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Sample records for modulate protein metabolism

  1. APL-1, the Alzheimer's Amyloid precursor protein in Caenorhabditis elegans, modulates multiple metabolic pathways throughout development.

    PubMed

    Ewald, Collin Y; Raps, Daniel A; Li, Chris

    2012-06-01

    Mutations in the amyloid precursor protein (APP) gene or in genes that process APP are correlated with familial Alzheimer's disease (AD). The biological function of APP remains unclear. APP is a transmembrane protein that can be sequentially cleaved by different secretases to yield multiple fragments, which can potentially act as signaling molecules. Caenorhabditis elegans encodes one APP-related protein, APL-1, which is essential for viability. Here, we show that APL-1 signaling is dependent on the activity of the FOXO transcription factor DAF-16 and the nuclear hormone receptor DAF-12 and influences metabolic pathways such as developmental progression, body size, and egg-laying rate. Furthermore, apl-1(yn5) mutants, which produce high levels of the extracellular APL-1 fragment, show an incompletely penetrant temperature-sensitive embryonic lethality. In a genetic screen to isolate mutants in which the apl-1(yn5) lethality rate is modified, we identified a suppressor mutation in MOA-1/R155.2, a receptor-protein tyrosine phosphatase, and an enhancer mutation in MOA-2/B0495.6, a protein involved in receptor-mediated endocytosis. Knockdown of apl-1 in an apl-1(yn5) background caused lethality and molting defects at all larval stages, suggesting that apl-1 is required for each transitional molt. We suggest that signaling of the released APL-1 fragment modulates multiple metabolic states and that APL-1 is required throughout development.

  2. Dengue Virus NS1 Protein Modulates Cellular Energy Metabolism by Increasing Glyceraldehyde-3-Phosphate Dehydrogenase Activity

    PubMed Central

    Allonso, Diego; Andrade, Iamara S.; Conde, Jonas N.; Coelho, Diego R.; Rocha, Daniele C. P.; da Silva, Manuela L.; Ventura, Gustavo T.

    2015-01-01

    ABSTRACT Dengue is one of the main public health concerns worldwide. Recent estimates indicate that over 390 million people are infected annually with the dengue virus (DENV), resulting in thousands of deaths. Among the DENV nonstructural proteins, the NS1 protein is the only one whose function during replication is still unknown. NS1 is a 46- to 55-kDa glycoprotein commonly found as both a membrane-associated homodimer and a soluble hexameric barrel-shaped lipoprotein. Despite its role in the pathogenic process, NS1 is essential for proper RNA accumulation and virus production. In the present study, we identified that glyceraldehyde-3-phosphate dehydrogenase (GAPDH) interacts with intracellular NS1. Molecular docking revealed that this interaction occurs through the hydrophobic protrusion of NS1 and the hydrophobic residues located at the opposite side of the catalytic site. Moreover, addition of purified recombinant NS1 enhanced the glycolytic activity of GAPDH in vitro. Interestingly, we observed that DENV infection promoted the relocalization of GAPDH to the perinuclear region, where NS1 is commonly found. Both DENV infection and expression of NS1 itself resulted in increased GAPDH activity. Our findings indicate that the NS1 protein acts to increase glycolytic flux and, consequently, energy production, which is consistent with the recent finding that DENV induces and requires glycolysis for proper replication. This is the first report to propose that NS1 is an important modulator of cellular energy metabolism. The data presented here provide new insights that may be useful for further drug design and the development of alternative antiviral therapies against DENV. IMPORTANCE Dengue represents a serious public health problem worldwide and is caused by infection with dengue virus (DENV). Estimates indicate that half of the global population is at risk of infection, with almost 400 million cases occurring per year. The NS1 glycoprotein is found in both the

  3. Pink-eyed Dilution Protein Modulates Arsenic Sensitivity and Intracellular Glutathione Metabolism

    PubMed Central

    Staleva, Liliana; Manga, Prashiela; Orlow, Seth J.

    2002-01-01

    Mutations in the mouse p (pink-eyed dilution) and human P genes lead to melanosomal defects and ocular developmental abnormalities. Despite the critical role played by the p gene product in controlling tyrosinase processing and melanosome biogenesis, its precise biological function is still not defined. We have expressed p heterologously in the yeast Saccharomyces cerevisiae to study its function in greater detail. Immunofluorescence studies revealed that p reaches the yeast vacuolar membrane via the prevacuolar compartment. Yeast cells expressing p exhibited increased sensitivity to a number of toxic compounds, including arsenicals. Similarly, cultured murine melanocytes expressing a functional p gene were also found to be more sensitive to arsenical compounds compared with p-null cell lines. Intracellular glutathione, known to play a role in detoxification of arsenicals, was diminished by 50% in p-expressing yeast. By using the glutathione-conjugating dye monochlorobimane, in combination with acivicin, an inhibitor of vacuolar gamma-glutamyl cysteine transpeptidase, involved in the breakdown of glutathione, we found that p facilitates the vacuolar accumulation of glutathione. Our data demonstrate that the pink-eyed dilution protein increases cellular sensitivity to arsenicals and other metalloids and can modulate intracellular glutathione metabolism. PMID:12475946

  4. Dynamic Evolution of Nitric Oxide Detoxifying Flavohemoglobins, a Family of Single-Protein Metabolic Modules in Bacteria and Eukaryotes.

    PubMed

    Wisecaver, Jennifer H; Alexander, William G; King, Sean B; Hittinger, Chris Todd; Rokas, Antonis

    2016-08-01

    Due to their functional independence, proteins that comprise standalone metabolic units, which we name single-protein metabolic modules, may be particularly prone to gene duplication (GD) and horizontal gene transfer (HGT). Flavohemoglobins (flavoHbs) are prime examples of single-protein metabolic modules, detoxifying nitric oxide (NO), a ubiquitous toxin whose antimicrobial properties many life forms exploit, to nitrate, a common source of nitrogen for organisms. FlavoHbs appear widespread in bacteria and have been identified in a handful of microbial eukaryotes, but how the distribution of this ecologically and biomedically important protein family evolved remains unknown. Reconstruction of the evolutionary history of 3,318 flavoHb protein sequences covering the family's known diversity showed evidence of recurrent HGT at multiple evolutionary scales including intrabacterial HGT, as well as HGT from bacteria to eukaryotes. One of the most striking examples of HGT is the acquisition of a flavoHb by the dandruff- and eczema-causing fungus Malassezia from Corynebacterium Actinobacteria, a transfer that growth experiments show is capable of mediating NO resistance in fungi. Other flavoHbs arose via GD; for example, many filamentous fungi possess two flavoHbs that are differentially targeted to the cytosol and mitochondria, likely conferring protection against external and internal sources of NO, respectively. Because single-protein metabolic modules such as flavoHb function independently, readily undergo GD and HGT, and are frequently involved in organismal defense and competition, we suggest that they represent "plug-and-play" proteins for ecological arms races. PMID:27189567

  5. UNCOUPLING PROTEIN-2 MODULATES THE LIPID METABOLIC RESPONSE TO FASTING IN MICE

    PubMed Central

    Sheets, Anthony R.; Fülöp, Péter; Derdák, Zoltán; Kassai, Andrea; Sabo, Edmond; Mark, Nicholas M.; Paragh, György; Wands, Jack R.; Baffy, György

    2008-01-01

    Uncoupling protein-2 (UCP2) regulates insulin secretion by controlling ATP levels in β cells. While UCP2 deficiency improves glycemic control in mice, increased expression of UCP2 interferes with glucose-stimulated insulin secretion. These observations link UCP2 to β cell dysfunction in type 2 diabetes with a perplexing evolutionary role. We found higher residual serum insulin levels and blunted lipid metabolic responses in fasted ucp2−/− mice, supporting the concept that UCP2 evolved to suppress insulin effects and to accommodate the fuel switch to fatty acids during starvation. In the absence of UCP2, fasting initially promotes peripheral lipolysis and hepatic fat accumulation at less than expected rates, but culminates in protracted steatosis indicating diminished hepatic utilization and clearance of fatty acids. We conclude that UCP2-mediated control of insulin secretion is a physiologically relevant mechanism of the metabolic response to fasting. PMID:18292186

  6. APL-1, the Alzheimer’s Amyloid Precursor Protein in Caenorhabditis elegans, Modulates Multiple Metabolic Pathways Throughout Development

    PubMed Central

    Ewald, Collin Y.; Raps, Daniel A.; Li, Chris

    2012-01-01

    Mutations in the amyloid precursor protein (APP) gene or in genes that process APP are correlated with familial Alzheimer’s disease (AD). The biological function of APP remains unclear. APP is a transmembrane protein that can be sequentially cleaved by different secretases to yield multiple fragments, which can potentially act as signaling molecules. Caenorhabditis elegans encodes one APP-related protein, APL-1, which is essential for viability. Here, we show that APL-1 signaling is dependent on the activity of the FOXO transcription factor DAF-16 and the nuclear hormone receptor DAF-12 and influences metabolic pathways such as developmental progression, body size, and egg-laying rate. Furthermore, apl-1(yn5) mutants, which produce high levels of the extracellular APL-1 fragment, show an incompletely penetrant temperature-sensitive embryonic lethality. In a genetic screen to isolate mutants in which the apl-1(yn5) lethality rate is modified, we identified a suppressor mutation in MOA-1/R155.2, a receptor-protein tyrosine phosphatase, and an enhancer mutation in MOA-2/B0495.6, a protein involved in receptor-mediated endocytosis. Knockdown of apl-1 in an apl-1(yn5) background caused lethality and molting defects at all larval stages, suggesting that apl-1 is required for each transitional molt. We suggest that signaling of the released APL-1 fragment modulates multiple metabolic states and that APL-1 is required throughout development. PMID:22466039

  7. Rice bran proteins and their hydrolysates modulate cholesterol metabolism in mice on hypercholesterolemic diets.

    PubMed

    Zhang, Huijuan; Wang, Jing; Liu, Yingli; Gong, Lingxiao; Sun, Baoguo

    2016-06-15

    The hypolipidemic properties of defatted rice bran protein (DRBP), fresh rice bran protein (FRBP), DRBP hydrolysates (DRBPH), and FRBP hydrolysates (FRBPH) were determined in mice on high fat diets for four weeks. Very low-density lipoprotein cholesterol (VLDL-C), low-density lipoprotein cholesterol (LDL-C) contents, and the hepatic total cholesterol content were reduced while fecal total cholesterol and total bile acid (TBA) contents were increased in the FRBPH diet group. The expression levels of hepatic genes for cholesterol biosynthesis HMG-CoAR and SREBP-2 were lowest in the FRBPH diet group. The mRNA level of HMG-CoAR was significantly positively correlated with the hepatic TG content (r = 0.82, P < 0.05). The mRNA levels of genes related to bile acid biosynthesis and cholesterol efflux, CYP7A1, ABCA1, and PPARγ were up-regulated in all test groups. The results suggest that FRBPH regulates cholesterol metabolism in mice fed the high fat and cholesterol diet by increasing fecal steroid excretion and expression levels of genes related to bile acid synthesis and cholesterol efflux, and the down-regulation of the expression levels of genes related to cholesterol biosynthesis. PMID:27216972

  8. Protein metabolism and requirements.

    PubMed

    Biolo, Gianni

    2013-01-01

    Skeletal muscle adaptation to critical illness includes insulin resistance, accelerated proteolysis, and increased release of glutamine and the other amino acids. Such amino acid efflux from skeletal muscle provides precursors for protein synthesis and energy fuel to the liver and to the rapidly dividing cells of the intestinal mucosa and the immune system. From these adaptation mechanisms, severe muscle wasting, glutamine depletion, and hyperglycemia, with increased patient morbidity and mortality, may ensue. Protein/amino acid nutrition, through either enteral or parenteral routes, plays a pivotal role in treatment of metabolic abnormalities in critical illness. In contrast to energy requirement, which can be accurately assessed by indirect calorimetry, methods to determine individual protein/amino acid needs are not currently available. In critical illness, a decreased ability of protein/amino acid intake to promote body protein synthesis is defined as anabolic resistance. This abnormality leads to increased protein/amino acid requirement and relative inefficiency of nutritional interventions. In addition to stress mediators, immobility and physical inactivity are key determinants of anabolic resistance. The development of mobility protocols in the intensive care unit should be encouraged to enhance the efficacy of nutrition. In critical illness, protein/amino acid requirement has been defined as the intake level associated with the lowest rate of catabolism. The optimal protein-sparing effects in patients receiving adequate energy are achieved when protein/amino acids are administered at rates between 1.3 and 1.5 g/kg/day. Extra glutamine supplementation is required in conditions of severe systemic inflammatory response. Protein requirement increases during hypocaloric feeding and in patients with acute renal failure on continuous renal replacement therapy. Evidence suggests that receiving adequate protein/amino acid intake may be more important than achieving

  9. Increasing protein intake modulates lipid metabolism in healthy young men and women consuming a high-fat hypercaloric diet.

    PubMed

    Rietman, Annemarie; Schwarz, Jessica; Blokker, Britt A; Siebelink, Els; Kok, Frans J; Afman, Lydia A; Tomé, Daniel; Mensink, Marco

    2014-08-01

    The objective of this study was to evaluate the effect of increasing protein intake, at the expense of carbohydrates, on intrahepatic lipids (IHLs), circulating triglycerides (TGs), and body composition in healthy humans consuming a high-fat, hypercaloric diet. A crossover randomized trial with a parallel control group was performed. After a 2-wk run-in period, participants were assigned to either the control diet [n = 10; 27.8 energy percent (en%) fat, 16.9 en% protein, 55.3 en% carbohydrates] for 4 wk or a high-fat, hypercaloric diet (n = 17; >2 MJ/d) crossover trial with 2 periods of 2 wk, with either high-protein (HP) (37.7 en% fat, 25.7 en% protein, 36.6 en% carbohydrates) or normal-protein (NP) (39.4 en% fat, 15.4 en% protein, 45.2 en% carbohydrates) content. Measurements were performed after 2 wk of run-in (baseline), 2 wk of intervention (period 1), and 4 wk of intervention (period 2). A trend toward lower IHL and plasma TG concentrations during the HP condition compared with the NP condition was observed (IHL: 0.35 ± 0.04% vs. 0.51 ± 0.08%, P = 0.08; TG: 0.65 ± 0.03 vs. 0.77 ± 0.05 mmol/L, P = 0.07, for HP and NP, respectively). Fat mass was significantly lower (10.6 ± 1.72 vs. 10.9 ± 1.73 kg; P = 0.02) with the HP diet than with the NP diet, whereas fat-free mass was higher (55.7 ± 2.79 vs. 55.2 ± 2.80 kg; P = 0.003). This study indicated that an HP, high-fat, hypercaloric diet affects lipid metabolism. It tends to lower the IHL and circulating TG concentrations and significantly lowers fat mass and increases fat-free mass compared with an NP, high-fat, hypercaloric diet. This trail was registered at www.clinicaltrials.gov as NCT01354626.

  10. The Copper Metabolism MURR1 Domain Protein 1 (COMMD1) Modulates the Aggregation of Misfolded Protein Species in a Client-Specific Manner

    PubMed Central

    Vonk, Willianne I. M.; Kakkar, Vaishali; Bartuzi, Paulina; Jaarsma, Dick; Berger, Ruud; Hofker, Marten H.; Klomp, Leo W. J.; Wijmenga, Cisca; Kampinga, Harm H.; van de Sluis, Bart

    2014-01-01

    The Copper Metabolism MURR1 domain protein 1 (COMMD1) is a protein involved in multiple cellular pathways, including copper homeostasis, NF-κB and hypoxia signalling. Acting as a scaffold protein, COMMD1 mediates the levels, stability and proteolysis of its substrates (e.g. the copper-transporters ATP7B and ATP7A, RELA and HIF-1α). Recently, we established an interaction between the Cu/Zn superoxide dismutase 1 (SOD1) and COMMD1, resulting in a decreased maturation and activation of SOD1. Mutations in SOD1, associated with the progressive neurodegenerative disorder Amyotrophic Lateral Sclerosis (ALS), cause misfolding and aggregation of the mutant SOD1 (mSOD1) protein. Here, we identify COMMD1 as a novel regulator of misfolded protein aggregation as it enhances the formation of mSOD1 aggregates upon binding. Interestingly, COMMD1 co-localizes to the sites of mSOD1 inclusions and forms high molecular weight complexes in the presence of mSOD1. The effect of COMMD1 on protein aggregation is client-specific as, in contrast to mSOD1, COMMD1 decreases the abundance of mutant Parkin inclusions, associated with Parkinson’s disease. Aggregation of a polyglutamine-expanded Huntingtin, causative of Huntington’s disease, appears unaltered by COMMD1. Altogether, this study offers new research directions to expand our current knowledge on the mechanisms underlying aggregation disease pathologies. PMID:24691167

  11. Pharmacologic inhibition of the CK2-mediated phosphorylation of B23/NPM in cancer cells selectively modulates genes related to protein synthesis, energetic metabolism, and ribosomal biogenesis.

    PubMed

    Perera, Yasser; Pedroso, Seidy; Borras-Hidalgo, Orlando; Vázquez, Dania M; Miranda, Jamilet; Villareal, Adelaida; Falcón, Viviana; Cruz, Luis D; Farinas, Hernán G; Perea, Silvio E

    2015-06-01

    B23/NPM is a multifunctional nucleolar protein frequently overexpressed, mutated, or rearranged in neoplastic tissues. B23/NPM is involved in diverse biological processes and is mainly regulated by heteroligomer association and posttranslational modification, phosphorylation being a major posttranslational event. While the role of B23/NPM in supporting and/or driving malignant transformation is widely recognized, the particular relevance of its CK2-mediated phosphorylation remains unsolved. Interestingly, the pharmacologic inhibition of such phosphorylation event by CIGB-300, a clinical-grade peptide drug, was previously associated to apoptosis induction in tumor cell lines. In this work, we sought to identify the biological processes modulated by CIGB-300 in a lung cancer cell line using subtractive suppression hybridization and subsequent functional annotation clustering. Our results indicate that CIGB-300 modulates a subset of genes involved in protein synthesis (ES = 8.4, p < 0.001), mitochondrial ATP metabolism (ES = 2.5, p < 0.001), and ribosomal biogenesis (ES = 1.5, p < 0.05). The impairment of these cellular processes by CIGB-300 was corroborated at the molecular and cellular levels by Western blot (P-S6/P-4EBP1, translation), confocal microscopy (JC-1, mitochondrial potential), qPCR (45SrRNA/p21, ribosome biogenesis), and electron microscopy (nucleolar structure, ribosome biogenesis). Altogether, our findings provide new insights on the potential relevance of the CK2-mediated phosphorylation of B23/NPM in cancer cells, revealing at the same time the potentialities of its pharmacological manipulation for cancer therapy. Finally, this work also suggests several candidate gene biomarkers to be evaluated during the clinical development of the anti-CK2 peptide CIGB-300.

  12. Epigenetic modulation of metabolic decisions.

    PubMed

    Öst, Anita; Pospisilik, John Andrew

    2015-04-01

    In the recent years there has been a tremendous increase in our understanding of chromatin, transcription and the importance of metabolites in their regulation. This review highlights what is currently sparse information that suggest existence of a refined system integrating metabolic and chromatin control. We indicate possible regulatory modes, such as feed forward amplification, that may help effect and stabilize long-lasting phenotypic decisions within and even across generations using adipogenesis as the primary context.

  13. Genetic and computational identification of a conserved bacterial metabolic module.

    PubMed

    Boutte, Cara C; Srinivasan, Balaji S; Flannick, Jason A; Novak, Antal F; Martens, Andrew T; Batzoglou, Serafim; Viollier, Patrick H; Crosson, Sean

    2008-12-01

    We have experimentally and computationally defined a set of genes that form a conserved metabolic module in the alpha-proteobacterium Caulobacter crescentus and used this module to illustrate a schema for the propagation of pathway-level annotation across bacterial genera. Applying comprehensive forward and reverse genetic methods and genome-wide transcriptional analysis, we (1) confirmed the presence of genes involved in catabolism of the abundant environmental sugar myo-inositol, (2) defined an operon encoding an ABC-family myo-inositol transmembrane transporter, and (3) identified a novel myo-inositol regulator protein and cis-acting regulatory motif that control expression of genes in this metabolic module. Despite being encoded from non-contiguous loci on the C. crescentus chromosome, these myo-inositol catabolic enzymes and transporter proteins form a tightly linked functional group in a computationally inferred network of protein associations. Primary sequence comparison was not sufficient to confidently extend annotation of all components of this novel metabolic module to related bacterial genera. Consequently, we implemented the Graemlin multiple-network alignment algorithm to generate cross-species predictions of genes involved in myo-inositol transport and catabolism in other alpha-proteobacteria. Although the chromosomal organization of genes in this functional module varied between species, the upstream regions of genes in this aligned network were enriched for the same palindromic cis-regulatory motif identified experimentally in C. crescentus. Transposon disruption of the operon encoding the computationally predicted ABC myo-inositol transporter of Sinorhizobium meliloti abolished growth on myo-inositol as the sole carbon source, confirming our cross-genera functional prediction. Thus, we have defined regulatory, transport, and catabolic genes and a cis-acting regulatory sequence that form a conserved module required for myo-inositol metabolism in

  14. On Functional Module Detection in Metabolic Networks

    PubMed Central

    Koch, Ina; Ackermann, Jörg

    2013-01-01

    Functional modules of metabolic networks are essential for understanding the metabolism of an organism as a whole. With the vast amount of experimental data and the construction of complex and large-scale, often genome-wide, models, the computer-aided identification of functional modules becomes more and more important. Since steady states play a key role in biology, many methods have been developed in that context, for example, elementary flux modes, extreme pathways, transition invariants and place invariants. Metabolic networks can be studied also from the point of view of graph theory, and algorithms for graph decomposition have been applied for the identification of functional modules. A prominent and currently intensively discussed field of methods in graph theory addresses the Q-modularity. In this paper, we recall known concepts of module detection based on the steady-state assumption, focusing on transition-invariants (elementary modes) and their computation as minimal solutions of systems of Diophantine equations. We present the Fourier-Motzkin algorithm in detail. Afterwards, we introduce the Q-modularity as an example for a useful non-steady-state method and its application to metabolic networks. To illustrate and discuss the concepts of invariants and Q-modularity, we apply a part of the central carbon metabolism in potato tubers (Solanum tuberosum) as running example. The intention of the paper is to give a compact presentation of known steady-state concepts from a graph-theoretical viewpoint in the context of network decomposition and reduction and to introduce the application of Q-modularity to metabolic Petri net models. PMID:24958145

  15. On functional module detection in metabolic networks.

    PubMed

    Koch, Ina; Ackermann, Jörg

    2013-08-12

    Functional modules of metabolic networks are essential for understanding the metabolism of an organism as a whole. With the vast amount of experimental data and the construction of complex and large-scale, often genome-wide, models, the computer-aided identification of functional modules becomes more and more important. Since steady states play a key role in biology, many methods have been developed in that context, for example, elementary flux modes, extreme pathways, transition invariants and place invariants. Metabolic networks can be studied also from the point of view of graph theory, and algorithms for graph decomposition have been applied for the identification of functional modules. A prominent and currently intensively discussed field of methods in graph theory addresses the Q-modularity. In this paper, we recall known concepts of module detection based on the steady-state assumption, focusing on transition-invariants (elementary modes) and their computation as minimal solutions of systems of Diophantine equations. We present the Fourier-Motzkin algorithm in detail. Afterwards, we introduce the Q-modularity as an example for a useful non-steady-state method and its application to metabolic networks. To illustrate and discuss the concepts of invariants and Q-modularity, we apply a part of the central carbon metabolism in potato tubers (Solanum tuberosum) as running example. The intention of the paper is to give a compact presentation of known steady-state concepts from a graph-theoretical viewpoint in the context of network decomposition and reduction and to introduce the application of Q-modularity to metabolic Petri net models.

  16. On functional module detection in metabolic networks.

    PubMed

    Koch, Ina; Ackermann, Jörg

    2013-01-01

    Functional modules of metabolic networks are essential for understanding the metabolism of an organism as a whole. With the vast amount of experimental data and the construction of complex and large-scale, often genome-wide, models, the computer-aided identification of functional modules becomes more and more important. Since steady states play a key role in biology, many methods have been developed in that context, for example, elementary flux modes, extreme pathways, transition invariants and place invariants. Metabolic networks can be studied also from the point of view of graph theory, and algorithms for graph decomposition have been applied for the identification of functional modules. A prominent and currently intensively discussed field of methods in graph theory addresses the Q-modularity. In this paper, we recall known concepts of module detection based on the steady-state assumption, focusing on transition-invariants (elementary modes) and their computation as minimal solutions of systems of Diophantine equations. We present the Fourier-Motzkin algorithm in detail. Afterwards, we introduce the Q-modularity as an example for a useful non-steady-state method and its application to metabolic networks. To illustrate and discuss the concepts of invariants and Q-modularity, we apply a part of the central carbon metabolism in potato tubers (Solanum tuberosum) as running example. The intention of the paper is to give a compact presentation of known steady-state concepts from a graph-theoretical viewpoint in the context of network decomposition and reduction and to introduce the application of Q-modularity to metabolic Petri net models. PMID:24958145

  17. SIRTUIN 1 AND SIRTUIN 3: PHYSIOLOGICAL MODULATORS OF METABOLISM

    PubMed Central

    Nogueiras, Ruben; Habegger, Kirk M.; Chaudhary, Nilika; Finan, Brian; Banks, Alexander S.; Dietrich, Marcelo O.; Horvath, Tamas L.; Sinclair, David A.; Pfluger, Paul T.; Tschöop, Matthias H.

    2013-01-01

    The sirtuins are a family of highly conserved NAD+-dependent deacetylases that act as cellular sensors to detect energy availability and modulate metabolic processes. Two sirtuins that are central to the control of metabolic processes are mammalian sirtuin 1 (SIRT1) and sirtuin 3 (SIRT3), which are localized to the nucleus and mitochondria, respectively. Both are activated by high NAD+ levels, a condition caused by low cellular energy status. By deacetylating a variety of proteins that induce catabolic processes while inhibiting anabolic processes, SIRT1 and SIRT3 coordinately increase cellular energy stores and ultimately maintain cellular energy homeostasis. Defects in the pathways controlled by SIRT1 and SIRT3 are known to result in various metabolic disorders. Consequently, activation of sirtuins by genetic or pharmacological means can elicit multiple metabolic benefits that protect mice from diet-induced obesity, type 2 diabetes, and nonalcoholic fatty liver disease. PMID:22811431

  18. Potassium Uptake Modulates Staphylococcus aureus Metabolism

    PubMed Central

    Gries, Casey M.; Sadykov, Marat R.; Bulock, Logan L.; Chaudhari, Sujata S.; Thomas, Vinai C.; Bose, Jeffrey L.

    2016-01-01

    ABSTRACT As a leading cause of community-associated and nosocomial infections, Staphylococcus aureus requires sophisticated mechanisms that function to maintain cellular homeostasis in response to its exposure to changing environmental conditions. The adaptation to stress and maintenance of homeostasis depend largely on membrane activity, including supporting electrochemical gradients and synthesis of ATP. This is largely achieved through potassium (K+) transport, which plays an essential role in maintaining chemiosmotic homeostasis, affects antimicrobial resistance, and contributes to fitness in vivo. Here, we report that S. aureus Ktr-mediated K+ uptake is necessary for maintaining cytoplasmic pH and the establishment of a proton motive force. Metabolite analyses revealed that K+ deficiency affects both metabolic and energy states of S. aureus by impairing oxidative phosphorylation and directing carbon flux toward substrate-level phosphorylation. Taken together, these results underline the importance of K+ uptake in maintaining essential components of S. aureus metabolism. IMPORTANCE Previous studies describing mechanisms for K+ uptake in S. aureus revealed that the Ktr-mediated K+ transport system was required for normal growth under alkaline conditions but not under neutral or acidic conditions. This work focuses on the effect of K+ uptake on S. aureus metabolism, including intracellular pH and carbon flux, and is the first to utilize a pH-dependent green fluorescent protein (GFP) to measure S. aureus cytoplasmic pH. These studies highlight the role of K+ uptake in supporting proton efflux under alkaline conditions and uncover a critical role for K+ uptake in establishing efficient carbon utilization. PMID:27340697

  19. Potassium Uptake Modulates Staphylococcus aureus Metabolism.

    PubMed

    Gries, Casey M; Sadykov, Marat R; Bulock, Logan L; Chaudhari, Sujata S; Thomas, Vinai C; Bose, Jeffrey L; Bayles, Kenneth W

    2016-01-01

    As a leading cause of community-associated and nosocomial infections, Staphylococcus aureus requires sophisticated mechanisms that function to maintain cellular homeostasis in response to its exposure to changing environmental conditions. The adaptation to stress and maintenance of homeostasis depend largely on membrane activity, including supporting electrochemical gradients and synthesis of ATP. This is largely achieved through potassium (K(+)) transport, which plays an essential role in maintaining chemiosmotic homeostasis, affects antimicrobial resistance, and contributes to fitness in vivo. Here, we report that S. aureus Ktr-mediated K(+) uptake is necessary for maintaining cytoplasmic pH and the establishment of a proton motive force. Metabolite analyses revealed that K(+) deficiency affects both metabolic and energy states of S. aureus by impairing oxidative phosphorylation and directing carbon flux toward substrate-level phosphorylation. Taken together, these results underline the importance of K(+) uptake in maintaining essential components of S. aureus metabolism. IMPORTANCE Previous studies describing mechanisms for K(+) uptake in S. aureus revealed that the Ktr-mediated K(+) transport system was required for normal growth under alkaline conditions but not under neutral or acidic conditions. This work focuses on the effect of K(+) uptake on S. aureus metabolism, including intracellular pH and carbon flux, and is the first to utilize a pH-dependent green fluorescent protein (GFP) to measure S. aureus cytoplasmic pH. These studies highlight the role of K(+) uptake in supporting proton efflux under alkaline conditions and uncover a critical role for K(+) uptake in establishing efficient carbon utilization. PMID:27340697

  20. Designed Proteins To Modulate Cellular Networks

    PubMed Central

    Cortajarena, Aitziber L.; Liu, Tina Y.; Hochstrasser, Mark; Regan, Lynne

    2012-01-01

    A major challenge of protein design is to create useful new proteins that interact specifically with biological targets in living cells. Such binding modules have many potential applications, including the targeted perturbation of protein networks. As a general approach to create such modules, we designed a library with approximately 109 different binding specificities based on a small 3-tetratricopeptide repeat (TPR) motif framework. We employed a novel strategy, based on split GFP reassembly, to screen the library for modules with the desired binding specificity. Using this approach, we identified modules that bind tightly and specifically to Dss1, a small human protein that interacts with the tumor suppressor protein BRCA2. We showed that these modules also bind the yeast homologue of Dss1, Sem1. Furthermore, we demonstrated that these modules inhibit Sem1 activity in yeast. This strategy will be generally applicable to make novel genetically encoded tools for systems/synthetic biology applications. PMID:20020775

  1. Snail modulates cell metabolism in MDCK cells

    SciTech Connect

    Haraguchi, Misako; Indo, Hiroko P.; Iwasaki, Yasumasa; Iwashita, Yoichiro; Fukushige, Tomoko; Majima, Hideyuki J.; Izumo, Kimiko; Horiuchi, Masahisa; Kanekura, Takuro; Furukawa, Tatsuhiko; Ozawa, Masayuki

    2013-03-22

    Highlights: ► MDCK/snail cells were more sensitive to glucose deprivation than MDCK/neo cells. ► MDCK/snail cells had decreased oxidative phosphorylation, O{sub 2} consumption and ATP content. ► TCA cycle enzyme activity, but not expression, was lower in MDCK/snail cells. ► MDCK/snail cells showed reduced PDH activity and increased PDK1 expression. ► MDCK/snail cells showed reduced expression of GLS2 and ACLY. -- Abstract: Snail, a repressor of E-cadherin gene transcription, induces epithelial-to-mesenchymal transition and is involved in tumor progression. Snail also mediates resistance to cell death induced by serum depletion. By contrast, we observed that snail-expressing MDCK (MDCK/snail) cells undergo cell death at a higher rate than control (MDCK/neo) cells in low-glucose medium. Therefore, we investigated whether snail expression influences cell metabolism in MDCK cells. Although gylcolysis was not affected in MDCK/snail cells, they did exhibit reduced pyruvate dehydrogenase (PDH) activity, which controls pyruvate entry into the tricarboxylic acid (TCA) cycle. Indeed, the activity of multiple enzymes involved in the TCA cycle was decreased in MDCK/snail cells, including that of mitochondrial NADP{sup +}-dependent isocitrate dehydrogenase (IDH2), succinate dehydrogenase (SDH), and electron transport Complex II and Complex IV. Consequently, lower ATP content, lower oxygen consumption and increased survival under hypoxic conditions was also observed in MDCK/snail cells compared to MDCK/neo cells. In addition, the expression and promoter activity of pyruvate dehydrogenase kinase 1 (PDK1), which phosphorylates and inhibits the activity of PDH, was increased in MDCK/snail cells, while expression levels of glutaminase 2 (GLS2) and ATP-citrate lyase (ACLY), which are involved in glutaminolysis and fatty acid synthesis, were decreased in MDCK/snail cells. These results suggest that snail modulates cell metabolism by altering the expression and activity of

  2. Module organization and variance in protein-protein interaction networks

    NASA Astrophysics Data System (ADS)

    Lin, Chun-Yu; Lee, Tsai-Ling; Chiu, Yi-Yuan; Lin, Yi-Wei; Lo, Yu-Shu; Lin, Chih-Ta; Yang, Jinn-Moon

    2015-03-01

    A module is a group of closely related proteins that act in concert to perform specific biological functions through protein-protein interactions (PPIs) that occur in time and space. However, the underlying module organization and variance remain unclear. In this study, we collected module templates to infer respective module families, including 58,041 homologous modules in 1,678 species, and PPI families using searches of complete genomic database. We then derived PPI evolution scores and interface evolution scores to describe the module elements, including core and ring components. Functions of core components were highly correlated with those of essential genes. In comparison with ring components, core proteins/PPIs were conserved across multiple species. Subsequently, protein/module variance of PPI networks confirmed that core components form dynamic network hubs and play key roles in various biological functions. Based on the analyses of gene essentiality, module variance, and gene co-expression, we summarize the observations of module organization and variance as follows: 1) a module consists of core and ring components; 2) core components perform major biological functions and collaborate with ring components to execute certain functions in some cases; 3) core components are more conserved and essential during organizational changes in different biological states or conditions.

  3. Posttranslational Protein Modifications in Plant Metabolism1

    PubMed Central

    Friso, Giulia; van Wijk, Klaas J.

