient polycationic non-viral transfectants. PEI architecture and size not only modulate transfection efficiency, but also cytotoxicity. However, the underlying mechanisms of PEI-induced multifaceted cell damage and death are largely unknown. Here, we demonstrate that the central mechanisms of PEI architecture- and size-dependent perturbations of integrated cellular metabolomics involve destabilization of plasma membrane and mitochondrial membranes with consequences on mitochondrial oxidative phosphorylation (OXPHOS), glycolytic flux and redox homeostasis that ultimately modulate cell death. In comparison to linear PEI, the branched architectures induced greater plasma membrane destabilization and were more detrimental to glycolytic activity and OXPHOS capacity as well as being a more potent inhibitor of the cytochrome c oxidase. Accordingly, the branched architectures caused a greater lactate dehydrogenase (LDH) and ATP depletion, activated AMP kinase (AMPK) and disturbed redox homeostasis through diminished availability of nicotinamide adenine dinucleotide phosphate (NADPH), reduced antioxidant capacity of glutathione (GSH) and increased burden of reactive oxygen species (ROS). The differences in metabolic and redox imprints were further reflected in the transfection performance of the polycations, but co-treatment with the GSH precursor N-acetyl-cysteine (NAC) counteracted redox dysregulation and increased the number of viable transfected cells. Integrated biomembrane integrity and metabolomic analysis provides a rapid approach for mechanistic understanding of multifactorial polycation-mediated cytotoxicity, and could form the basis for combinatorial throughput platforms for improved design and selection of safer polymeric vectors. Copyright © 2014 Elsevier B.V. All rights reserved.

Histone Deacetylase 3 Prepares Brown Adipose Tissue For Acute Thermogenic Challenge

PubMed Central

Emmett, Matthew J.; Lim, Hee-Woong; Jager, Jennifer; Richter, Hannah J.; Adlanmerini, Marine; Peed, Lindsey C.; Briggs, Erika R.; Steger, David J.; Ma, Tao; Sims, Carrie A.; Baur, Joseph A.; Pei, Liming; Won, Kyoung-Jae; Seale, Patrick; Gerhart-Hines, Zachary; Lazar, Mitchell A.

2017-01-01

Brown adipose tissue (BAT) is a thermogenic organ that dissipates chemical energy as heat to protect animals against hypothermia and to counteract metabolic disease1. However, the transcriptional mechanisms that determine BAT thermogenic capacity prior to environmental cold are unknown. Here we show that Histone Deacetylase 3 (HDAC3) is required to activate BAT enhancers to ensure thermogenic aptitude. Mice with BAT-specific genetic ablation of HDAC3 become severely hypothermic and succumb to acute cold exposure. UCP1 is nearly absent in BAT lacking HDAC3 and there is also marked down-regulation of mitochondrial oxidative phosphorylation (OXPHOS) genes resulting in diminished mitochondrial respiration. Remarkably, although HDAC3 acts canonically as a transcriptional corepressor2, it functions as a coactivator of Estrogen-Related Receptor α (ERRα) in BAT. HDAC3 coactivation of ERRα is mediated by deacetylation of PGC-1α and is required for the transcription of Ucp1, Pgc-1α and OXPHOS genes. Importantly, HDAC3 promotes the basal transcription of these genes independent of adrenergic stimulation. Thus, HDAC3 uniquely primes Ucp1 and the thermogenic transcriptional program to maintain a critical capacity for thermogenesis in BAT that can be rapidly engaged upon exposure to dangerously cold temperature. PMID:28614293

Mitophagy is required for mitochondrial biogenesis and myogenic differentiation of C2C12 myoblasts.

PubMed

Sin, Jon; Andres, Allen M; Taylor, David J R; Weston, Thomas; Hiraumi, Yoshimi; Stotland, Aleksandr; Kim, Brandon J; Huang, Chengqun; Doran, Kelly S; Gottlieb, Roberta A

2016-01-01

Myogenesis is a crucial process governing skeletal muscle development and homeostasis. Differentiation of primitive myoblasts into mature myotubes requires a metabolic switch to support the increased energetic demand of contractile muscle. Skeletal myoblasts specifically shift from a highly glycolytic state to relying predominantly on oxidative phosphorylation (OXPHOS) upon differentiation. We have found that this phenomenon requires dramatic remodeling of the mitochondrial network involving both mitochondrial clearance and biogenesis. During early myogenic differentiation, autophagy is robustly upregulated and this coincides with DNM1L/DRP1 (dynamin 1-like)-mediated fragmentation and subsequent removal of mitochondria via SQSTM1 (sequestosome 1)-mediated mitophagy. Mitochondria are then repopulated via PPARGC1A/PGC-1α (peroxisome proliferator-activated receptor gamma, coactivator 1 alpha)-mediated biogenesis. Mitochondrial fusion protein OPA1 (optic atrophy 1 [autosomal dominant]) is then briskly upregulated, resulting in the reformation of mitochondrial networks. The final product is a myotube replete with new mitochondria. Respirometry reveals that the constituents of these newly established mitochondrial networks are better primed for OXPHOS and are more tightly coupled than those in myoblasts. Additionally, we have found that suppressing autophagy with various inhibitors during differentiation interferes with myogenic differentiation. Together these data highlight the integral role of autophagy and mitophagy in myogenic differentiation.

miR-504 mediated down-regulation of nuclear respiratory factor 1 leads to radio-resistance in nasopharyngeal carcinoma

PubMed Central

Zhao, Luqing; Tang, Min; Hu, Zheyu; Yan, Bin; Pi, Weiwei; Li, Zhi; Zhang, Jing; Zhang, Liqin; Jiang, Wuzhong; Li, Guo; Qiu, Yuanzheng; Hu, Fang; Liu, Feng; Lu, Jingchen; Chen, Xue; Xiao, Lanbo; Xu, Zhijie; Tao, Yongguang; Yang, Lifang; Bode, Ann M.; Dong, Zigang; Zhou, Jian; Fan, Jia; Sun, Lunquan; Cao, Ya

2015-01-01

microRNAs (miRNAs) are involved in the various processes of DNA damage repair and play crucial roles in regulating response of tumors to radiation therapy. Here, we used nasopharyngeal carcinoma (NPC) radio-resistant cell lines as models and found that the expression of miR-504 was significantly up-regulated. In contrast, the expression of nuclear respiratory factor 1 (NRF1) and other mitochondrial metabolism factors, including mitochondrial transcription factor A (TFAM) and oxidative phosphorylation (OXPHOS) complex III were down-regulated in these cell lines. At the same time, the Seahorse cell mitochondrial stress test results indicated that the mitochondrial respiratory capacity was impaired in NPC radio-resistant cell lines and in a miR-504 over-expressing cell line. We also conducted dual luciferase reporter assays and verified that miR-504 could directly target NRF1. Additionally, miR-504 could down-regulate the expression of TFAM and OXPHOS complexes I, III, and IV and impaired the mitochondrial respiratory function of NPC cells. Furthermore, serum from NPC patients showed that miR-504 was up-regulated during different weeks of radiotherapy and correlated with tumor, lymph nodes and metastasis (TNM) stages and total tumor volume. The radio-therapeutic effect at three months after radiotherapy was evaluated. Results indicated that patients with high expression of miR-504 exhibited a relatively lower therapeutic effect ratio of complete response (CR), but a higher ratio of partial response (PR), compared to patients with low expression of miR-504. Taken together, these results demonstrated that miR-504 affected the radio-resistance of NPC by down-regulating the expression of NRF1 and disturbing mitochondrial respiratory function. Thus, miR-504 might become a promising biomarker of NPC radio-resistance and targeting miR-504 might improve tumor radiation response. PMID:26201446

Inhibition of oxidative phosphorylation suppresses the development of osimertinib resistance in a preclinical model of EGFR-driven lung adenocarcinoma.

PubMed

Martin, Matthew J; Eberlein, Cath; Taylor, Molly; Ashton, Susan; Robinson, David; Cross, Darren

2016-12-27

Metabolic plasticity is an emerging hallmark of cancer, and increased glycolysis is often observed in transformed cells. Small molecule inhibitors that target driver oncogenes can potentially inhibit the glycolytic pathway. Osimertinib (AZD9291) is a novel EGFR tyrosine kinase inhibitor (TKI) that is potent and selective for sensitising (EGFRm) and T790M resistance mutations. Clinical studies have shown osimertinib to be efficacious in patients with EGFRm/ T790M advanced NSCLC who have progressed after EGFR-TKI treatment. However experience with targeted therapies suggests that acquired resistance may emerge. Thus there is a need to characterize resistance mechanisms and to devise ways to prevent, delay or overcome osimertinib resistance. We show here that osimertinib suppresses glycolysis in parental EGFR-mutant lung adenocarcinoma lines, but has not in osimertinib-resistant cell lines. Critically, we show osimertinib treatment induces a strict dependence on mitochondrial oxidative phosphorylation (OxPhos), as OxPhos inhibitors significantly delay the long-term development of osimertinib resistance in osimertinib-sensitive lines. Accordingly, growth conditions which promote a less glycolytic phenotype confer a degree of osimertinib resistance. Our data support a model in which the combination of osimertinib and OxPhos inhibitors can delay or prevent resistance in osimertinib-naïve tumour cells, and represents a novel strategy that warrants further pre-clinical investigation.

Inhibition of oxidative phosphorylation suppresses the development of osimertinib resistance in a preclinical model of EGFR-driven lung adenocarcinoma

PubMed Central

Martin, Matthew J.; Eberlein, Cath; Taylor, Molly; Ashton, Susan; Robinson, David; Cross, Darren

2016-01-01

Metabolic plasticity is an emerging hallmark of cancer, and increased glycolysis is often observed in transformed cells. Small molecule inhibitors that target driver oncogenes can potentially inhibit the glycolytic pathway. Osimertinib (AZD9291) is a novel EGFR tyrosine kinase inhibitor (TKI) that is potent and selective for sensitising (EGFRm) and T790M resistance mutations. Clinical studies have shown osimertinib to be efficacious in patients with EGFRm/ T790M advanced NSCLC who have progressed after EGFR-TKI treatment. However experience with targeted therapies suggests that acquired resistance may emerge. Thus there is a need to characterize resistance mechanisms and to devise ways to prevent, delay or overcome osimertinib resistance. We show here that osimertinib suppresses glycolysis in parental EGFR-mutant lung adenocarcinoma lines, but has not in osimertinib-resistant cell lines. Critically, we show osimertinib treatment induces a strict dependence on mitochondrial oxidative phosphorylation (OxPhos), as OxPhos inhibitors significantly delay the long-term development of osimertinib resistance in osimertinib-sensitive lines. Accordingly, growth conditions which promote a less glycolytic phenotype confer a degree of osimertinib resistance. Our data support a model in which the combination of osimertinib and OxPhos inhibitors can delay or prevent resistance in osimertinib-naïve tumour cells, and represents a novel strategy that warrants further pre-clinical investigation. PMID:27861144

Mitochondrial transcription: Lessons from mouse models

PubMed Central

Peralta, Susana; Wang, Xiao; Moraes, Carlos T.

2012-01-01

Mammalian mitochondrial DNA (mtDNA) is a circular double-stranded DNA genome of ∼ 16.5 kilobase pairs (kb) that encodes 13 catalytic proteins of the ATP-producing oxidative phosphorylation system (OXPHOS), and the rRNAs and tRNAs required for the translation of the mtDNA transcripts. All the components needed for transcription and replication of the mtDNA are, therefore, encoded in the nuclear genome, as are the remaining components of the OXPHOS system and the mitochondrial translation machinery. Regulation of mtDNA gene expression is very important for modulating the OXPHOS capacity in response to metabolic requirements and in pathological processes. The combination of in vitro and in vivo studies has allowed the identification of the core machinery required for basal mtDNA transcription in mammals and a few proteins that regulate mtDNA transcription. Specifically, the generation of knockout mouse strains in the last several years, has been key to understanding the basis of mtDNA transcription in vivo. However, it is well accepted that many components of the transcription machinery are still unknown and little is known about mtDNA gene expression regulation under different metabolic requirements or disease processes. In this review we will focus on how the creation of knockout mouse models and the study of their phenotypes have contributed to the understanding of mitochondrial transcription in mammals. PMID:22120174

Cellular bioenergetics is impaired in patients with chronic fatigue syndrome.

PubMed

Tomas, Cara; Brown, Audrey; Strassheim, Victoria; Elson, Joanna L; Newton, Julia; Manning, Philip

2017-01-01

Chronic fatigue syndrome (CFS) is a highly debilitating disease of unknown aetiology. Abnormalities in bioenergetic function have been cited as one possible cause for CFS. Preliminary studies were performed to investigate cellular bioenergetic abnormalities in CFS patients. A series of assays were conducted using peripheral blood mononuclear cells (PBMCs) from CFS patients and healthy controls. These experiments investigated cellular patterns in oxidative phosphorylation (OXPHOS) and glycolysis. Results showed consistently lower measures of OXPHOS parameters in PBMCs taken from CFS patients compared with healthy controls. Seven key parameters of OXPHOS were calculated: basal respiration, ATP production, proton leak, maximal respiration, reserve capacity, non-mitochondrial respiration, and coupling efficiency. While many of the parameters differed between the CFS and control cohorts, maximal respiration was determined to be the key parameter in mitochondrial function to differ between CFS and control PBMCs due to the consistency of its impairment in CFS patients found throughout the study (p≤0.003). The lower maximal respiration in CFS PBMCs suggests that when the cells experience physiological stress they are less able to elevate their respiration rate to compensate for the increase in stress and are unable to fulfil cellular energy demands. The metabolic differences discovered highlight the inability of CFS patient PBMCs to fulfil cellular energetic demands both under basal conditions and when mitochondria are stressed during periods of high metabolic demand.

MITOCHONDRIAL DISEASES PART III: THERAPEUTIC INTERVENTIONS IN MOUSE MODELS OF OXPHOS DEFICIENCIES

PubMed Central

Peralta, Susana; Torraco, Alessandra; Iommarini, Luisa; Diaz, Francisca

2015-01-01

Mitochondrial defects are the cause of numerous disorders affecting the oxidative phosphorylation system (OXPHOS) in humans leading predominantly to neurological and muscular degeneration. The molecular origin, manifestations, and progression of mitochondrial diseases have a broad spectrum, which makes very challenging to find a globally effective therapy. The study of the molecular mechanisms underlying the mitochondrial dysfunction indicates that there is a wide range of pathways, enzymes and molecules that could be potentially targeted for therapeutic purpose. Therefore, focusing on the pathology of the disease is essential to design new treatments. In this review, we will summarize and discuss the different therapeutic interventions tested in some mouse models of mitochondrial diseases laying emphasis on the molecular mechanisms of action and their potential applications. PMID:25638392

Mitochondrial and glycolytic metabolic compartmentalization in diffuse large B-cell lymphoma.

PubMed

Gooptu, Mahasweta; Whitaker-Menezes, Diana; Sprandio, John; Domingo-Vidal, Marina; Lin, Zhao; Uppal, Guldeep; Gong, Jerald; Fratamico, Roberto; Leiby, Benjamin; Dulau-Florea, Alina; Caro, Jaime; Martinez-Outschoorn, Ubaldo

2017-06-01

Metabolic heterogeneity between neoplastic cells and surrounding stroma has been described in several epithelial malignancies; however, the metabolic phenotypes of neoplastic lymphocytes and neighboring stroma in diffuse large B-cell lymphoma (DLBCL) is unknown. We investigated the metabolic phenotypes of human DLBCL tumors by using immunohistochemical markers of glycolytic and mitochondrial oxidative phosphorylation (OXPHOS) metabolism. The lactate importer MCT4 is a marker of glycolysis, whereas the lactate importer MCT1 and TOMM20 are markers of OXPHOS metabolism. Staining patterns were assessed in 33 DLBCL samples as well as 18 control samples (non-neoplastic lymph nodes). TOMM20 and MCT1 were highly expressed in neoplastic lymphocytes, indicating an OXPHOS phenotype, whereas non-neoplastic lymphocytes in the control samples did not express these markers. Stromal cells in DLBCL samples strongly expressed MCT4, displaying a glycolytic phenotype, a feature not seen in stromal elements of non-neoplastic lymphatic tissue. Furthermore, the differential expression of lactate exporters (MCT4) on tumor-associated stroma and lactate importers (MCT1) on neoplastic lymphocytes support the hypothesis that neoplastic cells are metabolically linked to the stroma likely via mutually beneficial reprogramming. MCT4 is a marker of tumor-associated stroma in neoplastic tissue. Our findings suggest that disruption of neoplastic-stromal cell metabolic heterogeneity including MCT1 and MCT4 blockade should be studied to determine if it could represent a novel treatment target in DLBCL. Copyright © 2017 Elsevier Inc. All rights reserved.

The regulation of OXPHOS by extramitochondrial calcium.

PubMed

Gellerich, Frank N; Gizatullina, Zemfira; Trumbeckaite, Sonata; Nguyen, Huu P; Pallas, Thilo; Arandarcikaite, Odeta; Vielhaber, Stephan; Seppet, Enn; Striggow, Frank

2010-01-01

Despite extensive research, the regulation of mitochondrial function is still not understood completely. Ample evidence shows that cytosolic Ca2+ has a strategic task in co-ordinating the cellular work load and the regeneration of ATP by mitochondria. Currently, the paradigmatic view is that Cacyt2+ taken up by the Ca2+ uniporter activates the matrix enzymes pyruvate dehydrogenase, alpha-ketoglutarate dehydrogenase and isocitrate dehydrogenase. However, we have recently found that Ca2+ regulates the glutamate-dependent state 3 respiration by the supply of glutamate to mitochondria via aralar, a mitochondrial glutamate/aspartate carrier. Since this activation is not affected by ruthenium red, glutamate transport into mitochondria is controlled exclusively by extramitochondrial Ca2+. Therefore, this discovery shows that besides intramitochondrial also extramitochondrial Ca2+ regulates oxidative phosphorylation. This new mechanism acts as a mitochondrial "gas pedal", supplying the OXPHOS with substrate on demand. These results are in line with recent findings of Satrustegui and Palmieri showing that aralar as part of the malate-aspartate shuttle is involved in the Ca2+-dependent transport of reducing hydrogen equivalents (from NADH) into mitochondria. This review summarises results and evidence as well as hypothetical interpretations of data supporting the view that at the surface of mitochondria different regulatory Ca2+-binding sites exist and can contribute to cellular energy homeostasis. Moreover, on the basis of our own data, we propose that these surface Ca2+-binding sites may act as targets for neurotoxic proteins such as mutated huntingtin and others. The binding of these proteins to Ca2+-binding sites can impair the regulation by Ca2+, causing energetic depression and neurodegeneration. Copyright © 2010 Elsevier B.V. All rights reserved.

Hypoxic stress induces, but cannot sustain trophoblast stem cell differentiation to labyrinthine placenta due to mitochondrial insufficiency

PubMed Central

Xie, Yufen; Zhou, Sichang; Jiang, Zhongliang; Dai, Jing; Puscheck, Elizabeth E; Lee, Icksoo; Parker, Graham; Hüttemann, Maik; Rappolee, Daniel A

2014-01-01

Dysfunctional stem cell differentiation into placental lineages is associated with gestational diseases. Of the differentiated lineages available to trophoblast stem cells (TSC), elevated O2 and mitochondrial function are necessary to placental lineages at the maternal-placental surface and important in the etiology of preeclampsia. TSC lineage imbalance leads to embryonic failure during uterine implantation. Stress at implantation exacerbates stem cell depletion by decreasing proliferation and increasing differentiation. Implantation site O2 is normally ~2%. In culture, exposure to 2% O2 and fibroblast growth factor (FGF)4 enabled highest mouse TSC multipotency and proliferation. In contrast, hypoxic stress (0.5% O2) initiated the most TSC differentiation after 24 hr despite FGF4. However, hypoxic stress supported differentiation poorly after 4–7 days, despite FGF4 removal. At all tested O2 levels, FGF4 maintained Warburg metabolism; mitochondrial inactivity and aerobic glycolysis. However, hypoxic stress suppressed mitochondrial membrane potential, maintained low mitochondrial cytochrome c oxidase (oxidative phosphorylation/OxPhos), and high pyruvate kinase M2 (glycolysis) despite FGF4 removal. Inhibiting OxPhos inhibited differentiation at the differentiation optimum at 20% O2. Moreover, adding differentiation-inducing hyperosmolar stress failed to induce differentiation during hypoxia. Thus, differentiation depended on OxPhos at 20% O2; hypoxic and hyperosmolar stresses did not induce differentiation at 0.5% O2. Hypoxia-limited differentiation and mitochondrial inhibition and activation suggest that differentiation into two lineages of the labyrinthine placenta requires O2>0.5–2% and mitochondrial function. Stress-activated protein kinase increases an early lineage and suppresses later lineages in proportion to the deviation from optimal O2 for multipotency, thus it is the first enzyme reported to prioritize differentiation. PMID:25239494

Rosiglitazone Improves Stallion Sperm Motility, ATP Content, and Mitochondrial Function.

PubMed

Swegen, Aleona; Lambourne, Sarah Renay; Aitken, R John; Gibb, Zamira

2016-11-01

Media used for equine sperm storage often contain relatively high concentrations of glucose, even though stallion spermatozoa preferentially utilize oxidative phosphorylation (OXPHOS) over glycolysis to generate ATP and support motility. Rosiglitazone is an antidiabetic compound that enhances metabolic flexibility and glucose utilization in various cell types, but its effects on sperm metabolism are unknown. This study investigated the effects of rosiglitazone on stallion sperm function in vitro, along with the possible role of AMP-activated protein kinase (AMPK) in mediating these effects. Spermatozoa were incubated with or without rosiglitazone, GW9662 (an antagonist of peroxisome proliferator-activating receptor-gamma), and compound C (CC; an AMPK inhibitor). Sperm motility, viability, reactive oxygen species production, mitochondrial membrane potential (mMP), ATP content, and glucose uptake capacity were measured. Samples incubated with rosiglitazone displayed significantly higher motility, percentage of cells with normal mMP, ATP content, and glucose uptake capacity, while sperm viability was unaffected. The percentage of spermatozoa positive for mitochondrial ROS was also significantly lower in rosiglitazone-treated samples. AMPK localized to the sperm midpiece, and its phosphorylation, was increased in rosiglitazone-treated spermatozoa. CC decreased sperm AMPK phosphorylation and reduced sperm motility, and successfully inhibited the effects of rosiglitazone. Inclusion of rosiglitazone in a room temperature sperm storage medium maintained sperm motility above 60% for 6 days, attaining significantly higher motility than sperm stored in control media. The ability of rosiglitazone to substantially alleviate the time-dependent deterioration of stallion spermatozoa by diverting metabolism away from OXPHOS and toward glycolysis has novel implications for the long-term, functional preservation of these cells. © 2016 by the Society for the Study of Reproduction, Inc.

Defective mitochondrial rRNA methyltransferase MRM2 causes MELAS-like clinical syndrome

PubMed Central

Garone, Caterina; D’Souza, Aaron R; Dallabona, Cristina; Lodi, Tiziana; Rebelo-Guiomar, Pedro; Rorbach, Joanna; Donati, Maria Alice; Procopio, Elena; Montomoli, Martino; Guerrini, Renzo; Zeviani, Massimo; Calvo, Sarah E; Mootha, Vamsi K; DiMauro, Salvatore; Ferrero, Ileana; Minczuk, Michal

2017-01-01

Abstract Defects in nuclear-encoded proteins of the mitochondrial translation machinery cause early-onset and tissue-specific deficiency of one or more OXPHOS complexes. Here, we report a 7-year-old Italian boy with childhood-onset rapidly progressive encephalomyopathy and stroke-like episodes. Multiple OXPHOS defects and decreased mtDNA copy number (40%) were detected in muscle homogenate. Clinical features combined with low level of plasma citrulline were highly suggestive of mitochondrial encephalopathy, lactic acidosis and stroke-like episodes (MELAS) syndrome, however, the common m.3243 A > G mutation was excluded. Targeted exome sequencing of genes encoding the mitochondrial proteome identified a damaging mutation, c.567 G > A, affecting a highly conserved amino acid residue (p.Gly189Arg) of the MRM2 protein. MRM2 has never before been linked to a human disease and encodes an enzyme responsible for 2’-O-methyl modification at position U1369 in the human mitochondrial 16S rRNA. We generated a knockout yeast model for the orthologous gene that showed a defect in respiration and the reduction of the 2’-O-methyl modification at the equivalent position (U2791) in the yeast mitochondrial 21S rRNA. Complementation with the mrm2 allele carrying the equivalent yeast mutation failed to rescue the respiratory phenotype, which was instead completely rescued by expressing the wild-type allele. Our findings establish that defective MRM2 causes a MELAS-like phenotype, and suggests the genetic screening of the MRM2 gene in patients with a m.3243 A > G negative MELAS-like presentation. PMID:28973171

Defective mitochondrial rRNA methyltransferase MRM2 causes MELAS-like clinical syndrome.

PubMed

Garone, Caterina; D'Souza, Aaron R; Dallabona, Cristina; Lodi, Tiziana; Rebelo-Guiomar, Pedro; Rorbach, Joanna; Donati, Maria Alice; Procopio, Elena; Montomoli, Martino; Guerrini, Renzo; Zeviani, Massimo; Calvo, Sarah E; Mootha, Vamsi K; DiMauro, Salvatore; Ferrero, Ileana; Minczuk, Michal

2017-11-01

Defects in nuclear-encoded proteins of the mitochondrial translation machinery cause early-onset and tissue-specific deficiency of one or more OXPHOS complexes. Here, we report a 7-year-old Italian boy with childhood-onset rapidly progressive encephalomyopathy and stroke-like episodes. Multiple OXPHOS defects and decreased mtDNA copy number (40%) were detected in muscle homogenate. Clinical features combined with low level of plasma citrulline were highly suggestive of mitochondrial encephalopathy, lactic acidosis and stroke-like episodes (MELAS) syndrome, however, the common m.3243 A > G mutation was excluded. Targeted exome sequencing of genes encoding the mitochondrial proteome identified a damaging mutation, c.567 G > A, affecting a highly conserved amino acid residue (p.Gly189Arg) of the MRM2 protein. MRM2 has never before been linked to a human disease and encodes an enzyme responsible for 2'-O-methyl modification at position U1369 in the human mitochondrial 16S rRNA. We generated a knockout yeast model for the orthologous gene that showed a defect in respiration and the reduction of the 2'-O-methyl modification at the equivalent position (U2791) in the yeast mitochondrial 21S rRNA. Complementation with the mrm2 allele carrying the equivalent yeast mutation failed to rescue the respiratory phenotype, which was instead completely rescued by expressing the wild-type allele. Our findings establish that defective MRM2 causes a MELAS-like phenotype, and suggests the genetic screening of the MRM2 gene in patients with a m.3243 A > G negative MELAS-like presentation. © The Author 2017. Published by Oxford University Press.

The pathomechanism of cytochrome c oxidase deficiency includes nuclear DNA damage.

PubMed

Douiev, Liza; Saada, Ann

2018-06-07

Mitochondrial cytochrome c oxidase (COX, respiratory chain complex IV), contributes to ATP production via oxidative phosphorylation (OXPHOS). Clinical presentation of COX deficiency is heterogeneous ranging from mild to severe neuromuscular diseases. Anemia is among the symptoms and we have previously reported Fanconi anemia like features in COX4-1 deficiency, suggesting genomic instability and our preliminary results detected nuclear double stranded DNA breaks (DSB). We now quantified the DSB by phospho histone H2AX Ser139 staining of COX4-1 and COX6B1 deficient fibroblasts (225% and 215% of normal, respectively) and confirmed their occurrence by neutral comet assay. We further explored the mechanism of DNA damage by studying normal fibroblasts treated with micromolar concentrations of cyanide (KCN). Present results demonstrate elevated nuclear DSB in cells treated with 50 μM KCN for 24 h (170% of normal) in high-glucose medium conditions where ROS and ATP remain normal, although Glutathione content was partially decreased. In glucose-free and serum-free medium, where growth is hampered, DSB were not elevated. Additionally we demonstrate the benefit of nicotinamide riboside (NR) which ameliorated DSB in COX4-1, COX6B1 and KCN treated cells (130%, 154% and 87% of normal cells, respectively). Conversely a negative effect of a poly[ADP-ribose] polymerase (PARP) inhibitor was found. Although additional investigation is needed, our findings raise the possibility that the pathomechanism of COX deficiency and possibly also in other OXPHOS defects, include nuclear DNA damage resulting from nicotinamide adenine dinucleotide (NAD + ) deficit combined with a replicative state, rather than oxidative stress and energy depletion. Copyright © 2018 Elsevier B.V. All rights reserved.

Adaptive Patterns of Mitogenome Evolution Are Associated with the Loss of Shell Scutes in Turtles.

PubMed

Escalona, Tibisay; Weadick, Cameron J; Antunes, Agostinho

2017-10-01

The mitochondrial genome encodes several protein components of the oxidative phosphorylation (OXPHOS) pathway and is critical for aerobic respiration. These proteins have evolved adaptively in many taxa, but linking molecular-level patterns with higher-level attributes (e.g., morphology, physiology) remains a challenge. Turtles are a promising system for exploring mitochondrial genome evolution as different species face distinct respiratory challenges and employ multiple strategies for ensuring efficient respiration. One prominent adaptation to a highly aquatic lifestyle in turtles is the secondary loss of keratenized shell scutes (i.e., soft-shells), which is associated with enhanced swimming ability and, in some species, cutaneous respiration. We used codon models to examine patterns of selection on mitochondrial protein-coding genes along the three turtle lineages that independently evolved soft-shells. We found strong evidence for positive selection along the branches leading to the pig-nosed turtle (Carettochelys insculpta) and the softshells clade (Trionychidae), but only weak evidence for the leatherback (Dermochelys coriacea) branch. Positively selected sites were found to be particularly prevalent in OXPHOS Complex I proteins, especially subunit ND2, along both positively selected lineages, consistent with convergent adaptive evolution. Structural analysis showed that many of the identified sites are within key regions or near residues involved in proton transport, indicating that positive selection may have precipitated substantial changes in mitochondrial function. Overall, our study provides evidence that physiological challenges associated with adaptation to a highly aquatic lifestyle have shaped the evolution of the turtle mitochondrial genome in a lineage-specific manner. © The Author 2017. Published by Oxford University Press on behalf of the Society for Molecular Biology and Evolution. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.

Evaluating the Efficacy of GLUT Inhibitors Using a Seahorse Extracellular Flux Analyzer.

PubMed

Wei, Changyong; Heitmeier, Monique; Hruz, Paul W; Shanmugam, Mala

2018-01-01

Glucose is metabolized through anaerobic glycolysis and aerobic oxidative phosphorylation (OXPHOS). Perturbing glucose uptake and its subsequent metabolism can alter both glycolytic and OXPHOS pathways and consequently lactate and/or oxygen consumption. Production and secretion of lactate, as a consequence of glycolysis, leads to acidification of the extracellular medium. Molecular oxygen is the final electron acceptor in the electron transport chain, facilitating oxidative phosphorylation of ADP to ATP. The alterations in extracellular acidification and/or oxygen consumption can thus be used as indirect readouts of glucose metabolism and assessing the impact of inhibiting glucose transport through specific glucose transporters (GLUTs). The Seahorse bioenergetics analyzer can measure both the oxygen consumption rate (OCR) and extracellular acidification rate (ECAR). The proposed methodology affords a robust, high-throughput method to screen for GLUT inhibition in cells engineered to express specific GLUTs, providing live cell read-outs upon GLUT inhibition.

Optical imaging of metabolic adaptability in metastatic and non-metastatic breast cancer

NASA Astrophysics Data System (ADS)

Rebello, Lisa; Rajaram, Narasimhan

2018-02-01

Accurate methods for determining metastatic risk from the primary tumor are crucial for patient survival. Cell metabolism could potentially be used as a marker of metastatic risk. Optical imaging of the endogenous fluorescent molecules nicotinamide adenine dinucleotide (NADH) and flavin adenine dinucleotide (FAD) provides a non-destructive and label-free method for determining cell metabolism. The optical redox ratio (FAD/FAD+NADH) is sensitive to the balance between glycolysis and oxidative phosphorylation (OXPHOS). We have previously established that hypoxia-reoxygenation stress leads to metastatic potential-dependent changes in optical redox ratio. The objective of this study was to monitor the changes in optical redox ratio in breast cancer cells in response to different periods of hypoxic stress as well various levels of hypoxia to establish an optimal protocol. We measured the optical redox ratio of highly metastatic 4T1 murine breast cancer cells under normoxic conditions and after exposure to 30, 60, and 120 minutes of 0.5% O2. This was followed by an hour of reoxygenation. We found an increase in the optical redox ratio following reoxygenation from hypoxia for all durations. Statistically significant differences were observed at 60 and 120 minutes (p˂0.01) compared with normoxia, implying an ability to adapt to OXPHOS after reoxygenation. The switch to OXPHOS has been shown to be a key promoter of cell invasion. We will present our results from these investigations in human breast cancer cells as well as non-metastatic breast cancer cells exposed to various levels of hypoxia.

Novel insights into the functional metabolic impact of an apparent de novo m.8993T>G variant in the MT-ATP6 gene associated with maternally inherited form of Leigh Syndrome.

PubMed

Uittenbogaard, Martine; Brantner, Christine A; Fang, ZiShui; Wong, Lee-Jun C; Gropman, Andrea; Chiaramello, Anne

2018-03-27

In this study, we report a novel perpective of metabolic consequences for the m.8993T>G variant using fibroblasts from a proband with clinical symptoms compatible with Maternally Inherited Leigh Syndrome (MILS). Definitive diagnosis was corroborated by mitochondrial DNA testing for the pathogenic variant m.8993T>G in MT-ATP6 subunit by Sanger sequencing. The long-range PCR followed by massively parallel sequencing method detected the near homoplasmic m.8993T>G variant at 83% in the proband's fibroblasts and at 0.4% in the mother's fibroblasts. Our results are compatible with very low levels of germline heteroplasmy or an apparent de novo mutation. Our mitochondrial morphometric analysis reveals severe defects in mitochondrial cristae structure in the proband's fibroblasts. Our live-cell mitochondrial respiratory analyses show impaired oxidative phosphorylation with decreased spare respiratory capacity in response to energy stress in the proband's fibroblasts. We detected a diminished glycolysis with a lessened glycolytic capacity and reserve, revealing a stunted ability to switch to glycolysis upon full inhibition of OXPHOS activities. This dysregulated energy reprogramming results in a defective interplay between OXPHOS and glycolysis during an energy crisis. Our study sheds light on the potential pathophysiologic mechanism leading to chronic energy crisis in this MILS patient harboring the m.8993T>G variant. Copyright © 2018 Elsevier Inc. All rights reserved.

Hypoxic HepG2 cell adaptation decreases ATP synthase dimers and ATP production in inflated cristae by mitofilin down-regulation concomitant to MICOS clustering.

PubMed

Plecitá-Hlavatá, Lydie; Engstová, Hana; Alán, Lukáš; Špaček, Tomáš; Dlasková, Andrea; Smolková, Katarína; Špačková, Jitka; Tauber, Jan; Strádalová, Vendula; Malínský, Jan; Lessard, Mark; Bewersdorf, Joerg; Ježek, Petr

2016-05-01

The relationship of the inner mitochondrial membrane (IMM) cristae structure and intracristal space (ICS) to oxidative phosphorylation (oxphos) is not well understood. Mitofilin (subunit Mic60) of the mitochondrial contact site and cristae organizing system (MICOS) IMM complex is attached to the outer membrane (OMM) via the sorting and assembly machinery/topogenesis of mitochondrial outer membrane β-barrel proteins (SAM/TOB) complex and controls the shape of the cristae. ATP synthase dimers determine sharp cristae edges, whereas trimeric OPA1 tightens ICS outlets. Metabolism is altered during hypoxia, and we therefore studied cristae morphology in HepG2 cells adapted to 5% oxygen for 72 h. Three dimensional (3D), super-resolution biplane fluorescence photoactivation localization microscopy with Eos-conjugated, ICS-located lactamase-β indicated hypoxic ICS expansion with an unchanged OMM (visualized by Eos-mitochondrial fission protein-1). 3D direct stochastic optical reconstruction microscopy immunocytochemistry revealed foci of clustered mitofilin (but not MICOS subunit Mic19) in contrast to its even normoxic distribution. Mitofilin mRNA and protein decreased by ∼20%. ATP synthase dimers vs monomers and state-3/state-4 respiration ratios were lower during hypoxia. Electron microscopy confirmed ICS expansion (maximum in glycolytic cells), which was absent in reduced or OMM-detached cristae of OPA1- and mitofilin-silenced cells, respectively. Hypoxic adaptation is reported as rounding sharp cristae edges and expanding cristae width (ICS) by partial mitofilin/Mic60 down-regulation. Mitofilin-depleted MICOS detaches from SAM while remaining MICOS with mitofilin redistributes toward higher interdistances. This phenomenon causes partial oxphos dormancy in glycolytic cells via disruption of ATP synthase dimers.-Plecitá-Hlavatá, L., Engstová, H., Alán, L., Špaček, T., Dlasková, A., Smolková, K., Špačková, J., Tauber, J., Strádalová, V., Malínský, J., Lessard, M., Bewersdorf, J., Ježek, P. Hypoxic HepG2 cell adaptation decreases ATP synthase dimers and ATP production in inflated cristae by mitofilin down-regulation concomitant to MICOS clustering. © FASEB.

DOE Office of Scientific and Technical Information (OSTI.GOV)

Xu, Ting; Zhao, Jing; Hu, Ping

Pentachlorophenol (PCP) is a prevalent pollutant in the environment and has been demonstrated to be a serious toxicant to humans and animals. However, little is known regarding the molecular mechanism underlying its toxic effects on vertebrate early development. To explore the impacts and underlying mechanisms of PCP on early development, zebrafish (Danio rerio) embryos were exposed to PCP at concentrations of 0, 20 and 50 μg/L, and microscopic observation and cDNA microarray analysis were subsequently conducted at gastrulation stage. The morphological observations revealed that PCP caused a developmental delay of zebrafish embryos in a concentration-dependent manner. Transcriptomic data showed thatmore » 50 μg/L PCP treatment resulted in significant changes in gene expression level, and the genes involved in energy metabolism and cell behavior were identified based on gene functional enrichment analysis. The energy production of embryos was influenced by PCP via the activation of glycolysis along with the inhibition of oxidative phosphorylation (OXPHOS). The results suggested that PCP acts as an inhibitor of OXPHOS at 8 hpf (hours postfertilization). Consistent with the activated glycolysis, the cell cycle activity of PCP-treated embryos was higher than the controls. These characteristics are similar to the Warburg effect, which occurs in human tumors. The microinjection of exogenous ATP confirmed that an additional energy supply could rescue PCP-treated embryos from the developmental delay due to the energy deficit. Taken together, our results demonstrated that PCP causes a Warburg-like effect on zebrafish embryos during gastrulation, and the affected embryos had the phenotype of developmental delay. - Highlights: • We treat zebrafish embryos with PCP at gastrula stage. • PCP acts as an oxidative phosphorylation inhibitor, not an uncoupler, in gastrulation. • Exogenous ATP injection will rescue the development of effected embryos. • The transcriptome of PCP-treated embryo exhibits a Warburg-like effect in tumor cell.« less

Genetic disruption of lactate/H+ symporters (MCTs) and their subunit CD147/BASIGIN sensitizes glycolytic tumor cells to phenformin.

PubMed

Marchiq, Ibtissam; Le Floch, Renaud; Roux, Danièle; Simon, Marie-Pierre; Pouyssegur, Jacques

2015-01-01

Rapidly growing glycolytic tumors require energy and intracellular pH (pHi) homeostasis through the activity of two major monocarboxylate transporters, MCT1 and the hypoxia-inducible MCT4, in intimate association with the glycoprotein CD147/BASIGIN (BSG). To further explore and validate the blockade of lactic acid export as an anticancer strategy, we disrupted, via zinc finger nucleases, MCT4 and BASIGIN genes in colon adenocarcinoma (LS174T) and glioblastoma (U87) human cell lines. First, we showed that homozygous loss of MCT4 dramatically sensitized cells to the MCT1 inhibitor AZD3965. Second, we demonstrated that knockout of BSG leads to a decrease in lactate transport activity of MCT1 and MCT4 by 10- and 6-fold, respectively. Consequently, cells accumulated an intracellular pool of lactic and pyruvic acids, magnified by the MCT1 inhibitor decreasing further pHi and glycolysis. As a result, we found that these glycolytic/MCT-deficient cells resumed growth by redirecting their metabolism toward OXPHOS. Third, we showed that in contrast with parental cells, BSG-null cells became highly sensitive to phenformin, an inhibitor of mitochondrial complex I. Phenformin addition to these MCT-disrupted cells in normoxic and hypoxic conditions induced a rapid drop in cellular ATP-inducing cell death by "metabolic catastrophe." Finally, xenograft analysis confirmed the deleterious tumor growth effect of MCT1/MCT4 ablation, an action enhanced by phenformin treatment. Collectively, these findings highlight that inhibition of the MCT/BSG complexes alone or in combination with phenformin provides an acute anticancer strategy to target highly glycolytic tumors. This genetic approach validates the anticancer potential of the MCT1 and MCT4 inhibitors in current development. ©2014 American Association for Cancer Research.

Extracellular pH Modulates Neuroendocrine Prostate Cancer Cell Metabolism and Susceptibility to the Mitochondrial Inhibitor Niclosamide

PubMed Central

Ippolito, Joseph E.; Brandenburg, Matthew W.; Ge, Xia; Crowley, Jan R.; Kirmess, Kristopher M.; Som, Avik; D’Avignon, D. Andre; Arbeit, Jeffrey M.; Achilefu, Samuel; Yarasheski, Kevin E.; Milbrandt, Jeffrey

2016-01-01

Neuroendocrine prostate cancer is a lethal variant of prostate cancer that is associated with castrate-resistant growth, metastasis, and mortality. The tumor environment of neuroendocrine prostate cancer is heterogeneous and characterized by hypoxia, necrosis, and numerous mitoses. Although acidic extracellular pH has been implicated in aggressive cancer features including metastasis and therapeutic resistance, its role in neuroendocrine prostate cancer physiology and metabolism has not yet been explored. We used the well-characterized PNEC cell line as a model to establish the effects of extracellular pH (pH 6.5, 7.4, and 8.5) on neuroendocrine prostate cancer cell metabolism. We discovered that alkalinization of extracellular pH converted cellular metabolism to a nutrient consumption-dependent state that was susceptible to glucose deprivation, glutamine deprivation, and 2-deoxyglucose (2-DG) mediated inhibition of glycolysis. Conversely, acidic pH shifted cellular metabolism toward an oxidative phosphorylation (OXPHOS)-dependent state that was susceptible to OXPHOS inhibition. Based upon this mechanistic knowledge of pH-dependent metabolism, we identified that the FDA-approved anti-helminthic niclosamide depolarized mitochondrial potential and depleted ATP levels in PNEC cells whose effects were enhanced in acidic pH. To further establish relevance of these findings, we tested the effects of extracellular pH on susceptibility to nutrient deprivation and OXPHOS inhibition in a cohort of castrate-resistant prostate cancer cell lines C4-2B, PC-3, and PC-3M. We discovered similar pH-dependent toxicity profiles among all cell lines with these treatments. These findings underscore a potential importance to acidic extracellular pH in the modulation of cell metabolism in tumors and development of an emerging paradigm that exploits the synergy of environment and therapeutic efficacy in cancer. PMID:27438712

Cryopreservation of lipid bilayers by LEA proteins from Artemia franciscana and trehalose.

PubMed

Moore, Daniel S; Hand, Steven C

2016-10-01

The capacity of Late Embryogenesis Abundant (LEA) proteins and trehalose to protect liposomes against freezing-induced damage was examined by measuring the leakage of 5(6)-carboxyfluorescein (CF). Liposomes were prepared to simulate the lipid compositions of the inner leaflet of the plasma membrane, outer mitochondrial membrane (OMM), and inner mitochondrial membrane (IMM). Two recombinant LEA proteins belonging to Group 3 (AfrLEA2 and AfrLEA3m) were expressed and purified from embryos of Artemia franciscana. Only OMM-like liposomes were significantly protected by AfrLEA2 and AfrLEA3m against freeze-thaw damage; at the highest protein:lipid mass ratio tested, leakage of CF was 56.3% of control with AfrLEA3m and 29.3% with AfrLEA2. By comparison, trehalose provided protection to all compositional types. The greatest stabilization during freezing occurred when trehalose was present on both sides of the bilayer. When mitochondria isolated from rat liver were freeze-thawed in trehalose solution, the OMM remained intact based on the absence of increased oxygen consumption when cytochrome c was added during oxidative phosphorylation (OXPHOS). Respiratory control ratios (OXPHOS/LEAK) were depressed by only 30% after freeze-thawing in trehalose compared to non-frozen controls, which indicated some retention of OXPHOS capacity by the IMM. Trehalose then was loaded into the matrix (0.24 μmol/mg mitochondrial protein) by transient opening of the permeability transition pore, a procedure optimized for retention of OMM integrity. Surprisingly, respiratory control ratios were not improved after freeze-thawing with external plus matrix trehalose, when compared to external trehalose alone. This result could perhaps be explained by insufficient accumulation of matrix trehalose. Copyright © 2016 Elsevier Inc. All rights reserved.

Enterococcus faecalis infection causes inflammation, intracellular oxphos-independent ROS production, and DNA damage in human gastric cancer cells.

PubMed

Strickertsson, Jesper A B; Desler, Claus; Martin-Bertelsen, Tomas; Machado, Ana Manuel Dantas; Wadstrøm, Torkel; Winther, Ole; Rasmussen, Lene Juel; Friis-Hansen, Lennart

2013-01-01

Achlorhydria caused by e.g. atrophic gastritis allows for bacterial overgrowth, which induces chronic inflammation and damage to the mucosal cells of infected individuals driving gastric malignancies and cancer. Enterococcus faecalis (E. faecalis) can colonize achlohydric stomachs and we therefore wanted to study the impact of E. faecalis infection on inflammatory response, reactive oxygen species (ROS) formation, mitochondrial respiration, and mitochondrial genetic stability in gastric mucosal cells. To separate the changes induced by bacteria from those of the inflammatory cells we established an in vitro E. faecalis infection model system using the gastric carcinoma cell line MKN74. Total ROS and superoxide was measured by fluorescence microscopy. Cellular oxygen consumption was characterized non-invasively using XF24 microplate based respirometry. Gene expression was examined by microarray, and response pathways were identified by Gene Set Analysis (GSA). Selected gene transcripts were verified by quantitative real-time polymerase chain reaction (qRT-PCR). Mitochondrial mutations were determined by sequencing. Infection of MKN74 cells with E. faecalis induced intracellular ROS production through a pathway independent of oxidative phosphorylation (oxphos). Furthermore, E. faecalis infection induced mitochondrial DNA instability. Following infection, genes coding for inflammatory response proteins were transcriptionally up-regulated while DNA damage repair and cell cycle control genes were down-regulated. Cell growth slowed down when infected with viable E. faecalis and responded in a dose dependent manner to E. faecalis lysate. Infection by E. faecalis induced an oxphos-independent intracellular ROS response and damaged the mitochondrial genome in gastric cell culture. Finally the bacteria induced an NF-κB inflammatory response as well as impaired DNA damage response and cell cycle control gene expression. Array Express accession number E-MEXP-3496.

Inhibition of Mitochondrial Complex I Leads to Decreased Motility and Membrane Integrity Related to Increased Hydrogen Peroxide and Reduced ATP Production, while the Inhibition of Glycolysis Has Less Impact on Sperm Motility

PubMed Central

Plaza Davila, María; Martin Muñoz, Patricia; Tapia, Jose A.; Ortega Ferrusola, Cristina; Balao da Silva C, Carolina; Peña, Fernando J.

2015-01-01

Mitochondria have been proposed as the major source of reactive oxygen species in somatic cells and human spermatozoa. However, no data regarding the role of mitochondrial ROS production in stallion spermatozoa are available. To shed light on the role of the mitochondrial electron transport chain in the origin of oxidative stress in stallion spermatozoa, specific inhibitors of complex I (rotenone) and III (antimycin-A) were used. Ejaculates from seven Andalusian stallions were collected and incubated in BWW media at 37°C in the presence of rotenone, antimycin-A or control vehicle. Incubation in the presence of these inhibitors reduced sperm motility and velocity (CASA analysis) (p<0.01), but the effect was more evident in the presence of rotenone (a complex I inhibitor). These inhibitors also decreased ATP content. The inhibition of complexes I and III decreased the production of reactive oxygen species (p<0.01) as assessed by flow cytometry after staining with CellRox deep red. This observation suggests that the CellRox probe mainly identifies superoxide and that superoxide production may reflect intense mitochondrial activity rather than oxidative stress. The inhibition of complex I resulted in increased hydrogen peroxide production (p<0.01). The inhibition of glycolysis resulted in reduced sperm velocities (p<0.01) without an effect on the percentage of total motile sperm. Weak and moderate (but statistically significant) positive correlations were observed between sperm motility, velocity and membrane integrity and the production of reactive oxygen species. These results indicate that stallion sperm rely heavily on oxidative phosphorylation (OXPHOS) for the production of ATP for motility but also require glycolysis to maintain high velocities. These data also indicate that increased hydrogen peroxide originating in the mitochondria is a mechanism involved in stallion sperm senescence. PMID:26407142

Inhibition of Mitochondrial Complex I Leads to Decreased Motility and Membrane Integrity Related to Increased Hydrogen Peroxide and Reduced ATP Production, while the Inhibition of Glycolysis Has Less Impact on Sperm Motility.

PubMed

Plaza Davila, María; Martin Muñoz, Patricia; Tapia, Jose A; Ortega Ferrusola, Cristina; Balao da Silva C, Carolina; Peña, Fernando J

2015-01-01

Mitochondria have been proposed as the major source of reactive oxygen species in somatic cells and human spermatozoa. However, no data regarding the role of mitochondrial ROS production in stallion spermatozoa are available. To shed light on the role of the mitochondrial electron transport chain in the origin of oxidative stress in stallion spermatozoa, specific inhibitors of complex I (rotenone) and III (antimycin-A) were used. Ejaculates from seven Andalusian stallions were collected and incubated in BWW media at 37 °C in the presence of rotenone, antimycin-A or control vehicle. Incubation in the presence of these inhibitors reduced sperm motility and velocity (CASA analysis) (p<0.01), but the effect was more evident in the presence of rotenone (a complex I inhibitor). These inhibitors also decreased ATP content. The inhibition of complexes I and III decreased the production of reactive oxygen species (p<0.01) as assessed by flow cytometry after staining with CellRox deep red. This observation suggests that the CellRox probe mainly identifies superoxide and that superoxide production may reflect intense mitochondrial activity rather than oxidative stress. The inhibition of complex I resulted in increased hydrogen peroxide production (p<0.01). The inhibition of glycolysis resulted in reduced sperm velocities (p<0.01) without an effect on the percentage of total motile sperm. Weak and moderate (but statistically significant) positive correlations were observed between sperm motility, velocity and membrane integrity and the production of reactive oxygen species. These results indicate that stallion sperm rely heavily on oxidative phosphorylation (OXPHOS) for the production of ATP for motility but also require glycolysis to maintain high velocities. These data also indicate that increased hydrogen peroxide originating in the mitochondria is a mechanism involved in stallion sperm senescence.

Aim-less translation: loss of Saccharomyces cerevisiae mitochondrial translation initiation factor mIF3/Aim23 leads to unbalanced protein synthesis.

PubMed

Kuzmenko, Anton; Derbikova, Ksenia; Salvatori, Roger; Tankov, Stoyan; Atkinson, Gemma C; Tenson, Tanel; Ott, Martin; Kamenski, Piotr; Hauryliuk, Vasili

2016-01-05

The mitochondrial genome almost exclusively encodes a handful of transmembrane constituents of the oxidative phosphorylation (OXPHOS) system. Coordinated expression of these genes ensures the correct stoichiometry of the system's components. Translation initiation in mitochondria is assisted by two general initiation factors mIF2 and mIF3, orthologues of which in bacteria are indispensible for protein synthesis and viability. mIF3 was thought to be absent in Saccharomyces cerevisiae until we recently identified mitochondrial protein Aim23 as the missing orthologue. Here we show that, surprisingly, loss of mIF3/Aim23 in S. cerevisiae does not indiscriminately abrogate mitochondrial translation but rather causes an imbalance in protein production: the rate of synthesis of the Atp9 subunit of F1F0 ATP synthase (complex V) is increased, while expression of Cox1, Cox2 and Cox3 subunits of cytochrome c oxidase (complex IV) is repressed. Our results provide one more example of deviation of mitochondrial translation from its bacterial origins.

The Chemical Interplay between Nitric Oxide and Mitochondrial Cytochrome c Oxidase: Reactions, Effectors and Pathophysiology

PubMed Central

Sarti, Paolo; Forte, Elena; Giuffrè, Alessandro; Mastronicola, Daniela; Magnifico, Maria Chiara; Arese, Marzia

2012-01-01

Nitric oxide (NO) reacts with Complex I and cytochrome c oxidase (CcOX, Complex IV), inducing detrimental or cytoprotective effects. Two alternative reaction pathways (PWs) have been described whereby NO reacts with CcOX, producing either a relatively labile nitrite-bound derivative (CcOX-NO2  −, PW1) or a more stable nitrosyl-derivative (CcOX-NO, PW2). The two derivatives are both inhibited, displaying different persistency and O2 competitiveness. In the mitochondrion, during turnover with O2, one pathway prevails over the other one depending on NO, cytochrome c 2+ and O2 concentration. High cytochrome c 2+, and low O2 proved to be crucial in favoring CcOX nitrosylation, whereas under-standard cell-culture conditions formation of the nitrite derivative prevails. All together, these findings suggest that NO can modulate physiologically the mitochondrial respiratory/OXPHOS efficiency, eventually being converted to nitrite by CcOX, without cell detrimental effects. It is worthy to point out that nitrite, far from being a simple oxidation byproduct, represents a source of NO particularly important in view of the NO cell homeostasis, the NO production depends on the NO synthases whose activity is controlled by different stimuli/effectors; relevant to its bioavailability, NO is also produced by recycling cell/body nitrite. Bioenergetic parameters, such as mitochondrial ΔΨ, lactate, and ATP production, have been assayed in several cell lines, in the presence of endogenous or exogenous NO and the evidence collected suggests a crucial interplay between CcOX and NO with important energetic implications. PMID:22811713

Reciprocal transcriptional regulation of metabolic and signaling pathways correlates with disease severity in heart failure.

PubMed

Barth, Andreas S; Kumordzie, Ami; Frangakis, Constantine; Margulies, Kenneth B; Cappola, Thomas P; Tomaselli, Gordon F

2011-10-01

Systolic heart failure (HF) is a complex systemic syndrome that can result from a wide variety of clinical conditions and gene mutations. Despite phenotypic similarities, characterized by ventricular dilatation and reduced contractility, the extent of common and divergent gene expression between different forms of HF remains a matter of intense debate. Using a meta-analysis of 28 experimental (mouse, rat, dog) and human HF microarray studies, we demonstrate that gene expression changes are characterized by a coordinated and reciprocal regulation of major metabolic and signaling pathways. In response to a wide variety of stressors in animal models of HF, including ischemia, pressure overload, tachypacing, chronic isoproterenol infusion, Chagas disease, and transgenic mouse models, major metabolic pathways are invariably downregulated, whereas cell signaling pathways are upregulated. In contrast to this uniform transcriptional pattern that recapitulates a fetal gene expression program in experimental animal models of HF, human HF microarray studies displayed a greater heterogeneity, with some studies even showing upregulation of metabolic and downregulation of signaling pathways in end-stage human hearts. These discrepant results between animal and human studies are due to a number of factors, prominently cardiac disease and variable exposure to cold cardioplegic solution in nonfailing human samples, which can downregulate transcripts involved in oxidative phosphorylation (OXPHOS), thus mimicking gene expression patterns observed in failing samples. Additionally, β-blockers and ACE inhibitor use in end-stage human HF was associated with higher levels of myocardial OXPHOS transcripts, thus partially reversing the fetal gene expression pattern. In human failing samples, downregulation of metabolism was associated with hemodynamic markers of disease severity. Irrespective of the etiology, gene expression in failing myocardium is characterized by downregulation of metabolic transcripts and concomitant upregulation of cell signaling pathways. Gene expression changes along this metabolic-signaling axis in mammalian myocardium are a consistent feature in the heterogeneous transcriptional response observed in phenotypically similar models of HF.

Analysis of energy metabolism of HeLa cancer cells in vitro and in vivo using fluorescence lifetime microscopy

NASA Astrophysics Data System (ADS)

Lukina, Maria; Shirmanova, Marina; Dudenkova, Varvara; Druzhkova, Irina; Shumilova, Anastasia; Zagaynova, Elena

2016-04-01

The aim of the present work was to study energy metabolism in human cervical carcinoma (HeLa) cells in vitro and in vivo using two-photon FLIM. Cellular metabolism was examined by monitoring of the fluorescence lifetimes of free and protein-bound forms of NAD(P)H and FAD and their relative contributions. Two-photon fluorescence and second harmonic generation microscopy as well as standard histopathology with hematoxylin and eosin were used to characterize tissue structure. Cellular metabolism was analyzed in cancer cells co-cultured with human fibroblasts and in tumor xenografts transplanted to nude mice. In the HeLa-huFB co-culture we observed a metabolic shift from OXPHOS toward glycolysis in cancer cells, and from glycolysis to OXPHOS in fibroblasts, starting from Day 2 of co-culturing. In the tumor tissue we detected metabolic heterogeneity with more glycolytic metabolism of cancer cells in the stroma-rich zones. The results of the study are of a great importance for understanding metabolic behavior of tumors and for development of anticancer drugs targeted to metabolic pathways.

Mutations of the Mitochondrial Holocytochrome c–Type Synthase in X-Linked Dominant Microphthalmia with Linear Skin Defects Syndrome

PubMed Central

Wimplinger, Isabella; Morleo, Manuela; Rosenberger, Georg; Iaconis, Daniela; Orth, Ulrike; Meinecke, Peter; Lerer, Israela; Ballabio, Andrea; Gal, Andreas; Franco, Brunella; Kutsche, Kerstin

2006-01-01

The microphthalmia with linear skin defects syndrome (MLS, or MIDAS) is an X-linked dominant male-lethal disorder almost invariably associated with segmental monosomy of the Xp22 region. In two female patients, from two families, with MLS and a normal karyotype, we identified heterozygous de novo point mutations—a missense mutation (p.R217C) and a nonsense mutation (p.R197X)—in the HCCS gene. HCCS encodes the mitochondrial holocytochrome c–type synthase that functions as heme lyase by covalently adding the prosthetic heme group to both apocytochrome c and c1. We investigated a third family, displaying phenotypic variability, in which the mother and two of her daughters carry an 8.6-kb submicroscopic deletion encompassing part of the HCCS gene. Functional analysis demonstrates that both mutant proteins (R217C and Δ197–268) were unable to complement a Saccharomyces cerevisiae mutant deficient for the HCCS orthologue Cyc3p, in contrast to wild-type HCCS. Moreover, ectopically expressed HCCS wild-type and the R217C mutant protein are targeted to mitochondria in CHO-K1 cells, whereas the C-terminal–truncated Δ197–268 mutant failed to be sorted to mitochondria. Cytochrome c, the final product of holocytochrome c–type synthase activity, is implicated in both oxidative phosphorylation (OXPHOS) and apoptosis. We hypothesize that the inability of HCCS-deficient cells to undergo cytochrome c–mediated apoptosis may push cell death toward necrosis that gives rise to severe deterioration of the affected tissues. In summary, we suggest that disturbance of both OXPHOS and the balance between apoptosis and necrosis, as well as the X-inactivation pattern, may contribute to the variable phenotype observed in patients with MLS. PMID:17033964

Energy metabolism in neuroblastoma and Wilms tumor

PubMed Central

Aminzadeh, Sepideh; Vidali, Silvia; Sperl, Wolfgang; Feichtinger, René G.

2015-01-01

To support high proliferation, the majority of cancer cells undergo fundamental metabolic changes such as increasing their glucose uptake and shifting to glycolysis for ATP production at the expense of far more efficient mitochondrial energy production by oxidative phosphorylation (OXPHOS), which at first glance is a paradox. This phenomenon is known as the Warburg effect. However, enhanced glycolysis is necessary to provide building blocks for anabolic growth. Apart from the generation of ATP, intermediates of glycolysis serve as precursors for a variety of biosynthetic pathways essential for cell proliferation. In the last 10-15 years the field of tumor metabolism has experienced an enormous boom in interest. It is now well established that tumor suppressor genes and oncogenes often play a central role in the regulation of cellular metabolism. Therefore, they significantly contribute to the manifestation of the Warburg effect. While much attention has focused on adult solid tumors, so far there has been comparatively little effort directed at elucidation of the mechanism responsible for the Warburg effect in childhood cancers. In this review we focus on metabolic pathways in neuroblastoma (NB) and Wilms tumor (WT), the two most frequent solid tumors in children. Both tumor types show alterations of the OXPHOS system and glycolytic features. Chromosomal alterations and activation of oncogenes like MYC or inactivation of tumor suppressor genes like TP53 can in part explain the changes of energy metabolism in these cancers. The strict dependence of cancer cells on glucose metabolism is a fairly common feature among otherwise biologically diverse types of cancer. Therefore, inhibition of glycolysis or starvation of cancer cells through glucose deprivation via a high-fat low-carbohydrate diet may be a promising avenue for future adjuvant therapeutic strategies. PMID:26835356

Ischemic Preconditioning Protects Astrocytes against Oxygen Glucose Deprivation Via the Nuclear Erythroid 2-Related Factor 2 Pathway.

PubMed

Narayanan, Srinivasan V; Dave, Kunjan R; Perez-Pinzon, Miguel A

2018-04-01

Induction of ischemic preconditioning (IPC) represents a potential therapy against cerebral ischemia by activation of adaptive pathways and modulation of mitochondria to induce ischemic tolerance to various cells and tissues. Mitochondrial dysfunction has been ascribed to contribute to numerous neurodegenerative conditions and cerebral ischemia. Nuclear erythroid 2-related factor 2 (Nrf2) is a transcription factor that has traditionally been involved in upregulating cellular antioxidant systems to combat oxidative stress in the brain; however, the association of Nrf2 with mitochondria in the brain remains unclear. In the present study, we investigated the effects of Nrf2 on (i) IPC-induced protection of astrocytes; (ii) OXPHOS protein expression; and (iii) mitochondrial supercomplex formation.Oxygen-glucose deprivation (OGD) was used as an in vitro model of cerebral ischemia and IPC in cultured rodent astrocytes derived from WT C57Bl/6J and Nrf2 -/- mice. OXPHOS proteins were probed via western blotting, and supercomplexes were determined by blue native gel electrophoresis.IPC-induced cytoprotection in wild-type, but not Nrf2 -/- mouse astrocyte cultures following a lethal duration of OGD. In addition, our results suggest that Nrf2 localizes to the outer membrane in non-synaptic brain mitochondria, and that a lack of Nrf2 in vivo produces altered supercomplex formation in mitochondria.Our findings support a role of Nrf2 in mediating IPC-induced protection in astrocytes, which can profoundly impact the ischemic tolerance of neurons. In addition, we provide novel evidence for the association of Nrf2 to brain mitochondria and supercomplex formation. These studies offer new targets and pathways of Nrf2, which may be heavily implicated following cerebral ischemia.

Alterations in intrinsic mitochondrial function with aging are fiber type-specific and do not explain differential atrophy between muscles.

PubMed

Picard, Martin; Ritchie, Darmyn; Thomas, Melissa M; Wright, Kathryn J; Hepple, Russell T

2011-12-01

To determine whether mitochondrial dysfunction is causally related to muscle atrophy with aging, we examined respiratory capacity, H(2) O(2) emission, and function of the mitochondrial permeability transition pore (mPTP) in permeabilized myofibers prepared from four rat muscles that span a range of fiber type and degree of age-related atrophy. Muscle atrophy with aging was greatest in fast-twitch gastrocnemius (Gas) muscle (-38%), intermediate in both the fast-twitch extensor digitorum longus (EDL) and slow-twitch soleus (Sol) muscles (-21%), and non-existent in adductor longus (AL) muscle (+47%). In contrast, indices of mitochondrial dysfunction did not correspond to this differential degree of atrophy. Specifically, despite higher protein expression for oxidative phosphorylation (oxphos) system in fast Gas and EDL, state III respiratory capacity per myofiber wet weight was unchanged with aging, whereas the slow Sol showed proportional decreases in oxphos protein, citrate synthase activity, and state III respiration. Free radical leak (H(2) O(2) emission per O(2) flux) under state III respiration was higher with aging in the fast Gas, whereas state II free radical leak was higher in the slow AL. Only the fast muscles had impaired mPTP function with aging, with lower mitochondrial calcium retention capacity in EDL and shorter time to mPTP opening in Gas and EDL. Collectively, our results underscore that the age-related changes in muscle mitochondrial function depend largely upon fiber type and are unrelated to the severity of muscle atrophy, suggesting that intrinsic changes in mitochondrial function are unlikely to be causally involved in aging muscle atrophy. © 2011 The Authors. Aging Cell © 2011 Blackwell Publishing Ltd/Anatomical Society of Great Britain and Ireland.

Metabolome and proteome profiling of complex I deficiency induced by rotenone.

PubMed

Gielisch, Ina; Meierhofer, David

2015-01-02

Complex I (CI; NADH dehydrogenase) deficiency causes mitochondrial diseases, including Leigh syndrome. A variety of clinical symptoms of CI deficiency are known, including neurodegeneration. Here, we report an integrative study combining liquid chromatography-mass spectrometry (LC-MS)-based metabolome and proteome profiling in CI deficient HeLa cells. We report a rapid LC-MS-based method for the relative quantification of targeted metabolome profiling with an additional layer of confidence by applying multiple reaction monitoring (MRM) ion ratios for further identity confirmation and robustness. The proteome was analyzed by label-free quantification (LFQ). More than 6000 protein groups were identified. Pathway and network analyses revealed that the respiratory chain was highly deregulated, with metabolites such as FMN, FAD, NAD(+), and ADP, direct players of the OXPHOS system, and metabolites of the TCA cycle decreased up to 100-fold. Synthesis of functional iron-sulfur clusters, which are of central importance for the electron transfer chain, and degradation products like bilirubin were also significantly reduced. Glutathione metabolism on the pathway level, as well as individual metabolite components such as NADPH, glutathione (GSH), and oxidized glutathione (GSSG), was downregulated. Overall, metabolome and proteome profiles in CI deficient cells correlated well, supporting our integrated approach.

Adaptation of the Mitochondrial Genome in Cephalopods: Enhancing Proton Translocation Channels and the Subunit Interactions

PubMed Central

Almeida, Daniela; Maldonado, Emanuel; Vasconcelos, Vitor; Antunes, Agostinho

2015-01-01

Mitochondrial protein-coding genes (mt genes) encode subunits forming complexes of crucial cellular pathways, including those involved in the vital process of oxidative phosphorylation (OXPHOS). Despite the vital role of the mitochondrial genome (mt genome) in the survival of organisms, little is known with respect to its adaptive implications within marine invertebrates. The molluscan Class Cephalopoda is represented by a marine group of species known to occupy contrasting environments ranging from the intertidal to the deep sea, having distinct metabolic requirements, varied body shapes and highly advanced visual and nervous systems that make them highly competitive and successful worldwide predators. Thus, cephalopods are valuable models for testing natural selection acting on their mitochondrial subunits (mt subunits). Here, we used concatenated mt genes from 17 fully sequenced mt genomes of diverse cephalopod species to generate a robust mitochondrial phylogeny for the Class Cephalopoda. We followed an integrative approach considering several branches of interest–covering cephalopods with distinct morphologies, metabolic rates and habitats–to identify sites under positive selection and localize them in the respective protein alignment and/or tridimensional structure of the mt subunits. Our results revealed significant adaptive variation in several mt subunits involved in the energy production pathway of cephalopods: ND5 and ND6 from Complex I, CYTB from Complex III, COX2 and COX3 from Complex IV, and in ATP8 from Complex V. Furthermore, we identified relevant sites involved in protein-interactions, lining proton translocation channels, as well as disease/deficiencies related sites in the aforementioned complexes. A particular case, revealed by this study, is the involvement of some positively selected sites, found in Octopoda lineage in lining proton translocation channels (site 74 from ND5) and in interactions between subunits (site 507 from ND5) of Complex I. PMID:26285039

Oxygen and differentiation status modulate the effect of X-ray irradiation on physiology and mitochondrial proteome of human neuroblastoma cells.

PubMed

Džinić, Tamara; Hartwig, Sonja; Lehr, Stefan; Dencher, Norbert A

2016-12-01

Cytotoxic effects, including oxidative stress, of low linear energy transfer (LET)-ionizing radiation are often underestimated and studies of their mechanisms using cell culture models are widely conducted with cells cultivated at atmospheric oxygen that does not match its physiological levels in body tissues. Also, cell differentiation status plays a role in the outcome of experiments. We compared effects of 2 Gy X-ray irradiation on the physiology and mitochondrial proteome of nondifferentiated and human neuroblastoma (SH-SY5Y) cells treated with retinoic acid cultivated at 21% and 5% O 2 . Irradiation did not affect the amount of subunits of OxPhos complexes and other non-OxPhos mitochondrial proteins, except for heat shock protein 70, which was increased depending on oxygen level and differentiation status. These two factors were proven to modulate mitochondrial membrane potential and the bioenergetic status of cells. We suggest, moreover, that oxygen plays a role in the differentiation of human SH-SY5Y cells.

Electronic cigarette aerosols and copper nanoparticles induce mitochondrial stress and promote DNA fragmentation in lung fibroblasts.

PubMed

Lerner, Chad A; Rutagarama, Pierrot; Ahmad, Tanveer; Sundar, Isaac K; Elder, Alison; Rahman, Irfan

2016-09-02

Oxidants or nanoparticles have recently been identified as constituents of aerosols released from various styles of electronic cigarettes (E-cigs). Cells in the lung may be directly exposed to these constituents and harbor reactive properties capable of incurring acute cell injury. Our results show mitochondria are sensitive to both E-cig aerosols and aerosol containing copper nanoparticles when exposed to human lung fibroblasts (HFL-1) using an Air-Liquid Interface culture system, evident by elevated levels of mitochondrial ROS (mtROS). Increased mtROS after aerosol exposure is associated with reduced stability of OxPhos electron transport chain (ETC) complex IV subunit and nuclear DNA fragmentation. Increased levels of IL-8 and IL-6 in HFL-1 conditioned media were also observed. These findings reveal both mitochondrial, genotoxic, and inflammatory stresses are features of direct cell exposure to E-cig aerosols which are ensued by inflammatory duress, raising a concern on deleterious effect of vaping. Copyright © 2016 Elsevier Inc. All rights reserved.

Elevated Nicotinamide Phosphoribosyl Transferase in Skeletal Muscle Augments Exercise Performance and Mitochondrial Respiratory Capacity Following Exercise Training

PubMed Central

Brouwers, Bram; Stephens, Natalie A.; Costford, Sheila R.; Hopf, Meghan E.; Ayala, Julio E.; Yi, Fanchao; Xie, Hui; Li, Jian-Liang; Gardell, Stephen J.; Sparks, Lauren M.; Smith, Steven R.

2018-01-01

Mice overexpressing NAMPT in skeletal muscle (NamptTg mice) develop higher exercise endurance and maximal aerobic capacity (VO2max) following voluntary exercise training compared to wild-type (WT) mice. Here, we aimed to investigate the mechanisms underlying by determining skeletal muscle mitochondrial respiratory capacity in NamptTg and WT mice. Body weight and body composition, tissue weight (gastrocnemius, quadriceps, soleus, heart, liver, and epididymal white adipose tissue), skeletal muscle and liver glycogen content, VO2max, skeletal muscle mitochondrial respiratory capacity (measured by high-resolution respirometry), skeletal muscle gene expression (measured by microarray and qPCR), and skeletal muscle protein content (measured by Western blot) were determined following 6 weeks of voluntary exercise training (access to running wheel) in 13-week-old male NamptTg (exercised NamptTg) mice and WT (exercised WT) mice. Daily running distance and running time during the voluntary exercise training protocol were recorded. Daily running distance (p = 0.51) and running time (p = 0.85) were not significantly different between exercised NamptTg mice and exercised WT mice. VO2max was higher in exercised NamptTg mice compared to exercised WT mice (p = 0.02). Body weight (p = 0.92), fat mass (p = 0.49), lean mass (p = 0.91), tissue weight (all p > 0.05), and skeletal muscle (p = 0.72) and liver (p = 0.94) glycogen content were not significantly different between exercised NamptTg mice and exercised WT mice. Complex I oxidative phosphorylation (OXPHOS) respiratory capacity supported by fatty acid substrates (p < 0.01), maximal (complex I+II) OXPHOS respiratory capacity supported by glycolytic (p = 0.02) and fatty acid (p < 0.01) substrates, and maximal uncoupled respiratory capacity supported by fatty acid substrates (p < 0.01) was higher in exercised NamptTg mice compared to exercised WT mice. Transcriptomic analyses revealed differential expression for genes involved in oxidative metabolism in exercised NamptTg mice compared to exercised WT mice, specifically, enrichment for the gene set related to the SIRT3-mediated signaling pathway. SIRT3 protein content correlated with NAMPT protein content (r = 0.61, p = 0.04). In conclusion, NamptTg mice develop higher exercise capacity following voluntary exercise training compared to WT mice, which is paralleled by higher mitochondrial respiratory capacity in skeletal muscle. The changes in SIRT3 targets suggest that these effects are due to remodeling of mitochondrial function. PMID:29942262

Associations between fatty acid oxidation, hepatic mitochondrial function, and plasma acylcarnitine levels in mice.

PubMed

Bjørndal, Bodil; Alterås, Eva Katrine; Lindquist, Carine; Svardal, Asbjørn; Skorve, Jon; Berge, Rolf K

2018-01-01

The 4-thia fatty acid tetradecylthiopropionic acid (TTP) is known to inhibit mitochondrial β-oxidation, and can be used as chemically induced hepatic steatosis-model in rodents, while 3-thia fatty acid tetradecylthioacetic acid (TTA) stimulates fatty acid oxidation through activation of peroxisome proliferator activated receptor alpha (PPARα). We wished to determine how these two compounds affected in vivo respiration and mitochondrial efficiency, with an additional goal to elucidate whether mitochondrial function is reflected in plasma acylcarnitine levels. C57BL/6 mice were divided in 4 groups of 10 mice and fed a control low-fat diet, low-fat diets with 0.4% ( w /w) TTP, 0.4% TTA or a combination of these two fatty acids for three weeks ( n  = 10). At sacrifice, β-oxidation and oxidative phosphorylation (OXPHOS) capacity was analysed in fresh liver samples. Hepatic mitochondria were studied using transmission electron microscopy. Lipid classes were measured in plasma, heart and liver, acylcarnitines were measured in plasma, and gene expression was measured in liver. The TTP diet resulted in hepatic lipid accumulation, plasma L-carnitine and acetylcarnitine depletion and elevated palmitoylcarnitine and non-esterified fatty acid levels. No significant lipid accumulation was observed in heart. The TTA supplement resulted in enhanced hepatic β-oxidation, accompanied by an increased level of acetylcarnitine and palmitoylcarnitine in plasma. Analysis of mitochondrial respiration showed that TTP reduced oxidative phosphorylation, while TTA increased the maximum respiratory capacity of the electron transport system. Combined treatment with TTP and TTA resulted in a profound stimulation of genes involved in the PPAR-response and L-carnitine metabolism, and partly prevented triacylglycerol accumulation in the liver concomitant with increased peroxisomal β-oxidation and depletion of plasma acetylcarnitines. Despite an increased number of mitochondria in the liver of TTA + TTP fed mice, the OXPHOS capacity was significantly reduced. This study indicates that fatty acid β-oxidation directly affects mitochondrial respiratory capacity in liver. As plasma acylcarnitines reflected the reduced mitochondrial β-oxidation in TTP-fed mice, they could be useful tools to monitor mitochondrial function. As mitochondrial dysfunction is a major determinant of metabolic disease, this supports their use as plasma markers of cardiovascular risk in humans. Results however indicate that high PPAR activation obscures the interpretation of plasma acylcarnitine levels.

Energy transfer in “parasitic” cancer metabolism

PubMed Central

Martinez-Outschoorn, Ubaldo E; Pestell, Richard G; Howell, Anthony; Tykocinski, Mark L; Nagajyothi, Fnu; Machado, Fabiana S; Tanowitz, Herbert B

2011-01-01

It is now widely recognized that the tumor microenvironment promotes cancer cell growth and metastasis via changes in cytokine secretion and extra-cellular matrix remodeling. However, the role of tumor stromal cells in providing energy for epithelial cancer cell growth is a newly emerging paradigm. For example, we and others have recently proposed that tumor growth and metastasis is related to an energy imbalance. Host cells produce energy-rich nutrients via catabolism (through autophagy, mitophagy and aerobic glycolysis), which are then transferred to cancer cells, to fuel anabolic tumor growth. Stromal cell derived L-lactate is taken up by cancer cells and is used for mitochondrial oxidative phosphorylation (OXPHOS), to produce ATP efficiently. However, “parasitic” energy transfer may be a more generalized mechanism in cancer biology than previously appreciated. Two recent papers in Science and Nature Medicine now show that lipolysis in host tissues also fuels tumor growth. These studies demonstrate that free fatty acids produced by host cell lipolysis are re-used via β-oxidation (β-OX) in cancer cell mitochondria. Thus, stromal catabolites (such as lactate, ketones, glutamine and free fatty acids) promote tumor growth by acting as high-energy onco-metabolites. As such, host catabolism via autophagy, mitophagy and lipolysis may explain the pathogenesis of cancer-associated cachexia and provides exciting new druggable targets for novel therapeutic interventions. Taken together, these findings also suggest that tumor cells promote their own growth and survival by behaving as a “parasitic organism.” Hence, we propose the term “parasitic cancer metabolism” to describe this type of metabolic-coupling in tumors. Targeting tumor cell mitochondria (OXPHOS and β-OX) would effectively uncouple tumor cells from their hosts, leading to their acute starvation. In this context, we discuss new evidence that high-energy onco-metabolites (produced by the stroma) can confer drug resistance. Importantly, this metabolic chemo-resistance is reversed by blocking OXPHOS in cancer cell mitochondria, with drugs like Metformin, a mitochondrial “poison.” In summary, parasitic cancer metabolism is achieved architecturally by dividing tumor tissue into at least two well-defined opposing “metabolic compartments:” catabolic and anabolic. PMID:22033146

Changes in mitochondrial respiration in the human placenta over gestation.

PubMed

Holland, Olivia J; Hickey, Anthony J R; Alvsaker, Anna; Moran, Stephanie; Hedges, Christopher; Chamley, Lawrence W; Perkins, Anthony V

2017-09-01

Placental mitochondria are subjected to micro-environmental changes throughout gestation, in particular large variations in oxygen. How placental mitochondrial respiration adapts to changing oxygen concentrations remains unexplored. Additionally, placental tissue is often studied in culture; however, the effect of culture on placental mitochondria is unclear. Placental tissue was obtained from first trimester and term (laboured and non-laboured) pregnancies, and selectively permeabilized to access mitochondria. Respirometry was used to compare respiration states and substrate use in mitochondria. Additionally, explants of placental tissue were cultured for four, 12, 24, 48, or 96 h and respiration measured. Mitochondrial respiration decreased at 11 weeks compared to earlier gestations (p = 0.05-0.001), and mitochondrial content increased at 12-13 weeks compared to 7-10 weeks (p = 0.042). In term placentae, oxidative phosphorylation (OXPHOS) through mitochondrial complex IV (p < 0.001), the relative proportion of OXPHOS CI (p < 0.001), the total capacity of the respiratory system (p = 0.003), and mitochondrial content (p < 0.001) were higher compared to first trimester. Respiration was increased (p ≤ 0.006-0.001) in laboured compared to non-laboured placenta. After four hours of culture, respiration was depressed compared to fresh tissue from the same placenta and continued to decline with time in culture. Markers of apoptosis were increased, while markers of autophagy, mitochondrial biogenesis, and mitochondrial membrane potential were decreased after four hours of culture. Respiration and mitochondrial content alter over gestation/with labour. Decreased respiration at 11 weeks and increased mitochondrial content at 12-13 weeks may relate to onset of maternal blood flow, and increased respiration as a result of labour may be an adaptation to ischaemia-reperfusion. At term, mitochondria were more susceptible to changes in respiratory function relative to first trimester when cultured in vitro, perhaps reflecting changes in metabolic demands as gestation progresses. Metabolic plasticity of placental mitochondria has relevance to placenta-mediated diseases. Copyright © 2017 Elsevier Ltd. All rights reserved.

Bergamot natural products eradicate cancer stem cells (CSCs) by targeting mevalonate, Rho-GDI-signalling and mitochondrial metabolism.

PubMed

Fiorillo, Marco; Peiris-Pagès, Maria; Sanchez-Alvarez, Rosa; Bartella, Lucia; Di Donna, Leonardo; Dolce, Vincenza; Sindona, Giovanni; Sotgia, Federica; Cappello, Anna Rita; Lisanti, Michael P

2018-04-04

Here, we show that a 2:1 mixture of Brutieridin and Melitidin, termed "BMF", has a statin-like properties, which blocks the action of the rate-limiting enzyme for mevalonate biosynthesis, namely HMGR (3-hydroxy-3-methylglutaryl-CoA-reductase). Moreover, our results indicate that BMF functionally inhibits several key characteristics of CSCs. More specifically, BMF effectively i) reduced ALDH activity, ii) blocked mammosphere formation and iii) inhibited the activation of CSC-associated signalling pathways (STAT1/3, Notch and Wnt/beta-catenin) targeting Rho-GDI-signalling. In addition, BMF metabolically inhibited mitochondrial respiration (OXPHOS) and fatty acid oxidation (FAO). Importantly, BMF did not show the same toxic side-effects in normal fibroblasts that were observed with statins. Lastly, we show that high expression of the mRNA species encoding HMGR is associated with poor clinical outcome in breast cancer patients, providing a potential companion diagnostic for BMF-directed personalized therapy. Copyright © 2018 Elsevier B.V. All rights reserved.

Short-term sleep deprivation with exposure to nocturnal light alters mitochondrial bioenergetics in Drosophila.

PubMed

Rodrigues, Nathane Rosa; Macedo, Giulianna Echeverria; Martins, Illana Kemmerich; Gomes, Karen Kich; de Carvalho, Nélson Rodrigues; Posser, Thaís; Franco, Jeferson Luis

2018-05-20

Many studies have shown the effects of sleep deprivation in several aspects of health and disease. However, little is known about how mitochondrial bioenergetics function is affected under this condition. To clarify this, we developed a simple model of short-term sleep deprivation, in which fruit-flies were submitted to a nocturnal light condition and then mitochondrial parameters were assessed by high resolution respirometry (HRR). Exposure of flies to constant light was able to alter sleep patterns, causing locomotor deficits, increasing ROS production and lipid peroxidation, affecting mitochondrial activity, antioxidant defense enzymes and caspase activity. HRR analysis showed that sleep deprivation affected mitochondrial bioenergetics capacity, decreasing respiration at oxidative phosphorylation (OXPHOS) and electron transport system (ETS). In addition, the expression of genes involved in the response to oxidative stress and apoptosis were increased. Thus, our results suggest a connection between sleep deprivation and oxidative stress, pointing to mitochondria as a possible target of this relationship. Copyright © 2018 Elsevier Inc. All rights reserved.

Posttranscriptional Control of T Cell Effector Function by Aerobic Glycolysis

PubMed Central

Chang, Chih-Hao; Curtis, Jonathan D.; Maggi, Leonard B.; Faubert, Brandon; Villarino, Alejandro V.; O’Sullivan, David; Huang, Stanley Ching-Cheng; van der Windt, Gerritje J.W.; Blagih, Julianna; Qiu, Jing; Weber, Jason D.; Pearce, Edward J.; Jones, Russell G.; Pearce, Erika L.

2013-01-01

SUMMARY A “switch” from oxidative phosphorylation (OXPHOS) to aerobic glycolysis is a hallmark of T cell activation and is thought to be required to meet the metabolic demands of proliferation. However, why proliferating cells adopt this less efficient metabolism, especially in an oxygen-replete environment, remains incompletely understood. We show here that aerobic glycolysis is specifically required for effector function in T cells but that this pathway is not necessary for proliferation or survival. When activated T cells are provided with costimulation and growth factors but are blocked from engaging glycolysis, their ability to produce IFN-γ is markedly compromised. This defect is translational and is regulated by the binding of the glycolysis enzyme GAPDH to AU-rich elements within the 3′ UTR of IFN-γ mRNA. GAPDH, by engaging/disengaging glycolysis and through fluctuations in its expression, controls effector cytokine production. Thus, aerobic glycolysis is a metabolically regulated signaling mechanism needed to control cellular function. PMID:23746840

Loss of mitochondrial exo/endonuclease EXOG affects mitochondrial respiration and induces ROS-mediated cardiomyocyte hypertrophy.

PubMed

Tigchelaar, Wardit; Yu, Hongjuan; de Jong, Anne Margreet; van Gilst, Wiek H; van der Harst, Pim; Westenbrink, B Daan; de Boer, Rudolf A; Silljé, Herman H W

2015-01-15

Recently, a locus at the mitochondrial exo/endonuclease EXOG gene, which has been implicated in mitochondrial DNA repair, was associated with cardiac function. The function of EXOG in cardiomyocytes is still elusive. Here we investigated the role of EXOG in mitochondrial function and hypertrophy in cardiomyocytes. Depletion of EXOG in primary neonatal rat ventricular cardiomyocytes (NRVCs) induced a marked increase in cardiomyocyte hypertrophy. Depletion of EXOG, however, did not result in loss of mitochondrial DNA integrity. Although EXOG depletion did not induce fetal gene expression and common hypertrophy pathways were not activated, a clear increase in ribosomal S6 phosphorylation was observed, which readily explains increased protein synthesis. With the use of a Seahorse flux analyzer, it was shown that the mitochondrial oxidative consumption rate (OCR) was increased 2.4-fold in EXOG-depleted NRVCs. Moreover, ATP-linked OCR was 5.2-fold higher. This increase was not explained by mitochondrial biogenesis or alterations in mitochondrial membrane potential. Western blotting confirmed normal levels of the oxidative phosphorylation (OXPHOS) complexes. The increased OCR was accompanied by a 5.4-fold increase in mitochondrial ROS levels. These increased ROS levels could be normalized with specific mitochondrial ROS scavengers (MitoTEMPO, mnSOD). Remarkably, scavenging of excess ROS strongly attenuated the hypertrophic response. In conclusion, loss of EXOG affects normal mitochondrial function resulting in increased mitochondrial respiration, excess ROS production, and cardiomyocyte hypertrophy. Copyright © 2015 the American Physiological Society.

Novel large-range mitochondrial DNA deletions and fatal multisystemic disorder with prominent hepatopathy

DOE Office of Scientific and Technical Information (OSTI.GOV)

Bianchi, Marzia; Rizza, Teresa; Verrigni, Daniela

2011-11-18

Highlights: Black-Right-Pointing-Pointer Expanded array of mtDNA deletions. Black-Right-Pointing-Pointer Pearson syndrome with prominent hepatopathy associated with single mtDNA deletions. Black-Right-Pointing-Pointer Detection of deletions in fibroblasts and blood avoids muscle and liver biopsy. Black-Right-Pointing-Pointer Look for mtDNA deletions before to study nuclear genes related to mtDNA depletion. -- Abstract: Hepatic involvement in mitochondrial cytopathies rarely manifests in adulthood, but is a common feature in children. Multiple OXPHOS enzyme defects in children with liver involvement are often associated with dramatically reduced amounts of mtDNA. We investigated two novel large scale deletions in two infants with a multisystem disorder and prominent hepatopathy. Amount ofmore » mtDNA deletions and protein content were measured in different post-mortem tissues. The highest levels of deleted mtDNA were in liver, kidney, pancreas of both patients. Moreover, mtDNA deletions were detected in cultured skin fibroblasts in both patients and in blood of one during life. Biochemical analysis showed impairment of mainly complex I enzyme activity. Patients manifesting multisystem disorders in childhood may harbour rare mtDNA deletions in multiple tissues. For these patients, less invasive blood specimens or cultured fibroblasts can be used for molecular diagnosis. Our data further expand the array of deletions in the mitochondrial genomes in association with liver failure. Thus analysis of mtDNA should be considered in the diagnosis of childhood-onset hepatopathies.« less

The ADP/ATP Carrier and Its Relationship to Oxidative Phosphorylation in Ancestral Protist Trypanosoma brucei

PubMed Central

Gnipová, Anna; Šubrtová, Karolína; Panicucci, Brian; Horváth, Anton; Lukeš, Julius

2015-01-01

The highly conserved ADP/ATP carrier (AAC) is a key energetic link between the mitochondrial (mt) and cytosolic compartments of all aerobic eukaryotic cells, as it exchanges the ATP generated inside the organelle for the cytosolic ADP. Trypanosoma brucei, a parasitic protist of medical and veterinary importance, possesses a single functional AAC protein (TbAAC) that is related to the human and yeast ADP/ATP carriers. However, unlike previous studies performed with these model organisms, this study showed that TbAAC is most likely not a stable component of either the respiratory supercomplex III+IV or the ATP synthasome but rather functions as a physically separate entity in this highly diverged eukaryote. Therefore, TbAAC RNA interference (RNAi) ablation in the insect stage of T. brucei does not impair the activity or arrangement of the respiratory chain complexes. Nevertheless, RNAi silencing of TbAAC caused a severe growth defect that coincides with a significant reduction of mt ATP synthesis by both substrate and oxidative phosphorylation. Furthermore, TbAAC downregulation resulted in a decreased level of cytosolic ATP, a higher mt membrane potential, an elevated amount of reactive oxygen species, and a reduced consumption of oxygen in the mitochondria. Interestingly, while TbAAC has previously been demonstrated to serve as the sole ADP/ATP carrier for ADP influx into the mitochondria, our data suggest that a second carrier for ATP influx may be present and active in the T. brucei mitochondrion. Overall, this study provides more insight into the delicate balance of the functional relationship between TbAAC and the oxidative phosphorylation (OXPHOS) pathway in an early diverged eukaryote. PMID:25616281

Comparative Analysis of the Mitochondrial Physiology of Pancreatic β Cells

PubMed Central

Kim, Chul; Patel, Pinal; Gouvin, Lindsey M.; Brown, Melissa L.; Khalil, Ahmed; Henchey, Elizabeth M; Heuck, Alejandro P.; Yadava, Nagendra

2014-01-01

The mitochondrial metabolism of β cells is thought to be highly specialized. Its direct comparison with other cells using isolated mitochondria is limited by the availability of islets/β cells in sufficient quantity. In this study, we have compared mitochondrial metabolism of INS1E/β cells with other cells in intact and permeabilized states. To selectively permeabilize the plasma membrane, we have evaluated the use of perfringolysin-O (PFO) in conjunction with microplate-based respirometry. PFO is a protein that binds membranes based on a threshold level of active cholesterol. Therefore, unless active cholesterol reaches a threshold level in mitochondria, they are expected to remain untouched by PFO. Cytochrome c sensitivity tests showed that in PFO-permeabilized cells, the mitochondrial integrity was completely preserved. Our data show that a time-dependent decline of the oligomycin-insensitive respiration observed in INS1E cells was due to a limitation in substrate supply to the respiratory chain. We predict that it is linked with the β cell-specific metabolism involving metabolites shuttling between the cytoplasm and mitochondria. In permeabilized β cells, the Complex l-dependent respiration was either transient or absent because of the inefficient TCA cycle. The TCA cycle insufficiency was confirmed by analysis of the CO2 evolution. This may be linked with lower levels of NAD+, which is required as a co-factor for CO2 producing reactions of the TCA cycle. β cells showed comparable OxPhos and respiratory capacities that were not affected by the inorganic phosphate (Pi) levels in the respiration medium. They showed lower ADP-stimulation of the respiration on different substrates. We believe that this study will significantly enhance our understanding of the β cell mitochondrial metabolism. PMID:25309834

The evidence basis for coenzyme Q therapy in oxidative phosphorylation disease.

PubMed

Haas, Richard H

2007-06-01

The evidence supporting a treatment benefit for coenzyme Q10 (CoQ10) in primary mitochondrial disease (mitochondrial disease) whilst positive is limited. Mitochondrial disease in this context is defined as genetic disease causing an impairment in mitochondrial oxidative phosphorylation (OXPHOS). There are no treatment trials achieving the highest Level I evidence designation. Reasons for this include the relative rarity of mitochondrial disease, the heterogeneity of mitochondrial disease, the natural cofactor status and easy 'over the counter availability' of CoQ10 all of which make funding for the necessary large blinded clinical trials unlikely. At this time the best evidence for efficacy comes from controlled trials in common cardiovascular and neurodegenerative diseases with mitochondrial and OXPHOS dysfunction the etiology of which is most likely multifactorial with environmental factors playing on a background of genetic predisposition. There remain questions about dosing, bioavailability, tissue penetration and intracellular distribution of orally administered CoQ10, a compound which is endogenously produced within the mitochondria of all cells. In some mitochondrial diseases and other commoner disorders such as cardiac disease and Parkinson's disease low mitochondrial or tissue levels of CoQ10 have been demonstrated providing an obvious rationale for supplementation. This paper discusses the current state of the evidence supporting the use of CoQ10 in mitochondrial disease.

Overexpression of the human DEK oncogene reprograms cellular metabolism and promotes glycolysis

PubMed Central

Watanabe, Miki; Muraleedharan, Ranjithmenon; Lambert, Paul F.; Lane, Andrew N.; Romick-Rosendale, Lindsey E.; Wells, Susanne I.

2017-01-01

The DEK oncogene is overexpressed in many human malignancies including at early tumor stages. Our reported in vitro and in vivo models of squamous cell carcinoma have demonstrated that DEK contributes functionally to cellular and tumor survival and to proliferation. However, the underlying molecular mechanisms remain poorly understood. Based on recent RNA sequencing experiments, DEK expression was necessary for the transcription of several metabolic enzymes involved in anabolic pathways. This identified a possible mechanism whereby DEK may drive cellular metabolism to enable cell proliferation. Functional metabolic Seahorse analysis demonstrated increased baseline and maximum extracellular acidification rates, a readout of glycolysis, in DEK-overexpressing keratinocytes and squamous cell carcinoma cells. DEK overexpression also increased the maximum rate of oxygen consumption and therefore increased the potential for oxidative phosphorylation (OxPhos). To detect small metabolites that participate in glycolysis and the tricarboxylic acid cycle (TCA) that supplies substrate for OxPhos, we carried out NMR-based metabolomics studies. We found that high levels of DEK significantly reprogrammed cellular metabolism and altered the abundances of amino acids, TCA cycle intermediates and the glycolytic end products lactate, alanine and NAD+. Taken together, these data support a scenario whereby overexpression of the human DEK oncogene reprograms keratinocyte metabolism to fulfill energy and macromolecule demands required to enable and sustain cancer cell growth. PMID:28558019

Mitochondrial pyruvate carrier function determines cell stemness and metabolic reprogramming in cancer cells

PubMed Central

Li, Xiaoran; Kan, Quancheng; Fan, Zhirui; Li, Yaqing; Ji, Yasai; Zhao, Jing; Zhang, Mingzhi; Grigalavicius, Mantas; Berge, Viktor; Goscinski, Mariusz Adam; M. Nesland, Jahn; Suo, Zhenhe

2017-01-01

One of the remarkable features of cancer cells is aerobic glycolysis, a phenomenon known as the “Warburg Effect”, in which cells rely preferentially on glycolysis instead of oxidative phosphorylation (OXPHOS) as the main energy source even in the presence of high oxygen tension. Cells with dysfunctional mitochondria are unable to generate sufficient ATP from mitochondrial OXPHOS, and then are forced to rely on glycolysis for ATP generation. Here we report our results in a prostate cancer cell line in which the mitochondrial pyruvate carrier 1 (MPC1) gene was knockout. It was discovered that the MPC1 gene knockout cells revealed a metabolism reprogramming to aerobic glycolysis with reduced ATP production, and the cells became more migratory and resistant to both chemotherapy and radiotherapy. In addition, the MPC1 knockout cells expressed significantly higher levels of the stemness markers Nanog, Hif1α, Notch1, CD44 and ALDH. To further verify the correlation of MPC gene function and cell stemness/metabolic reprogramming, MPC inhibitor UK5099 was applied in two ovarian cancer cell lines and similar results were obtained. Taken together, our results reveal that functional MPC may determine the fate of metabolic program and the stemness status of cancer cells in vitro. PMID:28624784

Overexpression of the human DEK oncogene reprograms cellular metabolism and promotes glycolysis.

PubMed

Matrka, Marie C; Watanabe, Miki; Muraleedharan, Ranjithmenon; Lambert, Paul F; Lane, Andrew N; Romick-Rosendale, Lindsey E; Wells, Susanne I

2017-01-01

The DEK oncogene is overexpressed in many human malignancies including at early tumor stages. Our reported in vitro and in vivo models of squamous cell carcinoma have demonstrated that DEK contributes functionally to cellular and tumor survival and to proliferation. However, the underlying molecular mechanisms remain poorly understood. Based on recent RNA sequencing experiments, DEK expression was necessary for the transcription of several metabolic enzymes involved in anabolic pathways. This identified a possible mechanism whereby DEK may drive cellular metabolism to enable cell proliferation. Functional metabolic Seahorse analysis demonstrated increased baseline and maximum extracellular acidification rates, a readout of glycolysis, in DEK-overexpressing keratinocytes and squamous cell carcinoma cells. DEK overexpression also increased the maximum rate of oxygen consumption and therefore increased the potential for oxidative phosphorylation (OxPhos). To detect small metabolites that participate in glycolysis and the tricarboxylic acid cycle (TCA) that supplies substrate for OxPhos, we carried out NMR-based metabolomics studies. We found that high levels of DEK significantly reprogrammed cellular metabolism and altered the abundances of amino acids, TCA cycle intermediates and the glycolytic end products lactate, alanine and NAD+. Taken together, these data support a scenario whereby overexpression of the human DEK oncogene reprograms keratinocyte metabolism to fulfill energy and macromolecule demands required to enable and sustain cancer cell growth.

Bioenergetics of Mammalian Sperm Capacitation

PubMed Central

Ferramosca, Alessandra; Zara, Vincenzo

2014-01-01

After ejaculation, the mammalian male gamete must undergo the capacitation process, which is a prerequisite for egg fertilization. The bioenergetics of sperm capacitation is poorly understood despite its fundamental role in sustaining the biochemical and molecular events occurring during gamete activation. Glycolysis and mitochondrial oxidative phosphorylation (OXPHOS) are the two major metabolic pathways producing ATP which is the primary source of energy for spermatozoa. Since recent data suggest that spermatozoa have the ability to use different metabolic substrates, the main aim of this work is to present a broad overview of the current knowledge on the energy-producing metabolic pathways operating inside sperm mitochondria during capacitation in different mammalian species. Metabolism of glucose and of other energetic substrates, such as pyruvate, lactate, and citrate, is critically analyzed. Such knowledge, besides its obvious importance for basic science, could eventually translate into the development of novel strategies for treatment of male infertility, artificial reproduction, and sperm selection methods. PMID:24791005

Bioenergetics of mammalian sperm capacitation.

PubMed

Ferramosca, Alessandra; Zara, Vincenzo

2014-01-01

After ejaculation, the mammalian male gamete must undergo the capacitation process, which is a prerequisite for egg fertilization. The bioenergetics of sperm capacitation is poorly understood despite its fundamental role in sustaining the biochemical and molecular events occurring during gamete activation. Glycolysis and mitochondrial oxidative phosphorylation (OXPHOS) are the two major metabolic pathways producing ATP which is the primary source of energy for spermatozoa. Since recent data suggest that spermatozoa have the ability to use different metabolic substrates, the main aim of this work is to present a broad overview of the current knowledge on the energy-producing metabolic pathways operating inside sperm mitochondria during capacitation in different mammalian species. Metabolism of glucose and of other energetic substrates, such as pyruvate, lactate, and citrate, is critically analyzed. Such knowledge, besides its obvious importance for basic science, could eventually translate into the development of novel strategies for treatment of male infertility, artificial reproduction, and sperm selection methods.

Oxaloacetate Ameliorates Chemical Liver Injury via Oxidative Stress Reduction and Enhancement of Bioenergetic Fluxes.

PubMed

Kuang, Ye; Han, Xiaoyun; Xu, Mu; Wang, Yue; Zhao, Yuxiang; Yang, Qing

2018-05-31

Chemical injury is partly due to free radical lipid peroxidation, which can induce oxidative stress and produce a large number of reactive oxygen species (ROS). Oxaloacetic acid is an important intermediary in the tricarboxylic acid cycle (TCA cycle) and participates in metabolism and energy production. In our study, we found that oxaloacetate (OA) effectively alleviated liver injury which was induced by hydrogen peroxide (H₂O₂) in vitro and carbon tetrachloride (CCl₄) in vivo. OA scavenged ROS, prevented oxidative damage and maintained the normal structure of mitochondria. We further confirmed that OA increased adenosine triphosphate (ATP) by promoting the TCA production cycle and oxidative phosphorylation (OXPHOS). Finally, OA inhibited the mitogen-activated protein kinase (MAPK) and apoptotic pathways by suppressing tumor necrosis factor-α (TNF-α). Our findings reveal a mechanism for OA ameliorating chemical liver injury and suggest a possible implementation for preventing the chemical liver injury.

CLUH couples mitochondrial distribution to the energetic and metabolic status.

PubMed

Wakim, Jamal; Goudenege, David; Perrot, Rodolphe; Gueguen, Naig; Desquiret-Dumas, Valerie; Chao de la Barca, Juan Manuel; Dalla Rosa, Ilaria; Manero, Florence; Le Mao, Morgane; Chupin, Stephanie; Chevrollier, Arnaud; Procaccio, Vincent; Bonneau, Dominique; Logan, David C; Reynier, Pascal; Lenaers, Guy; Khiati, Salim

2017-06-01

Mitochondrial dynamics and distribution are critical for supplying ATP in response to energy demand. CLUH is a protein involved in mitochondrial distribution whose dysfunction leads to mitochondrial clustering, the metabolic consequences of which remain unknown. To gain insight into the role of CLUH on mitochondrial energy production and cellular metabolism, we have generated CLUH-knockout cells using CRISPR/Cas9. Mitochondrial clustering was associated with a smaller cell size and with decreased abundance of respiratory complexes, resulting in oxidative phosphorylation (OXPHOS) defects. This energetic impairment was found to be due to the alteration of mitochondrial translation and to a metabolic shift towards glucose dependency. Metabolomic profiling by mass spectroscopy revealed an increase in the concentration of some amino acids, indicating a dysfunctional Krebs cycle, and increased palmitoylcarnitine concentration, indicating an alteration of fatty acid oxidation, and a dramatic decrease in the concentrations of phosphatidylcholine and sphingomyeline, consistent with the decreased cell size. Taken together, our study establishes a clear function for CLUH in coupling mitochondrial distribution to the control of cell energetic and metabolic status. © 2017. Published by The Company of Biologists Ltd.

Screen for mitochondrial DNA copy number maintenance genes reveals essential role for ATP synthase

PubMed Central

Fukuoh, Atsushi; Cannino, Giuseppe; Gerards, Mike; Buckley, Suzanne; Kazancioglu, Selena; Scialo, Filippo; Lihavainen, Eero; Ribeiro, Andre; Dufour, Eric; Jacobs, Howard T

2014-01-01

The machinery of mitochondrial DNA (mtDNA) maintenance is only partially characterized and is of wide interest due to its involvement in disease. To identify novel components of this machinery, plus other cellular pathways required for mtDNA viability, we implemented a genome-wide RNAi screen in Drosophila S2 cells, assaying for loss of fluorescence of mtDNA nucleoids stained with the DNA-intercalating agent PicoGreen. In addition to previously characterized components of the mtDNA replication and transcription machineries, positives included many proteins of the cytosolic proteasome and ribosome (but not the mitoribosome), three proteins involved in vesicle transport, some other factors involved in mitochondrial biogenesis or nuclear gene expression, > 30 mainly uncharacterized proteins and most subunits of ATP synthase (but no other OXPHOS complex). ATP synthase knockdown precipitated a burst of mitochondrial ROS production, followed by copy number depletion involving increased mitochondrial turnover, not dependent on the canonical autophagy machinery. Our findings will inform future studies of the apparatus and regulation of mtDNA maintenance, and the role of mitochondrial bioenergetics and signaling in modulating mtDNA copy number. PMID:24952591

Determine the Role of Canonical Wnt Signaling in Ovarian Tumorigenesis

DTIC Science & Technology

2015-12-01

studies have shown that oxphos is increased during OIS, leading to an increase in oxygen consumption (Fig. 1).38,42,52 This is likely due to increased TCA...senescent cells display an increase in oxygen consumption but no appreciable increase in ATP levels. The mechanism by which increased fatty acid...benign ovarian tumors into invasive EOCs, and to investigate the effects of inhibition of the canonical Wnt signaling on malignant behavior of EOC cells

Acquired deficiency of tafazzin in the adult heart: Impact on mitochondrial function and response to cardiac injury.

PubMed

Szczepanek, Karol; Allegood, Jeremy; Aluri, Hema; Hu, Ying; Chen, Qun; Lesnefsky, Edward J

2016-04-01

The content and composition of cardiolipin (CL) is critical for preservation of mitochondrial oxidative phosphorylation (OXPHOS) and inner membrane integrity. Tafazzin (Taz) is an enzyme responsible for remodeling of immature CL containing mixed acyl groups into the mature tetralinoleyl form (C18:2)4-CL. We hypothesized that acquired defects in Taz in the mature heart would impact remodeling of CL and augment cardiac injury. The role of acquired Taz deficiency was studied using the inducible Taz knockdown (TazKD) mouse. Taz-specific shRNA is induced by doxycycline (DOX). One day of DOX intake decreased Taz mRNA in the heart to 20% vs. DOX-treated WT. Knockdown was initiated at an adult age and was stable during long term feeding. CL phenotype was assessed by (C18:2)4-CL content and was reduced 40% vs. WT at two months of DOX. TazKD showed increased production of reactive oxygen species and increased susceptibility to permeability transition pore opening at baseline. However, OXPHOS measured using the rate of oxygen consumption was unchanged in the setting of acquired Taz deficiency. Infarct size, measured in isolated buffer-perfused Langendorff hearts following 25min. Stop flow ischemia and 60min. Reperfusion was not altered in TazKD hearts. Thus, impaired Taz-function with onset at adult age does not enhance susceptibility to ischemia-reperfusion injury. Published by Elsevier B.V.

Linking mode of action of the model respiratory and photosynthesis uncoupler 3,5-dichlorophenol to adverse outcomes in Lemna minor.

PubMed

Xie, Li; Gomes, Tânia; Solhaug, Knut Asbjørn; Song, You; Tollefsen, Knut Erik

2018-04-01

Standard chemical toxicity testing guidelines using aquatic plant Lemna minor have been developed by several international standardisation organisations. Although being highly useful for regulatory purposes by focusing on traditional adverse endpoints, these tests provide limited information about the toxic mechanisms and modes of action (MoA). The present study aimed to use selected functional assays in L. minor after exposure to 3,5-dichlorophenol (3,5-DCP) as a model to characterise the toxic mechanisms causing growth inhibition and lethality in primary producers. The results demonstrated that 3,5-DCP caused concentration-dependent effects in chloroplasts and mitochondria. Uncoupling of oxidative phosphorylation (OXPHOS), reduction in chlorophyll (Chlorophyll a and b) content, reproduction rate and frond size were the most sensitive endpoints, followed by formation of reactive oxygen species (ROS), lipid peroxidation (LPO), reduction of carotenoid content and impairment of photosynthesis efficiency. Suppression of photosystem II (PSII) efficiency, electron transport rate (ETR), chlorophyll (a and b) contents and oxidative phosphorylation (OXPHOS) were closely correlated while ROS production and LPO were negative correlated with ETR, carotenoid content and growth parameters. A network of conceptual Adverse Outcome Pathways (AOPs) was developed to decipher the causal relationships between molecular, cellular, and apical adverse effects occurring in L. minor to form a basis for future studies with similar compounds. Copyright © 2018 Elsevier B.V. All rights reserved.

Hypoxia-induced IL-32β increases glycolysis in breast cancer cells.

PubMed

Park, Jeong Su; Lee, Sunyi; Jeong, Ae Lee; Han, Sora; Ka, Hye In; Lim, Jong-Seok; Lee, Myung Sok; Yoon, Do-Young; Lee, Jeong-Hyung; Yang, Young

2015-01-28

IL-32β is highly expressed and increases the migration and invasion of gastric, lung, and breast cancer cells. Since IL-32 enhances VEGF production under hypoxic conditions, whether IL-32β is regulated by hypoxia was examined. Hypoxic conditions and a mimetic chemical CoCl2 enhanced IL-32β production. When cells were treated with various inhibitors of ROS generation to prevent hypoxia-induced ROS function, IL-32β production was suppressed by both NADPH oxidase and mitochondrial ROS inhibitors. IL-32β translocated to the mitochondria under hypoxic conditions, where it was associated with mitochondrial biogenesis. Thus, whether hypoxia-induced IL-32β is associated with oxidative phosphorylation (OXPHOS) or glycolysis was examined. Glycolysis under aerobic and anaerobic conditions is impaired in IL-32β-depleted cells, and the hypoxia-induced IL-32β increased glycolysis through activation of lactate dehydrogenase. Src is also known to increase lactate dehydrogenase activity, and the hypoxia-induced IL-32β was found to stimulate Src activation by inhibiting the dephosphorylation of Src. These findings revealed that a hypoxia-ROS-IL-32β-Src-glycolysis pathway is associated with the regulation of cancer cell metabolism. Copyright © 2014 Elsevier Ireland Ltd. All rights reserved.

Maintained peak leg and pulmonary VO2 despite substantial reduction in muscle mitochondrial capacity.

PubMed

Boushel, R; Gnaiger, E; Larsen, F J; Helge, J W; González-Alonso, J; Ara, I; Munch-Andersen, T; van Hall, G; Søndergaard, H; Saltin, B; Calbet, J A L

2015-12-01

We recently reported the circulatory and muscle oxidative capacities of the arm after prolonged low-intensity skiing in the arctic (Boushel et al., 2014). In the present study, leg VO2 was measured by the Fick method during leg cycling while muscle mitochondrial capacity was examined on a biopsy of the vastus lateralis in healthy volunteers (7 male, 2 female) before and after 42 days of skiing at 60% HR max. Peak pulmonary VO2 (3.52 ± 0.18 L.min(-1) pre vs 3.52 ± 0.19 post) and VO2 across the leg (2.8 ± 0.4L.min(-1) pre vs 3.0 ± 0.2 post) were unchanged after the ski journey. Peak leg O2 delivery (3.6 ± 0.2 L.min(-1) pre vs 3.8 ± 0.4 post), O2 extraction (82 ± 1% pre vs 83 ± 1 post), and muscle capillaries per mm(2) (576 ± 17 pre vs 612 ± 28 post) were also unchanged; however, leg muscle mitochondrial OXPHOS capacity was reduced (90 ± 3 pmol.sec(-1) .mg(-1) pre vs 70 ± 2 post, P < 0.05) as was citrate synthase activity (40 ± 3 μmol.min(-1) .g(-1) pre vs 34 ± 3 vs P < 0.05). These findings indicate that peak muscle VO2 can be sustained with a substantial reduction in mitochondrial OXPHOS capacity. This is achieved at a similar O2 delivery and a higher relative ADP-stimulated mitochondrial respiration at a higher mitochondrial p50. These findings support the concept that muscle mitochondrial respiration is submaximal at VO2max , and that mitochondrial volume can be downregulated by chronic energy demand. © 2015 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

Intrinsic and extrinsic uncoupling of oxidative phosphorylation.

PubMed

Kadenbach, Bernhard

2003-06-05

This article reviews parameters of extrinsic uncoupling of oxidative phosphorylation (OxPhos) in mitochondria, based on induction of a proton leak across the inner membrane. The effects of classical uncouplers, fatty acids, uncoupling proteins (UCP1-UCP5) and thyroid hormones on the efficiency of OxPhos are described. Furthermore, the present knowledge on intrinsic uncoupling of cytochrome c oxidase (decrease of H(+)/e(-) stoichiometry=slip) is reviewed. Among the three proton pumps of the respiratory chain of mitochondria and bacteria, only cytochrome c oxidase is known to exhibit a slip of proton pumping. Intrinsic uncoupling was shown after chemical modification, by site-directed mutagenesis of the bacterial enzyme, at high membrane potential DeltaPsi, and in a tissue-specific manner to increase thermogenesis in heart and skeletal muscle by high ATP/ADP ratios, and in non-skeletal muscle tissues by palmitate. In addition, two mechanisms of respiratory control are described. The first occurs through the membrane potential DeltaPsi and maintains high DeltaPsi values (150-200 mV). The second occurs only in mitochondria, is suggested to keep DeltaPsi at low levels (100-150 mV) through the potential dependence of the ATP synthase and the allosteric ATP inhibition of cytochrome c oxidase at high ATP/ADP ratios, and is reversibly switched on by cAMP-dependent phosphorylation. Finally, the regulation of DeltaPsi and the production of reactive oxygen species (ROS) in mitochondria at high DeltaPsi values (150-200 mV) are discussed.

Increased oxidative phosphorylation in response to acute and chronic DNA damage

PubMed Central

Brace, Lear E; Vose, Sarah C; Stanya, Kristopher; Gathungu, Rose M; Marur, Vasant R; Longchamp, Alban; Treviño-Villarreal, Humberto; Mejia, Pedro; Vargas, Dorathy; Inouye, Karen; Bronson, Roderick T; Lee, Chih-Hao; Neilan, Edward; Kristal, Bruce S; Mitchell, James R

2016-01-01

Accumulation of DNA damage is intricately linked to aging, aging-related diseases and progeroid syndromes such as Cockayne syndrome (CS). Free radicals from endogenous oxidative energy metabolism can damage DNA, however the potential of acute or chronic DNA damage to modulate cellular and/or organismal energy metabolism remains largely unexplored. We modeled chronic endogenous genotoxic stress using a DNA repair-deficient Csa−/−|Xpa−/− mouse model of CS. Exogenous genotoxic stress was modeled in mice in vivo and primary cells in vitro treated with different genotoxins giving rise to diverse spectrums of lesions, including ultraviolet radiation, intrastrand crosslinking agents and ionizing radiation. Both chronic endogenous and acute exogenous genotoxic stress increased mitochondrial fatty acid oxidation (FAO) on the organismal level, manifested by increased oxygen consumption, reduced respiratory exchange ratio, progressive adipose loss and increased FAO in tissues ex vivo. In multiple primary cell types, the metabolic response to different genotoxins manifested as a cell-autonomous increase in oxidative phosphorylation (OXPHOS) subsequent to a transient decline in steady-state NAD+ and ATP levels, and required the DNA damage sensor PARP-1 and energy-sensing kinase AMPK. We conclude that increased FAO/OXPHOS is a general, beneficial, adaptive response to DNA damage on cellular and organismal levels, illustrating a fundamental link between genotoxic stress and energy metabolism driven by the energetic cost of DNA damage. Our study points to therapeutic opportunities to mitigate detrimental effects of DNA damage on primary cells in the context of radio/chemotherapy or progeroid syndromes. PMID:28721274

Oxidative phosphorylation versus glycolysis: what fuel do spermatozoa use?

PubMed Central

du Plessis, Stefan S; Agarwal, Ashok; Mohanty, Gayatri; van der Linde, Michelle

2015-01-01

Spermatozoa are highly specialized cells. Adenosine triphosphate (ATP), which provides the energy for supporting the key functions of the spermatozoa, is formed by 2 metabolic pathways, namely glycolysis and oxidative phosphorylation (OXPHOS). It is produced in the mitochondria through OXPHOS as well as in the head and principal piece of the flagellum through glycolysis. However, there is a great discrepancy as to which method of ATP production is primarily utilized by the spermatozoa for successful fertilization. Mitochondrial respiration is considered to be a more efficient metabolic process for ATP synthesis in comparison to glycolysis. However, studies have shown that the diffusion potential of ATP from the mitochondria to the distal end of the flagellum is not sufficient to support sperm motility, suggesting that glycolysis in the tail region is the preferred pathway for energy production. It is suggested by many investigators that although glycolysis forms the major source of ATP along the flagellum, energy required for sperm motility is mainly produced during mitochondrial respiration. Nevertheless, some studies have shown that when glycolysis is inhibited, proper functioning and motility of spermatozoa remains intact although it is unclear whether such motility can be sustained for prolonged periods of time, or is sufficiently vigorous to achieve optimal fertilization. The purpose of this article is to provide an overview of mammalian sperm energy metabolism and identify the preferred metabolic pathway for ATP generation which forms the basis of energy production in human spermatozoa during fertilization. PMID:25475660

Oxidative phosphorylation versus glycolysis: what fuel do spermatozoa use?

PubMed

du Plessis, Stefan S; Agarwal, Ashok; Mohanty, Gayatri; van der Linde, Michelle

2015-01-01

Spermatozoa are highly specialized cells. Adenosine triphosphate (ATP), which provides the energy for supporting the key functions of the spermatozoa, is formed by 2 metabolic pathways, namely glycolysis and oxidative phosphorylation (OXPHOS). It is produced in the mitochondria through OXPHOS as well as in the head and principal piece of the flagellum through glycolysis. However, there is a great discrepancy as to which method of ATP production is primarily utilized by the spermatozoa for successful fertilization. Mitochondrial respiration is considered to be a more efficient metabolic process for ATP synthesis in comparison to glycolysis. However, studies have shown that the diffusion potential of ATP from the mitochondria to the distal end of the flagellum is not sufficient to support sperm motility, suggesting that glycolysis in the tail region is the preferred pathway for energy production. It is suggested by many investigators that although glycolysis forms the major source of ATP along the flagellum, energy required for sperm motility is mainly produced during mitochondrial respiration. Nevertheless, some studies have shown that when glycolysis is inhibited, proper functioning and motility of spermatozoa remains intact although it is unclear whether such motility can be sustained for prolonged periods of time, or is sufficiently vigorous to achieve optimal fertilization. The purpose of this article is to provide an overview of mammalian sperm energy metabolism and identify the preferred metabolic pathway for ATP generation which forms the basis of energy production in human spermatozoa during fertilization.

Metabolic profiles of exercise in patients with McArdle disease or mitochondrial myopathy

PubMed Central

Sharma, Rohit; Tadvalkar, Laura; Clish, Clary B.; Haller, Ronald G.; Mootha, Vamsi K.

2017-01-01

McArdle disease and mitochondrial myopathy impair muscle oxidative phosphorylation (OXPHOS) by distinct mechanisms: the former by restricting oxidative substrate availability caused by blocked glycogen breakdown, the latter because of intrinsic respiratory chain defects. We applied metabolic profiling to systematically interrogate these disorders at rest, when muscle symptoms are typically minimal, and with exercise, when symptoms of premature fatigue and potential muscle injury are unmasked. At rest, patients with mitochondrial disease exhibit elevated lactate and reduced uridine; in McArdle disease purine nucleotide metabolites, including xanthine, hypoxanthine, and inosine are elevated. During exercise, glycolytic intermediates, TCA cycle intermediates, and pantothenate expand dramatically in both mitochondrial disease and control subjects. In contrast, in McArdle disease, these metabolites remain unchanged from rest; but urea cycle intermediates are increased, likely attributable to increased ammonia production as a result of exaggerated purine degradation. Our results establish skeletal muscle glycogen as the source of TCA cycle expansion that normally accompanies exercise and imply that impaired TCA cycle flux is a central mechanism of restricted oxidative capacity in this disorder. Finally, we report that resting levels of long-chain triacylglycerols in mitochondrial myopathy correlate with the severity of OXPHOS dysfunction, as indicated by the level of impaired O2 extraction from arterial blood during peak exercise. Our integrated analysis of exercise and metabolism provides unique insights into the biochemical basis of these muscle oxidative defects, with potential implications for their clinical management. PMID:28716914

Correlated FLIM and PLIM for cell metabolism

NASA Astrophysics Data System (ADS)

Rück, A.; Breymayer, J.; Kalinina, S.

2016-03-01

Correlated imaging of phosphorescence and fluorescence lifetime parameters of metabolic markers is a challenge for direct investigating mechanisms related to cell metabolism and oxygen tension. A large variety of clinical phenotypes is associated with mitochondrial defects accomplished with changes in cell metabolism. In many cases the hypoxic microenvironment of cancer cells shifts metabolism from oxidative phosphorylation (OXPHOS) to anaerobic or aerobic glycolysis, a process known as "Warburg" effect. Also during stem cell differentiation a switch in cell metabolism is observed. A defective mitochondrial function associated with hypoxia has been invoked in many complex disorders such as type 2 diabetes, Alzheimers disease, cardiac ischemia/reperfusion injury, tissue inflammation and cancer. Cellular responses to oxygen tension have been studied extensively, optical imaging techniques based on time correlated single photon counting (TCSPC) to detect the underlying metabolic mechanisms are therefore of prominent interest. They offer the possibility by inspecting fluorescence decay characteristics of intrinsic coenzymes to directly image metabolic pathways. Moreover oxygen tension can be determined by considering the phosphorescence lifetime of a phosphorescent probe. The combination of both fluorescence lifetime imaging (FLIM) of coenzymes like NADH and FAD and phosphorescence lifetime (PLIM) of phosphorescent dyes could provide valuable information about correlation of metabolic pathways and oxygen tension.

Alpha-lipoic acid attenuates endoplasmic reticulum stress-induced insulin resistance by improving mitochondrial function in HepG2 cells.

PubMed

Lei, Lin; Zhu, Yiwei; Gao, Wenwen; Du, Xiliang; Zhang, Min; Peng, Zhicheng; Fu, Shoupeng; Li, Xiaobing; Zhe, Wang; Li, Xinwei; Liu, Guowen

2016-10-01

Alpha-lipoic acid (ALA) has been reported to have beneficial effects for improving insulin sensitivity. However, the underlying molecular mechanism of the beneficial effects remains poorly understood. Endoplasmic reticulum (ER) stress and mitochondrial dysfunction are considered causal factors that induce insulin resistance. In this study, we investigated the effect of ALA on the modulation of insulin resistance in ER-stressed HepG2 cells, and we explored the potential mechanism of this effect. HepG2 cells were incubated with tunicamycin (Tun) for 6h to establish an ER stress cell model. Tun treatment induced ER stress, mitochondrial dysfunction and insulin resistance. Interestingly, ALA had no significant effect on ER stress signals. Pretreatment of the ER stress cell model with ALA for 24h improved insulin sensitivity, restored the expression levels of mitochondrial oxidative phosphorylation (OXPHOS) complexes and increased intracellular ATP production. Moreover, ALA augmented the β-oxidation capacity of the mitochondria. Importantly, ALA treatment could decrease oligomycin-induced mitochondrial dysfunction and then improved insulin resistance. Taken together, our data suggest that ALA prevents ER stress-induced insulin resistance by enhancing mitochondrial function. Copyright © 2016 Elsevier Inc. All rights reserved.

Drug Discovery & Development: State-of-the-Art and Future Directions” on the topic of “Targets”

PubMed Central

Cook, Gregory M.; Hards, Kiel; Dunn, Elyse; Heikal, Adam; Nakatani, Yoshio; Greening, Chris; Crick, Dean C.; Fontes, Fabio L.; Pethe, Kevin; Hasenoehrl, Erik; Berney, Michael

2017-01-01

Chapter summary The emergence and spread of drug-resistant pathogens, and our inability to develop new antimicrobials to combat resistance, has inspired scientists to seek out new targets for drug development. The Mycobacterium tuberculosis complex is a group of obligately aerobic bacteria that have specialized for inhabiting a wide range of intracellular and extracellular environments. Two fundamental features in this adaptation are the flexible utilization of energy sources and continued metabolism in the absence of growth. M. tuberculosis is an obligately aerobic heterotroph that depends on oxidative phosphorylation (OXPHOS) for growth and survival. However, several studies are redefining the metabolic breadth of the genus. Alternative electron donors and acceptors may provide the maintenance energy for the pathogen to maintain viability in hypoxic, nonreplicating states relevant to latent infection. This hidden metabolic flexibility may ultimately decrease the efficacy of drugs targeted against primary dehydrogenases and terminal oxidases. However, it may also open up opportunities to develop novel antimycobacterials targeting persister cells. In this review, we discuss the progress in understanding the role of energetic targets in mycobacterial physiology and pathogenesis, and the opportunities for drug discovery. PMID:28597820

Primary coenzyme Q10 (CoQ 10) deficiencies and related nephropathies.

PubMed

Ozaltin, Fatih

2014-06-01

Oxidative phosphorylation (OXPHOS) is a metabolic pathway that uses energy released by the oxidation of nutrients to generate adenosine triphosphate (ATP). Coenzyme Q10 (CoQ10), also known as ubiquinone, plays an essential role in the human body not only by generating ATP in the mitochondrial respiratory chain but also by providing protection from reactive oxygen species (ROS) and functioning in the activation of many mitochondrial dehydrogenases and enzymes required in pyrimidine nucleoside biosynthesis. The presentations of primary CoQ10 deficiencies caused by genetic mutations are very heterogeneous. The phenotypes related to energy depletion or ROS production may depend on the content of CoQ10 in the cell, which is determined by the severity of the mutation. Primary CoQ10 deficiency is unique among mitochondrial disorders because early supplementation with CoQ10 can prevent the onset of neurological and renal manifestations. In this review I summarize primary CoQ10 deficiencies caused by various genetic abnormalities, emphasizing its nephropathic form.

The accumulation of assembly intermediates of the mitochondrial complex I matrix arm is reduced by limiting glucose uptake in a neuronal-like model of MELAS syndrome.

PubMed

Geffroy, Guillaume; Benyahia, Rayane; Frey, Samuel; Desquiret-Dumas, Valerie; Gueguen, Naig; Bris, Celine; Belal, Sophie; Inisan, Aurore; Renaud, Aurelie; Chevrollier, Arnaud; Henrion, Daniel; Bonneau, Dominique; Letournel, Franck; Lenaers, Guy; Reynier, Pascal; Procaccio, Vincent

2018-05-01

Ketogenic diet (KD) which combined carbohydrate restriction and the addition of ketone bodies has emerged as an alternative metabolic intervention used as an anticonvulsant therapy or to treat different types of neurological or mitochondrial disorders including MELAS syndrome. MELAS syndrome is a severe mitochondrial disease mainly due to the m.3243A > G mitochondrial DNA mutation. The broad success of KD is due to multiple beneficial mechanisms with distinct effects of very low carbohydrates and ketones. To evaluate the metabolic part of carbohydrate restriction, transmitochondrial neuronal-like cybrid cells carrying the m.3243A > G mutation, shown to be associated with a severe complex I deficiency was exposed during 3 weeks to glucose restriction. Mitochondrial enzyme defects were combined with an accumulation of complex I (CI) matrix intermediates in the untreated mutant cells, leading to a drastic reduction in CI driven respiration. The severe reduction of CI was also paralleled in post-mortem brain tissue of a MELAS patient carrying high mutant load. Importantly, lowering significantly glucose concentration in cell culture improved CI assembly with a significant reduction of matrix assembly intermediates and respiration capacities were restored in a sequential manner. In addition, OXPHOS protein expression and mitochondrial DNA copy number were significantly increased in mutant cells exposed to glucose restriction. The accumulation of CI matrix intermediates appeared as a hallmark of MELAS pathophysiology highlighting a critical pathophysiological mechanism involving CI disassembly, which can be alleviated by lowering glucose fuelling and the induction of mitochondrial biogenesis, emphasizing the usefulness of metabolic interventions in MELAS syndrome. Copyright © 2018 Elsevier B.V. All rights reserved.

Mitochondrial pyruvate carrier function is negatively linked to Warburg phenotype in vitro and malignant features in esophageal squamous cell carcinomas

PubMed Central

Li, Yaqing; Li, Xiaoran; Kan, Quancheng; Zhang, Mingzhi; Li, Xiaoli; Xu, Ruiping; Wang, Junsheng; Yu, Dandan; Goscinski, Mariusz Adam; Wen, Jian-Guo; Nesland, Jahn M.; Suo, Zhenhe

2017-01-01

Aerobic glycolysis is one of the emerging hallmarks of cancer cells. In this study, we investigated the relationship between blocking mitochondrial pyruvate carrier (MPC) with MPC blocker UK5099 and the metabolic alteration as well as aggressive features of esophageal squamous carcinoma. It was found that blocking pyruvate transportation into mitochondria attenuated mitochondrial oxidative phosphorylation (OXPHOS) and triggered aerobic glycolysis, a feature of Warburg effect. In addition, the HIF-1α expression and ROS production were also activated upon UK5099 application. It was further revealed that the UK5099-treated cells became significantly more resistant to chemotherapy and radiotherapy, and the UK5099-treated tumor cells also exhibited stronger invasive capacity compared to the parental cells. In contrast to esophageal squamous epithelium cells, decreased MPC protein expression was observed in a series of 157 human squamous cell carcinomas, and low/negative MPC1 expression predicted an unfavorable clinical outcome. All these results together revealed the potential connection of altered MPC expression/activity with the Warburg metabolic reprogramming and tumor aggressiveness in cell lines and clinical samples. Collectively, our findings highlighted a therapeutic strategy targeting Warburg reprogramming of human esophageal squamous cell carcinomas. PMID:27911865

Naringin Ameliorates HIV-1 Nucleoside Reverse Transcriptase Inhibitors- Induced Mitochondrial Toxicity.

PubMed

Oluwafeyisetan, Adebiyi; Olubunmi, Adebiyi; Peter, Owira

2016-01-01

Mitochondrial reactive oxygen species (ROS) generation and defective oxidative phosphorylation (OXPHOS) have been proposed as possible mechanisms underlying the development of nucleoside reverse transcriptase inhibitors (NRTIs)-induced mitochondrial toxicities. Available options in managing these complications have, so far, produced controversial results, thus necessitating further research into newer agents with promise. Antioxidant and free-radical scavenging effects of naringin, a plant-derived flavonoid, have previously been demonstrated. This study was designed to investigate the effects of naringin on NRTIs-induced mitochondrial toxicity. Wistar rats were randomly divided into Zidovudine (AZT)-only (100 mg/kg body weight BW); AZT+Naringin (100+50 mg/kg BW); AZT+Vitamin E (100+100 mg/kg BW); Stavudine (d4T)- only (50 mg/kg BW); d4T+Naringin (50+50 mg/kg BW); d4T+Vitamin E (50+100 mg/kg BW) and Vehicle (3.0 mL/kg BW)-treated groups, respectively. After 56 days of oral daily dosing, rats were euthanized by halothane overdose, blood collected by cardiac puncture and livers promptly excised for further biochemical and ultrastructural analyses. </p> Results: AZT- or d4T-only caused significant mitochondrial dysfunction and mitochondrial ultrastructural damage compared to controls, while either naringin or vitamin E reversed indices of mitochondrial dysfunction evidenced by significantly reduced mitochondrial malondialdehyde (MDA) and blood lactate concentrations, increased liver manganese superoxide dismutase (MnSOD) activity and upregulate expression of mitochondrial-encoded subunit of electron transport chain (ETC) complex IV protein compared to AZT- or d4T-only treated rats. Furthermore, naringin or vitamin E, respectively, ameliorated mitochondrial damage observed in AZT- or d4T-only treated rats. Naringin ameliorated oxidative stress and NRTI-induced mitochondrial damage and might, therefore, be beneficial in managing toxicities and complications arising from NRTI use.

A novel mutation MT-COIII m.9267G>C and MT-COI m.5913G>A mutation in mitochondrial genes in a Tunisian family with maternally inherited diabetes and deafness (MIDD) associated with severe nephropathy.

PubMed

Tabebi, Mouna; Mkaouar-Rebai, Emna; Mnif, Mouna; Kallabi, Fakhri; Ben Mahmoud, Afif; Ben Saad, Wafa; Charfi, Nadia; Keskes-Ammar, Leila; Kamoun, Hassen; Abid, Mohamed; Fakhfakh, Faiza

2015-04-10

Mitochondrial diabetes (MD) is a heterogeneous disorder characterized by a chronic hyperglycemia, maternal transmission and its association with a bilateral hearing impairment. Several studies reported mutations in mitochondrial genes as potentially pathogenic for diabetes, since mitochondrial oxidative phosphorylation plays an important role in glucose-stimulated insulin secretion from beta cells. In the present report, we studied a Tunisian family with mitochondrial diabetes (MD) and deafness associated with nephropathy. The mutational analysis screening revealed the presence of a novel heteroplasmic mutation m.9276G>C in the mitochondrial COIII gene, detected in mtDNA extracted from leukocytes of a mother and her two daughters indicating that this mutation is maternally transmitted and suggest its implication in the observed phenotype. Bioinformatic tools showed that m.9267G>C mutation (p.A21P) is « deleterious » and it can modify the function and the stability of the MT-COIII protein by affecting the assembly of mitochondrial COX subunits and the translocation of protons then reducing the activity of the respective OXPHOS complexes of ATP synthesis. The nonsynonymous mutation (p.A21P) has not been reported before, it is the first mutation described in the COXIII gene which is related to insulin dependent mitochondrial diabetes and deafness and could be specific to the Tunisian population. The m.9267G>C mutation was present with a nonsynonymous inherited mitochondrial homoplasmic variation MT-COI m.5913 G>A (D4N) responsible of high blood pressure, a clinical feature detected in all explored patients. Copyright © 2015. Published by Elsevier Inc.