    2015-01-01

    Posttranslational modifications (PTMs) of proteins greatly expand proteome diversity, increase functionality, and allow for rapid responses, all at relatively low costs for the cell. PTMs play key roles in plants through their impact on signaling, gene expression, protein stability and interactions, and enzyme kinetics. Following a brief discussion of the experimental and bioinformatics challenges of PTM identification, localization, and quantification (occupancy), a concise overview is provided of the major PTMs and their (potential) functional consequences in plants, with emphasis on plant metabolism. Classic examples that illustrate the regulation of plant metabolic enzymes and pathways by PTMs and their cross talk are summarized. Recent large-scale proteomics studies mapped many PTMs to a wide range of metabolic functions. Unraveling of the PTM code, i.e. a predictive understanding of the (combinatorial) consequences of PTMs, is needed to convert this growing wealth of data into an understanding of plant metabolic regulation. PMID:26338952

  4. The AT-hook motif-encoding gene METABOLIC NETWORK MODULATOR 1 underlies natural variation in Arabidopsis primary metabolism

    PubMed Central

    Li, Baohua; Kliebenstein, Daniel J.

    2014-01-01

    Regulation of primary metabolism is a central mechanism by which plants coordinate their various responses to biotic and abiotic challenge. To identify genes responsible for natural variation in primary metabolism, we focused on cloning a locus from Arabidopsis thaliana that influences the level of TCA cycle metabolites in planta. We found that the Met.V.67 locus was controlled by natural variation in METABOLIC NETWORK MODULATOR 1 (MNM1), which encoded an AT-hook motif-containing protein that was unique to the Brassicales lineage. MNM1 had wide ranging effects on plant metabolism and displayed a tissue expression pattern that was suggestive of a function in sink tissues. Natural variation within MNM1 had differential effects during a diurnal time course, and this temporal dependency was supported by analysis of T-DNA insertion and over-expression lines for MNM1. Thus, the cloning of a natural variation locus specifically associated with primary metabolism allowed us to identify MNM1 as a lineage-specific modulator of primary metabolism, suggesting that the regulation of primary metabolism can change during evolution. PMID:25202318

  5. Quercetin modulates keratoconus metabolism in vitro

    PubMed Central

    McKay, Tina B; Sarker-Nag, Akhee; Lyon, Desiree’; Asara, John M; Karamichos, Dimitrios

    2016-01-01

    Corneal scarring is the result of a disease, infection or injury. The resulting scars cause significant loss of vision or even blindness. To-date, the most successful treatment is corneal transplantation, but it does not come without side effects. One of the corneal dystrophies that are correlated with corneal scarring is keratoconus (KC). The onset of the disease is still unknown; however, altered cellular metabolism has been linked to promoting the fibrotic phenotype and therefore scarring. We have previously shown that human keratoconus cells (HKCs) have altered metabolic activity when compared to normal human corneal fibroblasts (HCFs). In our current study, we present evidence that quercetin, a natural flavonoid, is a strong candidate for regulating metabolic activity of both HCFs and HKCs in vitro and therefore a potential therapeutic to target the altered cellular metabolism characteristic of HKCs. Targeted mass spectrometry-based metabolomics was performed on HCFs and HKCs with and without quercetin treatment in order to identify variations in metabolite flux. Overall, our study reveals a novel therapeutic target OF Quercetin on corneal stromal cell metabolism in both healthy and diseased states. Clearly, further studies are necessary in order to dissect the mechanism of action of quercetin. PMID:26173740

  6. Hierarchical decomposition of metabolic networks using k-modules.

    PubMed

    Reimers, Arne C

    2015-12-01

    The optimal solutions obtained by flux balance analysis (FBA) are typically not unique. Flux modules have recently been shown to be a very useful tool to simplify and decompose the space of FBA-optimal solutions. Since yield-maximization is sometimes not the primary objective encountered in vivo, we are also interested in understanding the space of sub-optimal solutions. Unfortunately, the flux modules are too restrictive and not suited for this task. We present a generalization, called k-module, which compensates the limited applicability of flux modules to the space of sub-optimal solutions. Intuitively, a k-module is a sub-network with low connectivity to the rest of the network. Recursive application of k-modules yields a hierarchical decomposition of the metabolic network, which is also known as branch decomposition in matroid theory. In particular, decompositions computed by existing methods, like the null-space-based approach, introduced by Poolman et al. [(2007) J. Theor. Biol. 249: , 691-705] can be interpreted as branch decompositions. With k-modules we can now compare alternative decompositions of metabolic networks to the classical sub-systems of glycolysis, tricarboxylic acid (TCA) cycle, etc. They can be used to speed up algorithmic problems [theoretically shown for elementary flux modes (EFM) enumeration] and have the potential to present computational solutions in a more intuitive way independently from the classical sub-systems.

  7. HDAC Inhibition Modulates Cardiac PPARs and Fatty Acid Metabolism in Diabetic Cardiomyopathy.

    PubMed

    Lee, Ting-I; Kao, Yu-Hsun; Tsai, Wen-Chin; Chung, Cheng-Chih; Chen, Yao-Chang; Chen, Yi-Jen

    2016-01-01

    Peroxisome proliferator-activated receptors (PPARs) regulate cardiac glucose and lipid homeostasis. Histone deacetylase (HDAC) inhibitor has anti-inflammatory effects which may play a key role in modulating PPARs and fatty acid metabolism. The aim of this study was to investigate whether HDAC inhibitor, MPT0E014, can modulate myocardial PPARs, inflammation, and fatty acid metabolism in diabetes mellitus (DM) cardiomyopathy. Electrocardiography, echocardiography, and western blotting were used to evaluate the electrophysiological activity, cardiac structure, fatty acid metabolism, inflammation, and PPAR isoform expressions in the control and streptozotocin-nicotinamide-induced DM rats with or without MPT0E014. Compared to control, DM and MPT0E014-treated DM rats had elevated blood glucose levels and lower body weights. However, MPT0E014-treated DM and control rats had smaller left ventricular end-diastolic diameter and shorter QT interval than DM rats. The control and MPT0E014-treated DM rats had greater cardiac PPAR-α and PPAR-δ protein expressions, but less cardiac PPAR-γ than DM rats. Moreover, control and MPT0E014-treated DM rats had lower concentrations of 5' adenosine monophosphate-activated protein kinase 2α, PPAR-γ coactivator 1α, phosphorylated acetyl CoA carboxylase, cluster of differentiation 36, diacylglycerol acyltransferase 1 (DGAT1), DGAT2, tumor necrosis factor-α, and interleukin-6 protein than DM rats. HDAC inhibition significantly attenuated DM cardiomyopathy through modulation of cardiac PPARS, fatty acid metabolism, and proinflammatory cytokines. PMID:27446205

  8. Positive modulator of bone morphogenic protein-2

    DOEpatents

    Zamora, Paul O.; Pena, Louis A.; Lin, Xinhua; Takahashi, Kazuyuki

    2009-01-27

    Compounds of the present invention of formula I and formula II are disclosed in the specification and wherein the compounds are modulators of Bone Morphogenic Protein activity. Compounds are synthetic peptides having a non-growth factor heparin binding region, a linker, and sequences that bind specifically to a receptor for Bone Morphogenic Protein. Uses of compounds of the present invention in the treatment of bone lesions, degenerative joint disease and to enhance bone formation are disclosed.

  9. Effects of metabolic rate on protein evolution.

    PubMed

    Gillooly, James F; McCoy, Michael W; Allen, Andrew P

    2007-12-22

    Since the modern evolutionary synthesis was first proposed early in the twentieth century, attention has focused on assessing the relative contribution of mutation versus natural selection on protein evolution. Here we test a model that yields general quantitative predictions on rates of protein evolution by combining principles of individual energetics with Kimura's neutral theory. The model successfully predicts much of the heterogeneity in rates of protein evolution for diverse eukaryotes (i.e. fishes, amphibians, reptiles, birds, mammals) from different thermal environments. Data also show that the ratio of non-synonymous to synonymous nucleotide substitution is independent of body size, and thus presumably of effective population size. These findings indicate that rates of protein evolution are largely controlled by mutation rates, which in turn are strongly influenced by individual metabolic rate.

  10. Complex Reconstitution from Individual Protein Modules.

    PubMed

    Basquin, Jérôme; Taschner, Michael; Lorentzen, Esben

    2016-01-01

    Cellular function relies on protein complexes that work as nano-machines. The structure and function of protein complexes is an outcome of the specific combination of protein subunits, or modules, within the complex. A major focus of molecular biology is thus to understand how protein subunits assemble to form complexes with distinct biological function. To this end, in vitro reconstitution of complexes from individual subunits to study their assembly, structure and activity is of central importance. With purified individual subunits and sub-modules at hand one can systematically dissect the hierarchical assembly of larger complexes using direct protein-protein interaction assays. Furthermore, activity assays can be carried out with individual subunits or smaller sub-complexes and compared to those of the fully assembled complex to precisely map functional sites and provide a molecular basis for in vivo observations. In this chapter we review methods for protein complex assembly from individual subunits and provide examples of advantages and potential pitfalls to this approach. PMID:27165333

  11. Chemoprotective activity of boldine: modulation of drug-metabolizing enzymes.

    PubMed

    Kubínová, R; Machala, M; Minksová, K; Neca, J; Suchý, V

    2001-03-01

    Possible chemoprotective effects of the naturally occurring alkaloid boldine, a major alkaloid of boldo (Peumus boldus Mol.) leaves and bark, including in vitro modulations of drug-metabolizing enzymes in mouse hepatoma Hepa-1 cell line and mouse hepatic microsomes, were investigated. Boldine manifested inhibition activity on hepatic microsomal CYP1A-dependent 7-ethoxyresorufin O-deethylase and CYP3A-dependent testosterone 6 beta-hydroxylase activities and stimulated glutathione S-transferase activity in Hepa-1 cells. In addition to the known antioxidant activity, boldine could decrease the metabolic activation of other xenobiotics including chemical mutagens. PMID:11265593

  12. Mitochondrial uncoupling proteins and energy metabolism

    PubMed Central

    Busiello, Rosa A.; Savarese, Sabrina; Lombardi, Assunta

    2015-01-01

    Understanding the metabolic factors that contribute to energy metabolism (EM) is critical for the development of new treatments for obesity and related diseases. Mitochondrial oxidative phosphorylation is not perfectly coupled to ATP synthesis, and the process of proton-leak plays a crucial role. Proton-leak accounts for a significant part of the resting metabolic rate (RMR) and therefore enhancement of this process represents a potential target for obesity treatment. Since their discovery, uncoupling proteins have stimulated great interest due to their involvement in mitochondrial-inducible proton-leak. Despite the widely accepted uncoupling/thermogenic effect of uncoupling protein one (UCP1), which was the first in this family to be discovered, the reactions catalyzed by its homolog UCP3 and the physiological role remain under debate. This review provides an overview of the role played by UCP1 and UCP3 in mitochondrial uncoupling/functionality as well as EM and suggests that they are a potential therapeutic target for treating obesity and its related diseases such as type II diabetes mellitus. PMID:25713540

  13. Protein acetylation in metabolism - metabolites and cofactors.

    PubMed

    Menzies, Keir J; Zhang, Hongbo; Katsyuba, Elena; Auwerx, Johan

    2016-01-01

    Reversible acetylation was initially described as an epigenetic mechanism regulating DNA accessibility. Since then, this process has emerged as a controller of histone and nonhistone acetylation that integrates key physiological processes such as metabolism, circadian rhythm and cell cycle, along with gene regulation in various organisms. The widespread and reversible nature of acetylation also revitalized interest in the mechanisms that regulate lysine acetyltransferases (KATs) and deacetylases (KDACs) in health and disease. Changes in protein or histone acetylation are especially relevant for many common diseases including obesity, diabetes mellitus, neurodegenerative diseases and cancer, as well as for some rare diseases such as mitochondrial diseases and lipodystrophies. In this Review, we examine the role of reversible acetylation in metabolic control and how changes in levels of metabolites or cofactors, including nicotinamide adenine dinucleotide, nicotinamide, coenzyme A, acetyl coenzyme A, zinc and butyrate and/or β-hydroxybutyrate, directly alter KAT or KDAC activity to link energy status to adaptive cellular and organismal homeostasis.

  14. Effect of acute heat stress on plant nutrient metabolism proteins

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Abrupt heating decreased the levels (per unit total root protein) of all but one of the nutrient metabolism proteins examined, and for most of the proteins, effects were greater for severe vs. moderate heat stress. For many of the nutrient metabolism proteins, initial effects of heat (1 d) were r...

  15. Metabolic Inflammation-Differential Modulation by Dietary Constituents

    PubMed Central

    Lyons, Claire L.; Kennedy, Elaine B.; Roche, Helen M.

    2016-01-01

    Obesity arises from a sustained positive energy balance which triggers a pro-inflammatory response, a key contributor to metabolic diseases such as T2D. Recent studies, focused on the emerging area of metabolic-inflammation, highlight that specific metabolites can modulate the functional nature and inflammatory phenotype of immune cells. In obesity, expanding adipose tissue attracts immune cells, creating an inflammatory environment within this fatty acid storage organ. Resident immune cells undergo both a pro-inflammatory and metabolic switch in their function. Inflammatory mediators, such as TNF-α and IL-1β, are induced by saturated fatty acids and disrupt insulin signaling. Conversely, monounsaturated and polyunsaturated fatty acids do not interrupt metabolism and inflammation to the same extent. AMPK links inflammation, metabolism and T2D, with roles to play in all and is influenced negatively by obesity. Lipid spillover results in hepatic lipotoxicity and steatosis. Also in skeletal muscle, excessive FFA can impede insulin’s action and promote inflammation. Ectopic fat can also affect pancreatic β-cell function, thereby contributing to insulin resistance. Therapeutics, lifestyle changes, supplements and dietary manipulation are all possible avenues to combat metabolic inflammation and the subsequent insulin resistant state which will be explored in the current review. PMID:27128935

  16. Metabolic Inflammation-Differential Modulation by Dietary Constituents.

    PubMed

    Lyons, Claire L; Kennedy, Elaine B; Roche, Helen M

    2016-01-01

    Obesity arises from a sustained positive energy balance which triggers a pro-inflammatory response, a key contributor to metabolic diseases such as T2D. Recent studies, focused on the emerging area of metabolic-inflammation, highlight that specific metabolites can modulate the functional nature and inflammatory phenotype of immune cells. In obesity, expanding adipose tissue attracts immune cells, creating an inflammatory environment within this fatty acid storage organ. Resident immune cells undergo both a pro-inflammatory and metabolic switch in their function. Inflammatory mediators, such as TNF-α and IL-1β, are induced by saturated fatty acids and disrupt insulin signaling. Conversely, monounsaturated and polyunsaturated fatty acids do not interrupt metabolism and inflammation to the same extent. AMPK links inflammation, metabolism and T2D, with roles to play in all and is influenced negatively by obesity. Lipid spillover results in hepatic lipotoxicity and steatosis. Also in skeletal muscle, excessive FFA can impede insulin's action and promote inflammation. Ectopic fat can also affect pancreatic β-cell function, thereby contributing to insulin resistance. Therapeutics, lifestyle changes, supplements and dietary manipulation are all possible avenues to combat metabolic inflammation and the subsequent insulin resistant state which will be explored in the current review. PMID:27128935

  17. Metabolic Inflammation-Differential Modulation by Dietary Constituents.

    PubMed

    Lyons, Claire L; Kennedy, Elaine B; Roche, Helen M

    2016-04-27

    Obesity arises from a sustained positive energy balance which triggers a pro-inflammatory response, a key contributor to metabolic diseases such as T2D. Recent studies, focused on the emerging area of metabolic-inflammation, highlight that specific metabolites can modulate the functional nature and inflammatory phenotype of immune cells. In obesity, expanding adipose tissue attracts immune cells, creating an inflammatory environment within this fatty acid storage organ. Resident immune cells undergo both a pro-inflammatory and metabolic switch in their function. Inflammatory mediators, such as TNF-α and IL-1β, are induced by saturated fatty acids and disrupt insulin signaling. Conversely, monounsaturated and polyunsaturated fatty acids do not interrupt metabolism and inflammation to the same extent. AMPK links inflammation, metabolism and T2D, with roles to play in all and is influenced negatively by obesity. Lipid spillover results in hepatic lipotoxicity and steatosis. Also in skeletal muscle, excessive FFA can impede insulin's action and promote inflammation. Ectopic fat can also affect pancreatic β-cell function, thereby contributing to insulin resistance. Therapeutics, lifestyle changes, supplements and dietary manipulation are all possible avenues to combat metabolic inflammation and the subsequent insulin resistant state which will be explored in the current review.

  18. Dysregulation of skeletal muscle protein metabolism by alcohol

    PubMed Central

    Steiner, Jennifer L.

    2015-01-01

    Alcohol abuse, either by acute intoxication or prolonged excessive consumption, leads to pathological changes in many organs and tissues including skeletal muscle. As muscle protein serves not only a contractile function but also as a metabolic reserve for amino acids, which are used to support the energy needs of other tissues, its content is tightly regulated and dynamic. This review focuses on the etiology by which alcohol perturbs skeletal muscle protein balance and thereby over time produces muscle wasting and weakness. The preponderance of data suggest that alcohol primarily impairs global protein synthesis, under basal conditions as well as in response to several anabolic stimuli including growth factors, nutrients, and muscle contraction. This inhibitory effect of alcohol is mediated, at least in part, by a reduction in mTOR kinase activity via a mechanism that remains poorly defined but likely involves altered protein-protein interactions within mTOR complex 1. Furthermore, alcohol can exacerbate the decrement in mTOR and/or muscle protein synthesis present in other catabolic states. In contrast, alcohol-induced changes in muscle protein degradation, either global or via specific modulation of the ubiquitin-proteasome or autophagy pathways, are relatively inconsistent and may be model dependent. Herein, changes produced by acute intoxication versus chronic ingestion are contrasted in relation to skeletal muscle metabolism, and limitations as well as opportunities for future research are discussed. As the proportion of more economically developed countries ages and chronic illness becomes more prevalent, a better understanding of the etiology of biomedical consequences of alcohol use disorders is warranted. PMID:25759394

  19. Foldons, Protein Structural Modules, and Exons

    NASA Astrophysics Data System (ADS)

    Panchenko, Anna R.; Luthey-Schulten, Zaida; Wolynes, Peter G.

    1996-03-01

    Foldons, which are kinetically competent, quasi-independently folding units of a protein, may be defined using energy landscape analysis. Foldons can be identified by maxima in a scan of the ratio of a contiguous segment's energetic stability gap to the energy variance of that segment's molten globule states, reflecting the requirement of minimal frustration. The predicted foldons are compared with the exons and structural modules for 16 of the 30 proteins studied. Statistical analysis indicates a strong correlation between the energetically determined foldons and Go's geometrically defined structural modules, but there are marked sequence-dependent effects. There is only a weak correlation of foldons to exons. For γ II-crystallin, myoglobin, barnase, α -lactalbumin, and cytochrome c the foldons and some noncontiguous clusters of foldons compare well with intermediates observed in experiment.

  20. Deciphering peculiar protein-protein interacting modules in Deinococcus radiodurans

    PubMed Central

    Mezhoud, Karim; Sghaier, Haïtham; Barkallah, Insaf

    2009-01-01

    Interactomes of proteins under positive selection from ionizing-radiation-resistant bacteria (IRRB) might be a part of the answer to the question as to how IRRB, particularly Deinococcus radiodurans R1 (Deira), resist ionizing radiation. Here, using the Database of Interacting Proteins (DIP) and the Protein Structural Interactome (PSI)-base server for PSI map, we have predicted novel interactions of orthologs of the 58 proteins under positive selection in Deira and other IRRB, but which are absent in IRSB. Among these, 18 domains and their interactomes have been identified in DNA checkpoint and repair; kinases pathways; energy and nucleotide metabolisms were the important biological processes that were found to be involved. This finding provides new clues to the cellular pathways that can to be important for ionizing-radiation resistance in Deira. PMID:19356244

  1. Apolipoprotein A-IV: a protein intimately involved in metabolism

    PubMed Central

    Wang, Fei; Kohan, Alison B.; Lo, Chun-Min; Liu, Min; Howles, Philip; Tso, Patrick

    2015-01-01

    The purpose of this review is to summarize our current understanding of the physiological roles of apoA-IV in metabolism, and to underscore the potential for apoA-IV to be a focus for new therapies aimed at the treatment of diabetes and obesity-related disorders. ApoA-IV is primarily synthesized by the small intestine, attached to chylomicrons by enterocytes, and secreted into intestinal lymph during fat absorption. In circulation, apoA-IV is associated with HDL and chylomicron remnants, but a large portion is lipoprotein free. Due to its anti-oxidative and anti-inflammatory properties, and because it can mediate reverse-cholesterol transport, proposed functions of circulating apoA-IV have been related to protection from cardiovascular disease. This review, however, focuses primarily on several properties of apoA-IV that impact other metabolic functions related to food intake, obesity, and diabetes. In addition to participating in triglyceride absorption, apoA-IV can act as an acute satiation factor through both peripheral and central routes of action. It also modulates glucose homeostasis through incretin-like effects on insulin secretion, and by moderating hepatic glucose production. While apoA-IV receptors remain to be conclusively identified, the latter modes of action suggest that this protein holds therapeutic promise for treating metabolic disease. PMID:25640749

  2. Pathogen mimicry of host protein-protein interfaces modulates immunity.

    PubMed

    Guven-Maiorov, Emine; Tsai, Chung-Jung; Nussinov, Ruth

    2016-10-01

    Signaling pathways shape and transmit the cell's reaction to its changing environment; however, pathogens can circumvent this response by manipulating host signaling. To subvert host defense, they beat it at its own game: they hijack host pathways by mimicking the binding surfaces of host-encoded proteins. For this, it is not necessary to achieve global protein homology; imitating merely the interaction surface is sufficient. Different protein folds often interact via similar protein-protein interface architectures. This similarity in binding surfaces permits the pathogenic protein to compete with a host target protein. Thus, rather than binding a host-encoded partner, the host protein hub binds the pathogenic surrogate. The outcome can be dire: rewiring or repurposing the host pathways, shifting the cell signaling landscape and consequently the immune response. They can also cause persistent infections as well as cancer by modulating key signaling pathways, such as those involving Ras. Mapping the rewired host-pathogen 'superorganism' interaction network - along with its structural details - is critical for in-depth understanding of pathogenic mechanisms and developing efficient therapeutics. Here, we overview the role of molecular mimicry in pathogen host evasion as well as types of molecular mimicry mechanisms that emerged during evolution.

  3. Metabolic pathways and activity-dependent modulation of glutamate concentration in the human brain.

    PubMed

    Mangia, Silvia; Giove, Federico; Dinuzzo, Mauro

    2012-11-01

    Glutamate is one of the most versatile molecules present in the human brain, involved in protein synthesis, energy production, ammonia detoxification, and transport of reducing equivalents. Aside from these critical metabolic roles, glutamate plays a major part in brain function, being not only the most abundant excitatory neurotransmitter, but also the precursor for γ-aminobutyric acid, the predominant inhibitory neurotransmitter. Regulation of glutamate levels is pivotal for normal brain function, as abnormal extracellular concentration of glutamate can lead to impaired neurotransmission, neurodegeneration and even neuronal death. Understanding how the neuron-astrocyte functional and metabolic interactions modulate glutamate concentration during different activation status and under physiological and pathological conditions is a challenging task, and can only be tentatively estimated from current literature. In this paper, we focus on describing the various metabolic pathways which potentially affect glutamate concentration in the brain, and emphasize which ones are likely to produce the variations in glutamate concentration observed during enhanced neuronal activity in human studies.

  4. HDAC Inhibition Modulates Cardiac PPARs and Fatty Acid Metabolism in Diabetic Cardiomyopathy

    PubMed Central

    Lee, Ting-I; Tsai, Wen-Chin; Chung, Cheng-Chih; Chen, Yao-Chang; Chen, Yi-Jen

    2016-01-01

    Peroxisome proliferator-activated receptors (PPARs) regulate cardiac glucose and lipid homeostasis. Histone deacetylase (HDAC) inhibitor has anti-inflammatory effects which may play a key role in modulating PPARs and fatty acid metabolism. The aim of this study was to investigate whether HDAC inhibitor, MPT0E014, can modulate myocardial PPARs, inflammation, and fatty acid metabolism in diabetes mellitus (DM) cardiomyopathy. Electrocardiography, echocardiography, and western blotting were used to evaluate the electrophysiological activity, cardiac structure, fatty acid metabolism, inflammation, and PPAR isoform expressions in the control and streptozotocin-nicotinamide-induced DM rats with or without MPT0E014. Compared to control, DM and MPT0E014-treated DM rats had elevated blood glucose levels and lower body weights. However, MPT0E014-treated DM and control rats had smaller left ventricular end-diastolic diameter and shorter QT interval than DM rats. The control and MPT0E014-treated DM rats had greater cardiac PPAR-α and PPAR-δ protein expressions, but less cardiac PPAR-γ than DM rats. Moreover, control and MPT0E014-treated DM rats had lower concentrations of 5′ adenosine monophosphate-activated protein kinase 2α, PPAR-γ coactivator 1α, phosphorylated acetyl CoA carboxylase, cluster of differentiation 36, diacylglycerol acyltransferase 1 (DGAT1), DGAT2, tumor necrosis factor-α, and interleukin-6 protein than DM rats. HDAC inhibition significantly attenuated DM cardiomyopathy through modulation of cardiac PPARS, fatty acid metabolism, and proinflammatory cytokines. PMID:27446205

  5. Modulation of tau protein fibrillization by oleocanthal.

    PubMed

    Monti, Maria Chiara; Margarucci, Luigi; Riccio, Raffaele; Casapullo, Agostino

    2012-09-28

    Among the phenolic compounds extracted from extra virgin olive oil, oleocanthal (1) has attracted considerable attention in the modulation of many human diseases, such as inflammation and Alzheimer's disease (AD). Indeed, 1 is capable of altering the fibrillization of tau protein, which is one of the key factors at the basis of neurodegenerative diseases, and of covalently reacting with lysine ε-amino groups of the tau fragment K18 in an unspecific fashion. In the present study, an investigation of the recognition process and the reaction profile between 1 and the wild-type tau protein has been conducted by a circular dichroism, surface plasmon resonance, fluorescence, and mass spectrometry combined approach. As a result, 1 has been found to interact with tau-441, inducing stable conformational modifications of the protein secondary structure and also interfering with tau aggregation. These findings provide experimental support for the potential reduced risk of AD and related neurodegenerative diseases associated with olive oil consumption and may offer a new chemical scaffold for the development of AD-modulating agents. PMID:22988908

  6. Protein acetylation in metabolism - metabolites and cofactors.

    PubMed

    Menzies, Keir J; Zhang, Hongbo; Katsyuba, Elena; Auwerx, Johan

    2016-01-01

    Reversible acetylation was initially described as an epigenetic mechanism regulating DNA accessibility. Since then, this process has emerged as a controller of histone and nonhistone acetylation that integrates key physiological processes such as metabolism, circadian rhythm and cell cycle, along with gene regulation in various organisms. The widespread and reversible nature of acetylation also revitalized interest in the mechanisms that regulate lysine acetyltransferases (KATs) and deacetylases (KDACs) in health and disease. Changes in protein or histone acetylation are especially relevant for many common diseases including obesity, diabetes mellitus, neurodegenerative diseases and cancer, as well as for some rare diseases such as mitochondrial diseases and lipodystrophies. In this Review, we examine the role of reversible acetylation in metabolic control and how changes in levels of metabolites or cofactors, including nicotinamide adenine dinucleotide, nicotinamide, coenzyme A, acetyl coenzyme A, zinc and butyrate and/or β-hydroxybutyrate, directly alter KAT or KDAC activity to link energy status to adaptive cellular and organismal homeostasis. PMID:26503676

  7. Metabolic effects of milk protein intake strongly depend on pre-existing metabolic and exercise status.

    PubMed

    Melnik, Bodo C; Schmitz, Gerd; John, Swen; Carrera-Bastos, Pedro; Lindeberg, Staffan; Cordain, Loren

    2013-01-01

    Milk protein intake has recently been suggested to improve metabolic health. This Perspective provides evidence that metabolic effects of milk protein intake have to be regarded in the context of the individual's pre-existing metabolic and exercise status. Milk proteins provide abundant branched-chain amino acids (BCAAs) and glutamine. Plasma BCAAs and glutamine are increased in obesity and insulin resistance, but decrease after gastric bypass surgery resulting in weight loss and improved insulin sensitivity. Milk protein consumption results in postprandial hyperinsulinemia in obese subjects, increases body weight of overweight adolescents and may thus deteriorate pre-existing metabolic disturbances of obese, insulin resistant individuals. PMID:24225036

  8. Metabolic effects of milk protein intake strongly depend on pre-existing metabolic and exercise status

    PubMed Central

    2013-01-01

    Milk protein intake has recently been suggested to improve metabolic health. This Perspective provides evidence that metabolic effects of milk protein intake have to be regarded in the context of the individual’s pre-existing metabolic and exercise status. Milk proteins provide abundant branched-chain amino acids (BCAAs) and glutamine. Plasma BCAAs and glutamine are increased in obesity and insulin resistance, but decrease after gastric bypass surgery resulting in weight loss and improved insulin sensitivity. Milk protein consumption results in postprandial hyperinsulinemia in obese subjects, increases body weight of overweight adolescents and may thus deteriorate pre-existing metabolic disturbances of obese, insulin resistant individuals. PMID:24225036

  9. Modulation of lipid metabolic defects rescues cleft palate in Tgfbr2 mutant mice.

    PubMed

    Iwata, Junichi; Suzuki, Akiko; Pelikan, Richard C; Ho, Thach-Vu; Sanchez-Lara, Pedro A; Chai, Yang

    2014-01-01

    Mutations in transforming growth factor beta (TGFβ) receptor type II (TGFBR2) cause Loeys-Dietz syndrome, characterized by craniofacial and cardiovascular abnormalities. Mice with a deletion of Tgfbr2 in cranial neural crest cells (Tgfbr2(fl/fl);Wnt1-Cre mice) develop cleft palate as the result of abnormal TGFβ signaling activation. However, little is known about metabolic processes downstream of TGFβ signaling during palatogenesis. Here, we show that Tgfbr2 mutant palatal mesenchymal cells spontaneously accumulate lipid droplets, resulting from reduced lipolysis activity. Tgfbr2 mutant palatal mesenchymal cells failed to respond to the cell proliferation stimulator sonic hedgehog, derived from the palatal epithelium. Treatment with p38 mitogen-activated protein kinase (MAPK) inhibitor or telmisartan, a modulator of p38 MAPK activation and lipid metabolism, blocked abnormal TGFβ-mediated p38 MAPK activation, restoring lipid metabolism and cell proliferation activity both in vitro and in vivo. Our results highlight the influence of alternative TGFβ signaling on lipid metabolic activities, as well as how lipid metabolic defects can affect cell proliferation and adversely impact palatogenesis. This discovery has broader implications for the understanding of metabolic defects and potential prevention of congenital birth defects. PMID:23975680

  10. Relationship between asparagine metabolism and protein concentration in soybean seed

    Technology Transfer Automated Retrieval System (TEKTRAN)

    The relationship between asparagine metabolism and protein concentration was investigated in soybean seed. Phenotyping of a population of recombinant inbred lines adapted to Illinois confirmed a positive correlation between free asparagine levels in developing seeds and protein concentration at matu...

  11. Type IV Pilin Proteins: Versatile Molecular Modules

    PubMed Central

    Giltner, Carmen L.; Nguyen, Ylan

    2012-01-01

    Summary: Type IV pili (T4P) are multifunctional protein fibers produced on the surfaces of a wide variety of bacteria and archaea. The major subunit of T4P is the type IV pilin, and structurally related proteins are found as components of the type II secretion (T2S) system, where they are called pseudopilins; of DNA uptake/competence systems in both Gram-negative and Gram-positive species; and of flagella, pili, and sugar-binding systems in the archaea. This broad distribution of a single protein family implies both a common evolutionary origin and a highly adaptable functional plan. The type IV pilin is a remarkably versatile architectural module that has been adopted widely for a variety of functions, including motility, attachment to chemically diverse surfaces, electrical conductance, acquisition of DNA, and secretion of a broad range of structurally distinct protein substrates. In this review, we consider recent advances in this research area, from structural revelations to insights into diversity, posttranslational modifications, regulation, and function. PMID:23204365

  12. Pharmacological Modulators of Endoplasmic Reticulum Stress in Metabolic Diseases

    PubMed Central

    Jung, Tae Woo; Choi, Kyung Mook

    2016-01-01

    The endoplasmic reticulum (ER) is the principal organelle responsible for correct protein folding, a step in protein synthesis that is critical for the functional conformation of proteins. ER stress is a primary feature of secretory cells and is involved in the pathogenesis of numerous human diseases, such as certain neurodegenerative and cardiometabolic disorders. The unfolded protein response (UPR) is a defense mechanism to attenuate ER stress and maintain the homeostasis of the organism. Two major degradation systems, including the proteasome and autophagy, are involved in this defense system. If ER stress overwhelms the capacity of the cell’s defense mechanisms, apoptotic death may result. This review is focused on the various pharmacological modulators that can protect cells from damage induced by ER stress. The possible mechanisms for cytoprotection are also discussed. PMID:26840310

  13. Vitamin A deficiency modulates iron metabolism via ineffective erythropoiesis.

    PubMed

    da Cunha, Marcela S B; Siqueira, Egle M A; Trindade, Luciano S; Arruda, Sandra F

    2014-10-01

    Vitamin A modulates inflammatory status, iron metabolism and erythropoiesis. Given that these factors modulate the expression of the hormone hepcidin (Hamp), we investigated the effect of vitamin A deficiency on molecular biomarkers of iron metabolism, the inflammatory response and the erythropoietic system. Five groups of male Wistar rats were treated: control (AIN-93G), the vitamin A-deficient (VAD) diet, the iron-deficient (FeD) diet, the vitamin A- and iron-deficient (VAFeD) diet or the diet with 12 mg atRA/kg diet replacing all-trans-retinyl palmitate by all-trans retinoic acid (atRA). Vitamin A deficiency reduced serum iron and transferrin saturation levels, increased spleen iron concentrations, reduced hepatic Hamp and kidney erythropoietin messenger RNA (mRNA) levels and up-regulated hepatic and spleen heme oxygenase-1 gene expression while reducing the liver HO-1 specific activity compared with the control. The FeD and VAFeD rats exhibited lower levels of serum iron and transferrin saturation, lower iron concentrations in tissues and lower hepatic Hamp mRNA levels compared with the control. The treatment with atRA resulted in lower serum iron and transferrin concentrations, an increased iron concentration in the liver, a decreased iron concentration in the spleen and in the gut, and decreased hepatic Hamp mRNA levels. In summary, these findings suggest that vitamin A deficiency leads to ineffective erythropoiesis by the down-regulation of renal erythropoietin expression in the kidney, resulting in erythrocyte malformation and the consequent accumulation of the heme group in the spleen. Vitamin A deficiency indirectly modulates systemic iron homeostasis by enhancing erythrophagocytosis of undifferentiated erythrocytes. PMID:24998947

  14. Vitamin A deficiency modulates iron metabolism via ineffective erythropoiesis.

    PubMed

    da Cunha, Marcela S B; Siqueira, Egle M A; Trindade, Luciano S; Arruda, Sandra F

    2014-10-01

    Vitamin A modulates inflammatory status, iron metabolism and erythropoiesis. Given that these factors modulate the expression of the hormone hepcidin (Hamp), we investigated the effect of vitamin A deficiency on molecular biomarkers of iron metabolism, the inflammatory response and the erythropoietic system. Five groups of male Wistar rats were treated: control (AIN-93G), the vitamin A-deficient (VAD) diet, the iron-deficient (FeD) diet, the vitamin A- and iron-deficient (VAFeD) diet or the diet with 12 mg atRA/kg diet replacing all-trans-retinyl palmitate by all-trans retinoic acid (atRA). Vitamin A deficiency reduced serum iron and transferrin saturation levels, increased spleen iron concentrations, reduced hepatic Hamp and kidney erythropoietin messenger RNA (mRNA) levels and up-regulated hepatic and spleen heme oxygenase-1 gene expression while reducing the liver HO-1 specific activity compared with the control. The FeD and VAFeD rats exhibited lower levels of serum iron and transferrin saturation, lower iron concentrations in tissues and lower hepatic Hamp mRNA levels compared with the control. The treatment with atRA resulted in lower serum iron and transferrin concentrations, an increased iron concentration in the liver, a decreased iron concentration in the spleen and in the gut, and decreased hepatic Hamp mRNA levels. In summary, these findings suggest that vitamin A deficiency leads to ineffective erythropoiesis by the down-regulation of renal erythropoietin expression in the kidney, resulting in erythrocyte malformation and the consequent accumulation of the heme group in the spleen. Vitamin A deficiency indirectly modulates systemic iron homeostasis by enhancing erythrophagocytosis of undifferentiated erythrocytes.