Low-dose rapamycin extends lifespan in a mouse model of mtDNA depletion syndrome

PubMed Central

Siegmund, Stephanie E; Yang, Hua; Sharma, Rohit; Javors, Martin; Skinner, Owen; Mootha, Vamsi; Hirano, Michio; Schon, Eric A

2017-01-01

Abstract Mitochondrial disorders affecting oxidative phosphorylation (OxPhos) are caused by mutations in both the nuclear and mitochondrial genomes. One promising candidate for treatment is the drug rapamycin, which has been shown to extend lifespan in multiple animal models, and which was previously shown to ameliorate mitochondrial disease in a knock-out mouse model lacking a nuclear-encoded gene specifying an OxPhos structural subunit (Ndufs4). In that model, relatively high-dose intraperitoneal rapamycin extended lifespan and improved markers of neurological disease, via an unknown mechanism. Here, we administered low-dose oral rapamycin to a knock-in (KI) mouse model of authentic mtDNA disease, specifically, progressive mtDNA depletion syndrome, resulting from a mutation in the mitochondrial nucleotide salvage enzyme thymidine kinase 2 (TK2). Importantly, low-dose oral rapamycin was sufficient to extend Tk2KI/KI mouse lifespan significantly, and did so in the absence of detectable improvements in mitochondrial dysfunction. We found no evidence that rapamycin increased survival by acting through canonical pathways, including mitochondrial autophagy. However, transcriptomics and metabolomics analyses uncovered systemic metabolic changes pointing to a potential ‘rapamycin metabolic signature.’ These changes also implied that rapamycin may have enabled the Tk2KI/KI mice to utilize alternative energy reserves, and possibly triggered indirect signaling events that modified mortality through developmental reprogramming. From a therapeutic standpoint, our results support the possibility that low-dose rapamycin, while not targeting the underlying mtDNA defect, could represent a crucial therapy for the treatment of mtDNA-driven, and some nuclear DNA-driven, mitochondrial diseases. PMID:28973153

Effects of Calorie Restriction and Fiber Type on Glucose Uptake and Abundance of Electron Transport Chain and Oxidative Phosphorylation Proteins in Single Fibers from Old Rats.

PubMed

Wang, Haiyan; Arias, Edward B; Yu, Carmen S; Verkerke, Anthony R P; Cartee, Gregory D

2017-11-09

Calorie restriction (CR; reducing calorie intake by ~40% below ad libitum) can increase glucose uptake by insulin-stimulated muscle. Because skeletal muscle is comprised of multiple, heterogeneous fiber types, our primary aim was to determine the effects of CR (initiated at 14 weeks old) and fiber type on insulin-stimulated glucose uptake by single fibers of diverse fiber types in 23-26-month-old rats. Isolated epitrochlearis muscles from AL and CR rats were incubated with [3H]-2-deoxyglucose ± insulin. Glucose uptake and fiber type were determined for single fibers dissected from the muscles. We also determined CR-effects on abundance of several key metabolic proteins in single fibers. CR resulted in: (a) significantly (p < .05 to .001) greater glucose uptake by insulin-stimulated type I, IIA, IIB, IIBX, and IIX fibers; (b) significantly (p < .05 to .001) reduced abundance of several mitochondrial electron transport chain (ETC) and oxidative phosphorylation (OxPhos) proteins in type I, IIA, and IIBX but not IIB and IIX fibers; and (c) unaltered hexokinase II abundance in each fiber type. These results demonstrate that CR can enhance glucose uptake in each fiber type of rat skeletal muscle in the absence of upregulation of the abundance of hexokinase II or key mitochondrial ETC and OxPhos proteins. © The Author 2017. Published by Oxford University Press on behalf of The Gerontological Society of America. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.

Variable toxicological response to the loss of OXPHOS through 1-methyl-4-phenylpyridinium-induced mitochondrial damage and anoxia in diverse neural immortal cell lines.

PubMed

Mazzio, Elizabeth A; Soliman, Youssef I; Soliman, Karam F A

2010-12-01

Immortal cell lines are used to investigate various aspects of neurodegeneration. These cells display high glycolytic turnover rate and produce an abundant amounts of lactate. Our previous studies indicate that these cells survive the loss of mitochondrial oxidative phosphorylation (OXPHOS) with ample glucose supply. In the current study, we investigate if cell type (w/variation in basal metabolic rate (MR)), can alter glucose utilization patterns which in turn may affect LC(50) for the mitochondrial toxin 1-methyl-4-phenylpyridinium (MPP(+)) in various cell lines. The data obtained indicate that cell lines MRs examined were generally consistent with the average of species adult body weight where mouse N-2A > rat-PC-12 > human SH-SY5Y. A higher MR was associated with accelerated utilization of glucose and earlier cell death with MPP(+): LC(50) mouse = 294 µM, rat = 695 µM, and human = 5.25 mM at 24 h. Cell death appears to be a function of the velocity by which glucose disappears, leading to the failure of glycolysis and subsequent halt of energy production. Similar effects were also observed at higher plating densities where the demand for glucose is amplified. A time-lapse study of MPP(+) toxicity (0-36 h) in N-2A cells indicates that an anaerobic shift occurs as early as 2 h (evidenced by a rise in lactate), followed by a descent in glucose concentrations at 4 h and exhaustion of glucose supplies at 22 h which was associated with the first detectable sign of cell death. It was also noted that MPP(+) toxicity was not associated with the generation of reactive oxygen species (O (2) (-) , H(2)0(2), and NO(2)) and was not attenuated by adding catalase or superoxide dismutase to the media. On the other hand, MPP(+) toxicity was reversed by providing additional supply of glucose, pyruvate ± mitochondrial monocarboxylate transporter blocker (α-cyano-4-HCA), or pyruvate ± pyruvate dehydrogenase inhibitor (octanoyl-CoA), suggesting that the exclusive anaerobic survival compensates for the loss of OXPHOS by MPP(+). To examine if neuroblastoma were capable of surviving the deprivation of O(2) for 24 h, a range of hypoxia to anoxia was established with various concentrations of dithionite. The data suggest that cell lines examined continue to thrive when incubated with high-glucose media (25 mM). In summary, vulnerability of immortal neuroblastoma cell lines to MPP(+) toxicity is dependent upon glucose concentrations within the media and cell MR, which indirectly dominates the velocity of glucose use and its end point disappearance, leading to cell death by ergogenic failure.

Mitochondrial Oxidative Phosphorylation Protein Levels in Peripheral Blood Mononuclear Cells Correlate with Levels in Subcutaneous Adipose Tissue within Samples Differing by HIV and Lipoatrophy Status

PubMed Central

Gerschenson, Mariana; Chow, Dominic; Libutti, Daniel E.; Willis, John H.; Murray, James; Capaldi, Roderick A.; Marusich, Michael

2008-01-01

Abstract Depletion of mitochondrial DNA (mtDNA) and mtDNA-encoded respiratory chain proteins in subcutaneous (SC) fat from patients with HIV lipoatrophy have clearly demonstrated the role of mitochondrial dysfunction in this syndrome. Research in HIV lipoatrophy, however, has been severely hampered by the lack of a suitable surrogate marker in blood or other easily obtained clinical specimens as fat biopsies are invasive and mtDNA levels in peripheral blood mononuclear cells (PBMC) do not consistently correlate with the disease process. We used a simple, rapid, quantitative 2-site dipstick immunoassay to measure OXPHOS enzymes Complex I (CI) and Complex IV (CIV), and rtPCR to measure mtDNA in 26 matched SC fat and PBMC specimens previously banked from individuals on potent antiretroviral (ARV) therapy with HIV lipoatrophy, on similar ARV therapy without lipoatrophy, and in HIV seronegative controls. Significant correlations were found between the respective PBMC and fat levels for both CI (r = 0.442, p = 0.024) and for CIV (r = 0.507, p = 0.008). Both CI and CIV protein levels were also significantly reduced in both PBMCs and fat in lipoatrophic subjects compared to HIV seronegative controls (p ≤ 0.05), while a comparative reduction in mtDNA levels in lipoatrophic subjects was observed only in fat. We conclude that CI and CIV levels in PBMCs correlate to their respective levels in fat and may have utility as surrogate markers of mitochondrial dysfunction in lipoatrophy. PMID:18844460

Correlated oxygen-sensing PLIM, cell metabolism FLIM and applications

NASA Astrophysics Data System (ADS)

Rück, A. C.; Kalinina, S.; Schäfer, P.; von Einem, B.; von Arnim, C.

2017-02-01

Correlated imaging of phosphorescence and fluorescence lifetime parameters of metabolic markers is a challenge for direct investigating mechanisms related to cell metabolism and oxygen tension. A large variety of clinical phenotypes is associated with mitochondrial defects accomplished with changes in cell metabolism. In many cases the hypoxic microenvironment of cancer cells shifts metabolism from oxidative phosphorylation (OXPHOS) to anaerobic or aerobic glycolysis, a process known as "Warburg" effect. Also during stem cell differentiation a switch in cell metabolism is observed. Mitochondrial dysfunction associated with hypoxia has been invoked in many complex disorders such as type 2 diabetes, Alzheimeŕs disease, cardiac ischemia/reperfusion injury, tissue inflammation and cancer. Cellular responses to oxygen tension have been studied extensively, optical imaging techniques based on time correlated single photon counting (TCSPC) to detect oxygen concentration and distribution are therefore of prominent interest. Moreover, they offer the possibility by inspecting fluorescence decay characteristics of intrinsic coenzymes to directly image metabolic pathways, whereas oxygen tension can be determined by considering the phosphorescence lifetime of a phosphorescent probe. The combination of both fluorescence lifetime imaging (FLIM) of coenzymes like NAD(P)H and FAD and phosphorescence lifetime (PLIM) of phosphorescent dyes could provide valuable information about correlation of metabolic pathways and oxygen tension.

Spatial Distribution of Cellular Function: The Partitioning of Proteins between Mitochondria and the Nucleus in MCF7 Breast Cancer Cells

PubMed Central

Qattan, Amal T.; Radulovic, Marko; Crawford, Mark; Godovac-Zimmermann, Jasminka

2014-01-01

Concurrent proteomics analysis of the nuclei and mitochondria of MCF7 breast cancer cells identified 985 proteins (40% of all detected proteins) present in both organelles. Numerous proteins from all five complexes involved in oxidative phosphorylation (e.g., NDUFA5, NDUFB10, NDUFS1, NDUF2, SDHA, UQRB, UQRC2, UQCRH, COX5A, COX5B, MT-CO2, ATP5A1, ATP5B, ATP5H, etc.), from the TCA-cycle (DLST, IDH2, IDH3A, OGDH, SUCLAG2, etc.), and from glycolysis (ALDOA, ENO1, FBP1, GPI, PGK1, TALDO1, etc.) were distributed to both the nucleus and mitochondria. In contrast, proteins involved in nuclear/mitochondrial RNA processing/translation and Ras/Rab signaling showed different partitioning patterns. The identity of the OxPhos, TCA-cycle, and glycolysis proteins distributed to both the nucleus and mitochondria provides evidence for spatio-functional integration of these processes over the two different subcellular organelles. We suggest that there are unrecognized aspects of functional coordination between the nucleus and mitochondria, that integration of core functional processes via wide subcellular distribution of constituent proteins is a common characteristic of cells, and that subcellular spatial integration of function may be a vital aspect of cancer. PMID:23051583

Maternal Betaine Supplementation during Gestation Enhances Expression of mtDNA-Encoded Genes through D-Loop DNA Hypomethylation in the Skeletal Muscle of Newborn Piglets.

PubMed

Jia, Yimin; Song, Haogang; Gao, Guichao; Cai, Demin; Yang, Xiaojing; Zhao, Ruqian

2015-11-25

Betaine has been widely used in animal and human nutrition to promote muscle growth and performance, yet it remains unknown whether maternal betaine supplementation during gestation affects the metabolic characteristics of neonatal skeletal muscles. In the present study, feeding sows with betaine-supplemented diets throughout gestation significantly upregulated the expression of mtDNA-encoded OXPHOS genes (p < 0.05), including COX1, COX2, and ND5, in the muscle of newborn piglets, which was associated with enhanced mitochondrial COX enzyme activity (p < 0.05). Concurrently, maternal betaine supplementation increased the plasma betaine concentration and muscle expression of methyl transfer enzymes (p < 0.05), BHMT and GNMT, in offspring piglets. Nevertheless, Dnmt3a was downregulated at the level of both mRNA and protein, which was associated with a hypomethylated mtDNA D-loop region (p < 0.05). These results suggest that maternal betaine supplementation during gestation enhances expression of mtDNA-encoded genes through D-loop DNA hypomethylation in the skeletal muscle of newborn piglets.

BCL2 and BCL(X)L selective inhibitors decrease mitochondrial ATP production in breast cancer cells and are synthetically lethal when combined with 2-deoxy-D-glucose.

PubMed

Lucantoni, Federico; Düssmann, Heiko; Llorente-Folch, Irene; Prehn, Jochen H M

2018-05-25

Cancer cells display differences regarding their engagement of glycolytic vs. mitochondrial oxidative phosphorylation (OXPHOS) pathway. Triple negative breast cancer, an aggressive form of breast cancer, is characterized by elevated glycolysis, while estrogen receptor positive breast cancer cells rely predominantly on OXPHOS. BCL2 proteins control the process of mitochondrial outer membrane permeabilization during apoptosis, but also regulate cellular bioenergetics. Because BCL2 proteins are overexpressed in breast cancer and targetable by selective antagonists, we here analysed the effect of BCL2 and BCL(X)L selective inhibitors, Venetoclax and WEHI-539, on mitochondrial bioenergetics and cell death. Employing single cell imaging using a FRET-based mitochondrial ATP sensor, we found that MCF7 breast cancer cells supplied with mitochondrial substrates reduced their mitochondrial ATP production when treated with Venetoclax or WEHI-539 at concentrations that per se did not induce cell death. Treatments with lower concentrations of both inhibitors also reduced the length of the mitochondrial network and the dynamics, as evaluated by quantitative confocal microscopy. We next tested the hypothesis that mitochondrial ATP production inhibition with BCL2 or BCL(X)L antagonists was synthetically lethal when combined with glycolysis inhibition. Treatment with 2-deoxy-D-glucose in combination with Venetoclax or WEHI-539 synergistically reduced the cellular bioenergetics of ER+ and TNBC breast cancer cells and abolished their clonogenic potential. Synthetic lethality was also observed when cultures were grown in 3D spheres. Our findings demonstrate that BCL2 antagonists exert potent effects on cancer metabolism independent of cell death-inducing effects, and demonstrate a synthetic lethality when these are applied in combination with glycolysis inhibitors.

BCL2 and BCL(X)L selective inhibitors decrease mitochondrial ATP production in breast cancer cells and are synthetically lethal when combined with 2-deoxy-D-glucose

PubMed Central

Lucantoni, Federico; Düssmann, Heiko; Llorente-Folch, Irene; Prehn, Jochen H.M.

2018-01-01

Cancer cells display differences regarding their engagement of glycolytic vs. mitochondrial oxidative phosphorylation (OXPHOS) pathway. Triple negative breast cancer, an aggressive form of breast cancer, is characterized by elevated glycolysis, while estrogen receptor positive breast cancer cells rely predominantly on OXPHOS. BCL2 proteins control the process of mitochondrial outer membrane permeabilization during apoptosis, but also regulate cellular bioenergetics. Because BCL2 proteins are overexpressed in breast cancer and targetable by selective antagonists, we here analysed the effect of BCL2 and BCL(X)L selective inhibitors, Venetoclax and WEHI-539, on mitochondrial bioenergetics and cell death. Employing single cell imaging using a FRET-based mitochondrial ATP sensor, we found that MCF7 breast cancer cells supplied with mitochondrial substrates reduced their mitochondrial ATP production when treated with Venetoclax or WEHI-539 at concentrations that per se did not induce cell death. Treatments with lower concentrations of both inhibitors also reduced the length of the mitochondrial network and the dynamics, as evaluated by quantitative confocal microscopy. We next tested the hypothesis that mitochondrial ATP production inhibition with BCL2 or BCL(X)L antagonists was synthetically lethal when combined with glycolysis inhibition. Treatment with 2-deoxy-D-glucose in combination with Venetoclax or WEHI-539 synergistically reduced the cellular bioenergetics of ER+ and TNBC breast cancer cells and abolished their clonogenic potential. Synthetic lethality was also observed when cultures were grown in 3D spheres. Our findings demonstrate that BCL2 antagonists exert potent effects on cancer metabolism independent of cell death-inducing effects, and demonstrate a synthetic lethality when these are applied in combination with glycolysis inhibitors. PMID:29899841

Analysis of the liver mitochondrial proteome in response to ethanol and S-adenosylmethionine treatments: novel molecular targets of disease and hepatoprotection.

PubMed

Andringa, Kelly K; King, Adrienne L; Eccleston, Heather B; Mantena, Sudheer K; Landar, Aimee; Jhala, Nirag C; Dickinson, Dale A; Squadrito, Giuseppe L; Bailey, Shannon M

2010-05-01

S-adenosylmethionine (SAM) minimizes alcohol hepatotoxicity; however, the molecular mechanisms responsible for SAM hepatoprotection remain unknown. Herein, we use proteomics to determine whether the hepatoprotective action of SAM against early-stage alcoholic liver disease is linked to alterations in the mitochondrial proteome. For this, male rats were fed control or ethanol-containing liquid diets +/- SAM and liver mitochondria were prepared for proteomic analysis. Two-dimensional isoelectric focusing (2D IEF/SDS-PAGE) and blue native gel electrophoresis (BN-PAGE) were used to determine changes in matrix and oxidative phosphorylation (OxPhos) proteins, respectively. SAM coadministration minimized alcohol-dependent inflammation and preserved mitochondrial respiration. SAM supplementation preserved liver SAM levels in ethanol-fed rats; however, mitochondrial SAM levels were increased by ethanol and SAM treatments. With use of 2D IEF/SDS-PAGE, 30 proteins showed significant changes in abundance in response to ethanol, SAM, or both. Classes of proteins affected by ethanol and SAM treatments were chaperones, beta oxidation proteins, sulfur metabolism proteins, and dehydrogenase enzymes involved in methionine, glycine, and choline metabolism. BN-PAGE revealed novel changes in the levels of 19 OxPhos proteins in response to ethanol, SAM, or both. Ethanol- and SAM-dependent alterations in the proteome were not linked to corresponding changes in gene expression. In conclusion, ethanol and SAM treatment led to multiple changes in the liver mitochondrial proteome. The protective effects of SAM against alcohol toxicity are mediated, in part, through maintenance of proteins involved in key mitochondrial energy conserving and biosynthetic pathways. This study demonstrates that SAM may be a promising candidate for treatment of alcoholic liver disease.

Effects of Tributyltin Chloride on Cybrids with or without an ATP Synthase Pathologic Mutation

PubMed Central

López-Gallardo, Ester; Llobet, Laura; Emperador, Sonia; Montoya, Julio; Ruiz-Pesini, Eduardo

2016-01-01

Background: The oxidative phosphorylation system (OXPHOS) includes nuclear chromosome (nDNA)– and mitochondrial DNA (mtDNA)–encoded polypeptides. Many rare OXPHOS disorders, such as striatal necrosis syndromes, are caused by genetic mutations. Despite important advances in sequencing procedures, causative mutations remain undetected in some patients. It is possible that etiologic factors, such as environmental toxins, are the cause of these cases. Indeed, the inhibition of a particular enzyme by a poison could imitate the biochemical effects of pathological mutations in that enzyme. Moreover, environmental factors can modify the penetrance or expressivity of pathological mutations. Objectives: We studied the interaction between mitochondrially encoded ATP synthase 6 (p.MT-ATP6) subunit and an environmental exposure that may contribute phenotypic differences between healthy individuals and patients suffering from striatal necrosis syndromes or other mitochondriopathies. Methods: We analyzed the effects of the ATP synthase inhibitor tributyltin chloride (TBTC), a widely distributed environmental factor that contaminates human food and water, on transmitochondrial cell lines with or without an ATP synthase mutation that causes striatal necrosis syndrome. Doses were selected based on TBTC concentrations previously reported in human whole blood samples. Results: TBTC modified the phenotypic effects caused by a pathological mtDNA mutation. Interestingly, wild-type cells treated with this xenobiotic showed similar bioenergetics when compared with the untreated mutated cells. Conclusions: In addition to the known genetic causes, our findings suggest that environmental exposure to TBTC might contribute to the etiology of striatal necrosis syndromes. Citation: López-Gallardo E, Llobet L, Emperador S, Montoya J, Ruiz-Pesini E. 2016. Effects of tributyltin chloride on cybrids with or without an ATP synthase pathologic mutation. Environ Health Perspect 124:1399–1405; http://dx.doi.org/10.1289/EHP182 PMID:27129022

Effects of Tributyltin Chloride on Cybrids with or without an ATP Synthase Pathologic Mutation.

PubMed

López-Gallardo, Ester; Llobet, Laura; Emperador, Sonia; Montoya, Julio; Ruiz-Pesini, Eduardo

2016-09-01

The oxidative phosphorylation system (OXPHOS) includes nuclear chromosome (nDNA)- and mitochondrial DNA (mtDNA)-encoded polypeptides. Many rare OXPHOS disorders, such as striatal necrosis syndromes, are caused by genetic mutations. Despite important advances in sequencing procedures, causative mutations remain undetected in some patients. It is possible that etiologic factors, such as environmental toxins, are the cause of these cases. Indeed, the inhibition of a particular enzyme by a poison could imitate the biochemical effects of pathological mutations in that enzyme. Moreover, environmental factors can modify the penetrance or expressivity of pathological mutations. We studied the interaction between mitochondrially encoded ATP synthase 6 (p.MT-ATP6) subunit and an environmental exposure that may contribute phenotypic differences between healthy individuals and patients suffering from striatal necrosis syndromes or other mitochondriopathies. We analyzed the effects of the ATP synthase inhibitor tributyltin chloride (TBTC), a widely distributed environmental factor that contaminates human food and water, on transmitochondrial cell lines with or without an ATP synthase mutation that causes striatal necrosis syndrome. Doses were selected based on TBTC concentrations previously reported in human whole blood samples. TBTC modified the phenotypic effects caused by a pathological mtDNA mutation. Interestingly, wild-type cells treated with this xenobiotic showed similar bioenergetics when compared with the untreated mutated cells. In addition to the known genetic causes, our findings suggest that environmental exposure to TBTC might contribute to the etiology of striatal necrosis syndromes. López-Gallardo E, Llobet L, Emperador S, Montoya J, Ruiz-Pesini E. 2016. Effects of tributyltin chloride on cybrids with or without an ATP synthase pathologic mutation. Environ Health Perspect 124:1399-1405; http://dx.doi.org/10.1289/EHP182.

Low-dose rapamycin extends lifespan in a mouse model of mtDNA depletion syndrome.

PubMed

Siegmund, Stephanie E; Yang, Hua; Sharma, Rohit; Javors, Martin; Skinner, Owen; Mootha, Vamsi; Hirano, Michio; Schon, Eric A

2017-12-01

Mitochondrial disorders affecting oxidative phosphorylation (OxPhos) are caused by mutations in both the nuclear and mitochondrial genomes. One promising candidate for treatment is the drug rapamycin, which has been shown to extend lifespan in multiple animal models, and which was previously shown to ameliorate mitochondrial disease in a knock-out mouse model lacking a nuclear-encoded gene specifying an OxPhos structural subunit (Ndufs4). In that model, relatively high-dose intraperitoneal rapamycin extended lifespan and improved markers of neurological disease, via an unknown mechanism. Here, we administered low-dose oral rapamycin to a knock-in (KI) mouse model of authentic mtDNA disease, specifically, progressive mtDNA depletion syndrome, resulting from a mutation in the mitochondrial nucleotide salvage enzyme thymidine kinase 2 (TK2). Importantly, low-dose oral rapamycin was sufficient to extend Tk2KI/KI mouse lifespan significantly, and did so in the absence of detectable improvements in mitochondrial dysfunction. We found no evidence that rapamycin increased survival by acting through canonical pathways, including mitochondrial autophagy. However, transcriptomics and metabolomics analyses uncovered systemic metabolic changes pointing to a potential 'rapamycin metabolic signature.' These changes also implied that rapamycin may have enabled the Tk2KI/KI mice to utilize alternative energy reserves, and possibly triggered indirect signaling events that modified mortality through developmental reprogramming. From a therapeutic standpoint, our results support the possibility that low-dose rapamycin, while not targeting the underlying mtDNA defect, could represent a crucial therapy for the treatment of mtDNA-driven, and some nuclear DNA-driven, mitochondrial diseases. © The Author 2017. Published by Oxford University Press.

Mechanisms of Attenuation of Pulmonary V’O2 Slow Component in Humans after Prolonged Endurance Training

PubMed Central

Zoladz, Jerzy A.; Majerczak, Joanna; Grassi, Bruno; Szkutnik, Zbigniew; Korostyński, Michał; Gołda, Sławomir; Grandys, Marcin; Jarmuszkiewicz, Wiesława; Kilarski, Wincenty; Karasinski, Janusz; Korzeniewski, Bernard

2016-01-01

In this study we have examined the effect of prolonged endurance training program on the pulmonary oxygen uptake (V’O2) kinetics during heavy-intensity cycling-exercise and its impact on maximal cycling and running performance. Twelve healthy, physically active men (mean±SD: age 22.33±1.44 years, V’O2peak 3198±458 mL ∙ min-1) performed an endurance training composed mainly of moderate-intensity cycling, lasting 20 weeks. Training resulted in a decrease (by ~5%, P = 0.027) in V’O2 during prior low-intensity exercise (20 W) and in shortening of τp of the V’O2 on-kinetics (30.1±5.9 s vs. 25.4±1.5 s, P = 0.007) during subsequent heavy-intensity cycling. This was accompanied by a decrease of the slow component of V’O2 on-kinetics by 49% (P = 0.001) and a decrease in the end-exercise V’O2 by ~5% (P = 0.005). An increase (P = 0.02) in the vascular endothelial growth factor receptor 2 mRNA level and a tendency (P = 0.06) to higher capillary-to-fiber ratio in the vastus lateralis muscle were found after training (n = 11). No significant effect of training on the V’O2peak was found (P = 0.12). However, the power output reached at the lactate threshold increased by 19% (P = 0.01). The power output obtained at the V’O2peak increased by 14% (P = 0.003) and the time of 1,500-m performance decreased by 5% (P = 0.001). Computer modeling of the skeletal muscle bioenergetic system suggests that the training-induced decrease in the slow component of V’O2 on-kinetics found in the present study is mainly caused by two factors: an intensification of the each-step activation (ESA) of oxidative phosphorylation (OXPHOS) complexes after training and decrease in the ‘‘additional” ATP usage rising gradually during heavy-intensity exercise. PMID:27104346

Evolutionary perspectives on the links between mitochondrial genotype and disease phenotype.

PubMed

Dowling, Damian K

2014-04-01

Disorders of the mitochondrial respiratory chain are heterogeneous in their symptoms and underlying genetics. Simple links between candidate mutations and expression of disease phenotype typically do not exist. It thus remains unclear how the genetic variation in the mitochondrial genome contributes to the phenotypic expression of complex traits and disease phenotypes. I summarize the basic genetic processes known to underpin mitochondrial disease. I highlight other plausible processes, drawn from the evolutionary biological literature, whose contribution to mitochondrial disease expression remains largely empirically unexplored. I highlight recent advances to the field, and discuss common-ground and -goals shared by researchers across medical and evolutionary domains. Mitochondrial genetic variance is linked to phenotypic variance across a variety of traits (e.g. reproductive function, life expectancy) fundamental to the upkeep of good health. Evolutionary theory predicts that mitochondrial genomes are destined to accumulate male-harming (but female-friendly) mutations, and this prediction has received proof-of-principle support. Furthermore, mitochondrial effects on the phenotype are typically manifested via interactions between mitochondrial and nuclear genes. Thus, whether a mitochondrial mutation is pathogenic in effect can depend on the nuclear genotype in which is it expressed. Many disease phenotypes associated with OXPHOS malfunction might be determined by the outcomes of mitochondrial-nuclear interactions, and by the evolutionary forces that historically shaped mitochondrial DNA (mtDNA) sequences. Concepts and results drawn from the evolutionary sciences can have broad, but currently under-utilized, applicability to the medical sciences and provide new insights into understanding the complex genetics of mitochondrial disease. This article is part of a Special Issue entitled Frontiers of Mitochondrial Research. Copyright © 2013. Published by Elsevier B.V.

mtDNA lineage analysis of mouse L-cell lines reveals the accumulation of multiple mtDNA mutants and intermolecular recombination

PubMed Central

Fan, Weiwei; Lin, Chun Shi; Potluri, Prasanth; Procaccio, Vincent; Wallace, Douglas C.

2012-01-01

The role of mitochondrial DNA (mtDNA) mutations and mtDNA recombination in cancer cell proliferation and developmental biology remains controversial. While analyzing the mtDNAs of several mouse L cell lines, we discovered that every cell line harbored multiple mtDNA mutants. These included four missense mutations, two frameshift mutations, and one tRNA homopolymer expansion. The LA9 cell lines lacked wild-type mtDNAs but harbored a heteroplasmic mixture of mtDNAs, each with a different combination of these variants. We isolated each of the mtDNAs in a separate cybrid cell line. This permitted determination of the linkage phase of each mtDNA and its physiological characteristics. All of the polypeptide mutations inhibited their oxidative phosphorylation (OXPHOS) complexes. However, they also increased mitochondrial reactive oxygen species (ROS) production, and the level of ROS production was proportional to the cellular proliferation rate. By comparing the mtDNA haplotypes of the different cell lines, we were able to reconstruct the mtDNA mutational history of the L–L929 cell line. This revealed that every heteroplasmic L-cell line harbored a mtDNA that had been generated by intracellular mtDNA homologous recombination. Therefore, deleterious mtDNA mutations that increase ROS production can provide a proliferative advantage to cancer or stem cells, and optimal combinations of mutant loci can be generated through recombination. PMID:22345519

Leptin Modulates Mitochondrial Function, Dynamics and Biogenesis in MCF-7 Cells.

PubMed

Blanquer-Rosselló, M Mar; Santandreu, Francisca M; Oliver, Jordi; Roca, Pilar; Valle, Adamo

2015-09-01

The adipokine leptin, known for its key role in the control of energy metabolism, has been shown to be involved in both normal and tumoral mammary growth. One of the hallmarks of cancer is an alteration of tumor metabolism since cancerous cells must rewire metabolism to satisfy the demands of growth and proliferation. Considering the sensibility of breast cancer cells to leptin, the objective of this study was to explore the effects of this adipokine on their metabolism. To this aim, we treated the MCF-7 breast cancer cell line with 50 ng/mL leptin and analyzed several features related to cellular and mitochondrial metabolism. As a result, leptin increased cell proliferation, shifted ATP production from glycolysis to mitochondria and decreased the levels of the glycolytic end-product lactate. We observed an improvement in ADP-dependent oxygen consumption and an amelioration of oxidative stress without changes in total mitochondrial mass or specific oxidative phosphorylation (OXPHOS) complexes. Furthermore, RT-PCR and western blot showed an up-regulation for genes and proteins related to biogenesis and mitochondrial dynamics. This expression signature, together with an increased mitophagy observed by confocal microscopy suggests that leptin may improve mitochondrial quality and function. Taken together, our results propose that leptin may improve bioenergetic efficiency by avoiding the production of reactive oxygen species (ROS) and conferring benefits for growth and survival of MCF-7 breast cancer cells. © 2015 Wiley Periodicals, Inc.

Bioenergetics of lung tumors: alteration of mitochondrial biogenesis and respiratory capacity.

PubMed

Bellance, N; Benard, G; Furt, F; Begueret, H; Smolková, K; Passerieux, E; Delage, J P; Baste, J M; Moreau, P; Rossignol, R

2009-12-01

Little is known on the metabolic profile of lung tumors and the reminiscence of embryonic features. Herein, we determined the bioenergetic profiles of human fibroblasts taken from lung epidermoid carcinoma (HLF-a) and fetal lung (MRC5). We also analysed human lung tumors and their surrounding healthy tissue from four patients with adenocarcinoma. On these different models, we measured functional parameters (cell growth rates in oxidative and glycolytic media, respiration, ATP synthesis and PDH activity) as well as compositional features (expression level of various energy proteins and upstream transcription factors). The results demonstrate that both the lung fetal and cancer cell lines produced their ATP predominantly by glycolysis, while oxidative phosphorylation was only capable of poor ATP delivery. This was explained by a decreased mitochondrial biogenesis caused by a lowered expression of PGC1alpha (as shown by RT-PCR and Western blot) and mtTFA. Consequently, the relative expression of glycolytic versus OXPHOS markers was high in these cells. Moreover, the re-activation of mitochondrial biogenesis with resveratrol induced cell death specifically in cancer cells. A consistent reduction of mitochondrial biogenesis and the subsequent alteration of respiratory capacity was also observed in lung tumors, associated with a lower expression level of bcl2. Our data give a better characterization of lung cancer cells' metabolic alterations which are essential for growth and survival. They designate mitochondrial biogenesis as a possible target for anti-cancer therapy.

Mitochondrial function is altered in horse atypical myopathy.

PubMed

Lemieux, Hélène; Boemer, François; van Galen, Gaby; Serteyn, Didier; Amory, Hélène; Baise, Etienne; Cassart, Dominique; van Loon, Gunther; Marcillaud-Pitel, Christel; Votion, Dominique-M

2016-09-01

Equine atypical myopathy in Europe is a fatal rhabdomyolysis syndrome that results from the ingestion of hypoglycin A contained in seeds and seedlings of Acer pseudoplatanus (sycamore maple). Acylcarnitine concentrations in serum and muscle OXPHOS capacity were determined in 15 atypical myopathy cases. All but one acylcarnitine were out of reference range and mitochondrial respiratory capacity was severely decreased up to 49% as compared to 10 healthy controls. The hallmark of atypical myopathy thus consists of a severe alteration in the energy metabolism including a severe impairment in muscle mitochondrial respiration that could contribute to its high death rate. Copyright © 2016 Elsevier B.V. and Mitochondria Research Society. All rights reserved.

Dysregulated autophagy in restrictive cardiomyopathy due to Pro209Leu mutation in BAG3.

PubMed

Schänzer, A; Rupp, S; Gräf, S; Zengeler, D; Jux, C; Akintürk, H; Gulatz, L; Mazhari, N; Acker, T; Van Coster, R; Garvalov, B K; Hahn, A

2018-03-01

Myofibrillary myopathies (MFM) are hereditary myopathies histologically characterized by degeneration of myofibrils and aggregation of proteins in striated muscle. Cardiomyopathy is common in MFM but the pathophysiological mechanisms are not well understood. The BAG3-Pro209Leu mutation is associated with early onset MFM and severe restrictive cardiomyopathy (RCM), often necessitating heart transplantation during childhood. We report on a young male patient with a BAG3-Pro209Leu mutation who underwent heart transplantation at eight years of age. Detailed morphological analyses of the explanted heart tissue showed intracytoplasmic inclusions, aggregation of BAG3 and desmin, disintegration of myofibers and Z-disk alterations. The presence of undegraded autophagosomes, seen by electron microscopy, as well as increased levels of p62, LC3-I and WIPI1, detected by immunohistochemistry and western blot analyses, indicated a dysregulation of autophagy. Parkin and PINK1, proteins involved in mitophagy, were slightly increased whereas mitochondrial OXPHOS activities were not altered. These findings indicate that altered autophagy plays a role in the pathogenesis and rapid progression of RCM in MFM caused by the BAG3-Pro209Leu mutation, which could have implications for future therapeutic strategies. Copyright © 2018 Elsevier Inc. All rights reserved.

Mitochondrial Molecular Abnormalities Revealed by Proteomic Analysis of Hippocampal Organelles of Mice Triple Transgenic for Alzheimer Disease

PubMed Central

Yu, Haitao; Lin, Xuemei; Wang, Dian; Zhang, Zaijun; Guo, Yi; Ren, Xiaohu; Xu, Benhong; Yuan, Jianhui; Liu, Jianjun; Spencer, Peter S.; Wang, Jian-Zhi; Yang, Xifei

2018-01-01

Mitochondrial dysfunction is implicated in the pathogenesis of Alzheimer’s disease (AD). However, the precise mitochondrial molecular deficits in AD remain poorly understood. Mitochondrial and nuclear proteomic analysis in mature male triple transgenic AD mice (PS1M146V/APPSwe/TauP301L) by two-dimensional fluorescence difference gel electrophoresis (2D-DIGE) coupled with MALDI-TOF-MS/MS, bio-informatics analysis and immunofluorescent staining were performed in this study. In addition to impaired spatial memory impairment and intracellular accumulation of amyloid 1–42 (Aβ1–42) in the 3xTg-AD mice, a well-accepted mouse model of the human disease, we also found significantly increased DNA oxidative damage in entorhinal cortex, hippocampal CA1, CA3 and dental gyrus (DG), as evidenced by the positive staining of 8-hydroxyguanosine, a biomarker of mild cognitive impairment early in AD. We identified significant differences in 27 hippocampal mitochondrial proteins (11 increased and 16 decreased), and 37 hippocampal nuclear proteins (12 increased and 25 decreased) in 3xTg-AD mice compared with the wild-type (WT) mice. Differentially expressed mitochondrial and nuclear proteins were mainly involved in energy metabolism (>55%), synapses, DNA damage, apoptosis and oxidative stress. Two proteins were differentially expressed in both hippocampal mitochondria and nuclei, namely electron transport chain (ETC)-related protein ATP synthase subunit d (ATP5H) was significantly decreased, and apoptosis-related dynamin-1 (DYN1), a pre-synaptic and mitochondrial division-regulated protein that was significantly increased. In sum, perturbations of hippocampus mitochondrial energy metabolism-related proteins responsible for ATP generation via oxidation phosphorylation (OXPHOS), especially nuclear-encoded OXPHOS proteins, correlated with the amyloid-associated cognitive deficits of this murine AD model. The molecular changes in respiratory chain-related proteins and DYN1 may represent novel biomarkers of AD. PMID:29593495

Mitochondrial DNA triplication and punctual mutations in patients with mitochondrial neuromuscular disorders

DOE Office of Scientific and Technical Information (OSTI.GOV)

Mkaouar-Rebai, Emna, E-mail: emna.mkaouar@gmail.com; Felhi, Rahma; Tabebi, Mouna

Mitochondrial diseases are a heterogeneous group of disorders caused by the impairment of the mitochondrial oxidative phosphorylation system which have been associated with various mutations of the mitochondrial DNA (mtDNA) and nuclear gene mutations. The clinical phenotypes are very diverse and the spectrum is still expanding. As brain and muscle are highly dependent on OXPHOS, consequently, neurological disorders and myopathy are common features of mtDNA mutations. Mutations in mtDNA can be classified into three categories: large-scale rearrangements, point mutations in tRNA or rRNA genes and point mutations in protein coding genes. In the present report, we screened mitochondrial genes ofmore » complex I, III, IV and V in 2 patients with mitochondrial neuromuscular disorders. The results showed the presence the pathogenic heteroplasmic m.9157G>A variation (A211T) in the MT-ATP6 gene in the first patient. We also reported the first case of triplication of 9 bp in the mitochondrial NC7 region in Africa and Tunisia, in association with the novel m.14924T>C in the MT-CYB gene in the second patient with mitochondrial neuromuscular disorder. - Highlights: • We reported 2 patients with mitochondrial neuromuscular disorders. • The heteroplasmic MT-ATP6 9157G>A variation was reported. • A triplication of 9 bp in the mitochondrial NC7 region was detected. • The m.14924T>C transition (S60P) in the MT-CYB gene was found.« less

Oncogenes induce the cancer-associated fibroblast phenotype

PubMed Central

Lisanti, Michael P; Martinez-Outschoorn, Ubaldo E; Sotgia, Federica

2013-01-01

Metabolic coupling, between mitochondria in cancer cells and catabolism in stromal fibroblasts, promotes tumor growth, recurrence, metastasis, and predicts anticancer drug resistance. Catabolic fibroblasts donate the necessary fuels (such as L-lactate, ketones, glutamine, other amino acids, and fatty acids) to anabolic cancer cells, to metabolize via their TCA cycle and oxidative phosphorylation (OXPHOS). This provides a simple mechanism by which metabolic energy and biomass are transferred from the host microenvironment to cancer cells. Recently, we showed that catabolic metabolism and “glycolytic reprogramming” in the tumor microenvironment are orchestrated by oncogene activation and inflammation, which originates in epithelial cancer cells. Oncogenes drive the onset of the cancer-associated fibroblast phenotype in adjacent normal fibroblasts via paracrine oxidative stress. This oncogene-induced transition to malignancy is “mirrored” by a loss of caveolin-1 (Cav-1) and an increase in MCT4 in adjacent stromal fibroblasts, functionally reflecting catabolic metabolism in the tumor microenvironment. Virtually identical findings were obtained using BRCA1-deficient breast and ovarian cancer cells. Thus, oncogene activation (RAS, NFkB, TGF-β) and/or tumor suppressor loss (BRCA1) have similar functional effects on adjacent stromal fibroblasts, initiating “metabolic symbiosis” and the cancer-associated fibroblast phenotype. New therapeutic strategies that metabolically uncouple oxidative cancer cells from their glycolytic stroma or modulate oxidative stress could be used to target this lethal subtype of cancers. Targeting “fibroblast addiction” in primary and metastatic tumor cells may expose a critical Achilles’ heel, leading to disease regression in both sporadic and familial cancers. PMID:23860382

3D aggregate culture improves metabolic maturation of human pluripotent stem cell derived cardiomyocytes.

PubMed

Correia, Cláudia; Koshkin, Alexey; Duarte, Patrícia; Hu, Dongjian; Carido, Madalena; Sebastião, Maria J; Gomes-Alves, Patrícia; Elliott, David A; Domian, Ibrahim J; Teixeira, Ana P; Alves, Paula M; Serra, Margarida

2018-03-01

Three-dimensional (3D) cultures of human pluripotent stem cell derived cardiomyocytes (hPSC-CMs) hold great promise for drug discovery, providing a better approximation to the in vivo physiology over standard two-dimensional (2D) monolayer cultures. However, the transition of CM differentiation protocols from 2D to 3D cultures is not straightforward. In this work, we relied on the aggregation of hPSC-derived cardiac progenitors and their culture under agitated conditions to generate highly pure cardiomyocyte aggregates. Whole-transcriptome analysis and 13 C-metabolic flux analysis allowed to demonstrate at both molecular and fluxome levels that such 3D culture environment enhances metabolic maturation of hiPSC-CMs. When compared to 2D, 3D cultures of hiPSC-CMs displayed down-regulation of genes involved in glycolysis and lipid biosynthesis and increased expression of genes involved in OXPHOS. Accordingly, 3D cultures of hiPSC-CMs had lower fluxes through glycolysis and fatty acid synthesis and increased TCA-cycle activity. Importantly, we demonstrated that the 3D culture environment reproducibly improved both CM purity and metabolic maturation across different hPSC lines, thereby providing a robust strategy to derive enriched hPSC-CMs with metabolic features closer to that of adult CMs. © 2017 Wiley Periodicals, Inc.

Acute exercise alters skeletal muscle mitochondrial respiration and H2O2 emission in response to hyperinsulinemic-euglycemic clamp in middle-aged obese men

PubMed Central

Trewin, Adam J.; Levinger, Itamar; Parker, Lewan; Shaw, Christopher S.; Serpiello, Fabio R.; Anderson, Mitchell J.; McConell, Glenn K.; Hare, David L.

2017-01-01

Obesity, sedentary lifestyle and aging are associated with mitochondrial dysfunction and impaired insulin sensitivity. Acute exercise increases insulin sensitivity in skeletal muscle; however, whether mitochondria are involved in these processes remains unclear. The aim of this study was to investigate the effects of insulin stimulation at rest and after acute exercise on skeletal muscle mitochondrial respiratory function (JO2) and hydrogen peroxide emission (JH2O2), and the associations with insulin sensitivity in obese, sedentary men. Nine men (means ± SD: 57 ± 6 years; BMI 33 ± 5 kg.m2) underwent hyperinsulinemic-euglycemic clamps in two separate trials 1–3 weeks apart: one under resting conditions, and another 1 hour after high-intensity exercise (4x4 min cycling at 95% HRpeak). Muscle biopsies were obtained at baseline, and pre/post clamp to measure JO2 with high-resolution respirometry and JH2O2 via Amplex UltraRed from permeabilized fibers. Post-exercise, both JO2 and JH2O2 during ADP stimulated state-3/OXPHOS respiration were lower compared to baseline (P<0.05), but not after subsequent insulin stimulation. JH2O2 was lower post-exercise and after subsequent insulin stimulation compared to insulin stimulation in the rest trial during succinate supported state-4/leak respiration (P<0.05). In contrast, JH2O2 increased during complex-I supported leak respiration with insulin after exercise compared with resting conditions (P<0.05). Resting insulin sensitivity and JH2O2 during complex-I leak respiration were positively correlated (r = 0.77, P<0.05). We conclude that in obese, older and sedentary men, acute exercise modifies skeletal muscle mitochondrial respiration and H2O2 emission responses to hyperinsulinemia in a respiratory state-specific manner, which may have implications for metabolic diseases involving insulin resistance. PMID:29161316

Codon usage bias and phylogenetic analysis of mitochondrial ND1 gene in pisces, aves, and mammals.

PubMed

Uddin, Arif; Choudhury, Monisha Nath; Chakraborty, Supriyo

2018-01-01

The mitochondrially encoded NADH:ubiquinone oxidoreductase core subunit 1 (MT-ND1) gene is a subunit of the respiratory chain complex I and involved in the first step of the electron transport chain of oxidative phosphorylation (OXPHOS). To understand the pattern of compositional properties, codon usage and expression level of mitochondrial ND1 genes in pisces, aves, and mammals, we used bioinformatic approaches as no work was reported earlier. In this study, a perl script was used for calculating nucleotide contents and different codon usage bias parameters. The codon usage bias of MT-ND1 was low but the expression level was high as revealed from high ENC and CAI value. Correspondence analysis (COA) suggests that the pattern of codon usage for MT-ND1 gene is not same across species and that compositional constraint played an important role in codon usage pattern of this gene among pisces, aves, and mammals. From the regression equation of GC12 on GC3, it can be inferred that the natural selection might have played a dominant role while mutation pressure played a minor role in influencing the codon usage patterns. Further, ND1 gene has a discrepancy with cytochrome B (CYB) gene in preference of codons as evident from COA. The codon usage bias was low. It is influenced by nucleotide composition, natural selection, mutation pressure, length (number) of amino acids, and relative dinucleotide composition. This study helps in understanding the molecular biology, genetics, evolution of MT-ND1 gene, and also for designing a synthetic gene.

Population mitogenomics provides insights into evolutionary history, source of invasions and diversifying selection in the House Crow (Corvus splendens).

PubMed

Krzemińska, Urszula; Morales, Hernán E; Greening, Chris; Nyári, Árpád S; Wilson, Robyn; Song, Beng Kah; Austin, Christopher M; Sunnucks, Paul; Pavlova, Alexandra; Rahman, Sadequr

2018-04-01

The House Crow (Corvus splendens) is a useful study system for investigating the genetic basis of adaptations underpinning successful range expansion. The species originates from the Indian subcontinent, but has successfully spread through a variety of thermal environments across Asia, Africa and Europe. Here, population mitogenomics was used to investigate the colonisation history and to test for signals of molecular selection on the mitochondrial genome. We sequenced the mitogenomes of 89 House Crows spanning four native and five invasive populations. A Bayesian dated phylogeny, based on the 13 mitochondrial protein-coding genes, supports a mid-Pleistocene (~630,000 years ago) divergence between the most distant genetic lineages. Phylogeographic patterns suggest that northern South Asia is the likely centre of origin for the species. Codon-based analyses of selection and assessments of changes in amino acid properties provide evidence of positive selection on the ND2 and ND5 genes against a background of purifying selection across the mitogenome. Protein homology modelling suggests that four amino acid substitutions inferred to be under positive selection may modulate coupling efficiency and proton translocation mediated by OXPHOS complex I. The identified substitutions are found within native House Crow lineages and ecological niche modelling predicts suitable climatic areas for the establishment of crow populations within the invasive range. Mitogenomic patterns in the invasive range of the species are more strongly associated with introduction history than climate. We speculate that invasions of the House Crow have been facilitated by standing genetic variation that accumulated due to diversifying selection within the native range.

Evolutionary implications of mitochondrial genetic variation: mitochondrial genetic effects on OXPHOS respiration and mitochondrial quantity change with age and sex in fruit flies.

PubMed

Wolff, J N; Pichaud, N; Camus, M F; Côté, G; Blier, P U; Dowling, D K

2016-04-01

The ancient acquisition of the mitochondrion into the ancestor of modern-day eukaryotes is thought to have been pivotal in facilitating the evolution of complex life. Mitochondria retain their own diminutive genome, with mitochondrial genes encoding core subunits involved in oxidative phosphorylation. Traditionally, it was assumed that there was little scope for genetic variation to accumulate and be maintained within the mitochondrial genome. However, in the past decade, mitochondrial genetic variation has been routinely tied to the expression of life-history traits such as fertility, development and longevity. To examine whether these broad-scale effects on life-history trait expression might ultimately find their root in mitochondrially mediated effects on core bioenergetic function, we measured the effects of genetic variation across twelve different mitochondrial haplotypes on respiratory capacity and mitochondrial quantity in the fruit fly, Drosophila melanogaster. We used strains of flies that differed only in their mitochondrial haplotype, and tested each sex separately at two different adult ages. Mitochondrial haplotypes affected both respiratory capacity and mitochondrial quantity. However, these effects were highly context-dependent, with the genetic effects contingent on both the sex and the age of the flies. These sex- and age-specific genetic effects are likely to resonate across the entire organismal life-history, providing insights into how mitochondrial genetic variation may contribute to sex-specific trajectories of life-history evolution. © 2016 European Society For Evolutionary Biology. Journal of Evolutionary Biology © 2016 European Society For Evolutionary Biology.

Positive selection on panpulmonate mitogenomes provide new clues on adaptations to terrestrial life.

PubMed

Romero, Pedro E; Weigand, Alexander M; Pfenninger, Markus

2016-08-22

Transitions from marine to intertidal and terrestrial habitats resulted in a significant adaptive radiation within the Panpulmonata (Gastropoda: Heterobranchia). This clade comprises several groups that invaded the land realm independently and in different time periods, e.g., Ellobioidea, Systellomatophora, and Stylommatophora. Thus, mitochondrial genomes of panpulmonate gastropods are promising to screen for adaptive molecular signatures related to land invasions. We obtained three complete mitochondrial genomes of terrestrial panpulmonates, i.e., the ellobiid Carychium tridentatum, and the stylommatophorans Arion rufus and Helicella itala. Our dataset consisted of 50 mitogenomes comprising almost all major panpulmonate lineages. The phylogenetic tree based on mitochondrial genes supports the monophyly of the clade Panpulmonata. Terrestrial lineages were sampled from Ellobioidea (1 sp.) and Stylommatophora (9 spp.). The branch-site test of positive selection detected significant non-synonymous changes in the terrestrial branches leading to Carychium (Ellobiodea) and Stylommatophora. These convergent changes occurred in the cob and nad5 genes (OXPHOS complex III and I, respectively). The convergence of the non-synonymous changes in cob and nad5 suggest possible ancient episodes of positive selection related to adaptations to non-marine habitats. The positively selected sites in our data are in agreement with previous results in vertebrates suggesting a general pattern of adaptation to the new metabolic requirements. The demand for energy due to the colonization of land (for example, to move and sustain the body mass in the new habitat) and the necessity to tolerate new conditions of abiotic stress may have changed the physiological constraints in the early terrestrial panpulmonates and triggered adaptations at the mitochondrial level.

Gait analysis in a mouse model resembling Leigh disease.

PubMed

de Haas, Ria; Russel, Frans G; Smeitink, Jan A

2016-01-01

Leigh disease (LD) is one of the clinical phenotypes of mitochondrial OXPHOS disorders and also known as sub-acute necrotizing encephalomyelopathy. The disease has an incidence of 1 in 77,000 live births. Symptoms typically begin early in life and prognosis for LD patients is poor. Currently, no clinically effective treatments are available. Suitable animal and cellular models are necessary for the understanding of the neuropathology and the development of successful new therapeutic strategies. In this study we used the Ndufs4 knockout (Ndufs4(-/-)) mouse, a model of mitochondrial complex I deficiency. Ndusf4(-/-) mice exhibit progressive neurodegeneration, which closely resemble the human LD phenotype. When dissecting behavioral abnormalities in animal models it is of great importance to apply translational tools that are clinically relevant. To distinguish gait abnormalities in patients, simple walking tests can be assessed, but in animals this is not easy. This study is the first to demonstrate automated CatWalk gait analysis in the Ndufs4(-/-) mouse model. Marked differences were noted between Ndufs4(-/-) and control mice in dynamic, static, coordination and support parameters. Variation of walking speed was significantly increased in Ndufs4(-/-) mice, suggesting hampered and uncoordinated gait. Furthermore, decreased regularity index, increased base of support and changes in support were noted in the Ndufs4(-/-) mice. Here, we report the ability of the CatWalk system to sensitively assess gait abnormalities in Ndufs4(-/-) mice. This objective gait analysis can be of great value for intervention and drug efficacy studies in animal models for mitochondrial disease. Copyright © 2015 Elsevier B.V. All rights reserved.

Activity-Based Protein Profiling Reveals Mitochondrial Oxidative Enzyme Impairment and Restoration in Diet-Induced Obese Mice

DOE Office of Scientific and Technical Information (OSTI.GOV)

Sadler, Natalie C.; Angel, Thomas E.; Lewis, Michael P.

High-fat diet (HFD) induced obesity and concomitant development of insulin resistance (IR) and type 2 diabetes mellitus have been linked to mitochondrial dysfunction. However, it is not clear whether mitochondrial dysfunction is a direct effect of a HFD or if the mitochondrial function is reduced with increased HFD duration. We hypothesized that the function of mitochondrial oxidative and lipid metabolism functions in skeletal muscle mitochondria for HFD mice are similar or elevated relative to standard diet (SD) mice, thereby IR is neither cause nor consequence of mitochondrial dysfunction. We applied a chemical probe approach to identify functionally reactive ATPases andmore » nucleotide-binding proteins in mitochondria isolated from skeletal muscle of C57Bl/6J mice fed HFD or SD chow for 2-, 8-, or 16-weeks; feeding time points known to induce IR. A total of 293 probe-labeled proteins were identified by mass spectrometry-based proteomics, of which 54 differed in abundance between HFD and SD mice. We found proteins associated with the TCA cycle, oxidative phosphorylation (OXPHOS), and lipid metabolism were altered in function when comparing SD to HFD fed mice at 2-weeks, however by 16-weeks HFD mice had TCA cycle, β-oxidation, and respiratory chain function at levels similar to or higher than SD mice.« less

Dietary fat overload reprograms brown fat mitochondria.

PubMed

Lettieri Barbato, Daniele; Tatulli, Giuseppe; Vegliante, Rolando; Cannata, Stefano M; Bernardini, Sergio; Ciriolo, Maria R; Aquilano, Katia

2015-01-01

Chronic nutrient overload accelerates the onset of several aging-related diseases reducing life expectancy. Although the mechanisms by which overnutrition affects metabolic processes in many tissues are known, its role on BAT physiology is still unclear. Herein, we investigated the mitochondrial responses in BAT of female mice exposed to high fat diet (HFD) at different steps of life. Although adult mice showed an unchanged mitochondrial amount, both respiration and OxPHOS subunits were strongly affected. Differently, offspring pups exposed to HFD during pregnancy and lactation displayed reduced mitochondrial mass but high oxidative efficiency that, however, resulted in increased bioenergetics state of BAT rather than augmented uncoupling respiration. Interestingly, the metabolic responses triggered by HFD were accompanied by changes in mitochondrial dynamics characterized by decreased content of the fragmentation marker Drp1 both in mothers and offspring pups. HFD-induced inactivation of the FoxO1 transcription factor seemed to be the up-stream modulator of Drp1 levels in brown fat cells. Furthermore, HFD offspring pups weaned with normal diet only partially reverted the mitochondrial dysfunctions caused by HFD. Finally these mice failed in activating the thermogenic program upon cold exposure. Collectively our findings suggest that maternal dietary fat overload irreversibly commits BAT unresponsiveness to physiological stimuli such as cool temperature and this dysfunction in the early stage of life might negatively modulate health and lifespan.

Oxidative stress, mitochondrial perturbations and fetal programming of renal disease induced by maternal smoking.

PubMed

Stangenberg, Stefanie; Nguyen, Long T; Chen, Hui; Al-Odat, Ibrahim; Killingsworth, Murray C; Gosnell, Martin E; Anwer, Ayad G; Goldys, Ewa M; Pollock, Carol A; Saad, Sonia

2015-07-01

An adverse in-utero environment is increasingly recognized to predispose to chronic disease in adulthood. Maternal smoking remains the most common modifiable adverse in-utero exposure leading to low birth weight, which is strongly associated with chronic kidney disease (CKD) in later life. In order to investigate underlying mechanisms for such susceptibility, female Balb/c mice were sham or cigarette smoke-exposed (SE) for 6 weeks before mating, throughout gestation and lactation. Offspring kidneys were examined for oxidative stress, expression of mitochondrial proteins, mitochondrial structure as well as renal functional parameters on postnatal day 1, day 20 (weaning) and week 13 (adult age). From birth throughout adulthood, SE offspring had increased renal levels of mitochondrial-derived reactive oxygen species (ROS), which left a footprint on DNA with increased 8-hydroxydeoxyguanosin (8-OHdG) in kidney tubular cells. Mitochondrial structural abnormalities were seen in SE kidneys at day 1 and week 13 along with a reduction in oxidative phosphorylation (OXPHOS) proteins and activity of mitochondrial antioxidant Manganese superoxide dismutase (MnSOD). Smoke exposure also resulted in increased mitochondrial DNA copy number (day 1-week 13) and lysosome density (day 1 and week 13). The appearance of mitochondrial defects preceded the onset of albuminuria at week 13. Thus, mitochondrial damage caused by maternal smoking may play an important role in development of CKD at adult life. Copyright © 2015 Elsevier Ltd. All rights reserved.

Mechanistic study on the nuclear modifier gene MSS1 mutation suppressing neomycin sensitivity of the mitochondrial 15S rRNA C1477G mutation in Saccharomyces cerevisiae.

PubMed

Zhou, Qiyin; Wang, Wei; He, Xiangyu; Zhu, Xiaoyu; Shen, Yaoyao; Yu, Zhe; Wang, Xuexiang; Qi, Xuchen; Zhang, Xuan; Fan, Mingjie; Dai, Yu; Yang, Shuxu; Yan, Qingfeng

2014-01-01

The phenotypic manifestation of mitochondrial DNA (mtDNA) mutations can be modulated by nuclear genes and environmental factors. However, neither the interaction among these factors nor their underlying mechanisms are well understood. The yeast Saccharomyces cerevisiae mtDNA 15S rRNA C1477G mutation (PR) corresponds to the human 12S rRNA A1555G mutation. Here we report that a nuclear modifier gene mss1 mutation suppresses the neomycin-sensitivity phenotype of a yeast C1477G mutant in fermentable YPD medium. Functional assays show that the mitochondrial function of the yeast C1477G mutant was impaired severely in YPD medium with neomycin. Moreover, the mss1 mutation led to a significant increase in the steady-state level of HAP5 (heme activated protein), which greatly up-regulated the expression of glycolytic transcription factors RAP1, GCR1, and GCR2 and thus stimulated glycolysis. Furthermore, the high expression of the key glycolytic enzyme genes HXK2, PFK1 and PYK1 indicated that enhanced glycolysis not only compensated for the ATP reduction from oxidative phosphorylation (OXPHOS) in mitochondria, but also ensured the growth of the mss1(PR) mutant in YPD medium with neomycin. This study advances our understanding of the phenotypic manifestation of mtDNA mutations.

The role of sodium hydrosulfide in attenuating the aging process via PI3K/AKT and CaMKKβ/AMPK pathways.

PubMed

Chen, Xubo; Zhao, Xueyan; Cai, Hua; Sun, Haiying; Hu, Yujuan; Huang, Xiang; Kong, Wen; Kong, Weijia

2017-08-01

Age-related dysfunction of the central auditory system, known as central presbycusis, is characterized by defects in speech perception and sound localization. It is important to determine the pathogenesis of central presbycusis in order to explore a feasible and effective intervention method. Recent work has provided fascinating insight into the beneficial function of H 2 S on oxidative stress and stress-related disease. In this study, we investigated the pathogenesis of central presbycusis and tried to explore the mechanism of H 2 S action on different aspects of aging by utilizing a mimetic aging rat and senescent cellular model. Our results indicate that NaHS decreased oxidative stress and apoptosis levels in an aging model via CaMKKβ and PI3K/AKT signaling pathways. Moreover, we found that NaHS restored the decreased activity of antioxidants such as GSH, SOD and CAT in the aging model in vivo and in vitro by regulating CaMKKβ and PI3K/AKT. Mitochondria function was preserved by NaHS, as indicated by the following: DNA POLG and OGG-1, the base excision repair enzymes in mitochondrial, were upregulated; OXPHOS activity was downregulated; mitochondrial membrane potential was restored; ATP production was increased; and mtDNA damage, indicated by the common deletion (CD), declined. These effects were also achieved by activating CaMKKβ/AMPK and PI3K/AKT signaling pathways. Lastly, protein homeostasis, indicated by HSP90 alpha, was strengthened by NaHS via CaMKKβ and PI3K/AKT. Our findings demonstrate that the ability to resist oxidative stress and mitochondria function are both decreased as aging developed; however, NaHS, a novel free radical scavenger and mitochondrial protective agent, precludes the process of oxidative damage by activating CaMKKβ and PI3K/AKT. This study might provide a therapeutic target for aging and age-related disease. Copyright © 2017 The Authors. Published by Elsevier B.V. All rights reserved.

Glucose deprivation increases monocarboxylate transporter 1 (MCT1) expression and MCT1-dependent tumor cell migration.

PubMed

De Saedeleer, C J; Porporato, P E; Copetti, T; Pérez-Escuredo, J; Payen, V L; Brisson, L; Feron, O; Sonveaux, P

2014-07-31

The glycolytic end-product lactate is a pleiotropic tumor growth-promoting factor. Its activities primarily depend on its uptake, a process facilitated by the lactate-proton symporter monocarboxylate transporter 1 (MCT1). Therefore, targeting the transporter or its chaperon protein CD147/basigin, itself involved in the aggressive malignant phenotype, is an attractive therapeutic option for cancer, but basic information is still lacking regarding the regulation of the expression, interaction and activities of both proteins. In this study, we found that glucose deprivation dose-dependently upregulates MCT1 and CD147 protein expression and their interaction in oxidative tumor cells. While this posttranslational induction could be recapitulated using glycolysis inhibition, hypoxia, oxidative phosphorylation (OXPHOS) inhibitor rotenone or hydrogen peroxide, it was blocked with alternative oxidative substrates and specific antioxidants, pointing out at a mitochondrial control. Indeed, we found that the stabilization of MCT1 and CD147 proteins upon glucose removal depends on mitochondrial impairment and the associated generation of reactive oxygen species. When glucose was a limited resource (a situation occurring naturally or during the treatment of many tumors), MCT1-CD147 heterocomplexes accumulated, including in cell protrusions of the plasma membrane. It endowed oxidative tumor cells with increased migratory capacities towards glucose. Migration increased in cells overexpressing MCT1 and CD147, but it was inhibited in glucose-starved cells provided with an alternative oxidative fuel, treated with an antioxidant, lacking MCT1 expression, or submitted to pharmacological MCT1 inhibition. While our study identifies the mitochondrion as a glucose sensor promoting tumor cell migration, MCT1 is also revealed as a transducer of this response, providing a new rationale for the use of MCT1 inhibitors in cancer.

Regulation of the oxidative balance with coenzyme Q10 sensitizes human glioblastoma cells to radiation and temozolomide.

PubMed

Frontiñán-Rubio, Javier; Santiago-Mora, Raquel María; Nieva-Velasco, Consuelo María; Ferrín, Gustavo; Martínez-González, Alicia; Gómez, María Victoria; Moreno, María; Ariza, Julia; Lozano, Eva; Arjona-Gutiérrez, Jacinto; Gil-Agudo, Antonio; De la Mata, Manuel; Pesic, Milica; Peinado, Juan Ramón; Villalba, José M; Pérez-Romasanta, Luis; Pérez-García, Víctor M; Alcaín, Francisco J; Durán-Prado, Mario

2018-05-18

To investigate how the modulation of the oxidative balance affects cytotoxic therapies in glioblastoma, in vitro. Human glioblastoma U251 and T98 cells and normal astrocytes C8D1A were loaded with coenzyme Q10 (CoQ). Mitochondrial superoxide ion (O 2 - ) and H 2 O 2 were measured by fluorescence microscopy. OXPHOS performance was assessed in U251 cells with an oxytherm Clark-type electrode. Radio- and chemotherapy cytotoxicity was assessed by immunostaining of γH2AX (24 h), annexin V and nuclei morphology, at short (72 h) and long (15 d) time. Hif-1α, SOD1, SOD2 and NQO1 were determined by immunolabeling. Catalase activity was measured by classic enzymatic assay. Glutathione levels and total antioxidant capacity were quantified using commercial kits. CoQ did not affect oxygen consumption but reduced the level of O 2 - and H 2 O 2 while shifted to a pro-oxidant cell status mainly due to a decrease in catalase activity and SOD2 level. Hif-1α was dampened, echoed by a decrease lactate and several key metabolites involved in glutathione synthesis. CoQ-treated cells were twofold more sensitive than control to radiation-induced DNA damage and apoptosis in short and long-term clonogenic assays, potentiating TMZ-induced cytotoxicity, without affecting non-transformed astrocytes. CoQ acts as sensitizer for cytotoxic therapies, disarming GBM cells, but not normal astrocytes, against further pro-oxidant injuries, being potentially useful in clinical practice for this fatal pathology. Copyright © 2018 Elsevier B.V. All rights reserved.

Association of cancer metabolism-related proteins with oral carcinogenesis – indications for chemoprevention and metabolic sensitizing of oral squamous cell carcinoma?

PubMed Central

2014-01-01

Background Tumor metabolism is a crucial factor for the carcinogenesis of oral squamous cell carcinoma (OSCC). Methods Expression of IGF-R1, glycolysis-related proteins (GLUT-1, HK 2, PFK-1, LDHA, TKTL1), mitochondrial enzymes (SDHA, SDHB, ATP synthase) were analyzed in normal oral mucosa (n = 5), oral precursor lesions (simple hyperplasia, n = 11; squamous intraepithelial neoplasia, SIN I-III, n = 35), and OSCC specimen (n = 42) by immunohistochemistry and real-time polymerase chain reaction (qPCR) analysis in OSCC cell lines. Metabolism-related proteins were correlated with proliferation activity (Ki-67) and apoptotic properties (TUNEL assay) in OSCC. Specificity of antibodies was confirmed by western blotting in cancer cell lines. Results Expression of IGF-R1, glycolysis-related proteins (GLUT-1, HK 2, LDHA, TKTL1), and mitochondrial enzymes (SDHA, SDHB, ATP synthase) were significantly increased in the carcinogenesis of OSCC. Metabolic active regions of OSCC were strongly correlated with proliferating cancer (Ki-67+) cells without detection of apoptosis (TUNEL assay). Conclusions This study provides the first evidence of the expression of IGF-R1, glycolysis-related proteins GLUT-1, HK 2, PFK-1, LDHA, and TKTL1, as well as mitochondrial enzymes SDHA, SDHB, and ATP synthase in the multi-step carcinogenesis of OSCC. Both, hypoxia-related glucose metabolism and mitochondrial oxidative phosphorylation characteristics are associated with the carcinogenesis of OSCC. Acidosis and OXPHOS may drive a metabolic shift towards the pentose phosphate pathway (PPP). Therefore, inhibition of the PPP, glycolysis, and targeted anti-mitochondrial therapies (ROS generation) by natural compounds or synthetic vitamin derivatives may act as sensitizer for apoptosis in cancer cells mediated by adjuvant therapies in OSCC. PMID:25048361

Electronic cigarette aerosols and copper nanoparticles induce mitochondrial stress and promote DNA fragmentation in lung fibroblasts

DOE Office of Scientific and Technical Information (OSTI.GOV)

Lerner, Chad A.; Rutagarama, Pierrot; Ahmad, Tanveer

Oxidants or nanoparticles have recently been identified as constituents of aerosols released from various styles of electronic cigarettes (E-cigs). Cells in the lung may be directly exposed to these constituents and harbor reactive properties capable of incurring acute cell injury. Our results show mitochondria are sensitive to both E-cig aerosols and aerosol containing copper nanoparticles when exposed to human lung fibroblasts (HFL-1) using an Air-Liquid Interface culture system, evident by elevated levels of mitochondrial ROS (mtROS). Increased mtROS after aerosol exposure is associated with reduced stability of OxPhos electron transport chain (ETC) complex IV subunit and nuclear DNA fragmentation. Increasedmore » levels of IL-8 and IL-6 in HFL-1 conditioned media were also observed. These findings reveal both mitochondrial, genotoxic, and inflammatory stresses are features of direct cell exposure to E-cig aerosols which are ensued by inflammatory duress, raising a concern on deleterious effect of vaping. - Graphical abstract: Oxidants and possibly reactive properties of metal particles in E-cig aerosols impart mitochondrial oxidative stress and DNA damage. These biological effects accompany inflammatory response which may raise concern regarding long term E-cig use. Mitochondria may be particularly sensitive to reactive properties of E-cig aerosols in addition to the potential for them to induce genotoxic stress by generating increased ROS. - Highlights: • Mitochondria are sensitive to both E-cig aerosols and metal nanoparticles. • Increased mtROS by E-cig aerosol is associated with disrupted mitochondrial energy. • E-cig causes nuclear DNA fragmentation. • E-cig aerosols induce pro-inflammatory response in human fibroblasts.« less

Low Concentrations of Cationic PAMAM Dendrimers Affect Lymphocyte Respiration in In vitro Studies.

PubMed

Labieniec-Watala, Magdalena; Szwed, Marzena; Hertel, Joanna; Wisnik, Ewelina

2017-01-01

In this study, the effect of low concentrations of poly(amido)amine dendrimers (G2-G4) on human lymphocytes was studied. Some works revealed that PAMAMs can adversely affect the morphology of blood components and mitochondria functions. In this context, the present report aimed to investigate the in vitro cationic dendrimers' effect on mitochondrial respiration and cell morphology in lymphocytes isolated from human blood. To monitor the mitochondrial changes, the high-resolution respirometer was used, whereas the cell morphology was analyzed using a flow cytometer and fluorescence microscopy. The concentration-dependent dendrimers' influence on lymphocytes morphology was shown. Changes in mitochondrial respiration revealed the concentration- and generation-dependent differences between dendrimer activity. There were no alterations in the routine respiration and in the state of the inner mitochondrial membrane (L/E), but decreased ADP- and FCCP-stimulated respirations were detected after treatment with G3 and G4 dendrimers. The markers of mitochondrial membrane integrity (RCR) and OXPHOS efficiency (P/E) significantly decreased regardless of the dendrimer generation used. Based on these in vitro evaluations, we state that cationic PAMAM dendrimers can impair both the morphology and the bioenergetics of human lymphocytes, even when used at low concentrations and in a short time (up to 1 h). However, these results do not imply that similar findings could be possible for in vivo observations. Copyright© Bentham Science Publishers; For any queries, please email at epub@benthamscience.org.

Metabolic adaptations to short-term every-other-day feeding in long-living Ames dwarf mice.

PubMed

Brown-Borg, Holly M; Rakoczy, Sharlene

2013-09-01

Restrictive dietary interventions exert significant beneficial physiological effects in terms of aging and age-related disease in many species. Every other day feeding (EOD) has been utilized in aging research and shown to mimic many of the positive outcomes consequent with dietary restriction. This study employed long living Ames dwarf mice subjected to EOD feeding to examine the adaptations of the oxidative phosphorylation and antioxidative defense systems to this feeding regimen. Every other day feeding lowered liver glutathione (GSH) concentrations in dwarf and wild type (WT) mice but altered GSH biosynthesis and degradation in WT mice only. The activities of liver OXPHOS enzymes and corresponding proteins declined in WT mice fed EOD while in dwarf animals, the levels were maintained or increased with this feeding regimen. Antioxidative enzymes were differentially affected depending on the tissue, whether proliferative or post-mitotic. Gene expression of components of liver methionine metabolism remained elevated in dwarf mice when compared to WT mice as previously reported however, enzymes responsible for recycling homocysteine to methionine were elevated in both genotypes in response to EOD feeding. The data suggest that the differences in anabolic hormone levels likely affect the sensitivity of long living and control mice to this dietary regimen, with dwarf mice exhibiting fewer responses in comparison to WT mice. These results provide further evidence that dwarf mice may be better protected against metabolic and environmental perturbations which may in turn, contribute to their extended longevity. © 2013.

A novel mutation MT-COIII m.9267G>C and MT-COI m.5913G>A mutation in mitochondrial genes in a Tunisian family with maternally inherited diabetes and deafness (MIDD) associated with sever nephropathy

DOE Office of Scientific and Technical Information (OSTI.GOV)

Tabebi, Mouna, E-mail: mouna.biologiste@yahoo.com; Mkaouar-Rebai, Emna; Mnif, Mouna

Mitochondrial diabetes (MD) is a heterogeneous disorder characterized by a chronic hyperglycemia, maternal transmission and its association with a bilateral hearing impairment. Several studies reported mutations in mitochondrial genes as potentially pathogenic for diabetes, since mitochondrial oxidative phosphorylation plays an important role in glucose-stimulated insulin secretion from beta cells. In the present report, we studied a Tunisian family with mitochondrial diabetes (MD) and deafness associated with nephropathy. The mutational analysis screening revealed the presence of a novel heteroplasmic mutation m.9276G>C in the mitochondrial COIII gene, detected in mtDNA extracted from leukocytes of a mother and her two daughters indicating thatmore » this mutation is maternally transmitted and suggest its implication in the observed phenotype. Bioinformatic tools showed that m.9267G>C mutation (p.A21P) is « deleterious » and it can modify the function and the stability of the MT-COIII protein by affecting the assembly of mitochondrial COX subunits and the translocation of protons then reducing the activity of the respective OXPHOS complexes of ATP synthesis. The nonsynonymous mutation (p.A21P) has not been reported before, it is the first mutation described in the COXIII gene which is related to insulin dependent mitochondrial diabetes and deafness and could be specific to the Tunisian population. The m.9267G>C mutation was present with a nonsynonymous inherited mitochondrial homoplasmic variation MT-COI m.5913 G>A (D4N) responsible of high blood pressure, a clinical feature detected in all explored patients. - Highlights: • MT-COX3 m.9267G>C (p.A21P), heteroplasmic substitution, is not reported in any database. • m.9267G>C can be responsible of the MIDD associated with nephropaty. • This substitution can modify the function and the stability of the MT-CO3 protein. • This substitution can modify MT-CO3 structure (2D and 3D). • MT-COX3 m.9267G>C is associated with MT-CO1 m.5913G>A a homoplasmic substitution.« less

Mitochondria in lung disease

PubMed Central

Cloonan, Suzanne M.; Choi, Augustine M.K.

2016-01-01

Mitochondria are a distinguishing feature of eukaryotic cells. Best known for their critical function in energy production via oxidative phosphorylation (OXPHOS), mitochondria are essential for nutrient and oxygen sensing and for the regulation of critical cellular processes, including cell death and inflammation. Such diverse functional roles for organelles that were once thought to be simple may be attributed to their distinct heteroplasmic genome, exclusive maternal lineage of inheritance, and ability to generate signals to communicate with other cellular organelles. Mitochondria are now thought of as one of the cell’s most sophisticated and dynamic responsive sensing systems. Specific signatures of mitochondrial dysfunction that are associated with disease pathogenesis and/or progression are becoming increasingly important. In particular, the centrality of mitochondria in the pathological processes and clinical phenotypes associated with a range of lung diseases is emerging. Understanding the molecular mechanisms regulating the mitochondrial processes of lung cells will help to better define phenotypes and clinical manifestations associated with respiratory disease and to identify potential diagnostic and therapeutic targets. PMID:26928034

Cancer metabolism, stemness and tumor recurrence: MCT1 and MCT4 are functional biomarkers of metabolic symbiosis in head and neck cancer.

PubMed

Curry, Joseph M; Tuluc, Madalina; Whitaker-Menezes, Diana; Ames, Julie A; Anantharaman, Archana; Butera, Aileen; Leiby, Benjamin; Cognetti, David M; Sotgia, Federica; Lisanti, Michael P; Martinez-Outschoorn, Ubaldo E

2013-05-01

Here, we interrogated head and neck cancer (HNSCC) specimens (n = 12) to examine if different metabolic compartments (oxidative vs. glycolytic) co-exist in human tumors. A large panel of well-established biomarkers was employed to determine the metabolic state of proliferative cancer cells. Interestingly, cell proliferation in cancer cells, as marked by Ki-67 immunostaining, was strictly correlated with oxidative mitochondrial metabolism (OXPHOS) and the uptake of mitochondrial fuels, as detected via MCT1 expression (p < 0.001). More specifically, three metabolic tumor compartments were delineated: (1) proliferative and mitochondrial-rich cancer cells (Ki-67+/TOMM20+/COX+/MCT1+); (2) non-proliferative and mitochondrial-poor cancer cells (Ki-67-/TOMM20-/COX-/MCT1-); and (3) non-proliferative and mitochondrial-poor stromal cells (Ki-67-/TOMM20-/COX-/MCT1-). In addition, high oxidative stress (MCT4+) was very specific for cancer tissues. Thus, we next evaluated the prognostic value of MCT4 in a second independent patient cohort (n = 40). Most importantly, oxidative stress (MCT4+) in non-proliferating epithelial cancer cells predicted poor clinical outcome (tumor recurrence; p < 0.0001; log-rank test), and was functionally associated with FDG-PET avidity (p < 0.04). Similarly, oxidative stress (MCT4+) in tumor stromal cells was specifically associated with higher tumor stage (p < 0.03), and was a highly specific marker for cancer-associated fibroblasts (p < 0.001). We propose that oxidative stress is a key hallmark of tumor tissues that drives high-energy metabolism in adjacent proliferating mitochondrial-rich cancer cells, via the paracrine transfer of mitochondrial fuels (such as L-lactate and ketone bodies). New antioxidants and MCT4 inhibitors should be developed to metabolically target "three-compartment tumor metabolism" in head and neck cancers. It is remarkable that two "non-proliferating" populations of cells (Ki-67-/MCT4+) within the tumor can actually determine clinical outcome, likely by providing high-energy mitochondrial "fuels" for proliferative cancer cells to burn. Finally, we also show that in normal mucosal tissue, the basal epithelial "stem cell" layer is hyper-proliferative (Ki-67+), mitochondrial-rich (TOMM20+/COX+) and is metabolically programmed to use mitochondrial fuels (MCT1+), such as ketone bodies and L-lactate. Thus, oxidative mitochondrial metabolism (OXPHOS) is a common feature of both (1) normal stem cells and (2) proliferating cancer cells. As such, we should consider metabolically treating cancer patients with mitochondrial inhibitors (such as Metformin), and/or with a combination of MCT1 and MCT4 inhibitors, to target "metabolic symbiosis."

Cancer metabolism, stemness and tumor recurrence

PubMed Central

Curry, Joseph M.; Tuluc, Madalina; Whitaker-Menezes, Diana; Ames, Julie A.; Anantharaman, Archana; Butera, Aileen; Leiby, Benjamin; Cognetti, David M.; Sotgia, Federica; Lisanti, Michael P.; Martinez-Outschoorn, Ubaldo E.

2013-01-01

Here, we interrogated head and neck cancer (HNSCC) specimens (n = 12) to examine if different metabolic compartments (oxidative vs. glycolytic) co-exist in human tumors. A large panel of well-established biomarkers was employed to determine the metabolic state of proliferative cancer cells. Interestingly, cell proliferation in cancer cells, as marked by Ki-67 immunostaining, was strictly correlated with oxidative mitochondrial metabolism (OXPHOS) and the uptake of mitochondrial fuels, as detected via MCT1 expression (p < 0.001). More specifically, three metabolic tumor compartments were delineated: (1) proliferative and mitochondrial-rich cancer cells (Ki-67+/TOMM20+/COX+/MCT1+); (2) non-proliferative and mitochondrial-poor cancer cells (Ki-67−/TOMM20−/COX−/MCT1−); and (3) non-proliferative and mitochondrial-poor stromal cells (Ki-67−/TOMM20−/COX−/MCT1−). In addition, high oxidative stress (MCT4+) was very specific for cancer tissues. Thus, we next evaluated the prognostic value of MCT4 in a second independent patient cohort (n = 40). Most importantly, oxidative stress (MCT4+) in non-proliferating epithelial cancer cells predicted poor clinical outcome (tumor recurrence; p < 0.0001; log-rank test), and was functionally associated with FDG-PET avidity (p < 0.04). Similarly, oxidative stress (MCT4+) in tumor stromal cells was specifically associated with higher tumor stage (p < 0.03), and was a highly specific marker for cancer-associated fibroblasts (p < 0.001). We propose that oxidative stress is a key hallmark of tumor tissues that drives high-energy metabolism in adjacent proliferating mitochondrial-rich cancer cells, via the paracrine transfer of mitochondrial fuels (such as L-lactate and ketone bodies). New antioxidants and MCT4 inhibitors should be developed to metabolically target “three-compartment tumor metabolism” in head and neck cancers. It is remarkable that two “non-proliferating” populations of cells (Ki-67−/MCT4+) within the tumor can actually determine clinical outcome, likely by providing high-energy mitochondrial “fuels” for proliferative cancer cells to burn. Finally, we also show that in normal mucosal tissue, the basal epithelial “stem cell” layer is hyper-proliferative (Ki-67+), mitochondrial-rich (TOMM20+/COX+) and is metabolically programmed to use mitochondrial fuels (MCT1+), such as ketone bodies and L-lactate. Thus, oxidative mitochondrial metabolism (OXPHOS) is a common feature of both (1) normal stem cells and (2) proliferating cancer cells. As such, we should consider metabolically treating cancer patients with mitochondrial inhibitors (such as Metformin), and/or with a combination of MCT1 and MCT4 inhibitors, to target “metabolic symbiosis.” PMID:23574725

Mitochondrial uncoupling reduces exercise capacity despite several skeletal muscle metabolic adaptations.

PubMed

Schlagowski, A I; Singh, F; Charles, A L; Gali Ramamoorthy, T; Favret, F; Piquard, F; Geny, B; Zoll, J

2014-02-15

The effects of mitochondrial uncoupling on skeletal muscle mitochondrial adaptation and maximal exercise capacity are unknown. In this study, rats were divided into a control group (CTL, n = 8) and a group treated with 2,4-dinitrophenol, a mitochondrial uncoupler, for 28 days (DNP, 30 mg·kg(-1)·day(-1) in drinking water, n = 8). The DNP group had a significantly lower body mass (P < 0.05) and a higher resting oxygen uptake (Vo2, P < 0.005). The incremental treadmill test showed that maximal running speed and running economy (P < 0.01) were impaired but that maximal Vo2 (Vo2max) was higher in the DNP-treated rats (P < 0.05). In skinned gastrocnemius fibers, basal respiration (V0) was higher (P < 0.01) in the DNP-treated animals, whereas the acceptor control ratio (ACR, Vmax/V0) was significantly lower (P < 0.05), indicating a reduction in OXPHOS efficiency. In skeletal muscle, DNP activated the mitochondrial biogenesis pathway, as indicated by changes in the mRNA expression of PGC1-α and -β, NRF-1 and -2, and TFAM, and increased the mRNA expression of cytochrome oxidase 1 (P < 0.01). The expression of two mitochondrial proteins (prohibitin and Ndufs 3) was higher after DNP treatment. Mitochondrial fission 1 protein (Fis-1) was increased in the DNP group (P < 0.01), but mitofusin-1 and -2 were unchanged. Histochemical staining for NADH dehydrogenase and succinate dehydrogenase activity in the gastrocnemius muscle revealed an increase in the proportion of oxidative fibers after DNP treatment. Our study shows that mitochondrial uncoupling induces several skeletal muscle adaptations, highlighting the role of mitochondrial coupling as a critical factor for maximal exercise capacities. These results emphasize the importance of investigating the qualitative aspects of mitochondrial function in addition to the amount of mitochondria.

Anemone rivularis inhibits pyruvate dehydrogenase kinase activity and tumor growth.

PubMed

Chung, Tae-Wook; Lee, Jung Hee; Choi, Hee-Jung; Park, Mi-Ju; Kim, Eun-Yeong; Han, Jung Ho; Jang, Se Bok; Lee, Syng-Ook; Lee, Sang Woo; Hang, Jin; Yi, Li Wan; Ha, Ki-Tae

2017-05-05

Anemone rivularis Buch.-Ham. ex DC. (Ranunculaceae) have been used as a traditional remedy for treatment of inflammation and cancer. However, there is no report demonstrating experimental evidence on anti-tumor action of A. rivularis. The Warburg's effect, preference of aerobic glycolysis rather than oxidative phosphorylation (OXPHOS) even in oxygen rich condition, is focused as one of major characteristics of malignant tumor. Thus, we investigated the effect of A. rivularis on the Pyruvate dehydrogenase (PDH) kinases (PDHKs), a major molecular targets for reducing aerobic glycolysis. The ethanol extract of whole plant of A. rivularis (ARE), fingerprinted by high performance liquid chromatography (HPLC), was applied to in vitro and cell-based PDHK activity assays. The effect of ARE on cell viabilities of several tumor cells was estimated by MTT assay. The expression of phosphor-PDH, PDH and PDHK1 were measured by Western blot analysis. The production of reactive oxygen species (ROS) and apoptosis was measured by fluorescence-activated cell sorting analysis, using 5-(and-6)-carboxy-2',7'-dichlorodihydrofluorescein diacetate (carboxy-H2DCFDA) and Annexin V/propidium iodide (PI) staining, respectively. Mitochondrial membrane potential was examined by tetramethylrhodamine methyl ester (TMRM) staining. In vivo anti-tumor efficacy of ARE was estimated by means of tumor volume and weight using allograft injection of murine Lewis lung carcinoma (LLC) cells to dorsa of C57BL/6 mice. ARE inhibited the viabilities of several cancer cells, including MDA-MB321, K562, HT29, Hep3B, DLD-1, and LLC. ARE suppressed PDHK activity in in vitro kinase assay, and also inhibited aerobic glycolysis by reducing phosphorylation of PDHA in human DLD-1 colon cancer and murine LLC cells. The expression of PDHK1, a major isoform of PDHKs in cancer, was not affected by ARE treatment. Moreover, ARE increased the both ROS production and mitochondrial damage. In addition, ARE suppressed the in vitro tumor growth through mitochondria-mediated apoptosis. The growth rates of allograft LLC cells were also reduced by ARE treatment. Here, we firstly report that ARE inhibits PDHK activity and growth of tumor in both in vitro and in vivo experiments. Therefore, we suggest ARE as a potential candidate for developing anti-cancer drugs. Copyright © 2017 Elsevier Ireland Ltd. All rights reserved.

Cancer-associated fibroblasts drive glycolysis in a targetable signaling loop implicated in head and neck squamous cell carcinoma progression.

PubMed

Kumar, Dhruv; New, Jacob; Vishwakarma, Vikalp; Joshi, Radhika; Enders, Jonathan; Lin, Fangchen; Dasari, Sumana; Gutierrez, Wade R; Leef, George; Ponnurangam, Sivapriya; Chavan, Hemantkumar; Ganaden, Lydia; Thornton, Mackenzie M; Dai, Hongying; Tawfik, Ossama; Straub, Jeffrey; Shnayder, Yelizaveta; Kakarala, Kiran; Tsue, Terance Ted; Girod, Douglas A; Van Houten, Bennett; Anant, Shrikant; Krishnamurthy, Partha; Thomas, Sufi Mary

2018-05-16

Despite aggressive therapies, head and neck squamous cell carcinoma (HNSCC) is associated with a less than 50% 5-year survival rate. Late stage HNSCC frequently consists of up to 80% cancer-associated fibroblasts (CAF). We previously reported that CAF-secreted hepatocyte growth factor (HGF) facilitates HNSCC progression, however very little is known about the role of CAFs in HNSCC metabolism. Here we demonstrate that CAF-secreted HGF increases extracellular lactate levels in HNSCC via upregulation of glycolysis. CAF-secreted HGF induced basic fibroblast growth factor (bFGF) secretion from HNSCC. CAFs were more efficient than HNSCC in using lactate as a carbon source. HNSCC-secreted bFGF increased mitochondrial oxidative phosphorylation (OXPHOS) and HGF secretion from CAFs. Combined inhibition of c-Met and FGFR significantly inhibited CAF-induced HNSCC growth in vitro and in vivo (p<0.001). Our cumulative findings underscore reciprocal signaling between CAF and HNSCC involving bFGF and HGF. This contributes to metabolic symbiosis and a targetable therapeutic axis involving c-Met and FGFR. Copyright ©2018, American Association for Cancer Research.

Hodgkin lymphoma: A complex metabolic ecosystem with glycolytic reprogramming of the tumor microenvironment.

PubMed

Mikkilineni, Lekha; Whitaker-Menezes, Diana; Domingo-Vidal, Marina; Sprandio, John; Avena, Paola; Cotzia, Paolo; Dulau-Florea, Alina; Gong, Jerald; Uppal, Guldeep; Zhan, Tingting; Leiby, Benjamin; Lin, Zhao; Pro, Barbara; Sotgia, Federica; Lisanti, Michael P; Martinez-Outschoorn, Ubaldo

2017-06-01

Twenty percent of patients with classical Hodgkin Lymphoma (cHL) have aggressive disease defined as relapsed or refractory disease to initial therapy. At present we cannot identify these patients pre-treatment. The microenvironment is very important in cHL because non-cancer cells constitute the majority of the cells in these tumors. Non-cancer intra-tumoral cells, such as tumor-associated macrophages (TAMs) have been shown to promote tumor growth in cHL via crosstalk with the cancer cells. Metabolic heterogeneity is defined as high mitochondrial metabolism in some tumor cells and glycolysis in others. We hypothesized that there are metabolic differences between cancer cells and non-cancer tumor cells, such as TAMs and tumor-infiltrating lymphocytes in cHL and that greater metabolic differences between cancer cells and TAMs are associated with poor outcomes. A case-control study was conducted with 22 tissue samples of cHL at diagnosis from a single institution. The case samples were from 11 patients with aggressive cHL who had relapsed after standard treatment with adriamycin, bleomycin, vinblastine, and dacarbazine (ABVD) or were refractory to this treatment. The control samples were from 11 patients with cHL who achieved a remission and never relapsed after ABVD. Reactive non-cancerous lymph nodes from four subjects served as additional controls. Samples were stained by immunohistochemistry for three metabolic markers: translocase of the outer mitochondrial membrane 20 (TOMM20), monocarboxylate transporter 1 (MCT1), and monocarboxylate transporter 4 (MCT4). TOMM20 is a marker of mitochondrial oxidative phosphorylation (OXPHOS) metabolism. Monocarboxylate transporter 1 (MCT1) is the main importer of lactate into cells and is a marker of OXPHOS. Monocarboxylate transporter 4 (MCT4) is the main lactate exporter out of cells and is a marker of glycolysis. The immunoreactivity for TOMM20, MCT1, and MCT4 was scored based on staining intensity and percentage of positive cells, as follows: 0 for no detectable staining in > 50% of cells; 1+ for faint to moderate staining in > 50% of cells, and 2+ for high or strong staining in > 50% of cells. TOMM20, MCT1, and MCT4 expression was significantly different in Hodgkin and Reed Sternberg (HRS) cells, which are the cancerous cells in cHL compared with TAMs and tumor-associated lymphocytes. HRS have high expression of TOMM20 and MCT1, while TAMs have absent expression of TOMM20 and MCT1 in all but two cases. Tumor-infiltrating lymphocytes have low TOMM20 expression and absent MCT1 expression. Conversely, high MCT4 expression was found in TAMs, but absent in HRS cells in all but one case. Tumor-infiltrating lymphocytes had absent MCT4 expression. Reactive lymph nodes in contrast to cHL tumors had low TOMM20, MCT1, and MCT4 expression in lymphocytes and macrophages. High TOMM20 and MCT1 expression in cancer cells with high MCT4 expression in TAMs is a signature of high metabolic heterogeneity between cancer cells and the tumor microenvironment. A high metabolic heterogeneity signature was associated with relapsed or refractory cHL with a hazard ratio of 5.87 (1.16-29.71; two-sided P < .05) compared with the low metabolic heterogeneity signature. Aggressive cHL exhibits features of metabolic heterogeneity with high mitochondrial metabolism in cancer cells and high glycolysis in TAMs, which is not seen in reactive lymph nodes. Future studies will need to confirm the value of these markers as prognostic and predictive biomarkers in clinical practice. Treatment intensity may be tailored in the future to the metabolic profile of the tumor microenvironment and drugs that target metabolic heterogeneity may be valuable in this disease. Copyright © 2017 Elsevier Inc. All rights reserved.

Altered Bioenergetics in Primary Dermal Fibroblasts from Adult Carriers of the FMR1 Premutation Before the Onset of the Neurodegenerative Disease Fragile X-Associated Tremor/Ataxia Syndrome.

PubMed

Napoli, Eleonora; Song, Gyu; Wong, Sarah; Hagerman, Randi; Giulivi, Cecilia

2016-10-01

Fragile X-associated tremor/ataxia syndrome (FXTAS) is a late onset neurodegenerative disorder, characterized by tremors, ataxia, impaired coordination, and cognitive decline. While all FXTAS individuals are carriers of a 55-200 CGG expansion at the 5'-UTR of the fragile X mental retardation gene (FMR1), also known as premutation, not all carriers develop FXTAS symptoms and some display other types of psychological/emotional disorders (e.g., autism, anxiety). The goal of this study was to investigate whether the mitochondrial dysfunction previously observed in fibroblasts from older premutation individuals (>60 years) was already present in younger (17-48 years), non-FXTAS-affected carriers and to identify the type and severity of the bioenergetic deficit. Since FXTAS affects mostly males, while females account for a small part of the FXTAS-affected population displaying less severe symptoms, only fibroblasts from males were evaluated in this study. Based on polarographic and enzymatic measurements, a generalized OXPHOS deficit was noted accompanied by increases in the matrix biomarker citrate synthase, oxidative stress (as increased mtDNA copy number and deletions), and mitochondrial network disruption/disorganization. Some of the outcomes (ATP-linked oxygen uptake, coupling, citrate synthase activity, and mitochondrial network organization) strongly correlated with the extent of the CGG expansion, with more severe deficits observed in cell lines carrying higher CGG number. Furthermore, mitochondrial outcomes can identify endophenotypes among carriers and are robust predictors of the premutation diagnosis before the onset of FXTAS, with the potential to be used as markers of prognosis and/or as readouts of pharmacological interventions.

Glycolysis, but not Mitochondria, responsible for intracellular ATP distribution in cortical area of podocytes.

PubMed

Ozawa, Shota; Ueda, Shuko; Imamura, Hiromi; Mori, Kiyoshi; Asanuma, Katsuhiko; Yanagita, Motoko; Nakagawa, Takahiko

2015-12-18

Differentiated podocytes, a type of renal glomerular cells, require substantial levels of energy to maintain glomerular physiology. Mitochondria and glycolysis are two major producers of ATP, but the precise roles of each in podocytes remain unknown. This study evaluated the roles of mitochondria and glycolysis in differentiated and differentiating podocytes. Mitochondria in differentiated podocytes are located in the central part of cell body while blocking mitochondria had minor effects on cell shape and migratory ability. In contrast, blocking glycolysis significantly reduced the formation of lamellipodia, a cortical area of these cells, decreased the cell migratory ability and induced the apoptosis. Consistently, the local ATP production in lamellipodia was predominantly regulated by glycolysis. In turn, synaptopodin expression was ameliorated by blocking either mitochondrial respiration or glycolysis. Similar to differentiated podocytes, the differentiating podocytes utilized the glycolysis for regulating apoptosis and lamellipodia formation while synaptopodin expression was likely involved in both mitochondrial OXPHOS and glycolysis. Finally, adult mouse podocytes have most of mitochondria predominantly in the center of the cytosol whereas phosphofructokinase, a rate limiting enzyme for glycolysis, was expressed in foot processes. These data suggest that mitochondria and glycolysis play parallel but distinct roles in differentiated and differentiating podocytes.

Glycolysis, but not Mitochondria, responsible for intracellular ATP distribution in cortical area of podocytes

PubMed Central

Ozawa, Shota; Ueda, Shuko; Imamura, Hiromi; Mori, Kiyoshi; Asanuma, Katsuhiko; Yanagita, Motoko; Nakagawa, Takahiko

2015-01-01

Differentiated podocytes, a type of renal glomerular cells, require substantial levels of energy to maintain glomerular physiology. Mitochondria and glycolysis are two major producers of ATP, but the precise roles of each in podocytes remain unknown. This study evaluated the roles of mitochondria and glycolysis in differentiated and differentiating podocytes. Mitochondria in differentiated podocytes are located in the central part of cell body while blocking mitochondria had minor effects on cell shape and migratory ability. In contrast, blocking glycolysis significantly reduced the formation of lamellipodia, a cortical area of these cells, decreased the cell migratory ability and induced the apoptosis. Consistently, the local ATP production in lamellipodia was predominantly regulated by glycolysis. In turn, synaptopodin expression was ameliorated by blocking either mitochondrial respiration or glycolysis. Similar to differentiated podocytes, the differentiating podocytes utilized the glycolysis for regulating apoptosis and lamellipodia formation while synaptopodin expression was likely involved in both mitochondrial OXPHOS and glycolysis. Finally, adult mouse podocytes have most of mitochondria predominantly in the center of the cytosol whereas phosphofructokinase, a rate limiting enzyme for glycolysis, was expressed in foot processes. These data suggest that mitochondria and glycolysis play parallel but distinct roles in differentiated and differentiating podocytes. PMID:26677804

Preimplantation death of xenomitochondrial mouse embryo harbouring bovine mitochondria

PubMed Central

Kawahara, Manabu; Koyama, Shiori; Iimura, Satomi; Yamazaki, Wataru; Tanaka, Aiko; Kohri, Nanami; Sasaki, Keisuke; Takahashi, Masashi

2015-01-01

Mitochondria, cellular organelles playing essential roles in eukaryotic cell metabolism, are thought to have evolved from bacteria. The organization of mtDNA is remarkably uniform across species, reflecting its vital and conserved role in oxidative phosphorylation (OXPHOS). Our objectives were to evaluate the compatibility of xenogeneic mitochondria in the development of preimplantation embryos in mammals. Mouse embryos harbouring bovine mitochondria (mtB-M embryos) were prepared by the cell-fusion technique employing the haemagglutinating virus of Japan (HVJ). The mtB-M embryos showed developmental delay at embryonic days (E) 3.5 after insemination. Furthermore, none of the mtB-M embryos could implant into the maternal uterus after embryo transfer, whereas control mouse embryos into which mitochondria from another mouse had been transferred developed as well as did non-manipulated embryos. When we performed quantitative PCR (qPCR) of mouse and bovine ND5, we found that the mtB-M embryos contained 8.3% of bovine mitochondria at the blastocyst stage. Thus, contamination with mitochondria from another species induces embryonic lethality prior to implantation into the maternal uterus. The heteroplasmic state of these xenogeneic mitochondria could have detrimental effects on preimplantation development, leading to preservation of species-specific mitochondrial integrity in mammals. PMID:26416548

Hypocapnic hypothesis of Leigh disease.

PubMed

Pronicka, Ewa

2017-04-01

Leigh syndrome (LS) is a neurogenetic disorder of children caused by mutations in at least 75 genes which impair mitochondrial bioenergetics. The changes have typical localization in basal ganglia and brainstem, and typical histological picture of spongiform appearance, vascular proliferation and gliosis. ATP deprivation, free radicals and lactate accumulation are suspected to be the causes. Hypocapnic hypothesis proposed in the paper questions the energy deprivation as the mechanism of LS. We assume that the primary harmful factor is hypocapnia (decrease in pCO 2 ) and respiratory alkalosis (increase in pH) due to hyperventilation, permanent or in response to stress. Inside mitochondria, the pH signal of high pH/low bicarbonate ion (HCO - 3 ) is transmitted by soluble adenyl cyclase (sAC) through cAMP dependent manner. The process can initiate brain lesions (necrosis, apoptosis, hypervascularity) in OXPHOS deficient cells residing at the LS area of the brain. The major message of the article is that it is not the ATP depletion but intracellular alkalization (and/or hyperoxia?) which seem to be the cause of LS. The paper includes suggestions concerning the methodology for further research on the LS mechanism and for therapeutic strategy. Copyright © 2017 The Author. Published by Elsevier Ltd.. All rights reserved.

Wnt modulates MCL1 to control cell survival in triple negative breast cancer

PubMed Central

2014-01-01

Background Triple negative breast cancer (TNBC) has higher rates of recurrence and distant metastasis, and poorer outcome as compared to non-TNBC. Aberrant activation of WNT signaling has been detected in TNBC, which might be important for triggering oncogenic conversion of breast epithelial cell. Therefore, we directed our focus on identifying the WNT ligand and its underlying mechanism in TNBC cells. Methods We performed large-scale analysis of public microarray data to screen the WNT ligands and the clinical significance of the responsible ligand in TNBC. WNT5B was identified and its overexpression in TNBC was confirmed by immunohistochemistry staining, Western blot and ELISA. ShRNA was used to knockdown WNT5B expression (shWNT5B). Cellular functional alteration with shWNT5B treatment was determined by using wound healing assay, mammosphere assay; while cell cycle and apoptosis were examined by flowcytometry. Mitochondrial morphology was photographed by electron microscope. Biological change of mitochondria was detected by RT-PCR and oxygen consumption assay. Activation of WNT pathway and its downstream targets were evaluated by liciferase assay, immunohistochemistry staining and immunoblot analysis. Statistical methods used in the experiments besides microarray analysis was two-tailed t-test. Results WNT5B was elevated both in the tumor and the patients’ serum. Suppression of WNT5B remarkably impaired cell growth, migration and mammosphere formation. Additionally, G0/G1 cell cycle arrest and caspase-independent apoptosis was observed. Study of the possible mechanism indicated that these effects occurred through suppression of mitochondrial biogenesis, as evidenced by reduced mitochondrial DNA (MtDNA) and compromised oxidative phosphorylation (OXPHOS). In Vivo and in vitro data uncovered that WNT5B modulated mitochondrial physiology was mediated by MCL1, which was regulated by WNT/β-catenin responsive gene, Myc. Clinic data analysis revealed that both WNT5B and MCL1 are associated with enhanced metastasis and decreased disease-free survival. Conclusions All our findings suggested that WNT5B/MCL1 cascade is critical for TNBC and understanding its regulatory apparatus provided valuable insight into the pathogenesis of the tumor development and the guidance for targeting therapeutics. PMID:24564888

Voluntary Running Aids to Maintain High Body Temperature in Rats Bred for High Aerobic Capacity

PubMed Central

Karvinen, Sira M.; Silvennoinen, Mika; Ma, Hongqiang; Törmäkangas, Timo; Rantalainen, Timo; Rinnankoski-Tuikka, Rita; Lensu, Sanna; Koch, Lauren G.; Britton, Steven L.; Kainulainen, Heikki

2016-01-01

The production of heat, i.e., thermogenesis, is a significant component of the metabolic rate, which in turn affects weight gain and health. Thermogenesis is linked to physical activity (PA) level. However, it is not known whether intrinsic exercise capacity, aging, and long-term voluntary running affect core body temperature. Here we use rat models selectively bred to differ in maximal treadmill endurance running capacity (Low capacity runners, LCR and High capacity Runners, HCR), that as adults are divergent for aerobic exercise capacity, aging, and metabolic disease risk to study the connection between PA and body temperature. Ten high capacity runner (HCR) and ten low capacity runner (LCR) female rats were studied between 9 and 21 months of age. Rectal body temperature of HCR and LCR rats was measured before and after 1-year voluntary running/control intervention to explore the effects of aging and PA. Also, we determined whether injected glucose and spontaneous activity affect the body temperature differently between LCR and HCR rats at 9 vs. 21 months of age. HCRs had on average 1.3°C higher body temperature than LCRs (p < 0.001). Aging decreased the body temperature level of HCRs to similar levels with LCRs. The opportunity to run voluntarily had a significant impact on the body temperature of HCRs (p < 0.001) allowing them to maintain body temperature at a similar level as when at younger age. Compared to LCRs, HCRs were spontaneously more active, had higher relative gastrocnemius muscle mass and higher UCP2, PGC-1α, cyt c, and OXPHOS levels in the skeletal muscle (p < 0.050). These results suggest that higher PA level together with greater relative muscle mass and higher mitochondrial content/function contribute to the accumulation of heat in the HCRs. Interestingly, neither aging nor voluntary training had a significant impact on core body temperature of LCRs. However, glucose injection resulted in a lowering of the body temperature of LCRs (p < 0.050), but not that of HCRs. In conclusion, rats born with high intrinsic capacity for aerobic exercise and better health have higher body temperature compared to rats born with low exercise capacity and disease risk. Voluntary running allowed HCRs to maintain high body temperature during aging, which suggests that high PA level was crucial in maintaining the high body temperature of HCRs. PMID:27504097

Voluntary Running Aids to Maintain High Body Temperature in Rats Bred for High Aerobic Capacity.

PubMed

Karvinen, Sira M; Silvennoinen, Mika; Ma, Hongqiang; Törmäkangas, Timo; Rantalainen, Timo; Rinnankoski-Tuikka, Rita; Lensu, Sanna; Koch, Lauren G; Britton, Steven L; Kainulainen, Heikki

2016-01-01

The production of heat, i.e., thermogenesis, is a significant component of the metabolic rate, which in turn affects weight gain and health. Thermogenesis is linked to physical activity (PA) level. However, it is not known whether intrinsic exercise capacity, aging, and long-term voluntary running affect core body temperature. Here we use rat models selectively bred to differ in maximal treadmill endurance running capacity (Low capacity runners, LCR and High capacity Runners, HCR), that as adults are divergent for aerobic exercise capacity, aging, and metabolic disease risk to study the connection between PA and body temperature. Ten high capacity runner (HCR) and ten low capacity runner (LCR) female rats were studied between 9 and 21 months of age. Rectal body temperature of HCR and LCR rats was measured before and after 1-year voluntary running/control intervention to explore the effects of aging and PA. Also, we determined whether injected glucose and spontaneous activity affect the body temperature differently between LCR and HCR rats at 9 vs. 21 months of age. HCRs had on average 1.3°C higher body temperature than LCRs (p < 0.001). Aging decreased the body temperature level of HCRs to similar levels with LCRs. The opportunity to run voluntarily had a significant impact on the body temperature of HCRs (p < 0.001) allowing them to maintain body temperature at a similar level as when at younger age. Compared to LCRs, HCRs were spontaneously more active, had higher relative gastrocnemius muscle mass and higher UCP2, PGC-1α, cyt c, and OXPHOS levels in the skeletal muscle (p < 0.050). These results suggest that higher PA level together with greater relative muscle mass and higher mitochondrial content/function contribute to the accumulation of heat in the HCRs. Interestingly, neither aging nor voluntary training had a significant impact on core body temperature of LCRs. However, glucose injection resulted in a lowering of the body temperature of LCRs (p < 0.050), but not that of HCRs. In conclusion, rats born with high intrinsic capacity for aerobic exercise and better health have higher body temperature compared to rats born with low exercise capacity and disease risk. Voluntary running allowed HCRs to maintain high body temperature during aging, which suggests that high PA level was crucial in maintaining the high body temperature of HCRs.

Functionalized active-nucleus complex sensor

DOEpatents

Pines, Alexander; Wemmer, David E.; Spence, Megan; Rubin, Seth

2003-11-25

A functionalized active-nucleus complex sensor that selectively associates with one or more target species, and a method for assaying and screening for one or a plurality of target species utilizing one or a plurality of functionalized active-nucleus complexes with at least two of the functionalized active-nucleus complexes having an attraction affinity to different corresponding target species. The functionalized active-nucleus complex has an active-nucleus and a targeting carrier. The method involves functionalizing an active-nucleus, for each functionalized active-nucleus complex, by incorporating the active-nucleus into a macromolucular or molecular complex that is capable of binding one of the target species and then bringing the macromolecular or molecular complexes into contact with the target species and detecting the occurrence of or change in a nuclear magnetic resonance signal from each of the active-nuclei in each of the functionalized active-nucleus complexes.

Acetylation of histone deacetylase 1 regulates NuRD corepressor complex activity.

PubMed

Yang, Tao; Jian, Wei; Luo, Yi; Fu, Xueqi; Noguchi, Constance; Bungert, Jörg; Huang, Suming; Qiu, Yi

2012-11-23

HDAC1-containing NuRD complex is required for GATA-1-mediated repression and activation. GATA-1 associated with acetylated HDAC1-containing NuRD complex, which has no deacetylase activity, for gene activation. Acetylated HDAC1 converts NuRD complex from a repressor to an activator during GATA-1-directed erythroid differentiation program. HDAC1 acetylation may function as a master regulator for the activity of HDAC1 containing complexes. Histone deacetylases (HDACs) play important roles in regulating cell proliferation and differentiation. The HDAC1-containing NuRD complex is generally considered as a corepressor complex and is required for GATA-1-mediated repression. However, recent studies also show that the NuRD complex is involved in GATA-1-mediated gene activation. We tested whether the GATA-1-associated NuRD complex loses its deacetylase activity and commits the GATA-1 complex to become an activator during erythropoiesis. We found that GATA-1-associated deacetylase activity gradually decreased upon induction of erythroid differentiation. GATA-1-associated HDAC1 is increasingly acetylated after differentiation. It has been demonstrated earlier that acetylated HDAC1 has no deacetylase activity. Indeed, overexpression of an HDAC1 mutant, which mimics acetylated HDAC1, promotes GATA-1-mediated transcription and erythroid differentiation. Furthermore, during erythroid differentiation, acetylated HDAC1 recruitment is increased at GATA-1-activated genes, whereas it is significantly decreased at GATA-1-repressed genes. Interestingly, deacetylase activity is not required for Mi2 remodeling activity, suggesting that remodeling activity may be required for both activation and repression. Thus, our data suggest that NuRD can function as a coactivator or repressor and that acetylated HDAC1 converts the NuRD complex from a repressor to an activator during GATA-1-directed erythroid differentiation.

Transcriptomic meta-analysis identifies gene expression characteristics in various samples of HIV-infected patients with nonprogressive disease.

PubMed

Zhang, Le-Le; Zhang, Zi-Ning; Wu, Xian; Jiang, Yong-Jun; Fu, Ya-Jing; Shang, Hong

2017-09-12

A small proportion of HIV-infected patients remain clinically and/or immunologically stable for years, including elite controllers (ECs) who have undetectable viremia (<50 copies/ml) and long-term nonprogressors (LTNPs) who maintain normal CD4 + T cell counts for prolonged periods (>10 years). However, the mechanism of nonprogression needs to be further resolved. In this study, a transcriptome meta-analysis was performed on nonprogressor and progressor microarray data to identify differential transcriptome pathways and potential biomarkers. Using the INMEX (integrative meta-analysis of expression data) program, we performed the meta-analysis to identify consistently differentially expressed genes (DEGs) in nonprogressors and further performed functional interpretation (gene ontology analysis and pathway analysis) of the DEGs identified in the meta-analysis. Five microarray datasets (81 cases and 98 controls in total), including whole blood, CD4 + and CD8 + T cells, were collected for meta-analysis. We determined that nonprogressors have reduced expression of important interferon-stimulated genes (ISGs), CD38, lymphocyte activation gene 3 (LAG-3) in whole blood, CD4 + and CD8 + T cells. Gene ontology (GO) analysis showed a significant enrichment in DEGs that function in the type I interferon signaling pathway. Upregulated pathways, including the PI3K-Akt signaling pathway in whole blood, cytokine-cytokine receptor interaction in CD4 + T cells and the MAPK signaling pathway in CD8 + T cells, were identified in nonprogressors compared with progressors. In each metabolic functional category, the number of downregulated DEGs was more than the upregulated DEGs, and almost all genes were downregulated DEGs in the oxidative phosphorylation (OXPHOS) and tricarboxylic acid (TCA) cycle in the three types of samples. Our transcriptomic meta-analysis provides a comprehensive evaluation of the gene expression profiles in major blood types of nonprogressors, providing new insights in the understanding of HIV pathogenesis and developing strategies to delay HIV disease progression.

Large-scale patterns formed by solar active regions during the ascending phase of cycle 21

NASA Astrophysics Data System (ADS)

Gaizauskas, V.; Harvey, K. L.; Harvey, J. W.; Zwaan, C.

1983-02-01

Synoptic maps of photospheric magnetic fields prepared at the Kitt Peak National Observatory are used in investigating large-scale patterns in the spatial and temporal distribution of solar active regions for 27 solar rotations between 1977 and 1979. The active regions are found to be distributed in 'complexes of activity' (Bumba and Howard, 1965). With the working definition of a complex of activity based on continuity and proximity of the constituent active regions, the phenomenology of complexes is explored. It is found that complexes of activity form within one month and that they are typically maintained for 3 to 6 solar rotations by fresh injections of magnetic flux. During the active lifetime of a complex of activity, the total magnetic flux in the complex remains steady to within a factor of 2. The magnetic polarities are closely balanced, and each complex rotates about the sun at its own special, constant rate. In certain cases, the complexes form two diverging branches.

Bioactive ruthenium(II)-arene complexes containing modified 18β-glycyrrhetinic acid ligands.

PubMed

Kong, Yaqiong; Chen, Feng; Su, Zhi; Qian, Yong; Wang, Fang-Xin; Wang, Xiuxiu; Zhao, Jing; Mao, Zong-Wan; Liu, Hong-Ke

2018-05-01

Metal-arene complexes containing bioactive natural-product derived ligands can have new and unusual properties. We report the synthesis, characterization and antiproliferative activity of two new Ru(II) arene complexes with imidazole (dichlorido complex 1) or bipyridyl (chlorido complex 2) ligands conjugated to 18β-glycyrrhetinic acid, an active triterpenoid metabolite of Glycyrrhiza glabra. In general, the conjugated ligands and complexes showed only moderate activity against HeLa (cervical), MCF-7 (breast) and A2780 (ovarian) cancer cells, although the activity of complex 2 in the former two cell lines approached that of the drug cisplatin. Complex 2 (in contrast to complex 1) also exhibited significant activity towards both Gram-positive S. aureus and Gram-negative E. coil bacteria. Complex 2 can induce condensation of DNA and enhances the generation of intracellular reactive oxygen species (ROS). The conjugation of natural products to ligands in organometallic half-sandwich complexes provides a strategy to enhance their biological activities. Copyright © 2018 Elsevier Inc. All rights reserved.

Diagnostic value of succinate ubiquinone reductase activity in the identification of patients with mitochondrial DNA depletion.

PubMed

Hargreaves, P; Rahman, S; Guthrie, P; Taanman, J W; Leonard, J V; Land, J M; Heales, S J R

2002-02-01

Mitochondrial DNA (mtDNA) depletion syndrome (McKusick 251880) is characterized by a progressive quantitative loss of mtDNA resulting in severe mitochondrial dysfunction. A diagnosis of mtDNA depletion can only be confirmed after Southern blot analysis of affected tissue. Only a limited number of centres have the facilities to offer this service, and this is frequently on an irregular basis. There is therefore a need for a test that can refine sample selection as well as complementing the molecular analysis. In this study we compared the activities of the nuclear-encoded succinate ubiquinone reductase (complex II) to the activities of the combined mitochondrial and nuclear-encoded mitochondrial electron transport chain (ETC) complexes; NADH:ubiquinone reductase (complex I), ubiquinol-cytochrome-c reductase (complex III), and cytochrome-c oxidase (complex IV), in skeletal muscle biopsies from 7 patients with confirmed mtDNA depletion. In one patient there was no evidence of an ETC defect. However, the remaining 6 patients exhibited reduced complex I and IV activities. Five of these patients also displayed reduced complex II-III (succinate:cytochrome-c reductase) activity. Individual measurement of complex II and complex III activities demonstrated normal levels of complex II activity compared to complex III, which was reduced in the 5 biopsies assayed. These findings suggest a possible diagnostic value for the detection of normal levels of complex II activity in conjunction with reduced complex I, III and IV activity in the identification of likely candidates for mtDNA depletion syndrome

Regulation of branched-chain amino acid catabolism in rat models for spontaneous type 2 diabetes mellitus.

PubMed

Kuzuya, Teiji; Katano, Yoshiaki; Nakano, Isao; Hirooka, Yoshiki; Itoh, Akihiro; Ishigami, Masatoshi; Hayashi, Kazuhiko; Honda, Takashi; Goto, Hidemi; Fujita, Yuko; Shikano, Rie; Muramatsu, Yuji; Bajotto, Gustavo; Tamura, Tomohiro; Tamura, Noriko; Shimomura, Yoshiharu

2008-08-15

The branched-chain alpha-keto acid dehydrogenase (BCKDH) complex is the most important regulatory enzyme in branched-chain amino acid (BCAA) catabolism. We examined the regulation of hepatic BCKDH complex activity in spontaneous type 2 diabetes Otsuka Long-Evans Tokushima Fatty (OLETF) rats and Zucker diabetic fatty rats. Hepatic BCKDH complex activity in these rats was significantly lower than in corresponding control rats. The amount of BCKDH complex in OLETF rats corresponded to the total activity of the complex. Activity and abundance of the bound form of BCKDH kinase, which is responsible for inactivation of the complex, showed an inverse correlation to BCKDH complex activity in OLETF rats. Dietary supplementation of 5% BCAAs for 10 weeks markedly increased BCKDH complex activity, and decreased the activity and bound form of BCKDH kinase in the rats. These results suggest that BCAA catabolism in type 2 diabetes is downregulated and enhanced by BCAA supplementation.

A Mitochondrial Paradigm of Metabolic and Degenerative Diseases, Aging, and Cancer: A Dawn for Evolutionary Medicine

PubMed Central

Wallace, Douglas C.

2005-01-01

Life is the interplay between structure and energy, yet the role of energy deficiency in human disease has been poorly explored by modern medicine. Since the mitochondria use oxidative phosphorylation (OXPHOS) to convert dietary calories into usable energy, generating reactive oxygen species (ROS) as a toxic by-product, I hypothesize that mitochondrial dysfunction plays a central role in a wide range of age-related disorders and various forms of cancer. Because mitochondrial DNA (mtDNA) is present in thousands of copies per cell and encodes essential genes for energy production, I propose that the delayed-onset and progressive course of the age-related diseases results from the accumulation of somatic mutations in the mtDNAs of post-mitotic tissues. The tissue-specific manifestations of these diseases may result from the varying energetic roles and needs of the different tissues. The variation in the individual and regional predisposition to degenerative diseases and cancer may result from the interaction of modern dietary caloric intake and ancient mitochondrial genetic polymorphisms. Therefore the mitochondria provide a direct link between our environment and our genes and the mtDNA variants that permitted our forbears to energetically adapt to their ancestral homes are influencing our health today. PMID:16285865

The phasor-FLIM fingerprints reveal shifts from OXPHOS to enhanced glycolysis in Huntington Disease

PubMed Central

Sameni, Sara; Syed, Adeela; Marsh, J. Lawrence; Digman, Michelle A.

2016-01-01

Huntington disease (HD) is an autosomal neurodegenerative disorder caused by the expansion of Polyglutamine (polyQ) in exon 1 of the Huntingtin protein. Glutamine repeats below 36 are considered normal while repeats above 40 lead to HD. Impairment in energy metabolism is a common trend in Huntington pathogenesis; however, this effect is not fully understood. Here, we used the phasor approach and Fluorescence Lifetime Imaging Microscopy (FLIM) to measure changes between free and bound fractions of NADH as a indirect measure of metabolic alteration in living cells. Using Phasor-FLIM, pixel maps of metabolic alteration in HEK293 cell lines and in transgenic Drosophila expressing expanded and unexpanded polyQ HTT exon1 in the eye disc were developed. We found a significant shift towards increased free NADH, indicating an increased glycolytic state for cells and tissues expressing the expanded polyQ compared to unexpanded control. In the nucleus, a further lifetime shift occurs towards higher free NADH suggesting a possible synergism between metabolic dysfunction and transcriptional regulation. Our results indicate that metabolic dysfunction in HD shifts to increased glycolysis leading to oxidative stress and cell death. This powerful label free method can be used to screen native HD tissue samples and for potential drug screening. PMID:27713486

The phasor-FLIM fingerprints reveal shifts from OXPHOS to enhanced glycolysis in Huntington Disease

NASA Astrophysics Data System (ADS)

Sameni, Sara; Syed, Adeela; Marsh, J. Lawrence; Digman, Michelle A.

2016-10-01

Huntington disease (HD) is an autosomal neurodegenerative disorder caused by the expansion of Polyglutamine (polyQ) in exon 1 of the Huntingtin protein. Glutamine repeats below 36 are considered normal while repeats above 40 lead to HD. Impairment in energy metabolism is a common trend in Huntington pathogenesis; however, this effect is not fully understood. Here, we used the phasor approach and Fluorescence Lifetime Imaging Microscopy (FLIM) to measure changes between free and bound fractions of NADH as a indirect measure of metabolic alteration in living cells. Using Phasor-FLIM, pixel maps of metabolic alteration in HEK293 cell lines and in transgenic Drosophila expressing expanded and unexpanded polyQ HTT exon1 in the eye disc were developed. We found a significant shift towards increased free NADH, indicating an increased glycolytic state for cells and tissues expressing the expanded polyQ compared to unexpanded control. In the nucleus, a further lifetime shift occurs towards higher free NADH suggesting a possible synergism between metabolic dysfunction and transcriptional regulation. Our results indicate that metabolic dysfunction in HD shifts to increased glycolysis leading to oxidative stress and cell death. This powerful label free method can be used to screen native HD tissue samples and for potential drug screening.

mtDNA Mutations and Their Role in Aging, Diseases and Forensic Sciences

PubMed Central

Zapico, Sara C.; Ubelaker, Douglas H.

2013-01-01

Mitochondria are independent organelles with their own DNA. As a primary function, mitochondria produce the energy for the cell through Oxidative Phosphorylation (OXPHOS) in the Electron Transport Chain (ETC). One of the toxic products of this process is Reactive Oxygen Species (ROS), which can induce oxidative damage in macromolecules like lipids, proteins and DNA. Mitochondrial DNA (mtDNA) is less protected and has fewer reparation mechanisms than nuclear DNA (nDNA), and as such is more exposed to oxidative, mutation-inducing damage. This review analyzes the causes and consequences of mtDNA mutations and their relationship with the aging process. Neurodegenerative diseases, related with the aging, are consequences of mtDNA mutations resulting in a decrease in mitochondrial function. Also described are “mitochondrial diseases”, pathologies produced by mtDNA mutations and whose symptoms are related with mitochondrial dysfunction. Finally, mtDNA haplogroups are defined in this review; these groups are important for determination of geographical origin of an individual. Additionally, different haplogroups exhibit variably longevity and risk of certain diseases. mtDNA mutations in aging and haplogroups are of special interest to forensic science research. Therefore this review will help to clarify the key role of mtDNA mutations in these processes and support further research in this area. PMID:24307969

Selective inhibition of deactivated mitochondrial complex I by biguanides.

PubMed

Matsuzaki, Satoshi; Humphries, Kenneth M

2015-03-24

Biguanides are widely used antihyperglycemic agents for diabetes mellitus and prediabetes treatment. Complex I is the rate-limiting step of the mitochondrial electron transport chain (ETC), a major source of mitochondrial free radical production, and a known target of biguanides. Complex I has two reversible conformational states, active and de-active. The deactivated state is promoted in the absence of substrates but is rapidly and fully reversed to the active state in the presence of NADH. The objective of this study was to determine the relative sensitivity of active/de-active complex I to biguanide-mediated inhibition and resulting superoxide radical (O₂(•⁻)) production. Using isolated rat heart mitochondria, we show that deactivation of complex I sensitizes it to metformin and phenformin (4- and 3-fold, respectively), but not to other known complex I inhibitors, such as rotenone. Mitochondrial O₂(•⁻) production by deactivated complex I was measured fluorescently by NADH-dependent 2-hydroxyethidium formation at alkaline pH to impede reactivation. Superoxide production was 260.4% higher than in active complex I at pH 9.4. However, phenformin treatment of de-active complex I decreased O₂(•⁻) production by 14.9%, while rotenone increased production by 42.9%. Mitochondria isolated from rat hearts subjected to cardiac ischemia, a condition known to induce complex I deactivation, were sensitized to phenformin-mediated complex I inhibition. This supports the idea that the effects of biguanides are likely to be influenced by the complex I state in vivo. These results demonstrate that the complex I active and de-active states are a determinant in biguanide-mediated inhibition.

Mitochondrial Complex 1 Activity Measured by Spectrophotometry Is Reduced across All Brain Regions in Ageing and More Specifically in Neurodegeneration.

PubMed

Pollard, Amelia Kate; Craig, Emma Louise; Chakrabarti, Lisa

2016-01-01

Mitochondrial function, in particular complex 1 of the electron transport chain (ETC), has been shown to decrease during normal ageing and in neurodegenerative disease. However, there is some debate concerning which area of the brain has the greatest complex 1 activity. It is important to identify the pattern of activity in order to be able to gauge the effect of age or disease related changes. We determined complex 1 activity spectrophotometrically in the cortex, brainstem and cerebellum of middle aged mice (70-71 weeks), a cerebellar ataxic neurodegeneration model (pcd5J) and young wild type controls. We share our updated protocol on the measurements of complex1 activity and find that mitochondrial fractions isolated from frozen tissues can be measured for robust activity. We show that complex 1 activity is clearly highest in the cortex when compared with brainstem and cerebellum (p<0.003). Cerebellum and brainstem mitochondria exhibit similar levels of complex 1 activity in wild type brains. In the aged brain we see similar levels of complex 1 activity in all three-brain regions. The specific activity of complex 1 measured in the aged cortex is significantly decreased when compared with controls (p<0.0001). Both the cerebellum and brainstem mitochondria also show significantly reduced activity with ageing (p<0.05). The mouse model of ataxia predictably has a lower complex 1 activity in the cerebellum, and although reductions are measured in the cortex and brain stem, the remaining activity is higher than in the aged brains. We present clear evidence that complex 1 activity decreases across the brain with age and much more specifically in the cerebellum of the pcd5j mouse. Mitochondrial impairment can be a region specific phenomenon in disease, but in ageing appears to affect the entire brain, abolishing the pattern of higher activity in cortical regions.

Selective Inhibition of Deactivated Mitochondrial Complex I by Biguanides †

PubMed Central

Matsuzaki, Satoshi; Humphries, Kenneth M.

2015-01-01

Biguanides are widely used antihyperglycemic agents for diabetes mellitus and prediabetes treatment. Complex I is the rate limiting step of the mitochondrial electron transport chain (ETC), a major source of mitochondrial free radical production, and a known target of biguanides. Complex I has two reversible conformational states, active and de-active. The deactivated state is promoted in the absence of substrates, but is rapidly and fully reversed to the active state in the presence of NADH. The objective of this study was to determine the relative sensitivity of active/de-active complex I to biguanide-mediated inhibition and resulting superoxide radical (O2•−) production. Using isolated rat heart mitochondria, we show that deactivation of complex I sensitizes it to metformin and phenformin (4- and 3-fold, respectively), but not to other known complex I inhibitors, such as rotenone. Mitochondrial O2•− production by deactivated complex I was measured fluorescently by the NADH-dependent 2-hydroxyethidium formation at alkaline pH to impede reactivation. Superoxide production was 260.4% higher than in active complex I at pH 9.4. However, phenformin treatment of de-active complex I decreased O2•− production by 14.9% while rotenone increased production by 42.9%. Mitochondria isolated from rat hearts subjected to cardiac ischemia, a condition known to induce complex I deactivation, were sensitized to phenformin:mediated complex I inhibition. This supports that the effects of biguanides are likely to be influenced by the complex I state in vivo. These results demonstrate that the complex I active/de-active states are a determinant in biguanide-mediated inhibition. PMID:25719498

Synthesis, structural elucidation, biological, antioxidant and nuclease activities of some 5-Fluorouracil-amino acid mixed ligand complexes

NASA Astrophysics Data System (ADS)

Shobana, Sutha; Subramaniam, Perumal; Mitu, Liviu; Dharmaraja, Jeyaprakash; Arvind Narayan, Sundaram

2015-01-01

Some biologically active mixed ligand complexes (1-9) have been synthesized from 5-Fluorouracil (5-FU; A) and amino acids (B) such as glycine (gly), L-alanine (ala) and L-valine (val) with Ni(II), Cu(II) and Zn(II) ions. The synthesized mixed ligand complexes (1-9) were characterized by various physico-chemical, spectral, thermal and morphological studies. 5-Fluorouracil and its mixed ligand complexes have been tested for their in vitro biological activities against some pathogenic bacterial and fungal species by the agar well diffusion method. The in vitro antioxidant activities of 5-Fluorouracil and its complexes have also been investigated by using the DPPH assay method. The results demonstrate that Cu(II) mixed ligand complexes (4-6) exhibit potent biological as well as antioxidant activities compared to 5-Fluorouracil and Ni(II) (1-3) and Zn(II) (7-9) mixed ligand complexes. Further, the cleaving activities of CT DNA under aerobic conditions show moderate activity with the synthesized Cu(II) and Ni(II) mixed ligand complexes (1-6) while no activity is seen with Zn(II) complexes (7-9). Binding studies of CT DNA with these complexes show a decrease in intensity of the charge transfer band to the extent of 5-15% along with a minor red shift. The free energy change values (Δ‡G) calculated from intrinsic binding constants indicate that the interaction between mixed ligand complex and DNA is spontaneous.

Comparative analysis of activator-Eσ54 complexes formed with nucleotide-metal fluoride analogues

PubMed Central

Burrows, Patricia C.; Joly, Nicolas; Nixon, B. Tracy; Buck, Martin

2009-01-01

Bacterial RNA polymerase (RNAP) containing the major variant σ54 factor forms open promoter complexes in a reaction in which specialized activator proteins hydrolyse ATP. Here we probe binding interactions between σ54-RNAP (Eσ54) and the ATPases associated with various cellular activities (AAA+) domain of the Escherichia coli activator protein, PspF, using nucleotide-metal fluoride (BeF and AlF) analogues representing ground and transition states of ATP, which allow complexes (that are otherwise too transient with ATP) to be captured. We show that the organization and functionality of the ADP–BeF- and ADP–AlF-dependent complexes greatly overlap. Our data support an activation pathway in which the initial ATP-dependent binding of the activator to the Eσ54 closed complex results in the re-organization of Eσ54 with respect to the transcription start-site. However, the nucleotide-dependent binding interactions between the activator and the Eσ54 closed complex are in themselves insufficient for forming open promoter complexes when linear double-stranded DNA is present in the initial closed complex. PMID:19553192

Prefrontal and parietal activity is modulated by the rule complexity of inductive reasoning and can be predicted by a cognitive model.

PubMed

Jia, Xiuqin; Liang, Peipeng; Shi, Lin; Wang, Defeng; Li, Kuncheng

2015-01-01

In neuroimaging studies, increased task complexity can lead to increased activation in task-specific regions or to activation of additional regions. How the brain adapts to increased rule complexity during inductive reasoning remains unclear. In the current study, three types of problems were created: simple rule induction (i.e., SI, with rule complexity of 1), complex rule induction (i.e., CI, with rule complexity of 2), and perceptual control. Our findings revealed that increased activations accompany increased rule complexity in the right dorsal lateral prefrontal cortex (DLPFC) and medial posterior parietal cortex (precuneus). A cognitive model predicted both the behavioral and brain imaging results. The current findings suggest that neural activity in frontal and parietal regions is modulated by rule complexity, which may shed light on the neural mechanisms of inductive reasoning. Copyright © 2014. Published by Elsevier Ltd.

Septin 7 reduces nonmuscle myosin IIA activity in the SNAP23 complex and hinders GLUT4 storage vesicle docking and fusion

DOE Office of Scientific and Technical Information (OSTI.GOV)

Wasik, Anita A.; Dumont, Vincent; Tienari, Jukka

Glomerular epithelial cells, podocytes, are insulin responsive and can develop insulin resistance. Here, we demonstrate that the small GTPase septin 7 forms a complex with nonmuscle myosin heavy chain IIA (NMHC-IIA; encoded by MYH9), a component of the nonmuscle myosin IIA (NM-IIA) hexameric complex. We observed that knockdown of NMHC-IIA decreases insulin-stimulated glucose uptake into podocytes. Both septin 7 and NM-IIA associate with SNAP23, a SNARE protein involved in GLUT4 storage vesicle (GSV) docking and fusion with the plasma membrane. We observed that insulin decreases the level of septin 7 and increases the activity of NM-IIA in the SNAP23 complex,more » as visualized by increased phosphorylation of myosin regulatory light chain. Also knockdown of septin 7 increases the activity of NM-IIA in the complex. The activity of NM-IIA is increased in diabetic rat glomeruli and cultured human podocytes exposed to macroalbuminuric sera from patients with type 1 diabetes. Collectively, the data suggest that the activity of NM-IIA in the SNAP23 complex plays a key role in insulin-stimulated glucose uptake into podocytes. Furthermore, we observed that septin 7 reduces the activity of NM-IIA in the SNAP23 complex and thereby hinders GSV docking and fusion with the plasma membrane. - Highlights: • Septin 7, nonmuscle myosin heavy chain IIA (NMHC-IIA) and SNAP23 form a complex. • Knockdown of septin 7 increases NM-IIA activity in the SNAP23 complex. • Insulin decreases septin 7 level and increases NM-IIA activity in the SNAP23 complex. • Septin 7 hinders GSV docking/fusion by reducing NM-IIA activity in the SNAP23 complex.« less

A Rhodium(III) Complex as an Inhibitor of Neural Precursor Cell Expressed, Developmentally Down-Regulated 8-Activating Enzyme with in Vivo Activity against Inflammatory Bowel Disease.