  15. Differential Protein Modulation in Midguts of Aedes aegypti Infected with Chikungunya and Dengue 2 Viruses

    PubMed Central

    Tchankouo-Nguetcheu, Stéphane; Khun, Huot; Pincet, Laurence; Roux, Pascal; Bahut, Muriel; Huerre, Michel; Guette, Catherine; Choumet, Valérie

    2010-01-01

    Background Arthropod borne virus infections cause several emerging and resurgent infectious diseases. Among the diseases caused by arboviruses, dengue and chikungunya are responsible for a high rate of severe human diseases worldwide. The midgut of mosquitoes is the first barrier for pathogen transmission and is a target organ where arboviruses must replicate prior to infecting other organs. A proteomic approach was undertaken to characterize the key virus/vector interactions and host protein modifications that happen in the midgut for viral transmission to eventually take place. Methodology and Principal Findings Using a proteomics differential approach with two-Dimensional Differential in-Gel Electrophoresis (2D-DIGE), we defined the protein modulations in the midgut of Aedes aegypti that were triggered seven days after an oral infection (7 DPI) with dengue 2 (DENV-2) and chikungunya (CHIKV) viruses. Gel profile comparisons showed that the level of 18 proteins was modulated by DENV-2 only and 12 proteins were modulated by CHIKV only. Twenty proteins were regulated by both viruses in either similar or different ways. Both viruses caused an increase of proteins involved in the generation of reactive oxygen species, energy production, and carbohydrate and lipid metabolism. Midgut infection by DENV-2 and CHIKV triggered an antioxidant response. CHIKV infection produced an increase of proteins involved in detoxification. Conclusion/Significance Our study constitutes the first analysis of the protein response of Aedes aegypti's midgut infected with viruses belonging to different families. It shows that the differentially regulated proteins in response to viral infection include structural, redox, regulatory proteins, and enzymes for several metabolic pathways. Some of these proteins like antioxidant are probably involved in cell protection. On the other hand, we propose that the modulation of other proteins like transferrin, hsp60 and alpha glucosidase, may favour

  16. Chemical Reporter for Visualizing Metabolic Cross-Talk between Carbohydrate Metabolism and Protein Modification

    PubMed Central

    2015-01-01

    Metabolic chemical reporters have been largely used to study posttranslational modifications. Generally, it was assumed that these reporters entered one biosynthetic pathway, resulting in labeling of one type of modification. However, because they are metabolized by cells before their addition onto proteins, metabolic chemical reporters potentially provide a unique opportunity to read-out on both modifications of interest and cellular metabolism. We report here the development of a metabolic chemical reporter 1-deoxy-N-pentynyl glucosamine (1-deoxy-GlcNAlk). This small-molecule cannot be incorporated into glycans; however, treatment of mammalian cells results in labeling of a variety proteins and enables their visualization and identification. Competition of this labeling with sodium acetate and an acetyltransferase inhibitor suggests that 1-deoxy-GlcNAlk can enter the protein acetylation pathway. These results demonstrate that metabolic chemical reporters have the potential to isolate and potentially discover cross-talk between metabolic pathways in living cells. PMID:25062036

  17. Chemical reporter for visualizing metabolic cross-talk between carbohydrate metabolism and protein modification.

    PubMed

    Zaro, Balyn W; Chuh, Kelly N; Pratt, Matthew R

    2014-09-19

    Metabolic chemical reporters have been largely used to study posttranslational modifications. Generally, it was assumed that these reporters entered one biosynthetic pathway, resulting in labeling of one type of modification. However, because they are metabolized by cells before their addition onto proteins, metabolic chemical reporters potentially provide a unique opportunity to read-out on both modifications of interest and cellular metabolism. We report here the development of a metabolic chemical reporter 1-deoxy-N-pentynyl glucosamine (1-deoxy-GlcNAlk). This small-molecule cannot be incorporated into glycans; however, treatment of mammalian cells results in labeling of a variety proteins and enables their visualization and identification. Competition of this labeling with sodium acetate and an acetyltransferase inhibitor suggests that 1-deoxy-GlcNAlk can enter the protein acetylation pathway. These results demonstrate that metabolic chemical reporters have the potential to isolate and potentially discover cross-talk between metabolic pathways in living cells.

  18. Protein Quality Control and Metabolism: Bidirectional Control in the Heart

    PubMed Central

    Wang, Zhao V.; Hill, Joseph A.

    2015-01-01

    The prevalence of heart disease, especially heart failure, continues to increase, and cardiovascular disease remains the leading cause of death worldwide. As cardiomyocytes are essentially irreplaceable, protein quality control is pivotal to cellular homeostasis and, ultimately, cardiac performance. Three evolutionarily conserved mechanisms – autophagy, the unfolded protein response, and the ubiquitin-proteasome system– act in concert to degrade misfolded proteins and eliminate defective organelles. Recent advances have revealed that these mechanisms are intimately associated with cellular metabolism. Going forward, comprehensive understanding of the role of protein quality control mechanisms in cardiac pathology will require integration of metabolic pathways and metabolic control. PMID:25651176

  19. Network integration of parallel metabolic and transcriptional data reveals metabolic modules that regulate macrophage polarization.

    PubMed

    Jha, Abhishek K; Huang, Stanley Ching-Cheng; Sergushichev, Alexey; Lampropoulou, Vicky; Ivanova, Yulia; Loginicheva, Ekaterina; Chmielewski, Karina; Stewart, Kelly M; Ashall, Juliet; Everts, Bart; Pearce, Edward J; Driggers, Edward M; Artyomov, Maxim N

    2015-03-17

    Macrophage polarization involves a coordinated metabolic and transcriptional rewiring that is only partially understood. By using an integrated high-throughput transcriptional-metabolic profiling and analysis pipeline, we characterized systemic changes during murine macrophage M1 and M2 polarization. M2 polarization was found to activate glutamine catabolism and UDP-GlcNAc-associated modules. Correspondingly, glutamine deprivation or inhibition of N-glycosylation decreased M2 polarization and production of chemokine CCL22. In M1 macrophages, we identified a metabolic break at Idh, the enzyme that converts isocitrate to alpha-ketoglutarate, providing mechanistic explanation for TCA cycle fragmentation. (13)C-tracer studies suggested the presence of an active variant of the aspartate-arginosuccinate shunt that compensated for this break. Consistently, inhibition of aspartate-aminotransferase, a key enzyme of the shunt, inhibited nitric oxide and interleukin-6 production in M1 macrophages, while promoting mitochondrial respiration. This systems approach provides a highly integrated picture of the physiological modules supporting macrophage polarization, identifying potential pharmacologic control points for both macrophage phenotypes. PMID:25786174

  20. Network integration of parallel metabolic and transcriptional data reveals metabolic modules that regulate macrophage polarization.

    PubMed

    Jha, Abhishek K; Huang, Stanley Ching-Cheng; Sergushichev, Alexey; Lampropoulou, Vicky; Ivanova, Yulia; Loginicheva, Ekaterina; Chmielewski, Karina; Stewart, Kelly M; Ashall, Juliet; Everts, Bart; Pearce, Edward J; Driggers, Edward M; Artyomov, Maxim N

    2015-03-17

    Macrophage polarization involves a coordinated metabolic and transcriptional rewiring that is only partially understood. By using an integrated high-throughput transcriptional-metabolic profiling and analysis pipeline, we characterized systemic changes during murine macrophage M1 and M2 polarization. M2 polarization was found to activate glutamine catabolism and UDP-GlcNAc-associated modules. Correspondingly, glutamine deprivation or inhibition of N-glycosylation decreased M2 polarization and production of chemokine CCL22. In M1 macrophages, we identified a metabolic break at Idh, the enzyme that converts isocitrate to alpha-ketoglutarate, providing mechanistic explanation for TCA cycle fragmentation. (13)C-tracer studies suggested the presence of an active variant of the aspartate-arginosuccinate shunt that compensated for this break. Consistently, inhibition of aspartate-aminotransferase, a key enzyme of the shunt, inhibited nitric oxide and interleukin-6 production in M1 macrophages, while promoting mitochondrial respiration. This systems approach provides a highly integrated picture of the physiological modules supporting macrophage polarization, identifying potential pharmacologic control points for both macrophage phenotypes.

  1. Protein Phosphorylation: A Major Switch Mechanism for Metabolic Regulation.

    PubMed

    Humphrey, Sean J; James, David E; Mann, Matthias

    2015-12-01

    Metabolism research is undergoing a renaissance because many diseases are increasingly recognized as being characterized by perturbations in intracellular metabolic regulation. Metabolic changes can be conferred through changes to the expression of metabolic enzymes, the concentrations of substrates or products that govern reaction kinetics, or post-translational modification (PTM) of the proteins that facilitate these reactions. On the 60th anniversary since its discovery, reversible protein phosphorylation is widely appreciated as an essential PTM regulating metabolism. With the ability to quantitatively measure dynamic changes in protein phosphorylation on a global scale - hereafter referred to as phosphoproteomics - we are now entering a new era in metabolism research, with mass spectrometry (MS)-based proteomics at the helm. PMID:26498855

  2. Protein design in systems metabolic engineering for industrial strain development.

    PubMed

    Chen, Zhen; Zeng, An-Ping

    2013-05-01

    Accelerating the process of industrial bacterial host strain development, aimed at increasing productivity, generating new bio-products or utilizing alternative feedstocks, requires the integration of complementary approaches to manipulate cellular metabolism and regulatory networks. Systems metabolic engineering extends the concept of classical metabolic engineering to the systems level by incorporating the techniques used in systems biology and synthetic biology, and offers a framework for the development of the next generation of industrial strains. As one of the most useful tools of systems metabolic engineering, protein design allows us to design and optimize cellular metabolism at a molecular level. Here, we review the current strategies of protein design for engineering cellular synthetic pathways, metabolic control systems and signaling pathways, and highlight the challenges of this subfield within the context of systems metabolic engineering.

  3. Ethanol Metabolism Modifies Hepatic Protein Acylation in Mice

    PubMed Central

    Fritz, Kristofer S.; Green, Michelle F.; Petersen, Dennis R.; Hirschey, Matthew D.

    2013-01-01

    Mitochondrial protein acetylation increases in response to chronic ethanol ingestion in mice, and is thought to reduce mitochondrial function and contribute to the pathogenesis of alcoholic liver disease. The mitochondrial deacetylase SIRT3 regulates the acetylation status of several mitochondrial proteins, including those involved in ethanol metabolism. The newly discovered desuccinylase activity of the mitochondrial sirtuin SIRT5 suggests that protein succinylation could be an important post-translational modification regulating mitochondrial metabolism. To assess the possible role of protein succinylation in ethanol metabolism, we surveyed hepatic sub-cellular protein fractions from mice fed a control or ethanol-supplemented diet for succinyl-lysine, as well as acetyl-, propionyl-, and butyryl-lysine post-translational modifications. We found mitochondrial protein propionylation increases, similar to mitochondrial protein acetylation. In contrast, mitochondrial protein succinylation is reduced. These mitochondrial protein modifications appear to be primarily driven by ethanol metabolism, and not by changes in mitochondrial sirtuin levels. Similar trends in acyl modifications were observed in the nucleus. However, comparatively fewer acyl modifications were observed in the cytoplasmic or the microsomal compartments, and were generally unchanged by ethanol metabolism. Using a mass spectrometry proteomics approach, we identified several candidate acetylated, propionylated, and succinylated proteins, which were enriched using antibodies against each modification. Additionally, we identified several acetyl and propionyl lysine residues on the same sites for a number of proteins and supports the idea of the overlapping nature of lysine-specific acylation. Thus, we show that novel post-translational modifications are present in hepatic mitochondrial, nuclear, cytoplasmic, and microsomal compartments and ethanol ingestion, and its associated metabolism, induce specific

  4. Carbon monoxide and mitochondria—modulation of cell metabolism, redox response and cell death

    PubMed Central

    Almeida, Ana S.; Figueiredo-Pereira, Cláudia; Vieira, Helena L. A.

    2015-01-01

    Carbon monoxide (CO) is an endogenously produced gasotransmitter, which is associated with cytoprotection and cellular homeostasis in several distinct cell types and tissues. CO mainly targets mitochondria because: (i) mitochondrial heme-proteins are the main potential candidates for CO to bind, (ii) many CO's biological actions are dependent on mitochondrial ROS signaling and (iii) heme is generated in the mitochondrial compartment. Mitochondria are the key cell energy factory, producing ATP through oxidative phosphorylation and regulating cell metabolism. These organelles are also implicated in many cell signaling pathways and the production of reactive oxygen species (ROS). Finally, mitochondria contain several factors activating programmed cell death pathways, which are released from the mitochondrial inter-membrane space upon mitochondrial membrane permeabilization. Therefore, disclosing CO mode of action at mitochondria opens avenues for deeper understanding CO's biological properties. Herein, it is discussed how CO affects the three main aspects of mitochondrial modulation of cell function: metabolism, redox response and cell death. PMID:25709582

  5. Regulators of G-protein-signaling proteins: negative modulators of G-protein-coupled receptor signaling.

    PubMed

    Woodard, Geoffrey E; Jardín, Isaac; Berna-Erro, A; Salido, Gines M; Rosado, Juan A

    2015-01-01

    Regulators of G-protein-signaling (RGS) proteins are a category of intracellular proteins that have an inhibitory effect on the intracellular signaling produced by G-protein-coupled receptors (GPCRs). RGS along with RGS-like proteins switch on through direct contact G-alpha subunits providing a variety of intracellular functions through intracellular signaling. RGS proteins have a common RGS domain that binds to G alpha. RGS proteins accelerate GTPase and thus enhance guanosine triphosphate hydrolysis through the alpha subunit of heterotrimeric G proteins. As a result, they inactivate the G protein and quickly turn off GPCR signaling thus terminating the resulting downstream signals. Activity and subcellular localization of RGS proteins can be changed through covalent molecular changes to the enzyme, differential gene splicing, and processing of the protein. Other roles of RGS proteins have shown them to not be solely committed to being inhibitors but behave more as modulators and integrators of signaling. RGS proteins modulate the duration and kinetics of slow calcium oscillations and rapid phototransduction and ion signaling events. In other cases, RGS proteins integrate G proteins with signaling pathways linked to such diverse cellular responses as cell growth and differentiation, cell motility, and intracellular trafficking. Human and animal studies have revealed that RGS proteins play a vital role in physiology and can be ideal targets for diseases such as those related to addiction where receptor signaling seems continuously switched on.

  6. Inflammation, oxidative stress and metabolic syndrome: dietary modulation.

    PubMed

    Fernández-García, José C; Cardona, Fernando; Tinahones, Francisco J

    2013-11-01

    The metabolic syndrome (MetS) is a cluster of risk factors for the development of cardiovascular disease and type 2 diabetes mellitus. These risk factors include raised blood pressure, dyslipidemia (raised triglycerides and lowered high-density lipoprotein cholesterol), raised fasting glucose, and central obesity. MetS has become a serious public health and clinical problem whose prevalence and incidence are increasing along with the worldwide rise in rates of obesity and sedentary lifestyles. A number of studies have shown that MetS is associated with a state of low-grade inflammation, characterized by abnormal pro-inflammatory cytokine production, increased acute-phase reactants, and activation of a network of inflammatory signalling pathways. Moreover, MetS has also been linked to oxidative stress, a consequence of a reduction in the antioxidant systems and an increase in the production of reactive oxygen species. Nevertheless, agreement exists that dietary intervention may modulate the pro-inflammatory state and lessen oxidative stress related to MetS, thereby decreasing the cardiovascular risk. In this review we address the current available evidence regarding dietary modulation of inflammation and oxidative stress associated with MetS. PMID:24168441

  7. BCL-2 family proteins as regulators of mitochondria metabolism.

    PubMed

    Gross, Atan

    2016-08-01

    The BCL-2 family proteins are major regulators of apoptosis, and one of their major sites of action are the mitochondria. Mitochondria are the cellular hubs for metabolism and indeed selected BCL-2 family proteins also possess roles related to mitochondria metabolism and dynamics. Here we discuss the link between mitochondrial metabolism/dynamics and the fate of stem cells, with an emphasis on the role of the BID-MTCH2 pair in regulating this link. We also discuss the possibility that BCL-2 family proteins act as metabolic sensors/messengers coming on and off of mitochondria to "sample" the cytosol and provide the mitochondria with up-to-date metabolic information. This article is part of a Special Issue entitled 'EBEC 2016: 19th European Bioenergetics Conference, Riva del Garda, Italy, July 2-6, 2016', edited by Prof. Paolo Bernardi.

  8. MANET: tracing evolution of protein architecture in metabolic networks

    PubMed Central

    Kim, Hee Shin; Mittenthal, Jay E; Caetano-Anollés, Gustavo

    2006-01-01

    Background Cellular metabolism can be characterized by networks of enzymatic reactions and transport processes capable of supporting cellular life. Our aim is to find evolutionary patterns and processes embedded in the architecture and function of modern metabolism, using information derived from structural genomics. Description The Molecular Ancestry Network (MANET) project traces evolution of protein architecture in biomolecular networks. We describe metabolic MANET, a database that links information in the Structural Classification of Proteins (SCOP), the Kyoto Encyclopedia of Genes and Genomes (KEGG), and phylogenetic reconstructions depicting the evolution of protein fold architecture. Metabolic MANET literally 'paints' the ancestries of enzymes derived from rooted phylogenomic trees directly onto over one hundred metabolic subnetworks, enabling the study of evolutionary patterns at global and local levels. An initial analysis of painted subnetworks reveals widespread enzymatic recruitment and an early origin of amino acid metabolism. Conclusion MANET maps evolutionary relationships directly and globally onto biological networks, and can generate and test hypotheses related to evolution of metabolism. We anticipate its use in the study of other networks, such as signaling and other protein-protein interaction networks. PMID:16854231

  9. Evolution of biomolecular networks: lessons from metabolic and protein interactions.

    PubMed

    Yamada, Takuji; Bork, Peer

    2009-11-01

    Despite only becoming popular at the beginning of this decade, biomolecular networks are now frameworks that facilitate many discoveries in molecular biology. The nodes of these networks are usually proteins (specifically enzymes in metabolic networks), whereas the links (or edges) are their interactions with other molecules. These networks are made up of protein-protein interactions or enzyme-enzyme interactions through shared metabolites in the case of metabolic networks. Evolutionary analysis has revealed that changes in the nodes and links in protein-protein interaction and metabolic networks are subject to different selection pressures owing to distinct topological features. However, many evolutionary constraints can be uncovered only if temporal and spatial aspects are included in the network analysis.

  10. Radioactive Lysine in Protein Metabolism Studies

    DOE R&D Accomplishments Database

    Miller, L. L.; Bale, W. F.; Yuile, C. L.; Masters, R. E.; Tishkoff, G. H.; Whipple,, G. H.

    1950-01-09

    Studies of incorporation of DL-lysine in various body proteins of the dog; the time course of labeled blood proteins; and apparent rate of disappearance of labeled plasma proteins for comparison of behavior of the plasma albumin and globulin fractions; shows more rapid turn over of globulin fraction.

  11. Protein and amino acid metabolism and requirements

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Proteins are the major structural and functional components of all cells of the body. Enzymes, membrane carriers, blood transport molecules, intracellular matrix, and even hair and fingernails are proteins, as are many hormones. Proteins also constitute a major portion of all membranes, and the cons...

  12. Predicting metabolic pathways of small molecules and enzymes based on interaction information of chemicals and proteins.

    PubMed

    Gao, Yu-Fei; Chen, Lei; Cai, Yu-Dong; Feng, Kai-Yan; Huang, Tao; Jiang, Yang

    2012-01-01

    Metabolic pathway analysis, one of the most important fields in biochemistry, is pivotal to understanding the maintenance and modulation of the functions of an organism. Good comprehension of metabolic pathways is critical to understanding the mechanisms of some fundamental biological processes. Given a small molecule or an enzyme, how may one identify the metabolic pathways in which it may participate? Answering such a question is a first important step in understanding a metabolic pathway system. By utilizing the information provided by chemical-chemical interactions, chemical-protein interactions, and protein-protein interactions, a novel method was proposed by which to allocate small molecules and enzymes to 11 major classes of metabolic pathways. A benchmark dataset consisting of 3,348 small molecules and 654 enzymes of yeast was constructed to test the method. It was observed that the first order prediction accuracy evaluated by the jackknife test was 79.56% in identifying the small molecules and enzymes in a benchmark dataset. Our method may become a useful vehicle in predicting the metabolic pathways of small molecules and enzymes, providing a basis for some further analysis of the pathway systems.

  13. Peptides as modulators of enzymes and regulatory proteins.

    PubMed

    Troitskaya, Larisa A; Kodadek, Thomas

    2004-04-01

    There is currently great interest in the development of methods to modulate the function of diverse classes of target proteins with chemicals (agonists or antagonists). These would be valuable reagents for biomedical research and some might serve as potential drug leads. Traditionally, most chemicals that modulate protein function have been enzyme inhibitors isolated in functional screens specific for the enzyme of interest. However, recent efforts from many laboratories have suggested that relatively simple binding assays may provide a more convenient and general route to chemical modulators. We review here this work with a particular emphasis on peptide modulators. PMID:15003603

  14. Modulation of phosphoinositide metabolism in aortic smooth muscle cells by allylamine

    SciTech Connect

    Cox, L.R.; Murphy, S.K.; Ramos, K. )

    1990-08-01

    Aortic smooth muscle cells (SMC) modulate from a contractile to a proliferative phenotype upon subchronic exposure to allylamine. The present studies were designed to determine if this phenotypic modulation is associated with alterations in the metabolism of membrane phosphoinositides. 32P incorporation into phosphatidylinositol 4-phosphate (PIP), phosphatidylinositol 4,5-bisphosphate (PIP2), and phosphatidic acid (PA) was lower by 31, 35, and 22%, respectively, in SMC from allylamine-treated animals relative to controls. In contrast, incorporation of (3H)myoinositol into inositol phosphates did not differ in allylamine cells relative to control cells. Exposure to dibutyryl (db) cAMP (0.2 mM) and theophylline (0.1 mM) reduced 32P incorporation into PIP and PIP2 in SMC from both experimental groups. Under these conditions, a decrease in (3H)myoinositol incorporation into inositol 1-phosphate was only observed in allylamine cells. The effects of db cAMP and theophylline in allylamine and control SMC correlated with a marked decrease in cellular proliferation. These results suggest that alterations in phosphoinositide synthesis and/or degradation contribute to the enhanced proliferation of SMC induced by allylamine. To further examine this concept, the effects of agents which modulate protein kinase C (PKC) activity were evaluated. Sphingosine (125-500 ng/ml), a PKC inhibitor, decreased SMC proliferation in allylamine, but not control cells. 12-O-Tetradecanoylphorbol-13-acetate (1-100 ng/ml), a PKC agonist, stimulated proliferation in control cells, but inhibited proliferation in cells from allylamine-treated animals. We conclude that allylamine-induced phenotypic modulation of SMC is associated with alterations in phosphoinositide metabolism.

  15. Effect of dietary protein restriction on renal ammonia metabolism.

    PubMed

    Lee, Hyun-Wook; Osis, Gunars; Handlogten, Mary E; Guo, Hui; Verlander, Jill W; Weiner, I David

    2015-06-15

    Dietary protein restriction has multiple benefits in kidney disease. Because protein intake is a major determinant of endogenous acid production, it is important that net acid excretion change in parallel during protein restriction. Ammonia is the primary component of net acid excretion, and inappropriate ammonia excretion can lead to negative nitrogen balance. Accordingly, we examined ammonia excretion in response to protein restriction and then we determined the molecular mechanism of the changes observed. Wild-type C57Bl/6 mice fed a 20% protein diet and then changed to 6% protein developed an 85% reduction in ammonia excretion within 2 days, which persisted during a 10-day study. The expression of multiple proteins involved in renal ammonia metabolism was altered, including the ammonia-generating enzymes phosphate-dependent glutaminase (PDG) and phosphoenolpyruvate carboxykinase (PEPCK) and the ammonia-metabolizing enzyme glutamine synthetase. Rhbg, an ammonia transporter, increased in expression in the inner stripe of outer medullary collecting duct intercalated cell (OMCDis-IC). However, collecting duct-specific Rhbg deletion did not alter the response to protein restriction. Rhcg deletion did not alter ammonia excretion in response to dietary protein restriction. These results indicate 1) dietary protein restriction decreases renal ammonia excretion through coordinated regulation of multiple components of ammonia metabolism; 2) increased Rhbg expression in the OMCDis-IC may indicate a biological role in addition to ammonia transport; and 3) Rhcg expression is not necessary to decrease ammonia excretion during dietary protein restriction. PMID:25925252

  16. Effect of dietary protein restriction on renal ammonia metabolism.

    PubMed

    Lee, Hyun-Wook; Osis, Gunars; Handlogten, Mary E; Guo, Hui; Verlander, Jill W; Weiner, I David

    2015-06-15

    Dietary protein restriction has multiple benefits in kidney disease. Because protein intake is a major determinant of endogenous acid production, it is important that net acid excretion change in parallel during protein restriction. Ammonia is the primary component of net acid excretion, and inappropriate ammonia excretion can lead to negative nitrogen balance. Accordingly, we examined ammonia excretion in response to protein restriction and then we determined the molecular mechanism of the changes observed. Wild-type C57Bl/6 mice fed a 20% protein diet and then changed to 6% protein developed an 85% reduction in ammonia excretion within 2 days, which persisted during a 10-day study. The expression of multiple proteins involved in renal ammonia metabolism was altered, including the ammonia-generating enzymes phosphate-dependent glutaminase (PDG) and phosphoenolpyruvate carboxykinase (PEPCK) and the ammonia-metabolizing enzyme glutamine synthetase. Rhbg, an ammonia transporter, increased in expression in the inner stripe of outer medullary collecting duct intercalated cell (OMCDis-IC). However, collecting duct-specific Rhbg deletion did not alter the response to protein restriction. Rhcg deletion did not alter ammonia excretion in response to dietary protein restriction. These results indicate 1) dietary protein restriction decreases renal ammonia excretion through coordinated regulation of multiple components of ammonia metabolism; 2) increased Rhbg expression in the OMCDis-IC may indicate a biological role in addition to ammonia transport; and 3) Rhcg expression is not necessary to decrease ammonia excretion during dietary protein restriction.

  17. Comparative genomics and evolution of proteins involved in RNA metabolism

    PubMed Central

    Anantharaman, Vivek; Koonin, Eugene V.; Aravind, L.

    2002-01-01

    RNA metabolism, broadly defined as the compendium of all processes that involve RNA, including transcription, processing and modification of transcripts, translation, RNA degradation and its regulation, is the central and most evolutionarily conserved part of cell physiology. A comprehensive, genome-wide census of all enzymatic and non-enzymatic protein domains involved in RNA metabolism was conducted by using sequence profile analysis and structural comparisons. Proteins related to RNA metabolism comprise from 3 to 11% of the complete protein repertoire in bacteria, archaea and eukaryotes, with the greatest fraction seen in parasitic bacteria with small genomes. Approximately one-half of protein domains involved in RNA metabolism are present in most, if not all, species from all three primary kingdoms and are traceable to the last universal common ancestor (LUCA). The principal features of LUCA’s RNA metabolism system were reconstructed by parsimony-based evolutionary analysis of all relevant groups of orthologous proteins. This reconstruction shows that LUCA possessed not only the basal translation system, but also the principal forms of RNA modification, such as methylation, pseudouridylation and thiouridylation, as well as simple mechanisms for polyadenylation and RNA degradation. Some of these ancient domains form paralogous groups whose evolution can be traced back in time beyond LUCA, towards low-specificity proteins, which probably functioned as cofactors for ribozymes within the RNA world framework. The main lineage-specific innovations of RNA metabolism systems were identified. The most notable phase of innovation in RNA metabolism coincides with the advent of eukaryotes and was brought about by the merge of the archaeal and bacterial systems via mitochondrial endosymbiosis, but also involved emergence of several new, eukaryote-specific RNA-binding domains. Subsequent, vast expansions of these domains mark the origin of alternative splicing in animals

  18. Can We Prevent Obesity-Related Metabolic Diseases by Dietary Modulation of the Gut Microbiota?

    PubMed

    Brahe, Lena K; Astrup, Arne; Larsen, Lesli H

    2016-01-01

    Obesity increases the risk of type 2 diabetes, cardiovascular diseases, and certain cancers, which are among the leading causes of death worldwide. Obesity and obesity-related metabolic diseases are characterized by specific alterations in the human gut microbiota. Experimental studies with gut microbiota transplantations in mice and in humans indicate that a specific gut microbiota composition can be the cause and not just the consequence of the obese state and metabolic disease, which suggests a potential for gut microbiota modulation in prevention and treatment of obesity-related metabolic diseases. In addition, dietary intervention studies have suggested that modulation of the gut microbiota can improve metabolic risk markers in humans, but a causal role of the gut microbiota in such studies has not yet been established. Here, we review and discuss the role of the gut microbiota in obesity-related metabolic diseases and the potential of dietary modulation of the gut microbiota in metabolic disease prevention and treatment. PMID:26773017

  19. Can We Prevent Obesity-Related Metabolic Diseases by Dietary Modulation of the Gut Microbiota?

    PubMed

    Brahe, Lena K; Astrup, Arne; Larsen, Lesli H

    2016-01-01

    Obesity increases the risk of type 2 diabetes, cardiovascular diseases, and certain cancers, which are among the leading causes of death worldwide. Obesity and obesity-related metabolic diseases are characterized by specific alterations in the human gut microbiota. Experimental studies with gut microbiota transplantations in mice and in humans indicate that a specific gut microbiota composition can be the cause and not just the consequence of the obese state and metabolic disease, which suggests a potential for gut microbiota modulation in prevention and treatment of obesity-related metabolic diseases. In addition, dietary intervention studies have suggested that modulation of the gut microbiota can improve metabolic risk markers in humans, but a causal role of the gut microbiota in such studies has not yet been established. Here, we review and discuss the role of the gut microbiota in obesity-related metabolic diseases and the potential of dietary modulation of the gut microbiota in metabolic disease prevention and treatment.

  20. Leucine and protein metabolism in obese zucker rats

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Branched-chain amino acids (BCAAs) are circulating nutrient signals for protein accretion, however they increase in obesity and appear to prognosticate diabetes onset. To understand the mechanisms whereby obesity affects BCAAs and protein metabolism, we employed metabolomics and measured rates of [1...

  1. Protein and amino acid metabolism in the human newborn.

    PubMed

    Kalhan, Satish C; Bier, Dennis M

    2008-01-01

    Birth and adaptation to extrauterine life involve major shifts in the protein and energy metabolism of the human newborn. These include a shift from a state of continuous supply of nutrients including amino acids from the mother to cyclic periodic oral intake, a change in the redox state of organs, thermogenesis, and a significant change in the mobilization and use of oxidative substrates. The development of safe, stable isotopic tracer methods has allowed the study of protein and amino acid metabolism not only in the healthy newborn but also in those born prematurely and of low birth weight. These studies have identified the unique and quantitative aspects of amino acid/protein metabolism in the neonate, thus contributing to rational nutritional care of these babies. The present review summarizes the contemporary data on some of the significant developments in essential and dispensable amino acids and their relationship to overall protein metabolism. Specifically, the recent data of kinetics of leucine, phenylalanine, glutamine, sulfur amino acid, and threonine and their relation to whole-body protein turnover are presented. Finally, the physiological rationale and the impact of nutrient (amino acids) interventions on the dynamics of protein metabolism are discussed.

  2. Circadian Clocks as Modulators of Metabolic Comorbidity in Psychiatric Disorders.

    PubMed

    Barandas, Rita; Landgraf, Dominic; McCarthy, Michael J; Welsh, David K

    2015-12-01

    Psychiatric disorders such as schizophrenia, bipolar disorder, and major depressive disorder are often accompanied by metabolic dysfunction symptoms, including obesity and diabetes. Since the circadian system controls important brain systems that regulate affective, cognitive, and metabolic functions, and neuropsychiatric and metabolic diseases are often correlated with disturbances of circadian rhythms, we hypothesize that dysregulation of circadian clocks plays a central role in metabolic comorbidity in psychiatric disorders. In this review paper, we highlight the role of circadian clocks in glucocorticoid, dopamine, and orexin/melanin-concentrating hormone systems and describe how a dysfunction of these clocks may contribute to the simultaneous development of psychiatric and metabolic symptoms. PMID:26483181

  3. Adjustments of Protein Metabolism in Fasting Arctic Charr, Salvelinus alpinus

    PubMed Central

    Cassidy, Alicia A.; Saulnier, Roxanne J.; Lamarre, Simon G.

    2016-01-01

    Protein metabolism, including the interrelated processes of synthesis and degradation, mediates the growth of an animal. In ectothermic animals, protein metabolism is responsive to changes in both biotic and abiotic conditions. This study aimed to characterise responses of protein metabolism to food deprivation that occur in the coldwater salmonid, Arctic charr, Salvelinus alpinus. We compared two groups of Arctic charr: one fed continuously and the other deprived of food for 36 days. We measured the fractional rate of protein synthesis (KS) in individuals from the fed and fasted groups using a flooding dose technique modified for the use of deuterium-labelled phenylalanine. The enzyme activities of the three major protein degradation pathways (ubiquitin proteasome, lysosomal cathepsins and the calpain systems) were measured in the same fish. This study is the first to measure both KS and the enzymatic activity of protein degradation in the same fish, allowing us to examine the apparent contribution of different protein degradation pathways to protein turnover in various tissues (red and white muscle, liver, heart and gills). KS was lower in the white muscle and in liver of the fasted fish compared to the fed fish. There were no observable effects of food deprivation on the protease activities in any of the tissues with the exception of liver, where the ubiquitin proteasome pathway seemed to be activated during fasting conditions. Lysosomal proteolysis appears to be the primary degradation pathway for muscle protein, while the ubiquitin proteasome pathway seems to predominate in the liver. We speculate that Arctic charr regulate protein metabolism during food deprivation to conserve proteins. PMID:27096948

  4. Co-evolution of metabolism and protein sequences.

    PubMed

    Schütte, Moritz; Klitgord, Niels; Segrè, Daniel; Ebenhöh, Oliver

    2010-01-01

    The set of chemicals producible and usable by metabolic pathways must have evolved in parallel with the enzymes that catalyze them. One implication of this common historical path should be a correspondence between the innovation steps that gradually added new metabolic reactions to the biosphere-level biochemical toolkit, and the gradual sequence changes that must have slowly shaped the corresponding enzyme structures. However, global signatures of a long-term co-evolution have not been identified. Here we search for such signatures by computing correlations between inter-reaction distances on a metabolic network, and sequence distances of the corresponding enzyme proteins. We perform our calculations using the set of all known metabolic reactions, available from the KEGG database. Reaction-reaction distance on the metabolic network is computed as the length of the shortest path on a projection of the metabolic network, in which nodes are reactions and edges indicate whether two reactions share a common metabolite, after removal of cofactors. Estimating the distance between enzyme sequences in a meaningful way requires some special care: for each enzyme commission (EC) number, we select from KEGG a consensus set of protein sequences using the cluster of orthologous groups of proteins (COG) database. We define the evolutionary distance between protein sequences as an asymmetric transition probability between two enzymes, derived from the corresponding pair-wise BLAST scores. By comparing the distances between sequences to the minimal distances on the metabolic reaction graph, we find a small but statistically significant correlation between the two measures. This suggests that the evolutionary walk in enzyme sequence space has locally mirrored, to some extent, the gradual expansion of metabolism. PMID:20238426

  5. Processing incommensurately modulated protein diffraction data with Eval15

    SciTech Connect

    Porta, Jason; Lovelace, Jeffrey J.; Schreurs, Antoine M. M.; Kroon-Batenburg, Loes M. J.; Borgstahl, Gloria E. O.

    2011-07-01

    Data processing of an incommensurately modulated profilin–actin crystal is described. Recent challenges in biological X-ray crystallography include the processing of modulated diffraction data. A modulated crystal has lost its three-dimensional translational symmetry but retains long-range order that can be restored by refining a periodic modulation function. The presence of a crystal modulation is indicated by an X-ray diffraction pattern with periodic main reflections flanked by off-lattice satellite reflections. While the periodic main reflections can easily be indexed using three reciprocal-lattice vectors a*, b*, c*, the satellite reflections have a non-integral relationship to the main lattice and require a q vector for indexing. While methods for the processing of diffraction intensities from modulated small-molecule crystals are well developed, they have not been applied in protein crystallography. A recipe is presented here for processing incommensurately modulated data from a macromolecular crystal using the Eval program suite. The diffraction data are from an incommensurately modulated crystal of profilin–actin with single-order satellites parallel to b*. The steps taken in this report can be used as a guide for protein crystallographers when encountering crystal modulations. To our knowledge, this is the first report of the processing of data from an incommensurately modulated macromolecular crystal.

  6. Glucocorticoids and 11β-HSD1 are major regulators of intramyocellular protein metabolism

    PubMed Central

    Hassan-Smith, Zaki K; Doig, Craig L; Sherlock, Mark; Stewart, Paul M; Lavery, Gareth G

    2016-01-01

    The adverse metabolic effects of prescribed and endogenous glucocorticoid excess, ‘Cushing’s syndrome’, create a significant health burden. While skeletal muscle atrophy and resultant myopathy is a clinical feature, the molecular mechanisms underpinning these changes are not fully defined. We have characterized the impact of glucocorticoids upon key metabolic pathways and processes regulating muscle size and mass including: protein synthesis, protein degradation, and myoblast proliferation in both murine C2C12 and human primary myotube cultures. Furthermore, we have investigated the role of pre-receptor modulation of glucocorticoid availability by 11β-hydroxysteroid dehydrogenase type 1 (11β-HSD1) in these processes. Corticosterone (CORT) decreased myotube area, decreased protein synthesis, and increased protein degradation in murine myotubes. This was supported by decreased mRNA expression of insulin-like growth factor (IGF1), decreased activating phosphorylation of mammalian target of rapamycin (mTOR), decreased phosphorylation of 4E binding protein 1 (4E-BP1), and increased mRNA expression of key atrophy markers including: atrogin-1, forkhead box O3a (FOXO3a), myostatin (MSTN), and muscle-ring finger protein-1 (MuRF1). These findings were endorsed in human primary myotubes, where cortisol also decreased protein synthesis and increased protein degradation. The effects of 11-dehydrocorticosterone (11DHC) (in murine myotubes) and cortisone (in human myotubes) on protein metabolism were indistinguishable from that of CORT/cortisol treatments. Selective 11β-HSD1 inhibition blocked the decrease in protein synthesis, increase in protein degradation, and reduction in myotube area induced by 11DHC/cortisone. Furthermore, CORT/cortisol, but not 11DHC/cortisone, decreased murine and human myoblast proliferative capacity. Glucocorticoids are potent regulators of skeletal muscle protein homeostasis and myoblast proliferation. Our data underscores the potential use

  7. Functional module identification in protein interaction networks by interaction patterns

    PubMed Central

    Wang, Yijie; Qian, Xiaoning

    2014-01-01

    Motivation: Identifying functional modules in protein–protein interaction (PPI) networks may shed light on cellular functional organization and thereafter underlying cellular mechanisms. Many existing module identification algorithms aim to detect densely connected groups of proteins as potential modules. However, based on this simple topological criterion of ‘higher than expected connectivity’, those algorithms may miss biologically meaningful modules of functional significance, in which proteins have similar interaction patterns to other proteins in networks but may not be densely connected to each other. A few blockmodel module identification algorithms have been proposed to address the problem but the lack of global optimum guarantee and the prohibitive computational complexity have been the bottleneck of their applications in real-world large-scale PPI networks. Results: In this article, we propose a novel optimization formulation LCP2 (low two-hop conductance sets) using the concept of Markov random walk on graphs, which enables simultaneous identification of both dense and sparse modules based on protein interaction patterns in given networks through searching for LCP2 by random walk. A spectral approximate algorithm SLCP2 is derived to identify non-overlapping functional modules. Based on a bottom-up greedy strategy, we further extend LCP2 to a new algorithm (greedy algorithm for LCP2) GLCP2 to identify overlapping functional modules. We compare SLCP2 and GLCP2 with a range of state-of-the-art algorithms on synthetic networks and real-world PPI networks. The performance evaluation based on several criteria with respect to protein complex prediction, high level Gene Ontology term prediction and especially sparse module detection, has demonstrated that our algorithms based on searching for LCP2 outperform all other compared algorithms. Availability and implementation: All data and code are available at http://www.cse.usf.edu/∼xqian/fmi/slcp2hop

  8. The metabolically-modulated stem cell niche: a dynamic scenario regulating cancer cell phenotype and resistance to therapy

    PubMed Central

    Rovida, Elisabetta; Peppicelli, Silvia; Bono, Silvia; Bianchini, Francesca; Tusa, Ignazia; Cheloni, Giulia; Marzi, Ilaria; Cipolleschi, Maria Grazia; Calorini, Lido; Sbarba, Persio Dello

    2014-01-01

    This Perspective addresses the interactions of cancer stem cells (CSC) with environment which result in the modulation of CSC metabolism, and thereby of CSC phenotype and resistance to therapy. We considered first as a model disease chronic myeloid leukemia (CML), which is triggered by a well-identified oncogenetic protein (BCR/Abl) and brilliantly treated with tyrosine kinase inhibitors (TKi). However, TKi are extremely effective in inducing remission of disease, but unable, in most cases, to prevent relapse. We demonstrated that the interference with cell metabolism (oxygen/glucose shortage) enriches cells exhibiting the leukemia stem cell (LSC) phenotype and, at the same time, suppresses BCR/Abl protein expression. These LSC are therefore refractory to the TKi Imatinib-mesylate, pointing to cell metabolism as an important factor controlling the onset of TKi-resistant minimal residual disease (MRD) of CML and the related relapse. Studies of solid neoplasias brought another player into the control of MRD, low tissue pH, which often parallels cancer growth and progression. Thus, a 3-party scenario emerged for the regulation of CSC/LSC maintenance, MRD induction and disease relapse: the “hypoxic” versus the “ischemic” vs. the “acidic” environment. As these environments are unlikely constrained within rigid borders, we named this model the “metabolically-modulated stem cell niche.” PMID:25485495

  9. Root carbon and protein metabolism associated with heat tolerance.

    PubMed

    Huang, Bingru; Rachmilevitch, Shimon; Xu, Jichen

    2012-05-01

    Extensive past efforts have been taken toward understanding heat tolerance mechanisms of the aboveground organs. Root systems play critical roles in whole-plant adaptation to heat stress, but are less studied. This review discusses recent research results revealing some critical physiological and metabolic factors underlying root thermotolerance, with a focus on temperate perennial grass species. Comparative analysis of differential root responses to supraoptimal temperatures by a heat-adapted temperate C3 species, Agrostis scabra, which can survive high soil temperatures up to 45 °C in geothermal areas in Yellow Stone National Park, and a heat-sensitive cogeneric species, Agrostis stolonifera, suggested that efficient carbon and protein metabolism is critical for root thermotolerance. Superior root thermotolerance in a perennial grass was associated with a greater capacity to control respiratory costs through respiratory acclimation, lowering carbon investment in maintenance for protein turnover, and efficiently partitioning carbon into different metabolic pools and alternative respiration pathways. Proteomic analysis demonstrated that root thermotolerance was associated with an increased maintenance of stability and less degradation of proteins, particularly those important for metabolism and energy production. In addition, thermotolerant roots are better able to maintain growth and activity during heat stress by activating stress defence proteins such as those participating in antioxidant defence (i.e. superoxide dismutase, peroxidase, glutathione S-transferase) and chaperoning protection (i.e. heat shock protein).

  10. Root carbon and protein metabolism associated with heat tolerance.

    PubMed

    Huang, Bingru; Rachmilevitch, Shimon; Xu, Jichen

    2012-05-01

    Extensive past efforts have been taken toward understanding heat tolerance mechanisms of the aboveground organs. Root systems play critical roles in whole-plant adaptation to heat stress, but are less studied. This review discusses recent research results revealing some critical physiological and metabolic factors underlying root thermotolerance, with a focus on temperate perennial grass species. Comparative analysis of differential root responses to supraoptimal temperatures by a heat-adapted temperate C3 species, Agrostis scabra, which can survive high soil temperatures up to 45 °C in geothermal areas in Yellow Stone National Park, and a heat-sensitive cogeneric species, Agrostis stolonifera, suggested that efficient carbon and protein metabolism is critical for root thermotolerance. Superior root thermotolerance in a perennial grass was associated with a greater capacity to control respiratory costs through respiratory acclimation, lowering carbon investment in maintenance for protein turnover, and efficiently partitioning carbon into different metabolic pools and alternative respiration pathways. Proteomic analysis demonstrated that root thermotolerance was associated with an increased maintenance of stability and less degradation of proteins, particularly those important for metabolism and energy production. In addition, thermotolerant roots are better able to maintain growth and activity during heat stress by activating stress defence proteins such as those participating in antioxidant defence (i.e. superoxide dismutase, peroxidase, glutathione S-transferase) and chaperoning protection (i.e. heat shock protein). PMID:22328905

  11. Therapeutic modulation of cannabinoid lipid signaling: metabolic profiling of a novel antinociceptive cannabinoid-2 receptor agonist

    PubMed Central

    Wood, JodiAnne T.; Smith, Dustin M.; Janero, David R.; Zvonok, Alexander M.; Makriyannis, Alexandros

    2012-01-01

    Aims AM-1241, a novel, racemic cannabinoid-2 receptor (CB2) ligand, is the primary experimental agonist used to characterize the role of CB2-mediated lipid signaling in health and disease, including substance abuse disorders. In vivo pharmacological effects have been used as indirect proxies for AM-1241 biotransformation processes that could modulate activity. We report the initial pre-clinical characterization of AM-1241 biotransformation and in vivo distribution. Main methods AM-1241 metabolism was characterized in a variety of predictive in vitro systems (Caco-2 cells, mouse, rat and human microsomes) and in the mouse in vivo. Liquid chromatography and mass spectrometry techniques were used to quantify AM-1241 tissue distribution and metabolic conversion. Key findings AM-1241 bound extensively to plasma protein/albumin. A pharmacological AM-1241 dose (25 mg/kg, i.v.) was administered to mice for direct determination of its plasma half-life (37 min), following which AM-1241 was quantified in brain, spleen, liver, and kidney. After p.o. administration, AM-1241 was detected in plasma, spleen, and kidney; its oral bioavailability was ~21%. From Caco-2 permeability studies and microsomal-based hepatic clearance estimates, in vivo AM-1241 absorption was moderate. Hepatic microsomal metabolism of AM-1241 in vitro generated hydroxylation and demethylation metabolites. Species-dependent differences were discovered in AM-1241’s predicted hepatic clearance. Our data demonstrate that AM-1241 has the following characteristics: a) short plasma half-life; b) limited oral bioavailability; c) extensive plasma/albumin binding; d) metabolic substrate for hepatic hydroxylation and demethylation; e) moderate hepatic clearance. Significance These results should help inform the design, optimization, and pre-clinical profiling of CB2 ligands as pharmacological tools and medicines. PMID:22749867

  12. Processing incommensurately modulated protein diffraction data with Eval15.

    PubMed

    Porta, Jason; Lovelace, Jeffrey J; Schreurs, Antoine M M; Kroon-Batenburg, Loes M J; Borgstahl, Gloria E O

    2011-07-01

    Recent challenges in biological X-ray crystallography include the processing of modulated diffraction data. A modulated crystal has lost its three-dimensional translational symmetry but retains long-range order that can be restored by refining a periodic modulation function. The presence of a crystal modulation is indicated by an X-ray diffraction pattern with periodic main reflections flanked by off-lattice satellite reflections. While the periodic main reflections can easily be indexed using three reciprocal-lattice vectors a*, b*, c*, the satellite reflections have a non-integral relationship to the main lattice and require a q vector for indexing. While methods for the processing of diffraction intensities from modulated small-molecule crystals are well developed, they have not been applied in protein crystallography. A recipe is presented here for processing incommensurately modulated data from a macromolecular crystal using the Eval program suite. The diffraction data are from an incommensurately modulated crystal of profilin-actin with single-order satellites parallel to b*. The steps taken in this report can be used as a guide for protein crystallographers when encountering crystal modulations. To our knowledge, this is the first report of the processing of data from an incommensurately modulated macromolecular crystal.

  13. DAPIT Over-Expression Modulates Glucose Metabolism and Cell Behaviour in HEK293T Cells

    PubMed Central

    Kontro, Heidi; Cannino, Giuseppe; Rustin, Pierre; Dufour, Eric; Kainulainen, Heikki

    2015-01-01

    Introduction Diabetes Associated Protein in Insulin-sensitive Tissues (DAPIT) is a subunit of mitochondrial ATP synthase and has also been found to associate with the vacuolar H+-ATPase. Its expression is particularly high in cells with elevated aerobic metabolism and in epithelial cells that actively transport nutrients and ions. Deletion of DAPIT is known to induce loss of mitochondrial ATP synthase but the effects of its over-expression are obscure. Results In order to study the consequences of high expression of DAPIT, we constructed a transgenic cell line that constitutively expressed DAPIT in human embryonal kidney cells, HEK293T. Enhanced DAPIT expression decreased mtDNA content and mitochondrial mass, and saturated respiratory chain by decreasing H+-ATP synthase activity. DAPIT over-expression also increased mitochondrial membrane potential and superoxide level, and translocated the transcription factors hypoxia inducible factor 1α (Hif1α) and β-catenin to the nucleus. Accordingly, cells over-expressing DAPIT used more glucose and generated a larger amount of lactate compared to control cells. Interestingly, these changes were associated with an epithelial to mesenchymal (EMT)-like transition by changing E-cadherin to N-cadherin and up-regulating several key junction/adhesion proteins. At physiological level, DAPIT over-expression slowed down cell growth by G1 arrest and migration, and enhanced cell detachment. Several cancers also showed an increase in genomic copy number of Usmg5 (gene encoding DAPIT), thereby providing strong correlative evidence for DAPIT possibly having oncogenic function in cancers. Conclusions DAPIT over-expression thus appears to modulate mitochondrial functions and alter cellular regulations, promote anaerobic metabolism and induce EMT-like transition. We propose that DAPIT over-expression couples the changes in mitochondrial metabolism to physiological and pathophysiological regulations, and suggest it could play a critical

  14. Regulation of intestinal protein metabolism by amino acids.

    PubMed

    Bertrand, Julien; Goichon, Alexis; Déchelotte, Pierre; Coëffier, Moïse

    2013-09-01

    Gut homeostasis plays a major role in health and may be regulated by quantitative and qualitative food intake. In the intestinal mucosa, an intense renewal of proteins occurs, at approximately 50% per day in humans. In some pathophysiological conditions, protein turnover is altered and may contribute to intestinal or systemic diseases. Amino acids are key effectors of gut protein turnover, both as constituents of proteins and as regulatory molecules limiting intestinal injury and maintaining intestinal functions. Many studies have focused on two amino acids: glutamine, known as the preferential substrate of rapidly dividing cells, and arginine, another conditionally essential amino acid. The effects of glutamine and arginine on protein synthesis appear to be model and condition dependent, as are the involved signaling pathways. The regulation of gut protein degradation by amino acids has been minimally documented until now. This review will examine recent data, helping to better understand how amino acids regulate intestinal protein metabolism, and will explore perspectives for future studies.

  15. Protein homeostasis disorders of key enzymes of amino acids metabolism: mutation-induced protein kinetic destabilization and new therapeutic strategies.

    PubMed

    Pey, Angel L

    2013-12-01

    Many inborn errors of amino acids metabolism are caused by single point mutations affecting the ability of proteins to fold properly (i.e., protein homeostasis), thus leading to enzyme loss-of-function. Mutations may affect protein homeostasis by altering intrinsic physical properties of the polypeptide (folding thermodynamics, and rates of folding/unfolding/misfolding) as well as the interaction of partially folded states with elements of the protein homeostasis network (such as molecular chaperones and proteolytic machineries). Understanding these mutational effects on protein homeostasis is required to develop new therapeutic strategies aimed to target specific features of the mutant polypeptide. Here, I review recent work in three different diseases of protein homeostasis associated to inborn errors of amino acids metabolism: phenylketonuria, inherited homocystinuria and primary hyperoxaluria type I. These three different genetic disorders involve proteins operating in different cell organelles and displaying different structural complexities. Mutations often decrease protein kinetic stability of the native state (i.e., its half-life for irreversible denaturation), which can be studied using simple kinetic models amenable to biophysical and biochemical characterization. Natural ligands and pharmacological chaperones are shown to stabilize mutant enzymes, thus supporting their therapeutic application to overcome protein kinetic destabilization. The role of molecular chaperones in protein folding and misfolding is also discussed as well as their potential pharmacological modulation as promising new therapeutic approaches. Since current available treatments for these diseases are either burdening or only successful in a fraction of patients, alternative treatments must be considered covering studies from protein structure and biophysics to studies in animal models and patients.

  16. Protein-Tyrosine Phosphatase 1B Substrates and Metabolic Regulation

    PubMed Central

    Bakke, Jesse; Haj, Fawaz G.

    2014-01-01

    Metabolic homeostasis requires integration of complex signaling networks which, when deregulated, contribute to metabolic syndrome and related disorders. Protein-tyrosine phosphatase 1B (PTP1B) has emerged as a key regulator of signaling networks that are implicated in metabolic diseases such as obesity and type 2 diabetes. In this review, we examine mechanisms that regulate PTP1B-substrate interaction, enzymatic activity and experimental approaches to identify PTP1B substrates. We then highlight findings that implicate PTP1B in metabolic regulation. In particular, insulin and leptin signaling are discussed as well as recently identified PTP1B substrates that are involved in endoplasmic reticulum stress response, cell-cell communication, energy balance and vesicle trafficking. In summary, PTP1B exhibits exquisite substrate specificity and is an outstanding pharmaceutical target for obesity and type 2 diabetes. PMID:25263014

  17. Expanding Metabolic Engineering Algorithms Using Feasible Space and Shadow Price Constraint Modules

    PubMed Central

    Tervo, Christopher J.; Reed, Jennifer L.

    2014-01-01

    While numerous computational methods have been developed that use genome-scale models to propose mutants for the purpose of metabolic engineering, they generally compare mutants based on a single criteria (e.g., production rate at a mutant’s maximum growth rate). As such, these approaches remain limited in their ability to include multiple complex engineering constraints. To address this shortcoming, we have developed feasible space and shadow price constraint (FaceCon and ShadowCon) modules that can be added to existing mixed integer linear adaptive evolution metabolic engineering algorithms, such as OptKnock and OptORF. These modules allow strain designs to be identified amongst a set of multiple metabolic engineering algorithm solutions that are capable of high chemical production while also satisfying additional design criteria. We describe the various module implementations and their potential applications to the field of metabolic engineering. We then incorporated these modules into the OptORF metabolic engineering algorithm. Using an Escherichia coli genome-scale model (iJO1366), we generated different strain designs for the anaerobic production of ethanol from glucose, thus demonstrating the tractability and potential utility of these modules in metabolic engineering algorithms. PMID:25478320

  18. The Ribosomal Protein-Mdm2-p53 Pathway and Energy Metabolism

    PubMed Central

    Deisenroth, Chad; Zhang, Yanping

    2011-01-01

    Cellular growth and division are two fundamental processes that are exquisitely sensitive and responsive to environmental fluctuations. One of the most energetically demanding functions of these processes is ribosome biogenesis, the key component to regulating overall protein synthesis and cell growth. Perturbations to ribosome biogenesis have been demonstrated to induce an acute stress response leading to p53 activation through the inhibition of Mdm2 by a number of ribosomal proteins. The energy status of a cell is a highly dynamic variable that naturally contributes to metabolic fluctuations, which can affect both the rates of ribosome biogenesis and p53 function. This, in turn, determines whether a cell is in an anabolic, growth-promoting state or a catabolic, growth-suppressing state. Here the authors integrate the known functions of p53 to postulate how changes in nutrient availability may induce the ribosomal protein–Mdm2-p53 signaling pathway to modulate p53-dependent metabolic regulation. PMID:21779508

  19. [Review: plant polyphenols modulate lipid metabolism and related molecular mechanism].

    PubMed

    Dai, Yan-li; Zou, Yu-xiao; Liu, Fan; Li, Hong-zhi

    2015-11-01

    Lipid metabolism disorder is an important risk factor to obesity, hyperlipidemia and type 2 diabetes as well as other chronic metabolic disease. It is also a key target in preventing metabolic syndrome, chronic disease prevention. Plant polyphenol plays an important role in maintaining or improving lipid profile in a variety of ways. including regulating cholesterol absorption, inhibiting synthesis and secretion of triglyceride, and lowering plasma low density lipoprotein oxidation, etc. The purpose of this article is to review the lipid regulation effects of plant polyphenols and its related mechanisms.

  20. Dietary beta-carotene and lutein metabolism is modulated by the APOE genotype.

    PubMed

    Huebbe, Patricia; Lange, Jennifer; Lietz, Georg; Rimbach, Gerald

    2016-07-01

    The human apolipoprotein E (APOE) genotype has been suggested to interact with nutrient metabolism particularly with lipid soluble vitamins. Plasma carotenoid levels are determined by numerous dietary and genetic factors with high inter-individual variation; however, the APOE genotype has not been systematically examined so far. Our aim was to investigate the effect of the APOE genotype on dietary carotenoid metabolism with special regard to transcriptional regulation of carotenoid absorption, cleavage and adipocyte fat storage. We supplemented targeted replacement mice expressing human APOE3 and APOE4 isoforms with dietary beta-carotene (BC) and lutein (LUT) for 8 weeks. Plasma BC and adipose tissue BC and LUT levels were in trend lower in APOE4 than APOE3 mice, while hepatic expression of the beta-carotene oxygenases BCO1 and BCO2 was significantly higher. In contrast to the liver, mRNA levels of proteins involved in carotenoid absorption and cleavage in the small intestinal mucosa as well as of adipogenic markers in the adipose tissue were not different between APOE3 and APOE4 mice. Our data suggest that the hepatic carotenoid cleavage activity is higher in APOE4 mice partially reducing the circulation and extra-hepatic accumulation of intact carotenoids as compared to APOE3. Therefore we suggest considering the APOE genotype as modulator of carotenoid status in the future. © 2016 BioFactors, 42(4):388-396, 2016. PMID:27040933

  1. Low Concentrations of Nitric Oxide Modulate Streptococcus pneumoniae Biofilm Metabolism and Antibiotic Tolerance

    PubMed Central

    Allan, Raymond N.; Morgan, Samantha; Brito-Mutunayagam, Sanjita; Skipp, Paul; Feelisch, Martin; Hayes, Stephen M.; Hellier, William; Clarke, Stuart C.; Stoodley, Paul; Burgess, Andrea; Ismail-Koch, Hasnaa; Salib, Rami J.; Webb, Jeremy S.; Hall-Stoodley, Luanne

    2016-01-01

    Streptococcus pneumoniae is one of the key pathogens responsible for otitis media (OM), the most common infection in children and the largest cause of childhood antibiotic prescription. Novel therapeutic strategies that reduce the overall antibiotic consumption due to OM are required because, although widespread pneumococcal conjugate immunization has controlled invasive pneumococcal disease, overall OM incidence has not decreased. Biofilm formation represents an important phenotype contributing to the antibiotic tolerance and persistence of S. pneumoniae in chronic or recurrent OM. We investigated the treatment of pneumococcal biofilms with nitric oxide (NO), an endogenous signaling molecule and therapeutic agent that has been demonstrated to trigger biofilm dispersal in other bacterial species. We hypothesized that addition of low concentrations of NO to pneumococcal biofilms would improve antibiotic efficacy and that higher concentrations exert direct antibacterial effects. Unlike in many other bacterial species, low concentrations of NO did not result in S. pneumoniae biofilm dispersal. Instead, treatment of both in vitro biofilms and ex vivo adenoid tissue samples (a reservoir for S. pneumoniae biofilms) with low concentrations of NO enhanced pneumococcal killing when combined with amoxicillin-clavulanic acid, an antibiotic commonly used to treat chronic OM. Quantitative proteomic analysis using iTRAQ (isobaric tag for relative and absolute quantitation) identified 13 proteins that were differentially expressed following low-concentration NO treatment, 85% of which function in metabolism or translation. Treatment with low-concentration NO, therefore, appears to modulate pneumococcal metabolism and may represent a novel therapeutic approach to reduce antibiotic tolerance in pneumococcal biofilms. PMID:26856845

  2. Total Cellular RNA Modulates Protein Activity.

    PubMed

    Majumder, Subhabrata; DeMott, Christopher M; Reverdatto, Sergey; Burz, David S; Shekhtman, Alexander

    2016-08-16

    RNA constitutes up to 20% of a cell's dry weight, corresponding to ∼20 mg/mL. This high concentration of RNA facilitates low-affinity protein-RNA quinary interactions, which may play an important role in facilitating and regulating biological processes. In the yeast Pichia pastoris, the level of ubiquitin-RNA colocalization increases when cells are grown in the presence of dextrose and methanol instead of methanol as the sole carbon source. Total RNA isolated from cells grown in methanol increases β-galactosidase activity relative to that seen with RNA isolated from cells grown in the presence of dextrose and methanol. Because the total cellular RNA content changes with growth medium, protein-RNA quinary interactions can alter in-cell protein biochemistry and may play an important role in cell adaptation, critical to many physiological and pathological states. PMID:27456029

  3. Metabolic engineering of Escherichia coli to improve recombinant protein production.

    PubMed

    Liu, Min; Feng, Xinjun; Ding, Yamei; Zhao, Guang; Liu, Huizhou; Xian, Mo

    2015-12-01

    Escherichia coli is one of the most widely used strains for recombinant protein production. However, obstacles also exist in both academic researches and industrial applications, such as the metabolic burden, the carbon source waste, and the cells' physiological deterioration. This article reviews recent approaches for improving recombinant protein production in metabolic engineering, including workhorse selection, stress factor application, and carbon flux regulation. Selecting a suitable host is the first key point for recombinant protein production. In general, it all depends on characteristics of the strains and the target proteins. It will be triggered cells physiological deterioration when the medium is significantly different from the cell's natural environment. Coexpression of stress factors can help proteins to fold into their native conformation. Carbon flux regulation is a direct approach for redirecting more carbon flux toward the desirable pathways and products. However, some undesirable consequences are usually found in metabolic engineering, such as glucose transport inhibition, cell growth retardation, and useless metabolite accumulation. More efficient regulators and platform cell factories should be explored to meet a variety of production demands.

  4. Intron positions correlate with module boundaries in ancient proteins

    PubMed Central

    de Souza, Sandro Jose; Long, Manyuan; Schoenbach, Lloyd; Roy, Scott William; Gilbert, Walter

    1996-01-01

    We analyze the three-dimensional structure of proteins by a computer program that finds regions of sequence that contain module boundaries, defining a module as a segment of polypeptide chain bounded in space by a specific given distance. The program defines a set of “linker regions” that have the property that if an intron were to be placed into each linker region, the protein would be dissected into a set of modules all less than the specified diameter. We test a set of 32 proteins, all of ancient origin, and a corresponding set of 570 intron positions, to ask if there is a statistically significant excess of intron positions within the linker regions. For 28-Å modules, a standard size used historically, we find such an excess, with P < 0.003. This correlation is neither due to a compositional or sequence bias in the linker regions nor to a surface bias in intron positions. Furthermore, a subset of 20 introns, which can be putatively identified as old, lies even more explicitly within the linker regions, with P < 0.0003. Thus, there is a strong correlation between intron positions and three-dimensional structural elements of ancient proteins as expected by the introns-early approach. We then study a range of module diameters and show that, as the diameter varies, significant peaks of correlation appear for module diameters centered at 21.7, 27.6, and 32.9 Å. These preferred module diameters roughly correspond to predicted exon sizes of 15, 22, and 30 residues. Thus, there are significant correlations between introns, modules, and a quantized pattern of the lengths of polypeptide chains, which is the prediction of the “Exon Theory of Genes.” PMID:8962105

  5. Intron positions correlate with module boundaries in ancient proteins.

    PubMed

    de Souza, S J; Long, M; Schoenbach, L; Roy, S W; Gilbert, W

    1996-12-10

    We analyze the three-dimensional structure of proteins by a computer program that finds regions of sequence that contain module boundaries, defining a module as a segment of polypeptide chain bounded in space by a specific given distance. The program defines a set of "linker regions" that have the property that if an intron were to be placed into each linker region, the protein would be dissected into a set of modules all less than the specified diameter. We test a set of 32 proteins, all of ancient origin, and a corresponding set of 570 intron positions, to ask if there is a statistically significant excess of intron positions within the linker regions. For 28-A modules, a standard size used historically, we find such an excess, with P < 0.003. This correlation is neither due to a compositional or sequence bias in the linker regions nor to a surface bias in intron positions. Furthermore, a subset of 20 introns, which can be putatively identified as old, lies even more explicitly within the linker regions, with P < 0.0003. Thus, there is a strong correlation between intron positions and three-dimensional structural elements of ancient proteins as expected by the introns-early approach. We then study a range of module diameters and show that, as the diameter varies, significant peaks of correlation appear for module diameters centered at 21.7, 27.6, and 32.9 A. These preferred module diameters roughly correspond to predicted exon sizes of 15, 22, and 30 residues. Thus, there are significant correlations between introns, modules, and a quantized pattern of the lengths of polypeptide chains, which is the prediction of the "Exon Theory of Genes."

  6. Engineering and Assembly of Protein Modules into Functional Molecular Systems.

    PubMed

    Hirschi, Stephan; Stauffer, Mirko; Harder, Daniel; Müller, Daniel J; Meier, Wolfgang; Fotiadis, Dimitrios

    2016-01-01

    Synthetic biology approaches range from the introduction of unique features into organisms to the assembly of isolated biomacromolecules or synthetic building blocks into artificial biological systems with biomimetic or completely novel functionalities. Simple molecular systems can be based on containers on the nanoscale that are equipped with tailored functional modules for various applications in healthcare, industry or biological and medical research. The concept, or vision, of assembling native or engineered proteins and/or synthetic components as functional modules into molecular systems is discussed. The main focus is laid on the engineering of energizing modules generating chemical energy, transport modules using this energy to translocate molecules between compartments of a molecular system, and catalytic modules (bio-)chemically processing the molecules. Further key aspects of this discourse are possible approaches for the assembly of simple nanofactories and their applications in biotechnology and medical health. PMID:27363367

  7. The roles of lipid and glucose metabolism in modulation of β-amyloid, tau, and neurodegeneration in the pathogenesis of Alzheimer disease

    PubMed Central

    Sato, Naoyuki; Morishita, Ryuichi

    2015-01-01

    Diabetes is a risk factor for Alzheimer disease (AD). Apolipoprotein E (ApoE) and several genes related to AD have recently been identified by genome-wide association studies (GWAS) as being closely linked to lipid metabolism. Lipid metabolism and glucose-energy metabolism are closely related. Here, we review the emerging evidence regarding the roles of lipid and glucose metabolism in the modulation of β-amyloid, tau, and neurodegeneration during the pathogenesis of AD. Disruption of homeostasis of lipid and glucose metabolism affects production and clearance of β-amyloid and tau phosphorylation, and induces neurodegeneration. A more integrated understanding of the interactions among lipid, glucose, and protein metabolism is required to elucidate the pathogenesis of AD and to develop next-generation therapeutic options. PMID:26557086

  8. Dimethyl fumarate modulates antioxidant and lipid metabolism in oligodendrocytes.

    PubMed

    Huang, He; Taraboletti, Alexandra; Shriver, Leah P

    2015-08-01

    Oxidative stress contributes to pathology associated with inflammatory brain disorders and therapies that upregulate antioxidant pathways may be neuroprotective in diseases such as multiple sclerosis. Dimethyl fumarate, a small molecule therapeutic for multiple sclerosis, activates cellular antioxidant signaling pathways and may promote myelin preservation. However, it is still unclear what mechanisms may underlie this neuroprotection and whether dimethyl fumarate affects oligodendrocyte responses to oxidative stress. Here, we examine metabolic alterations in oligodendrocytes treated with dimethyl fumarate by using a global metabolomic platform that employs both hydrophilic interaction liquid chromatography-mass spectrometry and shotgun lipidomics. Prolonged treatment of oligodendrocytes with dimethyl fumarate induces changes in citric acid cycle intermediates, glutathione, and lipids, indicating that this compound can directly impact oligodendrocyte metabolism. These metabolic alterations are also associated with protection from oxidant challenge. This study provides insight into the mechanisms by which dimethyl fumarate could preserve myelin integrity in patients with multiple sclerosis. PMID:25967672

  9. Homocysteine Metabolism in Children with Down Syndrome: In Vitro Modulation

    PubMed Central

    Pogribna, Marta; Melnyk, Stepan; Pogribny, Igor; Chango, Abalo; Yi, Ping; James, S. Jill

    2001-01-01

    The gene for cystathionine β-synthase (CBS) is located on chromosome 21 and is overexpressed in children with Down syndrome (DS), or trisomy 21. The dual purpose of the present study was to evaluate the impact of overexpression of the CBS gene on homocysteine metabolism in children with DS and to determine whether the supplementation of trisomy 21 lymphoblasts in vitro with selected nutrients would shift the genetically induced metabolic imbalance. Plasma samples were obtained from 42 children with karyotypically confirmed full trisomy 21 and from 36 normal siblings (mean age 7.4 years). Metabolites involved in homocysteine metabolism were measured and compared to those of normal siblings used as controls. Lymphocyte DNA methylation status was determined as a functional endpoint. The results indicated that plasma levels of homocysteine, methionine, S-adenosylhomocysteine, and S-adenosylmethionine were all significantly decreased in children with DS and that their lymphocyte DNA was hypermethylated relative to that in normal siblings. Plasma levels of cystathionine and cysteine were significantly increased, consistent with an increase in CBS activity. Plasma glutathione levels were significantly reduced in the children with DS and may reflect an increase in oxidative stress due to the overexpression of the superoxide dismutase gene, also located on chromosome 21. The addition of methionine, folinic acid, methyl-B12, thymidine, or dimethylglycine to the cultured trisomy 21 lymphoblastoid cells improved the metabolic profile in vitro. The increased activity of CBS in children with DS significantly alters homocysteine metabolism such that the folate-dependent resynthesis of methionine is compromised. The decreased availability of homocysteine promotes the well-established “folate trap,” creating a functional folate deficiency that may contribute to the metabolic pathology of this complex genetic disorder. PMID:11391481

  10. Myocardial Reloading After Extracorporeal Membrane Oxygenation Alters Substrate Metabolism While Promoting Protein Synthesis

    PubMed Central

    Kajimoto, Masaki; O'Kelly Priddy, Colleen M.; Ledee, Dolena R.; Xu, Chun; Isern, Nancy; Olson, Aaron K.; Rosiers, Christine Des; Portman, Michael A.