PubMed

Zhong, Hai-Jing; Wang, Wanhe; Kang, Tian-Shu; Yan, Hui; Yang, Yali; Xu, Lipeng; Wang, Yuqiang; Ma, Dik-Lung; Leung, Chung-Hang

2017-01-12

We report herein the identification of the rhodium(III) complex [Rh(phq) 2 (MOPIP)] + (1) as a potent and selective ATP-competitive neural precursor cell expressed, developmentally down-regulated 8 (NEDD8)-activating enzyme (NAE) inhibitor. Structure-activity relationship analysis indicated that the overall organometallic design of complex 1 was important for anti-inflammatory activity. Complex 1 showed promising anti-inflammatory activity in vivo for the potential treatment of inflammatory bowel disease.

Ursolic Acid Hydrazide Based Organometallic Complexes: Synthesis, Characterization, Antibacterial, Antioxidant and Docking Studies

NASA Astrophysics Data System (ADS)

Jabeen, Muafia; Ahmad, Sajjad; Shahid, Khadija; Sadiq, Abdul; Rashid, Umer

2018-03-01

In the current research work,eleven metal complexes were synthesized from the hydrazide derivative of ursolic acid. Metal complexes of tin, antimony and iron were synthesized and characterized by FT-IR and NMR spectroscopy. The antibacterial and antioxidant activities were performed for these complexes, which revealed that the metal complexes synthesized are more potent than their parent compounds. We observed that antioxidant activity showed by triphenyltin complex was significant and least activity have been shown by antimony trichloride complex.The synthesized metal complexes were then evaluated against two Gram-negative and two Gram-positive bacterial strains. Triphenyl tin complex emerged as potent antibacterial agent with MIC value of 8 μg/ml each against Shigellaspp, S. typhi and S. aureus. While, the MIC value againstS. pneumoniae is 4 μg/ml.Computational docking studies were carried out on molecular targets to interpret the results of antioxidant and antibacterial activities. Based on the results, it may be inferred that the metal complexes of ursolic acid are more active as compared to the parent drug and may be proved for some other pharmacological potential by further analysis.

Synthesis, characterization, cytotoxic and antitubercular activities of new gold(I) and gold(III) complexes containing ligands derived from carbohydrates.

PubMed

Chaves, Joana Darc Souza; Damasceno, Jaqueline Lopes; Paula, Marcela Cristina Ferreira; de Oliveira, Pollyanna Francielli; Azevedo, Gustavo Chevitarese; Matos, Renato Camargo; Lourenço, Maria Cristina S; Tavares, Denise Crispim; Silva, Heveline; Fontes, Ana Paula Soares; de Almeida, Mauro Vieira

2015-10-01

Novel gold(I) and gold(III) complexes containing derivatives of D-galactose, D-ribose and D-glucono-1,5-lactone as ligands were synthesized and characterized by IR, (1)H, and (13)C NMR, high resolution mass spectra and cyclic voltammetry. The compounds were evaluated in vitro for their cytotoxicity against three types of tumor cells: cervical carcinoma (HeLa) breast adenocarcinoma (MCF-7) and glioblastoma (MO59J) and one non-tumor cell line: human lung fibroblasts (GM07492A). Their antitubercular activity was evaluated as well expressed as the minimum inhibitory concentration (MIC90) in μg/mL. In general, the gold(I) complexes were more active than gold(III) complexes, for example, the gold(I) complex (1) was about 8.8 times and 7.6 times more cytotoxic than gold(III) complex (8) in MO59J and MCF-7 cells, respectively. Ribose and alkyl phosphine derivative complexes were more active than galactose and aryl phosphine complexes. The presence of a thiazolidine ring did not improve the cytotoxicity. The study of the cytotoxic activity revealed effective antitumor activities for the gold(I) complexes, being more active than cisplatin in all the tested tumor cell lines. Gold(I) compounds (1), (2), (3), (4) and (6) exhibited relevant antitubercular activity even when compared with first line drugs such as rifampicin.

Abi1 is essential for the formation and activation of a WAVE2 signalling complex.

PubMed

Innocenti, Metello; Zucconi, Adriana; Disanza, Andrea; Frittoli, Emanuela; Areces, Liliana B; Steffen, Anika; Stradal, Theresia E B; Di Fiore, Pier Paolo; Carlier, Marie-France; Scita, Giorgio

2004-04-01

WAVE2 belongs to a family of proteins that mediates actin reorganization by relaying signals from Rac to the Arp2/3 complex, resulting in lamellipodia protrusion. WAVE2 displays Arp2/3-dependent actin nucleation activity in vitro, and does not bind directly to Rac. Instead, it forms macromolecular complexes that have been reported to exert both positive and negative modes of regulation. How these complexes are assembled, localized and activated in vivo remains to be established. Here we use tandem mass spectrometry to identify an Abi1-based complex containing WAVE2, Nap1 (Nck-associated protein) and PIR121. Abi1 interacts directly with the WHD domain of WAVE2, increases WAVE2 actin polymerization activity and mediates the assembly of a WAVE2-Abi1-Nap1-PIR121 complex. The WAVE2-Abi1-Nap1-PIR121 complex is as active as the WAVE2-Abi1 sub-complex in stimulating Arp2/3, and after Rac activation it is re-localized to the leading edge of ruffles in vivo. Consistently, inhibition of Abi1 by RNA interference (RNAi) abrogates Rac-dependent lamellipodia protrusion. Thus, Abi1 orchestrates the proper assembly of the WAVE2 complex and mediates its activation at the leading edge in vivo.

Acetylation of Histone Deacetylase 1 Regulates NuRD Corepressor Complex Activity*

PubMed Central

Yang, Tao; Jian, Wei; Luo, Yi; Fu, Xueqi; Noguchi, Constance; Bungert, Jörg; Huang, Suming; Qiu, Yi

2012-01-01

Histone deacetylases (HDACs) play important roles in regulating cell proliferation and differentiation. The HDAC1-containing NuRD complex is generally considered as a corepressor complex and is required for GATA-1-mediated repression. However, recent studies also show that the NuRD complex is involved in GATA-1-mediated gene activation. We tested whether the GATA-1-associated NuRD complex loses its deacetylase activity and commits the GATA-1 complex to become an activator during erythropoiesis. We found that GATA-1-associated deacetylase activity gradually decreased upon induction of erythroid differentiation. GATA-1-associated HDAC1 is increasingly acetylated after differentiation. It has been demonstrated earlier that acetylated HDAC1 has no deacetylase activity. Indeed, overexpression of an HDAC1 mutant, which mimics acetylated HDAC1, promotes GATA-1-mediated transcription and erythroid differentiation. Furthermore, during erythroid differentiation, acetylated HDAC1 recruitment is increased at GATA-1-activated genes, whereas it is significantly decreased at GATA-1-repressed genes. Interestingly, deacetylase activity is not required for Mi2 remodeling activity, suggesting that remodeling activity may be required for both activation and repression. Thus, our data suggest that NuRD can function as a coactivator or repressor and that acetylated HDAC1 converts the NuRD complex from a repressor to an activator during GATA-1-directed erythroid differentiation. PMID:23014989

Role and structural mechanism of WASP-triggered conformational changes in branched actin filament nucleation by Arp2/3 complex.

PubMed

Rodnick-Smith, Max; Luan, Qing; Liu, Su-Ling; Nolen, Brad J

2016-07-05

The Arp2/3 (Actin-related proteins 2/3) complex is activated by WASP (Wiskott-Aldrich syndrome protein) family proteins to nucleate branched actin filaments that are important for cellular motility. WASP recruits actin monomers to the complex and stimulates movement of Arp2 and Arp3 into a "short-pitch" conformation that mimics the arrangement of actin subunits within filaments. The relative contribution of these functions in Arp2/3 complex activation and the mechanism by which WASP stimulates the conformational change have been unknown. We purified budding yeast Arp2/3 complex held in or near the short-pitch conformation by an engineered covalent cross-link to determine if the WASP-induced conformational change is sufficient for activity. Remarkably, cross-linked Arp2/3 complex bypasses the need for WASP in activation and is more active than WASP-activated Arp2/3 complex. These data indicate that stimulation of the short-pitch conformation is the critical activating function of WASP and that monomer delivery is not a fundamental requirement for nucleation but is a specific requirement for WASP-mediated activation. During activation, WASP limits nucleation rates by releasing slowly from nascent branches. The cross-linked complex is inhibited by WASP's CA region, even though CA potently stimulates cross-linking, suggesting that slow WASP detachment masks the activating potential of the short-pitch conformational switch. We use structure-based mutations and WASP-Arp fusion chimeras to determine how WASP stimulates movement toward the short-pitch conformation. Our data indicate that WASP displaces the autoinhibitory Arp3 C-terminal tail from a hydrophobic groove at Arp3's barbed end to destabilize the inactive state, providing a mechanism by which WASP stimulates the short-pitch conformation and activates Arp2/3 complex.

Synthesis, structural and biochemical activity studies of a new hexadentate Schiff base ligand and its Cu(II), Ni(II), and Co(II) complexes

NASA Astrophysics Data System (ADS)

Ekmekcioglu, Pinar; Karabocek, Nevin; Karabocek, Serdar; Emirik, Mustafa

2015-11-01

A new Schiff base ligand (H2L) and its metal complexes have been prepared and characterized by elemental analysis, magnetic moment and spectral studies. The comparative in-vitro antimicrobial activities against various pathogens with reference to known antibiotics activity under the standard control of different concentrations revealed that the metal complexes (6-8) showed enhanced antimicrobial activities in general as compared to free ligand. As an exception, the free ligand showed better activity against Trichoderma. The antifungal activity experiments were performed in triplicate. The order of biochemical activity for metal complexes were observed as in the following. CuL > CoL > NiL, which is exactly same as the order of stability constants of these complexes. Additionally, we performed DFT and TD-DFT calculation for free ligand and Cu(II) complex to support the experimental data. The geometries of the Cu(II) complex have been optimized using the B3LYP level of theory. The theoretical calculations confirm that the copper (II) center exhibits a distorted square pyramidal geometry which is favored by experimental results.

The Differential Gibbs Free Energy of Activation and its Implications in the Transition-State of Enzymatic Reactions

NASA Astrophysics Data System (ADS)

Maggi, F.; Riley, W. J.

2016-12-01

We propose a mathematical framework to introduce the concept of differential free energy of activation in enzymatically catalyzed reactions, and apply it to N uptake by microalgae and bacteria. This framework extends the thermodynamic capabilities of the classical transition-state theory in and harmonizes the consolidated definitions of kinetic parameters with their thermodynamic and physical meaning. Here, the activation energy is assumed to be a necessary energetic level for equilibrium complexation between reactants and activated complex; however, an additional energy contribution is required for the equilibrium activated complex to release reaction products. We call this "differential free energy of activation"; it can be described by a Boltzmann distribution, and corresponds to a free energy level different from that of complexation. Whether this level is above or below the free energy of activation depends on the reaction, and defines energy domains that correspond to "superactivated", "activated", and "subactivated" complexes. The activated complex reaching one of those states will eventually release the products from an energy level different than that of activation. The concept of differential free energy of activation was tested on 57 independent experiments of NH­4+ and NO3- uptake by various microalgae and bacteria at temperatures ranging between 1 and 45oC. Results showed that the complexation equilibrium always favored the activated complex, but the differential energy of activation led to an apparent energy barrier consistent with observations. Temperature affected all energy levels within this framework but did not alter substantially these thermodynamic features. Overall the approach: (1) provides a thermodynamic and mathematical link between Michaelis-Menten and rate constants; (2) shows that both kinetic parameters can be described or approximated by Arrhenius' like equations; (3) describes the likelihood of formation of sub-, super-, and activated complexes; and (4) shows direction and thermodynamic likelihood of each reaction branch within the transition state. The approach suites particularly well for calibration of kinetic parameters against experimentally acquired reaction dynamics measurements of nutrient biogeochemical cycles.

Clofibric acid stimulates branched-chain amino acid catabolism by three mechanisms.

PubMed

Kobayashi, Rumi; Murakami, Taro; Obayashi, Mariko; Nakai, Naoya; Jaskiewicz, Jerzy; Fujiwara, Yoko; Shimomura, Yoshiharu; Harris, Robert A

2002-11-15

Clofibrate promotes catabolism of branched-chain amino acids by increasing the activity of the branched-chain alpha-keto acid dehydrogenase [BCKDH] complex. Depending upon the sex of the rats, nutritional state, and tissue being studied, clofibrate can affect BCKDH complex activity by three different mechanisms. First, by directly inhibiting BCKDH kinase activity, clofibrate can increase the proportion of the BCKDH complex in the active, dephosphorylated state. This occurs in situations in which the BCKDH complex is largely inactive due to phosphorylation, e.g., in the skeletal muscle of chow-fed rats or in the liver of female rats late in the light cycle. Second, by increasing the levels at which the enzyme components of the BCKDH complex are expressed, clofibrate can increase the total enzymatic activity of the BCKDH complex. This is readily demonstrated in livers of rats fed a low-protein diet, a nutritional condition that induces a decrease in the level of expression of the BCKDH complex. Third, by decreasing the amount of BCKDH kinase expressed and therefore its activity, clofibrate induces an increase in the percentage of the BCKDH complex in the active, dephosphorylated state. This occurs in the livers of rats fed a low-protein diet, a nutritional condition that causes inactivation of the BCKDH complex due to upregulation of the amount of BCKDH kinase. WY-14,643, which, like clofibric acid, is a ligand for the peroxisome-proliferator-activated receptor alpha [PPARalpha], does not directly inhibit BCKDH kinase but produces the same long-term effects as clofibrate on expression of the BCKDH complex and its kinase. Thus, clofibrate is unique in its capacity to stimulate BCAA oxidation through inhibition of BCKDH kinase activity, whereas PPARalpha activators in general promote BCAA oxidation by increasing expression of components of the BCKDH complex and decreasing expression of the BCKDH kinase.

[Study on molecular recognition technology in active constituents extracted and isolated from Aconitum pendulum].

PubMed

Ma, Xue-Qin; Li, Guo-Shan; Fu, Xue-Yan; Ma, Jing-Zu

2011-03-01

To investigate CD molecular recognition technology applied in active constituents extracted and isolated from traditional Chinese medicine--Aconitum pendulum. The inclusion constant and form probability of the inclusion complex of Aconitum pendulum with p-CD was calculated by UV spectra method. The active constituents of Aconitum pendulum were extracted and isolated by molecular recognition technology. The inclusion complex was identified by UV. The chemical constituents of Aconitum pendulum and inclusion complex was determined by HPLC. The analgesic effects of inclusion complex was investigated by experiment of intraperitoneal injection of acetic acid in rats. The inclusion complex was identified and confirmed by UV spectra method, the chemical components of inclusion complex were simple, and the content of active constituents increased significantly, the analgesic effects of inclusion complex was well. The molecular recognition technology can be used for extracting and isolating active constituents of Aconitum pendulum, and the effects are obvious.

Antimalarial, antimicrobial, cytotoxic, DNA interaction and SOD like activities of tetrahedral copper(II) complexes

NASA Astrophysics Data System (ADS)

Mehta, Jugal V.; Gajera, Sanjay B.; Patel, Mohan N.

2015-02-01

The mononuclear copper(II) complexes with P, O-donor ligand and different fluoroquinolones have been synthesized and characterized by elemental analysis, electronic spectra, TGA, EPR, FT-IR and LC-MS spectroscopy. An antimicrobial efficiency of the complexes has been tested against five different microorganisms in terms of minimum inhibitory concentration (MIC) and displays very good antimicrobial activity. The binding strength and binding mode of the complexes with Herring Sperm DNA (HS DNA) have been investigated by absorption titration and viscosity measurement studies. The studies suggest the classical intercalative mode of DNA binding. Gel electrophoresis assay determines the ability of the complexes to cleave the supercoiled form of pUC19 DNA. Synthesized complexes have been tested for their SOD mimic activity using nonenzymatic NBT/NADH/PMS system and found to have good antioxidant activity. All the complexes show good cytotoxic and in vitro antimalarial activities.

Complement activating properties of complexes containing rheumatoid factor in synovial fluids and sera from patients with rheumatoid arthritis.

PubMed Central

Elson, C J; Carter, S D; Cottrell, B J; Scott, D G; Bacon, P A; Wallington, T B

1985-01-01

The relationship between complexes containing rheumatoid factor and complexes activating complement was examined in synovial fluids and sera from patients with rheumatoid arthritis (RA). In each case this was performed by quantifying the amount of rheumatoid factor bound by solid phase Fab'2 anti-C3 and/or solid phase conglutinin. Both anti-C3 coated and conglutinin coated microtitre plates bound high levels of complexes containing rheumatoid factor from sera of RA patients with vasculitis. Unexpectedly, these complexes were detected in synovial fluids from only a minority of RA patients with synovitis. However, RA synovial fluids did contain other complexes as shown by the presence of complement consuming activity, C1q binding material and immunoglobulin attaching to conglutinin. It is considered that in RA synovial fluids the complexes containing RF and those activating complement are not necessarily the same whilst in vasculitic sera the complexes containing rheumatoid factor also activate complement. PMID:3978872

Synthesis, crystal structures, molecular docking, and in vitro biological activities evaluation of transition metal complexes with 4-(3,4-dichlorophenyl) piperazine-1-carboxylic acid

NASA Astrophysics Data System (ADS)

Chen, Zhi-Jian; Chen, Ya-Na; Xu, Chun-Na; Zhao, Shan-Shan; Cao, Qi-Yue; Qian, Shao-Song; Qin, Jie; Zhu, Hai-Liang

2016-08-01

Three novel mononuclear complexes, [MⅡ(L)2·2H2O], (M = Cu, Ni or Cd; HL = 4-(3,4-dichlorophenyl)piperazine-1-carboxylic acid)were synthesized and structurally determined by single-crystal X-ray diffraction. Molecular docking study preliminarily revealed that complex 1 had potential urease inhibitory activity. In accordance with the result of calculation, in vitro tests of the inhibitory activities of complexes 1-3 against jack bean urease showed complex 1 (IC50 = 8.17 ± 0.91 μM) had better inhibitory activities than the positive reference acetohydroxamic acid (AHA) (IC50 = 26.99 ± 1.43 μM), while complexes 2 and 3 showed no inhibitory activities., kinetics study was carried out to explore the mechanism of the inhibiting of the enzyme, and the result indicated that complex 1 was a competitive inhibitor of urease. Albumin binding experiment and in vitro toxicity evaluation of complex 1 were implemented to explore its Pharmacological properties.

Cobalt Complexes as Antiviral and Antibacterial Agents

DTIC Science & Technology

2010-01-01

observed. Complex 26 has antibacterial activity against E. coli, S. aureus and Micrococcus lysodeikiticus, showing better growth inhibitory activity in...complexes exhibited activity towards E. coli, B. subtilis, S. aureus and Micrococcus lysodeikiticus. Figure 15. Selenium containing and

The assembly, activation, and substrate specificity of Cyclin D1/Cdk2 complexes

PubMed Central

Jahn, Stephan C.; Law, Mary E.; Corsino, Patrick E.; Rowe, Thomas C.; Davis, Bradley J.; Law, Brian K.

2013-01-01

Previous studies have shown conflicting data regarding Cyclin D1/Cdk2 complexes and, considering the widespread overexpression of Cyclin D1 in cancer, it is important to fully understand their relevance. While many have shown Cyclin D1/Cdk2 complexes to form active complexes, others have failed to show activity or association. Here, using a novel p21-PCNA fusion protein as well as p21 mutant proteins, we show that p21 is a required scaffolding protein, with Cyclin D1 and Cdk2 failing to complex in its absence. These p21/Cyclin D1/Cdk2 complexes are active and also bind the trimeric PCNA complex, with each trimer capable of independently binding distinct Cyclin/Cdk complexes. We also show that increased p21 levels due to treatment with chemotherapeutic agents result in increased formation and kinase activity of Cyclin D1/Cdk2 complexes, and that Cyclin D1/Cdk2 complexes are able to phosphorylate a number of substrates in addition to Rb. Nucleophosmin and Cdh1, two proteins important for centrosome replication and implicated in the chromosomal instability of cancer are shown to be phosphorylated by Cyclin D1/Cdk2 complexes. Additionally, PSF is identified as a novel Cdk2 substrate, being phosphorylated by Cdk2 complexed with either Cyclin E or Cyclin D1, and given the many functions of PSF, it could have important implications on cellular activity. PMID:23627734

THE BIOLOGICAL ACTIVITY OF SOLUBLE ANTIGEN-ANTIBODY COMPLEXES

PubMed Central

Ishizaka, Kimishige; Ishizaka, Teruko; Campbell, Dan H.

1959-01-01

Soluble BSA-anti-BSA complexes, formed in antigen excess, give immediate skin reactions in normal guinea pigs. The mechanism of the reaction is not that of passive or reversed passive anaphylaxis. The complex itself is toxic. Skin activity of the complex depends on its composition. It has become obvious that the complex composed of two antigen molecules and one antibody molecule, (Ag2Ab), does not have the activity, whereas, Ag3Ab2 and more complicated complexes do. The role of complement as well as speculation on the structural changes of antibody-antigen complexes is presented. PMID:13620844

Polyelectrolyte complexes between (cross-linked) N-carboxyethylchitosan and (quaternized) poly[2-(dimethylamino)ethyl methacrylate]: preparation, characterization, and antibacterial properties.

PubMed

Yancheva, Elena; Paneva, Dilyana; Maximova, Vera; Mespouille, Laetitia; Dubois, Philippe; Manolova, Nevena; Rashkov, Iliya

2007-03-01

Novel polyelectrolyte complexes (PECs) between N-carboxyethylchitosan (CECh) and well-defined (quaternized) poly[2-(dimethylamino)ethyl methacrylate] (PDMAEMA) have been obtained. The modification of chitosan into CECh allows the preparation of PECs in a pH range in which chitosan cannot form complexes. The CECh/PDMAEMA complex is formed in a narrow pH range around 7. The quaternization of the tertiary amino groups of PDMAEMA enables complex formation with CECh both in neutral and in alkaline medium. Cross-linked CECh is also capable of forming complexes with (quaternized) PDMAEMA. The antibacterial activity of (cross-linked) CECh, (quaternized) PDMAEMA, and their complexes against Escherichia coli has been evaluated. In contrast to (quaternized) PDMAEMA, (cross-linked) CECh exhibits no antibacterial activity. The complex formation between cross-linked CECh and (quaternized) PDMAEMA results in a loss of the inherent antibacterial activity of the latter in neutral medium. In acidic medium, the complexes exhibit strong antibacterial activity due to complex disintegration and release of (quaternized) PDMAEMA.

A Damaged Oxidative Phosphorylation Mechanism Is Involved in the Antifungal Activity of Citral against Penicillium digitatum

PubMed Central

OuYang, Qiuli; Tao, Nengguo; Zhang, Miaoling

2018-01-01

Citral exhibits strong antifungal activity against Penicillium digitatum. In this study, 41 over-expressed and 84 repressed proteins in P. digitatum after 1.0 μL/mL of citral exposure for 30 min were identified by the iTRAQ technique. The proteins were closely related with oxidative phosphorylation, the TCA cycle and RNA transport. The mitochondrial complex I, complex II, complex III, complex IV and complex V, which are involved in oxidative phosphorylation were drastically affected. Among of them, the activities of mitochondrial complex I and complex IV were apparently suppressed, whereas those of mitochondrial complex II, complex III and complex V were significantly induced. Meanwhile, citral apparently triggered a reduction in the intracellular ATP, the mitochondrial membrane potential (MMP) and glutathione content, in contrast to an increase in the glutathione S-transferase activity and the accumulation of reactive oxygen species (ROS). Addition of exogenous cysteine decreased the antifungal activity. In addition, cysteine maintained the basal ROS level, deferred the decrease of MMP and the membrane damage. These results indicate that citral inhibited the growth of P. digitatum by damaging oxidative phosphorylation and cell membranes through the massive accumulation of ROS. PMID:29503638

A Damaged Oxidative Phosphorylation Mechanism Is Involved in the Antifungal Activity of Citral against Penicillium digitatum.

PubMed

OuYang, Qiuli; Tao, Nengguo; Zhang, Miaoling

2018-01-01

Citral exhibits strong antifungal activity against Penicillium digitatum . In this study, 41 over-expressed and 84 repressed proteins in P. digitatum after 1.0 μL/mL of citral exposure for 30 min were identified by the iTRAQ technique. The proteins were closely related with oxidative phosphorylation, the TCA cycle and RNA transport. The mitochondrial complex I, complex II, complex III, complex IV and complex V, which are involved in oxidative phosphorylation were drastically affected. Among of them, the activities of mitochondrial complex I and complex IV were apparently suppressed, whereas those of mitochondrial complex II, complex III and complex V were significantly induced. Meanwhile, citral apparently triggered a reduction in the intracellular ATP, the mitochondrial membrane potential (MMP) and glutathione content, in contrast to an increase in the glutathione S-transferase activity and the accumulation of reactive oxygen species (ROS). Addition of exogenous cysteine decreased the antifungal activity. In addition, cysteine maintained the basal ROS level, deferred the decrease of MMP and the membrane damage. These results indicate that citral inhibited the growth of P. digitatum by damaging oxidative phosphorylation and cell membranes through the massive accumulation of ROS.

In vitro reconstitution and characterization of the yeast mitochondrial degradosome complex unravels tight functional interdependence.

PubMed

Malecki, Michal; Jedrzejczak, Robert; Stepien, Piotr P; Golik, Pawel

2007-09-07

The mitochondrial degradosome (mtEXO), the main RNA-degrading complex of yeast mitochondria, is composed of two subunits: an exoribonuclease encoded by the DSS1 gene and an RNA helicase encoded by the SUV3 gene. We expressed both subunits of the yeast mitochondrial degradosome in Escherichia coli, reconstituted the complex in vitro and analyzed the RNase, ATPase and helicase activities of the two subunits separately and in complex. The results reveal a very strong functional interdependence. For every enzymatic activity, we observed significant changes when the relevant protein was present in the complex, compared to the activity measured for the protein alone. The ATPase activity of Suv3p is stimulated by RNA and its background activity in the absence of RNA is reduced greatly when the protein is in the complex with Dss1p. The Suv3 protein alone does not display RNA-unwinding activity and the 3' to 5' directional helicase activity requiring a free 3' single-stranded substrate becomes apparent only when Suv3p is in complex with Dss1p. The Dss1 protein alone does have some basal exoribonuclease activity, which is not ATP-dependent, but in the presence of Suv3p the activity of the entire complex is enhanced greatly and is entirely ATP-dependent, with no residual activity observed in the absence of ATP. Such absolute ATP-dependence is unique among known exoribonuclease complexes. On the basis of these results, we propose a model in which the Suv3p RNA helicase acts as a molecular motor feeding the substrate to the catalytic centre of the RNase subunit.

Proteolytic turnover of the Gal4 transcription factor is not required for function in vivo.

PubMed

Nalley, Kip; Johnston, Stephen Albert; Kodadek, Thomas

2006-08-31

Transactivator-promoter complexes are essential intermediates in the activation of eukaryotic gene expression. Recent studies of these complexes have shown that some are quite dynamic in living cells owing to rapid and reversible disruption of activator-promoter complexes by molecular chaperones, or a slower, ubiquitin-proteasome-pathway-mediated turnover of DNA-bound activator. These mechanisms may act to ensure continued responsiveness of activators to signalling cascades by limiting the lifetime of the active protein-DNA complex. Furthermore, the potency of some activators is compromised by proteasome inhibition, leading to the suggestion that periodic clearance of activators from a promoter is essential for high-level expression. Here we describe a variant of the chromatin immunoprecipitation assay that has allowed direct observation of the kinetic stability of native Gal4-promoter complexes in yeast. Under non-inducing conditions, the complex is dynamic, but on induction the Gal4-promoter complexes 'lock in' and exhibit long half-lives. Inhibition of proteasome-mediated proteolysis had little or no effect on Gal4-mediated gene expression. These studies, combined with earlier data, show that the lifetimes of different transactivator-promoter complexes in vivo can vary widely and that proteasome-mediated turnover is not a general requirement for transactivator function.

Synthesis and antimalarial activity of metal complexes of cross-bridged tetraazamacrocyclic ligands.

PubMed

Hubin, Timothy J; Amoyaw, Prince N-A; Roewe, Kimberly D; Simpson, Natalie C; Maples, Randall D; Carder Freeman, TaRynn N; Cain, Amy N; Le, Justin G; Archibald, Stephen J; Khan, Shabana I; Tekwani, Babu L; Khan, M O Faruk

2014-07-01

Using transition metals such as manganese(II), iron(II), cobalt(II), nickel(II), copper(II), and zinc(II), several new metal complexes of cross-bridged tetraazamacrocyclic chelators namely, cyclen- and cyclam-analogs with benzyl groups, were synthesized and screened for in vitro antimalarial activity against chloroquine-resistant (W2) and chloroquine-sensitive (D6) strains of Plasmodium falciparum. The metal-free chelators tested showed little or no antimalarial activity. All the metal complexes of the dibenzyl cross-bridged cyclam ligand exhibited potent antimalarial activity. The Mn(2+) complex of this ligand was the most potent with IC50s of 0.127 and 0.157μM against the chloroquine-sensitive (D6) and chloroquine-resistant (W2) P. falciparum strains, respectively. In general, the dibenzyl hydrophobic ligands showed better anti-malarial activity compared to the activity of monobenzyl ligands, potentially because of their higher lipophilicity and thus better cell penetration ability. The higher antimalarial activity displayed by the manganese complex for the cyclam ligand in comparison to that of the cyclen, correlates with the larger pocket of cyclam compared to that of cyclen which produces a more stable complex with the Mn(2+). Few of the Cu(2+) and Fe(2+) complexes also showed improvement in activity but Ni(2+), Co(2+) and Zn(2+) complexes did not show any improvement in activity upon the metal-free ligands for anti-malarial development. Published by Elsevier Ltd.

The dermatomyositis-specific autoantigen Mi2 is a component of a complex containing histone deacetylase and nucleosome remodeling activities.

PubMed

Zhang, Y; LeRoy, G; Seelig, H P; Lane, W S; Reinberg, D

1998-10-16

Histone acetylation and deacetylation were found to be catalyzed by structurally distinct, multisubunit complexes that mediate, respectively, activation and repression of transcription. ATP-dependent nucleosome remodeling, mediated by different multisubunit complexes, was thought to be involved only in transcription activation. Here we report the isolation of a protein complex that contains both histone deacetylation and ATP-dependent nucleosome remodeling activities. The complex contains the histone deacetylases HDAC1/2, histone-binding proteins, the dermatomyositis-specific autoantigen Mi2beta, a polypeptide related to the metastasis-associated protein 1, and a novel polypeptide of 32 kDa. Patients with dermatomyositis have a high rate of malignancy. The finding that Mi2beta exists in a complex containing histone deacetylase and nucleosome remodeling activities suggests a role for chromatin reorganization in cancer metastasis.

Developmental and hormone-induced changes of mitochondrial electron transport chain enzyme activities during the last instar larval development of maize stem borer, Chilo partellus (Lepidoptera: Crambidae).

PubMed

VenkatRao, V; Chaitanya, R K; Naresh Kumar, D; Bramhaiah, M; Dutta-Gupta, A

2016-12-01

The energy demand for structural remodelling in holometabolous insects is met by cellular mitochondria. Developmental and hormone-induced changes in the mitochondrial respiratory activity during insect metamorphosis are not well documented. The present study investigates activities of enzymes of mitochondrial electron transport chain (ETC) namely, NADH:ubiquinone oxidoreductase or complex I, Succinate: ubiquinone oxidoreductase or complex II, Ubiquinol:ferricytochrome c oxidoreductase or complex III, cytochrome c oxidase or complex IV and F 1 F 0 ATPase (ATPase), during Chilo partellus development. Further, the effect of juvenile hormone (JH) analog, methoprene, and brain and corpora-allata-corpora-cardiaca (CC-CA) homogenates that represent neurohormones, on the ETC enzyme activities was monitored. The enzymatic activities increased from penultimate to last larval stage and thereafter declined during pupal development with an exception of ATPase which showed high enzyme activity during last larval and pupal stages compared to the penultimate stage. JH analog, methoprene differentially modulated ETC enzyme activities. It stimulated complex I and IV enzyme activities, but did not alter the activities of complex II, III and ATPase. On the other hand, brain homogenate declined the ATPase activity while the injected CC-CA homogenate stimulated complex I and IV enzyme activities. Cumulatively, the present study is the first to show that mitochondrial ETC enzyme system is under hormone control, particularly of JH and neurohormones during insect development. Copyright © 2015 Elsevier Inc. All rights reserved.

Quercetin-Iron Complex: Synthesis, Characterization, Antioxidant, DNA Binding, DNA Cleavage, and Antibacterial Activity Studies.

PubMed

Raza, Aun; Xu, Xiuquan; Xia, Li; Xia, Changkun; Tang, Jian; Ouyang, Zhen

2016-11-01

Quercetin-iron (II) complex was synthesized and characterized by elemental analysis, ultraviolet-visible spectrophotometry, fourier transform infrared spectroscopy, mass spectrometry, proton nuclear magnetic resonance spectroscopy, thermogravimetry and differential scanning calorimetry, scanning electron micrography and molar conductivity. The low molar conductivity value investigates the non-electrolyte nature of the complex. The elemental analysis and other physical and spectroscopic methods reveal the 1:2 stoichiometric ratio (metal:ligand) of the complex. Antioxidant study of the quercetin and its metal complex against 2, 2-di-phenyl-1-picryl hydrazyl radical showed that the complex has much more radical scavenging activity than free quercetin. The interaction of quercetin-iron (II) complex with DNA was determined using ultraviolet visible spectra, fluorescence spectra and agarose gel electrophoresis. The results showed that quercetin-iron (II) complex can intercalate moderately with DNA, quench a strong intercalator ethidium bromide and compete for the intercalative binding sites. The complex showed significant cleavage of pBR 322 DNA from supercoiled form to nicked circular form and these cleavage effects were dose-dependent. Moreover, the mechanism of DNA cleavage indicated that it was an oxidative cleavage pathway. These results revealed the potential nuclease activity of complex to cleave DNA. In addition, antibacterial activity of complex on E.coli and S. aureus was also investigated. The results showed that complex has higher antibacterial activity than ligand.

Cannabinoid-Induced Changes in the Activity of Electron Transport Chain Complexes of Brain Mitochondria.

PubMed

Singh, Namrata; Hroudová, Jana; Fišar, Zdeněk

2015-08-01

The aim of this study was to investigate changes in the activity of individual mitochondrial respiratory chain complexes (I, II/III, IV) and citrate synthase induced by pharmacologically different cannabinoids. In vitro effects of selected cannabinoids on mitochondrial enzymes were measured in crude mitochondrial fraction isolated from pig brain. Both cannabinoid receptor agonists, Δ(9)-tetrahydrocannabinol, anandamide, and R-(+)-WIN55,212-2, and antagonist/inverse agonists of cannabinoid receptors, AM251, and cannabidiol were examined in pig brain mitochondria. Different effects of these cannabinoids on mitochondrial respiratory chain complexes and citrate synthase were found. Citrate synthase activity was decreased only by Δ(9)-tetrahydrocannabinol and AM251. Significant increase in the complex I activity was induced by anandamide. At micromolar concentration, all the tested cannabinoids inhibited the activity of electron transport chain complexes II/III and IV. Stimulatory effect of anandamide on activity of complex I may participate on distinct physiological effects of endocannabinoids compared to phytocannabinoids or synthetic cannabinoids. Common inhibitory effect of cannabinoids on activity of complex II/III and IV confirmed a non-receptor-mediated mechanism of cannabinoid action on individual components of system of oxidative phosphorylation.

Optimization of WAVE2 complex-induced actin polymerization by membrane-bound IRSp53, PIP(3), and Rac.

PubMed

Suetsugu, Shiro; Kurisu, Shusaku; Oikawa, Tsukasa; Yamazaki, Daisuke; Oda, Atsushi; Takenawa, Tadaomi

2006-05-22

WAVE2 activates the actin-related protein (Arp) 2/3 complex for Rac-induced actin polymerization during lamellipodium formation and exists as a large WAVE2 protein complex with Sra1/PIR121, Nap1, Abi1, and HSPC300. IRSp53 binds to both Rac and Cdc42 and is proposed to link Rac to WAVE2. We found that the knockdown of IRSp53 by RNA interference decreased lamellipodium formation without a decrease in the amount of WAVE2 complex. Localization of WAVE2 at the cell periphery was retained in IRSp53 knockdown cells. Moreover, activated Cdc42 but not Rac weakened the association between WAVE2 and IRSp53. When we measured Arp2/3 activation in vitro, the WAVE2 complex isolated from the membrane fraction of cells was fully active in an IRSp53-dependent manner but WAVE2 isolated from the cytosol was not. Purified WAVE2 and purified WAVE2 complex were activated by IRSp53 in a Rac-dependent manner with PIP(3)-containing liposomes. Therefore, IRSp53 optimizes the activity of the WAVE2 complex in the presence of activated Rac and PIP(3).

Pyrrolidine dithiocarbamate-zinc(II) and -copper(II) complexes induce apoptosis in tumor cells by inhibiting the proteasomal activity☆

PubMed Central

Milacic, Vesna; Chen, Di; Giovagnini, Lorena; Diez, Alejandro; Fregona, Dolores; Dou, Q. Ping

2013-01-01

Zinc and copper are trace elements essential for proper folding, stabilization and catalytic activity of many metalloenzymes in living organisms. However, disturbed zinc and copper homeostasis is reported in many types of cancer. We have previously demonstrated that copper complexes induced proteasome inhibition and apoptosis in cultured human cancer cells. In the current study we hypothesized that zinc complexes could also inhibit the proteasomal chymotrypsin-like activity responsible for subsequent apoptosis induction. We first showed that zinc(II) chloride was able to inhibit the chymotrypsin-like activity of a purified 20S proteasome with an IC50 value of 13.8 μM, which was less potent than copper(II) chloride (IC50 5.3 μM). We then compared the potencies of a pyrrolidine dithiocarbamate (PyDT)-zinc(II) complex and a PyDT-copper(II) complex to inhibit cellular proteasomal activity, suppress proliferation and induce apoptosis in various human breast and prostate cancer cell lines. Consistently, zinc complex was less potent than copper complex in inhibiting the proteasome and inducing apoptosis. Additionally, zinc and copper complexes appear to use somewhat different mechanisms to kill tumor cells. Zinc complexes were able to activate calpain-, but not caspase-3-dependent pathway, while copper complexes were able to induce activation of both proteases. Furthermore, the potencies of these PyDT-metal complexes depend on the nature of metals and also on the ratio of PyDT to the metal ion within the complex, which probably affects their stability and availability for interacting with and inhibiting the proteasome in tumor cells. PMID:18501397

Heterotropic Effect of β-lactam Antibiotics on Antioxidant Property of Haptoglobin (2-2)-Hemoglobin Complex.

PubMed

Tayari, Masoumeh; Moosavi-Nejad, Zahra; Moosavi Nejad, Fatemeh; Rezaei-Tavirani, Mostafa; Dehghan Shasaltaneh, Marzieh

2011-01-01

Haptoglobin (Hp) is a mammalian serum glycoprotein showing a genetic polymorphism with three types, 1-1, 2-2 and 1-2. Hp appears to conserve the recycling of heme-iron by forming an essentially irreversible but non-covalent complex with hemoglobin which is released into the plasma by erythrocyte lysis. As an important consequence, Haptoglobin-Hemoglobin complex (Hp-Hb) shows considerable antioxidant property. In this study, antioxidant activity of Hp (2-2)-Hb complex on hydrogen peroxide has been studied and analyzed in the absence and presence of two beta-lactam antibiotics in-vitro. For this purpose, non-Michaelis behavior of peroxidase activity of Hp (2-2)-Hb complex was analyzed using Eadie-Hofstee, Clearance and Hill plots, in the absence and presence of pharmaceutical dose of ampicillin and coamoxiclav. The results have shown that peroxidase activity of Hp (2-2)-Hb complex is modulated via homotropic effect of hydrogen peroxide as an allostric substrate. On the other hand antioxidant property of Hp (2-2)-Hb complex increased via heterotropic effect of both antibiotics on the peroxidase activity of the complex. Both drugs also have mild effect on quality of homotropic property of the peroxidase activity of Hp (2-2)-Hb complex. Therefore, it can be concluded from our study that both beta-lactam antibiotics can increase peroxidase activity of Hp (2-2)-Hb complex via heterotropic effect. Thus, the two antibiotics (especially ampicillin) may help those individuals with Hp (2-2) phenotype to improve the Hp-Hb complex efficiency of removing hydrogen peroxide from serum under oxidative stress. This can be important in the individuals with phenotype Hp 2-2 who have less antioxidant activity relative to other phenotypes and are susceptible to cardiovascular disorders, as has been reported by other researchers.

Investigating the effect of gallium curcumin and gallium diacetylcurcumin complexes on the structure, function and oxidative stability of the peroxidase enzyme and their anticancer and antibacterial activities.

PubMed

Jahangoshaei, Parisa; Hassani, Leila; Mohammadi, Fakhrossadat; Hamidi, Akram; Mohammadi, Khosro

2015-10-01

Curcumin has a wide spectrum of biological and pharmacological activities including anti-inflammatory, antioxidant, antiproliferative, antimicrobial and anticancer activities. Complexation of curcumin with metals has gained attention in recent years for improvement of its stability. In this study, the effect of gallium curcumin and gallium diacetylcurcumin on the structure, function and oxidative stability of horseradish peroxidase (HRP) enzyme were evaluated by spectroscopic techniques. In addition to the enzymatic investigation, the cytotoxic effect of the complexes was assessed on bladder, MCF-7 breast cancer and LNCaP prostate carcinoma cell lines by MTT assay. Furthermore, antibacterial activity of the complexes against S. aureus and E. coli was explored by dilution test method. The results showed that the complexes improve activity of HRP and also increase its tolerance against the oxidative condition. After addition of the complexes, affinity of HRP for hydrogen peroxide substrate decreases, while the affinity increases for phenol substrate. Circular dichroism, intrinsic and synchronous fluorescence spectra showed that the enzyme structure around the catalytic heme group becomes less compact and also the distance between the heme group and tryptophan residues increases due to binding of the complexes to HRP. On the whole, it can be concluded that the change in the enzyme structure upon binding to the gallium curcumin and gallium diacetylcurcumin complexes results in an increase in the antioxidant efficiency and activity of the peroxidise enzyme. The result of anticancer and antibacterial activities suggested that the complexes exhibit the potential for cancer treatment, but they have no significant antibacterial activity.

Highly Reactive Scandium Phosphinoalkylidene Complex: C-H and H-H Bonds Activation.

PubMed

Mao, Weiqing; Xiang, Li; Alvarez Lamsfus, Carlos; Maron, Laurent; Leng, Xuebing; Chen, Yaofeng

2017-01-25

The first scandium phosphinoalkylidene complex was synthesized and structurally characterized. The complex has the shortest Sc-C bond lengths reported to date (2.089(3) Å). DFT calculations reveal the presence of a three center π interaction in the complex. This scandium phosphinoalkylidene complex undergoes intermolecular C-H bond activation of pyridine, 4-dimethylamino pyridine and 1,3-dimethylpyrazole at room temperature. Furthermore, the complex rapidly activates H 2 under mild conditions. DFT calculations also demonstrate that the C-H activation of 1,3-dimethylpyrazole is selective for thermodynamic reasons and the relatively slow reaction is due to the need of fully breaking the chelating effect of the phosphino group to undergo the reaction whereas this is not the case for H 2 .

Design of copper DNA intercalators with leishmanicidal activity.

PubMed

Navarro, Maribel; Cisneros-Fajardo, Efrén José; Sierralta, Aníbal; Fernández-Mestre, Mercedes; Silva, Pedro; Arrieche, Dwight; Marchán, Edgar

2003-04-01

The complexes [Cu(dppz)(NO(3))]NO(3) (1), [Cu(dppz)(2)(NO(3))]NO(3) (2), [Cu(dpq)(NO(3))]NO(3) (3), and [Cu(dpq)(2)(NO(3))]NO(3) (4) were synthesized and characterized by elemental analysis, FAB-mass spectrometry, EPR, UV, and IR spectroscopies, and molar conductivity. DNA interaction studies showed that intercalation is an important way of interacting with DNA for these complexes. The biological activity of these copper complexes was evaluated on Leishmania braziliensis promastigotes, and the results showed leishmanicidal activity. Preliminary ultrastructural studies with the most active complex (2) at 1 h revealed parasite swelling and binucleated cells. This finding suggests that the leishmanicidal activity of the copper complexes could be associated with their interaction with the parasitic DNA.

Antivirion Effects of Streptovaricin Complex Against Friend Virus

PubMed Central

Horoszewicz, Julius S.; Leong, Susan S.; Byrd, Daniel M.; Carter, William A.

1974-01-01

The in vitro antivirion activities of five different streptovaricin complex lots against the polycythemic strain of the Friend virus were evaluated. The assay system was based on the inhibition of the Friend virus-induced spleen foci. The virus inactivation process was shown to be susceptible to variation in temperature, pH, and time. The antivirion activity and the acute toxicity for mice, as well as the optical properties of these streptovaricin complexes, do not co-vary; this suggests that their biological activities are not associated with a single molecular structure. In addition, the antivirion activity of the five preparations of streptovaricin complex differs about 30-fold, indicating that this activity does not reside in a major component of the complex. PMID:15825311

Role and structural mechanism of WASP-triggered conformational changes in branched actin filament nucleation by Arp2/3 complex

PubMed Central

Rodnick-Smith, Max; Luan, Qing; Liu, Su-Ling; Nolen, Brad J.

2016-01-01

The Arp2/3 (Actin-related proteins 2/3) complex is activated by WASP (Wiskott–Aldrich syndrome protein) family proteins to nucleate branched actin filaments that are important for cellular motility. WASP recruits actin monomers to the complex and stimulates movement of Arp2 and Arp3 into a “short-pitch” conformation that mimics the arrangement of actin subunits within filaments. The relative contribution of these functions in Arp2/3 complex activation and the mechanism by which WASP stimulates the conformational change have been unknown. We purified budding yeast Arp2/3 complex held in or near the short-pitch conformation by an engineered covalent cross-link to determine if the WASP-induced conformational change is sufficient for activity. Remarkably, cross-linked Arp2/3 complex bypasses the need for WASP in activation and is more active than WASP-activated Arp2/3 complex. These data indicate that stimulation of the short-pitch conformation is the critical activating function of WASP and that monomer delivery is not a fundamental requirement for nucleation but is a specific requirement for WASP-mediated activation. During activation, WASP limits nucleation rates by releasing slowly from nascent branches. The cross-linked complex is inhibited by WASP’s CA region, even though CA potently stimulates cross-linking, suggesting that slow WASP detachment masks the activating potential of the short-pitch conformational switch. We use structure-based mutations and WASP–Arp fusion chimeras to determine how WASP stimulates movement toward the short-pitch conformation. Our data indicate that WASP displaces the autoinhibitory Arp3 C-terminal tail from a hydrophobic groove at Arp3′s barbed end to destabilize the inactive state, providing a mechanism by which WASP stimulates the short-pitch conformation and activates Arp2/3 complex. PMID:27325766

Pib2 and the EGO complex are both required for activation of TORC1.

PubMed

Varlakhanova, Natalia V; Mihalevic, Michael J; Bernstein, Kara A; Ford, Marijn G J

2017-11-15

The TORC1 complex is a key regulator of cell growth and metabolism in Saccharomyces cerevisiae The vacuole-associated EGO complex couples activation of TORC1 to the availability of amino acids, specifically glutamine and leucine. The EGO complex is also essential for reactivation of TORC1 following rapamycin-induced growth arrest and for its distribution on the vacuolar membrane. Pib2, a FYVE-containing phosphatidylinositol 3-phosphate (PI3P)-binding protein, is a newly discovered and poorly characterized activator of TORC1. Here, we show that Pib2 is required for reactivation of TORC1 following rapamycin-induced growth arrest. Pib2 is required for EGO complex-mediated activation of TORC1 by glutamine and leucine as well as for redistribution of Tor1 on the vacuolar membrane. Therefore, Pib2 and the EGO complex cooperate to activate TORC1 and connect phosphoinositide 3-kinase (PI3K) signaling and TORC1 activity. © 2017. Published by The Company of Biologists Ltd.

Roles of mono-ubiquitinated Smad4 in the formation of Smad transcriptional complexes

DOE Office of Scientific and Technical Information (OSTI.GOV)

Wang Bei; Suzuki, Hiroyuki; Kato, Mitsuyasu

2008-11-14

TGF-{beta} activates receptor-regulated Smad (R-Smad) through phosphorylation by type I receptors. Activated R-Smad binds to Smad4 and the complex translocates into the nucleus and stimulates the transcription of target genes through association with co-activators including p300. It is not clear, however, how activated Smad complexes are removed from target genes. In this study, we show that TGF-{beta} enhances the mono-ubiquitination of Smad4. Smad4 mono-ubiquitination was promoted by p300 and suppressed by the c-Ski co-repressor. Smad4 mono-ubiquitination disrupted the interaction with Smad2 in the presence of constitutively active TGF-{beta} type I receptor. Furthermore, mono-ubiquitinated Smad4 was not found in DNA-binding Smadmore » complexes. A Smad4-Ubiquitin fusion protein, which mimics mono-ubiquitinated Smad4, enhanced localization to the cytoplasm. These results suggest that mono-ubiquitination of Smad4 occurs in the transcriptional activator complex and facilitates the turnover of Smad complexes at target genes.« less

Roles of mono-ubiquitinated Smad4 in the formation of Smad transcriptional complexes.

PubMed

Wang, Bei; Suzuki, Hiroyuki; Kato, Mitsuyasu

2008-11-14

TGF-beta activates receptor-regulated Smad (R-Smad) through phosphorylation by type I receptors. Activated R-Smad binds to Smad4 and the complex translocates into the nucleus and stimulates the transcription of target genes through association with co-activators including p300. It is not clear, however, how activated Smad complexes are removed from target genes. In this study, we show that TGF-beta enhances the mono-ubiquitination of Smad4. Smad4 mono-ubiquitination was promoted by p300 and suppressed by the c-Ski co-repressor. Smad4 mono-ubiquitination disrupted the interaction with Smad2 in the presence of constitutively active TGF-beta type I receptor. Furthermore, mono-ubiquitinated Smad4 was not found in DNA-binding Smad complexes. A Smad4-Ubiquitin fusion protein, which mimics mono-ubiquitinated Smad4, enhanced localization to the cytoplasm. These results suggest that mono-ubiquitination of Smad4 occurs in the transcriptional activator complex and facilitates the turnover of Smad complexes at target genes.

Antimalarial and antimicrobial activities of 8-Aminoquinoline-Uracils metal complexes

PubMed Central

Phopin, Kamonrat; Sinthupoom, Nujarin; Treeratanapiboon, Lertyot; Kunwittaya, Sarun; Prachayasittikul, Supaluk; Ruchirawat, Somsak; Prachayasittikul, Virapong

2016-01-01

8-Aminoquinoline (8AQ) derivatives have been reported to have antimalarial, anticancer, and antioxidant activities. This study investigated the potency of 8AQ-5-substituted (iodo and nitro) uracils metal (Mn, Cu, Ni) complexes (1-6) as antimalarial and antimicrobial agents. Interestingly, all of these metal complexes (1-6) showed fair antimalarial activities. Moreover, Cu complexes 2 (8AQ-Cu-5Iu) and 5 (8AQ-Cu-5Nu) exerted antimicrobial activities against Gram-negative bacteria including P. shigelloides and S. dysenteriae. The results reveal application of 8AQ and its metal complexes as potential compounds to be further developed as novel antimalarial and antibacterial agents. PMID:27103894

Antimalarial and antimicrobial activities of 8-Aminoquinoline-Uracils metal complexes.

PubMed

Phopin, Kamonrat; Sinthupoom, Nujarin; Treeratanapiboon, Lertyot; Kunwittaya, Sarun; Prachayasittikul, Supaluk; Ruchirawat, Somsak; Prachayasittikul, Virapong

2016-01-01

8-Aminoquinoline (8AQ) derivatives have been reported to have antimalarial, anticancer, and antioxidant activities. This study investigated the potency of 8AQ-5-substituted (iodo and nitro) uracils metal (Mn, Cu, Ni) complexes (1-6) as antimalarial and antimicrobial agents. Interestingly, all of these metal complexes (1-6) showed fair antimalarial activities. Moreover, Cu complexes 2 (8AQ-Cu-5Iu) and 5 (8AQ-Cu-5Nu) exerted antimicrobial activities against Gram-negative bacteria including P. shigelloides and S. dysenteriae. The results reveal application of 8AQ and its metal complexes as potential compounds to be further developed as novel antimalarial and antibacterial agents.

Studies on Some Biologically Cobalt(II), Copper(II) and Zinc(II) Complexes With ONO, NNO and SNO Donor Pyrazinoylhydrazine-Derived Ligands

PubMed Central

Praveen, Marapaka; Sherazi, Syed K. A.

1998-01-01

Biologically active complexes of Co(II), Ni(II), Cu(II) and Zn(II) with novel ONO, NNO and SNO donor pyrazinoylhydrazine-derived compounds have been prepared and characterized on the basis of analytical data and various physicochemical studies. Distorted octahedral structures for all the complexes have been proposed. The synthesized ligands and their complexes have been screened for their antibacterial activity against bacterial species Escherichia coli, Pseudomonas aeruginosa, Staphylococcus aureus and Klebsiella pneumonae. The activity data show the metal complexes to be more active than the parent free ligands against one or more bacterial species. PMID:18475857

Development of Novel DNA Cleavage Systems Based on Copper Complexes. Synthesis and Characterisation of Cu(II) Complexes of Hydroxyflavones

PubMed Central

el Amrani, F. Ben-Allal; Perelló, L.; Torres, L.

2000-01-01

Copper(II) complexes of several hydroxyflavones were prepared and characterised through their physico-chemical properties. The nuclease activity of three synthesised complexes is reported. These copper(II) complexes present more nuclease activity than the ligands and the copper(II) ion. PMID:18475969

Control of C-H Bond Activation by Mo-Oxo Complexes: pKa or Bond Dissociation Free Energy (BDFE)?

PubMed

Nazemi, Azadeh; Cundari, Thomas R

2017-10-16

A density functional theory (DFT) study (BMK/6-31+G(d)) was initiated to investigate the activation of benzylic carbon-hydrogen bonds by a molybdenum-oxo complex with a potentially redox noninnocent supporting ligand-a simple mimic of the active species of the enzyme ethylbenzene dehydrogenase (EBDH)-through deprotonation (C-H bond heterolysis) or hydrogen atom abstraction (C-H bond homolysis) routes. Activation free-energy barriers for neutral and anionic Mo-oxo complexes were high, but lower for anionic complexes than neutral complexes. Interesting trends as a function of substituents were observed that indicated significant H δ+ character in the transition states (TS), which was further supported by the preference for [2 + 2] addition over HAA for most complexes. Hence, it was hypothesized that C-H activation by these EBDH mimics is controlled more by the pK a than by the bond dissociation free energy of the C-H bond being activated. Therefore, the results suggest promising pathways for designing more efficient and selective catalysts for hydrocarbon oxidation based on EBDH active-site mimics.

Complement activation by ligand-driven juxtaposition of discrete pattern recognition complexes

PubMed Central

Degn, Søren E.; Kjaer, Troels R.; Kidmose, Rune T.; Jensen, Lisbeth; Hansen, Annette G.; Tekin, Mustafa; Jensenius, Jens C.; Andersen, Gregers R.; Thiel, Steffen

2014-01-01

Defining mechanisms governing translation of molecular binding events into immune activation is central to understanding immune function. In the lectin pathway of complement, the pattern recognition molecules (PRMs) mannan-binding lectin (MBL) and ficolins complexed with the MBL-associated serine proteases (MASP)-1 and MASP-2 cleave C4 and C2 to generate C3 convertase. MASP-1 was recently found to be the exclusive activator of MASP-2 under physiological conditions, yet the predominant oligomeric forms of MBL carry only a single MASP homodimer. This prompted us to investigate whether activation of MASP-2 by MASP-1 occurs through PRM-driven juxtaposition on ligand surfaces. We demonstrate that intercomplex activation occurs between discrete PRM/MASP complexes. PRM ligand binding does not directly escort the transition of MASP from zymogen to active enzyme in the PRM/MASP complex; rather, clustering of PRM/MASP complexes directly causes activation. Our results support a clustering-based mechanism of activation, fundamentally different from the conformational model suggested for the classical pathway of complement. PMID:25197071

Synthesis, crystal structure and spectroscopy of bioactive Cd(II) polymeric complex of the non-steroidal anti-inflammatory drug diclofenac sodium: Antiproliferative and biological activity

NASA Astrophysics Data System (ADS)

Tabrizi, Leila; Chiniforoshan, Hossein; McArdle, Patrick

2015-02-01

The interaction of Cd(II) with the non-steroidal anti-inflammatory drug diclofenac sodium (Dic) leads to the formation of the complex [Cd2(L)41.5(MeOH)2(H2O)]n(L = Dic), 1, which has been isolated and structurally characterized by X-ray crystallography. Diclofenac sodium and its metal complex 1 have also been evaluated for antiproliferative activity in vitro against the cells of three human cancer cell lines, MCF-7 (breast cancer cell line), T24 (bladder cancer cell line), A-549 (non-small cell lung carcinoma), and a mouse fibroblast L-929 cell line. The results of cytotoxic activity in vitro expressed as IC50 values indicated the diclofenac sodium and cadmium chloride are non active or less active than the metal complex of diclofenac (1). Complex 1 was also found to be a more potent cytotoxic agent against T-24 and MCF-7 cancer cell lines than the prevalent benchmark metallodrug, cisplatin, under the same experimental conditions. The superoxide dismutase activity was measured by Fridovich test which showed that complex 1 shows a low value in comparison with Cu complexes. The binding properties of this complex to biomolecules, bovine or human serum albumin, are presented and evaluated. Antibacterial and growth inhibitory activity is also higher than that of the parent ligand compound.

DNA Cleavage, Cytotoxic Activities, and Antimicrobial Studies of Ternary Copper(II) Complexes of Isoxazole Schiff Base and Heterocyclic Compounds

PubMed Central

Chityala, Vijay Kumar; Sathish Kumar, K.; Macha, Ramesh; Tigulla, Parthasarathy; Shivaraj

2014-01-01

Novel mixed ligand bivalent copper complexes [Cu. L. A. ClO 4] and [Cu. L. A] where “L” is Schiff bases, namely 2-((3,4-dimethylisoxazol-5-ylimino)methyl)-4-bromophenol (DMIIMBP)/2-((3,4-dimethylisoxazol-5-ylimino)methyl)-4-chlorophenol (DMIIMCP), and “A” is heterocyclic compound, such as 1,10-phenanthroline (phen)/2,21-bipyridyl (bipy)/8-hydroxyquinoline (oxine)/5-chloro-8-hydroxyquinoline (5-Cl-oxine), have been synthesized. These complexes have been characterized by IR, UV-Vis, ESR, elemental analysis, magnetic moments, TG, and DTA. On the basis of spectral studies and analytical data, five-coordinated square pyramidal/four-coordinated square planar geometry is assigned to all complexes. The ligands and their ternary complexes with Cu(II) have been screened for antimicrobial activity against bacteria and fungi by paper disc method. The antimicrobial studies of Schiff bases and their metal complexes showed significant activity and further it is observed that the metal complexes showed more activity than corresponding Schiff bases. In vitro antitumor activity of Cu(II) complexes was assayed against human cervical carcinoma (HeLa) cancer cells and it was observed that few complexes exhibit good antitumor activity on HeLa cell lines. The DNA cleavage studies have also been carried out on pBR 322 and it is observed that these Cu(II) complexes are capable of cleaving supercoiled plasmid DNA in the presence of H2O2 and UV light. PMID:24895493

Mononuclear nonheme iron(III) complexes that show superoxide dismutase-like activity and antioxidant effects against menadione-mediated oxidative stress.

PubMed

Hitomi, Yutaka; Iwamoto, Yuji; Kashida, Akihiro; Kodera, Masahito

2015-05-21

This communication describes the superoxide dismutase (SOD)-like activity of mononuclear iron(III) complexes with pentadentate monocarboxylamido ligands. The SOD activity can be controlled by the electronic nature of the substituent group on the ligand. The nitro-substituted complex showed clear cytoprotective activity against menadione-mediated oxidative stress in cultured cells.

Using activity theory to study cultural complexity in medical education.

PubMed

Frambach, Janneke M; Driessen, Erik W; van der Vleuten, Cees P M

2014-06-01

There is a growing need for research on culture, cultural differences and cultural effects of globalization in medical education, but these are complex phenomena to investigate. Socio-cultural activity theory seems a useful framework to study cultural complexity, because it matches current views on culture as a dynamic process situated in a social context, and has been valued in diverse fields for yielding rich understandings of complex issues and key factors involved. This paper explains how activity theory can be used in (cross-)cultural medical education research. We discuss activity theory's theoretical background and principles, and we show how these can be applied to the cultural research practice by discussing the steps involved in a cross-cultural study that we conducted, from formulating research questions to drawing conclusions. We describe how the activity system, the unit of analysis in activity theory, can serve as an organizing principle to grasp cultural complexity. We end with reflections on the theoretical and practical use of activity theory for cultural research and note that it is not a shortcut to capture cultural complexity: it is a challenge for researchers to determine the boundaries of their study and to analyze and interpret the dynamics of the activity system.

HTLV-1 Tax Induces Formation of the Active Macromolecular IKK Complex by Generating Lys63- and Met1-Linked Hybrid Polyubiquitin Chains.

PubMed

Shibata, Yuri; Tokunaga, Fuminori; Goto, Eiji; Komatsu, Ginga; Gohda, Jin; Saeki, Yasushi; Tanaka, Keiji; Takahashi, Hirotaka; Sawasaki, Tatsuya; Inoue, Satoshi; Oshiumi, Hiroyuki; Seya, Tsukasa; Nakano, Hiroyasu; Tanaka, Yuetsu; Iwai, Kazuhiro; Inoue, Jun-Ichiro

2017-01-01

The Tax protein of human T-cell leukemia virus type 1 (HTLV-1) is crucial for the development of adult T-cell leukemia (ATL), a highly malignant CD4+ T cell neoplasm. Among the multiple aberrant Tax-induced effects on cellular processes, persistent activation of transcription factor NF-κB, which is activated only transiently upon physiological stimulation, is essential for leukemogenesis. We and others have shown that Tax induces activation of the IκB kinase (IKK) complex, which is a critical step in NF-κB activation, by generating Lys63-linked polyubiquitin chains. However, the molecular mechanism underlying Tax-induced IKK activation is controversial and not fully understood. Here, we demonstrate that Tax recruits linear (Met1-linked) ubiquitin chain assembly complex (LUBAC) to the IKK complex and that Tax fails to induce IKK activation in cells that lack LUBAC activity. Mass spectrometric analyses revealed that both Lys63-linked and Met1-linked polyubiquitin chains are associated with the IKK complex. Furthermore, treatment of the IKK-associated polyubiquitin chains with Met1-linked-chain-specific deubiquitinase (OTULIN) resulted in the reduction of high molecular weight polyubiquitin chains and the generation of short Lys63-linked ubiquitin chains, indicating that Tax can induce the generation of Lys63- and Met1-linked hybrid polyubiquitin chains. We also demonstrate that Tax induces formation of the active macromolecular IKK complex and that the blocking of Tax-induced polyubiquitin chain synthesis inhibited formation of the macromolecular complex. Taken together, these results lead us to propose a novel model in which the hybrid-chain-dependent oligomerization of the IKK complex triggered by Tax leads to trans-autophosphorylation-mediated IKK activation.

HTLV-1 Tax Induces Formation of the Active Macromolecular IKK Complex by Generating Lys63- and Met1-Linked Hybrid Polyubiquitin Chains

PubMed Central

Tokunaga, Fuminori; Goto, Eiji; Komatsu, Ginga; Saeki, Yasushi; Tanaka, Keiji; Takahashi, Hirotaka; Sawasaki, Tatsuya; Inoue, Satoshi; Oshiumi, Hiroyuki; Seya, Tsukasa; Nakano, Hiroyasu; Tanaka, Yuetsu; Iwai, Kazuhiro

2017-01-01

The Tax protein of human T-cell leukemia virus type 1 (HTLV-1) is crucial for the development of adult T-cell leukemia (ATL), a highly malignant CD4+ T cell neoplasm. Among the multiple aberrant Tax-induced effects on cellular processes, persistent activation of transcription factor NF-κB, which is activated only transiently upon physiological stimulation, is essential for leukemogenesis. We and others have shown that Tax induces activation of the IκB kinase (IKK) complex, which is a critical step in NF-κB activation, by generating Lys63-linked polyubiquitin chains. However, the molecular mechanism underlying Tax-induced IKK activation is controversial and not fully understood. Here, we demonstrate that Tax recruits linear (Met1-linked) ubiquitin chain assembly complex (LUBAC) to the IKK complex and that Tax fails to induce IKK activation in cells that lack LUBAC activity. Mass spectrometric analyses revealed that both Lys63-linked and Met1-linked polyubiquitin chains are associated with the IKK complex. Furthermore, treatment of the IKK-associated polyubiquitin chains with Met1-linked-chain-specific deubiquitinase (OTULIN) resulted in the reduction of high molecular weight polyubiquitin chains and the generation of short Lys63-linked ubiquitin chains, indicating that Tax can induce the generation of Lys63- and Met1-linked hybrid polyubiquitin chains. We also demonstrate that Tax induces formation of the active macromolecular IKK complex and that the blocking of Tax-induced polyubiquitin chain synthesis inhibited formation of the macromolecular complex. Taken together, these results lead us to propose a novel model in which the hybrid-chain-dependent oligomerization of the IKK complex triggered by Tax leads to trans-autophosphorylation-mediated IKK activation. PMID:28103322

The effect of menadione on glutathione S-transferase A1 (GSTA1): c-Jun N-terminal kinase (JNK) complex dissociation in human colonic adenocarcinoma Caco-2 cells.