    2013-01-01

    Background Extracorporeal membrane oxygenation (ECMO) unloads the heart, providing a bridge to recovery in children after myocardial stunning. ECMO also induces stress which can adversely affect the ability to reload or wean the heart from the circuit. Metabolic impairments induced by altered loading and/or stress conditions may impact weaning. However, cardiac substrate and amino acid requirements upon weaning are unknown. We assessed the hypothesis that ventricular reloading with ECMO modulates both substrate entry into the citric acid cycle (CAC) and myocardial protein synthesis. Methods and Results Sixteen immature piglets (7.8 to 15.6 kg) were separated into 2 groups based on ventricular loading status: 8‐hour ECMO (UNLOAD) and postwean from ECMO (RELOAD). We infused into the coronary artery [2‐13C]‐pyruvate as an oxidative substrate and [13C6]‐L‐leucine as an indicator for amino acid oxidation and protein synthesis. Upon RELOAD, each functional parameter, which were decreased substantially by ECMO, recovered to near‐baseline level with the exclusion of minimum dP/dt. Accordingly, myocardial oxygen consumption was also increased, indicating that overall mitochondrial metabolism was reestablished. At the metabolic level, when compared to UNLOAD, RELOAD altered the contribution of various substrates/pathways to tissue pyruvate formation, favoring exogenous pyruvate versus glycolysis, and acetyl‐CoA formation, shifting away from pyruvate decarboxylation to endogenous substrate, presumably fatty acids. Furthermore, there was also a significant increase of tissue concentrations for all CAC intermediates (≈80%), suggesting enhanced anaplerosis, and of fractional protein synthesis rates (>70%). Conclusions RELOAD alters both cytosolic and mitochondrial energy substrate metabolism, while favoring leucine incorporation into protein synthesis rather than oxidation in the CAC. Improved understanding of factors governing these metabolic perturbations may

  11. Size matters: plasticity in metabolic scaling shows body-size may modulate responses to climate change.

    PubMed

    Carey, Nicholas; Sigwart, Julia D

    2014-08-01

    Variability in metabolic scaling in animals, the relationship between metabolic rate ( R: ) and body mass ( M: ), has been a source of debate and controversy for decades. R: is proportional to MB: , the precise value of B: much debated, but historically considered equal in all organisms. Recent metabolic theory, however, predicts B: to vary among species with ecology and metabolic level, and may also vary within species under different abiotic conditions. Under climate change, most species will experience increased temperatures, and marine organisms will experience the additional stressor of decreased seawater pH ('ocean acidification'). Responses to these environmental changes are modulated by myriad species-specific factors. Body-size is a fundamental biological parameter, but its modulating role is relatively unexplored. Here, we show that changes to metabolic scaling reveal asymmetric responses to stressors across body-size ranges; B: is systematically decreased under increasing temperature in three grazing molluscs, indicating smaller individuals were more responsive to warming. Larger individuals were, however, more responsive to reduced seawater pH in low temperatures. These alterations to the allometry of metabolism highlight abiotic control of metabolic scaling, and indicate that responses to climate warming and ocean acidification may be modulated by body-size.

  12. Size matters: plasticity in metabolic scaling shows body-size may modulate responses to climate change

    PubMed Central

    Carey, Nicholas; Sigwart, Julia D.

    2014-01-01

    Variability in metabolic scaling in animals, the relationship between metabolic rate (R) and body mass (M), has been a source of debate and controversy for decades. R is proportional to Mb, the precise value of b much debated, but historically considered equal in all organisms. Recent metabolic theory, however, predicts b to vary among species with ecology and metabolic level, and may also vary within species under different abiotic conditions. Under climate change, most species will experience increased temperatures, and marine organisms will experience the additional stressor of decreased seawater pH (‘ocean acidification’). Responses to these environmental changes are modulated by myriad species-specific factors. Body-size is a fundamental biological parameter, but its modulating role is relatively unexplored. Here, we show that changes to metabolic scaling reveal asymmetric responses to stressors across body-size ranges; b is systematically decreased under increasing temperature in three grazing molluscs, indicating smaller individuals were more responsive to warming. Larger individuals were, however, more responsive to reduced seawater pH in low temperatures. These alterations to the allometry of metabolism highlight abiotic control of metabolic scaling, and indicate that responses to climate warming and ocean acidification may be modulated by body-size. PMID:25122741

  13. Size matters: plasticity in metabolic scaling shows body-size may modulate responses to climate change.

    PubMed

    Carey, Nicholas; Sigwart, Julia D

    2014-08-01

    Variability in metabolic scaling in animals, the relationship between metabolic rate ( R: ) and body mass ( M: ), has been a source of debate and controversy for decades. R: is proportional to MB: , the precise value of B: much debated, but historically considered equal in all organisms. Recent metabolic theory, however, predicts B: to vary among species with ecology and metabolic level, and may also vary within species under different abiotic conditions. Under climate change, most species will experience increased temperatures, and marine organisms will experience the additional stressor of decreased seawater pH ('ocean acidification'). Responses to these environmental changes are modulated by myriad species-specific factors. Body-size is a fundamental biological parameter, but its modulating role is relatively unexplored. Here, we show that changes to metabolic scaling reveal asymmetric responses to stressors across body-size ranges; B: is systematically decreased under increasing temperature in three grazing molluscs, indicating smaller individuals were more responsive to warming. Larger individuals were, however, more responsive to reduced seawater pH in low temperatures. These alterations to the allometry of metabolism highlight abiotic control of metabolic scaling, and indicate that responses to climate warming and ocean acidification may be modulated by body-size. PMID:25122741

  14. Protein and leucine metabolism in maple syrup urine disease

    SciTech Connect

    Thompson, G.N.; Bresson, J.L.; Pacy, P.J.; Bonnefont, J.P.; Walter, J.H.; Leonard, J.V.; Saudubray, J.M.; Halliday, D. )

    1990-04-01

    Constant infusions of (13C)leucine and (2H5)phenylalanine were used to trace leucine and protein kinetics, respectively, in seven children with maple syrup urine disease (MSUD) and eleven controls matched for age and dietary protein intake. Despite significant elevations of plasma leucine (mean 351 mumol/l, range 224-477) in MSUD subjects, mean whole body protein synthesis (3.78 +/- 0.42 (SD) g.kg-1. 24 h-1) and catabolism (4.07 +/- 0.46) were similar to control values (3.69 +/- 0.50 and 4.09 +/- 0.50, respectively). The relationship between phenylalanine and leucine fluxes was also similar in MSUD subjects (mean phenylalanine-leucine flux ratio 0.35 +/- 0.07) and previously reported adult controls (0.33 +/- 0.02). Leucine oxidation was undetectable in four of the MSUD subjects and very low in the other three (less than 4 mumol.kg-1.h-1; controls 13-20). These results show that persistent elevation in leucine concentration has no effect on protein synthesis. The marked disturbance in leucine metabolism in MSUD did not alter the relationship between rates of catabolism of protein to phenylalanine and leucine, which provides further support for the validity of the use of a single amino acid to trace whole body protein metabolism. The minimal leucine oxidation in MSUD differs from findings in other inborn metabolic errors and indicates that in patients with classical MSUD there is no significant route of leucine disposal other than through protein synthesis.

  15. Osmolalities of selected enteral products and carbohydrate modules used to treat inherited metabolic disorders.

    PubMed

    Martin, S B; Acosta, P B

    1987-01-01

    Osmolalities of selected defined-formula products for use in treatment of inherited disorders of amino acid metabolism were measured at 12 energy concentrations. Osmotic behaviors of six carbohydrate modules as components of L-amino acid formulas were also studied. Osmolality measurements were made using a Wescor vapor pressure depression osmometer (model 5100 C). Phenyl-Free at concentrations greater than 10 kcal/oz yielded high osmolalities that exceeded the recommended level for infants. Lofenalac, Low Phe/Tyr Diet Powder, and MSUD Diet Powder at concentrations up to but no greater than 20 kcal/oz exerted osmolalities acceptable for use with infants. Low Methionine Diet Powder produced the lowest osmolality of the products tested. Differences among products can be explained by the formulations of the products, with sources of nitrogen and carbohydrate and percents of protein, carbohydrate, and fat considered. Carbohydrate type significantly affected formula osmolality; differences among carbohydrate sources can be attributed to their molecular sizes. Formulas that contained glucose exerted the highest osmolalities, while those with corn syrup or sucrose yielded the next highest. Protein Free Diet Powder, Polycose, and Moducal exerted reasonably low osmolalities.

  16. Alterations in protein metabolism during space flight and inactivity

    NASA Technical Reports Server (NTRS)

    Ferrando, Arny A.; Paddon-Jones, Doug; Wolfe, Robert R.

    2002-01-01

    Space flight and the accompanying diminished muscular activity lead to a loss of body nitrogen and muscle function. These losses may affect crew capabilities and health in long-duration missions. Space flight alters protein metabolism such that the body is unable to maintain protein synthetic rates. A concomitant hypocaloric intake and altered anabolic/catabolic hormonal profiles may contribute to or exacerbate this problem. The inactivity associated with bedrest also reduces muscle and whole-body protein synthesis. For this reason, bedrest provides a good model for the investigation of potential exercise and nutritional countermeasures to restore muscle protein synthesis. We have demonstrated that minimal resistance exercise preserves muscle protein synthesis throughout bedrest. In addition, ongoing work indicates that an essential amino acid and carbohydrate supplement may ameliorate the loss of lean body mass and muscle strength associated with 28 d of bedrest. The investigation of inactivity-induced alterations in protein metabolism, during space flight or prolonged bedrest, is applicable to clinical populations and, in a more general sense, to the problems associated with the decreased activity that occur with aging.

  17. Dopamine modulates metabolic rate and temperature sensitivity in Drosophila melanogaster.

    PubMed

    Ueno, Taro; Tomita, Jun; Kume, Shoen; Kume, Kazuhiko

    2012-01-01

    Homeothermal animals, such as mammals, maintain their body temperature by heat generation and heat dissipation, while poikilothermal animals, such as insects, accomplish it by relocating to an environment of their favored temperature. Catecholamines are known to regulate thermogenesis and metabolic rate in mammals, but their roles in other animals are poorly understood. The fruit fly, Drosophila melanogaster, has been used as a model system for the genetic studies of temperature preference behavior. Here, we demonstrate that metabolic rate and temperature sensitivity of some temperature sensitive behaviors are regulated by dopamine in Drosophila. Temperature-sensitive molecules like dTrpA1 and shi(ts) induce temperature-dependent behavioral changes, and the temperature at which the changes are induced were lowered in the dopamine transporter-defective mutant, fumin. The mutant also displays a preference for lower temperatures. This thermophobic phenotype was rescued by the genetic recovery of the dopamine transporter in dopamine neurons. Flies fed with a dopamine biosynthesis inhibitor (3-iodo-L-tyrosine), which diminishes dopamine signaling, exhibited preference for a higher temperature. Furthermore, we found that the metabolic rate is up-regulated in the fumin mutant. Taken together, dopamine has functions in the temperature sensitivity of behavioral changes and metabolic rate regulation in Drosophila, as well as its previously reported functions in arousal/sleep regulation.

  18. Tumor microenvironment derived exosomes pleiotropically modulate cancer cell metabolism

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Cancer-associated fibroblasts (CAFs) are a major cellular component of tumor microenvironment in most solid cancers. Altered cellular metabolism is a hallmark of cancer, and much of the published literature has focused on neoplastic cell-autonomous processes for these adaptations. We demonstrate tha...

  19. Role of acetylcholinesterase inhibitors in the metabolism of amyloid precursor protein.

    PubMed

    Pakaski, M; Kasa, P

    2003-06-01

    Potentiation of central cholinergic activity has been proposed as a therapeutic approach for improving the cognitive function in patients with Alzheimer's disease (AD). Increasing the acetylcholine concentration in the brain by modulating acetylcholine-sterase (AChE) activity is among the most promising therapeutic strategies. Efforts to treat the underlying pathology based on the modulation of amyloid precursor protein (APP) processing in order to decrease the accumulation of beta-amyloid are also very important. Alterations in APP metabolism have recently been proposed to play a key role in the long-lasting effects of AChE inhibitors. This review surveys recent data from in vivo and in vitro studies that have contributed to our understanding of the role of AChE inhibitors in APP processing. The regulatory mechanisms relating to the muscarinic agonist effect, protein kinase C activation and mitogen-activated protein kinase phosphorylation, involving the alpha-secretase or the 5 -UTR region of the APP gene, are also discussed. Further work is warranted to elucidate the exact roles in APP metabolism of the AChE inhibitors used in AD therapy at present. PMID:12769797

  20. Modulation of Alloimmunity by Heat Shock Proteins.

    PubMed

    Borges, Thiago J; Lang, Benjamin J; Lopes, Rafael L; Bonorino, Cristina

    2016-01-01

    The immunological mechanisms that evolved for host defense against pathogens and injury are also responsible for transplant rejection. Host rejection of foreign tissue was originally thought to be mediated mainly by T cell recognition of foreign MHC alleles. Management of solid organ transplant rejection has thus focused mainly on inhibition of T cell function and matching MHC alleles between donor and host. Recently, however, it has been demonstrated that the magnitude of the initial innate immune responses upon transplantation has a decisive impact on rejection. The exact mechanisms underlying this phenomenon have yet to be characterized. Ischemic cell death and inflammation that occur upon transplantation are synonymous with extracellular release of various heat shock proteins (Hsps), many of which have been shown to have immune-modulatory properties. Here, we review the impact of Hsps upon alloimmunity and discuss the potential use of Hsps as accessory agents to improve solid organ transplant outcomes. PMID:27555846

  1. Modulation of Alloimmunity by Heat Shock Proteins

    PubMed Central

    Borges, Thiago J.; Lang, Benjamin J.; Lopes, Rafael L.; Bonorino, Cristina

    2016-01-01

    The immunological mechanisms that evolved for host defense against pathogens and injury are also responsible for transplant rejection. Host rejection of foreign tissue was originally thought to be mediated mainly by T cell recognition of foreign MHC alleles. Management of solid organ transplant rejection has thus focused mainly on inhibition of T cell function and matching MHC alleles between donor and host. Recently, however, it has been demonstrated that the magnitude of the initial innate immune responses upon transplantation has a decisive impact on rejection. The exact mechanisms underlying this phenomenon have yet to be characterized. Ischemic cell death and inflammation that occur upon transplantation are synonymous with extracellular release of various heat shock proteins (Hsps), many of which have been shown to have immune-modulatory properties. Here, we review the impact of Hsps upon alloimmunity and discuss the potential use of Hsps as accessory agents to improve solid organ transplant outcomes. PMID:27555846

  2. Modulation of Alloimmunity by Heat Shock Proteins.

    PubMed

    Borges, Thiago J; Lang, Benjamin J; Lopes, Rafael L; Bonorino, Cristina

    2016-01-01

    The immunological mechanisms that evolved for host defense against pathogens and injury are also responsible for transplant rejection. Host rejection of foreign tissue was originally thought to be mediated mainly by T cell recognition of foreign MHC alleles. Management of solid organ transplant rejection has thus focused mainly on inhibition of T cell function and matching MHC alleles between donor and host. Recently, however, it has been demonstrated that the magnitude of the initial innate immune responses upon transplantation has a decisive impact on rejection. The exact mechanisms underlying this phenomenon have yet to be characterized. Ischemic cell death and inflammation that occur upon transplantation are synonymous with extracellular release of various heat shock proteins (Hsps), many of which have been shown to have immune-modulatory properties. Here, we review the impact of Hsps upon alloimmunity and discuss the potential use of Hsps as accessory agents to improve solid organ transplant outcomes.

  3. Expression data on liver metabolic pathway genes and proteins

    PubMed Central

    Raja Gopal Reddy, Mooli; Pavan Kumar, Chodisetti; Mahesh, Malleswarapu; Sravan Kumar, Manchiryala; Jeyakumar, Shanmugam M.

    2016-01-01

    Here, we present the expression data on various metabolic pathways of liver with special emphasize on lipid and carbohydrate metabolism and long chain polyunsaturated fatty acid (PUFA) synthesis, both at gene and protein levels. The data were obtained to understand the effect of vitamin A deficiency on the expression status (both gene and protein levels) of some of the key factors involved in lipogenesis, fatty acid oxidation, triglyceride secretion, long chain PUFA, resolvin D1 synthesis, glucose transport and glycogen synthesis of liver, using modern biology tools, such as quantitative real-time PCR (RT-PCR) and immunoblotting techniques. This data article provides the supporting evidence to the article “Vitamin A deficiency suppresses high fructose-induced triglyceride synthesis and elevates resolvin D1 levels” [1] and therefore, these data may be referred back, for comprehensive understanding and interpretations and for future studies. PMID:26909377

  4. The Involvement of Transport Proteins in Transcriptional and Metabolic Regulation

    PubMed Central

    Västermark, Åke; Saier, Milton H.

    2014-01-01

    Transport proteins have sometimes gained secondary regulatory functions that influence gene expression and metabolism. These functions allow communication with the external world via mechanistically distinctive signal transduction pathways. In this brief review we focus on three transport systems in Escherichia coli that control and coordinate carbon, exogenous hexose-phosphate and phosphorous metabolism. The transport proteins that play central roles in these processes are (1) the phosphoenolpyruvate (PEP)-dependent phosphotransferase system, PTS, (2) the glucose-6-phosphate receptor, UhpC, and (3) the phosphate-specific transporter, PstSABC, respectively. While the PTS participates in multiple complex regulatory processes, three of which are discussed here, UhpC and the Pst transporters exemplify differing strategies. PMID:24513656

  5. AMP-activated protein kinase and metabolic control

    PubMed Central

    Viollet, Benoit; Andreelli, Fabrizio

    2011-01-01

    AMP-activated protein kinase (AMPK), a phylogenetically conserved serine/threonine protein kinase, is a major regulator of cellular and whole-body energy homeostasis that coordinates metabolic pathways in order to balance nutrient supply with energy demand. It is now recognized that pharmacological activation of AMPK improves blood glucose homeostasis, lipid profile and blood pressure in insulin-resistant rodents. Indeed, AMPK activation mimics the beneficial effects of physical activity or those of calorie restriction by acting on multiple cellular targets. In addition it is now demonstrated that AMPK is one of the probable (albeit indirect) targets of major antidiabetic drugs including, the biguanides (metformin) and thiazolidinediones, as well as of insulin sensitizing adipokines (e.g., adiponectin). Taken together, such findings highlight the logic underlying the concept of targeting the AMPK pathway for the treatment of metabolic syndrome and type 2 diabetes. PMID:21484577

  6. SIZ1-Dependent Post-Translational Modification by SUMO Modulates Sugar Signaling and Metabolism in Arabidopsis thaliana.

    PubMed

    Castro, Pedro Humberto; Verde, Nuno; Lourenço, Tiago; Magalhães, Alexandre Papadopoulos; Tavares, Rui Manuel; Bejarano, Eduardo Rodríguez; Azevedo, Herlânder

    2015-12-01

    Post-translational modification mechanisms function as switches that mediate the balance between optimum growth and the response to environmental stimuli, by regulating the activity of key proteins. SUMO (small ubiquitin-like modifier) attachment, or sumoylation, is a post-translational modification that is essential for the plant stress response, also modulating hormonal circuits to co-ordinate developmental processes. The Arabidopsis SUMO E3 ligase SAP and Miz 1 (SIZ1) is the major SUMO conjugation enhancer in response to stress, and is implicated in several aspects of plant development. Here we report that known SUMO targets are over-represented in multiple carbohydrate-related proteins, suggesting a functional link between sumoylation and sugar metabolism and signaling in plants. We subsequently observed that SUMO-conjugated proteins accumulate in response to high doses of sugar in a SIZ1-dependent manner, and that the null siz1 mutant displays increased expression of sucrose and starch catabolic genes and shows reduced starch levels. We demonstrated that SIZ1 controls germination time and post-germination growth via osmotic and sugar-dependent signaling, respectively. Glucose was specifically linked to SUMO-sugar interplay, with high levels inducing root growth inhibition and aberrant root hair morphology in siz1. The use of sugar analogs and sugar marker gene expression analysis allowed us to implicate SIZ1 in a signaling pathway dependent on glucose metabolism, probably involving modulation of SNF1-related kinase 1 (SnRK1) activity.

  7. The tumor suppressor WW domain-containing oxidoreductase modulates cell metabolism

    PubMed Central

    Abu-Remaileh, Muhannad

    2015-01-01

    The WW domain-containing oxidoreductase (WWOX) encodes a tumor suppressor that is frequently altered in cancer. WWOX binds several proteins and thus is postulated to be involved in a variety of cellular processes. Interestingly, Wwox-knockout mice develop normally in utero but succumb to hypoglycemia and other metabolic defects early in life resulting in their death by 3–4 weeks of age. Cumulative evidence has linked WWOX with cellular metabolism including steroid metabolism, high-density lipoprotein cholesterol (HDL-C) metabolism, bone metabolism and, more recently, glucose metabolism. In this review, we discuss these evolving functions for WWOX and how its deletion affects cellular metabolism and neoplastic progression. PMID:25491415

  8. Whole-organism screening for modulators of fasting metabolism using transgenic zebrafish.

    PubMed

    Gut, Philipp; Stainier, Didier Y R

    2015-01-01

    Organismal energy homeostasis is maintained by complex interorgan communications making the discovery of novel drugs against metabolic diseases challenging using traditional high-throughput approaches in vitro. Here, we describe a method that rapidly identifies small molecules with an impact on organismal energy balance in vivo. Specifically, we developed a whole-organism screen for modulators of fasting metabolism using transgenic bioluminescence-reporter zebrafish for the gluconeogenic gene phosphoenolpyruvate-carboxykinase 1 (pck1).

  9. Genetic ablation of carotene oxygenases and consumption of lycopene or tomato powder diets modulates carotenoid and lipid metabolism in mice

    PubMed Central

    Ford, Nikki A.; Elsen, Amy C.; Erdman, John W.

    2013-01-01

    Carotene-15,15'-monooxygenase (CMO-I) cleaves β-carotene to form vitamin A while carotene-9’,10’-monooxygenase (CMO-II) preferentially cleaves non-provitamin A carotenoids. Recent reports indicate that beta-carotene metabolites regulate dietary lipid uptake while lycopene regulates peroxisome-proliferated activator receptor (PPAR) expression. To determine the physiologic consequences of carotenoids and their interactions with CMO-I and CMO-II, we characterized mammalian carotenoid metabolism, metabolic perturbations and lipid metabolism in female CMO-I−/− and CMO-II−/− mice fed lycopene or tomato-containing diets for 30 days. We hypothesized that there would be significant interactions between diet and genotype on carotenoid accumulation and lipid parameters. CMO-I−/− mice had higher levels of leptin, insulin and hepatic lipidosis, but lower levels of serum cholesterol. CMO-II−/− mice had increased tissue lycopene and phytofluene accumulation, reduced IGF-1 levels and cholesterol levels, but elevated liver lipids and cholesterol compared with WT mice. The diets did not modulate these genotypic perturbations, but lycopene and tomato powder did significantly decrease serum insulin-like growth factor-I. Tomato powder also reduced hepatic PPAR expression, independent of genotype. These data point to the pleiotropic actions of CMO-I and CMO-II supporting a strong role of these proteins in regulating tissue carotenoid accumulation and the lipid metabolic phenotype, as well as tomato carotenoid-independent regulation of lipid metabolism. PMID:24034573

  10. Genetic ablation of carotene oxygenases and consumption of lycopene or tomato powder diets modulate carotenoid and lipid metabolism in mice.

    PubMed

    Ford, Nikki A; Elsen, Amy C; Erdman, John W

    2013-09-01

    Carotene-15,15'-monooxygenase (CMO-I) cleaves β-carotene to form vitamin A, whereas carotene-9',10'-monooxygenase (CMO-II) preferentially cleaves non-provitamin A carotenoids. Recent reports indicate that β-carotene metabolites regulate dietary lipid uptake, whereas lycopene regulates peroxisome proliferator-activated receptor expression. To determine the physiologic consequences of carotenoids and their interactions with CMO-I and CMO-II, we characterized mammalian carotenoid metabolism, metabolic perturbations, and lipid metabolism in female CMO-I(-/-) and CMO-II(-/-) mice fed lycopene or tomato-containing diets for 30 days. We hypothesized that there would be significant interactions between diet and genotype on carotenoid accumulation and lipid parameters. CMO-I(-/-) mice had higher levels of leptin, insulin, and hepatic lipidosis but lower levels of serum cholesterol. CMO-II(-/-) mice had increased tissue lycopene and phytofluene accumulation, reduced insulin-like growth factor 1 levels and cholesterol levels, but elevated liver lipids and cholesterol compared with wild-type mice. The diets did not modulate these genotypic perturbations, but lycopene and tomato powder significantly decreased serum insulin-like growth factor 1. Tomato powder also increased hepatic peroxisome proliferator-activated receptor expression, independent of genotype. These data point to the pleiotropic actions of CMO-I and CMO-II supporting a strong role of these proteins in regulating tissue carotenoid accumulation and the lipid metabolic phenotype as well as tomato carotenoid-independent regulation of lipid metabolism. PMID:24034573

  11. Perilipin-related protein regulates lipid metabolism in C. elegans

    PubMed Central

    Chughtai, Ahmed Ali; Kaššák, Filip; Kostrouchová, Markéta; Novotný, Jan Philipp; Krause, Michael W.; Kostrouch, Zdenek

    2015-01-01

    Perilipins are lipid droplet surface proteins that contribute to fat metabolism by controlling the access of lipids to lipolytic enzymes. Perilipins have been identified in organisms as diverse as metazoa, fungi, and amoebas but strikingly not in nematodes. Here we identify the protein encoded by the W01A8.1 gene in Caenorhabditis elegans as the closest homologue and likely orthologue of metazoan perilipin. We demonstrate that nematode W01A8.1 is a cytoplasmic protein residing on lipid droplets similarly as human perilipins 1 and 2. Downregulation or elimination of W01A8.1 affects the appearance of lipid droplets resulting in the formation of large lipid droplets localized around the dividing nucleus during the early zygotic divisions. Visualization of lipid containing structures by CARS microscopy in vivo showed that lipid-containing structures become gradually enlarged during oogenesis and relocate during the first zygotic division around the dividing nucleus. In mutant embryos, the lipid containing structures show defective intracellular distribution in subsequent embryonic divisions and become gradually smaller during further development. In contrast to embryos, lipid-containing structures in enterocytes and in epidermal cells of adult animals are smaller in mutants than in wild type animals. Our results demonstrate the existence of a perilipin-related regulation of fat metabolism in nematodes and provide new possibilities for functional studies of lipid metabolism. PMID:26357594

  12. Angiogenesis and vascular functions in modulation of obesity, adipose metabolism, and insulin sensitivity.

    PubMed

    Cao, Yihai

    2013-10-01

    White and brown adipose tissues are hypervascularized and the adipose vasculature displays phenotypic and functional plasticity to coordinate with metabolic demands of adipocytes. Blood vessels not only supply nutrients and oxygen to nourish adipocytes, they also serve as a cellular reservoir to provide adipose precursor and stem cells that control adipose tissue mass and function. Multiple signaling molecules modulate the complex interplay between the vascular system and the adipocytes. Understanding fundamental mechanisms by which angiogenesis and vasculatures modulate adipocyte functions may provide new therapeutic options for treatment of obesity and metabolic disorders by targeting the adipose vasculature.

  13. Myocardial Reloading after Extracorporeal Membrane Oxygenation Alters Substrate Metabolism While Promoting Protein Synthesis

    SciTech Connect

    Kajimoto, Masaki; Priddy, Colleen M.; Ledee, Dolena; Xu, Chun; Isern, Nancy G.; Olson, Aaron; Des Rosiers, Christine; Portman, Michael A.

    2013-08-19

    Extracorporeal membrane oxygenation (ECMO) unloads the heart providing a bridge to recovery in children after myocardial stunning. Mortality after ECMO remains high.Cardiac substrate and amino acid requirements upon weaning are unknown and may impact recovery. We assessed the hypothesis that ventricular reloading modulates both substrate entry into the citric acid cycle (CAC) and myocardial protein synthesis. Fourteen immature piglets (7.8-15.6 kg) were separated into 2 groups based on ventricular loading status: 8 hour-ECMO (UNLOAD) and post-wean from ECMO (RELOAD). We infused [2-13C]-pyruvate as an oxidative substrate and [13C6]-L-leucine, as a tracer of amino acid oxidation and protein synthesis into the coronary artery. RELOAD showed marked elevations in myocardial oxygen consumption above baseline and UNLOAD. Pyruvate uptake was markedly increased though RELOAD decreased pyruvate contribution to oxidative CAC metabolism.RELOAD also increased absolute concentrations of all CAC intermediates, while maintaining or increasing 13C-molar percent enrichment. RELOAD also significantly increased cardiac fractional protein synthesis rates by >70% over UNLOAD. Conclusions: RELOAD produced high energy metabolic requirement and rebound protein synthesis. Relative pyruvate decarboxylation decreased with RELOAD while promoting anaplerotic pyruvate carboxylation and amino acid incorporation into protein rather than to the CAC for oxidation. These perturbations may serve as therapeutic targets to improve contractile function after ECMO.

  14. Metabolic impact assessment for heterologous protein production in Streptomyces lividans based on genome-scale metabolic network modeling.

    PubMed

    Lule, Ivan; D'Huys, Pieter-Jan; Van Mellaert, Lieve; Anné, Jozef; Bernaerts, Kristel; Van Impe, Jan

    2013-11-01

    The metabolic impact exerted on a microorganism due to heterologous protein production is still poorly understood in Streptomyces lividans. In this present paper, based on exometabolomic data, a proposed genome-scale metabolic network model is used to assess this metabolic impact in S. lividans. Constraint-based modeling results obtained in this work revealed that the metabolic impact due to heterologous protein production is widely distributed in the genome of S. lividans, causing both slow substrate assimilation and a shift in active pathways. Exchange fluxes that are critical for model performance have been identified for metabolites of mouse tumor necrosis factor, histidine, valine and lysine, as well as biomass. Our results unravel the interaction of heterologous protein production with intracellular metabolism of S. lividans, thus, a possible basis for further studies in relieving the metabolic burden via metabolic or bioprocess engineering.

  15. Tumor microenvironment derived exosomes pleiotropically modulate cancer cell metabolism

    PubMed Central

    Zhao, Hongyun; Yang, Lifeng; Baddour, Joelle; Achreja, Abhinav; Bernard, Vincent; Moss, Tyler; Marini, Juan C; Tudawe, Thavisha; Seviour, Elena G; San Lucas, F Anthony; Alvarez, Hector; Gupta, Sonal; Maiti, Sourindra N; Cooper, Laurence; Peehl, Donna; Ram, Prahlad T; Maitra, Anirban; Nagrath, Deepak

    2016-01-01

    Cancer-associated fibroblasts (CAFs) are a major cellular component of tumor microenvironment in most solid cancers. Altered cellular metabolism is a hallmark of cancer, and much of the published literature has focused on neoplastic cell-autonomous processes for these adaptations. We demonstrate that exosomes secreted by patient-derived CAFs can strikingly reprogram the metabolic machinery following their uptake by cancer cells. We find that CAF-derived exosomes (CDEs) inhibit mitochondrial oxidative phosphorylation, thereby increasing glycolysis and glutamine-dependent reductive carboxylation in cancer cells. Through 13C-labeled isotope labeling experiments we elucidate that exosomes supply amino acids to nutrient-deprived cancer cells in a mechanism similar to macropinocytosis, albeit without the previously described dependence on oncogenic-Kras signaling. Using intra-exosomal metabolomics, we provide compelling evidence that CDEs contain intact metabolites, including amino acids, lipids, and TCA-cycle intermediates that are avidly utilized by cancer cells for central carbon metabolism and promoting tumor growth under nutrient deprivation or nutrient stressed conditions. DOI: http://dx.doi.org/10.7554/eLife.10250.001 PMID:26920219

  16. Tumor microenvironment derived exosomes pleiotropically modulate cancer cell metabolism.

    PubMed

    Zhao, Hongyun; Yang, Lifeng; Baddour, Joelle; Achreja, Abhinav; Bernard, Vincent; Moss, Tyler; Marini, Juan C; Tudawe, Thavisha; Seviour, Elena G; San Lucas, F Anthony; Alvarez, Hector; Gupta, Sonal; Maiti, Sourindra N; Cooper, Laurence; Peehl, Donna; Ram, Prahlad T; Maitra, Anirban; Nagrath, Deepak

    2016-01-01

    Cancer-associated fibroblasts (CAFs) are a major cellular component of tumor microenvironment in most solid cancers. Altered cellular metabolism is a hallmark of cancer, and much of the published literature has focused on neoplastic cell-autonomous processes for these adaptations. We demonstrate that exosomes secreted by patient-derived CAFs can strikingly reprogram the metabolic machinery following their uptake by cancer cells. We find that CAF-derived exosomes (CDEs) inhibit mitochondrial oxidative phosphorylation, thereby increasing glycolysis and glutamine-dependent reductive carboxylation in cancer cells. Through 13C-labeled isotope labeling experiments we elucidate that exosomes supply amino acids to nutrient-deprived cancer cells in a mechanism similar to macropinocytosis, albeit without the previously described dependence on oncogenic-Kras signaling. Using intra-exosomal metabolomics, we provide compelling evidence that CDEs contain intact metabolites, including amino acids, lipids, and TCA-cycle intermediates that are avidly utilized by cancer cells for central carbon metabolism and promoting tumor growth under nutrient deprivation or nutrient stressed conditions. PMID:26920219

  17. Ethanol impairs post-prandial hepatic protein metabolism.

    PubMed Central

    De Feo, P; Volpi, E; Lucidi, P; Cruciani, G; Monacchia, F; Reboldi, G; Santeusanio, F; Bolli, G B; Brunetti, P

    1995-01-01

    The effects of acute ethanol ingestion on whole body and hepatic protein metabolism in humans are not known. To simulate social drinking, we compared the effects of the association of a mixed meal (632 kcal, 17% amino acids, 50% glucose, 33% lipids) with a bottle of either table wine (ethanol content 71 g) or water on the estimates ([1-14C]-leucine infusion) of whole body protein breakdown, oxidation, and synthesis, and on the intravascular fractional secretory rates (FSR) of hepatically (albumin, fibrinogen) and extrahepatically (IgG) synthesized plasma proteins in two randomized groups (ethanol n = 7, water n = 7) of healthy nonalcoholic volunteers. Each study was carried out for 8 h. Protein kinetics were measured in the overnight post-absorptive state, over the first 4 h, and during a meal infusion (via a nasogastric feeding tube at constant rate) combined with the oral ingestion of wine or water, over the last 4 h. When compared with water, wine ingestion during the meal reduced (P < 0.03) by 24% the rate of leucine oxidation, did not modify the estimates of whole body protein breakdown and synthesis, reduced (P < 0.01) by approximately 30% the FSR of albumin and fibrinogen, but did not affect IgG FSR. In conclusion, 70 g of ethanol, an amount usual among social drinkers, impairs hepatic protein metabolism. The habitual consumption of such amounts by reducing the synthesis and/or secretion of hepatic proteins might lead to the progressive development of liver injury and to hypoalbuminemia also in the absence of protein malnutrition. PMID:7706451

  18. Silent information regulator 1 modulator resveratrol increases brain lactate production and inhibits mitochondrial metabolism, whereas SRT1720 increases oxidative metabolism.

    PubMed

    Rowlands, Benjamin D; Lau, Chew Ling; Ryall, James G; Thomas, Donald S; Klugmann, Matthias; Beart, Philip M; Rae, Caroline D

    2015-07-01

    Silent information regulators (SIRTs) have been shown to deacetylate a range of metabolic enzymes, including those in glycolysis and the Krebs cycle, and thus alter their activity. SIRTs require NAD(+) for their activity, linking cellular energy status to enzyme activity. To examine the impact of SIRT1 modulation on oxidative metabolism, this study tests the effect of ligands that are either SIRT-activating compounds (resveratrol and SRT1720) or SIRT inhibitors (EX527) on the metabolism of (13)C-enriched substrates by guinea pig brain cortical tissue slices with (13)C and (1)H nuclear magnetic resonance spectroscopy. Resveratrol increased lactate labeling but decreased incorporation of (13)C into Krebs cycle intermediates, consistent with effects on AMPK and inhibition of the F0/F1-ATPase. By testing with resveratrol that was directly applied to astrocytes with a Seahorse analyzer, increased glycolytic shift and increased mitochondrial proton leak resulting from interactions of resveratrol with the mitochondrial electron transport chain were revealed. SRT1720, by contrast, stimulated incorporation of (13)C into Krebs cycle intermediates and reduced incorporation into lactate, although the inhibitor EX527 paradoxically also increased Krebs cycle (13)C incorporation. In summary, the various SIRT1 modulators show distinct acute effects on oxidative metabolism. The strong effects of resveratrol on the mitochondrial respiratory chain and on glycolysis suggest that caution should be used in attempts to increase bioavailability of this compound in the CNS.

  19. Modulation of apoptosis by V protein mumps virus

    PubMed Central

    2011-01-01

    Background The Urabe AM9 vaccine strain of mumps virus contains two variants of V protein: VWT (of HN-A1081 viral population) and VGly (of HN-G1081). The V protein is a promoting factor of viral replication by blocking the IFN antiviral pathway. Findings We studied the relationship between V protein variants and IFN-α2b-induced apoptosis. V proteins decrease activation of the extrinsic IFN-α2b-induced apoptotic pathway monitored by the caspase 8 activity, being the effect greater with the VWT protein. Both V proteins decrease the activity of caspase 9 of the intrinsic apoptotic pathway. In a system without IFN, the VWT and VGly proteins expression promotes activation of caspases 3 and 7. However, when the cellular system was stimulated with IFN-α, this activity decreased partially. TUNEL assay shows that for treatment with IFN-α and ibuprofen of cervical adenocarcinoma cells there is nuclear DNA fragmentation but the V protein expression reduces this process. Conclusions The reduction in the levels of caspases and DNA fragmentation, suggesting that V protein, particularly VWT protein of Urabe AM9 vaccine strain, modulates apoptosis. In addition, the VWT protein shows a protective role for cell proliferation in the presence of antiproliferative signals. PMID:21569530

  20. Environmentally modulated phosphorylation and dynamics of proteins in photosynthetic membranes.

    PubMed

    Vener, Alexander V

    2007-06-01

    Recent advances in vectorial proteomics of protein domains exposed to the surface of photosynthetic thylakoid membranes of plants and the green alga Chlamydomonas reinhardtii allowed mapping of in vivo phosphorylation sites in integral and peripheral membrane proteins. In plants, significant changes of thylakoid protein phosphorylation are observed in response to stress, particularly in photosystem II under high light or high temperature stress. Thylakoid protein phosphorylation in the algae is much more responsive to the ambient redox and light conditions, as well as to CO(2) availability. The light-dependent multiple and differential phosphorylation of CP29 linker protein in the green algae is suggested to control photosynthetic state transitions and uncoupling of light harvesting proteins from photosystem II under high light. The similar role for regulation of the dynamic distribution of light harvesting proteins in plants is proposed for the TSP9 protein, which together with other recently discovered peripheral proteins undergoes specific environment- and redox-dependent phosphorylation at the thylakoid surface. This review focuses on the environmentally modulated reversible phosphorylation of thylakoid proteins related to their membrane dynamics and affinity towards particular photosynthetic protein complexes. PMID:17184728

  1. MAPK14/p38α-dependent modulation of glucose metabolism affects ROS levels and autophagy during starvation

    PubMed Central

    Desideri, Enrico; Vegliante, Rolando; Cardaci, Simone; Nepravishta, Ridvan; Paci, Maurizio; Ciriolo, Maria Rosa

    2014-01-01

    Increased glycolytic flux is a common feature of many cancer cells, which have adapted their metabolism to maximize glucose incorporation and catabolism to generate ATP and substrates for biosynthetic reactions. Indeed, glycolysis allows a rapid production of ATP and provides metabolic intermediates required for cancer cells growth. Moreover, it makes cancer cells less sensitive to fluctuations of oxygen tension, a condition usually occurring in a newly established tumor environment. Here, we provide evidence for a dual role of MAPK14 in driving a rearrangement of glucose metabolism that contributes to limiting reactive oxygen species (ROS) production and autophagy activation in condition of nutrient deprivation. We demonstrate that MAPK14 is phosphoactivated during nutrient deprivation and affects glucose metabolism at 2 different levels: on the one hand, it increases SLC2A3 mRNA and protein levels, resulting in a higher incorporation of glucose within the cell. This event involves the MAPK14-mediated enhancement of HIF1A protein stability. On the other hand, MAPK14 mediates a metabolic shift from glycolysis to the pentose phosphate pathway (PPP) through the modulation of PFKFB3 (6-phosphofructo-2-kinase/fructose 2,6-bisphosphatase 3) degradation by the proteasome. This event requires the presence of 2 distinct degradation sequences, KEN box and DSG motif Ser273, which are recognized by 2 different E3 ligase complexes. The mutation of either motif increases PFKFB3 resistance to starvation-induced degradation. The MAPK14-driven metabolic reprogramming sustains the production of NADPH, an important cofactor for many reduction reactions and for the maintenance of the proper intracellular redox environment, resulting in reduced levels of ROS. The final effect is a reduced activation of autophagy and an increased resistance to nutrient deprivation. PMID:25046111

  2. A worm of one's own: how helminths modulate host adipose tissue function and metabolism.

    PubMed

    Guigas, Bruno; Molofsky, Ari B

    2015-09-01

    Parasitic helminths have coexisted with human beings throughout time. Success in eradicating helminths has limited helminth-induced morbidity and mortality but is also correlated with increasing rates of 'western' diseases, including metabolic syndrome and type 2 diabetes. Recent studies in mice describe how type 2 immune cells, traditionally associated with helminth infection, maintain adipose tissue homeostasis and promote adipose tissue beiging, protecting against obesity and metabolic dysfunction. Here, we review these studies and discuss how helminths and helminth-derived molecules may modulate these physiologic pathways to improve metabolic functions in specific tissues, such as adipose and liver, as well as at the whole-organism level.

  3. Protein complexes and functional modules in molecular networks

    NASA Astrophysics Data System (ADS)

    Spirin, Victor; Mirny, Leonid A.

    2003-10-01

    Proteins, nucleic acids, and small molecules form a dense network of molecular interactions in a cell. Molecules are nodes of this network, and the interactions between them are edges. The architecture of molecular networks can reveal important principles of cellular organization and function, similarly to the way that protein structure tells us about the function and organization of a protein. Computational analysis of molecular networks has been primarily concerned with node degree [Wagner, A. & Fell, D. A. (2001) Proc. R. Soc. London Ser. B 268, 1803-1810; Jeong, H., Tombor, B., Albert, R., Oltvai, Z. N. & Barabasi, A. L. (2000) Nature 407, 651-654] or degree correlation [Maslov, S. & Sneppen, K. (2002) Science 296, 910-913], and hence focused on single/two-body properties of these networks. Here, by analyzing the multibody structure of the network of protein-protein interactions, we discovered molecular modules that are densely connected within themselves but sparsely connected with the rest of the network. Comparison with experimental data and functional annotation of genes showed two types of modules: (i) protein complexes (splicing machinery, transcription factors, etc.) and (ii) dynamic functional units (signaling cascades, cell-cycle regulation, etc.). Discovered modules are highly statistically significant, as is evident from comparison with random graphs, and are robust to noise in the data. Our results provide strong support for the network modularity principle introduced by Hartwell et al. [Hartwell, L. H., Hopfield, J. J., Leibler, S. & Murray, A. W. (1999) Nature 402, C47-C52], suggesting that found modules constitute the "building blocks" of molecular networks.

  4. In Absence of the Cellular Prion Protein, Alterations in Copper Metabolism and Copper-Dependent Oxidase Activity Affect Iron Distribution

    PubMed Central

    Gasperini, Lisa; Meneghetti, Elisa; Legname, Giuseppe; Benetti, Federico

    2016-01-01

    Essential elements as copper and iron modulate a wide range of physiological functions. Their metabolism is strictly regulated by cellular pathways, since dysregulation of metal homeostasis is responsible for many detrimental effects. Neurodegenerative disorders such as Alzheimer's disease, Parkinson's disease and prion diseases are characterized by alterations of metal ions. These neurodegenerative maladies involve proteins that bind metals and mediate their metabolism through not well-defined mechanisms. Prion protein, for instance, interacts with divalent cations via multiple metal-binding sites and it modulates several metal-dependent physiological functions, such as S-nitrosylation of NMDA receptors. In this work we focused on the effect of prion protein absence on copper and iron metabolism during development and adulthood. In particular, we investigated copper and iron functional values in serum and several organs such as liver, spleen, total brain and isolated hippocampus. Our results show that iron content is diminished in prion protein-null mouse serum, while it accumulates in liver and spleen. Our data suggest that these alterations can be due to impairments in copper-dependent cerulopalsmin activity which is known to affect iron mobilization. In prion protein-null mouse total brain and hippocampus, metal ion content shows a fluctuating trend, suggesting the presence of homeostatic compensatory mechanisms. However, copper and iron functional values are likely altered also in these two organs, as indicated by the modulation of metal-binding protein expression levels. Altogether, these results reveal that the absence of the cellular prion protein impairs copper metabolism and copper-dependent oxidase activity, with ensuing alteration of iron mobilization from cellular storage compartments. PMID:27729845

  5. Cardioprotection by Phytochemicals via Antiplatelet Effects and Metabolism Modulations.

    PubMed

    Zhang, Pei-Ying

    2015-11-01

    The multi-factorial aetiology is the characteristic element of cardiac disorders. Further scientific research had recognized for a long time that platelet function is related to the risk of developing atherosclerosis. Activated blood platelets play a central role in this chronic inflammatory condition as they contribute to plaque formation within blood vessels in the early stages of atherogenesis. The present review article summarizes the cardioprotective role played by Phytochemicals via antiplatelet effects. Also, various metabolic modifications have been included that have protective effect during cardiac pathology. PMID:27352325

  6. Hyperhomocysteinemia and bleomycin hydrolase modulate the expression of mouse brain proteins involved in neurodegeneration.

    PubMed

    Suszyńska-Zajczyk, Joanna; Luczak, Magdalena; Marczak, Lukasz; Jakubowski, Hieronim

    2014-01-01

    Homocysteine (Hcy) is a risk factor for Alzheimer's disease (AD). Bleomycin hydrolase (BLMH) participates in Hcy metabolism and is also linked to AD. The inactivation of the Blmh gene in mice causes accumulation of Hcy-thiolactone in the brain and increases susceptibility to Hcy-thiolactone-induced seizures. To gain insight into brain-related Blmh function, we used two-dimensional IEF/SDS-PAGE gel electrophoresis and MALDI-TOF/TOF mass spectrometry to examine brain proteomes of Blmh-/- mice and their Blmh+/+ littermates fed with a hyperhomocysteinemic high-Met or a control diet. We found that: (1) proteins involved in brain-specific function (Ncald, Nrgn, Stmn1, Stmn2), antioxidant defenses (Aop1), cell cycle (RhoGDI1, Ran), and cytoskeleton assembly (Tbcb, CapZa2) were differentially expressed in brains of Blmh-null mice; (2) hyperhomocysteinemia amplified effects of the Blmh-/- genotype on brain protein expression; (3) proteins involved in brain-specific function (Pebp1), antioxidant defenses (Sod1, Prdx2, DJ-1), energy metabolism (Atp5d, Ak1, Pgam-B), and iron metabolism (Fth) showed differential expression in Blmh-null brains only in hyperhomocysteinemic animals; (4) most proteins regulated by the Blmh-/- genotype were also regulated by high-Met diet, albeit in the opposite direction; and (5) the differentially expressed proteins play important roles in neural development, learning, plasticity, and aging and are linked to neurodegenerative diseases, including AD. Taken together, our findings suggest that Blmh interacts with diverse cellular processes from energy metabolism and anti-oxidative defenses to cell cycle, cytoskeleton dynamics, and synaptic plasticity essential for normal brain homeostasis and that modulation of these interactions by hyperhomocysteinemia underlies the involvement of Hcy in AD.

  7. The gut microbiota modulates host amino acid and glutathione metabolism in mice.

    PubMed

    Mardinoglu, Adil; Shoaie, Saeed; Bergentall, Mattias; Ghaffari, Pouyan; Zhang, Cheng; Larsson, Erik; Bäckhed, Fredrik; Nielsen, Jens

    2015-10-16

    The gut microbiota has been proposed as an environmental factor that promotes the progression of metabolic diseases. Here, we investigated how the gut microbiota modulates the global metabolic differences in duodenum, jejunum, ileum, colon, liver, and two white adipose tissue depots obtained from conventionally raised (CONV-R) and germ-free (GF) mice using gene expression data and tissue-specific genome-scale metabolic models (GEMs). We created a generic mouse metabolic reaction (MMR) GEM, reconstructed 28 tissue-specific GEMs based on proteomics data, and manually curated GEMs for small intestine, colon, liver, and adipose tissues. We used these functional models to determine the global metabolic differences between CONV-R and GF mice. Based on gene expression data, we found that the gut microbiota affects the host amino acid (AA) metabolism, which leads to modifications in glutathione metabolism. To validate our predictions, we measured the level of AAs and N-acetylated AAs in the hepatic portal vein of CONV-R and GF mice. Finally, we simulated the metabolic differences between the small intestine of the CONV-R and GF mice accounting for the content of the diet and relative gene expression differences. Our analyses revealed that the gut microbiota influences host amino acid and glutathione metabolism in mice.

  8. The gut microbiota modulates host amino acid and glutathione metabolism in mice

    PubMed Central

    Mardinoglu, Adil; Shoaie, Saeed; Bergentall, Mattias; Ghaffari, Pouyan; Zhang, Cheng; Larsson, Erik; Bäckhed, Fredrik; Nielsen, Jens

    2015-01-01

    The gut microbiota has been proposed as an environmental factor that promotes the progression of metabolic diseases. Here, we investigated how the gut microbiota modulates the global metabolic differences in duodenum, jejunum, ileum, colon, liver, and two white adipose tissue depots obtained from conventionally raised (CONV-R) and germ-free (GF) mice using gene expression data and tissue-specific genome-scale metabolic models (GEMs). We created a generic mouse metabolic reaction (MMR) GEM, reconstructed 28 tissue-specific GEMs based on proteomics data, and manually curated GEMs for small intestine, colon, liver, and adipose tissues. We used these functional models to determine the global metabolic differences between CONV-R and GF mice. Based on gene expression data, we found that the gut microbiota affects the host amino acid (AA) metabolism, which leads to modifications in glutathione metabolism. To validate our predictions, we measured the level of AAs and N-acetylated AAs in the hepatic portal vein of CONV-R and GF mice. Finally, we simulated the metabolic differences between the small intestine of the CONV-R and GF mice accounting for the content of the diet and relative gene expression differences. Our analyses revealed that the gut microbiota influences host amino acid and glutathione metabolism in mice. PMID:26475342

  9. SIRT3 regulates cellular iron metabolism and cancer growth by repressing iron regulatory protein 1.

    PubMed

    Jeong, S M; Lee, J; Finley, L W S; Schmidt, P J; Fleming, M D; Haigis, M C

    2015-04-16

    Iron metabolism is essential for many cellular processes, including oxygen transport, respiration and DNA synthesis, and many cancer cells exhibit dysregulation in iron metabolism. Maintenance of cellular iron homeostasis is regulated by iron regulatory proteins (IRPs), which control the expression of iron-related genes by binding iron-responsive elements (IREs) of target mRNAs. Here, we report that mitochondrial SIRT3 regulates cellular iron metabolism by modulating IRP1 activity. SIRT3 loss increases reactive oxygen species production, leading to elevated IRP1 binding to IREs. As a consequence, IRP1 target genes, such as the transferrin receptor (TfR1), a membrane-associated glycoprotein critical for iron uptake and cell proliferation, are controlled by SIRT3. Importantly, SIRT3 deficiency results in a defect in cellular iron homeostasis. SIRT3 null cells contain high levels of iron and lose iron-dependent TfR1 regulation. Moreover, SIRT3 null mice exhibit higher levels of iron and TfR1 expression in the pancreas. We found that the regulation of iron uptake and TfR1 expression contribute to the tumor-suppressive activity of SIRT3. Indeed, SIRT3 expression is negatively correlated with TfR1 expression in human pancreatic cancers. SIRT3 overexpression decreases TfR1 expression by inhibiting IRP1 and represses proliferation in pancreatic cancer cells. Our data uncover a novel role of SIRT3 in cellular iron metabolism through IRP1 regulation and suggest that SIRT3 functions as a tumor suppressor, in part, by modulating cellular iron metabolism. PMID:24909164

  10. The ATP-binding cassette transporter-2 (ABCA2) regulates esterification of plasma membrane cholesterol by modulation of sphingolipid metabolism

    PubMed Central

    Davis, Warren

    2015-01-01

    The ATP-binding cassette transporters are a large family (~ 48 genes divided into seven families A–G) of proteins that utilize the energy of ATP-hydrolysis to pump substrates across lipid bilayers against a concentration gradient. The ABC “A” subfamily is comprised of 13 members and transport sterols, phospholipids and bile acids. ABCA2 is the most abundant ABC transporter in human and rodent brain with highest expression in oligodendrocytes, although it is also expressed in neurons. Several groups have studied a possible connection between ABCA2 and Alzheimer’s disease as well as early atherosclerosis. ABCA2 expression levels have been associated with changes in cholesterol and sphingolipid metabolism. In this paper, we hypothesized that ABCA2 expression level may regulate esterification of plasma membrane-derived cholesterol by modulation of sphingolipid metabolism. ABCA2 overexpression in N2a neuroblastoma cells was associated with an altered bilayer distribution of the sphingolipid ceramide that inhibited acylCoA:cholesterol acyltransferase (ACAT) activity and cholesterol esterification. In contrast, depletion of endogenous ABCA2 in the rat schwannoma cell line D6P2T increased esterification of plasma membrane cholesterol following treatment with exogenous bacterial sphingomyelinase. These findings suggest that control of ABCA2 expression level may be a key locus of regulation for esterification of plasma membrane-derived cholesterol through modulation of sphingolipid metabolism. PMID:24201375

  11. Modulation of fructokinase activity of potato (Solanum tuberosum) results in substantial shifts in tuber metabolism.

    PubMed

    Davies, Howard V; Shepherd, Louise V T; Burrell, Michael M; Carrari, Fernando; Urbanczyk-Wochniak, Ewa; Leisse, Andrea; Hancock, Robert D; Taylor, Mark; Viola, Roberto; Ross, Heather; McRae, Diane; Willmitzer, Lothar; Fernie, Alisdair R

    2005-07-01

    Potato plants (Solanum tuberosum L. cvs Desiree and Record) transformed with sense and antisense constructs of a cDNA encoding the potato fructokinase StFK1 exhibited altered transcription of this gene, altered amount of protein and altered enzyme activities. Measurement of the maximal catalytic activity of fructokinase revealed a 2-fold variation in leaf (from 90 to 180% of wild type activity) and either a 10- or 30-fold variation in tuber (from 10 or 30% to 300% in Record and Desiree, respectively) activity. The comparative effect of the antisense construct in leaf and tuber tissue suggests that this isoform is only a minor contributor to the total fructokinase activity in the leaf but the predominant isoform in the tuber. Antisense inhibition of the fructokinase resulted in a reduced tuber yield; however, its overexpression had no impact on this parameter. The modulation of fructokinase activity had few, consistent effects on carbohydrate levels, with the exception of a general increase in glucose content in the antisense lines, suggesting that this enzyme is not important for the control of starch synthesis. However, when metabolic fluxes were estimated, it became apparent that the transgenic lines display a marked shift in metabolism, with the rate of redistribution of radiolabel to sucrose markedly affected by the activity of fructokinase. These data suggest an important role for fructokinase, acting in concert with sucrose synthase, in maintaining a balance between sucrose synthesis and degradation by a mechanism independent of that controlled by the hexose phosphate-mediated activation of sucrose phosphate synthase. PMID:15890680

  12. Modulation of fructokinase activity of potato (Solanum tuberosum) results in substantial shifts in tuber metabolism.

    PubMed

    Davies, Howard V; Shepherd, Louise V T; Burrell, Michael M; Carrari, Fernando; Urbanczyk-Wochniak, Ewa; Leisse, Andrea; Hancock, Robert D; Taylor, Mark; Viola, Roberto; Ross, Heather; McRae, Diane; Willmitzer, Lothar; Fernie, Alisdair R

    2005-07-01

    Potato plants (Solanum tuberosum L. cvs Desiree and Record) transformed with sense and antisense constructs of a cDNA encoding the potato fructokinase StFK1 exhibited altered transcription of this gene, altered amount of protein and altered enzyme activities. Measurement of the maximal catalytic activity of fructokinase revealed a 2-fold variation in leaf (from 90 to 180% of wild type activity) and either a 10- or 30-fold variation in tuber (from 10 or 30% to 300% in Record and Desiree, respectively) activity. The comparative effect of the antisense construct in leaf and tuber tissue suggests that this isoform is only a minor contributor to the total fructokinase activity in the leaf but the predominant isoform in the tuber. Antisense inhibition of the fructokinase resulted in a reduced tuber yield; however, its overexpression had no impact on this parameter. The modulation of fructokinase activity had few, consistent effects on carbohydrate levels, with the exception of a general increase in glucose content in the antisense lines, suggesting that this enzyme is not important for the control of starch synthesis. However, when metabolic fluxes were estimated, it became apparent that the transgenic lines display a marked shift in metabolism, with the rate of redistribution of radiolabel to sucrose markedly affected by the activity of fructokinase. These data suggest an important role for fructokinase, acting in concert with sucrose synthase, in maintaining a balance between sucrose synthesis and degradation by a mechanism independent of that controlled by the hexose phosphate-mediated activation of sucrose phosphate synthase.

  13. Hydrogen sulfide-releasing aspirin modulates xenobiotic metabolizing enzymes in vitro and in vivo.

    PubMed

    Chattopadhyay, Mitali; Kodela, Ravinder; Nath, Niharika; Street, Cherease R; Velázquez-Martínez, Carlos A; Boring, Daniel; Kashfi, Khosrow

    2012-03-15

    The balance between phase-I carcinogen-activating and phase-II detoxifying xenobiotic metabolizing enzymes is critical to determining an individual's risk for cancer. We evaluated the effect of Hydrogen sulfide-releasing aspirin (HS-ASA) on xenobiotic metabolizing enzymes in HT-29 human colon and Hepa 1c1c7 mouse liver adenocarcinoma cells and in Wistar rats. HS-ASA inhibited the growth of HT-29 and Hepa 1c1c7 cells, with an IC(50) of 3.2 ± 0.3 μM and 4.2 ± 0.4 μM, respectively. The IC(50) for ASA in both cell lines was greater than 5000 μM at 24h. In these cell lines, HS-ASA caused a dose-dependent increase in activity and expression of the phase-II enzymes glutathione S-transferase (GST) and NAD(P)H:quinoneoxireductase (NQO1). It also caused an increase in UDP-glucuronosyltransferase (UGT) expression. The levels of CYP 1A1 a phase-I enzyme was increased by HS-ASA in both cell lines. Pretreatment of cells with NaF, an esterase inhibitor, abrogated the HS-ASA-mediated increases in NQO1 enzyme activity. HS-ASA increased the protein levels of the transcription factor Nrf2, which is a regulator of the phase-II enzymes. In vivo, HS-ASA at 100mg/kg/day had no effect on rat's weights; it induced a 3.4-fold and 1.4-fold increase in hepatic GST and NQO1 enzyme activities, respectively. GST and NQO1 protein levels were also increased. In contrast to that in cultured cells, CYP 1A1 protein levels were not altered in vivo. Therefore, HS-ASA induces phase-II enzymes, at least in part, through the action of H(2)S and by modulating Nrf2; these effects may be part of its mechanism of action against carcinogenesis.

  14. The Effect of Protein Mass Modulation on Human Dihydrofolate Reductase.

    PubMed

    Francis, Kevin; Sapienza, Paul J; Lee, Andrew L; Kohen, Amnon

    2016-02-23

    Dihydrofolate reductase (DHFR) from Escherichia coli has long served as a model enzyme with which to elucidate possible links between protein dynamics and the catalyzed reaction. Such physical properties of its human counterpart have not been rigorously studied so far, but recent computer-based simulations suggest that these two DHFRs differ significantly in how closely coupled the protein dynamics and the catalyzed C-H → C hydride transfer step are. To test this prediction, two contemporary probes for studying the effect of protein dynamics on catalysis were combined here: temperature dependence of intrinsic kinetic isotope effects (KIEs), which are sensitive to the physical nature of the chemical step, and protein mass modulation, which slows down fast dynamics (femto- to picosecond time scale) throughout the protein. The intrinsic H/T KIEs of human DHFR, like those of E. coli DHFR, are shown to be temperature-independent in the range from 5 to 45 °C, indicating fast sampling of donor and acceptor distances (DADs) at the reaction's transition state (or tunneling ready state, TRS). Mass modulation of these enzymes through isotopic labeling with (13)C, (15)N, and (2)H at nonexchangeable hydrogens yields an 11% heavier enzyme. The additional mass has no effect on the intrinsic KIEs of the human enzyme. This finding indicates that the mass modulation of the human DHFR affects neither DAD distribution nor the DAD's conformational sampling dynamics. Furthermore, reduction in the enzymatic turnover number and the dissociation rate constant for the product indicate that the isotopic substitution affects kinetic steps that are not the catalyzed C-H → C hydride transfer. The findings are discussed in terms of fast dynamics and their role in catalysis, the comparison of calculations and experiments, and the interpretation of isotopically modulated heavy enzymes in general.

  15. The Effect of Protein Mass Modulation on Human Dihydrofolate Reductase.

    PubMed

    Francis, Kevin; Sapienza, Paul J; Lee, Andrew L; Kohen, Amnon

    2016-02-23

    Dihydrofolate reductase (DHFR) from Escherichia coli has long served as a model enzyme with which to elucidate possible links between protein dynamics and the catalyzed reaction. Such physical properties of its human counterpart have not been rigorously studied so far, but recent computer-based simulations suggest that these two DHFRs differ significantly in how closely coupled the protein dynamics and the catalyzed C-H → C hydride transfer step are. To test this prediction, two contemporary probes for studying the effect of protein dynamics on catalysis were combined here: temperature dependence of intrinsic kinetic isotope effects (KIEs), which are sensitive to the physical nature of the chemical step, and protein mass modulation, which slows down fast dynamics (femto- to picosecond time scale) throughout the protein. The intrinsic H/T KIEs of human DHFR, like those of E. coli DHFR, are shown to be temperature-independent in the range from 5 to 45 °C, indicating fast sampling of donor and acceptor distances (DADs) at the reaction's transition state (or tunneling ready state, TRS). Mass modulation of these enzymes through isotopic labeling with (13)C, (15)N, and (2)H at nonexchangeable hydrogens yields an 11% heavier enzyme. The additional mass has no effect on the intrinsic KIEs of the human enzyme. This finding indicates that the mass modulation of the human DHFR affects neither DAD distribution nor the DAD's conformational sampling dynamics. Furthermore, reduction in the enzymatic turnover number and the dissociation rate constant for the product indicate that the isotopic substitution affects kinetic steps that are not the catalyzed C-H → C hydride transfer. The findings are discussed in terms of fast dynamics and their role in catalysis, the comparison of calculations and experiments, and the interpretation of isotopically modulated heavy enzymes in general. PMID:26813442

  16. Modulation of gut microbiota in the management of metabolic disorders: the prospects and challenges.

    PubMed

    Erejuwa, Omotayo O; Sulaiman, Siti A; Ab Wahab, Mohd S

    2014-01-01

    The gut microbiota plays a number of important roles including digestion, metabolism, extraction of nutrients, synthesis of vitamins, prevention against pathogen colonization, and modulation of the immune system. Alterations or changes in composition and biodiversity of the gut microbiota have been associated with many gastrointestinal tract (GIT) disorders such as inflammatory bowel disease and colon cancer. Recent evidence suggests that altered composition and diversity of gut microbiota may play a role in the increased prevalence of metabolic diseases. This review article has two main objectives. First, it underscores approaches (such as probiotics, prebiotics, antimicrobial agents, bariatric surgery, and weight loss strategies) and their prospects in modulating the gut microbiota in the management of metabolic diseases. Second, it highlights some of the current challenges and discusses areas of future research as it relates to the gut microbiota and metabolic diseases. The prospect of modulating the gut microbiota seems promising. However, considering that research investigating the role of gut microbiota in metabolic diseases is still in its infancy, more rigorous and well-designed in vitro, animal and clinical studies are needed. PMID:24608927

  17. Modulation of Gut Microbiota in the Management of Metabolic Disorders: The Prospects and Challenges

    PubMed Central

    Erejuwa, Omotayo O.; Sulaiman, Siti A.; Ab Wahab, Mohd S.

    2014-01-01

    The gut microbiota plays a number of important roles including digestion, metabolism, extraction of nutrients, synthesis of vitamins, prevention against pathogen colonization, and modulation of the immune system. Alterations or changes in composition and biodiversity of the gut microbiota have been associated with many gastrointestinal tract (GIT) disorders such as inflammatory bowel disease and colon cancer. Recent evidence suggests that altered composition and diversity of gut microbiota may play a role in the increased prevalence of metabolic diseases. This review article has two main objectives. First, it underscores approaches (such as probiotics, prebiotics, antimicrobial agents, bariatric surgery, and weight loss strategies) and their prospects in modulating the gut microbiota in the management of metabolic diseases. Second, it highlights some of the current challenges and discusses areas of future research as it relates to the gut microbiota and metabolic diseases. The prospect of modulating the gut microbiota seems promising. However, considering that research investigating the role of gut microbiota in metabolic diseases is still in its infancy, more rigorous and well-designed in vitro, animal and clinical studies are needed. PMID:24608927

  18. Metabolic control in a state of decreased activation: modulation of red cell metabolism.

    PubMed

    Jevning, R; Wilson, A F; Pirkle, H; O'Halloran, J P; Walsh, R N

    1983-11-01

    Very little is known in depth of the biochemical and physiological changes induced at the cellular level by human behavioral states. For study of the physiology of behavior at this level, the erythrocyte may be useful, because it is readily available and its metabolism and metabolic control are comparatively well understood. In this report we describe a marked decline of red cell glycolytic rate induced by the transcendental meditation technique (TM). This decline was significantly correlated with decreased plasma lactate concentration and with relaxation as indicated by electrodermal response. The occurrence of sleep was not correlated with the metabolic changes. The observed lack of variation of blood pH, blood gases, glucose, and hematocrit in this behavior implies that the decrease of erythrocyte metabolism is not an epiphenomenon of respiratory change or substrate availability. Based upon further measurements indicating persisting alteration of the red blood cell, we suggest the possibility of attachment of a humoral agent(s) to the cell in the mechanism of this effect. This behavioral effect is unique, and the effector(s) responsible may increase our understanding of metabolic control of the erythrocyte and of TM.

  19. Alternative modulation of protein–protein interactions by small molecules

    PubMed Central

    Fischer, Gerhard; Rossmann, Maxim; Hyvönen, Marko

    2015-01-01

    Protein–protein interactions (PPI) have become increasingly popular drug targets, with a number of promising compounds currently in clinical trials. Recent research shows, that PPIs can be modulated in more ways than direct inhibition, where novel non-competitive modes of action promise a solution for the difficult nature of PPI drug discovery. Here, we review recently discovered PPI modulators in light of their mode of action and categorise them as disrupting versus stabilising, orthosteric versus allosteric and by their ability to affect the proteins’ dynamics. We also give recent examples of compounds successful in the clinic, analyse their physicochemical properties and discuss how to overcome the hurdles in discovering alternative modes of modulation. PMID:25935873

  20. Protein S-glutathionlyation links energy metabolism to redox signaling in mitochondria

    PubMed Central

    Mailloux, Ryan J.; Treberg, Jason R.