PubMed

Adnan, Humaira; Antenos, Monica; Kirby, Gordon M

2012-10-02

Glutathione S-transferases (GSTs) act as modulators of mitogen-activated protein kinase signal transduction pathways via a mechanism involving protein-protein interactions. We have demonstrated that GSTA1 forms complexes with JNK and modifies JNK activation during cellular stress, but the factors that influence complex association and dissociation are unknown. We hypothesized that menadione causes dissociation of GSTA1-JNK complexes, activates JNK, and the consequences of menadione exposure depend on GSTA1 expression. We demonstrate that menadione causes GSTA1-JNK dissociation and JNK activation in preconfluent Caco-2 cells, whereas postconfluent cells are resistant to this effect. Moreover, preconfluent cells are more sensitive than postconfluent cells to menadione-induced cytotoxicity. Activation of JNK is transient since removal of menadione causes GSTA1 to re-associate with JNK reducing cytotoxicity. Over-expression and knockdown of GSTA1 did not alter JNK activation by menadione or sensitivity to menadione-induced cytotoxicity. These results indicate that GSTA1-JNK complex integrity does not affect the ability of menadione to activate JNK. N-acetyl cysteine prevents GSH depletion and blocks menadione-induced complex dissociation, JNK activation and inhibits menadione-induced cytotoxicity. JNK activation and inhibits menadione-induced cytotoxicity. The data suggest that the mechanism of menadione-induced JNK activation involves the production of reactive oxygen species, likely superoxide anion, and intracellular GSH levels play an important role in preventing GSTA1-JNK complex dissociation, subsequent JNK activation and induction of cytotoxicity. Copyright © 2012 Elsevier Ireland Ltd. All rights reserved.

Single and double C-Cl-activation of methylene chloride by P,N-ligand coordinated rhodium complexes.

PubMed

Blank, Benoît; Glatz, Germund; Kempe, Rhett

2009-02-02

Two in one: The simultaneous formation of bimetallic mu-methylene bridged Rh(III) complexes as well as dimeric Rh(III) complexes with terminal chloromethyl groups is observed for P,N-ligand stabilized Rh(I) complexes by C-Cl bond activation of methylene chloride. A mechanistic proposal for the formation of both activation products is also discussed. The synthesis of Rh(I) complexes with P-functionalized aminopyridine ligands is reported as well as the first simultaneous observation of a single and double activation of C-Cl bonds of methylene chloride affording both a dimeric Rh(III) complex bearing terminal CH(2)Cl groups in addition to a binuclear Rh(III) complex with a bridging mu-CH(2) group. The structures of the oxidative addition products were obtained by X-ray diffraction studies and NMR experiments were performed to elucidate some aspects of the reaction pathway.

Oxidation-promoted activation of a ferrocene C-H bond by a rhodium complex.

PubMed

Labande, Agnès; Debono, Nathalie; Sournia-Saquet, Alix; Daran, Jean-Claude; Poli, Rinaldo

2013-05-14

The oxidation of a rhodium(I) complex containing a ferrocene-based heterodifunctional phosphine N-heterocyclic carbene (NHC) ligand produces a stable, planar chiral rhodium(III) complex with an unexpected C-H activation on ferrocene. The oxidation of rhodium(I) to rhodium(III) may be accomplished by initial oxidation of ferrocene to ferrocenium and subsequent electron transfer from rhodium to ferrocenium. Preliminary catalytic tests showed that the rhodium(III) complex is active for the Grignard-type arylation of 4-nitrobenzaldehyde via C-H activation of 2-phenylpyridine.

Synthesis and antitumor activity of seleno- and thio-purines complexed with cis-diamminoplatinum (II).

PubMed

Maeda, M; Abiko, N; Sasaki, T

1982-02-01

cis-Diamminoplatinum (II) complexes with selenoguanine, thioguanine, 6-thioxanthine, or 6-mercaptopurine were synthesized by the reaction of stoichiometric amounts of selenopurine or thiopurine with aquated cis-dichlorodimmineplatinum (II) in slightly acidic medium, and their antitumor activity was studied against L1210 cells in mice. These compounds exhibited a medium antitumor activity with very low toxicity. The antitumor activity was dependent on the nature of the purine ligand. These complexes were very stable in various aqueous solvents at 37 degrees C for 10 d but not in the presence of mouse serum. The mechanism of the action effected by the complex is not clear. However, the slow release of an antitumor active purine from the complex, SeG-Pt (NH3)2, was observed.

In vitro inhibition of hyaluronidase by sodium copper chlorophyllin complex and chlorophyllin analogs

PubMed Central

McCook, John P; Dorogi, Peter L; Vasily, David B; Cefalo, Dustin R

2015-01-01

Background Inhibitors of hyaluronidase are potent agents that maintain hyaluronic acid homeostasis and may serve as anti-aging, anti-inflammatory, and anti-microbial agents. Sodium copper chlorophyllin complex is being used therapeutically as a component in anti-aging cosmeceuticals, and has been shown to have anti-hyaluronidase activity. In this study we evaluated various commercial lots of sodium copper chlorophyllin complex to identify the primary small molecule constituents, and to test various sodium copper chlorophyllin complexes and their small molecule analog compounds for hyaluronidase inhibitory activity in vitro. Ascorbate analogs were tested in combination with copper chlorophyllin complexes for potential additive or synergistic activity. Materials and methods For hyaluronidase activity assays, dilutions of test materials were evaluated for hydrolytic activity of hyaluronidase by precipitation of non-digested hyaluronate by measuring related turbidity at 595 nm. High-performance liquid chromatography and mass spectroscopy was used to analyze and identify the primary small molecule constituents in various old and new commercial lots of sodium copper chlorophyllin complex. Results The most active small molecule component of sodium copper chlorophyllin complex was disodium copper isochlorin e4, followed by oxidized disodium copper isochlorin e4. Sodium copper chlorophyllin complex and copper isochlorin e4 disodium salt had hyaluronidase inhibitory activity down to 10 µg/mL. The oxidized form of copper isochlorin e4 disodium salt had substantial hyaluronidase inhibitory activity at 100 µg/mL but not at 10 µg/mL. Ascorbate derivatives did not enhance the hyaluronidase inhibitory activity of sodium copper chlorophyllin. Copper isochlorin e4 analogs were always the dominant components of the small molecule content of the commercial lots tested; oxidized copper isochlorin e4 was found in increased concentrations in older compared to newer lots tested. Conclusion These results support the concept of using the hyaluronidase inhibitory activity of sodium copper chlorophyllin complex to increase the hyaluronic acid level of the dermal extracellular matrix for the improvement of the appearance of aging facial skin. PMID:26300653

In vitro inhibition of hyaluronidase by sodium copper chlorophyllin complex and chlorophyllin analogs.

PubMed

McCook, John P; Dorogi, Peter L; Vasily, David B; Cefalo, Dustin R

2015-01-01

Inhibitors of hyaluronidase are potent agents that maintain hyaluronic acid homeostasis and may serve as anti-aging, anti-inflammatory, and anti-microbial agents. Sodium copper chlorophyllin complex is being used therapeutically as a component in anti-aging cosmeceuticals, and has been shown to have anti-hyaluronidase activity. In this study we evaluated various commercial lots of sodium copper chlorophyllin complex to identify the primary small molecule constituents, and to test various sodium copper chlorophyllin complexes and their small molecule analog compounds for hyaluronidase inhibitory activity in vitro. Ascorbate analogs were tested in combination with copper chlorophyllin complexes for potential additive or synergistic activity. For hyaluronidase activity assays, dilutions of test materials were evaluated for hydrolytic activity of hyaluronidase by precipitation of non-digested hyaluronate by measuring related turbidity at 595 nm. High-performance liquid chromatography and mass spectroscopy was used to analyze and identify the primary small molecule constituents in various old and new commercial lots of sodium copper chlorophyllin complex. The most active small molecule component of sodium copper chlorophyllin complex was disodium copper isochlorin e4, followed by oxidized disodium copper isochlorin e4. Sodium copper chlorophyllin complex and copper isochlorin e4 disodium salt had hyaluronidase inhibitory activity down to 10 µg/mL. The oxidized form of copper isochlorin e4 disodium salt had substantial hyaluronidase inhibitory activity at 100 µg/mL but not at 10 µg/mL. Ascorbate derivatives did not enhance the hyaluronidase inhibitory activity of sodium copper chlorophyllin. Copper isochlorin e4 analogs were always the dominant components of the small molecule content of the commercial lots tested; oxidized copper isochlorin e4 was found in increased concentrations in older compared to newer lots tested. These results support the concept of using the hyaluronidase inhibitory activity of sodium copper chlorophyllin complex to increase the hyaluronic acid level of the dermal extracellular matrix for the improvement of the appearance of aging facial skin.

Anticancer, antibacterial and antifungal activity of new ni (ii) and cu (ii) complexes of imidazole-phenanthroline derivatives.

PubMed

Moghadam, Mahboube Eslami; Divsalar, Adeleh; Zare, Marziye Shahraki; Gholizadeh, Roghayeh; Mahalleh, Doran; Saghatforosh, Lotfali; Sanati, Soheila

2017-11-02

Two new nickel(II) and copper(II) complexes of 2-(Furan-2-yl)-1H-Imidazo[4,5-f][1,10]Phenanthroline (FIP) and 2-(thiophen-2-yl)-1H-imidazo[4,5-f][1,10]phenanthroline (TIP), imidazophen derivatives were synthesized. The structures of the compounds were determined by UV-visible and FT-IR spectroscopic methods and elemental analysis. The biological activities of Ni and Cu complexes, as anticancer agents, were tested against chronic myelogenous leukemia cell line, K562, at micromolar concentration. The MTT studies showed Cc 50 values are 21 and 160 µM for Cu and Ni(II) complexes, respectively; suggesting that Ni (II) complex has Cc 50 almost seven times of that obtained for cisplatin. Biological activity of the Ni(II) and Cu(II) complexes were also assayed against selective microorganisms by disc diffusion method. These results showed that the Cu(II) complex is antifungal agent but Ni(II) complex has antibacterial activity.

The Candida albicans Inhibitory Activity of the Extract from Papaya (Carica papaya L.) Seed Relates to Mitochondria Dysfunction.

PubMed

Zhang, Tao; Chen, Weijun

2017-08-25

The inhibitory activity of the papaya seed extract (PSE) on Candida albicans ( C. albicans ) was determined by turbidimetry method. The inhibitory mechanisms were also evaluated from the prospective of reactive oxygen species (ROS) generation, mitochondrial membrane potential (MMP) decrease, and the activities of four complex enzymes in mitochondria respiratory chain. Results obtained from this study indicated that the PSE exhibited an effective inhibitory activity on C. albicans and induced significant accumulation of ROS and collapse of MMP. The Complex I and Complex III exhibited continues significant decrease in mitochondrial enzyme activity assays, but the Complex II and Complex IV activities were not positively correlated. Furthermore, the GC-MS analysis demonstrated that the PSE represents a rich and high-purity source of benzyl isothiocyanate (BITC), which indicated the BITC may be responsible for the mitochondrial dysfunction.

NADH-ubiquinone oxidoreductase activity in the kinetoplasts of the plant trypanosomatid Phytomonas serpens.

PubMed

González-Halphen, Diego; Maslov, Dmitri A

2004-03-01

NADH-ubiquinone oxidoreductase activity is present in mitochondrial lysates of Phytomonas serpens. Rotenone at 2-10 microM inhibited the activity 50-75%, indicating that it belongs to respiratory complex I. The activity was also inhibited 50-60% in the presence of 10-30 nM atovaquone suggesting that inhibition of complex I represents a likely mechanism of the known antileishmanial activity of this drug. The complex was partially purified by chromatography on DEAE-Sepharose CL-6B and gel-filtration on Sepharose CL-2B. The NADH:ubiquinone oxidoreductase activity in this preparation was completely inactivated by 20 nM atovaquone. The partially purified complex was present in a low amount and its subunits could not be discerned by staining with Coomassie. However, one of its components, a homologue of the 39 kDa subunit of the bovine complex I, was identified immunochemically in the original lysate and in the partially purified material.

Changes in complex spike activity during classical conditioning

PubMed Central

Rasmussen, Anders; Jirenhed, Dan-Anders; Wetmore, Daniel Z.; Hesslow, Germund

2014-01-01

The cerebellar cortex is necessary for adaptively timed conditioned responses (CRs) in eyeblink conditioning. During conditioning, Purkinje cells acquire pause responses or “Purkinje cell CRs” to the conditioned stimuli (CS), resulting in disinhibition of the cerebellar nuclei (CN), allowing them to activate motor nuclei that control eyeblinks. This disinhibition also causes inhibition of the inferior olive (IO), via the nucleo-olivary pathway (N-O). Activation of the IO, which relays the unconditional stimulus (US) to the cortex, elicits characteristic complex spikes in Purkinje cells. Although Purkinje cell activity, as well as stimulation of the CN, is known to influence IO activity, much remains to be learned about the way that learned changes in simple spike firing affects the IO. In the present study, we analyzed changes in simple and complex spike firing, in extracellular Purkinje cell records, from the C3 zone, in decerebrate ferrets undergoing training in a conditioning paradigm. In agreement with the N-O feedback hypothesis, acquisition resulted in a gradual decrease in complex spike activity during the conditioned stimulus, with a delay that is consistent with the long N-O latency. Also supporting the feedback hypothesis, training with a short interstimulus interval (ISI), which does not lead to acquisition of a Purkinje cell CR, did not cause a suppression of complex spike activity. In contrast, observations that extinction did not lead to a recovery in complex spike activity and the irregular patterns of simple and complex spike activity after the conditioned stimulus are less conclusive. PMID:25140129

Respiratory chain supercomplexes associate with the cysteine desulfurase complex of the iron–sulfur cluster assembly machinery

PubMed Central

Böttinger, Lena; Mårtensson, Christoph U.; Song, Jiyao; Zufall, Nicole; Wiedemann, Nils; Becker, Thomas

2018-01-01

Mitochondria are the powerhouses of eukaryotic cells. The activity of the respiratory chain complexes generates a proton gradient across the inner membrane, which is used by the F1FO-ATP synthase to produce ATP for cellular metabolism. In baker’s yeast, Saccharomyces cerevisiae, the cytochrome bc1 complex (complex III) and cytochrome c oxidase (complex IV) associate in respiratory chain supercomplexes. Iron–sulfur clusters (ISC) form reactive centers of respiratory chain complexes. The assembly of ISC occurs in the mitochondrial matrix and is essential for cell viability. The cysteine desulfurase Nfs1 provides sulfur for ISC assembly and forms with partner proteins the ISC-biogenesis desulfurase complex (ISD complex). Here, we report an unexpected interaction of the active ISD complex with the cytochrome bc1 complex and cytochrome c oxidase. The individual deletion of complex III or complex IV blocks the association of the ISD complex with respiratory chain components. We conclude that the ISD complex binds selectively to respiratory chain supercomplexes. We propose that this molecular link contributes to coordination of iron–sulfur cluster formation with respiratory activity. PMID:29386296

Synthesis, characterization and antimicrobial studies of Schiff base complexes

NASA Astrophysics Data System (ADS)

Zafar, Hina; Ahmad, Anis; Khan, Asad U.; Khan, Tahir Ali

2015-10-01

The Schiff base complexes, MLCl2 [M = Fe(II), Co(II), Ni(II), Cu(II) and Zn(II)] have been synthesized by the template reaction of respective metal ions with 2-acetylpyrrole and 1,3-diaminopropane in 1:2:1 M ratio. The complexes have been characterized by elemental analyses, ESI - mass, NMR (1H and 13C), IR, XRD, electronic and EPR spectral studies, magnetic susceptibility and molar conductance measurements. These studies show that all the complexes have octahedral arrangement around the metal ions. The molar conductance measurements of all the complexes in DMSO indicate their non-electrolytic nature. The complexes were screened for their antibacterial activity in vitro against Gram-positive (Streptococcus pyogenes) and Gram-negative (Klebsiella pneumoniae) bacteria. Among the metal complexes studied the copper complex [CuLCl2], showed highest antibacterial activity nearly equal to standard drug ciprofloxacin. Other complexes also showed considerable antibacterial activity. The relative order of activity against S. Pyogenes is as Cu(II) > Zn(II) > Co(II) = Fe(II) > Ni(II) and with K. Pneumonia is as Cu(II) > Co(II) > Zn(II) > Fe(II) > Ni(II).

Synthesis, crystal structures, molecular docking, and in vitro biological activities of transition metals with 4-(2,3-dichlorophenyl)piperazine-1-carboxylic acid.

PubMed

Yang, Dan-Dan; Chen, Ya-Nan; Wu, Yu-Shan; Wang, Rui; Chen, Zhi-Jian; Qin, Jie; Qian, Shao-Song; Zhu, Hai-Liang

2016-07-15

Four novel mononuclear complexes, [Cd(L)2·2H2O] (1), [Ni(L)2·2H2O] (2) [Cu(L)2·H2O] (3), and [Zn(L)2·2H2O] (4) (CCDC numbers: 1444630-1444633 for complexes 1-4) (HL=4-(2,3-dichlorophenyl)piperazine-1-carboxylic acid) were synthesized, and have been characterized by IR spectroscopy, elemental analysis, and X-ray crystallography. Molecular docking study preliminarily revealed that complex 1 had potential telomerase inhibitory activity. In accordance with the result of calculation, in vitro tests of the inhibitory activities of complex 1 against telomerase showed complex 1 (IC50=8.17±0.91μM) had better inhibitory activities, while complexes 2, 3 and 4 showed no inhibitory activities. Antiproliferative activity in human cancer cell line HepG2 was further determined by MTT assays. The IC50 value (6.5±0.2μM) for the complex 1 having good inhibitory activity against HepG2 was at the same micromolar concentrations with cis-platinum (2.2±1.2μM). While the IC50 value for the metal-free ligand, complex 2, 3 and 4 was more than 100μM. These results indicated that telomerase was potentially an anticancer drug target and showed that complex 1 was a potent inhibitor of human telomerase as well as an antiproliferative compound. Copyright © 2016 Elsevier Ltd. All rights reserved.

Synthesis, spectroscopic, anticancer, antibacterial and antifungal studies of Ni(II) and Cu(II) complexes with hydrazine carboxamide, 2-[3-methyl-2-thienyl methylene

NASA Astrophysics Data System (ADS)

Chandra, Sulekh; Vandana; Kumar, Suresh

2015-01-01

Schiff's base ligand(L) hydrazine carboxamide, 2-[3-methyl-2-thienyl methylene] and its metal complexes have been synthesized and characterized by elemental analysis, molar conductance, various spectroscopic techniques such as electronic, IR, 1H NMR, mass, EPR. Molar conductance of complexes in DMF solution corresponds to non-electrolyte. Complexes have general composition [M(L)2X2], where M = Ni(II) and Cu(II), X = Cl-, NO3-, CH3COO- and ½SO42-. On the basis of above spectral studies, an octahedral geometry has been assigned for Ni(II) complexes and tetragonal geometry for Cu(II) complexes except [Cu(L)2SO4] which possesses five coordinated trigonal bipyramidal geometry. These metal complexes were also tested for their anticancer, antibacterial and antifungal activities to assess their inhibition potential. Anticancer activity of ligand and its metal complexes were evaluated using SRB fluorometric assay and Adriamycin (ADR) was applied as positive control. Schiff's base ligand and its metal complexes were screened for their antibacterial and antifungal activity against Escherichia coli, Bacillus cereus and Aspergillus niger, Aspergillus flavus, respectively. Kirby-Bauer single disk susceptibility test was used for antibacterial activity and well diffusion method for antifungal activity of the compounds on the used fungi.

Budding Yeast Silencing Complexes and Regulation of Sir2 Activity by Protein-Protein Interactions

PubMed Central

Tanny, Jason C.; Kirkpatrick, Donald S.; Gerber, Scott A.; Gygi, Steven P.; Moazed, Danesh

2004-01-01

Gene silencing in the budding yeast Saccharomyces cerevisiae requires the enzymatic activity of the Sir2 protein, a highly conserved NAD-dependent deacetylase. In order to study the activity of native Sir2, we purified and characterized two budding yeast Sir2 complexes: the Sir2/Sir4 complex, which mediates silencing at mating-type loci and at telomeres, and the RENT complex, which mediates silencing at the ribosomal DNA repeats. Analyses of the protein compositions of these complexes confirmed previously described interactions. We show that the assembly of Sir2 into native silencing complexes does not alter its selectivity for acetylated substrates, nor does it allow the deacetylation of nucleosomal histones. The inability of Sir2 complexes to deacetylate nucleosomes suggests that additional factors influence Sir2 activity in vivo. In contrast, Sir2 complexes show significant enhancement in their affinities for acetylated substrates and their sensitivities to the physiological inhibitor nicotinamide relative to recombinant Sir2. Reconstitution experiments showed that, for the Sir2/Sir4 complex, these differences stem from the physical interaction of Sir2 with Sir4. Finally, we provide evidence that the different nicotinamide sensitivities of Sir2/Sir4 and RENT in vitro could contribute to locus-specific differences in how Sir2 activity is regulated in vivo. PMID:15282295

Drug Delivery Systems For Anti-Cancer Active Complexes of Some Coinage Metals.

PubMed

Zhang, Ming; Saint-Germain, Camille; He, Guiling; Sun, Raymond Wai-Yin

2018-02-12

Although cisplatin and a number of platinum complexes have widely been used for the treatment of neoplasia, patients receiving these treatments have frequently suffered from their severe toxic side effects, the development of resistance with consequent relapse. In the recent decades, numerous complexes of coinage metals including that of gold, copper and silver have been reported to display promising in vitro and/or in vivo anti-cancer activities as well as potent activities towards cisplatin-resistant tumors. Nevertheless, the medical development of these metal complexes has been hampered by their instability in aqueous solutions and the nonspecific binding in biological systems. One of the approaches to overcome these problems is to design and develop adequate drug delivery systems (DDSs) for the transport of these complexes. By functionalization, encapsulation or formulation of the metal complexes, several types of DDSs have been reported to improve the desired pharmacological profile of the metal complexes, improving their overall stability, bioavailability, anti-cancer activity and reducing their toxicity towards normal cells. In this review, we summarized the recent findings for different DDSs for various anti- cancer active complexes of some coinage metals. Copyright© Bentham Science Publishers; For any queries, please email at epub@benthamscience.org.

Lessons from isolable nickel(I) precursor complexes for small molecule activation.

PubMed

Yao, Shenglai; Driess, Matthias

2012-02-21

Small-molecule activation by transition metals is essential to numerous organic transformations, both biological and industrial. Creating useful metal-mediated activation systems often depends on stabilizing the metal with uncommon low oxidation states and low coordination numbers. This provides a redox-active metal center with vacant coordination sites well suited for interacting with small molecules. Monovalent nickel species, with their d(9) electronic configuration, are moderately strong one-electron reducing agents that are synthetically attractive if they can be isolated. They represent suitable reagents for closing the knowledge gap in nickel-mediated activation of small molecules. Recently, the first strikingly stable dinuclear β-diketiminate nickel(I) precursor complexes were synthesized, proving to be suitable promoters for small-molecule binding and activation. They have led to many unprecedented nickel complexes bearing activated small molecules in different reduction stages. In this Account, we describe selected achievements in the activation of nitrous oxide (N(2)O), O(2), the heavier chalcogens (S, Se, and Te), and white phosphorus (P(4)) through this β-diketiminatonickel(I) precursor species. We emphasize the reductive activation of O(2), owing to its promise in oxidation processes. The one-electron-reduced O(2) activation product, that is, the corresponding β-diketiminato-supported Ni-O(2) complex, is a genuine superoxonickel(II) complex, representing an important intermediate in the early stages of O(2) activation. It selectively acts as an oxygen-atom transfer agent, hydrogen-atom scavenger, or both towards exogenous organic substrates to yield oxidation products. The one-electron reduction of the superoxonickel(II) moiety was examined by using elemental potassium, β-diketiminatozinc(II) chloride, and β-diketiminatoiron(I) complexes, affording the first heterobimetallic complexes featuring a [NiO(2)M] subunit (M is K, Zn, or Fe). Through density functional theory (DFT) calculations, the geometric and electronic structures of these complexes were established and their distinctive reactivity, including the unprecedented monooxygenase-like activity of a bis(μ-oxo)nickel-iron complex, was studied. The studies have further led to other heterobimetallic complexes containing a [NiO(2)M] core, which are useful for understanding the influence of the heterometal on structure-reactivity relationships. The activation of N(2)O led directly to the hydrogen-atom abstraction product bis(μ-hydroxo)nickel(II) species and prevented isolation of any intermediate. In contrast, the activation of elemental S, Se, and Te with the same nickel(I) reagent furnished activation products with superchalcogenido E(2)(-) (E is S, Se, or Te) and dichalcogenido E(2)(2-) ligand in different activation stages. The isolable supersulfidonickel(II) subunit may serve as a versatile building block for the synthesis of heterobimetallic disulfidonickel(II) complexes with a [NiS(2)M] core. In the case of white phosphorus, the P(4) molecule has been coordinated to the nickel(I) center of dinuclear β-diketiminatonickel(I) precursor complexes; however, the whole P(4) subunit is a weaker electron acceptor than the dichalcogen ligands E(2), thus remaining unreduced. This P(4) binding mode is rare and could open new doors for subsequent functionalization of P(4). Our advances in understanding how these small molecules are bound to a nickel(I) center and are activated for further transformation offer promise for designing new catalysts. These nickel-containing complexes offer exceptional potential for nickel-mediated transformations of organic molecules and as model compounds for mimicking active sites of nickel-containing metalloenzymes.

Synthesis, spectroscopic characterization, electrochemistry and biological evaluation of some binuclear transition metal complexes of bicompartmental ONO donor ligands containing benzo[b]thiophene moiety

NASA Astrophysics Data System (ADS)

Mahendra Raj, K.; Vivekanand, B.; Nagesh, G. Y.; Mruthyunjayaswamy, B. H. M.

2014-02-01

A series of new binucleating Cu(II), Co(II), Ni(II) and Zn(II) complexes of bicompartmental ligands with ONO donor were synthesized. The ligands were obtained by the condensation of 3-chloro-6-substituted benzo[b]thiophene-2-carbohydrazides and 4,6-diacetylresorcinol. The synthesized ligands and their complexes were characterized by elemental analysis and various spectroscopic techniques. Elemental analysis, IR, 1H NMR, ESI-mass, UV-Visible, TG-DTA, magnetic measurements, molar conductance and powder-XRD data has been used to elucidate their structures. The bonding sites are the oxygen atom of amide carbonyl, azomethine nitrogen and phenolic oxygen for ligands 1 and 2. The binuclear nature of the complexes was confirmed by ESR spectral data. TG-DTA studies for some complexes showed the presence of coordinated water molecules and the final product is the metal oxide. All the complexes were investigated for their electrochemical activity, only the Cu(II) complexes showed the redox property. Cu(II) complexes were square planar, whereas Co(II), Ni(II) and Zn(II) complexes were octahedral. Powder-XRD pattern have been studied in order to test the degree of crystallinity of the complexes and unit cell calculations were made. In order to evaluate the effect of antimicrobial activity of metal ions upon chelation, both the ligands and their metal complexes were screened for their antibacterial and antifungal activities by minimum inhibitory concentration (MIC) method. The results showed that the metal complexes were found to be more active than free ligands. The DNA cleaving capacities of all the complexes were analyzed by agarose gel electrophoresis method against supercoiled plasmid DNA. Among the compounds tested for antioxidant capacity, ligand 1 displayed excellent activity than its metal complexes.

The cellulose-binding activity of the PsB multiprotein complex is required for proper assembly of the spore coat and spore viability in Dictyostelium discoideum.

PubMed

Srinivasan, S; Griffiths, K R; McGuire, V; Champion, A; Williams, K L; Alexander, S

2000-08-01

The terminal event of spore differentiation in the cellular slime mould Dictyostelium discoideum is the assembly of the spore coat, which surrounds the dormant amoeba and allows the organism to survive during extended periods of environmental stress. The spore coat is a polarized extracellular matrix composed of glycoproteins and cellulose. The process of spore coat formation begins by the regulated secretion of spore coat proteins from the prespore vesicles (PSVs). Four of the major spore coat proteins (SP96, PsB/SP85, SP70 and SP60) exist as a preassembled multiprotein complex within the PSVs. This complete complex has an endogenous cellulose-binding activity. Mutant strains lacking either the SP96 or SP70 proteins produce partial complexes that do not have cellulose-binding activity, while mutants lacking SP60 produce a partial complex that retains this activity. Using a combination of immunofluorescence microscopy and biochemical methods we now show that the lack of cellulose-binding activity in the SP96 and SP70 mutants results in abnormally assembled spore coats and spores with greatly reduced viability. In contrast, the SP60 mutant, in which the PsB complex retains its cellulose-binding activity, produces spores with apparently unaltered structure and viability. Thus, it is the loss of the cellulose-binding activity of the PsB complex, rather than the mere loss of individual spore coat proteins, that results in compromised spore coat structure. These results support the idea that the cellulose-binding activity associated with the complete PsB complex plays an active role in the assembly of the spore coat.

Prompt and easy activation by specific thioredoxins of calvin cycle enzymes of Arabidopsis thaliana associated in the GAPDH/CP12/PRK supramolecular complex.

PubMed

Marri, Lucia; Zaffagnini, Mirko; Collin, Valérie; Issakidis-Bourguet, Emmanuelle; Lemaire, Stéphane D; Pupillo, Paolo; Sparla, Francesca; Miginiac-Maslow, Myroslawa; Trost, Paolo

2009-03-01

The Calvin cycle enzymes glyceraldehyde-3-phosphate dehydrogenase (GAPDH) and phosphoribulokinase (PRK) can form under oxidizing conditions a supramolecular complex with the regulatory protein CP12. Both GAPDH and PRK activities are inhibited within the complex, but they can be fully restored by reduced thioredoxins (TRXs). We have investigated the interactions of eight different chloroplast thioredoxin isoforms (TRX f1, m1, m2, m3, m4, y1, y2, x) with GAPDH (A(4), B(4), and B(8) isoforms), PRK and CP12 (isoform 2), all from Arabidopsis thaliana. In the complex, both A(4)-GAPDH and PRK were promptly activated by TRX f1, or more slowly by TRXs m1 and m2, but all other TRXs were ineffective. Free PRK was regulated by TRX f1, m1, or m2, while B(4)- and B(8)-GAPDH were absolutely specific for TRX f1. Interestingly, reductive activation of PRK caged in the complex was much faster than reductive activation of free oxidized PRK, and activation of A(4)-GAPDH in the complex was much faster (and less demanding in terms of reducing potential) than activation of free oxidized B(4)- or B(8)-GAPDH. It is proposed that CP12-assembled supramolecular complex may represent a reservoir of inhibited enzymes ready to be released in fully active conformation following reduction and dissociation of the complex by TRXs upon the shift from dark to low light. On the contrary, autonomous redox-modulation of GAPDH (B-containing isoforms) would be more suited to conditions of very active photosynthesis.

Stabilization and activation of alpha-chymotrypsin in water-organic solvent systems by complex formation with oligoamines.

PubMed

Kudryashova, Elena V; Artemova, Tatiana M; Vinogradov, Alexei A; Gladilin, Alexander K; Mozhaev, Vadim V; Levashov, Andrey V

2003-04-01

Formation of enzyme-oligoamine complexes was suggested as an approach to obtain biocatalysts with enhanced resistance towards inactivation in water-organic media. Complex formation results in broadening (by 20-40% v/v ethanol) of the range of cosolvent concentrations where the enzyme retains its catalytic activity (stabilization effect). At moderate cosolvent concentrations (20-40% v/v) complex formation activates the enzyme (by 3-6 times). The magnitude of activation and stabilization effects increases with the number of possible electrostatic contacts between the protein surface and the molecules of oligoamines (OA). Circular dichroism spectra in the far-UV region show that complex formation stabilizes protein conformation and prevents aggregation in water-organic solvent mixtures. Two populations of the complexes with different thermodynamic stabilities were found in alpha-chymotrypsin (CT)-OA systems depending on the CT/OA ratio. The average dissociation constants and stoichiometries of both low- and high-affinity populations of the complexes were estimated. It appears that it is the low-affinity sites on the CT surface that are responsible for the activation effect.

Analysis of the NuRD subunits reveals a histone deacetylase core complex and a connection with DNA methylation

PubMed Central

Zhang, Yi; Ng, Huck-Hui; Erdjument-Bromage, Hediye; Tempst, Paul; Bird, Adrian; Reinberg, Danny

1999-01-01

ATP-dependent nucleosome remodeling and core histone acetylation and deacetylation represent mechanisms to alter nucleosome structure. NuRD is a multisubunit complex containing nucleosome remodeling and histone deacetylase activities. The histone deacetylases HDAC1 and HDAC2 and the histone binding proteins RbAp48 and RbAp46 form a core complex shared between NuRD and Sin3-histone deacetylase complexes. The histone deacetylase activity of the core complex is severely compromised. A novel polypeptide highly related to the metastasis-associated protein 1, MTA2, and the methyl-CpG-binding domain-containing protein, MBD3, were found to be subunits of the NuRD complex. MTA2 modulates the enzymatic activity of the histone deacetylase core complex. MBD3 mediates the association of MTA2 with the core histone deacetylase complex. MBD3 does not directly bind methylated DNA but is highly related to MBD2, a polypeptide that binds to methylated DNA and has been reported to possess demethylase activity. MBD2 interacts with the NuRD complex and directs the complex to methylated DNA. NuRD may provide a means of gene silencing by DNA methylation. PMID:10444591

Synthesis, photoluminescence and biological properties of terbium(III) complexes with hydroxyketone and nitrogen containing heterocyclic ligands

NASA Astrophysics Data System (ADS)

Poonam; Kumar, Rajesh; Boora, Priti; Khatkar, Anurag; Khatkar, S. P.; Taxak, V. B.

2016-01-01

The ternary terbium(III) complexes [Tb(HDAP)3ṡbiq], [Tb(HDAP)3ṡdmph] and [Tb(HDAP)3ṡbathophen] were prepared by using methoxy substituted hydroxyketone ligand HDAP (2-hydroxy-4,6-dimethoxyacetophenone) and an ancillary ligand 2,2-biquinoline or 5,6-dimethyl-1,10-phenanthroline or bathophenanthroline respectively. The ligand and synthesized complexes were characterised based on elemental analysis, FT-IR and 1H NMR. Thermal behaviour of the synthesized complexes illustrates the general decomposition patterns of the complexes by thermogravimetric analysis. Photophysical properties such as excitation spectra, emission spectra and luminescence decay curves of the complexes were investigated in detail. The main green emitting peak at 548 nm can be attributed to 5D4 → 7F5 of Tb3+ ion. Thus, these complexes might be used to make a bright green light-emitting diode for display purpose. In addition the in vitro antibacterial activities of HDAP and its Tb(III) complexes against Bacillus subtilis, Staphylococcus aureus, Escherichia coli and antifungal activities against Candida albicans and Aspergillus niger are reported. The Tb3+ complexes were found to be more potent antimicrobial agent as compared to the ligand. Among all these complexes, [Tb(HDAP)3ṡbathophen] exhibited excellent antimicrobial activity which proves its potential usefulness as an antimicrobial agent. Furthermore, in vitro antioxidant activity tests were carried out by using DPPH method which indicates that the complexes have considerable antioxidant activity when compared with the standard ascorbic acid.

Optimization of WAVE2 complex–induced actin polymerization by membrane-bound IRSp53, PIP3, and Rac

PubMed Central

Suetsugu, Shiro; Kurisu, Shusaku; Oikawa, Tsukasa; Yamazaki, Daisuke; Oda, Atsushi; Takenawa, Tadaomi

2006-01-01

WAVE2 activates the actin-related protein (Arp) 2/3 complex for Rac-induced actin polymerization during lamellipodium formation and exists as a large WAVE2 protein complex with Sra1/PIR121, Nap1, Abi1, and HSPC300. IRSp53 binds to both Rac and Cdc42 and is proposed to link Rac to WAVE2. We found that the knockdown of IRSp53 by RNA interference decreased lamellipodium formation without a decrease in the amount of WAVE2 complex. Localization of WAVE2 at the cell periphery was retained in IRSp53 knockdown cells. Moreover, activated Cdc42 but not Rac weakened the association between WAVE2 and IRSp53. When we measured Arp2/3 activation in vitro, the WAVE2 complex isolated from the membrane fraction of cells was fully active in an IRSp53-dependent manner but WAVE2 isolated from the cytosol was not. Purified WAVE2 and purified WAVE2 complex were activated by IRSp53 in a Rac-dependent manner with PIP3-containing liposomes. Therefore, IRSp53 optimizes the activity of the WAVE2 complex in the presence of activated Rac and PIP3. PMID:16702231

Antiplasmodial activities of gold(I) complexes involving functionalized N-heterocyclic carbenes.

PubMed

Hemmert, Catherine; Ramadani, Arba Pramundita; Boselli, Luca; Fernández Álvarez, Álvaro; Paloque, Lucie; Augereau, Jean-Michel; Gornitzka, Heinz; Benoit-Vical, Françoise

2016-07-01

A series of twenty five molecules, including imidazolium salts functionalized by N-, O- or S-containing groups and their corresponding cationic, neutral or anionic gold(I) complexes were evaluated on Plasmodium falciparum in vitro and then on Vero cells to determine their selectivity. Among them, eight new compounds were synthesized and fully characterized by spectroscopic methods. The X-ray structures of three gold(I) complexes are presented. Except one complex (18), all the cationic gold(I) complexes show potent antiplasmodial activity with IC50 in the micro- and submicromolar range, correlated with their lipophilicity. Structure-activity relationships enable to evidence a lead-complex (21) displaying a good activity (IC50=210nM) close to the value obtained with chloroquine (IC50=514nM) and a weak cytotoxicity. Copyright © 2016 Elsevier Ltd. All rights reserved.

Pharmacological Evaluation of Naproxen Metal Complexes on Antinociceptive, Anxiolytic, CNS Depressant, and Hypoglycemic Properties

PubMed Central

Das, Narhari; Abdur Rahman, S. M.

2016-01-01

Purpose. The present study was designed to investigate the antinociceptive, anxiolytic, CNS depressant, and hypoglycemic effects of the naproxen metal complexes. Methods. The antinociceptive activity was evaluated by acetic acid-induced writhing method and radiant heat tail-flick method while anxiolytic activity was evaluated by elevated plus maze model. The CNS depressant activity of naproxen metal complexes was assessed using phenobarbitone-induced sleeping time test and the hypoglycemic test was performed using oral glucose tolerance test. Results. Metal complexes significantly (P < 0.001) reduced the number of abdominal muscle contractions induced by 0.7% acetic acid solution in a dose dependent manner. At the dose of 25 mg/kg body weight p.o. copper, cobalt, and zinc complexes exhibited higher antinociceptive activity having 59.15%, 60.56%, and 57.75% of writhing inhibition, respectively, than the parent ligand naproxen (54.93%). In tail-flick test, at both doses of 25 and 50 mg/kg, the copper, cobalt, silver, and zinc complexes showed higher antinociceptive activity after 90 minutes than the parent drug naproxen. In elevated plus maze (EPM) model the cobalt and zinc complexes of naproxen showed significant anxiolytic effects in dose dependent manner, while the copper, cobalt, and zinc complexes showed significant CNS depressant and hypoglycemic activity. Conclusion. The present study demonstrated that copper, cobalt, and zinc complexes possess higher antinociceptive, anxiolytic, CNS depressant, and hypoglycemic properties than the parent ligand. PMID:27478435

Synthesis, spectral characterization and catalytic activity of Co(II) complexes of drugs: crystal structure of Co(II)-trimethoprim complex.

PubMed

Madhupriya, Selvaraj; Elango, Kuppanagounder P

2014-01-24

New Co(II) complexes with drugs such as trimethoprim (TMP), cimetidine (CTD), niacinamide (NAM) and ofloxacin (OFL) as ligands were synthesized. The complexes were characterized by analytical analysis, various spectral techniques such as FT-IR, UV-Vis, magnetic measurements and molar conductivity. The magnetic susceptibility results coupled with the electronic spectra suggested a tetrahedral geometry for the complexes. The coordination mode of trimethoprim ligand and geometry of the complex were confirmed by single crystal X-ray studies. In this complex the metal ion possesses a tetrahedral geometry with two nitrogen atom from two TMP ligands and two chloride ions coordinated to it. The catalytic activity of the complexes in aryl-aryl coupling reaction was screened and the results indicated that among the four complexes [Co(OFL)Cl(H2O)] exhibited excellent catalytic activity. Copyright © 2013 Elsevier B.V. All rights reserved.

Toxin activity assays, devices, methods and systems therefor

DOEpatents

Koh, Chung-Yan; Schaff, Ulrich Y.; Sommer, Gregory Jon

2016-04-05

Embodiments of the present invention are directed toward devices, system and method for conducting toxin activity assay using sedimentation. The toxin activity assay may include generating complexes which bind to a plurality of beads in a fluid sample. The complexes may include a target toxin and a labeling agent, or may be generated due to presence of active target toxin and/or labeling agent designed to be incorporated into complexes responsive to the presence of target active toxin. The plurality of beads including the complexes may be transported through a density media, wherein the density media has a lower density than a density of the beads and higher than a density of the fluid sample, and wherein the transporting occurs, at least in part, by sedimentation. Signal may be detected from the labeling agents of the complexes.

Synthesis, characterization and biological activities of semicarbazones and their copper complexes.

PubMed

Venkatachalam, Taracad K; Bernhardt, Paul V; Noble, Chris J; Fletcher, Nicholas; Pierens, Gregory K; Thurecht, Kris J; Reutens, David C

2016-09-01

Substituted semicarbazones/thiosemicarbazones and their copper complexes have been prepared and several single crystal structures examined. The copper complexes of these semicarbazone/thiosemicarbazones were prepared and several crystal structures examined. The single crystal X-ray structure of the pyridyl-substituted semicarbazone showed two types of copper complexes, a monomer and a dimer. We also found that the p-nitrophenyl semicarbazone formed a conventional 'magic lantern' acetate-bridged dimer. Electron Paramagnetic Resonance (EPR) of several of the copper complexes was consistent with the results of single crystal X-ray crystallography. The EPR spectra of the p-nitrophenyl semicarbazone copper complex in dimethylsulfoxide (DMSO) showed the presence of two species, confirming the structural information. Since thiosemicarbazones and semicarbazones have been reported to exhibit anticancer activity, we examined the anticancer activity of several of the derivatives reported in the present study and interestingly only the thiosemicarbazone showed activity while the semicarbazones were not active indicating that introduction of sulphur atom alters the biological profile of these thiosemicarbazones. Copyright © 2016 Elsevier Inc. All rights reserved.

Generalization of the Activated Complex Theory of Reaction Rates. I. Quantum Mechanical Treatment

DOE R&D Accomplishments Database

Marcus, R. A.

1964-01-01

In its usual form activated complex theory assumes a quasi-equilibrium between reactants and activated complex, a separable reaction coordinate, a Cartesian reaction coordinate, and an absence of interaction of rotation with internal motion in the complex. In the present paper a rate expression is derived without introducing the Cartesian assumption. The expression bears a formal resemblance to the usual one and reduces to it when the added assumptions of the latter are introduced.

Competition between thyroid hormone receptor-associated protein (TRAP) 220 and transcriptional intermediary factor (TIF) 2 for binding to nuclear receptors. Implications for the recruitment of TRAP and p160 coactivator complexes.

PubMed

Treuter, E; Johansson, L; Thomsen, J S; Wärnmark, A; Leers, J; Pelto-Huikko, M; Sjöberg, M; Wright, A P; Spyrou, G; Gustafsson, J A

1999-03-05

Transcriptional activation by nuclear receptors (NRs) involves the concerted action of coactivators, chromatin components, and the basal transcription machinery. Crucial NR coactivators, which target primarily the conserved ligand-regulated activation (AF-2) domain, include p160 family members, such as TIF2, as well as p160-associated coactivators, such as CBP/p300. Because these coactivators possess intrinsic histone acetyltransferase activity, they are believed to function mainly by regulating chromatin-dependent transcriptional activation. Recent evidence suggests the existence of an additional NR coactivator complex, referred to as the thyroid hormone receptor-associated protein (TRAP) complex, which may function more directly as a bridging complex to the basal transcription machinery. TRAP220, the 220-kDa NR-binding subunit of the complex, has been identified in independent studies using both biochemical and genetic approaches. In light of the functional differences identified between p160 and TRAP coactivator complexes in NR activation, we have attempted to compare interaction and functional characteristics of TIF 2 and TRAP220. Our findings imply that competition between the NR-binding subunits of distinct coactivator complexes may act as a putative regulatory step in establishing either a sequential activation cascade or the formation of independent coactivator complexes.

Antibacterial characteristics of newly developed amphiphilic lipids and DNA-lipid complexes against bacteria.

PubMed

Inoue, Y; Fukushima, T; Hayakawa, T; Takeuchi, H; Kaminishi, H; Miyazaki, K; Okahata, Y

2003-05-01

The purpose of this study was to investigate the antibacterial activity of newly developed amphiphilic lipids and DNA/lipid complexes against two types of oral bacteria and two types of hospital infection bacteria. Nine amphiphilic lipids were quantitatively prepared from the reaction of n-alkyl alcohol, alpha-amino acids, and p-toluenesulfonic acid. Nine DNA-lipid complexes were prepared by the simple mixing of DNA and amphiphilic lipids. The DNA-lipid complexes were insoluble in water. The antibacterial activity of lipids and DNA-lipid complexes against Porphyromonas gingivalis, Streptococcus mutans, Staphylococcus aureus, and Pseudomonas aeruginosa were evaluated by the disk-diffusion method. Seven artificial lipids showed antibacterial behavior; in particular, the lipids prepared from n-decyl alcohol and glycine and from n-decyl alcohol and L-alanine showed antibacterial activity against the four bacterial strains used in this study. On the other hand, the lipids of glutamic acid derivatives did not show any antibacterial activity against the four bacteria strains except for the lipid with an n-octyl group. Five DNA-lipid complexes also had an antibacterial effect. The complex prepared from DNA and glycine decyl ester p-toluenesulfonic acid salt exhibited antibacterial activity against the four types of bacteria strains. In this study it was found that lipids and DNA-lipid complexes with a mono-decyl group or a mono-dodecyl group have more favorable antibacterial activity. Copyright 2003 Wiley Periodicals, Inc.

Signals of monocyte activation in patients with SLE.

PubMed Central

Kávai, M; Zsindely, A; Sonkoly, I; Major, M; Demján, I; Szegedi, G

1983-01-01

The Fc receptor mediated reaction, the beta-glucuronidase and the lactic dehydrogenase activities of monocytes and the serum lysozyme level were tested together with the circulating immune complex content of patients with systemic lupus erythematosus. Simultaneously with the increasing FC receptor-mediated reaction and the elevated enzyme activities of patient monocytes, the secretion of lysozyme and the immune complex content of the sera were higher than those of the controls. A positive correlation was demonstrated between the Fc receptor-mediated reaction, the beta-glucuronidase activity, the lysozyme secretion and the immune complex content of the sera. Thus, the monocytes of patients appeared to be activated by the circulating immune complexes. PMID:6839541

Can spectro-temporal complexity explain the autistic pattern of performance on auditory tasks?

PubMed

Samson, Fabienne; Mottron, Laurent; Jemel, Boutheina; Belin, Pascal; Ciocca, Valter

2006-01-01

To test the hypothesis that level of neural complexity explain the relative level of performance and brain activity in autistic individuals, available behavioural, ERP and imaging findings related to the perception of increasingly complex auditory material under various processing tasks in autism were reviewed. Tasks involving simple material (pure tones) and/or low-level operations (detection, labelling, chord disembedding, detection of pitch changes) show a superior level of performance and shorter ERP latencies. In contrast, tasks involving spectrally- and temporally-dynamic material and/or complex operations (evaluation, attention) are poorly performed by autistics, or generate inferior ERP activity or brain activation. Neural complexity required to perform auditory tasks may therefore explain pattern of performance and activation of autistic individuals during auditory tasks.

Plasticity of TOM complex assembly in skeletal muscle mitochondria in response to chronic contractile activity.

PubMed

Joseph, Anna-Maria; Hood, David A

2012-03-01

We investigated the assembly of the TOM complex within skeletal muscle under conditions of chronic contractile activity-induced mitochondrial biogenesis. Tom40 import into mitochondria was increased by chronic contractile activity, as was its time-dependent assembly into the TOM complex. These changes coincided with contractile activity-induced augmentations in the expression of key protein import machinery components Tim17, Tim23, and Tom22, as well as the cytosolic chaperone Hsp90. These data indicate the adaptability of the TOM protein import complex and suggest a regulatory role for the assembly of this complex in exercise-induced mitochondrial biogenesis. Copyright © 2011 Elsevier B.V. and Mitochondria Research Society. All rights reserved. All rights reserved.

Nickel(II) and palladium(II) triphenylphosphine complexes incorporating tridentate Schiff base ligands: Synthesis, characterization and biocidal activities

NASA Astrophysics Data System (ADS)

Shabbir, Muhammad; Akhter, Zareen; Ashraf, Ahmad Raza; Ismail, Hammad; Habib, Anum; Mirza, Bushra

2017-12-01

Nickel(II) and palladium(II) triphenylphosphine complexes incorporating tridentate Schiff bases have been prepared and characterized by elemental analysis as well as by spectroscopic techniques (FTIR & NMR). The synthesized compounds were assessed to check their potential biocidal activity by using different biological assays (brine shrimp cytotoxicity, antimicrobial, antioxidant, antitumor and drug-DNA interaction). Results of brine shrimp cytotoxicity assay showed that ligand molecules are more bioactive than metal complexes with LD50 as low as 12.4 μg/mL. The prominent antitumor activity was shown by nickel complexes while the palladium complexes exhibited moderate activity. The synthesized compounds have shown high propensity for DNA binding either through intercalation or groove binding which represents the mechanism of antitumor effect of these compounds. Additionally, ligand molecules and nickel metal complexes showed significant antioxidant activity with IC50 values as low as 3.1 μg/mL and 18.9 μg/mL respectively while palladium complexes exhibited moderate activity. Moreover, in antimicrobial assays H2L1, Ni(L1)PPh3 and H2L3 showed dual inhibition against bacterial and fungal strains while for the rest of the compounds varying degree of activity was recorded against different strains. Overall comparison of results suggests that the synthesized compounds can be promising candidate for drug formulation and development.

Activation of the DnaK-ClpB Complex is Regulated by the Properties of the Bound Substrate.

PubMed

Fernández-Higuero, Jose Angel; Aguado, Alejandra; Perales-Calvo, Judit; Moro, Fernando; Muga, Arturo

2018-04-11

The chaperone ClpB in bacteria is responsible for the reactivation of aggregated proteins in collaboration with the DnaK system. Association of these chaperones at the aggregate surface stimulates ATP hydrolysis, which mediates substrate remodeling. However, a question that remains unanswered is whether the bichaperone complex can be selectively activated by substrates that require remodeling. We find that large aggregates or bulky, native-like substrates activates the complex, whereas a smaller, permanently unfolded protein or extended, short peptides fail to stimulate it. Our data also indicate that ClpB interacts differently with DnaK in the presence of aggregates or small peptides, displaying a higher affinity for aggregate-bound DnaK, and that DnaK-ClpB collaboration requires the coupled ATPase-dependent remodeling activities of both chaperones. Complex stimulation is mediated by residues at the β subdomain of DnaK substrate binding domain, which become accessible to the disaggregase when the lid is allosterically detached from the β subdomain. Complex activation also requires an active NBD2 and the integrity of the M domain-ring of ClpB. Disruption of the M-domain ring allows the unproductive stimulation of the DnaK-ClpB complex in solution. The ability of the DnaK-ClpB complex to discrimínate different substrate proteins might allow its activation when client proteins require remodeling.

Suppression of BRCA2 by Mutant Mitochondrial DNA in Prostate Cancer

DTIC Science & Technology

2011-05-01

Briefly, the electron transfer activities of complex I/III (NADH dehydrogenase/cytochrome bc1 complex: catalyzes the electron transfer from NADH to...ferricytochrome c) and complex II/III (succinate dehydrogenase/cytochrome bc1 complex: catalyzes the electron transfer from succinate to ferricytochrome...The electron transfer activity of complex IV (cytochrome c oxidase: catalyzes the final step of the respiratory chain by transferring electrons from

No evidence of a role for mitochondrial complex I in Helicobacter pylori pathogenesis.

PubMed

Ng, Garrett Z; Ke, Bi-Xia; Laskowski, Adrienne; Thorburn, David R; Sutton, Philip

2017-06-01

Complex I is the first enzyme complex in the mitochondrial respiratory chain, responsible for generating a large fraction of energy during oxidative phosphorylation. Recently, it has been identified that complex I deficiency can result in increased inflammation due to the generation of reactive oxygen species by innate immune cells. As a reduction in complex I activity has been demonstrated in human stomachs with atrophic gastritis, we investigated whether complex I deficiency could influence Helicobacter pylori pathogenesis. Ndufs6 gt/gt mice have a partial complex I deficiency. Complex I activity was quantified in the stomachs and immune cells of Ndufs6 gt/gt mice by spectrophotometric assays. Ndufs6 gt/gt mice were infected with H. pylori and bacterial colonization assessed by colony-forming assay, gastritis assessed histologically, and H. pylori -specific humoral response quantified by ELISA. The immune cells and stomachs of Ndufs6 gt/gt mice were found to have significantly decreased complex I activity, validating the model for assessing the effects of complex I deficiency in H. pylori infection. However, there was no observable effect of complex I deficiency on either H. pylori colonization, the resulting gastritis, or the humoral response. Although complex I activity is described to suppress innate immune responses and is decreased during atrophic gastritis in humans, our data suggest it does not affect H. pylori pathogenesis. © 2017 John Wiley & Sons Ltd.

Myoglobin and the regulation of mitochondrial respiratory chain complex IV.

PubMed

Yamada, Tatsuya; Takakura, Hisashi; Jue, Thomas; Hashimoto, Takeshi; Ishizawa, Rie; Furuichi, Yasuro; Kato, Yukio; Iwanaka, Nobumasa; Masuda, Kazumi

2016-01-15

Mitochondrial respiration is regulated by multiple elaborate mechanisms. It has been shown that muscle specific O2 binding protein, Myoglobin (Mb), is localized in mitochondria and interacts with respiratory chain complex IV, suggesting that Mb could be a factor that regulates mitochondrial respiration. Here, we demonstrate that muscle mitochondrial respiration is improved by Mb overexpression via up-regulation of complex IV activity in cultured myoblasts; in contrast, suppression of Mb expression induces a decrease in complex IV activity and mitochondrial respiration compared with the overexpression model. The present data are the first to show the biological significance of mitochondrial Mb as a potential modulator of mitochondrial respiratory capacity. Mitochondria are important organelles for metabolism, and their respiratory capacity is a primary factor in the regulation of energy expenditure. Deficiencies of cytochrome c oxidase complex IV, which reduces O2 in mitochondria, are linked to several diseases, such as mitochondrial myopathy. Moreover, mitochondrial respiration in skeletal muscle tissue tends to be susceptible to complex IV activity. Recently, we showed that the muscle-specific protein myoglobin (Mb) interacts with complex IV. The precise roles of mitochondrial Mb remain unclear. Here, we demonstrate that Mb facilitates mitochondrial respiratory capacity in skeletal muscles. Although mitochondrial DNA copy numbers were not altered in Mb-overexpressing myotubes, O2 consumption was greater in these myotubes than that in mock cells (Mock vs. Mb-Flag::GFP: state 4, 1.00 ± 0.09 vs. 1.77 ± 0.34; state 3, 1.00 ± 0.29; Mock: 1.60 ± 0.53; complex 2-3-4: 1.00 ± 0.30 vs. 1.50 ± 0.44; complex IV: 1.00 ± 0.14 vs. 1.87 ± 0.27). This improvement in respiratory capacity could be because of the activation of enzymatic activity of respiratory complexes. Moreover, mitochondrial respiration was up-regulated in myoblasts transiently overexpressing Mb; complex IV activity was solely activated in Mb-overexpressing myoblasts, and complex IV activity was decreased in the myoblasts in which Mb expression was suppressed by Mb-siRNA transfection (Mb vector transfected vs. Mb vector, control siRNA transfected vs. Mb vector, Mb siRNA transfected: 0.15 vs. 0.15 vs. 0.06). Therefore, Mb enhances the enzymatic activity of complex IV to ameliorate mitochondrial respiratory capacity, and could play a pivotal role in skeletal muscle metabolism. © 2015 The Authors. The Journal of Physiology © 2015 The Physiological Society.

Effect of nitroso complexes of some transition metals on the activity of soluble guanylate cyclase.

PubMed

Severina, I S; Bussygina, O G; Grigorjev, N B

1992-03-01

Effects of nitroso complexes of some transition metals (Fe, Co, Cr), differing in the character of NO oxidation on the activity of human and rat platelet guanylate cyclase were studied. 3 types of nitroso complexes were used: (1) NO group carries a positive charge--a nitrosonium cation (Na2[FeNO + (CN)5]-nitroprusside); (2) NO is neutral--(K3[CrNO(CN)5 and [CoNO(NH3)5]SO4) and (3) NO is coordinated as anion NO- (K3[CoNO-(CN)5]. It is shown that the highest stimulatory effect is produced by sodium nitroprusside, whose activating action is due to the interaction of its NO group with the guanylate cyclase heme. Nitroso complexes (Co and Cr) the NO group of which is neutral stimulated guanylate cyclase activity insignificantly and this activation was not guanylate cyclase heme directed. Nitroso complex (Co) with NO coordinated as anion NO(-)--is a guanylate cyclase inhibitor. In contrast to nitroprusside, the nitroso complexes used (Co and Cr) have no hypotensive effect. It was concluded that the essential requirement for the realization of the hypotensive effect of transition metals' nitroso complexes is the ability of these compounds to activate soluble guanylate cyclase solely by the heme-dependent mechanism.

Synthesis, spectroscopic characterization and biological activities of N4O2 Schiff base ligand and its metal complexes of Co(II), Ni(II), Cu(II) and Zn(II)

NASA Astrophysics Data System (ADS)

Al-Resayes, Saud I.; Shakir, Mohammad; Abbasi, Ambreen; Amin, Kr. Mohammad Yusuf; Lateef, Abdul

The Schiff base ligand, bis(indoline-2-one)triethylenetetramine (L) obtained from condensation of triethylenetetramine and isatin was used to synthesize the complexes of type, [ML]Cl2 [M = Co(II), Ni(II), Cu(II) and Zn(II)]. L was characterized on the basis of the results of elemental analysis, FT-IR, 1H and 13C NMR, mass spectroscopic studies. The stoichiometry, bonding and stereochemistries of complexes were ascertained on the basis of results of elemental analysis, magnetic susceptibility values, molar conductance and various spectroscopic studies. EPR, UV-vis and magnetic moments revealed an octahedral geometry for complexes. L and its Cu(II) and Zn(II) complexes were screened for their antibacterial activity. Analgesic activity of Cu(II) and Zn(II) complexes was also tested in rats by tail flick method. Both complexes were found to possess good antibacterial and moderate analgesic activity.

Ru(II) complexes of N 4 and N 2O 2 macrocyclic Schiff base ligands: Their antibacterial and antifungal studies

NASA Astrophysics Data System (ADS)

Shanker, Kanne; Rohini, Rondla; Ravinder, Vadde; Reddy, P. Muralidhar; Ho, Yen-Peng

2009-07-01

Reactions of [RuCl 2(DMSO) 4] with some of the biologically active macrocyclic Schiff base ligands containing N 4 and N 2O 2 donor group yielded a number of stable complexes, effecting complete displacement of DMSO groups from the complex. The interaction of tetradentate ligand with [RuCl 2(DMSO) 4] gave neutral complexes of the type [RuCl 2(L)] [where L = tetradentate macrocyclic ligand]. These complexes were characterized by elemental, IR, 1H, 13C NMR, mass, electronic, thermal, molar conductance and magnetic susceptibility measurements. An octahedral geometry has been proposed for all complexes. All the macrocycles and macrocyclic Ru(II) complexes along with existing antibacterial drugs were screened for antibacterial activity against Gram +ve ( Bacillus subtilis, Staphylococcus aureus) and Gram -ve ( Escherichia coli, Klebsiella pneumonia) bacteria. All these compounds were found to be more active when compared to streptomycin and ampicillin. The representative macrocyclic Schiff bases and their complexes were also tested in vitro to evaluate their activity against fungi, namely, Aspergillus flavus and Fusarium species.

Active remodelling of the TIM23 complex during translocation of preproteins into mitochondria.

PubMed

Popov-Celeketić, Dusan; Mapa, Koyeli; Neupert, Walter; Mokranjac, Dejana

2008-05-21

The TIM23 (translocase of the mitochondrial inner membrane) complex mediates translocation of preproteins across and their insertion into the mitochondrial inner membrane. How the translocase mediates sorting of preproteins into the two different subcompartments is poorly understood. In particular, it is not clear whether association of two operationally defined parts of the translocase, the membrane-integrated part and the import motor, depends on the activity state of the translocase. We established conditions to in vivo trap the TIM23 complex in different translocation modes. Membrane-integrated part of the complex and import motor were always found in one complex irrespective of whether an arrested preprotein was present or not. Instead, we detected different conformations of the complex in response to the presence and, importantly, the type of preprotein being translocated. Two non-essential subunits of the complex, Tim21 and Pam17, modulate its activity in an antagonistic manner. Our data demonstrate that the TIM23 complex acts as a single structural and functional entity that is actively remodelled to sort preproteins into different mitochondrial subcompartments.

Active remodelling of the TIM23 complex during translocation of preproteins into mitochondria

PubMed Central

Popov-Čeleketić, Dus̆an; Mapa, Koyeli; Neupert, Walter; Mokranjac, Dejana

2008-01-01

The TIM23 (translocase of the mitochondrial inner membrane) complex mediates translocation of preproteins across and their insertion into the mitochondrial inner membrane. How the translocase mediates sorting of preproteins into the two different subcompartments is poorly understood. In particular, it is not clear whether association of two operationally defined parts of the translocase, the membrane-integrated part and the import motor, depends on the activity state of the translocase. We established conditions to in vivo trap the TIM23 complex in different translocation modes. Membrane-integrated part of the complex and import motor were always found in one complex irrespective of whether an arrested preprotein was present or not. Instead, we detected different conformations of the complex in response to the presence and, importantly, the type of preprotein being translocated. Two non-essential subunits of the complex, Tim21 and Pam17, modulate its activity in an antagonistic manner. Our data demonstrate that the TIM23 complex acts as a single structural and functional entity that is actively remodelled to sort preproteins into different mitochondrial subcompartments. PMID:18418384

Structural characterization and antioxidant properties of Cu(II) and Ni(II) complexes derived from dicyandiamide

NASA Astrophysics Data System (ADS)

Kertmen, Seda Nur; Gonul, Ilyas; Kose, Muhammet

2018-01-01

New Cu(II) and Ni(II) complexes derived from dicyandiamide were synthesized and characterised by spectroscopic and analytical methods. Molecular structures of the complexes were determined by single crystal X-ray diffraction studies. In the complexes, the Cu(II) or Ni(II) ions are four-coordinate with a slight distorted square planar geometry. The ligands (L-nPen and L-iPen) derived from dicyandiamide formed via nucleophilic addition of alcohol solvent molecule in the presence Cu(II) or Ni(II) ions. Complexes were stabilised by intricate array of hydrogen bonding interactions. Antioxidant activity of the complexes was evaluated by DPPH radical scavenging and CUPRAC methods. The complexes exhibit antioxidant activity, however, their activities were much lower than standard antioxidants (Vitamin C and trolox).

The Nun protein of bacteriophage HK022 inhibits translocation of Escherichia coli RNA polymerase without abolishing its catalytic activities

PubMed Central

Hung, Siu Chun; Gottesman, Max E.

1997-01-01

Bacteriophage HK022 Nun protein blocks transcription elongation by Escherichia coli RNA polymerase in vitro without dissociating the transcription complex. Nun is active on complexes located at any template site tested. Ultimately, only the 3′-OH terminal nucleotide of the nascent transcript in an arrested complex can turn over; it is removed by pyrophosphate and restored with NTPs. This suggests that Nun inhibits the translocation of RNA polymerase without abolishing its catalytic activities. Unlike spontaneously arrested complexes, Nun-arrested complexes cannot be reactivated by transcription factor GreB. The various complexes show distinct patterns of nucleotide incorporation and pyrophosphorolysis before or after treatment with Nun, suggesting that the configuration of RNAP, transcript, and template DNA is different in each complex. PMID:9334329

Human T-Cell Leukemia Virus Type 1 (HTLV-1) Tax Requires CADM1/TSLC1 for Inactivation of the NF-κB Inhibitor A20 and Constitutive NF-κB Signaling

PubMed Central

Thomas, Remy; van der Weyden, Louise; Rauch, Dan; Ratner, Lee; Nyborg, Jennifer K.; Ramos, Juan Carlos; Takai, Yoshimi; Shembade, Noula

2015-01-01

Persistent activation of NF-κB by the Human T-cell leukemia virus type 1 (HTLV-1) oncoprotein, Tax, is vital for the development and pathogenesis of adult T-cell leukemia (ATL) and HTLV-1-associated myelopathy/tropical spastic paraparesis (HAM/TSP). K63-linked polyubiquitinated Tax activates the IKK complex in the plasma membrane-associated lipid raft microdomain. Tax also interacts with TAX1BP1 to inactivate the NF-κB negative regulatory ubiquitin-editing A20 enzyme complex. However, the molecular mechanisms of Tax-mediated IKK activation and A20 protein complex inactivation are poorly understood. Here, we demonstrated that membrane associated CADM1 (Cell adhesion molecule1) recruits Ubc13 to Tax, causing K63-linked polyubiquitination of Tax, and IKK complex activation in the membrane lipid raft. The c-terminal cytoplasmic tail containing PDZ binding motif of CADM1 is critical for Tax to maintain persistent NF-κB activation. Finally, Tax failed to inactivate the NF-κB negative regulator ubiquitin-editing enzyme A20 complex, and activate the IKK complex in the lipid raft in absence of CADM1. Our results thus indicate that CADM1 functions as a critical scaffold molecule for Tax and Ubc13 to form a cellular complex with NEMO, TAX1BP1 and NRP, to activate the IKK complex in the plasma membrane-associated lipid rafts, to inactivate NF-κB negative regulators, and maintain persistent NF-κB activation in HTLV-1 infected cells. PMID:25774694

Activity of Pure Streptovaricins and Fractionated Streptovaricin Complex Against Friend Virus

PubMed Central

Horoszewicz, Julius S.; Rinehart, Kenneth L.; Leong, Susan S.; Carter, William A.