    2015-01-01

    At its core mitochondrial function relies on redox reactions. Electrons stripped from nutrients are used to form NADH and NADPH, electron carriers that are similar in structure but support different functions. NADH supports ATP production but also generates reactive oxygen species (ROS), superoxide (O2·-) and hydrogen peroxide (H2O2). NADH-driven ROS production is counterbalanced by NADPH which maintains antioxidants in an active state. Mitochondria rely on a redox buffering network composed of reduced glutathione (GSH) and peroxiredoxins (Prx) to quench ROS generated by nutrient metabolism. As H2O2 is quenched, NADPH is expended to reactivate antioxidant networks and reset the redox environment. Thus, the mitochondrial redox environment is in a constant state of flux reflecting changes in nutrient and ROS metabolism. Changes in redox environment can modulate protein function through oxidation of protein cysteine thiols. Typically cysteine oxidation is considered to be mediated by H2O2 which oxidizes protein thiols (SH) forming sulfenic acid (SOH). However, problems begin to emerge when one critically evaluates the regulatory function of SOH. Indeed SOH formation is slow, non-specific, and once formed SOH reacts rapidly with a variety of molecules. By contrast, protein S-glutathionylation (PGlu) reactions involve the conjugation and removal of glutathione moieties from modifiable cysteine residues. PGlu reactions are driven by fluctuations in the availability of GSH and oxidized glutathione (GSSG) and thus should be exquisitely sensitive to changes ROS flux due to shifts in the glutathione pool in response to varying H2O2 availability. Here, we propose that energy metabolism-linked redox signals originating from mitochondria are mediated indirectly by H2O2 through the GSH redox buffering network in and outside mitochondria. This proposal is based on several observations that have shown that unlike other redox modifications PGlu reactions fulfill the requisite

  1. Protein S-glutathionlyation links energy metabolism to redox signaling in mitochondria.

    PubMed

    Mailloux, Ryan J; Treberg, Jason R

    2016-08-01

    At its core mitochondrial function relies on redox reactions. Electrons stripped from nutrients are used to form NADH and NADPH, electron carriers that are similar in structure but support different functions. NADH supports ATP production but also generates reactive oxygen species (ROS), superoxide (O2(·-)) and hydrogen peroxide (H2O2). NADH-driven ROS production is counterbalanced by NADPH which maintains antioxidants in an active state. Mitochondria rely on a redox buffering network composed of reduced glutathione (GSH) and peroxiredoxins (Prx) to quench ROS generated by nutrient metabolism. As H2O2 is quenched, NADPH is expended to reactivate antioxidant networks and reset the redox environment. Thus, the mitochondrial redox environment is in a constant state of flux reflecting changes in nutrient and ROS metabolism. Changes in redox environment can modulate protein function through oxidation of protein cysteine thiols. Typically cysteine oxidation is considered to be mediated by H2O2 which oxidizes protein thiols (SH) forming sulfenic acid (SOH). However, problems begin to emerge when one critically evaluates the regulatory function of SOH. Indeed SOH formation is slow, non-specific, and once formed SOH reacts rapidly with a variety of molecules. By contrast, protein S-glutathionylation (PGlu) reactions involve the conjugation and removal of glutathione moieties from modifiable cysteine residues. PGlu reactions are driven by fluctuations in the availability of GSH and oxidized glutathione (GSSG) and thus should be exquisitely sensitive to changes ROS flux due to shifts in the glutathione pool in response to varying H2O2 availability. Here, we propose that energy metabolism-linked redox signals originating from mitochondria are mediated indirectly by H2O2 through the GSH redox buffering network in and outside mitochondria. This proposal is based on several observations that have shown that unlike other redox modifications PGlu reactions fulfill the requisite

  2. Metabolic Flux Ratio Analysis of Genetic and Environmental Modulations of Escherichia coli Central Carbon Metabolism

    PubMed Central

    Sauer, Uwe; Lasko, Daniel R.; Fiaux, Jocelyne; Hochuli, Michel; Glaser, Ralf; Szyperski, Thomas; Wüthrich, Kurt; Bailey, James E.

    1999-01-01

    The response of Escherichia coli central carbon metabolism to genetic and environmental manipulation has been studied by use of a recently developed methodology for metabolic flux ratio (METAFoR) analysis; this methodology can also directly reveal active metabolic pathways. Generation of fluxome data arrays by use of the METAFoR approach is based on two-dimensional 13C-1H correlation nuclear magnetic resonance spectroscopy with fractionally labeled biomass and, in contrast to metabolic flux analysis, does not require measurements of extracellular substrate and metabolite concentrations. METAFoR analyses of E. coli strains that moderately overexpress phosphofructokinase, pyruvate kinase, pyruvate decarboxylase, or alcohol dehydrogenase revealed that only a few flux ratios change in concert with the overexpression of these enzymes. Disruption of both pyruvate kinase isoenzymes resulted in altered flux ratios for reactions connecting the phosphoenolpyruvate (PEP) and pyruvate pools but did not significantly alter central metabolism. These data indicate remarkable robustness and rigidity in central carbon metabolism in the presence of genetic variation. More significant physiological changes and flux ratio differences were seen in response to altered environmental conditions. For example, in ammonia-limited chemostat cultures, compared to glucose-limited chemostat cultures, a reduced fraction of PEP molecules was derived through at least one transketolase reaction, and there was a higher relative contribution of anaplerotic PEP carboxylation than of the tricarboxylic acid (TCA) cycle for oxaloacetate synthesis. These two parameters also showed significant variation between aerobic and anaerobic batch cultures. Finally, two reactions catalyzed by PEP carboxykinase and malic enzyme were identified by METAFoR analysis; these had previously been considered absent in E. coli cells grown in glucose-containing media. Backward flux from the TCA cycle to glycolysis, as

  3. Arabidopsis ribosomal proteins control vacuole trafficking and developmental programs through the regulation of lipid metabolism.

    PubMed

    Li, Ruixi; Sun, Ruobai; Hicks, Glenn R; Raikhel, Natasha V

    2015-01-01

    The vacuole is the most prominent compartment in plant cells and is important for ion and protein storage. In our effort to search for key regulators in the plant vacuole sorting pathway, ribosomal large subunit 4 (rpl4d) was identified as a translational mutant defective in both vacuole trafficking and normal development. Polysome profiling of the rpl4d mutant showed reduction in polysome-bound mRNA compared with wild-type, but no significant change in the general mRNA distribution pattern. Ribsomal profiling data indicated that genes in the lipid metabolism pathways were translationally down-regulated in the rpl4d mutant. Live imaging studies by Nile red staining suggested that both polar and nonpolar lipid accumulation was reduced in meristem tissues of rpl4d mutants. Pharmacological evidence showed that sterol and sphingolipid biosynthetic inhibitors can phenocopy the defects of the rpl4d mutant, including an altered vacuole trafficking pattern. Genetic evidence from lipid biosynthetic mutants indicates that alteration in the metabolism of either sterol or sphingolipid biosynthesis resulted in vacuole trafficking defects, similar to the rpl4d mutant. Tissue-specific complementation with key enzymes from lipid biosynthesis pathways can partially rescue both vacuole trafficking and auxin-related developmental defects in the rpl4d mutant. These results indicate that lipid metabolism modulates auxin-mediated tissue differentiation and endomembrane trafficking pathways downstream of ribosomal protein function.

  4. The modulation of leaf metabolism plays a role in salt tolerance of Cymodocea nodosa exposed to hypersaline stress in mesocosms.

    PubMed

    Piro, Amalia; Marín-Guirao, Lázaro; Serra, Ilia A; Spadafora, Antonia; Sandoval-Gil, José M; Bernardeau-Esteller, Jaime; Fernandez, Juan M R; Mazzuca, Silvia

    2015-01-01

    Applying proteomics, we tested the physiological responses of the euryhaline seagrass Cymodocea nodosa to deliberate manipulation of salinity in a mesocosm system. Plants were subjected to a chronic hypersaline condition (43 psu) to compare protein expression and plant photochemistry responses after 15 and 30 days of exposure with those of plants cultured under normal/ambient saline conditions (37 psu). Results showed a general decline in the expression level of leaf proteins in hypersaline stressed plants, with more intense reductions after long-lasting exposure. Specifically, the carbon-fixing enzyme RuBisCo displayed a lower accumulation level in stressed plants relative to controls. In contrast, the key enzymes involved in the regulation of glycolysis, cytosolic glyceraldehyde-3-phosphate dehydrogenase, enolase 2 and triose-phosphate isomerase, showed significantly higher accumulation levels. These responses suggested a shift in carbon metabolism in stressed plants. Hypersaline stress also induced a significant alteration of the photosynthetic physiology of C. nodosa by means of a down-regulation in structural proteins and enzymes of both PSII and PSI. However we found an over-expression of the cytochrome b559 alpha subunit of the PSII initial complex, which is a receptor for the PSII core proteins involved in biogenesis or repair processes and therefore potentially involved in the absence of effects at the photochemical level of stressed plants. As expected hypersalinity also affects vacuolar metabolism by increasing the leaf cell turgor pressure and enhancing the up-take of Na(+) by over-accumulating the tonoplast specific intrinsic protein pyrophosphate-energized inorganic pyrophosphatase (H(+)-PPase) coupled to the Na(+)/H(+)-antiporter. The modulation of carbon metabolism and the enhancement of vacuole capacity in Na(+) sequestration and osmolarity changes are discussed in relation to salt tolerance of C. nodosa.

  5. The modulation of leaf metabolism plays a role in salt tolerance of Cymodocea nodosa exposed to hypersaline stress in mesocosms

    PubMed Central

    Piro, Amalia; Marín-Guirao, Lázaro; Serra, Ilia A.; Spadafora, Antonia; Sandoval-Gil, José M.; Bernardeau-Esteller, Jaime; Fernandez, Juan M. R.; Mazzuca, Silvia

    2015-01-01

    Applying proteomics, we tested the physiological responses of the euryhaline seagrass Cymodocea nodosa to deliberate manipulation of salinity in a mesocosm system. Plants were subjected to a chronic hypersaline condition (43 psu) to compare protein expression and plant photochemistry responses after 15 and 30 days of exposure with those of plants cultured under normal/ambient saline conditions (37 psu). Results showed a general decline in the expression level of leaf proteins in hypersaline stressed plants, with more intense reductions after long-lasting exposure. Specifically, the carbon-fixing enzyme RuBisCo displayed a lower accumulation level in stressed plants relative to controls. In contrast, the key enzymes involved in the regulation of glycolysis, cytosolic glyceraldehyde-3-phosphate dehydrogenase, enolase 2 and triose-phosphate isomerase, showed significantly higher accumulation levels. These responses suggested a shift in carbon metabolism in stressed plants. Hypersaline stress also induced a significant alteration of the photosynthetic physiology of C. nodosa by means of a down-regulation in structural proteins and enzymes of both PSII and PSI. However we found an over-expression of the cytochrome b559 alpha subunit of the PSII initial complex, which is a receptor for the PSII core proteins involved in biogenesis or repair processes and therefore potentially involved in the absence of effects at the photochemical level of stressed plants. As expected hypersalinity also affects vacuolar metabolism by increasing the leaf cell turgor pressure and enhancing the up-take of Na+ by over-accumulating the tonoplast specific intrinsic protein pyrophosphate-energized inorganic pyrophosphatase (H(+)-PPase) coupled to the Na+/H+-antiporter. The modulation of carbon metabolism and the enhancement of vacuole capacity in Na+ sequestration and osmolarity changes are discussed in relation to salt tolerance of C. nodosa. PMID:26167167

  6. Transcriptional Profiles of Drought-Related Genes in Modulating Metabolic Processes and Antioxidant Defenses in Lolium multiflorum

    PubMed Central

    Pan, Ling; Zhang, Xinquan; Wang, Jianping; Ma, Xiao; Zhou, Meiliang; Huang, LinKai; Nie, Gang; Wang, Pengxi; Yang, Zhongfu; Li, Ji

    2016-01-01

    Drought is a major environmental stress that limits growth and development of cool-season annual grasses. Drought transcriptional profiles of resistant and susceptible lines were studied to understand the molecular mechanisms of drought tolerance in annual ryegrass (Lolium multiflorum L.). A total of 4718 genes exhibited significantly differential expression in two L. multiflorum lines. Additionally, up-regulated genes associated with drought response in the resistant lines were compared with susceptible lines. Gene ontology enrichment and pathway analyses revealed that genes partially encoding drought-responsive proteins as key regulators were significantly involved in carbon metabolism, lipid metabolism, and signal transduction. Comparable gene expression was used to identify the genes that contribute to the high drought tolerance in resistant lines of annual ryegrass. Moreover, we proposed the hypothesis that short-term drought have a beneficial effect on oxidation stress, which may be ascribed to a direct effect on the drought tolerance of annual ryegrass. Evidence suggests that some of the genes encoding antioxidants (HPTs, GGT, AP, 6-PGD, and G6PDH) function as antioxidant in lipid metabolism and signal transduction pathways, which have indispensable and promoting roles in drought resistance. This study provides the first transcriptome data on the induction of drought-related gene expression in annual ryegrass, especially via modulation of metabolic homeostasis, signal transduction, and antioxidant defenses to improve drought tolerance response to short-term drought stress. PMID:27200005

  7. Novel Allosteric Modulators of G Protein-coupled Receptors*

    PubMed Central

    Gentry, Patrick R.; Sexton, Patrick M.; Christopoulos, Arthur

    2015-01-01

    G protein-coupled receptors (GPCRs) are allosteric proteins, because their signal transduction relies on interactions between topographically distinct, yet conformationally linked, domains. Much of the focus on GPCR allostery in the new millennium, however, has been on modes of targeting GPCR allosteric sites with chemical probes due to the potential for novel therapeutics. It is now apparent that some GPCRs possess more than one targetable allosteric site, in addition to a growing list of putative endogenous modulators. Advances in structural biology are also shedding new insights into mechanisms of allostery, although the complexities of candidate allosteric drugs necessitate rigorous biological characterization. PMID:26100627

  8. Physiological and Pathogenic Roles of Prolyl Isomerase Pin1 in Metabolic Regulations via Multiple Signal Transduction Pathway Modulations

    PubMed Central

    Nakatsu, Yusuke; Matsunaga, Yasuka; Yamamotoya, Takeshi; Ueda, Koji; Inoue, Yuki; Mori, Keiichi; Sakoda, Hideyuki; Fujishiro, Midori; Ono, Hiraku; Kushiyama, Akifumi; Asano, Tomoichiro

    2016-01-01

    Prolyl isomerases are divided into three groups, the FKBP family, Cyclophilin and the Parvulin family (Pin1 and Par14). Among these isomerases, Pin1 is a unique prolyl isomerase binding to the motif including pSer/pThr-Pro that is phosphorylated by kinases. Once bound, Pin1 modulates the enzymatic activity, protein stability or subcellular localization of target proteins by changing the cis- and trans-formations of proline. Several studies have examined the roles of Pin1 in the pathogenesis of cancers and Alzheimer’s disease. On the other hand, recent studies have newly demonstrated Pin1 to be involved in regulating glucose and lipid metabolism. Interestingly, while Pin1 expression is markedly increased by high-fat diet feeding, Pin1 KO mice are resistant to diet-induced obesity, non-alcoholic steatohepatitis and diabetic vascular dysfunction. These phenomena result from the binding of Pin1 to several key factors regulating metabolic functions, which include insulin receptor substrate-1, AMPK, Crtc2 and NF-κB p65. In this review, we focus on recent advances in elucidating the physiological roles of Pin1 as well as the pathogenesis of disorders involving this isomerase, from the viewpoint of the relationships between signal transductions and metabolic functions. PMID:27618008

  9. Physiological and Pathogenic Roles of Prolyl Isomerase Pin1 in Metabolic Regulations via Multiple Signal Transduction Pathway Modulations.

    PubMed

    Nakatsu, Yusuke; Matsunaga, Yasuka; Yamamotoya, Takeshi; Ueda, Koji; Inoue, Yuki; Mori, Keiichi; Sakoda, Hideyuki; Fujishiro, Midori; Ono, Hiraku; Kushiyama, Akifumi; Asano, Tomoichiro

    2016-09-07

    Prolyl isomerases are divided into three groups, the FKBP family, Cyclophilin and the Parvulin family (Pin1 and Par14). Among these isomerases, Pin1 is a unique prolyl isomerase binding to the motif including pSer/pThr-Pro that is phosphorylated by kinases. Once bound, Pin1 modulates the enzymatic activity, protein stability or subcellular localization of target proteins by changing the cis- and trans-formations of proline. Several studies have examined the roles of Pin1 in the pathogenesis of cancers and Alzheimer's disease. On the other hand, recent studies have newly demonstrated Pin1 to be involved in regulating glucose and lipid metabolism. Interestingly, while Pin1 expression is markedly increased by high-fat diet feeding, Pin1 KO mice are resistant to diet-induced obesity, non-alcoholic steatohepatitis and diabetic vascular dysfunction. These phenomena result from the binding of Pin1 to several key factors regulating metabolic functions, which include insulin receptor substrate-1, AMPK, Crtc2 and NF-κB p65. In this review, we focus on recent advances in elucidating the physiological roles of Pin1 as well as the pathogenesis of disorders involving this isomerase, from the viewpoint of the relationships between signal transductions and metabolic functions.

  10. Physiological and Pathogenic Roles of Prolyl Isomerase Pin1 in Metabolic Regulations via Multiple Signal Transduction Pathway Modulations.

    PubMed

    Nakatsu, Yusuke; Matsunaga, Yasuka; Yamamotoya, Takeshi; Ueda, Koji; Inoue, Yuki; Mori, Keiichi; Sakoda, Hideyuki; Fujishiro, Midori; Ono, Hiraku; Kushiyama, Akifumi; Asano, Tomoichiro

    2016-01-01

    Prolyl isomerases are divided into three groups, the FKBP family, Cyclophilin and the Parvulin family (Pin1 and Par14). Among these isomerases, Pin1 is a unique prolyl isomerase binding to the motif including pSer/pThr-Pro that is phosphorylated by kinases. Once bound, Pin1 modulates the enzymatic activity, protein stability or subcellular localization of target proteins by changing the cis- and trans-formations of proline. Several studies have examined the roles of Pin1 in the pathogenesis of cancers and Alzheimer's disease. On the other hand, recent studies have newly demonstrated Pin1 to be involved in regulating glucose and lipid metabolism. Interestingly, while Pin1 expression is markedly increased by high-fat diet feeding, Pin1 KO mice are resistant to diet-induced obesity, non-alcoholic steatohepatitis and diabetic vascular dysfunction. These phenomena result from the binding of Pin1 to several key factors regulating metabolic functions, which include insulin receptor substrate-1, AMPK, Crtc2 and NF-κB p65. In this review, we focus on recent advances in elucidating the physiological roles of Pin1 as well as the pathogenesis of disorders involving this isomerase, from the viewpoint of the relationships between signal transductions and metabolic functions. PMID:27618008

  11. Leucine and Protein Metabolism in Obese Zucker Rats

    PubMed Central

    She, Pengxiang; Olson, Kristine C.; Kadota, Yoshihiro; Inukai, Ayami; Shimomura, Yoshiharu; Hoppel, Charles L.; Adams, Sean H.; Kawamata, Yasuko; Matsumoto, Hideki; Sakai, Ryosei; Lang, Charles H.; Lynch, Christopher J.

    2013-01-01

    Branched-chain amino acids (BCAAs) are circulating nutrient signals for protein accretion, however, they increase in obesity and elevations appear to be prognostic of diabetes. To understand the mechanisms whereby obesity affects BCAAs and protein metabolism, we employed metabolomics and measured rates of [1-14C]-leucine metabolism, tissue-specific protein synthesis and branched-chain keto-acid (BCKA) dehydrogenase complex (BCKDC) activities. Male obese Zucker rats (11-weeks old) had increased body weight (BW, 53%), liver (107%) and fat (∼300%), but lower plantaris and gastrocnemius masses (−21–24%). Plasma BCAAs and BCKAs were elevated 45–69% and ∼100%, respectively, in obese rats. Processes facilitating these rises appeared to include increased dietary intake (23%), leucine (Leu) turnover and proteolysis [35% per g fat free mass (FFM), urinary markers of proteolysis: 3-methylhistidine (183%) and 4-hydroxyproline (766%)] and decreased BCKDC per g kidney, heart, gastrocnemius and liver (−47–66%). A process disposing of circulating BCAAs, protein synthesis, was increased 23–29% by obesity in whole-body (FFM corrected), gastrocnemius and liver. Despite the observed decreases in BCKDC activities per gm tissue, rates of whole-body Leu oxidation in obese rats were 22% and 59% higher normalized to BW and FFM, respectively. Consistently, urinary concentrations of eight BCAA catabolism-derived acylcarnitines were also elevated. The unexpected increase in BCAA oxidation may be due to a substrate effect in liver. Supporting this idea, BCKAs were elevated more in liver (193–418%) than plasma or muscle, and per g losses of hepatic BCKDC activities were completely offset by increased liver mass, in contrast to other tissues. In summary, our results indicate that plasma BCKAs may represent a more sensitive metabolic signature for obesity than BCAAs. Processes supporting elevated BCAA]BCKAs in the obese Zucker rat include increased dietary intake, Leu and

  12. Molecular tweezers modulate 14-3-3 protein-protein interactions.

    PubMed

    Bier, David; Rose, Rolf; Bravo-Rodriguez, Kenny; Bartel, Maria; Ramirez-Anguita, Juan Manuel; Dutt, Som; Wilch, Constanze; Klärner, Frank-Gerrit; Sanchez-Garcia, Elsa; Schrader, Thomas; Ottmann, Christian

    2013-03-01

    Supramolecular chemistry has recently emerged as a promising way to modulate protein functions, but devising molecules that will interact with a protein in the desired manner is difficult as many competing interactions exist in a biological environment (with solvents, salts or different sites for the target biomolecule). We now show that lysine-specific molecular tweezers bind to a 14-3-3 adapter protein and modulate its interaction with partner proteins. The tweezers inhibit binding between the 14-3-3 protein and two partner proteins--a phosphorylated (C-Raf) protein and an unphosphorylated one (ExoS)--in a concentration-dependent manner. Protein crystallography shows that this effect arises from the binding of the tweezers to a single surface-exposed lysine (Lys214) of the 14-3-3 protein in the proximity of its central channel, which normally binds the partner proteins. A combination of structural analysis and computer simulations provides rules for the tweezers' binding preferences, thus allowing us to predict their influence on this type of protein-protein interactions. PMID:23422566

  13. Interleukin-6 receptor gene polymorphism modulates interleukin-6 levels and the metabolic syndrome: GBCS-CVD.

    PubMed

    Jiang, Chao Qiang; Lam, Tai Hing; Liu, Bin; Lin, Jie Ming; Yue, Xiao Jun; Jin, Ya Li; Cheung, Bernard M Y; Thomas, G Neil

    2010-10-01

    Interleukin-6 (IL-6) is a key pleiotropic cytokine that modulates the inflammatory response. Single-nucleotide polymorphisms (SNPs) within associated genes may contribute to the metabolic syndrome (MES). We examined the role of the IL-6 (rs1524107-C/T) and IL-6 receptor (IL-6R, rs8192284-A/C, Asp358Ala) SNPs in modulating IL-6 levels and the syndrome. A total of 1,979 older Chinese subjects aged 50-92 years from Guangzhou Biobank Cohort Study (GBCS) were recruited. SNPs were detected using Taqman SNP genotyping kits. IL-6 was measured using enzyme-linked immunosorbent assay. The genotype frequencies were 4.9, 33.9, and 61.3% for the IL-6 CC, CT, and TT, and 12.0, 44.9, and 43.1% for the IL-6R CC, AC, and AA, respectively. Both SNPs were in Hardy-Weinberg equilibrium. The IL-6 SNP was not associated with IL-6 levels or the MES, but was dose-dependently associated with fibrinogen levels, P = 0.049. IL-6 levels significantly decreased with increasing proportions of the IL-6R A-allele 9.8 ± 4.9, 9.3 ± 4.8, and 8.4 ± 4.3, respectively, P = 0.001. Conversely, the A-allele was associated with elevated triglyceride, P = 0.009, C-reactive protein, P = 0.047, and potentially with fasting glucose levels, P = 0.077. There was an increasing prevalence of the MES in those carrying the IL-6R CC, AC, and AA genotypes at 18.1, 21.5, 25.2%, respectively, P = 0.010. The SNP was a significant independent predictor of the MES after adjusting for general obesity, age, gender and lifestyle, and socioeconomic parameters, P = 0.023. These data, which are in accord with studies from white populations suggest the IL-6R SNP may play a role in the pathogenesis of the MES possibly through modulating IL-6 levels.

  14. Protein dynamics modulated electron transfer kinetics in early stage photosynthesis

    NASA Astrophysics Data System (ADS)

    Kundu, Prasanta; Dua, Arti

    2013-01-01

    A recent experiment has probed the electron transfer kinetics in the early stage of photosynthesis in Rhodobacter sphaeroides for the reaction center of wild type and different mutants [Science 316, 747 (2007)]. By monitoring the changes in the transient absorption of the donor-acceptor pair at 280 and 930 nm, both of which show non-exponential temporal decay, the experiment has provided a strong evidence that the initial electron transfer kinetics is modulated by the dynamics of protein backbone. In this work, we present a model where the electron transfer kinetics of the donor-acceptor pair is described along the reaction coordinate associated with the distance fluctuations in a protein backbone. The stochastic evolution of the reaction coordinate is described in terms of a non-Markovian generalized Langevin equation with a memory kernel and Gaussian colored noise, both of which are completely described in terms of the microscopics of the protein normal modes. This model provides excellent fits to the transient absorption signals at 280 and 930 nm associated with protein distance fluctuations and protein dynamics modulated electron transfer reaction, respectively. In contrast to previous models, the present work explains the microscopic origins of the non-exponential decay of the transient absorption curve at 280 nm in terms of multiple time scales of relaxation of the protein normal modes. Dynamic disorder in the reaction pathway due to protein conformational fluctuations which occur on time scales slower than or comparable to the electron transfer kinetics explains the microscopic origin of the non-exponential nature of the transient absorption decay at 930 nm. The theoretical estimates for the relative driving force for five different mutants are in close agreement with the experimental estimates obtained using electrochemical measurements.

  15. Ecology drives the distribution of specialized tyrosine metabolism modules in fungi.

    PubMed

    Greene, George H; McGary, Kriston L; Rokas, Antonis; Slot, Jason C

    2014-01-01

    Gene clusters encoding accessory or environmentally specialized metabolic pathways likely play a significant role in the evolution of fungal genomes. Two such gene clusters encoding enzymes associated with the tyrosine metabolism pathway (KEGG #00350) have been identified in the filamentous fungus Aspergillus fumigatus. The l-tyrosine degradation (TD) gene cluster encodes a functional module that facilitates breakdown of the phenolic amino acid, l-tyrosine through a homogentisate intermediate, but is also involved in the production of pyomelanin, a fungal pathogenicity factor. The gentisate catabolism (GC) gene cluster encodes a functional module likely involved in phenolic compound degradation, which may enable metabolism of biphenolic stilbenes in multiple lineages. Our investigation of the evolution of the TD and GC gene clusters in 214 fungal genomes revealed spotty distributions partially shaped by gene cluster loss and horizontal gene transfer (HGT). Specifically, a TD gene cluster shows evidence of HGT between the extremophilic, melanized fungi Exophiala dermatitidis and Baudoinia compniacensis, and a GC gene cluster shows evidence of HGT between Sordariomycete and Dothideomycete grass pathogens. These results suggest that the distribution of specialized tyrosine metabolism modules is influenced by both the ecology and phylogeny of fungal species. PMID:24391152

  16. Ecology Drives the Distribution of Specialized Tyrosine Metabolism Modules in Fungi

    PubMed Central

    Greene, George H.; McGary, Kriston L.; Rokas, Antonis; Slot, Jason C.

    2014-01-01

    Gene clusters encoding accessory or environmentally specialized metabolic pathways likely play a significant role in the evolution of fungal genomes. Two such gene clusters encoding enzymes associated with the tyrosine metabolism pathway (KEGG #00350) have been identified in the filamentous fungus Aspergillus fumigatus. The l-tyrosine degradation (TD) gene cluster encodes a functional module that facilitates breakdown of the phenolic amino acid, l-tyrosine through a homogentisate intermediate, but is also involved in the production of pyomelanin, a fungal pathogenicity factor. The gentisate catabolism (GC) gene cluster encodes a functional module likely involved in phenolic compound degradation, which may enable metabolism of biphenolic stilbenes in multiple lineages. Our investigation of the evolution of the TD and GC gene clusters in 214 fungal genomes revealed spotty distributions partially shaped by gene cluster loss and horizontal gene transfer (HGT). Specifically, a TD gene cluster shows evidence of HGT between the extremophilic, melanized fungi Exophiala dermatitidis and Baudoinia compniacensis, and a GC gene cluster shows evidence of HGT between Sordariomycete and Dothideomycete grass pathogens. These results suggest that the distribution of specialized tyrosine metabolism modules is influenced by both the ecology and phylogeny of fungal species. PMID:24391152

  17. Modulators of Hepatic Lipoprotein Metabolism Identified in a Search for Small-Molecule Inducers of Tribbles Pseudokinase 1 Expression

    PubMed Central

    Nagiec, Marek M.; Skepner, Adam P.; Negri, Joseph; Eichhorn, Michelle; Kuperwasser, Nicolas; Comer, Eamon; Muncipinto, Giovanni; Subramanian, Aravind; Clish, Clary; Musunuru, Kiran; Duvall, Jeremy R.; Foley, Michael; Perez, Jose R.; Palmer, Michelle A. J.

    2015-01-01

    Recent genome wide association studies have linked tribbles pseudokinase 1 (TRIB1) to the risk of coronary artery disease (CAD). Based on the observations that increased expression of TRIB1 reduces secretion of VLDL and is associated with lower plasma levels of LDL cholesterol and triglycerides, higher plasma levels of HDL cholesterol and reduced risk for myocardial infarction, we carried out a high throughput phenotypic screen based on quantitative RT-PCR assay to identify compounds that induce TRIB1 expression in human HepG2 hepatoma cells. In a screen of a collection of diversity-oriented synthesis (DOS)-derived compounds, we identified a series of benzofuran-based compounds that upregulate TRIB1 expression and phenocopy the effects of TRIB1 cDNA overexpression, as they inhibit triglyceride synthesis and apoB secretion in cells. In addition, the compounds downregulate expression of MTTP and APOC3, key components of the lipoprotein assembly pathway. However, CRISPR-Cas9 induced chromosomal disruption of the TRIB1 locus in HepG2 cells, while confirming its regulatory role in lipoprotein metabolism, demonstrated that the effects of benzofurans persist in TRIB1-null cells indicating that TRIB1 is sufficient but not necessary to transmit the effects of the drug. Remarkably, active benzofurans, as well as natural products capable of TRIB1 upregulation, also modulate hepatic cell cholesterol metabolism by elevating the expression of LDLR transcript and LDL receptor protein, while reducing the levels of PCSK9 transcript and secreted PCSK9 protein and stimulating LDL uptake. The effects of benzofurans are not masked by cholesterol depletion and are independent of the SREBP-2 regulatory circuit, indicating that these compounds represent a novel class of chemically tractable small-molecule modulators that shift cellular lipoprotein metabolism in HepG2 cells from lipogenesis to scavenging. PMID:25811180

  18. Exosome engineering for efficient intracellular delivery of soluble proteins using optically reversible protein-protein interaction module.

    PubMed

    Yim, Nambin; Ryu, Seung-Wook; Choi, Kyungsun; Lee, Kwang Ryeol; Lee, Seunghee; Choi, Hojun; Kim, Jeongjin; Shaker, Mohammed R; Sun, Woong; Park, Ji-Ho; Kim, Daesoo; Heo, Won Do; Choi, Chulhee

    2016-01-01

    Nanoparticle-mediated delivery of functional macromolecules is a promising method for treating a variety of human diseases. Among nanoparticles, cell-derived exosomes have recently been highlighted as a new therapeutic strategy for the in vivo delivery of nucleotides and chemical drugs. Here we describe a new tool for intracellular delivery of target proteins, named 'exosomes for protein loading via optically reversible protein-protein interactions' (EXPLORs). By integrating a reversible protein-protein interaction module controlled by blue light with the endogenous process of exosome biogenesis, we are able to successfully load cargo proteins into newly generated exosomes. Treatment with protein-loaded EXPLORs is shown to significantly increase intracellular levels of cargo proteins and their function in recipient cells in vitro and in vivo. These results clearly indicate the potential of EXPLORs as a mechanism for the efficient intracellular transfer of protein-based therapeutics into recipient cells and tissues. PMID:27447450

  19. Salacia oblonga root improves cardiac lipid metabolism in Zucker diabetic fatty rats: modulation of cardiac PPAR-alpha-mediated transcription of fatty acid metabolic genes.

    PubMed

    Huang, Tom Hsun-Wei; Yang, Qinglin; Harada, Masaki; Uberai, Jasna; Radford, Jane; Li, George Q; Yamahara, Johji; Roufogalis, Basil D; Li, Yuhao

    2006-01-01

    Excess cardiac triglyceride accumulation in diabetes and obesity induces lipotoxicity, which predisposes the myocytes to death. On the other hand, increased cardiac fatty acid (FA) oxidation plays a role in the development of myocardial dysfunction in diabetes. PPAR-alpha plays an important role in maintaining homeostasis of lipid metabolism. We have previously demonstrated that the extract from Salacia oblonga root (SOE), an Ayurvedic anti-diabetic and anti-obesity medicine, improves hyperlipidemia in Zucker diabetic fatty (ZDF) rats (a genetic model of type 2 diabetes and obesity) and possesses PPAR-alpha activating properties. Here we demonstrate that chronic oral administration of SOE reduces cardiac triglyceride and FA contents and decreases the Oil red O-stained area in the myocardium of ZDF rats, which parallels the effects on plasma triglyceride and FA levels. Furthermore, the treatment suppressed cardiac overexpression of both FA transporter protein-1 mRNA and protein in ZDF rats, suggesting inhibition of increased cardiac FA uptake as the basis for decreased cardiac FA levels. Additionally, the treatment also inhibited overexpression in ZDF rat heart of PPAR-alpha mRNA and protein and carnitine palmitoyltransferase-1, acyl-CoA oxidase and 5'-AMP-activated protein kinase mRNAs and restored the downregulated acetyl-CoA carboxylase mRNA. These results suggest that SOE inhibits cardiac FA oxidation in ZDF rats. Thus, our findings suggest that improvement by SOE of excess cardiac lipid accumulation and increased cardiac FA oxidation in diabetes and obesity occurs by reduction of cardiac FA uptake, thereby modulating cardiac PPAR-alpha-mediated FA metabolic gene transcription. PMID:16129467

  20. Salacia oblonga root improves cardiac lipid metabolism in Zucker diabetic fatty rats: Modulation of cardiac PPAR-{alpha}-mediated transcription of fatty acid metabolic genes

    SciTech Connect

    Huang, Tom H.-W.; Yang Qinglin; Harada, Masaki; Uberai, Jasna; Radford, Jane; Li, George Q.; Yamahara, Johji; Roufogalis, Basil D.; Li Yuhao . E-mail: yuhao@pharm.usyd.edu.au

    2006-01-15

    Excess cardiac triglyceride accumulation in diabetes and obesity induces lipotoxicity, which predisposes the myocytes to death. On the other hand, increased cardiac fatty acid (FA) oxidation plays a role in the development of myocardial dysfunction in diabetes. PPAR-{alpha} plays an important role in maintaining homeostasis of lipid metabolism. We have previously demonstrated that the extract from Salacia oblonga root (SOE), an Ayurvedic anti-diabetic and anti-obesity medicine, improves hyperlipidemia in Zucker diabetic fatty (ZDF) rats (a genetic model of type 2 diabetes and obesity) and possesses PPAR-{alpha} activating properties. Here we demonstrate that chronic oral administration of SOE reduces cardiac triglyceride and FA contents and decreases the Oil red O-stained area in the myocardium of ZDF rats, which parallels the effects on plasma triglyceride and FA levels. Furthermore, the treatment suppressed cardiac overexpression of both FA transporter protein-1 mRNA and protein in ZDF rats, suggesting inhibition of increased cardiac FA uptake as the basis for decreased cardiac FA levels. Additionally, the treatment also inhibited overexpression in ZDF rat heart of PPAR-{alpha} mRNA and protein and carnitine palmitoyltransferase-1, acyl-CoA oxidase and 5'-AMP-activated protein kinase mRNAs and restored the downregulated acetyl-CoA carboxylase mRNA. These results suggest that SOE inhibits cardiac FA oxidation in ZDF rats. Thus, our findings suggest that improvement by SOE of excess cardiac lipid accumulation and increased cardiac FA oxidation in diabetes and obesity occurs by reduction of cardiac FA uptake, thereby modulating cardiac PPAR-{alpha}-mediated FA metabolic gene transcription.