1975-01-01

Chromatographic fractionation of streptovaricin complex yields two stable components enriched (4- to 16-fold) in activity directed against the polycythemic strain of Friend virus; both components apparently contain no streptovaricins. When compared with their unfractionated parent streptovaricin complex, eight individual intact streptovaricins (A through G and J) show at least a 30-fold reduction in antiviral activity. These results further support the conclusion that the diversified biological properties of streptovaricin complex probably reside in different molecular structures. PMID:237470

Modulation of Mitochondrial Complex I Activity Averts Cognitive Decline in Multiple Animal Models of Familial Alzheimer's Disease

PubMed Central

Zhang, Liang; Zhang, Song; Maezawa, Izumi; Trushin, Sergey; Minhas, Paras; Pinto, Matthew; Jin, Lee-Way; Prasain, Keshar; Nguyen, Thi D.T.; Yamazaki, Yu; Kanekiyo, Takahisa; Bu, Guojun; Gateno, Benjamin; Chang, Kyeong-Ok; Nath, Karl A.; Nemutlu, Emirhan; Dzeja, Petras; Pang, Yuan-Ping; Hua, Duy H.; Trushina, Eugenia

2015-01-01

Development of therapeutic strategies to prevent Alzheimer's disease (AD) is of great importance. We show that mild inhibition of mitochondrial complex I with small molecule CP2 reduces levels of amyloid beta and phospho-Tau and averts cognitive decline in three animal models of familial AD. Low-mass molecular dynamics simulations and biochemical studies confirmed that CP2 competes with flavin mononucleotide for binding to the redox center of complex I leading to elevated AMP/ATP ratio and activation of AMP-activated protein kinase in neurons and mouse brain without inducing oxidative damage or inflammation. Furthermore, modulation of complex I activity augmented mitochondrial bioenergetics increasing coupling efficiency of respiratory chain and neuronal resistance to stress. Concomitant reduction of glycogen synthase kinase 3β activity and restoration of axonal trafficking resulted in elevated levels of neurotrophic factors and synaptic proteins in adult AD mice. Our results suggest that metabolic reprogramming induced by modulation of mitochondrial complex I activity represents promising therapeutic strategy for AD. PMID:26086035

GINS complex protein Sld5 recruits SIK1 to activate MCM helicase during DNA replication.

PubMed

Joshi, Kiranmai; Shah, Varun Jayeshkumar; Maddika, Subbareddy

2016-12-01

In eukaryotes, proper loading and activation of MCM helicase at chromosomal origins plays a central role in DNA replication. Activation of MCM helicase requires its association with CDC45-GINS complex, but the mechanism of how this complex activates MCM helicase is poorly understood. Here we identified SIK1 (salt-inducible kinase 1), an AMPK related protein kinase, as a molecular link that connects GINS complex with MCM helicase activity. We demonstrated that Sld5 a component of GINS complex interacts with SIK1 and recruits it to the sites of DNA replication at the onset of S phase. Depletion of SIK1 leads to defective DNA replication. Further, we showed that SIK1 phosphorylates MCM2 at five conserved residues at its N-terminus, which is essential for the activation of MCM helicase. Collectively, our results suggest SIK1 as a novel integral component of CMG replicative helicase during eukaryotic DNA replication. Copyright © 2016 Elsevier Inc. All rights reserved.

Substitutional impact on biological activity of new water soluble Ni(II) complexes: Preparation, spectral characterization, X-ray crystallography, DNA/protein binding, antibacterial activity and in vitro cytotoxicity.

PubMed

Umadevi, C; Kalaivani, P; Puschmann, H; Murugan, S; Mohan, P S; Prabhakaran, R

2017-02-01

A series of new water soluble nickel(II) complexes containing triphenylphosphine and 4-methoxysalicylaldehyde-4(N)-substituted thiosemicarbazones were synthesized and characterized. Crystallographic investigations confirmed the structure of the complexes (1-4) having the general structure [Ni(4-Msal-Rtsc)(PPh 3 )] (Where R=H (1); CH 3 (2); C 2 H 5 (3); C 6 H 5 (4)) which showed that thiosemicarbazone ligands coordinated to nickel(II) ion as ONS tridentate bibasic donor. DNA/BSA protein binding ability of the ligands and their new complexes were studied by taking calf-thymus DNA (CT-DNA) and Bovine serum albumin (BSA) through absorption and emission titrations. Ethidium bromide (EB) displacement study showed the intercalative binding trend of the complexes to DNA. From the albumin binding studies, the mechanism of quenching was found as static and the alterations in the secondary structure of BSA by the compounds were confirmed with synchronous spectral studies. The binding affinity of the complexes to CT-DNA and BSA has the order of [Ni(4-Msal-etsc)(PPh 3 )] (3) >[Ni(4-Msal-mtsc)(PPh 3 )] (2) >[Ni(4-Msal-tsc)(PPh 3 )] (1) >[Ni(4-Msal-ptsc)(PPh 3 )] (4). In vitro cytotoxicity of the complexes was tested on human lung cancer cells (A549), human cervical cancer cells (HeLa), human liver carcinoma cells (Hep G2). All the complexes exhibited significant activity against three cancer cells. Among them, complex 4 exhibited almost 2.5 fold activity than cisplatin in A549 and HepG2 cell lines. In HeLa cell line, the complexes exhibited significant activity which is less than cisplatin. While comparing the activity of the complexes in A549 and HepG2 cell lines it falls in the order 4>1>2>3>cisplatin. The results obtained from DNA, protein binding and cytotoxicity studies, it is concluded that the cytotoxicity of the complexes as determined by MTT assay were not unduly influenced by the complexes having different binding efficiency with DNA and protein. The complexes exhibited good spectrum of antibacterial activity against four pathogenic bacteria such as E. faecalis (gram +ve), S. aureus (gram +ve), E. coli (gram -ve) and P. aeruginosa (gram -ve). Copyright © 2016 Elsevier B.V. All rights reserved.

Activated Prothrombin Complex Concentrate Versus 4-Factor Prothrombin Complex Concentrate for Vitamin K-Antagonist Reversal.

PubMed

Rowe, A Shaun; Dietrich, Scott K; Phillips, John W; Foster, Kaci E; Canter, Joshua R

2018-06-01

To compare the international normalized ratio normalization efficacy of activated prothrombin complex concentrates and 4-factor prothrombin complex concentrates and to evaluate the thrombotic complications in patients treated with these products for warfarin-associated hemorrhage. Retrospective, Multicenter Cohort. Large, Community, Teaching Hospital. Patients greater than 18 years old and received either activated prothrombin complex concentrate or 4-factor prothrombin complex concentrate for the treatment of warfarin-associated hemorrhage. We excluded those patients who received either agent for an indication other than warfarin-associated hemorrhage, pregnant, had a baseline international normalized ratio of less than 2, received a massive transfusion as defined by hospital protocol, received plasma for treatment of warfarin-associated hemorrhage, or were treated for an acute warfarin ingestion. Patients in the activated prothrombin complex concentrate group (enrolled from one hospital) with an international normalized ratio of less than 5 received 500 IU and those with an international normalized ratio greater than 5 received 1,000 IU. Patients in the 4-factor prothrombin complex concentrate (enrolled from a separate hospital) group received the Food and Drug Administration approved dosing algorithm. A total of 158 patients were included in the final analysis (activated prothrombin complex concentrate = 118; 4-factor prothrombin complex concentrate = 40). Those in the 4-factor prothrombin complex concentrate group had a higher pretreatment international normalized ratio (2.7 ± 1.8 vs 3.5 ± 2.9; p = 0.0164). However, the posttreatment international normalized ratio was similar between the groups. In addition, even when controlling for differences in the pretreatment international normalized ratio, there was no difference in the ability to achieve a posttreatment international normalized ratio of less than 1.4 (odds ratio, 0.753 [95% CI, 0.637-0.890]; p = 0.0009). Those in the activated prothrombin complex concentrate group did have higher odds of achieving a posttreatment international normalized ratio of less than 1.2 (odds ratio, 3.23 [95% CI, 1.34-7.81]; p = 0.0088). There was only one posttreatment thrombotic complication reported. A low, fixed dose of activated prothrombin complex concentrate was as effective as standard dose 4-factor prothrombin complex concentrate for normalization of international normalized ratio. In addition, we did not see an increase in thrombotic events.

Biotinylated platinum(IV) complexes designed to target cancer cells.

PubMed

Zhao, Jian; Hua, Wuyang; Xu, Gang; Gou, Shaohua

2017-11-01

Three biotinylated platinum(IV) complexes (1-3) were designed and synthesized. The resulting platinum(IV) complexes exhibited effective cytotoxicity against the tested cancer cell lines, especially complex 1, which was 2.0-9.6-fold more potent than cisplatin. These complexes were found to be rapidly reduced to their activated platinum(II) counterparts by glutathione or ascorbic acid under biologically relevant condition. Additional molecular docking studies revealed that the biotin moieties of all Pt(IV) complexes can effectively bind with the streptavidin through the noncovalent interactions. Besides, introduction of the biotin group can obviously promote the cancer cell uptake of platinum when treated with complex 1, particularly in cisplatin-resistant SGC-7901/Cis cancer cells. Further mechanistic studies on complex 1 indicated that it activated the expression of Bax, and induced cytochrome c release from the mitochondria, and finally activated caspase-3. Copyright © 2017 Elsevier Inc. All rights reserved.

Symmetrical and unsymmetrical pincer complexes with group 10 metals: synthesis via aryl C-H activation and some catalytic applications.

PubMed

Niu, Jun-Long; Hao, Xin-Qi; Gong, Jun-Fang; Song, Mao-Ping

2011-05-21

Aryl-based pincer metal complexes with anionic terdentate ligands have been widely applied in organic synthesis, organometallic catalysis and other related areas. Synthetically, the most simple and convenient method for the construction of these complexes is the direct metal-induced C(aryl)-H bond activation, which can be fulfilled by choosing the appropriate functional donor groups in the two side arms of the aryl-based pincer preligands. In this perspective, we wish to summarize some results achieved by our group in this context. Successful examples include symmetrical chiral bis(imidazoline) NCN pincer complexes with Ni(II), Pd(II) and Pt(II), bis(phosphinite) and bis(phosphoramidite) PCP pincer Pd(II) complexes, unsymmetrical (pyrazolyl)phosphinite, (amino)phosphinite and (imino)phosphinite PCN pincer Pd(II) complexes, chiral (imidazolinyl)phosphinite and (imidazolinyl)phosphoramidite PCN pincer complexes with Ni(II) and Pd(II) as well as unsymmetrical (oxazolinyl)amine and (oxazolinyl)pyrazole NCN' pincer Pd(II) complexes. Among them, the P-donor containing complexes are efficiently synthesized by the "one-pot phosphorylation/metalation" method. The obtained symmetrical and unsymmetrical pincer complexes have been used as catalysts in Suzuki-Miyaura reaction (Pd), asymmetric Friedel-Crafts alkylation of indole with trans-β-nitrostyrene (Pt) as well as in asymmetric allylation of aldehyde and sulfonimine (Pd). In the Suzuki couplings conducted at 40-50 °C, some unsymmetrical Pd complexes exhibit much higher activity than the related symmetrical ones which can be attributed to their faster release of active Pd(0) species resulting from the hemilabile coordination of the ligands. Literature results on the synthesis of some related pincer complexes as well as their activities in the above catalytic reactions are also presented.

Complexing of Green Tea Catechins with Food Constituents and Degradation of the Complexes by Lactobacillus plantarum

PubMed Central

HAYASHI, Taeko; UEDA, Shuhei; TSURUTA, Hiroki; KUWAHARA, Hiroshige; OSAWA, Ro

2012-01-01

Complexing of green tea catechins with food constituents and their hydrolysis by tannase-producing Lactobacillus plantarum strains, were investigated. Our observations indicated that 1) epigallocatechin gallate (EGCg) and other catechin galloyl esters bound with food ingredients (i.e., proteins) to form a complex that is likely to be unabsorbable through the intestinal wall, whereas most catechins not esterified with gallic acid (GA) remain in free form, not complexing with food ingredients; 2) tannase activity of L. plantarum is strain dependent, possibly grouped into those with high tannase activity hydrolyzing EGCg to epigallocatechin and GA and those with the low activity; and 3) L. plantarum strains with high tannase activity are capable of hydrolyzing not only intact EGCg but also EGCg and other catechin galloyl esters complexed with dietary proteins to free non-galloyl ester catechins and GA. The evidence suggests that L. plantarum with high tannase activity, if it colonizes the human intestine, would release free non-galloyl-ester catechins and GA that are readily absorbed through the human intestinal epithelia from the complexes, thereby ensuring maximum delivery of the bioactive polyphenols of green tea to the host. PMID:24936346

Nickel(II) Complex of Polyhydroxybenzaldehyde N4-Thiosemicarbazone Exhibits Anti-Inflammatory Activity by Inhibiting NF-κB Transactivation

PubMed Central

Loh, Sheng Wei; Looi, Chung Yeng; Hassandarvish, Pouya; Phan, Alicia Yi Ling; Wong, Won Fen; Wang, Hao; Paterson, Ian C.; Ea, Chee Kwee; Mustafa, Mohd Rais; Maah, Mohd Jamil

2014-01-01

Background The biological properties of thiosemicarbazone have been widely reported. The incorporation of some transition metals such as Fe, Ni and Cu to thiosemicarbazone complexes is known to enhance its biological effects. In this study, we incorporated nickel(II) ions into thiosemicarbazone with N4-substitution groups H3L (H; H3L1, CH3; H3L2, C6H5; H3L3 and C2H5; H3L4) and examined its potential anti-inflammatory activity. Methodology/Principal Findings Four ligands (1–4) and their respective nickel-containing complexes (5–8) were synthesized and characterized. The compounds synthesized were tested for their effects on NF-κB nuclear translocation, pro-inflammatory cytokines secretion and NF-κB transactivation activity. The active compound was further evaluated on its ability to suppress carrageenan-induced acute inflammation in vivo. A potential binding target of the active compound was also predicted by molecular docking analysis. Conclusions/Significance Among all synthesized compounds tested, we found that complex [Ni(H2L1)(PPh3)]Cl (5) (complex 5), potently inhibited IκBα degradation and NF-κB p65 nuclear translocation in LPS-stimulated RAW264.7 cells as well as TNFα-stimulated HeLa S3 cells. In addition, complex 5 significantly down-regulated LPS- or TNFα-induced transcription of NF-κB target genes, including genes that encode the pro-inflammatory cytokines TNFα, IFNβ and IL6. Luciferase reporter assays confirmed that complex 5 inhibited the transactivation activity of NF-κB. Furthermore, the anti-inflammatory effect of complex 5 was also supported by its suppressive effect on carrageenan-induced paw edema formation in wild type C57BL/6 mice. Interestingly, molecular docking study showed that complex 5 potentially interact with the active site of IKKβ. Taken together, we suggest complex 5 as a novel NF-κB inhibitor with potent anti-inflammatory effects. PMID:24977407

Complexation of carbendazim with hydroxypropyl-β-cyclodextrin to improve solubility and fungicidal activity.

PubMed

Ge, Xia; Huang, Zheng; Tian, Shilong; Huang, Yulong; Zeng, Chaozhen

2012-06-05

The effect of hydroxypropyl-β-cyclodextrin (HPβCD) on the improvement of the solubility and fungicidal activity of carbendazim (MBC) has been investigated. The inclusion complexation of HPβCD with MBC has been prepared and characterized by phase solubility diagram, fluorescence, (1)H NMR, ROESY and FT-IR spectra. The stoichiometric ratio and stability constant were determined by Job's plot and phase solubility studies, respectively. The inclusion complex MBC·HPβCD has exhibited different properties from MBC. The obtained inclusion complex was found to significantly improve the water solubility of MBC. In addition, the biological activity indicated that the complex displayed the better fungicidal activity than MBC. The present study provided useful information for a more rational application of MBC. Copyright © 2012 Elsevier Ltd. All rights reserved.

Complexation of sesquiterpene lactones with cyclodextrins: synthesis and effects on their activities on parasitic weeds.

PubMed

Cala, Antonio; Molinillo, José M G; Fernández-Aparicio, Mónica; Ayuso, Jesús; Álvarez, José A; Rubiales, Diego; Macías, Francisco A

2017-08-09

Allelochemicals are safer, more selective and more active alternatives than synthetic agrochemicals for weed control. However, the low solubility of these compounds in aqueous media limits their use as agrochemicals. Herein, we propose the application of α-, β- and γ-cyclodextrins to improve the physicochemical properties and biological activities of three sesquiterpene lactones: dehydrocostuslactone, costunolide and (-)-α-santonin. Complexation was achieved by kneading and coprecipitation methods. Aqueous solubility was increased in the range 100-4600% and the solubility-phase diagrams suggested that complex formation had been successful. The results of the PM3 semiempirical calculations were consistent with the experimental results. The activities on etiolated wheat coleoptiles, Standard Target Species and parasitic weeds were improved. Cyclodextrins preserved or enhanced the activity of the three sesquiterpene lactones. Free cyclodextrins did not show significant activity and therefore the enhancement in activity was due to complexation. These results are promising for applications in agrochemical design.

Dorso-medial and ventro-lateral functional specialization of the human retrosplenial complex in spatial updating and orienting.

PubMed

Burles, Ford; Slone, Edward; Iaria, Giuseppe

2017-04-01

The retrosplenial complex is a region within the posterior cingulate cortex implicated in spatial navigation. Here, we investigated the functional specialization of this large and anatomically heterogeneous region using fMRI and resting-state functional connectivity combined with a spatial task with distinct phases of spatial 'updating' (i.e., integrating and maintaining object locations in memory during spatial displacement) and 'orienting' (i.e., recalling unseen locations from current position in space). Both spatial 'updating' and 'orienting' produced bilateral activity in the retrosplenial complex, among other areas. However, spatial 'updating' produced slightly greater activity in ventro-lateral portions, of the retrosplenial complex, whereas spatial 'orienting' produced greater activity in a more dorsal and medial portion of it (both regions localized along the parieto-occipital fissure). At rest, both ventro-lateral and dorso-medial subregions of the retrosplenial complex were functionally connected to the hippocampus and parahippocampus, regions both involved in spatial orientation and navigation. However, the ventro-lateral subregion of the retrosplenial complex displayed more positive functional connectivity with ventral occipital and temporal object recognition regions, whereas the dorso-medial subregion activity was more correlated to dorsal activity and frontal activity, as well as negatively correlated with more ventral parietal structures. These findings provide evidence for a dorso-medial to ventro-lateral functional specialization within the human retrosplenial complex that may shed more light on the complex neural mechanisms underlying spatial orientation and navigation in humans.

Antibacterial activity and spectral studies of trivalent chromium, manganese, iron macrocyclic complexes derived from oxalyldihydrazide and glyoxal.

PubMed

Singh, D P; Kumar, Ramesh; Singh, Jitender

2009-06-01

A new series of complexes is synthesized by template condensation of oxalyldihydrazide and glyoxal in methanolic medium in the presence of trivalent chromium, manganese and iron salts forming complexes of the type: [M(C(8)H(8)N(8)O(4))X]X(2) where M = Cr(III), Mn(III), Fe(III) and X = Cl(-1), NO(-1)(3), CH(3)COO(-1). The complexes have been characterized with the help of elemental analyses, conductance measurements, magnetic susceptibility measurements, electronic, NMR, infrared and far infrared spectral studies. On the basis of these studies, a five coordinate square pyramidal geometry for these complexes has been proposed. The biological activities of the metal complexes were tested in vitro against a number of pathogenic bacteria and some of the complexes exhibited remarkable antibacterial activities.

Studies on DNA binding behaviour of biologically active transition metal complexes of new tetradentate N2O2 donor Schiff bases: inhibitory activity against bacteria.

PubMed

Sobha, S; Mahalakshmi, R; Raman, N

2012-06-15

A series of Cu(II), Ni(II) and Zn(II) complexes of the type ML have been synthesized with Schiff bases derived from o-acetoacetotoluidide, 2-hydroxybenzaldehyde and o-phenylenediamine/1,4-diaminobutane. The complexes are insoluble in common organic solvents but soluble in DMF and DMSO. The measured molar conductance values in DMSO indicate that the complexes are non-electrolytic in nature. All the six metal complexes have been fully characterized with the help of elemental analyses, molecular weights, molar conductance values, magnetic moments and spectroscopic data. The analytical data helped to elucidate the structure of the metal complexes. The Schiff bases are found to act as tetradentate ligands using N(2)O(2) donor set of atoms leading to a square-planar geometry for the complexes around all the metal ions. The binding properties of metal complexes with DNA were investigated by absorption spectra, viscosity measurements and cyclic voltammetry. Detailed analysis reveals that the metal complexes intercalate into the DNA base stack as intercalators. All the metal complexes cleave the pUC19 DNA in presence of H(2)O(2.) The Schiff bases and their complexes have been screened for their antibacterial activity against five bacterial strains (Staphylococcus aureus, Pseudomonas aeruginosa, Escherichia coli, Staphylococcus epidermidis, Klebsiella pneumoniae) by disk diffusion method. All the metal complexes have potent biocidal activity than the free ligands. Copyright © 2012 Elsevier B.V. All rights reserved.

New bioactive silver(I) complexes: Synthesis, characterization, anticancer, antibacterial and anticarbonic anhydrase II activities

NASA Astrophysics Data System (ADS)

Ozdemir, Ummuhan O.; Ozbek, Neslihan; Genc, Zuhal Karagoz; İlbiz, Firdevs; Gündüzalp, Ayla Balaban

2017-06-01

Silver(I) complexes of alkyl sulfonic acide hydrazides were newly synthesized as homologous series. Methanesulfonic acide hydrazide (L1), ethanesulfonic acide hydrazide (L2), propanesulfonic acide hydrazide (L3) and butanesulfonic acide hydrazide (L4) were used for complexation with Ag(I) ions. The silver complexes obtained in the mol ratio of 1:2 have the structural formula as Ag(L1)2NO3 (I), Ag(L2)2NO3 (II), Ag(L3)2NO3(III), (Ag(L4)2NO3 (IV). The Ag(I) complexes exhibit distorted linear two-fold coordination in [AgL2]+ cations with uncoordinated nitrates. Ligands are chelated with silver(I) ions through unsubstituted primary nitrogen in hydrazide group. Ag(I) complexes were characterized by using elemental analysis, spectroscopic methods (FT-IR, LC-MS), magnetic susceptibility and conductivity measurements. Silver(I) complexes were optimized using PBEPBE/LanL2DZ/DEF2SV basic set performed by DFT method with the Gaussian 09 program package. The geometrical parameters, frontier molecular orbitals (HOMOs and LUMOs) and molecular electrostatic potential (MEP) mapped surfaces of the optimized geometries were also determined by this quantum set. The anticancer activities of silver(I) complexes on MCF-7 human breast cancer cell line were investigated by comparing IC50 values. The antibacterial activities of complexes were studied against Gram positive bacteria; S. aureus ATCC 6538, B. subtilis ATCC 6633, B. cereus NRRL-B-3711, E. faecalis ATCC 29212 and Gram negative bacteria; E. coli ATCC 11230, P. aeruginosa ATCC 15442, K. pneumonia ATCC 70063 by using disc diffusion method. The inhibition activities of Ag(I) complexes on carbonic anhydrase II enzyme (hCA II) were also investigated by comparing IC50 and Ki values. The biological activity screening shows that Ag(I) complex of butanesulfonicacidehydrazide (IV) has the highest activity against tested breast cancer cell lines MCF-7, Gram positive/Gram negative bacteria and carbonic anhydrase II (hCA II) isoenzyme.

Neural correlates in the processing of phoneme-level complexity in vowel production.

PubMed

Park, Haeil; Iverson, Gregory K; Park, Hae-Jeong

2011-12-01

We investigated how articulatory complexity at the phoneme level is manifested neurobiologically in an overt production task. fMRI images were acquired from young Korean-speaking adults as they pronounced bisyllabic pseudowords in which we manipulated phonological complexity defined in terms of vowel duration and instability (viz., COMPLEX: /tiɯi/ > MID-COMPLEX: /tiye/ > SIMPLE: /tii/). Increased activity in the left inferior frontal gyrus (Brodmann Areas (BA) 44 and 47), supplementary motor area and anterior insula was observed for the articulation of COMPLEX sequences relative to MID-COMPLEX; this was the case with the articulation of MID-COMPLEX relative to SIMPLE, except that the pars orbitalis (BA 47) was dominantly identified in the Broca's area. The differentiation indicates that phonological complexity is reflected in the neural processing of distinct phonemic representations, both by recruiting brain regions associated with retrieval of phonological information from memory and via articulatory rehearsal for the production of COMPLEX vowels. In addition, the finding that increased complexity engages greater areas of the brain suggests that brain activation can be a neurobiological measure of articulo-phonological complexity, complementing, if not substituting for, biomechanical measurements of speech motor activity. 2011 Elsevier Inc. All rights reserved.

Comparative study of copper(II)-curcumin complexes as superoxide dismutase mimics and free radical scavengers.

PubMed

Barik, Atanu; Mishra, Beena; Kunwar, Amit; Kadam, Ramakant M; Shen, Liang; Dutta, Sabari; Padhye, Subhash; Satpati, Ashis K; Zhang, Hong-Yu; Indira Priyadarsini, K

2007-04-01

Two stoichiometrically different copper(II) complexes of curcumin (stoichiometry, 1:1 and 1:2 for copper:curcumin), were examined for their superoxide dismutase (SOD) activity, free radical-scavenging ability and antioxidant potential. Both the complexes are soluble in lipids and DMSO. The formation constants of the complexes were determined by voltammetry. EPR spectra of the complexes in DMSO at 77K showed that the 1:2 Cu(II)-curcumin complex is square planar and the 1:1 Cu(II)-curcumin complex is distorted orthorhombic. Cu(II)-curcumin complex (1:1) with larger distortion from square planar structure shows higher SOD activity. These complexes inhibit gamma-radiation induced lipid peroxidation in liposomes and react with DPPH acting as free radical scavengers. One-electron oxidation of the two complexes by radiolytically generated azide radicals in Tx-100 micellar solutions produced phenoxyl radicals, indicating that the phenolic moiety of curcumin in the complexes participates in free radical reactions. Depending on the structure, these two complexes possess different SOD activities, free radical neutralizing abilities and antioxidant potentials. In addition, quantum chemical calculations with density functional theory have been performed to support the experimental observations.

Synthesis, spectroscopic characterization and in vitro cytotoxicities of new organometallic palladium complexes with biologically active β-diketones; Biological evaluation probing of the interaction mechanism with DNA/Protein and molecular docking

NASA Astrophysics Data System (ADS)

Karami, Kazem; Rafiee, Mina; Lighvan, Zohreh Mehri; Zakariazadeh, Mostafa; Faal, Ali Yeganeh; Esmaeili, Seyed-Alireza; Momtazi-Borojeni, Amir Abbas

2018-02-01

[Pd{(C,N)sbnd C6H4CH (CH3)NH}(CUR)] (3) and [Pd2{(C,N)sbnd C6H4CH(CH3)NH2}2(μ-N3CS2)] (4) [cur = 1,7-bis(4-hydroxy-3-methoxyphenyl)-1,6-heptadiene-3,5-dion] novel organometallic complexes with biologically active ligands have been prepared and characterized via elemental analysis, multinuclear spectroscopic techniques (1H, and 13C NMR and IR) and their biological activities, including antitumoral activity and DNA-protein interactions have been investigated. Fluorescence spectroscopy used to study the interaction of the complexes with BSA have shown the affinity of the complexes for these proteins with relatively high binding constant values and the changed secondary structure of BSA in the presence of the complexes. In the meantime, spectroscopy and competitive titration have been applied to investigate the interaction of complexes with Warfarin and Ibuprofen site markers for sites I and II, respectively, with BSA. The results have suggested that the locations of complexes 3 and 4 are sites II and I, respectively. UV-Vis spectroscopy, emission titration and helix melting methods have been used to study the interaction of these complexes with CT-DNA, indicating that complexes are bound to CT-DNA by intercalation binding mode. In addition, good cytotoxic activity against MCF-7 (human breast cancer) and JURKAT (human leukemia) cell line has been shown by both complexes whereas low cytotoxicity was exerted on normal peripheral blood mononuclear cells.

Photo-activated luminescence sensor and method of detecting trichloroethylene and related volatile organochloride compounds

DOEpatents

Dinh, Tuan V.

1996-01-01

A sensor for detecting trichloroethylene and related volatile organochloride compounds uses a photo-activator that produces a photo-product complex with the contaminant. Characteristics of the light emitted from the complex will indicate the presence of the contaminant. A probe containing the photo-activator has an excitation light interface and a contaminant interface. One particular embodiment uses a porous membrane as the contaminant interface, so that the contaminant can migrate therethrough to the photo-activator and thereby form the complex.

Staphylococcus-mediated T-cell activation and spontaneous natural killer cell activity in the absence of major histocompatibility complex class II molecules

NASA Technical Reports Server (NTRS)

Chapes, S. K.; Hoynowski, S. M.; Woods, K. M.; Armstrong, J. W.; Beharka, A. A.; Iandolo, J. J.; Spooner, B. S. (Principal Investigator)

1993-01-01

We used major histocompatibility complex class II antigen-deficient transgenic mice to show that in vitro natural killer cell cytotoxicity and T-cell activation by staphylococcal exotoxins (superantigens) are not dependent upon the presence of major histocompatibility complex class II molecules. T cells can be activated by exotoxins in the presence of exogenously added interleukin 1 or 2 or in the presence of specific antibody without exogenously added cytokines.

Metal complexes of diisopropylthiourea: synthesis, characterization and antibacterial studies.

PubMed

Ajibade, Peter A; Zulu, Nonkululeko H

2011-01-01

Co(II), Cu(II), Zn(II) and Fe(III) complexes of diisopropylthiourea have been synthesized and characterized by elemental analyses, molar conductivity, magnetic susceptibility, FTIR and electronic spectroscopy. The compounds are non-electrolytes in solution and spectroscopic data of the complexes are consistent with 4-coordinate geometry for the metal(II) complexes and six coordinate octahedral for Fe(III) complex. The complexes were screened for their antibacterial activities against six bacteria: Escherichia coli, Pseudomonas auriginosa, Klebsiella pneumoniae, Bacillus cereus, Staphylococcus aureus and Bacillus pumilus. The complexes showed varied antibacterial activities and their minimum inhibitory concentrations (MICs) were determined.

Synthesis, spectroscopic characterization, electrochemistry and biological evaluation of some metal (II) complexes with ONO donor ligand containing benzo[b]thiophene and coumarin moieties

NASA Astrophysics Data System (ADS)

Mahendra Raj, K.; Mruthyunjayaswamy, B. H. M.

2014-09-01

Schiff base ligand 3-chloro-N‧-((7-hydroxy-4-methyl-2-oxo-2H-chromen-8-yl)methylene)benzo[b]thiophene-2-carbohydrazide and its Cu(II), Co(II), Ni(II) and Zn(II) complexes were synthesized, characterized by elemental analysis and various physico-chemical techniques like, IR, 1H NMR, ESI-mass, UV-Visible, thermogravimetry - differential thermal analysis, magnetic measurements and molar conductance. Spectral analysis indicates octahedral geometry for all the complexes. Cu(II) complex have 1:1 stoichiometry of the type [M(L)(Cl)(H2O)2], whereas Co(II), Ni(II) and Zn(II) complexes have 1:2 stoichiometric ratio of the type [M(L)2]. The bonding sites are the oxygen atom of amide carbonyl, nitrogen of azomethine function and phenolic oxygen of the Schiff base ligand via deprotonation. The thermogravimetry - differential thermal analysis studies gave evidence for the presence of coordinated water molecules in the composition of Cu(II) complex which was further supported by IR measurements. All the complexes were investigated for their electrochemical activity, but only the Cu(II) complex showed the redox property. In order to evaluate the effect of antimicrobial potency of metal ions upon chelation, ligand and its metal complexes along with their respective metal chlorides were screened for their antibacterial and antifungal activities by minimum inhibitory concentration (MIC) method. The results showed that the metal complexes were found to be more active than free ligand. Ligand and its complexes were screened for free radical scavenging activity by DPPH method and DNA cleavage activity using Calf-thymus DNA (Cat. No-105850).

Regulation of succinate-ubiquinone reductase and fumarate reductase activities in human complex II by phosphorylation of its flavoprotein subunit.

PubMed

Tomitsuka, Eriko; Kita, Kiyoshi; Esumi, Hiroyasu

2009-01-01

Complex II (succinate-ubiquinone reductase; SQR) is a mitochondrial respiratory chain enzyme that is directly involved in the TCA cycle. Complex II exerts a reverse reaction, fumarate reductase (FRD) activity, in various species such as bacteria, parasitic helminths and shellfish, but the existence of FRD activity in humans has not been previously reported. Here, we describe the detection of FRD activity in human cancer cells. The activity level was low, but distinct, and it increased significantly when the cells were cultured under hypoxic and glucose-deprived conditions. Treatment with phosphatase caused the dephosphorylation of flavoprotein subunit (Fp) with a concomitant increase in SQR activity, whereas FRD activity decreased. On the other hand, treatment with protein kinase caused an increase in FRD activity and a decrease in SQR activity. These data suggest that modification of the Fp subunit regulates both the SQR and FRD activities of complex II and that the phosphorylation of Fp might be important for maintaining mitochondrial energy metabolism within the tumor microenvironment.

Differential responses of targeted lung redox enzymes to rat exposure to 60 or 85% oxygen

PubMed Central

Gan, Zhuohui; Roerig, David L.; Clough, Anne V.

2011-01-01

Rat exposure to 60% O2 (hyper-60) or 85% O2 (hyper-85) for 7 days confers susceptibility or tolerance, respectively, of the otherwise lethal effects of exposure to 100% O2. The objective of this study was to determine whether activities of the antioxidant cytosolic enzyme NAD(P)H:quinone oxidoreductase 1 (NQO1) and mitochondrial complex III are differentially altered in hyper-60 and hyper-85 lungs. Duroquinone (DQ), an NQO1 substrate, or its hydroquinone (DQH2), a complex III substrate, was infused into the arterial inflow of isolated, perfused lungs, and the venous efflux rates of DQH2 and DQ were measured. Based on inhibitor effects and kinetic modeling, capacities of NQO1-mediated DQ reduction (Vmax1) and complex III-mediated DQH2 oxidation (Vmax2) increased by ∼140 and ∼180% in hyper-85 lungs, respectively, compared with rates in lungs of rats exposed to room air (normoxic). In hyper-60 lungs, Vmax1 increased by ∼80%, with no effect on Vmax2. Additional studies revealed that mitochondrial complex I activity in hyper-60 and hyper-85 lung tissue homogenates was ∼50% lower than in normoxic lung homogenates, whereas mitochondrial complex IV activity was ∼90% higher in only hyper-85 lung tissue homogenates. Thus NQO1 activity increased in both hyper-60 and hyper-85 lungs, whereas complex III activity increased in hyper-85 lungs only. This increase, along with the increase in complex IV activity, may counter the effects the depression in complex I activity might have on tissue mitochondrial function and/or reactive oxygen species production and may be important to the tolerance of 100% O2 observed in hyper-85 rats. PMID:21551015

Differential responses of targeted lung redox enzymes to rat exposure to 60 or 85% oxygen.

PubMed

Gan, Zhuohui; Roerig, David L; Clough, Anne V; Audi, Said H

2011-07-01

Rat exposure to 60% O(2) (hyper-60) or 85% O(2) (hyper-85) for 7 days confers susceptibility or tolerance, respectively, of the otherwise lethal effects of exposure to 100% O(2). The objective of this study was to determine whether activities of the antioxidant cytosolic enzyme NAD(P)H:quinone oxidoreductase 1 (NQO1) and mitochondrial complex III are differentially altered in hyper-60 and hyper-85 lungs. Duroquinone (DQ), an NQO1 substrate, or its hydroquinone (DQH(2)), a complex III substrate, was infused into the arterial inflow of isolated, perfused lungs, and the venous efflux rates of DQH(2) and DQ were measured. Based on inhibitor effects and kinetic modeling, capacities of NQO1-mediated DQ reduction (V(max1)) and complex III-mediated DQH(2) oxidation (V(max2)) increased by ∼140 and ∼180% in hyper-85 lungs, respectively, compared with rates in lungs of rats exposed to room air (normoxic). In hyper-60 lungs, V(max1) increased by ∼80%, with no effect on V(max2). Additional studies revealed that mitochondrial complex I activity in hyper-60 and hyper-85 lung tissue homogenates was ∼50% lower than in normoxic lung homogenates, whereas mitochondrial complex IV activity was ∼90% higher in only hyper-85 lung tissue homogenates. Thus NQO1 activity increased in both hyper-60 and hyper-85 lungs, whereas complex III activity increased in hyper-85 lungs only. This increase, along with the increase in complex IV activity, may counter the effects the depression in complex I activity might have on tissue mitochondrial function and/or reactive oxygen species production and may be important to the tolerance of 100% O(2) observed in hyper-85 rats.

Pharmacologically significant complexes of Mn(II), Co(II), Ni(II), Cu(II) and Zn(II) of novel Schiff base ligand, (E)-N-(furan-2-yl methylene) quinolin-8-amine: Synthesis, spectral, XRD, SEM, antimicrobial, antioxidant and in vitro cytotoxic studies

NASA Astrophysics Data System (ADS)

Shakir, M.; Hanif, Summaiya; Sherwani, Mohd. Asif; Mohammad, Owais; Al-Resayes, Saud I.

2015-07-01

A novel series of metal complexes of the types, [ML2(H2O)2]Cl2 and [ML2]Cl2 [M = Mn(II), 1; Co(II), 2; Ni(II), 3; Cu(II), 4; and Zn(II), 5] were synthesized by the interaction of ligand, L (E)-N-(furan-2-yl methylene) quinolin-8-amine, derived from the condensation of 2-furaldehyde and 8-aminoquinoline. The synthesized ligand and its metal complexes were characterized on the basis of results obtained from elemental analysis, ESI-MS, XRD, SEM, TGA/DTA, FT-IR, UV-Vis, magnetic moment and 1H and 13C NMR spectroscopic studies. EPR parameters were recorded in case of complex 4. The comparative in-vitro antimicrobial activities against various pathogens with reference to known antibiotics and antioxidant activity against standard control at variable concentrations revealed that the metal complexes show enhanced antimicrobial and free radical scavenging activities in general as compared to free ligand. However, the complexes 1 and 5 have shown best antioxidant activity among all the metal complexes. Furthermore, comparative in-vitro antiproliferative activity on ligand and its metal chelates performed on MDA-MB-231 (breast carcinoma), KCL22 (blood lymphoid carcinoma), HeLa (cervical carcinoma) cell lines and normal cells (PBMC) revealed that metal chelates show moderate to good activity as compared to ligand where as complex 1 seems to be the most promising one possessing a broad spectrum of activity against all the selected cancer cell lines with IC50 < 2.10 μM.

A supramolecular complex between proteinases and beta-cyclodextrin that preserves enzymatic activity: physicochemical characterization.

PubMed

Denadai, Angelo M L; Santoro, Marcelo M; Lopes, Miriam T P; Chenna, Angélica; de Sousa, Frederico B; Avelar, Gabriela M; Gomes, Marco R Túlio; Guzman, Fanny; Salas, Carlos E; Sinisterra, Rubén D

2006-01-01

Cyclodextrins are suitable drug delivery systems because of their ability to subtly modify the physical, chemical, and biological properties of guest molecules through labile interactions by formation of inclusion and/or association complexes. Plant cysteine proteinases from Caricaceae and Bromeliaceae are the subject of therapeutic interest, because of their anti-inflammatory, antitumoral, immunogenic, and wound-healing properties. In this study, we analyzed the association between beta-cyclodextrin (betaCD) and fraction P1G10 containing the bioactive proteinases from Carica candamarcensis, and described the physicochemical nature of the solid-state self-assembled complexes by Fourier transform infrared (FTIR) spectroscopy, thermogravimetry (TG), differential scanning calorimetry (DSC), X-ray powder diffraction (XRD), and nuclear magnetic resonance (NMR), as well as in solution by circular dichroism (CD), isothermal titration calorimetry (ITC), and amidase activity. The physicochemical analyses suggest the formation of a complex between P1G10 and betaCD. Higher secondary interactions, namely hydrophobic interactions, hydrogen bonding and van der Waals forces were observed at higher P1G10 : betaCD mass ratios. These results provide evidence of the occurrence of strong solid-state supramolecular non-covalent interactions between P1G10 and betaCD. Microcalorimetric analysis demonstrates that complexation results in a favorable enthalpic contribution, as has already been described during formation of similar betaCD inclusion compounds. The amidase activity of the complex shows that the enzyme activity is not readily available at 24 hours after dissolution of the complex in aqueous buffer; the proteinase becomes biologically active by the second day and remains stable until day 16, when a gradual decrease occurs, with basal activity attained by day 29. The reported results underscore the potential for betaCDs as candidates for complexing cysteine proteinases, resulting in supramolecular arrays with sustained proteolytic activity.

DNA Binding, Cleavage and Antibacterial Activity of Mononuclear Cu(II), Ni(II) and Co(II) Complexes Derived from Novel Benzothiazole Schiff Bases.

PubMed

Vamsikrishna, Narendrula; Kumar, Marri Pradeep; Tejaswi, Somapangu; Rambabu, Aveli; Shivaraj

2016-07-01

A series of novel bivalent metal complexes M(L1)2 and M(L2)2 where M = Cu(II), Ni(II), Co(II) and L1 = 2-((benzo [d] thiazol-6-ylimino)methyl)-4-bromophenol [BTEMBP], L2 = 1-((benzo [d] thiazol-6-ylimino)methyl) naphthalen-2-ol [BTEMNAPP] were synthesized. All the compounds have been characterized by elemental analysis, SEM, Mass, (1)H NMR, (13)C NMR, UV-Vis, IR, ESR, spectral data and magnetic susceptibility measurements. Based on the analytical and spectral data four-coordinated square planar geometry is assigned to all the complexes. DNA binding properties of these complexes have been investigated by electronic absorption spectroscopy, fluorescence and viscosity measurements. It is observed that these binary complexes strongly bind to calf thymus DNA by an intercalation mode. DNA cleavage efficacy of these complexes was tested in presence of H2O2 and UV light by gel electrophoresis and found that all the complexes showed better nuclease activity. Finally the compounds were screened for antibacterial activity against few pathogens and found that the complexes have potent biocidal activity than their free ligands.

Syntheses, Characterization, Resolution, and Biological Studies of Coordination Compounds of Aspartic Acid and Glycine

PubMed Central

Akinkunmi, Ezekiel; Ojo, Isaac; Adebajo, Clement; Isabirye, David

2017-01-01

Enantiomerically enriched coordination compounds of aspartic acid and racemic mixtures of coordination compounds of glycine metal-ligand ratio 1 : 3 were synthesized and characterized using infrared and UV-Vis spectrophotometric techniques and magnetic susceptibility measurements. Five of the complexes were resolved using (+)-cis-dichlorobis(ethylenediamine)cobalt(III) chloride, (+)-bis(glycinato)(1,10-phenanthroline)cobalt(III) chloride, and (+)-tris(1,10-phenanthroline)nickel(II) chloride as resolving agents. The antimicrobial and cytotoxic activities of these complexes were then determined. The results obtained indicated that aspartic acid and glycine coordinated in a bidentate fashion. The enantiomeric purity of the compounds was in the range of 22.10–32.10%, with (+)-cis-dichlorobis(ethylenediamine)cobalt(III) complex as the more efficient resolving agent. The resolved complexes exhibited better activity in some cases compared to the parent complexes for both biological activities. It was therefore inferred that although the increase in the lipophilicity of the complexes may assist in the permeability of the complexes through the cell membrane of the pathogens, the enantiomeric purity of the complexes is also of importance in their activity as antimicrobial and cytotoxic agents. PMID:28293149

Synthesis, structures and urease inhibitory activity of cobalt(III) complexes with Schiff bases.

PubMed

Jing, Changling; Wang, Cunfang; Yan, Kai; Zhao, Kedong; Sheng, Guihua; Qu, Dan; Niu, Fang; Zhu, Hailiang; You, Zhonglu

2016-01-15

A series of new cobalt(III) complexes were prepared. They are [CoL(1)(py)3]·NO3 (1), [CoL(2)(bipy)(N3)]·CH3OH (2), [CoL(3)(HL(3))(N3)]·NO3 (3), and [CoL(4)(MeOH)(N3)] (4), where L(1), L(2), L(3) and L(4) are the deprotonated form of N'-(2-hydroxy-5-methoxybenzylidene)-3-methylbenzohydrazide, N'-(2-hydroxybenzylidene)-3-hydroxylbenzohydrazide, 2-[(2-dimethylaminoethylimino)methyl]-4-methylphenol, and N,N'-bis(5-methylsalicylidene)-o-phenylenediamine, respectively, py is pyridine, and bipy is 2,2'-bipyridine. The complexes were characterized by infrared and UV-Vis spectra, and single crystal X-ray diffraction. The Co atoms in the complexes are in octahedral coordination. Complexes 1 and 4 show effective urease inhibitory activities, with IC50 values of 4.27 and 0.35 μmol L(-1), respectively. Complex 2 has medium activity against urease, with IC50 value of 68.7 μmol L(-1). While complex 3 has no activity against urease. Molecular docking study of the complexes with Helicobacter pylori urease was performed. Copyright © 2015 Elsevier Ltd. All rights reserved.

Template engineered biopotent macrocyclic complexes involving furan moiety: Molecular modeling and molecular docking

NASA Astrophysics Data System (ADS)

Rathi, Parveen; Singh, D. P.

2015-08-01

Bioactive cobalt(II), nickel(II), copper(II) and zinc(II) complexes of octaazamacrocycle, 19, 20-dioxa-2,3,5,6,11,12,14,15-octaazatricyclo[14.2.1.1]icosa-1,6,8,10,15,17-hexaene-4,13-dithione, derived from furan-2,5-dione and thiocarbonohydrazide in the mole ratio 2:2:1 have been engineered via template methodology. The synthesized metal complexes have also been structurally characterized in the light of various physicochemical techniques and evaluated for antimicrobial and antioxidant activities. All these studies point toward the formation of divalent macrocyclic complexes possessing distorted octahedral geometry and having significant antimicrobial and antioxidant properties as compared to the starting precursors. Virtual screening of a representative complex was done through docking to the binding site of COX-2 to evaluate the anti-inflammatory activity of the series. Non-electrolytic nature of the complexes has been predicted on the basis of low value of molar conductivity in DMSO. All the complexes were having notable activities against pathogenic microbes as compared to precursors-thiocarbonohydrazide and furan-2,5-dione however, the complex 5, [Ni (C10H8N8O2S2) (NO3)2], shows the best antimicrobial activity.

Fe (III), Co(II), Ni(II), Cu(II) and Zn(II) complexes of schiff bases based-on glycine and phenylalanine: Synthesis, magnetic/thermal properties and antimicrobial activity

NASA Astrophysics Data System (ADS)

Sevgi, Fatih; Bagkesici, Ugur; Kursunlu, Ahmed Nuri; Guler, Ersin

2018-02-01

Zinc (II), copper (II), nickel (II), cobalt (II) and iron (III) complexes of Schiff bases (LG, LP) derived from 2-hydroxynaphthaldehyde with glycine and phenylalanine were reported and characterized by 1H NMR, 13C NMR, elemental analyses, melting point, FT-IR, magnetic susceptibility and thermal analyses (TGA). TGA data show that iron and cobalt include to the coordinated water and metal:ligand ratio is 1:2 while the complex stoichiometry for Ni (II), Cu (II) and Zn (II) complexes is 1:1. As expected, Ni (II) and Zn (II) complexes are diamagnetic; Cu (II), Co (II) and Fe (III) complexes are paramagnetic character due to a strong ligand of LG and LP. The LG, LP and their metal complexes were screened for their antimicrobial activities against five Gram-positive (Staphylococcus aureus, Methicillin resistant Staphylococcus aureus (MRSA), Bacillus cereus, Streptococcus mutans and Enterococcus faecalis) and three Gram-negative (Escherichia coli, Klebsiella pneumoniae and Pseudomonas aeruginosa) and one fungi (Candida albicans) by using broth microdilution techniques. The activity data show that ligands and their metal complexes exhibited moderate to good activity against Gram-positive bacteria and fungi.

Metal complexes of 3-(4-bromophenyl)-1-pyridin-2-ylprop-2-en-1-one thiosemicarbazone: cytotoxic activity and investigation on the mode of action of the gold(III) complex.

PubMed

Sâmia, Luciana B P; Parrilha, Gabrieli L; Da Silva, Jeferson G; Ramos, Jonas P; Souza-Fagundes, Elaine M; Castelli, Silvia; Vutey, Venn; Desideri, Alessandro; Beraldo, Heloisa

2016-06-01

Complexes [Au(PyCT4BrPh)Cl]Cl (1), [Pt(PyCT4BrPh)Cl]0.5KCl (2), and [Pd(PyCT4BrPh)Cl]KCl (3) were obtained with 3-(4-bromophenyl)-1-pyridin-2-ylprop-2-en-1-one thiosemicarbazone (HPyCT4BrPh). Although complexes (2) and (3) did not exhibit potent cytotoxic activity, HPyCT4BrPh and its gold(III) complex (1) proved to be highly cytotoxic against HL-60 (human promyelocytic leukemia) and THP-1 (human monocytic leukemia) cells, and against MDA-MB 231 and MCF-7 (human breast adenocarcinoma) solid tumor cells. Except for HL-60 cells, upon coordination to gold(III) a 2- to 3-fold increase in the cytotoxic effect was observed. An investigation on the possible biological targets of the gold(III) complex was carried out. Complex (1) but not the free thiosemicarbazone inhibits the enzymatic activity of thioredoxin reductase (TrxR). The affinity of 1 for TrxR suggests metal binding to a selenol residue in the active site of the enzyme. While HPyCT4BrPh was inactive, 1 was able to inhibit topoisomerase IB (Topo IB) activity. Hence, inhibition of TrxR and Topo IB could contribute to the mechanism of cytotoxic action of complex (1).

GA binding protein augments autophagy via transcriptional activation of BECN1-PIK3C3 complex genes

PubMed Central

Zhu, Wan; Swaminathan, Gayathri; Plowey, Edward D

2014-01-01

Macroautophagy is a vesicular catabolic trafficking pathway that is thought to protect cells from diverse stressors and to promote longevity. Recent studies have revealed that transcription factors play important roles in the regulation of autophagy. In this study, we have identified GA binding protein (GABP) as a transcriptional regulator of the combinatorial expression of BECN1-PIK3C3 complex genes involved in autophagosome initiation. We performed bioinformatics analyses that demonstrated highly conserved putative GABP sites in genes that encode BECN1/Beclin 1, several BECN1 interacting proteins, and downstream autophagy proteins including the ATG12–ATG5-ATG16L1 complex. We demonstrate that GABP binds to the promoter regions of BECN1-PIK3C3 complex genes and activates their transcriptional activities. Knockdown of GABP reduced BECN1-PIK3C3 complex transcripts, BECN1-PIK3C3 complex protein levels and autophagy in cultured cells. Conversely, overexpression of GABP increased autophagy. Nutrient starvation increased GABP-dependent transcriptional activity of BECN1-PIK3C3 complex gene promoters and increased the recruitment of GABP to the BECN1 promoter. Our data reveal a novel function of GABP in the regulation of autophagy via transcriptional activation of the BECN1-PIK3C3 complex. PMID:25046113

Carbohydrate linked organotin(IV) complexes as human topoisomerase Iα inhibitor and their antiproliferative effects against the human carcinoma cell line.

PubMed

Khan, Rais Ahmad; Yadav, Shipra; Hussain, Zahid; Arjmand, Farukh; Tabassum, Sartaj

2014-02-14

Dimethyltin(IV) complexes with ethanolamine (1) and biologically significant N-glycosides (2 and 3) were designed and synthesized. The structural elucidation of complexes 1-3 was done using elemental and spectroscopic methods; in addition, complex 1 was studied by single crystal X-ray diffraction studies. The in vitro DNA binding profile of complexes 2 and 3 was carried out by employing different biophysical methods to ascertain the feasibility of glycosylated complexes. Further, the cleaving ability of 2 and 3 was investigated by the agarose gel electrophoretic mobility assay with supercoiled pBR322 DNA, and demonstrated significantly good nuclease activity. Furthermore, both the complexes exhibited significant inhibitory effects on the catalytic activity of human Topo I at lower concentration than standard drugs. Computer-aided molecular docking techniques were used to ascertain the mode and mechanism of action towards the molecular target DNA and Topo I. The cytotoxicity of 2 and 3 against human hepatoma cancer cells (Huh7) was evaluated, which revealed significant regression in cancerous cells as compared with the standard drug. The antiproliferative activities of 2 and 3 were tested against human hepatoma cancer cells (Huh7), and results showed significantly good activity. Additionally, to validate the remarkable antiproliferative activity of complexes 2 and 3, specific regulatory gene expression (MMP-2 and TGF-β) was obtained by real time PCR.

Structural characterization of 1,8-naphthalimides and in vitro microbiological activity of their Cu(II) and Zn(II) complexes

NASA Astrophysics Data System (ADS)

Grabchev, Ivo; Yordanova, Stanislava; Bosch, Paula; Vasileva-Tonkova, Evgenia; Kukeva, Rositsa; Stoyanov, Stanimir; Stoyanova, Radostina

2017-02-01

Two new 1,8-naphthalimide derivatives (NI1 and NI2) have been synthesized and characterized. The photophysical properties of the new compounds have been investigated in organic solvents of different polarity. It has been shown that both compounds are solvent depended. Cu(II) and Zn(II) complexes of NI2 were obtained and characterized by IR-NMR, fluorescence and EPR spectroscopy. The influence of different metal cations on the fluorescence intensity has been investigated in acetonitrile solution. Antimicrobial composite PLA-metal complexes materials have been obtained for the first time. Microbiological activity of both metal complexes has been investigated in vitro against different Gram-positive and Gram-negative bacteria and two yeasts. The various antimicrobial activities and the minimum inhibitory concentrations (MICs) of both complexes have been determined. The microbiological activity of composite materials PLA-metal complexes in thin polymeric film has also been investigated. The results suggest that the new metal complexes could find application in designing new antimicrobial preparations to control the spread of infections.

Synthesis, characterization, and anti-cancer activity of emodin-Mn(II) metal complex.

PubMed

Yang, Li; Tan, Jun; Wang, Bo-Chu; Zhu, Lian-Cai

2014-12-01

To synthesize and characterize a novel metal complex of Mn (II) with emodin, and evaluate its anti-cancer activity. The elemental analyses, IR, UV-vis, atomic absorption spectroscopy, TG-DSC, (1)H NMR, and (13)C NMR data were used to characterize the structure of the complex. The cytotoxicity of the complex against the human cancer cell lines HepG2, HeLa, MCF-7, B16, and MDA-MB-231 was tested by the MTT assay and flow cytometry. Emodin was coordinated with Mn(II) through the 9-C=O and 1-OH, and the general formula of the complex was Mn(II) (emodin)2·2H2O. In studies of the cytotoxicity, the complex exhibited significant activity, and the IC50 values of the complex against five cancer cell lines improved approximately three-fold compared with those of emodin. The complex could induce cell morphological changes, decrease the percentage of viability, and induce G0/G1 phase arrest and apoptosis in cancer cells. The coordination of emodin with Mn(II) can improve its anticancer activity, and the complex Mn(II) (emodin)2·2H2O could be studied further as a promising anticancer drug. Copyright © 2014 China Pharmaceutical University. Published by Elsevier B.V. All rights reserved.

Spectral studies, thermal investigation and biological activity of some metal complexes derived from (E)-N‧-(1-(4-aminophenyl)ethylidene)morpholine-4-carbothiohydrazide

NASA Astrophysics Data System (ADS)

El-Samanody, El-Sayed A.; Polis, Magdy W.; Emara, Esam M.

2017-09-01

A new series of biologically active Co(II), Ni(II), Cu(II), Zn(II) and Cd(II) complexes derived from the novel thiosemicarbazone ligand; (E)-N‧-(1-(4-aminophenyl)ethylidene)morpholine-4-carbothiohydrazide (HL) were synthesized. The mode of bonding of the ligand and the geometrical structures of its metal complexes were achieved by different analytical and spectral methods. The ligand coordinated with metal ions in a neutral bidentate fashion through the thione sulfur and azomethine nitrogen atoms. All metal complexes adopted octahedral geometry, except Cu(II) complexes (3, 6, 7) which have a square planar structure. The general thermal decomposition pathways of the ligand along with its metal complexes were explained. The thermal stability of the complexes is controlled by the number of outer and inner sphere water molecules, ionic radii and the steric hindrance. The activation thermodynamic parameters; (activation energy (E*), enthalpy of activation (ΔH*), entropy of activation (ΔS*) and Gibbs free energy (ΔG*)) along with order of reaction (n) were estimated from DTG curves. The ESR spectra of Cu(II) complexes indicated that (dx2-y2)1 is the ground state with covalence character of metal-ligand bonds. The molluscicidal and biochemical effects of the ligand and its Ni(II); Cu(II) complexes (2; 3, 5, 7) along with their combinations with metaldehyde were screened in vitro on the mucous gland of Eobania vermiculata. The tested compounds exhibited a significant toxicity against the tested animals and have almost the same toxic effect of metaldehyde which increases the mucous secretion of the snails and leads to death.

Pentamethylcyclopentadienyl-rhodium and iridium complexes containing (N^N and N^O) bound chloroquine analogue ligands: synthesis, characterization and antimalarial properties.

PubMed

Ekengard, Erik; Kumar, Kamlesh; Fogeron, Thibault; de Kock, Carmen; Smith, Peter J; Haukka, Matti; Monari, Magda; Nordlander, Ebbe

2016-03-07

The synthesis and characterization of twenty new pentamethylcyclopentadienyl-rhodium and iridium complexes containing N^N and N^O-chelating chloroquine analogue ligands are described. The in vitro antimalarial activity of the new ligands as well as the complexes was evaluated against the chloroquine sensitive (CQS) NF54 and the chloroquine resistant (CQR) Dd2 strains of Plasmodium falciparum. The antimalarial activity was found to be good to moderate; although all complexes are less active than artesunate, some of the ligands and complexes showed better activity than chloroquine (CQ). In particular, rhodium complexes were found to be considerably more active than iridium complexes against the CQS NF54 strain. Salicylaldimine Schiff base ligands having electron-withdrawing groups (F, Cl, Br, I and NO2) in para position of the salicyl moiety and their rhodium complexes showed good antiplasmodial activity against both the CQS-NF54 and the CQR-Dd2 strains. The crystal structures of (η(5)-pentamethylcyclopentadienyl){N(1)-(7-chloroquinolin-4-yl)-N(2)-(pyridin-2-ylmethyl)ethane-1,2-diamine)} chlororhodium(III) chloride and (η(5)-pentamethylcyclopentadienyl){(4-chloro-2-(((2-((7-chloroquinolin-4-yl)amino)ethyl)imino)methyl)phenolate)}chlororhodium(III) chloride are reported. The crystallization of the amino-pyridyl complex (η(5)-pentamethylcyclopentadienyl){(N(1)-(7-chloroquinolin-4-yl)-N(2)-(pyridin-2-ylmethyl)ethane-1,2-diamine)}chloroiridium(III) chloride in acetone resulted in the formation of the imino-pyridyl derivative (η(5)-pentamethylcyclopentadienyl){(N1-(7-chloroquinolin-4-yl)-N2-(pyridin-2-ylmethylene)ethane-1,2-diamine)}chloroiridium(III) chloride, the crystal structure of which is also reported.

Biotinylated Rh(III) complexes in engineered streptavidin for accelerated asymmetric C-H activation.

PubMed

Hyster, Todd K; Knörr, Livia; Ward, Thomas R; Rovis, Tomislav

2012-10-26

Enzymes provide an exquisitely tailored chiral environment to foster high catalytic activities and selectivities, but their native structures are optimized for very specific biochemical transformations. Designing a protein to accommodate a non-native transition metal complex can broaden the scope of enzymatic transformations while raising the activity and selectivity of small-molecule catalysis. Here, we report the creation of a bifunctional artificial metalloenzyme in which a glutamic acid or aspartic acid residue engineered into streptavidin acts in concert with a docked biotinylated rhodium(III) complex to enable catalytic asymmetric carbon-hydrogen (C-H) activation. The coupling of benzamides and alkenes to access dihydroisoquinolones proceeds with up to nearly a 100-fold rate acceleration compared with the activity of the isolated rhodium complex and enantiomeric ratios as high as 93:7.

Antagonizing STAT3 dimerization with a rhodium(III) complex.

PubMed

Ma, Dik-Lung; Liu, Li-Juan; Leung, Ka-Ho; Chen, Yen-Ting; Zhong, Hai-Jing; Chan, Daniel Shiu-Hin; Wang, Hui-Min David; Leung, Chung-Hang

2014-08-25

Kinetically inert metal complexes have arisen as promising alternatives to existing platinum and ruthenium chemotherapeutics. Reported herein, to our knowledge, is the first example of a substitutionally inert, Group 9 organometallic compound as a direct inhibitor of signal transducer and activator of transcription 3 (STAT3) dimerization. From a series of cyclometalated rhodium(III) and iridium(III) complexes, a rhodium(III) complex emerged as a potent inhibitor of STAT3 that targeted the SH2 domain and inhibited STAT3 phosphorylation and dimerization. Significantly, the complex exhibited potent anti-tumor activities in an in vivo mouse xenograft model of melanoma. This study demonstrates that rhodium complexes may be developed as effective STAT3 inhibitors with potent anti-tumor activity. © 2014 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Hairpin-shaped tetranuclear palladium(II) complex: synthesis, crystal structure, DNA binding and cytotoxicity activity studies.

PubMed

Gao, En-Jun; Wang, Ke-Hua; Zhu, Ming-Chang; Liu, Lei

2010-07-01

A novel tetranuclear palladium(II) complex [Pd(4)(phen)(4) (micro-pydc)(4)].10H(2)O (phen = 1,10-phenanthroline, pydc = pyridine-3,4-dicarboxylate) has been synthesized and characterized. In the tetranuclear complex, two pairs of dipalladated [Pd(phen)] moieties are bridged together by four pydc, presenting a hairpin molecular shape. The binding of the title complex with fish sperm DNA (FS-DNA) has been investigated by UV spectrum and fluorescence spectrum. All the results indicate that the complex bind to DNA in an intercalative mode and considerating the molecular shape and size, the dipalladated phenanthroline moieties bisintercalate to the base pairs of DNA. Agarose gel electrophoresis assay demonstrates the ability of the complex to cleave the pBR322 plasmid DNA. Cytotoxic activity studies show the complex exhibited good cytotoxic activity against four different cancer cell lines. Crown Copyright (c) 2010. Published by Elsevier Masson SAS. All rights reserved.

Brilliant Sm, Eu, Tb, and Dy Chiral Lanthanide Complexes with Strong Circularly Polarized Luminescence

PubMed Central

Petoud, Stéphane; Muller, Gilles; Moore, Evan G.; Xu, Jide; Sokolnicki, Jurek; Riehl, James P.; Le, Uyen N.; Cohen, Seth M.; Raymond, Kenneth N.

2009-01-01

The synthesis, characterization, and luminescent behavior of trivalent Sm, Eu, Dy, and Tb complexes of two enantiomeric, octadentate, chiral, 2-hydroxyisophthalamide ligands are reported. These complexes are highly luminescent in solution. Functionalization of the achiral parent ligand with a chiral 1-phenylethylamine substituent on the open face of the complex in close proximity to the metal center yields complexes with strong circularly polarized luminescence (CPL) activity. This appears to be the first example of a system utilizing the same ligand architecture to sensitize four different lanthanide cations and display CPL activity. The luminescence dissymmetry factor, glum, recorded for the Eu(III) complex is one of the highest values reported, and this is the first time the CPL effect has been demonstrated for a Sm(III) complex with a chiral ligand. The combination of high luminescence intensity with CPL activity should enable new bioanalytical applications of macromolecules in chiral environments. PMID:17199285

Complex-valued multistate associative memory with nonlinear multilevel functions for gray-level image reconstruction.

PubMed

Tanaka, Gouhei; Aihara, Kazuyuki

2009-09-01

A widely used complex-valued activation function for complex-valued multistate Hopfield networks is revealed to be essentially based on a multilevel step function. By replacing the multilevel step function with other multilevel characteristics, we present two alternative complex-valued activation functions. One is based on a multilevel sigmoid function, while the other on a characteristic of a multistate bifurcating neuron. Numerical experiments show that both modifications to the complex-valued activation function bring about improvements in network performance for a multistate associative memory. The advantage of the proposed networks over the complex-valued Hopfield networks with the multilevel step function is more outstanding when a complex-valued neuron represents a larger number of multivalued states. Further, the performance of the proposed networks in reconstructing noisy 256 gray-level images is demonstrated in comparison with other recent associative memories to clarify their advantages and disadvantages.

Synthesis and spectroscopic studies of biologically active compounds derived from oxalyldihydrazide and benzil, and their Cr(III), Fe(III) and Mn(III) complexes.

PubMed

Singh, D P; Kumar, Ramesh; Singh, Jitender

2009-04-01

A new series of complexes have been synthesized by template condensation of oxalyldihydrazide and benzil in methanolic medium in the presence of trivalent chromium, manganese and iron salts forming complexes of the type [M(C(32)H(24)N(8)O(4))X]X(2) where M = Cr(III), Mn(III), Fe(III) and X = Cl(-1), NO(3)(-1), CH(3)COO(-1). The complexes have been characterized with the help of elemental analyses, conductance measurements, magnetic susceptibility measurements, electronic, NMR, infrared and far infrared spectral studies. On the basis of these studies, a five coordinate square pyramidal geometry has been proposed for all these complexes. The biological activities of the metal complexes have been tested in vitro against a number of pathogenic bacteria to assess their inhibiting potential. Some of these complexes have been found to exhibit remarkable antibacterial activities.

Development of nanoscale structure in LAT-based signaling complexes

PubMed Central

2016-01-01

ABSTRACT The adapter molecule linker for activation of T cells (LAT) plays a crucial role in forming signaling complexes induced by stimulation of the T cell receptor (TCR). These multi-molecular complexes are dynamic structures that activate highly regulated signaling pathways. Previously, we have demonstrated nanoscale structure in LAT-based complexes where the adapter SLP-76 (also known as LCP2) localizes to the periphery of LAT clusters. In this study, we show that initially LAT and SLP-76 are randomly dispersed throughout the clusters that form upon TCR engagement. The segregation of LAT and SLP-76 develops near the end of the spreading process. The local concentration of LAT also increases at the same time. Both changes require TCR activation and an intact actin cytoskeleton. These results demonstrate that the nanoscale organization of LAT-based signaling complexes is dynamic and indicates that different kinds of LAT-based complexes appear at different times during T cell activation. PMID:27875277

Tiam1 interaction with the PAR complex promotes talin-mediated Rac1 activation during polarized cell migration

PubMed Central

Wang, Shujie; Watanabe, Takashi; Matsuzawa, Kenji; Katsumi, Akira; Kakeno, Mai; Matsui, Toshinori; Ye, Feng; Sato, Kazuhide; Murase, Kiyoko; Sugiyama, Ikuko; Kimura, Kazushi; Mizoguchi, Akira; Ginsberg, Mark H.; Collard, John G.

2012-01-01

Migrating cells acquire front-rear polarity with a leading edge and a trailing tail for directional movement. The Rac exchange factor Tiam1 participates in polarized cell migration with the PAR complex of PAR3, PAR6, and atypical protein kinase C. However, it remains largely unknown how Tiam1 is regulated and contributes to the establishment of polarity in migrating cells. We show here that Tiam1 interacts directly with talin, which binds and activates integrins to mediate their signaling. Tiam1 accumulated at adhesions in a manner dependent on talin and the PAR complex. The interactions of talin with Tiam1 and the PAR complex were required for adhesion-induced Rac1 activation, cell spreading, and migration toward integrin substrates. Furthermore, Tiam1 acted with talin to regulate adhesion turnover. Thus, we propose that Tiam1, with the PAR complex, binds to integrins through talin and, together with the PAR complex, thereby regulates Rac1 activity and adhesion turnover for polarized migration. PMID:23071154

Anti-trypanosomal activity of cationic N-heterocyclic carbene gold(I) complexes.

PubMed

Winter, Isabel; Lockhauserbäumer, Julia; Lallinger-Kube, Gertrud; Schobert, Rainer; Ersfeld, Klaus; Biersack, Bernhard

2017-06-01

Two gold(I) N-heterocyclic carbene complexes 1a and 1b were tested for their anti-trypanosomal activity against Trypanosoma brucei parasites. Both gold compounds exhibited excellent anti-trypanosomal activity (IC 50 =0.9-3.0nM). The effects of the gold complexes 1a and 1b on the T. b. brucei cytoskeleton were evaluated. Rapid detachment of the flagellum from the cell body occurred after treatment with the gold complexes. In addition, a quick and complete degeneration of the parasitic cytoskeleton was induced by the gold complexes, only the microtubules of the detached flagellum remained intact. Both gold compounds 1a and 1b feature selective anti-trypanosomal agents and were distinctly more active against T. b. brucei cells than against human HeLa cells. Thus, the gold complexes 1a and 1b feature promising drug candidates for the treatment of trypanosome infections such as sleeping sickness (human African Trypanosomiasis caused by Trypanosoma brucei parasites). Copyright © 2017 Elsevier B.V. All rights reserved.

Syntheses of new rare earth complexes with carboxymethylated polysaccharides and evaluation of their in vitro antifungal activities.

PubMed

Sun, Xiaobo; Jin, Xiaozhe; Pan, Wei; Wang, Jinping

2014-11-26

In the present paper, La, Eu and Yb were selected to represent light, middle and heavy rare earths to form complexes with polysaccharides through chelating coordination of carboxyl groups, which were added into polysaccharide chains by means of carboxymethylation. Their antifungal activities against plant pathogenic fungi were evaluated using growth rate method. These rare earth complexes exhibited various antifungal activities against the tested fungi, depending on rare earth elements, polysaccharide types and fungal species. Among these three metal elements (i.e. La, Eu and Yb), Yb formed the complexes with the most effective antifungal properties. Furthermore, the results showed that ligands of carboxymethylated polysaccharides played a key role in promoting cytotoxicity of the rare earth complexes. Carboxymethylated Ganoderma applanatum polysaccharide (CGAP) was found to be the most effective ligand to form complexes with antifungal activities, followed by carboxymethylated lentinan (CLNT) and carboxymethylated Momordica charantia polysaccharide (CMCP). Copyright © 2014 Elsevier Ltd. All rights reserved.

Structure of the active form of human origin recognition complex and its ATPase motor module

DOE Office of Scientific and Technical Information (OSTI.GOV)

Tocilj, Ante; On, Kin Fan; Yuan, Zuanning

Binding of the Origin Recognition Complex (ORC) to origins of replication marks the first step in the initiation of replication of the genome in all eukaryotic cells. Here, we report the structure of the active form of human ORC determined by X-ray crystallography and cryo-electron microscopy. The complex is composed of an ORC1/4/5 motor module lobe in an organization reminiscent of the DNA polymerase clamp loader complexes. A second lobe contains the ORC2/3 subunits. The complex is organized as a double-layered shallow corkscrew, with the AAA+ and AAA+-like domains forming one layer, and the winged-helix domains (WHDs) forming a topmore » layer. CDC6 fits easily between ORC1 and ORC2, completing the ring and the DNA-binding channel, forming an additional ATP hydrolysis site. Analysis of the ATPase activity of the complex provides a basis for understanding ORC activity as well as molecular defects observed in Meier-Gorlin Syndrome mutations.« less

Mitochondrial Dysfunction in Schizophrenia: Determination of Mitochondrial Respiratory Activity in a Two-Hit Mouse Model.

PubMed

Monpays, Cécile; Deslauriers, Jessica; Sarret, Philippe; Grignon, Sylvain

2016-08-01

Schizophrenia is a chronic mental illness in which mitochondrial dysfunction has been suggested. Our laboratory recently developed a juvenile murine two-hit model (THM) of schizophrenia based on the combination of gestational inflammation, followed by juvenile restraint stress. We previously reported that relevant behaviors and neurochemical disturbances, including oxidative stress, were reversed by the antioxidant lipoic acid (LA), thereby pointing to the central role played by oxidative abnormalities and prompting us to investigate mitochondrial function. Mitochondrial activity was determined with the MitoXpress® commercial kit in two schizophrenia-relevant regions (prefrontal cortex (PFC) and striatum). Measurements were performed in state 3, with substrates for complex I- and complex II-induced respiratory activity (IRA). We observed an increase in complex I IRA in the PFC and striatum in both sexes but an increase in complex II activity only in males. LA treatment prevented this increase only in complex II IRA in males. Expression levels of the different respiratory chain complexes, as well as fission/fusion proteins and protein carbonylation, were unchanged. In conclusion, our juvenile schizophrenia THM shows an increase in mitochondrial activity reversed by LA, specifically in complex II IRA in males. Further investigations are required to determine the mechanisms of these modifications.

IN VITRO AND IN VIVO ACTIVITY OF A LYMPHOCYTE AND IMMUNE COMPLEX-DEPENDENT CHEMOTACTIC FACTOR FOR EOSINOPHILS

PubMed Central

Cohen, Stanley; Ward, Peter A.

1971-01-01

When cultured in the presence of specific antigen, lymphocytes from delayed-hypersensitive guinea pigs release a number of biologically active substances into the culture medium. Such active supernatants can react with immune complexes in vitro to generate a factor which is chemotactic for eosinophils. The factor involved is unique, since previously described chemotactic factors for other cell types require for their generation either immune complexes or substances released into lymphocyte culture, but not both. In the case of the eosinophil chemotactic factor, the interaction between the substance elaborated by the lymphocytes and the immune complexes appears to be specific in that the immune complexes must contain the same antigen as that used to activate the lymphocyte cultures. Although this factor was generated in an in vitro system, it has been shown to possess in vivo as well as in vitro activity. It is therefore possible that this factor may be of biological significance in situations where eosinophils are participants in inflammatory or immunologic reactions. PMID:5099667

From Synthesis to Biological Impact of Pd (II) Complexes: Synthesis, Characterization, and Antimicrobial and Scavenging Activity

PubMed Central

Sharma, Nitin Kumar; Ameta, Rakesh Kumar; Singh, Man

2016-01-01

The Pd (II) complexes with a series of halosubstituted benzylamine ligands (BLs) have been synthesized and characterized with different spectroscopic technique such as FTIR, UV/Vis, LCMS, 1H, and 13C NMR. Their molecular sustainability in different solvents such as DMSO, DMSO : H2O, and DMSO : PBS at physiological condition (pH 7.2) was determined by UV/Vis spectrophotometer. The in vitro antibacterial and antifungal activities of the complexes were investigated against Gram-positive and Gram-negative microbes and two different fungi indicated their significant biological potential. Additionally, their antioxidant activity has been analyzed with DPPH• free radical through spectrophotometric method and the result inferred them as an antioxidant. The stronger antibacterial and antioxidant activities of the synthesized complexes suggested them as a stronger antimicrobial agent. Our study advances the biological importance of palladium (II) amine complexes in the field of antimicrobial and antioxidant activities. PMID:27119023

Structural characterization and antimicrobial activities of transition metal complexes of a hydrazone ligand

NASA Astrophysics Data System (ADS)

Bakale, Raghavendra P.; Naik, Ganesh N.; Machakanur, Shrinath S.; Mangannavar, Chandrashekhar V.; Muchchandi, Iranna S.; Gudasi, Kalagouda B.

2018-02-01

A hydrazone ligand has been synthesized by the condensation of 2-nitrobenzaldehyde and hydralazine, and its Co(II), Ni(II), Cu(II) and Zn(II) complexes have been reported. Structural characterization of the ligand and its metal complexes has been performed by various spectroscopic [IR, NMR, UV-Vis, Mass], thermal and other physicochemical methods. The structure of the ligand and its Ni(II) complex has been characterized by single crystal X-ray diffraction studies. All the synthesized compounds have been screened for in vitro antimicrobial activity. The antibacterial activity is tested against Gram-positive strains Enterococcus faecalis, Streptococcus mutans and Staphylococcus aureus and Gram-negative strains Escherichia coli, Pseudomonas aeruginosa and Klebsiella pneumoniae using ciprofloxacin as the reference standard. Antifungal activity is tested against Candida albicans, Aspergillus fumigatus and Aspergillus niger using ketoconazole as the reference standard. The minimum inhibitory concentration (MIC) was determined for test compounds as well as for reference standard. Ligand, Cu(II) and Zn(II) complexes have shown excellent activity against Candida albicans.

Reactivity of dinuclear copper(II) complexes towards melanoma cells: Correlation with its stability, tyrosinase mimicking and nuclease activity.

PubMed

Nunes, Cléia Justino; Borges, Beatriz Essenfelder; Nakao, Lia Sumie; Peyroux, Eugénie; Hardré, Renaud; Faure, Bruno; Réglier, Marius; Giorgi, Michel; Prieto, Marcela Bach; Oliveira, Carla Columbano; Da Costa Ferreira, Ana M

2015-08-01

In this work, the influence of two new dinuclear copper(II) complexes in the viability of melanoma cells (B16F10 and TM1MNG3) was investigated, with the aim of verifying possible correlations between their cytotoxicity and their structure. One of the complexes had a polydentate dinucleating amine-imine ligand (complex 2), and the other a tridentate imine and a diamine-bridging ligand (complex 4). The analogous mononuclear copper(II) species (complexes 1 and 3, respectively) were also prepared for comparative studies. Crystal structure determination of complex 2 indicated a square-based pyramidal geometry around each copper, coordinated to three N atoms from the ligand and the remaining sites being occupied by either solvent molecules or counter-ions. Complex 4 has a tetragonal geometry. Interactions of these complexes with human albumin protein (HSA) allowed an estimation of their relative stabilities. Complementary studies of their reactivity towards DNA indicated that all of them are able of causing significant oxidative damage, with single and double strand cleavages, in the presence of hydrogen peroxide. However, nuclease activity of the dinuclear species was very similar and much higher than that of the corresponding mononuclear compounds. Although complex 2, with a more flexible structure, exhibits a much higher tyrosinase activity than complex 4, having a more rigid environment around the metal ion, both complexes showed comparable cytotoxicity towards melanoma cells. Corresponding mononuclear complexes showed to be remarkably less reactive as tyrosinase mimics as well as cytotoxic agents. Moreover, the dinuclear complexes showed higher cytotoxicity towards more melanogenic cells. The obtained results indicated that the structure of these species is decisive for its activity towards the malignant tumor cells tested. Copyright © 2015 Elsevier Inc. All rights reserved.

Neural Responses to Complex Auditory Rhythms: The Role of Attending

PubMed Central

Chapin, Heather L.; Zanto, Theodore; Jantzen, Kelly J.; Kelso, Scott J. A.; Steinberg, Fred; Large, Edward W.

2010-01-01

The aim of this study was to explore the role of attention in pulse and meter perception using complex rhythms. We used a selective attention paradigm in which participants attended to either a complex auditory rhythm or a visually presented word list. Performance on a reproduction task was used to gauge whether participants were attending to the appropriate stimulus. We hypothesized that attention to complex rhythms – which contain no energy at the pulse frequency – would lead to activations in motor areas involved in pulse perception. Moreover, because multiple repetitions of a complex rhythm are needed to perceive a pulse, activations in pulse-related areas would be seen only after sufficient time had elapsed for pulse perception to develop. Selective attention was also expected to modulate activity in sensory areas specific to the modality. We found that selective attention to rhythms led to increased BOLD responses in basal ganglia, and basal ganglia activity was observed only after the rhythms had cycled enough times for a stable pulse percept to develop. These observations suggest that attention is needed to recruit motor activations associated with the perception of pulse in complex rhythms. Moreover, attention to the auditory stimulus enhanced activity in an attentional sensory network including primary auditory cortex, insula, anterior cingulate, and prefrontal cortex, and suppressed activity in sensory areas associated with attending to the visual stimulus. PMID:21833279

Selective ligand activity at Nur/retinoid X receptor complexes revealed by dimer-specific bioluminescence resonance energy transfer-based sensors

PubMed Central

Giner, Xavier C; Cotnoir-White, David; Mader, Sylvie; Lévesque, Daniel

2017-01-01

Retinoid X receptors (RXR) play a role as master regulators due to their capacity to form heterodimers with other nuclear receptors. Accordingly, retinoid signaling is involved in multiple biological processes, including development, cell differentiation, metabolism and cell death. However, the role and functions of RXR in different heterodimer complexes remain unsolved, mainly because most RXR drugs (called rexinoids) are not selective to specific heterodimer complexes. This also strongly limits the use of rexinoids for specific therapeutic approaches. In order to better characterize rexinoids at specific nuclear receptor complexes, we have developed and optimized luciferase protein complementation-based Bioluminescence Resonance Energy Transfer (BRET) assays, which can directly measure recruitment of a co-activator motif fused to yellow fluorescent protein (YFP) by specific nuclear receptor dimers. To validate the assays, we compared rexinoid modulation of co-activator recruitment by RXR homodimer, and heterodimers Nur77/RXR and Nurr1/RXR. Results reveal that some rexinoids display selective co-activator recruitment activities with homo- or hetero-dimer complexes. In particular, SR11237 (BMS649) has increased potency for recruitment of co-activator motif and transcriptional activity with the Nur77/RXR heterodimer compared to other complexes. This technology should prove useful to identify new compounds with specificity for individual dimeric species formed by nuclear receptors. PMID:26148973

Mitochondrial phenotype during torpor: Modulation of mitochondrial electron transport system in the Chilean mouse-opossum Thylamys elegans.

PubMed

Cortes, Pablo A; Bozinovic, Francisco; Blier, Pierre U

2018-07-01

Mammalian torpor is a phenotype characterized by a controlled decline of metabolic rate, generally followed by a reduction in body temperature. During arousal from torpor, both metabolic rate and body temperature rapidly returns to resting levels. Metabolic rate reduction experienced by torpid animals is triggered by active suppression of mitochondrial respiration, which is rapidly reversed during rewarming process. In this study, we analyzed the changes in the maximal activity of key enzymes related to electron transport system (complexes I, III and IV) in six tissues of torpid, arousing and euthermic Chilean mouse-opossums (Thylamys elegans). We observed higher maximal activities of complexes I and IV during torpor in brain, heart and liver, the most metabolically active organs in mammals. On the contrary, higher enzymatic activities of complexes III were observed during torpor in kidneys and lungs. Moreover, skeletal muscle was the only tissue without significant differences among stages in all complexes evaluated, suggesting no modulation of oxidative capacities of electron transport system components in this thermogenic tissue. In overall, our data suggest that complexes I and IV activity plays a major role in initiation and maintenance of metabolic suppression during torpor in Chilean mouse-opossum, whereas improvement of oxidative capacities in complex III might be critical to sustain metabolic machinery in organs that remains metabolically active during torpor. Copyright © 2018 Elsevier Inc. All rights reserved.

Alterations in mitochondrial DNA copy number and the activities of electron transport chain complexes and pyruvate dehydrogenase in the frontal cortex from subjects with autism

PubMed Central

Gu, F; Chauhan, V; Kaur, K; Brown, W T; LaFauci, G; Wegiel, J; Chauhan, A

2013-01-01

Autism is a neurodevelopmental disorder associated with social deficits and behavioral abnormalities. Recent evidence suggests that mitochondrial dysfunction and oxidative stress may contribute to the etiology of autism. This is the first study to compare the activities of mitochondrial electron transport chain (ETC) complexes (I–V) and pyruvate dehydrogenase (PDH), as well as mitochondrial DNA (mtDNA) copy number in the frontal cortex tissues from autistic and age-matched control subjects. The activities of complexes I, V and PDH were most affected in autism (n=14) being significantly reduced by 31%, 36% and 35%, respectively. When 99% confidence interval (CI) of control group was taken as a reference range, impaired activities of complexes I, III and V were observed in 43%, 29% and 43% of autistic subjects, respectively. Reduced activities of all five ETC complexes were observed in 14% of autistic cases, and the activities of multiple complexes were decreased in 29% of autistic subjects. These results suggest that defects in complexes I and III (sites of mitochondrial free radical generation) and complex V (adenosine triphosphate synthase) are more prevalent in autism. PDH activity was also reduced in 57% of autistic subjects. The ratios of mtDNA of three mitochondrial genes ND1, ND4 and Cyt B (that encode for subunits of complexes I and III) to nuclear DNA were significantly increased in autism, suggesting a higher mtDNA copy number in autism. Compared with the 95% CI of the control group, 44% of autistic children showed higher copy numbers of all three mitochondrial genes examined. Furthermore, ND4 and Cyt B deletions were observed in 44% and 33% of autistic children, respectively. This study indicates that autism is associated with mitochondrial dysfunction in the brain. PMID:24002085

The ‘active life’ of Hsp90 complexes☆

PubMed Central

Prodromou, Chrisostomos

2012-01-01

Hsp90 forms a variety of complexes differing both in clientele and co-chaperones. Central to the role of co-chaperones in the formation of Hsp90 complexes is the delivery of client proteins and the regulation of the ATPase activity of Hsp90. Determining the mechanisms by which co-chaperones regulate Hsp90 is essential in understanding the assembly of these complexes and the activation and maturation of Hsp90's clientele. Mechanistically, co-chaperones alter the kinetics of the ATP-coupled conformational changes of Hsp90. The structural changes leading to the formation of a catalytically active unit involve all regions of the Hsp90 dimer. Their complexity has allowed different orthologues of Hsp90 to evolve kinetically in slightly different ways. The interaction of the cytosolic Hsp90 with a variety of co-chaperones lends itself to a complex set of different regulatory mechanisms that modulate Hsp90's conformation and ATPase activity. It also appears that the conformational switches of Hsp90 are not necessarily coupled under all circumstances. Here, I described different co-chaperone complexes and then discuss in detail the mechanisms and role that specific co-chaperones play in this. I will also discuss emerging evidence that post-translational modifications also affect the ATPase activity of Hsp90, and thus complex formation. Finally, I will present evidence showing how Hsp90's active site, although being highly conserved, can be altered to show resistance to drug binding, but still maintain ATP binding and ATPase activity. Such changes are therefore unlikely to significantly alter Hsp90's interactions with client proteins and co-chaperones. This article is part of a Special Issue entitled: Heat Shock Protein 90 (HSP90) PMID:21840346

Synthesis, spectral, thermal and antimicrobial studies of transition metal complexes of 14-membered tetraaza[N4] macrocyclic ligand

NASA Astrophysics Data System (ADS)

Shankarwar, Sunil G.; Nagolkar, Bhagwat B.; Shelke, Vinod A.; Chondhekar, Trimbak K.

2015-06-01

A series of metal complexes of Mn(II), Co(II), Ni(II), Cu(II), have been synthesized with newly synthesized biologically active macrocyclic ligand. The ligand was synthesized by condensation of β-diketone 1-(4-chlorophenyl)-3-(2-hydroxyphenyl)propane-1,3-dione and o-phenylene diamine. All the complexes were characterized by elemental analysis, molar conductivity, magnetic susceptibility, thermal analysis, X-ray diffraction, IR, 1H-NMR, UV-Vis spectroscopy and mass spectroscopy. From the analytical data, stoichiometry of the complexes was found to be 1:2 (metal:ligand). Thermal behavior (TG/DTA) and kinetic parameters suggest more ordered activated state in complex formation. All the complexes are of high spin type and six coordinated. On the basis of IR, electronic spectral studies and magnetic behavior, an octahedral geometry has been assigned to these complexes. The antibacterial and antifungal activities of the ligand and its metal complexes, has been screened in vitro against Staphylococcus aureus, Escherichia coli and Aspergillus niger, Trichoderma respectively.

Effects of Preretirement Work Complexity and Postretirement Leisure Activity on Cognitive Aging

PubMed Central

Finkel, Deborah; Pedersen, Nancy L.

2016-01-01

Objectives: We examined the influence of postretirement leisure activity on longitudinal associations between work complexity in main lifetime occupation and trajectories of cognitive change before and after retirement. Methods: Information on complexity of work with data, people, and things, leisure activity participation in older adulthood, and four cognitive factors (verbal, spatial, memory, and speed) was available from 421 individuals in the longitudinal Swedish Adoption/Twin Study of Aging. Participants were followed for an average of 14.2 years (SD = 7.1 years) and up to 23 years across eight cognitive assessments. Most of the sample (88.6%) completed at least three cognitive assessments. Results: Results of growth curve analyses indicated that higher complexity of work with people significantly attenuated cognitive aging in verbal skills, memory, and speed of processing controlling for age, sex, and education. When leisure activity was added, greater cognitive and physical leisure activity was associated with reduced cognitive aging in verbal skills, speed of processing, and memory (for cognitive activity only). Discussion: Engagement in cognitive or physical leisure activities in older adulthood may compensate for cognitive disadvantage potentially imposed by working in occupations that offer fewer cognitive challenges. These results may provide a platform to encourage leisure activity participation in those retiring from less complex occupations. PMID:25975289

Regulation of NADH/CoQ oxidoreductase: do phosphorylation events affect activity?

PubMed

Maj, Mary C; Raha, Sandeep; Myint, Tomoko; Robinson, Brian H

2004-01-01

We had previously suggested that phosphorylation of proteins by mitochondrial kinases regulate the activity of NADH/CoQ oxidoreductase. Initial data showed that pyruvate dehydrogenase kinase (PDK) and cAMP-dependent protein kinase A (PKA) phosphorylate mitochondrial membrane proteins. Upon phosphorylation with crude PDK, mitochondria appeared to be deficient in NADH/cytochrome c reductase activity associated with increased superoxide production. Conversely, phosphorylation by PKA resulted in increased NADH/cytochrome c reductase activity and decreased superoxide formation. Current data confirms PKA involvement in regulating Complex I activity through phosphorylation of an 18 kDa subunit. Beef heart NADH/ cytochrome c reductase activity increases to 150% of control upon incubation with PKA and ATP-gamma-S. We have cloned the four human isoforms of PDK and purified beef heart Complex I. Incubation of mitochondria with PDK isoforms and ATP did not alter Complex I activity or superoxide production. Radiolabeling of mitochondria and purified Complex I with PDK failed to reveal phosphorylated proteins.

Mitochondrial markers predict recurrence, metastasis and tamoxifen-resistance in breast cancer patients: Early detection of treatment failure with companion diagnostics.

PubMed

Sotgia, Federica; Fiorillo, Marco; Lisanti, Michael P

2017-09-15

Here, we used a data-mining and informatics approach to discover new biomarkers of resistance to hormonal therapy in breast cancer. More specifically, we investigated whether nuclear-encoded genes associated with mitochondrial biogenesis can be used to predict tumor recurrence, distant metastasis and treatment failure in high-risk breast cancer patients. Overall, this strategy allowed us to directly provide in silico validation of the prognostic value of these mitochondrial components in large and clinically relevant patient populations, with >15 years of follow-up data. For this purpose, we employed a group of 145 ER(+) luminal A breast cancer patients, with lymph-node (LN) metastasis at diagnosis, that were treated with tamoxifen, but not any chemotherapy agents. Using this approach, we identified >60 new individual mitochondrial biomarkers that predicted treatment failure and tumor recurrence, with hazard-ratios (HR) of up to 4.17 ( p =2.2e-07). These include mitochondrial chaperones (HSPD1, HSPA9), membrane proteins (VDAC2, TOMM70A) and anti-oxidants (SOD2), as well as 18 different mitochondrial ribosomal proteins (MRPs) and >20 distinct components of the OXPHOS complexes. In addition, we combined 4 mitochondrial proteins (HSPD1, UQCRB, MRPL15, COX17), to generate a compact mitochondrial gene signature, associated with a HR of 5.34 ( p =1e-09). This signature also successfully predicted distant metastasis and was effective in larger groups of ER(+) ( N =2,447), basal ( N =540) and HER2(+) ( N =193) breast cancers. It was also effective in all breast cancers ( N =3,180), if considered together as a single group. Based on this analysis, we conclude that mitochondrial biogenesis should be considered as a new therapeutic target for overcoming tumor recurrence, distant metastasis and treatment failure in patients with breast cancer. In summary, we identified individual mitochondrial biomarkers and 2 compact mitochondrial gene signatures that can be used to predict tamoxifen-resistance and tumor recurrence, at their initial diagnosis, in patients with advanced breast cancer. In the long-term, these mitochondrial biomarkers could provide a new companion diagnostics platform to help clinicians to accurately predict the response to hormonal therapy in ER(+) breast cancer patients, facilitating more personalized and effective treatment. Similarly, these mitochondrial markers could be used as companion diagnostics, to determine which breast cancer patients would benefit most from clinical treatments with mitochondrially-targeted anti-cancer therapeutics. Finally, we also showed that these mitochondrial markers are superior when directly compared with conventional biomarkers, such as Ki67 and PCNA.

Gold(I)-Triphenylphosphine Complexes with Hypoxanthine-Derived Ligands: In Vitro Evaluations of Anticancer and Anti-Inflammatory Activities

PubMed Central

Křikavová, Radka; Hošek, Jan; Vančo, Ján; Hutyra, Jakub; Dvořák, Zdeněk; Trávníček, Zdeněk

2014-01-01

A series of gold(I) complexes involving triphenylphosphine (PPh3) and one N-donor ligand derived from deprotonated mono- or disubstituted hypoxanthine (HLn) of the general composition [Au(Ln)(PPh3)] (1–9) is reported. The complexes were thoroughly characterized, including multinuclear high resolution NMR spectroscopy as well as single crystal X-ray analysis (for complexes 1 and 3). The complexes were screened for their in vitro cytotoxicity against human cancer cell lines MCF7 (breast carcinoma), HOS (osteosarcoma) and THP-1 (monocytic leukaemia), which identified the complexes 4–6 as the most promising representatives, who antiproliferative activity was further tested against A549 (lung adenocarcinoma), G-361 (melanoma), HeLa (cervical cancer), A2780 (ovarian carcinoma), A2780R (ovarian carcinoma resistant to cisplatin), 22Rv1 (prostate cancer) cell lines. Complexes 4–6 showed a significantly higher in vitro anticancer effect against the employed cancer cells, except for G-361, as compared with the commercially used anticancer drug cisplatin, with IC50 ≈ 1–30 µM. Anti-inflammatory activity was evaluated in vitro by the assessment of the ability of the complexes to modulate secretion of the pro-inflammatory cytokines, i.e. tumour necrosis factor-α (TNF-α) and interleukin-1β (IL-1β), in the lipopolysaccharide-activated macrophage-like THP-1 cell model. The results of this study identified the complexes as auspicious anti-inflammatory agents with similar or better activity as compared with the clinically applied gold-based antiarthritic drug Auranofin. In an effort to explore the possible mechanisms responsible for the biological effect, the products of interactions of selected complexes with sulfur-containing biomolecules (L-cysteine and reduced glutathione) were studied by means of the mass-spectrometry study. PMID:25226034

Spectral characterization, cyclic voltammetry, morphology, biological activities and DNA cleaving studies of amino acid Schiff base metal(II) complexes

NASA Astrophysics Data System (ADS)

Neelakantan, M. A.; Rusalraj, F.; Dharmaraja, J.; Johnsonraja, S.; Jeyakumar, T.; Sankaranarayana Pillai, M.

2008-12-01

Metal complexes are synthesized with Schiff bases derived from o-phthalaldehyde (opa) and amino acids viz., glycine (gly) L-alanine (ala), L-phenylalanine (pal). Metal ions coordinate in a tetradentate or hexadentate manner with these N 2O 2 donor ligands, which are characterized by elemental analysis, molar conductance, magnetic moments, IR, electronic, 1H NMR and EPR spectral studies. The elemental analysis suggests the stoichiometry to be 1:1 (metal:ligand). Based on EPR studies, spin-Hamiltonian and bonding parameters have been calculated. The g-values calculated for copper complexes at 300 K and in frozen DMSO (77 K) indicate the presence of the unpaired electron in the d orbital. The evaluated metal-ligand bonding parameters showed strong in-plane σ- and π-bonding. X-ray diffraction (XRD) and scanning electron micrography (SEM) analysis provide the crystalline nature and the morphology of the metal complexes. The cyclic voltammograms of the Cu(II)/Mn(II)/VO(II) complexes investigated in DMSO solution exhibit metal centered electroactivity in the potential range -1.5 to +1.5 V. The electrochemical data obtained for Cu(II) complexes explains the change of structural arrangement of the ligand around Cu(II) ions. The biological activity of the complexes has been tested on eight bacteria and three fungi. Cu(II) and Ni(II) complexes show an increased activity in comparison to the controls. The metal complexes of opapal Schiff base were evaluated for their DNA cleaving activities with calf-thymus DNA (CT DNA) under aerobic conditions. Cu(II) and VO(II) complexes show more pronounced activity in presence of the oxidant.

On the nature of carbon-hydrogen bond activation at rhodium and related reactions.

PubMed

Jones, William D

2005-06-27

Over the past 20 years, substantial progress has been made in the understanding of the activation of C-H and other strong bonds by reactive metal complexes in low oxidation states. This paper will present an overview of the use of pentamethylcyclopentadienyl and trispyrazolylborate rhodium complexes for the activation of arene and alkane C-H bonds. Insights into bond strengths, kinetic and thermodynamic selectivities, and the nature of the intermediates involved will be reviewed. The role of eta-2 arene complexes will be shown to be critical to the C-H activation reactions. Some information about the fleeting alkane sigma-complexes will also be presented. In addition, use of these complexes with thiophenes has shown the ability to cleave C-S bonds. Mechanistic information has been obtained indicating coordination through sulfur prior to cleavage. Relevant examples of nickel-based C-S cleavage will also be given.

Mice Lacking TR4 Nuclear Receptor Develop Mitochondrial Myopathy with Deficiency in Complex I

PubMed Central

Liu, Su; Lee, Yi-Fen; Chou, Samuel; Uno, Hideo; Li, Gonghui; Brookes, Paul; Massett, Michael P.; Wu, Qiao; Chen, Lu-Min

2011-01-01

The estimated incidence of mitochondrial diseases in humans is approximately 1:5000 to 1:10,000, whereas the molecular mechanisms for more than 50% of human mitochondrial disease cases still remain unclear. Here we report that mice lacking testicular nuclear receptor 4 (TR4−/−) suffered mitochondrial myopathy, and histological examination of TR4−/− soleus muscle revealed abnormal mitochondrial accumulation. In addition, increased serum lactate levels, decreased mitochondrial ATP production, and decreased electron transport chain complex I activity were found in TR4−/− mice. Restoration of TR4 into TR4−/− myoblasts rescued mitochondrial ATP generation capacity and complex I activity. Further real-time PCR quantification and promoter studies found TR4 could modulate complex I activity via transcriptionally regulating the complex I assembly factor NDUFAF1, and restoration of NDUFAF1 level in TR4−/− myoblasts increased mitochondrial ATP generation capacity and complex I activity. Together, these results suggest that TR4 plays vital roles in mitochondrial function, which may help us to better understand the pathogenesis of mitochondrial myopathy, and targeting TR4 via its ligands/activators may allow us to develop better therapeutic approaches. PMID:21622535

Comparing anti-hyperglycemic activity and acute oral toxicity of three different trivalent chromium complexes in mice.

PubMed

Li, Fang; Wu, Xiangyang; Zou, Yanmin; Zhao, Ting; Zhang, Min; Feng, Weiwei; Yang, Liuqing

2012-05-01

Three different ligands (rutin, folate and stachyose) of chromium(III) complexes were compared to examine whether they have similar effect on anti-hyperglycemic activity as well as the acute toxicity status. Anti-hyperglycemic activities of chromium rutin complex (CrRC), chromium folate complex (CrFC) and chromium stachyose complex (CrSC) were examined in alloxan-induced diabetic mice with daily oral gavage for a period of 2 weeks at the dose of 0.5-3.0 mg Cr/kg. Acute toxicities of CrRC and CrFC were tested using ICR mice at the dose of 1.0-5.0 g/kg with a single oral gavage and observed for a period of 2 weeks. Biological activities results indicated that only CrRC and CrFC could decrease blood glucose level, reduce the activities of aspartate transaminase, alanine transaminase, alkaline phosphatase, and increase liver glycogen level. In acute toxicity study, LD(50) values for both CrRC and CrFC were above 5.0 g/kg. The minimum lethal dose for CrFC was above 5.0 g/kg, while that for CrRC was 1.0 g/kg. Anti-diabetic activity of those chromium complexes was not similar and their acute toxicities were also different. CrFC represent an optimal chromium supplement among those chromium complexes with potential therapeutic value to control blood glucose in diabetes and non-toxicity in acute toxicity. Copyright © 2012 Elsevier Ltd. All rights reserved.

Activation of Hsp90/NOS and increased NO generation does not impair mitochondrial respiratory chain by competitive binding at cytochrome C Oxidase in low oxygen concentrations

PubMed Central

Presley, Tennille; Vedam, Kaushik; Liu, Xiaoping; Zweier, Jay L.

2009-01-01

Nitric oxide (NO) is known to regulate mitochondrial respiration, especially during metabolic stress and disease, by nitrosation of the mitochondrial electron transport chain (ETC) complexes (irreversible) and by a competitive binding at O2 binding site of cytochrome c oxidase (CcO) in complex IV (reversible). In this study, by using bovine aortic endothelial cells, we demonstrate that the inhibitory effect of endogenously generated NO by nitric oxide synthase (NOS) activation, by either NOS stimulators or association with heat shock protein 90 (Hsp90), is significant only at high prevailing pO2 through nitrosation of mitochondrial ETC complexes, but it does not inhibit the respiration by competitive binding at CcO at very low pO2. ETC complexes activity measurements confirmed that significant reduction in complex IV activity was noticed at higher pO2, but it was unaffected at low pO2 in these cells. This was further extended to heat-shocked cells, where NOS was activated by the induction/activation of (Hsp90) through heat shock at an elevated temperature of 42°C. From these results, we conclude that the entire attenuation of respiration by endogenous NO is due to irreversible inhibition by nitrosation of ETC complexes but not through reversible inhibition by competing with O2 binding at CcO at complex IV. PMID:19412660

Subunits of ADA-two-A-containing (ATAC) or Spt-Ada-Gcn5-acetyltrasferase (SAGA) Coactivator Complexes Enhance the Acetyltransferase Activity of GCN5.

PubMed

Riss, Anne; Scheer, Elisabeth; Joint, Mathilde; Trowitzsch, Simon; Berger, Imre; Tora, László

2015-11-27

Histone acetyl transferases (HATs) play a crucial role in eukaryotes by regulating chromatin architecture and locus specific transcription. GCN5 (KAT2A) is a member of the GNAT (Gcn5-related N-acetyltransferase) family of HATs. In metazoans this enzyme is found in two functionally distinct coactivator complexes, SAGA (Spt Ada Gcn5 acetyltransferase) and ATAC (Ada Two A-containing). These two multiprotein complexes comprise complex-specific and shared subunits, which are organized in functional modules. The HAT module of ATAC is composed of GCN5, ADA2a, ADA3, and SGF29, whereas in the SAGA HAT module ADA2b is present instead of ADA2a. To better understand how the activity of human (h) hGCN5 is regulated in the two related, but different, HAT complexes we carried out in vitro HAT assays. We compared the activity of hGCN5 alone with its activity when it was part of purified recombinant hATAC or hSAGA HAT modules or endogenous hATAC or hSAGA complexes using histone tail peptides and full-length histones as substrates. We demonstrated that the subunit environment of the HAT complexes into which GCN5 incorporates determines the enhancement of GCN5 activity. On histone peptides we show that all the tested GCN5-containing complexes acetylate mainly histone H3K14. Our results suggest a stronger influence of ADA2b as compared with ADA2a on the activity of GCN5. However, the lysine acetylation specificity of GCN5 on histone tails or full-length histones was not changed when incorporated in the HAT modules of ATAC or SAGA complexes. Our results thus demonstrate that the catalytic activity of GCN5 is stimulated by subunits of the ADA2a- or ADA2b-containing HAT modules and is further increased by incorporation of the distinct HAT modules in the ATAC or SAGA holo-complexes. © 2015 by The American Society for Biochemistry and Molecular Biology, Inc.

Photo-activated luminescence sensor and method of detecting trichloroethylene and related volatile organochloride compounds

DOEpatents

Dinh, T.V.

1996-06-11

A sensor for detecting trichloroethylene and related volatile organochloride compounds uses a photo-activator that produces a photo-product complex with the contaminant. Characteristics of the light emitted from the complex will indicate the presence of the contaminant. A probe containing the photo-activator has an excitation light interface and a contaminant interface. One particular embodiment uses a porous membrane as the contaminant interface, so that the contaminant can migrate there through to the photo-activator and thereby form the complex. 23 figs.

Synthesis of trimethoprim metal complexes: Spectral, electrochemical, thermal, DNA-binding and surface morphology studies.

PubMed

Demirezen, Nihat; Tarınç, Derya; Polat, Duygu; Ceşme, Mustafa; Gölcü, Ayşegül; Tümer, Mehmet

2012-08-01

Complexes of trimethoprim (TMP), with Cu(II), Zn(II), Pt(II), Ru(III) and Fe(III) have been synthesized. Then, these complexes have been characterized by spectroscopic techniques involving UV-vis, IR, mass and (1)H NMR. CHN elemental analysis, electrochemical and thermal behavior of complexes have also been investigated. The electrochemical properties of all complexes have been investigated by cyclic voltammetry (CV) using glassy carbon electrode. The biological activity of the complexes has been evaluated by examining their ability to bind to calf-thymus DNA (CT DNA) with UV spectroscopy and cyclic voltammetry. UV studies of the interaction of the complexes with DNA have shown that these compounds can bind to CT DNA. The binding constants of the complexes with CT DNA have also been calculated. The cyclic voltammograms of the complexes in the presence of CT DNA have shown that the complexes can bind to CT DNA by both the intercalative and the electrostatic binding mode. The antimicrobial activity of these complexes has been evaluated against three Gram-positive and four Gram-negative bacteria. Antifungal activity against two different fungi has been evaluated and compared with the reference drug TMP. Almost all types of complexes show excellent activity against all type of bacteria and fungi. The morphology of the CT DNA, TMP, metal ions and metal complexes has been investigated by scanning electron microscopy (SEM). To get the SEM images, the interaction of compounds with CT DNA has been studied by means of differential pulse voltammetry (DPV) at CT DNA modified pencil graphite electrode (PGE). The decrease in intensity of the guanine oxidation signals has been used as an indicator for the interaction mechanism. Copyright © 2012 Elsevier B.V. All rights reserved.

Mechanism Underlying IκB Kinase Activation Mediated by the Linear Ubiquitin Chain Assembly Complex

PubMed Central

Fujita, Hiroaki; Akita, Mariko; Kato, Ryuichi; Sasaki, Yoshiteru; Wakatsuki, Soichi

2014-01-01

The linear ubiquitin chain assembly complex (LUBAC) ligase, consisting of HOIL-1L, HOIP, and SHARPIN, specifically generates linear polyubiquitin chains. LUBAC-mediated linear polyubiquitination has been implicated in NF-κB activation. NEMO, a component of the IκB kinase (IKK) complex, is a substrate of LUBAC, but the precise molecular mechanism underlying linear chain-mediated NF-κB activation has not been fully elucidated. Here, we demonstrate that linearly polyubiquitinated NEMO activates IKK more potently than unanchored linear chains. In mutational analyses based on the crystal structure of the complex between the HOIP NZF1 and NEMO CC2-LZ domains, which are involved in the HOIP-NEMO interaction, NEMO mutations that impaired linear ubiquitin recognition activity and prevented recognition by LUBAC synergistically suppressed signal-induced NF-κB activation. HOIP NZF1 bound to NEMO and ubiquitin simultaneously, and HOIP NZF1 mutants defective in interaction with either NEMO or ubiquitin could not restore signal-induced NF-κB activation. Furthermore, linear chain-mediated activation of IKK2 involved homotypic interaction of the IKK2 kinase domain. Collectively, these results demonstrate that linear polyubiquitination of NEMO plays crucial roles in IKK activation and that this modification involves the HOIP NZF1 domain and recognition of NEMO-conjugated linear ubiquitin chains by NEMO on another IKK complex. PMID:24469399

Chitosan-Copper (II) complex as antibacterial agent: synthesis, characterization and coordinating bond- activity correlation study

NASA Astrophysics Data System (ADS)

Mekahlia, S.; Bouzid, B.

2009-11-01

The antimicrobial activity of chitosan is unstable and sensitive to many factors such as molecular weight. Recent investigations showed that low molecular weight chitosan exhibited strong bactericidal activities compared to chitosan with high molecular weight. Since chitosan degradation can be caused by the coordinating bond, we attempt to synthesize and characterize the chitosan-Cu (II) complex, and thereafter study the coordinating bond effect on its antibacterial activity against Salmonella enteritidis. Seven chitosan-copper complexes with different copper contents were prepared and characterized by FT-IR, UV-vis, XRD and atomic absorption spectrophotometry (AAS). Results indicated that for chitosan-Cu (II) complexes with molar ratio close to 1:1, the inhibition rate reached 100%.

The antioxidant effects of complexes of tilapia fish skin collagen and different marine oligosaccharides

NASA Astrophysics Data System (ADS)

Ren, Shuwen; Li, Jing; Guan, Huashi

2010-12-01

An excess of reactive oxygen species (ROS) leads to a variety of chronic health problems. As potent antioxidants, marine bioactive extracts containing oligosaccharides and peptides have been extensively studied. Recently, there is a growing interest in protein-polysaccharide complexes because of their potential uses in pharmaceutical and food industries. However, only few studies are available on the antioxidant activities of such complexes, in terms of their ROS scavenging capability. In this study, we combined different marine oligosaccharides (isolated and purified) with collagen peptides derived from tilapia fish skin, and evaluated the antioxidant activity of the marine peptide-oligosaccharide complexes vis-à-vis the activity of their original component molecules. Biochemical and cellular assays were performed to measure the scavenging effects on 1, 1-diphenyl-2-picrylhydrazyl (DPPH), hydroxyl and superoxide radicals, and to evaluate the influences on the activities of superoxide dismutase (SOD), glutathione peroxidase (GSH-Px) and the level of malondialdehyde (MDA) in UV-induced photoaging models. The results indicated that the antioxidant activities of all the complexes were stronger than those of their individual components. Among the 11 complexes tested, two complexes, namely MA1000+CP and κ-ca3000+CP, turned out to be highly effective antioxidants. Although the detailed mechanisms of this improved scavenging ability are not fully understood, this work provides insights into the design of highly efficient peptide-oligosaccharide complexes for potential applications in pharmaceutical, cosmetics and food industries.

Ruthenium(II) Complexes with 2-Phenylimidazo[4,5-f][1,10]phenanthroline Derivatives that Strongly Combat Cisplatin-Resistant Tumor Cells

NASA Astrophysics Data System (ADS)

Zeng, Leli; Chen, Yu; Liu, Jiangping; Huang, Huaiyi; Guan, Ruilin; Ji, Liangnian; Chao, Hui

2016-01-01

Cisplatin was the first metal-based therapeutic agent approved for the treatment of human cancers, but its clinical activity is greatly limited by tumor drug resistance. This work utilized the parent complex [Ru(phen)2(PIP)]2+ (1) to develop three Ru(II) complexes (2-4) with different positional modifications. These compounds exhibited similar or superior cytotoxicities compared to cisplatin in HeLa, A549 and multidrug-resistant (A549R) tumor cell lines. Complex 4, the most potent member of the series, was highly active against A549R cancer cells (IC50 = 0.8 μM). This complex exhibited 178-fold better activity than cisplatin (IC50 = 142.5 μM) in A549R cells. 3D multicellular A549R tumor spheroids were also used to confirm the high proliferative and cytotoxic activity of complex 4. Complex 4 had the greatest cellular uptake and had a tendency to accumulate in the mitochondria of A549R cells. Further mechanistic studies showed that complex 4 induced A549R cell apoptosis via inhibition of thioredoxin reductase (TrxR), elevated intracellular ROS levels, mitochondrial dysfunction and cell cycle arrest, making it an outstanding candidate for overcoming cisplatin resistance.

N-((5-chloropyridin-2-yl)carbamothioyl)furan-2-carboxamide and its Co(II), Ni(II) and Cu(II) complexes: Synthesis, characterization, DFT computations, thermal decomposition, antioxidant and antitumor activity

NASA Astrophysics Data System (ADS)

Yeşilkaynak, Tuncay; Özpınar, Celal; Emen, Fatih Mehmet; Ateş, Burhan; Kaya, Kerem

2017-02-01

N-((5-chloropyridin-2-yl)carbamothioyl)furan-2-carboxamide (HL: C11H8ClN3O2S) and its Co(II), Ni(II) and Cu(II) complexes have been synthesized and characterized by elemental analysis, FT-IR,1H NMR and HR-MS methods. The HL was characterized by single crystal X-ray diffraction technique. It crystallizes in the monoclinic system. The HL has the space group P 1 21/c 1, Z = 4, and its unit cell parameters are a = 4.5437(5) Å, b = 22.4550(3) Å, c = 11.8947(14) Å. The ligand coordinates the metal ions as bidentate and thus essentially yields neutral complexes of the [ML2] type. ML2 complex structures were optimized using B97D/TZVP level. Molecular orbitals of both HL ligand were calculated at the same level. Thermal decomposition of the complexes has been investigated by thermogravimetry. The complexes were screened for their anticancer and antioxidant activities. Antioxidant activity of the complexes was determined by using the DPPH and ABTS assays. The anticancer activity of the complexes was studied by using MTT assay in MCF-7 breast cancer cells.

Enhanced anti-cancer activities of a gold(III) pyrrolidinedithiocarbamato complex incorporated in a biodegradable metal-organic framework.

PubMed

Sun, Raymond Wai-Yin; Zhang, Ming; Li, Dan; Li, Mian; Wong, Alice Sze-Tsai

2016-10-01

An anti-cancer active gold(III) pyrrolidinedithiocarbamato complex [(PDTC)Au III Cl 2 ] (1) has been synthesized and characterized by means of X-ray crystallography. Compared to the pyrrolidinedithiocarbamate ligand itself, this gold(III) complex displays an up to 33-fold higher anti-cancer potency towards a panel of cancer cell lines including the cisplatin-resistant ovarian carcinoma cell line (A2780cis). As demonstrated by a set of Transwell® assay-based cytotoxicity experiments, incorporating this gold(III) complex in a zinc-based biodegradable metal-organic framework (MOF) displays a significant enhancement in anti-cancer activity towards A2780cis than the gold(III) complex alone. Copyright © 2016 Elsevier Inc. All rights reserved.

High temperature ethylene polymerization catalyzed by titanium(IV) complexes with tetradentate aminophenolate ligands in cis-O, N, N chelating mode.

PubMed

Zhao, Ruiguo; Liu, Taotao; Wang, Liying; Ma, Haiyan

2014-09-07

A series of titanium trichloride complexes , ligated with claw-type tetradentate aminophenolate ligands were synthesized from the direct reaction of TiCl4(THF)2 with 1 equiv. of the corresponding aminophenol in the presence of triethylamine. For comparison purposes, titanium isopropoxide complexes were also synthesized via the reaction of Ti(O(i)Pr)4 and 1 equiv. of the proligand. Similar reactions of ZrCl4(THF)2 with the corresponding aminophenol ligands in the presence of triethylamine only allowed the isolation of zirconium complex . The X-ray diffraction studies reveal that titanium trichloride complexes , and titanium triisopropoxide complex all possess a distorted octahedral geometry with the tetradentate aminophenolate ligand in cis-O, N, N chelating mode, where the methoxy group of the aryl unit does not coordinate with the metal center in the solid state. Upon activation with MMAO, these titanium and zirconium(iv) complexes exhibited moderate to high catalytic activities for ethylene polymerization at 30-120 °C, producing high-molecular-weight polyethylenes with broad distributions (Mw/Mn = 10.2-34.8). The activities of titanium trichloride complexes are significantly higher than those of titanium isopropoxide and zirconium trichloride complexes at high temperatures. The highest activity of 15 456 kg (mol-Ti h)(-1) could be achieved by titanium trichloride complex with bromo groups on both ortho- and para-positions of the phenolate ring of the ligand at 120 °C.

Synthetic bioactive novel ether based Schiff bases and their copper(II) complexes

NASA Astrophysics Data System (ADS)

Shabbir, Muhammad; Akhter, Zareen; Ismail, Hammad; Mirza, Bushra

2017-10-01

Novel ether based Schiff bases (HL1- HL4) were synthesized from 5-chloro-2-hydroxy benzaldehyde and primary amines (1-amino-4-phenoxybenzene, 4-(4-aminophenyloxy) biphenyl, 1-(4-aminophenoxy) naphthalene and 2-(4-aminophenoxy) naphthalene). From these Schiff bases copper(II) complexes (Cu(L1)2-Cu(L4)2)) were synthesized and characterized by elemental analysis and spectroscopic (FTIR, NMR) techniques. The synthesized Schiff bases and copper(II) complexes were further assessed for various biological studies. In brine shrimp assay the copper(II) complexes revealed 4-fold higher activity (LD50 3.8 μg/ml) as compared with simple ligands (LD50 12.4 μg/ml). Similar findings were observed in potato disc antitumor assay with higher activities for copper(II) complexes (IC50 range 20.4-24.1 μg/ml) than ligands (IC50 range 40.5-48.3 μg/ml). DPPH assay was performed to determine the antioxidant potential of the compounds. Significant antioxidant activity was shown by the copper(II) complexes whereas simple ligands have shown no activity. In DNA protection assay significant protection behavior was exhibited by simple ligand molecules while copper(II) complexes showed neutral behavior (neither protective nor damaging).

Positional effects of hydroxy groups on catalytic activity of proton-responsive half-sandwich Cp*Iridium(III) complexes

DOE PAGES

Suna, Yuki; Fujita, Etsuko; Ertem, Mehmed Z.; ...

2014-11-12

Proton-responsive half-sandwich Cp*Ir(III) complexes possessing a bipyridine ligand with two hydroxy groups at the 3,3'-, 4,4'-, 5,5'- or 6,6'-positions (3DHBP, 4DHBP, 5DHBP, or 6DHBP) were systematically investigated. UV-vis titration data provided average pK a values of the hydroxy groups on the ligands. Both hydroxy groups were found to deprotonate in the pH 4.6–5.6 range for the 4–6DHBP complexes. One of the hydroxy groups of the 3DHBP complex exhibited the low pK a value of < 0.4 because the deprotonation is facilitated by the strong intramolecular hydrogen bond formed between the generated oxyanion and the remaining hydroxy group, which in turnmore » leads to an elevated pK a value of ~13.6 for the second deprotonation step. The crystal structures of the 4– and 6DHBP complexes obtained from basic aqueous solutions revealed their deprotonated forms. The intramolecular hydrogen bond in the 3DHBP complex was also observed in the crystal structures. The catalytic activities of these complexes in aqueous phase reactions, at appropriate pH, for hydrogenation of carbon dioxide (pH 8.5), dehydrogenation of formic acid (pH 1.8), transfer hydrogenation reactions using formic acid/formate as a hydrogen source (pH 7.2 and 2.6) were investigated to compare the positional effects of the hydroxy groups. The 4– and 6DHBP complexes exhibited remarkably enhanced catalytic activities under basic conditions because of the resonance effect of the strong electrondonating oxyanions, whereas the 5DHBP complex exhibited negligible activity despite the presence of electron-donating groups. The 3DHBP complex exhibited relatively high catalytic activity at low pH owing to the one strong electron-donating oxyanion group stabilized by the intramolecular hydrogen bond. DFT calculations were employed to study the mechanism of CO₂ hydrogenation by the 4DHBP and 6DHBP complexes, and comparison of the activation free energies of the H₂ heterolysis and CO₂ insertion steps indicated that H₂ heterolysis is the rate-determining step for both complexes. The presence of a pendent base in the 6DHBP complex was found to facilitate the rate-determining step, and renders 6DHBP a more effective catalyst for formate production.« less

Adenosine 3′,5′-cyclic monophosphate (cAMP)-dependent phosphoregulation of mitochondrial complex I is inhibited by nucleoside reverse transcriptase inhibitors

PubMed Central

Lund, Kaleb C.; Wallace, Kendall B.

2008-01-01

Nucleoside analog reverse transcriptase inhibitors (NRTI) are known to directly inhibit mitochondrial complex I activity as well as various mitochondrial kinases. Recent observations that complex I activity and superoxide production are modulated through cAMP-dependent phosphorylation suggests a mechanism through which NRTIs may affect mitochondrial respiration via kinase-dependent protein phosphorylation. In the current study we examine the potential for NRTIs to inhibit the cAMP-dependent phosphorylation of complex I and the associated NADH:CoQ oxidoreductase activities and rates of superoxide production using HepG2 cells. Phosphoprotein staining of immunocaptured complex I revealed that 3′-azido-3′-deoxythymidine (AZT; 10 and 50 μM), AZT monophosphate (150 μM), and 2′,3′-dideoxycytidine (ddC; 1μM) prevented the phosphorylation of the NDUFB11 subunit of complex I. This was associated with a decrease in complex I activity with AZT and AZT monophosphate only. In the presence of succinate, superoxide production was increased with 2′,3′-dideoxyinosine (ddI; 10 μM) and ddC (1 μM). In the presence of succinate + cAMP AZT showed an inverse dose-dependent effect on superoxide production. None of the NRTIs examined inhibit PKA activity suggesting that the observed effects are due to a direct interaction with complex I. These data demonstrate a direct effect of NRTIs on cAMP-dependent regulation of mitochondrial bioenergetics independent of DNA polymerase-γ activity; in the case of AZT these observations may provide a mechanism for the observed long-term toxicity with this drug. PMID:17904600

Effect of mitochondrial complex I inhibition on Fe-S cluster protein activity

DOE Office of Scientific and Technical Information (OSTI.GOV)

Mena, Natalia P.; Millennium Institute of Cell Dynamics and Biotechnology, Santiago; Bulteau, Anne Laure

2011-06-03

Highlights: {yields} Mitochondrial complex I inhibition resulted in decreased activity of Fe-S containing enzymes mitochondrial aconitase and cytoplasmic aconitase and xanthine oxidase. {yields} Complex I inhibition resulted in the loss of Fe-S clusters in cytoplasmic aconitase and of glutamine phosphoribosyl pyrophosphate amidotransferase. {yields} Consistent with loss of cytoplasmic aconitase activity, an increase in iron regulatory protein 1 activity was found. {yields} Complex I inhibition resulted in an increase in the labile cytoplasmic iron pool. -- Abstract: Iron-sulfur (Fe-S) clusters are small inorganic cofactors formed by tetrahedral coordination of iron atoms with sulfur groups. Present in numerous proteins, these clusters aremore » involved in key biological processes such as electron transfer, metabolic and regulatory processes, DNA synthesis and repair and protein structure stabilization. Fe-S clusters are synthesized mainly in the mitochondrion, where they are directly incorporated into mitochondrial Fe-S cluster-containing proteins or exported for cytoplasmic and nuclear cluster-protein assembly. In this study, we tested the hypothesis that inhibition of mitochondrial complex I by rotenone decreases Fe-S cluster synthesis and cluster content and activity of Fe-S cluster-containing enzymes. Inhibition of complex I resulted in decreased activity of three Fe-S cluster-containing enzymes: mitochondrial and cytosolic aconitases and xanthine oxidase. In addition, the Fe-S cluster content of glutamine phosphoribosyl pyrophosphate amidotransferase and mitochondrial aconitase was dramatically decreased. The reduction in cytosolic aconitase activity was associated with an increase in iron regulatory protein (IRP) mRNA binding activity and with an increase in the cytoplasmic labile iron pool. Since IRP activity post-transcriptionally regulates the expression of iron import proteins, Fe-S cluster inhibition may result in a false iron deficiency signal. Given that inhibition of complex I and iron accumulation are hallmarks of idiopathic Parkinson's disease, the findings reported here may have relevance for understanding the pathophysiology of this disease.« less

Comprehensive chemical characterization of industrial PM2.5 from steel industry activities

NASA Astrophysics Data System (ADS)

Sylvestre, Alexandre; Mizzi, Aurélie; Mathiot, Sébastien; Masson, Fanny; Jaffrezo, Jean L.; Dron, Julien; Mesbah, Boualem; Wortham, Henri; Marchand, Nicolas

2017-03-01

Industrial sources are among the least documented PM (Particulate Matter) source in terms of chemical composition, which limits our understanding of their effective impact on ambient PM concentrations. We report 4 chemical emission profiles of PM2.5 for multiple activities located in a vast metallurgical complex. Emissions profiles were calculated as the difference of species concentrations between an upwind and a downwind site normalized by the absolute PM2.5 enrichment between both sites. We characterized the PM2.5 emissions profiles of the industrial activities related to the cast iron (complex 1) and the iron ore conversion processes (complex 2), as well as 2 storage areas: a blast furnace slag area (complex 3) and an ore terminal (complex 4). PM2.5 major fractions (Organic Carbon (OC) and Elemental Carbon (EC), major ions), organic markers as well as metals/trace elements are reported for the 4 industrial complexes. Among the trace elements, iron is the most emitted for the complex 1 (146.0 mg g-1 of PM2.5), the complex 2 (70.07 mg g-1) and the complex 3 (124.4 mg g-1) followed by Al, Mn and Zn. A strong emission of Polycyclic Aromatic Hydrocarbons (PAH), representing 1.3% of the Organic Matter (OM), is observed for the iron ore transformation complex (complex 2) which merges the activities of coke and iron sinter production and the blast furnace processes. In addition to unsubstituted PAHs, sulfur containing PAHs (SPAHs) are also significantly emitted (between 0.011 and 0.068 mg g-1) by the complex 2 and could become very useful organic markers of steel industry activities. For the complexes 1 and 2 (cast iron and iron ore converters), a strong fraction of sulfate ranging from 0.284 to 0.336 g g-1) and only partially neutralized by ammonium, is observed indicating that sulfates, if not directly emitted by the industrial activity, are formed very quickly in the plume. Emission from complex 4 (Ore terminal) are characterized by high contribution of Al (125.7 mg g-1 of PM2.5) but also, in a lesser extent, of Fe, Mn, Ti and Zn. We also highlighted high contribution of calcium ranging from 0.123 to 0.558 g g-1 for all of the industrial complexes under study. Since calcium is also widely used as a proxy of the dust contributions in source apportionment studies, our results suggest that this assumption should be reexamined in environments impacted by industrial emissions.

The role of activity complexes in the distribution of solar magnetic fields.

NASA Astrophysics Data System (ADS)

García de La Rosa, J. I.; Reyes, R. C.

Using published data on the large-scale distribution of solar activity, the authors conclude that the longlived coronal holes are formed and maintained by the unbalanced magnetic flux which developes at both extremes of the complexes of activity.

Development of an efficient E. coli expression and purification system for a catalytically active, human Cullin3-RINGBox1 protein complex and elucidation of its quaternary structure with Keap1

DOE Office of Scientific and Technical Information (OSTI.GOV)

Small, Evan; Eggler, Aimee; Mesecar, Andrew D., E-mail: amesecar@purdue.edu

2010-10-01

Research highlights: {yields} A novel expression strategy was used to purify Cul3-Rbx1 from E. coli. {yields} The Cul3-Rbx1 complex is fully active and catalyzes ubiquitination of Nrf2 in vitro. {yields} Cul3, Rbx1, and Keap1 form a complex with unique stoichiometry of 1:1:2. -- Abstract: The Cullin3-based E3 ubiquitin ligase complex is thought to play an important role in the cellular response to oxidative stress and xenobiotic assault. While limited biochemical studies of the ligase's role in these complex signaling pathways are beginning to emerge, structural studies are lagging far behind due to the inability to acquire sufficient quantities of full-length,more » highly pure and active Cullin3. Here we describe the design and construction of an optimized expression and purification system for the full-length, human Cullin3-RINGBox 1 (Rbx1) protein complex from Escherichia coli. The dual-expression system is comprised of codon-optimized Cullin3 and Rbx1 genes co-expressed from a single pET-Duet-1 plasmid. Rapid purification of the Cullin3-Rbx1 complex is achieved in two steps via an affinity column followed by size-exclusion chromatography. Approximately 15 mg of highly pure and active Cullin3-Rbx1 protein from 1 L of E. coli culture can be achieved. Analysis of the quaternary structure of the Cullin3-Rbx1 and Cullin3-Rbx1-Keap1 complexes by size-exclusion chromatography and analytical ultracentrifugation indicates a 1:1 stoichiometry for the Cullin3-Rbx1 complex (MW = 111 kDa), and a 1:1:2 stoichiometry for the Cullin3-Rbx1-Keap1 complex (MW = 280 kDa). This latter complex has a novel quaternary structural organization for cullin E3 ligases, and it is fully active based on an in vitro Cullin3-Rbx1-Keap1-Nrf2 ubiquitination activity assay that was developed and optimized in this study.« less

Parallel effects of memory set activation and search on timing and working memory capacity.

PubMed

Schweickert, Richard; Fortin, Claudette; Xi, Zhuangzhuang; Viau-Quesnel, Charles

2014-01-01

Accurately estimating a time interval is required in everyday activities such as driving or cooking. Estimating time is relatively easy, provided a person attends to it. But a brief shift of attention to another task usually interferes with timing. Most processes carried out concurrently with timing interfere with it. Curiously, some do not. Literature on a few processes suggests a general proposition, the Timing and Complex-Span Hypothesis: A process interferes with concurrent timing if and only if process performance is related to complex span. Complex-span is the number of items correctly recalled in order, when each item presented for study is followed by a brief activity. Literature on task switching, visual search, memory search, word generation and mental time travel supports the hypothesis. Previous work found that another process, activation of a memory set in long term memory, is not related to complex-span. If the Timing and Complex-Span Hypothesis is true, activation should not interfere with concurrent timing in dual-task conditions. We tested such activation in single-task memory search task conditions and in dual-task conditions where memory search was executed with concurrent timing. In Experiment 1, activating a memory set increased reaction time, with no significant effect on time production. In Experiment 2, set size and memory set activation were manipulated. Activation and set size had a puzzling interaction for time productions, perhaps due to difficult conditions, leading us to use a related but easier task in Experiment 3. In Experiment 3 increasing set size lengthened time production, but memory activation had no significant effect. Results here and in previous literature on the whole support the Timing and Complex-Span Hypotheses. Results also support a sequential organization of activation and search of memory. This organization predicts activation and set size have additive effects on reaction time and multiplicative effects on percent correct, which was found.

Synthesis and spectroscopic characterization of gallic acid and some of its azo complexes

NASA Astrophysics Data System (ADS)

Masoud, Mamdouh S.; Hagagg, Sawsan S.; Ali, Alaa E.; Nasr, Nessma M.

2012-04-01

A series of gallic acid and azo gallic acid complexes were prepared and characterized by elemental analysis, IR, electronic spectra and magnetic susceptibility. The complexes were of different geometries: Octahedral, Tetrahedral and Square Planar. ESR was studied for copper complexes. All of the prepared complexes were of isotropic nature. The thermal analyses of the complexes were studied by DTA and DSC techniques. The thermodynamic parameters and the thermal transitions, such as glass transitions, crystallization and melting temperatures for some ligands and their complexes were evaluated and discussed. The entropy change values, ΔS#, showed that the transition states are more ordered than the reacting complexes. The biological activities of some ligands and their complexes are tested against Gram positive and Gram negative bacteria. The results showed that some complexes have a well considerable activity against different organisms.

Inhibition of nuclear factor kappaB proteins-platinated DNA interactions correlates with cytotoxic effectiveness of the platinum complexes

PubMed Central

Brabec, Viktor; Kasparkova, Jana; Kostrhunova, Hana; Farrell, Nicholas P.

2016-01-01

Nuclear DNA is the target responsible for anticancer activity of platinum anticancer drugs. Their activity is mediated by altered signals related to programmed cell death and the activation of various signaling pathways. An example is activation of nuclear factor kappaB (NF-κB). Binding of NF-κB proteins to their consensus sequences in DNA (κB sites) is the key biochemical activity responsible for the biological functions of NF-κB. Using gel-mobility-shift assays and surface plasmon resonance spectroscopy we examined the interactions of NF-κB proteins with oligodeoxyribonucleotide duplexes containing κB site damaged by DNA adducts of three platinum complexes. These complexes markedly differed in their toxic effects in tumor cells and comprised highly cytotoxic trinuclear platinum(II) complex BBR3464, less cytotoxic conventional cisplatin and ineffective transplatin. The results indicate that structurally different DNA adducts of these platinum complexes exhibit a different efficiency to affect the affinity of the platinated DNA (κB sites) to NF-κB proteins. Our results support the hypothesis that structural perturbations induced in DNA by platinum(II) complexes correlate with their higher efficiency to inhibit binding of NF-κB proteins to their κB sites and cytotoxicity as well. However, the full generalization of this hypothesis will require to evaluate a larger series of platinum(II) complexes. PMID:27574114

Inhibition of nuclear factor kappaB proteins-platinated DNA interactions correlates with cytotoxic effectiveness of the platinum complexes.

PubMed

Brabec, Viktor; Kasparkova, Jana; Kostrhunova, Hana; Farrell, Nicholas P

2016-08-30

Nuclear DNA is the target responsible for anticancer activity of platinum anticancer drugs. Their activity is mediated by altered signals related to programmed cell death and the activation of various signaling pathways. An example is activation of nuclear factor kappaB (NF-κB). Binding of NF-κB proteins to their consensus sequences in DNA (κB sites) is the key biochemical activity responsible for the biological functions of NF-κB. Using gel-mobility-shift assays and surface plasmon resonance spectroscopy we examined the interactions of NF-κB proteins with oligodeoxyribonucleotide duplexes containing κB site damaged by DNA adducts of three platinum complexes. These complexes markedly differed in their toxic effects in tumor cells and comprised highly cytotoxic trinuclear platinum(II) complex BBR3464, less cytotoxic conventional cisplatin and ineffective transplatin. The results indicate that structurally different DNA adducts of these platinum complexes exhibit a different efficiency to affect the affinity of the platinated DNA (κB sites) to NF-κB proteins. Our results support the hypothesis that structural perturbations induced in DNA by platinum(II) complexes correlate with their higher efficiency to inhibit binding of NF-κB proteins to their κB sites and cytotoxicity as well. However, the full generalization of this hypothesis will require to evaluate a larger series of platinum(II) complexes.