  1. Experimental study on trace chemical contaminant generation rates of human metabolism in spacecraft crew module

    NASA Astrophysics Data System (ADS)

    Lihua, Guo; Xinxing, He; Guoxin, Xu; Xin, Qi

    2012-12-01

    Trace chemical contaminants generated by human metabolism is a major source of contamination in spacecraft crew module. In this research, types and generation rates of pollutants from human metabolism were determined in the Chinese diets. Expired air, skin gas, and sweat of 20 subjects were analyzed at different exercise states in a simulated module. The exercise states were designed according to the basic activities in the orbit of astronauts. Qualitative and quantitative analyses of contaminants generated by human metabolic were performed with gas chromatography/mass spectrometry, gas chromatography and UV spectrophotometer. Sixteen chemical compounds from metabolic sources were found. With the increase in physical load, the concentrations of chemical compounds from human skin and expired air correspondingly increased. The species and the offgassing rates of pollutants from human metabolism are different among the Chinese, Americans and the Russians due to differences in ethnicity and dietary customs. This research provides data to aid in the design, development and operation of China's long duration space mission.

  2. Red wine polyphenols modulate fecal microbiota and reduce markers of the metabolic syndrome in obese patients.

    PubMed

    Moreno-Indias, Isabel; Sánchez-Alcoholado, Lidia; Pérez-Martínez, Pablo; Andrés-Lacueva, Cristina; Cardona, Fernando; Tinahones, Francisco; Queipo-Ortuño, María Isabel

    2016-04-01

    This study evaluated the possible prebiotic effect of a moderate intake of red wine polyphenols on the modulation of the gut microbiota composition and the improvement in the risk factors for the metabolic syndrome in obese patients. Ten metabolic syndrome patients and ten healthy subjects were included in a randomized, crossover, controlled intervention study. After a washout period, the subjects consumed red wine and de-alcoholized red wine over a 30 day period for each. The dominant bacterial composition did not differ significantly between the study groups after the two red wine intake periods. In the metabolic syndrome patients, red wine polyphenols significantly increased the number of fecal bifidobacteria and Lactobacillus (intestinal barrier protectors) and butyrate-producing bacteria (Faecalibacterium prausnitzii and Roseburia) at the expense of less desirable groups of bacteria such as LPS producers (Escherichia coli and Enterobacter cloacae). The changes in gut microbiota in these patients could be responsible for the improvement in the metabolic syndrome markers. Modulation of the gut microbiota by using red wine could be an effective strategy for managing metabolic diseases associated with obesity.

  3. Text mining for metabolic pathways, signaling cascades, and protein networks.

    PubMed

    Hoffmann, Robert; Krallinger, Martin; Andres, Eduardo; Tamames, Javier; Blaschke, Christian; Valencia, Alfonso

    2005-05-10

    The complexity of the information stored in databases and publications on metabolic and signaling pathways, the high throughput of experimental data, and the growing number of publications make it imperative to provide systems to help the researcher navigate through these interrelated information resources. Text-mining methods have started to play a key role in the creation and maintenance of links between the information stored in biological databases and its original sources in the literature. These links will be extremely useful for database updating and curation, especially if a number of technical problems can be solved satisfactorily, including the identification of protein and gene names (entities in general) and the characterization of their types of interactions. The first generation of openly accessible text-mining systems, such as iHOP (Information Hyperlinked over Proteins), provides additional functions to facilitate the reconstruction of protein interaction networks, combine database and text information, and support the scientist in the formulation of novel hypotheses. The next challenge is the generation of comprehensive information regarding the general function of signaling pathways and protein interaction networks. PMID:15886388

  4. Changes in contralateral protein metabolism following unilateral sciatic nerve section

    SciTech Connect

    Menendez, J.A.; Cubas, S.C.

    1990-03-01

    Changes in nerve biochemistry, anatomy, and function following injuries to the contralateral nerve have been repeatedly reported, though their significance is unknown. The most likely mechanisms for their development are either substances carried by axoplasmic flow or electrically transmitted signals. This study analyzes which mechanism underlies the development of a contralateral change in protein metabolism. The incorporation of labelled amino acids (AA) into proteins of both sciatic nerves was assessed by liquid scintillation after an unilateral section. AA were offered locally for 30 min to the distal stump of the sectioned nerves and at homologous levels of the intact contralateral nerves. At various times, from 1 to 24 h, both sciatic nerves were removed and the proteins extracted with trichloroacetic acid (TCA). An increase in incorporation was found in both nerves 14-24 h after section. No difference existed between sectioned and intact nerves, which is consistent with the contralateral effect. Lidocaine, but not colchicine, when applied previously to the nerves midway between the sectioning site and the spinal cord, inhibited the contralateral increase in AA incorporation. It is concluded that electrical signals, crossing through the spinal cord, are responsible for the development of the contralateral effect. Both the nature of the proteins and the significance of the contralateral effect are matters for speculation.

  5. Cell-specific modulation of surfactant proteins by ambroxol treatment

    SciTech Connect

    Seifart, Carola . E-mail: zwiebel@mailer.uni-marburg.de; Clostermann, Ursula; Seifart, Ulf

    2005-02-15

    Ambroxol [trans-4-(2-amino-3,5-dibromobenzylamino)-cyclohexanole hydrochloride], a mucolytic agent, was postulated to provide surfactant stimulatory properties and was previously used to prevent surfactant deficiency. Currently, the underlying mechanisms are not exactly clear. Because surfactant homeostasis is regulated by surfactant-specific proteins (SP), we analyzed protein amount and mRNA expression in whole lung tissue, isolated type II pneumocytes and bronchoalveolar lavage of Sprague-Dawley rats treated with ambroxol i.p. (75 mg/kg body weight, twice a day [every 12 h]). The methods used included competitive polymerase chain reaction (RT-PCR), Northern blotting, Western immunoblotting, and immunohistochemistry. In isolated type II pneumocytes of ambroxol-treated animals, SP-C protein and mRNA content were increased, whereas SP-A, -B and -D protein, mRNA, and immunoreactivity remained unaffected. However, ambroxol treatment resulted in a significant increase of SP-B and in a decrease of SP-D in whole lung tissue with enhanced immunostaining for SP-B in Clara Cells. SP-A and SP-D were significantly decreased in BAL fluid of ambroxol-treated animals. The data suggest that surfactant protein expression is modulated in a cell-specific manner by ambroxol, as type II pneumocytes exhibited an increase in SP-C, whereas Clara cells exhibited an increase in the immunoreactivity for SP-B accounting for the increased SP-B content of whole lung tissue. The results indicate that ambroxol may exert its positive effects, observed in the treatment of diseases related to surfactant deficiency, via modulation of surfactant protein expression.

  6. Cell-specific modulation of surfactant proteins by ambroxol treatment.

    PubMed

    Seifart, Carola; Clostermann, Ursula; Seifart, Ulf; Müller, Bernd; Vogelmeier, Claus; von Wichert, Peter; Fehrenbach, Heinz

    2005-02-15

    Ambroxol [trans-4-(2-amino-3,5-dibromobenzylamino)-cyclohexanole hydrochloride], a mucolytic agent, was postulated to provide surfactant stimulatory properties and was previously used to prevent surfactant deficiency. Currently, the underlying mechanisms are not exactly clear. Because surfactant homeostasis is regulated by surfactant-specific proteins (SP), we analyzed protein amount and mRNA expression in whole lung tissue, isolated type II pneumocytes and bronchoalveolar lavage of Sprague-Dawley rats treated with ambroxol i.p. (75 mg/kg body weight, twice a day [every 12 h]). The methods used included competitive polymerase chain reaction (RT-PCR), Northern blotting, Western immunoblotting, and immunohistochemistry. In isolated type II pneumocytes of ambroxol-treated animals, SP-C protein and mRNA content were increased, whereas SP-A, -B and -D protein, mRNA, and immunoreactivity remained unaffected. However, ambroxol treatment resulted in a significant increase of SP-B and in a decrease of SP-D in whole lung tissue with enhanced immunostaining for SP-B in Clara Cells. SP-A and SP-D were significantly decreased in BAL fluid of ambroxol-treated animals. The data suggest that surfactant protein expression is modulated in a cell-specific manner by ambroxol, as type II pneumocytes exhibited an increase in SP-C, whereas Clara cells exhibited an increase in the immunoreactivity for SP-B accounting for the increased SP-B content of whole lung tissue. The results indicate that ambroxol may exert its positive effects, observed in the treatment of diseases related to surfactant deficiency, via modulation of surfactant protein expression.

  7. Modulation of arachidonic acid metabolism by Rous sarcoma virus

    SciTech Connect

    Barker, K.; Aderem, A.; Hanafusa, H. )

    1989-07-01

    Arachidonic acid (C{sub 20:4}) metabolites were released constitutively from wild-type Rous sarcoma virus-transformed chicken embryo fibroblasts (CEF). {sup 3}H-labeled C{sub 20:4} and its metabolites were released from unstimulated and uninfected CEF only in response to stimuli such as serum, phorbol ester, or the calcium ionophore A23187. High-pressure liquid chromatography analysis showed that the radioactivity released from ({sup 3}H)arachidonate-labeled transformed cells was contained in free arachidonate and in the cyclooxygenase products prostaglandin E{sub 2} and prostaglandin F{sub 2} alpha; no lipoxygenase products were identified. The release of C{sub 20:4} and its metabolites from CEF infected with pp60{sup src} deletion mutants was correlated with serum-independent DNA synthesis and with the expression of the mRNA for 9E3, a gene expressed in Rous sarcoma virus-transformed cells which has homology with several mitogenic and inflammatory peptides. {sup 3}H-labeled C{sub 20:4} release was not correlated with p36 phosphorylation, which argues against a role for this protein as a phospholipase A{sub 2} inhibitor. CEF infected with other oncogenic viruses encoding a tyrosine kinase also released C{sub 20:4}, as did CEF infected with viruses that contained mos and ras; however, infection with a crk-containing virus did not result in stimulation of {sup 3}H-labeled C{sub 20:4} release, suggesting that utilization of this signaling pathway is specific for particular transformation stimuli.

  8. Acute responses of muscle protein metabolism to reduced blood flow reflect metabolic priorities for homeostasis.

    PubMed

    Zhang, Xiao-Jun; Irtun, Oivind; Chinkes, David L; Wolfe, Robert R

    2008-03-01

    The present experiment was designed to measure the synthetic and breakdown rates of muscle protein in the hindlimb of rabbits with or without clamping the femoral artery. l-[ring-(13)C(6)]phenylalanine was infused as a tracer for measurement of muscle protein kinetics by means of an arteriovenous model, tracer incorporation, and tracee release methods. The ultrasonic flowmeter, dye dilution, and microsphere methods were used to determine the flow rates in the femoral artery, in the leg, and in muscle capillary, respectively. The femoral artery flow accounted for 65% of leg flow. A 50% reduction in the femoral artery flow reduced leg flow by 28% and nutritive flow by 26%, which did not change protein synthetic or breakdown rate in leg muscle. Full clamp of the femoral artery reduced leg flow by 42% and nutritive flow by 59%, which decreased (P < 0.05) both the fractional synthetic rate from 0.19 +/- 0.05 to 0.14 +/- 0.03%/day and fractional breakdown rate from 0.28 +/- 0.07 to 0.23 +/- 0.09%/day of muscle protein. Neither the partial nor full clamp reduced (P = 0.27-0.39) the intracellular phenylalanine concentration or net protein balance in leg muscle. We conclude that the flow threshold to cause a fall of protein turnover rate in leg muscle was a reduction of 30-40% of the leg flow. The acute responses of muscle protein kinetics to the reductions in blood flow reflected the metabolic priorities to maintain muscle homeostasis. These findings cannot be extrapolated to more chronic conditions without experimental validation. PMID:18089763

  9. Modulator of Apoptosis 1 (MOAP-1) Is a Tumor Suppressor Protein Linked to the RASSF1A Protein*

    PubMed Central

    Law, Jennifer; Salla, Mohamed; Zare, Alaa; Wong, Yoke; Luong, Le; Volodko, Natalia; Svystun, Orysya; Flood, Kayla; Lim, Jonathan; Sung, Miranda; Dyck, Jason R. B.; Tan, Chong Teik; Su, Yu-Chin; Yu, Victor C.; Mackey, John; Baksh, Shairaz

    2015-01-01

    Modulator of apoptosis 1 (MOAP-1) is a BH3-like protein that plays key roles in cell death or apoptosis. It is an integral partner to the tumor suppressor protein, Ras association domain family 1A (RASSF1A), and functions to activate the Bcl-2 family pro-apoptotic protein Bax. Although RASSF1A is now considered a bona fide tumor suppressor protein, the role of MOAP-1 as a tumor suppressor protein has yet to be determined. In this study, we present several lines of evidence from cancer databases, immunoblotting of cancer cells, proliferation, and xenograft assays as well as DNA microarray analysis to demonstrate the role of MOAP-1 as a tumor suppressor protein. Frequent loss of MOAP-1 expression, in at least some cancers, appears to be attributed to mRNA down-regulation and the rapid proteasomal degradation of MOAP-1 that could be reversed utilizing the proteasome inhibitor MG132. Overexpression of MOAP-1 in several cancer cell lines resulted in reduced tumorigenesis and up-regulation of genes involved in cancer regulatory pathways that include apoptosis (p53, Fas, and MST1), DNA damage control (poly(ADP)-ribose polymerase and ataxia telangiectasia mutated), those within the cell metabolism (IR-α, IR-β, and AMP-activated protein kinase), and a stabilizing effect on microtubules. The loss of RASSF1A (an upstream regulator of MOAP-1) is one of the earliest detectable epigenetically silenced tumor suppressor proteins in cancer, and we speculate that the additional loss of function of MOAP-1 may be a second hit to functionally compromise the RASSF1A/MOAP-1 death receptor-dependent pathway and drive tumorigenesis. PMID:26269600

  10. Functional modulation of G-protein coupled receptors during Parkinson disease-like neurodegeneration.

    PubMed

    Jenkins, Bruce G; Zhu, Aijun; Poutiainen, Pekka; Choi, Ji-Kyung; Kil, Kun-Eek; Zhang, Zhaoda; Kuruppu, Darshini; Aytan, Nurgul; Dedeoglu, Alpaslan; Brownell, Anna-Liisa

    2016-09-01

    G-protein coupled dopamine and metabotropic glutamate receptors (mGlu) can modulate neurotransmission during Parkinson's disease (PD)-like neurodegeneration. PET imaging studies in a unilateral dopamine denervation model (6-OHDA) showed a significant inverse correlation of presynaptic mGlu4 and postsynaptic mGlu5 expression in the striatum and rapidly declining mGlu4 and enhanced mGlu5 expression in the hippocampus during progressive degeneration over time. Immunohistochemical studies verified the decreased mGlu4 expression in the hippocampus on the lesion side but did not show difference in mGlu5 expression between lesion and control side. Pharmacological MRI studies showed enhanced hemodynamic response in several brain areas on the lesion side compared to the control side after challenge with mGlu4 positive allosteric modulator or mGlu5 negative allosteric modulator. However, mGlu4 response was biphasic having short enhancement followed by negative response on both sides of brain. Studies in mGlu4 expressing cells demonstrated that glutamate induces cooperative increase in binding of mGlu4 ligands - especially at high glutamate levels consistent with in vivo concentration. This suggests that mGlu allosteric modulators as drug candidates will be highly sensitive to changes in glutamate concentration and hence metabolic state. These experiments demonstrate the importance of the longitudinal imaging studies to investigate temporal changes in receptor functions to obtain individual response for experimental drugs. PMID:26581500

  11. Non-Genomic Origins of Proteins and Metabolism

    NASA Technical Reports Server (NTRS)

    Pohorille, Andrew

    2003-01-01

    It is proposed that evolution of inanimate matter to cells endowed with a nucleic acid- based coding of genetic information was preceded by an evolutionary phase, in which peptides not coded by nucleic acids were able to self-organize into networks capable of evolution towards increasing metabolic complexity. Recent findings that truly different, simple peptides (Keefe and Szostak, 2001) can perform the same function (such as ATP binding) provide experimental support for this mechanism of early protobiological evolution. The central concept underlying this mechanism is that the reproduction of cellular functions alone was sufficient for self-maintenance of protocells, and that self- replication of macromolecules was not required at this stage of evolution. The precise transfer of information between successive generations of the earliest protocells was unnecessary and, possibly, undesirable. The key requirement in the initial stage of protocellular evolution was an ability to rapidly explore a large number of protein sequences in order to discover a set of molecules capable of supporting self- maintenance and growth of protocells. Undoubtedly, the essential protocellular functions were carried out by molecules not nearly as efficient or as specific as contemporary proteins. Many, potentially unrelated sequences could have performed each of these functions at an evolutionarily acceptable level. As evolution progressed, however proteins must have performed their functions with increasing efficiency and specificity. This, in turn, put additional constraints on protein sequences and the fraction of proteins capable of performing their functions at the required level decreased. At some point, the likelihood of generating a sufficiently efficient set of proteins through a non-coded synthesis was so small that further evolution was not possible without storing information about the sequences of these proteins. Beyond this point, further evolution required coupling between

  12. Glucocorticoid Modulation of Mitochondrial Function in Hepatoma Cells Requires the Mitochondrial Fission Protein Drp1

    PubMed Central

    Hernández-Alvarez, María Isabel; Paz, José C.; Sebastián, David; Muñoz, Juan Pablo; Liesa, Marc; Segalés, Jessica; Palacín, Manuel

    2013-01-01

    Abstract Aims: Glucocorticoids, such as dexamethasone, enhance hepatic energy metabolism and gluconeogenesis partly through changes in mitochondrial function. Mitochondrial function is influenced by the balance between mitochondrial fusion and fission events. However, whether glucocorticoids modulate mitochondrial function through the regulation of mitochondrial dynamics is currently unknown. Results: Here, we report that the effects of dexamethasone on mitochondrial function and gluconeogenesis in hepatoma cells are dependent on the mitochondrial fission protein dynamin-related protein 1 (Drp1). Dexamethasone increased routine oxygen consumption, maximal respiratory capacity, superoxide anion, proton leak, and gluconeogenesis in hepatoma cells. Under these conditions, dexamethasone altered mitochondrial morphology, which was paralleled by a large increase in Drp1 expression, and reduced mitofusin 1 (Mfn1) and Mfn2. In vivo dexamethasone treatment also enhanced Drp1 expression in mouse liver. On the basis of these observations, we analyzed the dependence on the Drp1 function of dexamethasone effects on mitochondrial respiration and gluconeogenesis. We show that the increase in mitochondrial respiration and gluconeogenesis induced by dexamethasone are hampered by the inhibition of Drp1 function. Innovation: Our findings provide the first evidence that the effects of glucocorticoids on hepatic metabolism require the mitochondrial fission protein Drp1. Conclusion: In summary, we demonstrate that the mitochondrial effects of dexamethasone both on mitochondrial respiration and on the gluconeogenic pathway depend on Drp1. Antioxid. Redox Signal. 19, 366–378. PMID:22703557

  13. Integrated pathway modules using time-course metabolic profiles and EST data from Milnesium tardigradum

    PubMed Central

    2012-01-01

    Background Tardigrades are multicellular organisms, resistant to extreme environmental changes such as heat, drought, radiation and freezing. They outlast these conditions in an inactive form (tun) to escape damage to cellular structures and cell death. Tardigrades are apparently able to prevent or repair such damage and are therefore a crucial model organism for stress tolerance. Cultures of the tardigrade Milnesium tardigradum were dehydrated by removing the surrounding water to induce tun formation. During this process and the subsequent rehydration, metabolites were measured in a time series by GC-MS. Additionally expressed sequence tags are available, especially libraries generated from the active and inactive state. The aim of this integrated analysis is to trace changes in tardigrade metabolism and identify pathways responsible for their extreme resistance against physical stress. Results In this study we propose a novel integrative approach for the analysis of metabolic networks to identify modules of joint shifts on the transcriptomic and metabolic levels. We derive a tardigrade-specific metabolic network represented as an undirected graph with 3,658 nodes (metabolites) and 4,378 edges (reactions). Time course metabolite profiles are used to score the network nodes showing a significant change over time. The edges are scored according to information on enzymes from the EST data. Using this combined information, we identify a key subnetwork (functional module) of concerted changes in metabolic pathways, specific for de- and rehydration. The module is enriched in reactions showing significant changes in metabolite levels and enzyme abundance during the transition. It resembles the cessation of a measurable metabolism (e.g. glycolysis and amino acid anabolism) during the tun formation, the production of storage metabolites and bioprotectants, such as DNA stabilizers, and the generation of amino acids and cellular components from monosaccharides as carbon and

  14. Deciphering the biological effects of acupuncture treatment modulating multiple metabolism pathways

    PubMed Central

    Zhang, Aihua; Yan, Guangli; Sun, Hui; Cheng, Weiping; Meng, Xiangcai; Liu, Li; Xie, Ning; Wang, Xijun

    2016-01-01

    Acupuncture is an alternative therapy that is widely used to treat various diseases. However, detailed biological interpretation of the acupuncture stimulations is limited. We here used metabolomics and proteomics technology, thereby identifying the serum small molecular metabolites into the effect and mechanism pathways of standardized acupuncture treatments at ‘Zusanli’ acupoint which was the most often used acupoint in previous reports. Comprehensive overview of serum metabolic profiles during acupuncture stimulation was investigated. Thirty-four differential metabolites were identified in serum metabolome and associated with ten metabolism pathways. Importantly, we have found that high impact glycerophospholipid metabolism, fatty acid metabolism, ether lipid metabolism were acutely perturbed by acupuncture stimulation. As such, these alterations may be useful to clarify the biological mechanism of acupuncture stimulation. A series of differentially expressed proteins were identified and such effects of acupuncture stimulation were found to play a role in transport, enzymatic activity, signaling pathway or receptor interaction. Pathway analysis further revealed that most of these proteins were found to play a pivotal role in the regulation of multiple metabolism pathways. It demonstrated that the metabolomics coupled with proteomics as a powerful approach for potential applications in understanding the biological effects of acupuncture stimulation. PMID:26879284

  15. Yeast product supplementation modulated feeding behavior and metabolism in transition dairy cows.

    PubMed

    Yuan, K; Liang, T; Muckey, M B; Mendonça, L G D; Hulbert, L E; Elrod, C C; Bradford, B J

    2015-01-01

    Yeast supplementation has been shown to increase feed intake and production in some studies with early lactation dairy cows, but the mechanisms underlying such an effect remain unknown. The objective of this study was to assess the effects of supplementing a yeast product derived from Saccharomyces cerevisiae on production, feeding behavior, and metabolism in cows during the transition to lactation. Forty multiparous Holstein cows were blocked by expected calving date and randomly assigned within block to 1 of 4 treatments (n=10) from 21 d before expected calving to 42 d postpartum. Rations were top-dressed with a yeast culture plus enzymatically hydrolyzed yeast (YC-EHY; Celmanax, Vi-COR Inc., Mason City, IA) at the rate of 0, 30, 60, or 90g/d throughout the experiment. Dry matter and water intake, feeding behavior, and milk production were monitored. Plasma samples collected on -21, -7, 1, 4, 7, 14, 21, and 35 d relative to calving were analyzed for glucose, β-hydroxybutyrate, and nonesterified fatty acids. Data were analyzed using mixed models with repeated measures over time. Pre- or postpartum dry matter intake and water intake did not differ among treatments. Quadratic dose effects were observed for prepartum feeding behavior, reflecting decreased meal size, meal length, and intermeal interval, and increased meal frequency for cows received 30 and 60g/d of YC-EHY. Postpartum feeding behavior was unaffected by treatments. Milk yields were not affected (45.3, 42.6, 47.8, and 46.7kg/d for 0, 30, 60, and 90g/d, respectively) by treatments. Tendencies for increased percentages of milk fat, protein, and lactose were detected for cows receiving YC-EHY. Supplementing YC-EHY increased plasma β-hydroxybutyrate and tended to decrease (quadratic dose effect) glucose but did not affect nonesterified fatty acids. Yeast product supplementation during the transition period did not affect milk production and dry matter intake but modulated feeding behavior and metabolism

  16. Modulation of B-cell exosome proteins by gamma herpesvirus infection

    PubMed Central

    Meckes, David G.; Gunawardena, Harsha P.; Dekroon, Robert M.; Heaton, Phillip R.; Edwards, Rachel H.; Ozgur, Sezgin; Griffith, Jack D.; Damania, Blossom; Raab-Traub, Nancy

    2013-01-01

    The human gamma herpesviruses, Kaposi sarcoma-associated virus (KSHV) and EBV, are associated with multiple cancers. Recent evidence suggests that EBV and possibly other viruses can manipulate the tumor microenvironment through the secretion of specific viral and cellular components into exosomes, small endocytically derived vesicles that are released from cells. Exosomes produced by EBV-infected nasopharyngeal carcinoma cells contain high levels of the viral oncogene latent membrane protein 1 and viral microRNAs that activate critical signaling pathways in recipient cells. In this study, to determine the effects of EBV and KSHV on exosome content, quantitative proteomics techniques were performed on exosomes purified from 11 B-cell lines that are uninfected, infected with EBV or with KSHV, or infected with both viruses. Using mass spectrometry, 871 proteins were identified, of which ∼360 were unique to the viral exosomes. Analysis by 2D difference gel electrophoresis and spectral counting identified multiple significant changes compared with the uninfected control cells and between viral groups. These data predict that both EBV and KSHV exosomes likely modulate cell death and survival, ribosome function, protein synthesis, and mammalian target of rapamycin signaling. Distinct viral-specific effects on exosomes suggest that KSHV exosomes would affect cellular metabolism, whereas EBV exosomes would activate cellular signaling mediated through integrins, actin, IFN, and NFκB. The changes in exosome content identified in this study suggest ways that these oncogenic viruses modulate the tumor microenvironment and may provide diagnostic markers specific for EBV and KSHV associated malignancies. PMID:23818640

  17. Periodic and stochastic thermal modulation of protein folding kinetics

    SciTech Connect

    Platkov, Max; Gruebele, Martin

    2014-07-21

    Chemical reactions are usually observed either by relaxation of a bulk sample after applying a sudden external perturbation, or by intrinsic fluctuations of a few molecules. Here we show that the two ideas can be combined to measure protein folding kinetics, either by periodic thermal modulation, or by creating artificial thermal noise that greatly exceeds natural thermal fluctuations. We study the folding reaction of the enzyme phosphoglycerate kinase driven by periodic temperature waveforms. As the temperature waveform unfolds and refolds the protein, its fluorescence color changes due to FRET (Förster resonant Energy Transfer) of two donor/acceptor fluorophores labeling the protein. We adapt a simple model of periodically driven kinetics that nicely fits the data at all temperatures and driving frequencies: The phase shifts of the periodic donor and acceptor fluorescence signals as a function of driving frequency reveal reaction rates. We also drive the reaction with stochastic temperature waveforms that produce thermal fluctuations much greater than natural fluctuations in the bulk. Such artificial thermal noise allows the recovery of weak underlying signals due to protein folding kinetics. This opens up the possibility for future detection of a stochastic resonance for protein folding subject to noise with controllable amplitude.

  18. Periodic and stochastic thermal modulation of protein folding kinetics

    NASA Astrophysics Data System (ADS)

    Platkov, Max; Gruebele, Martin

    2014-07-01

    Chemical reactions are usually observed either by relaxation of a bulk sample after applying a sudden external perturbation, or by intrinsic fluctuations of a few molecules. Here we show that the two ideas can be combined to measure protein folding kinetics, either by periodic thermal modulation, or by creating artificial thermal noise that greatly exceeds natural thermal fluctuations. We study the folding reaction of the enzyme phosphoglycerate kinase driven by periodic temperature waveforms. As the temperature waveform unfolds and refolds the protein, its fluorescence color changes due to FRET (Förster resonant Energy Transfer) of two donor/acceptor fluorophores labeling the protein. We adapt a simple model of periodically driven kinetics that nicely fits the data at all temperatures and driving frequencies: The phase shifts of the periodic donor and acceptor fluorescence signals as a function of driving frequency reveal reaction rates. We also drive the reaction with stochastic temperature waveforms that produce thermal fluctuations much greater than natural fluctuations in the bulk. Such artificial thermal noise allows the recovery of weak underlying signals due to protein folding kinetics. This opens up the possibility for future detection of a stochastic resonance for protein folding subject to noise with controllable amplitude.

  19. Mitochondrial mayhem: the mitochondrion as a modulator of iron metabolism and its role in disease.

    PubMed

    Huang, Michael Li-Hsuan; Lane, Darius J R; Richardson, Des R

    2011-12-15

    The mitochondrion plays vital roles in various aspects of cellular metabolism, ranging from energy transduction and apoptosis to the synthesis of important molecules such as heme. Mitochondria are also centrally involved in iron metabolism, as exemplified by disruptions in mitochondrial proteins that lead to perturbations in whole-cell iron processing. Recent investigations have identified a host of mitochondrial proteins (e.g., mitochondrial ferritin; mitoferrins 1 and 2; ABCBs 6, 7, and 10; and frataxin) that may play roles in the homeostasis of mitochondrial iron. These mitochondrial proteins appear to participate in one or more processes of iron storage, iron uptake, and heme and iron-sulfur cluster synthesis. In this review, we present and critically discuss the evidence suggesting that the mitochondrion may contribute to the regulation of whole-cell iron metabolism. Further, human diseases that arise from a dysregulation of these mitochondrial molecules reveal the ability of the mitochondrion to communicate with cytosolic iron metabolism to coordinate whole-cell iron processing and to fulfill the high demands of this organelle for iron. This review highlights new advances in understanding iron metabolism in terms of novel molecular players and diseases associated with its dysregulation.

  20. A Flavonoid Compound Promotes Neuronal Differentiation of Embryonic Stem Cells via PPAR-β Modulating Mitochondrial Energy Metabolism.

    PubMed

    Mei, Yu-Qin; Pan, Zong-Fu; Chen, Wen-Teng; Xu, Min-Hua; Zhu, Dan-Yan; Yu, Yong-Ping; Lou, Yi-Jia

    2016-01-01

    Relatively little is known regarding mitochondrial metabolism in neuronal differentiation of embryonic stem (ES) cells. By using a small molecule, present research has investigated the pattern of cellular energy metabolism in neural progenitor cells derived from mouse ES cells. Flavonoid compound 4a faithfully facilitated ES cells to differentiate into neurons morphologically and functionally. The expression and localization of peroxisome proliferator-activated receptors (PPARs) were examined in neural progenitor cells. PPAR-β expression showed robust upregulation compared to solvent control. Treatment with PPAR-β agonist L165041 alone or together with compound 4a significantly promoted neuronal differentiation, while antagonist GSK0660 blocked the neurogenesis-promoting effect of compound 4a. Consistently, knockdown of PPAR-β in ES cells abolished compound 4a-induced neuronal differentiation. Interestingly, we found that mitochondrial fusion protein Mfn2 was also abolished by sh-PPAR-β, resulting in abnormal mitochondrial Ca2+ ([Ca2+]M) transients as well as impaired mitochondrial bioenergetics. In conclusion, we demonstrated that by modulating mitochondrial energy metabolism through Mfn2 and mitochondrial Ca2+, PPAR-β took an important role in neuronal differentiation induced by flavonoid compound 4a. PMID:27315062

  1. 27-Hydroxycholesterol contributes to disruptive effects on learning and memory by modulating cholesterol metabolism in the rat brain.

    PubMed

    Zhang, D-D; Yu, H-L; Ma, W-W; Liu, Q-R; Han, J; Wang, H; Xiao, R

    2015-08-01

    Cholesterol metabolism is important for neuronal function in the central nervous system (CNS). The oxysterol 27-hydroxycholesterol (27-OHC) is a cholesterol metabolite that crosses the blood-brain barrier (BBB) and may be a useful substitutive marker for neurodegenerative diseases. However, the effects of 27-OHC on learning and memory and the underlying mechanisms are unclear. To determine this mechanism, we investigated learning and memory and cholesterol metabolism in rat brain following the injection of various doses of 27-OHC into the caudal vein. We found that 27-OHC increased cholesterol levels and upregulated the expression of liver X receptor-α (LXR-α) and adenosine triphosphate (ATP)-binding cassette transporter protein family member A1 (ABCA1). In addition, 27-OHC decreased the expression of 3-hydroxy-3-methylglutaryl-CoA reductase (HMG-CR) and low-density lipoprotein receptor (LDLR) in rat brain tissues. These findings suggest that 27-OHC may negatively modulate cognitive effects and cholesterol metabolism in the brain.

  2. A Flavonoid Compound Promotes Neuronal Differentiation of Embryonic Stem Cells via PPAR-β Modulating Mitochondrial Energy Metabolism

    PubMed Central

    Mei, Yu-qin; Pan, Zong-fu; Chen, Wen-teng; Xu, Min-hua; Zhu, Dan-yan; Yu, Yong-ping; Lou, Yi-jia

    2016-01-01

    Relatively little is known regarding mitochondrial metabolism in neuronal differentiation of embryonic stem (ES) cells. By using a small molecule, present research has investigated the pattern of cellular energy metabolism in neural progenitor cells derived from mouse ES cells. Flavonoid compound 4a faithfully facilitated ES cells to differentiate into neurons morphologically and functionally. The expression and localization of peroxisome proliferator-activated receptors (PPARs) were examined in neural progenitor cells. PPAR-β expression showed robust upregulation compared to solvent control. Treatment with PPAR-β agonist L165041 alone or together with compound 4a significantly promoted neuronal differentiation, while antagonist GSK0660 blocked the neurogenesis-promoting effect of compound 4a. Consistently, knockdown of PPAR-β in ES cells abolished compound 4a-induced neuronal differentiation. Interestingly, we found that mitochondrial fusion protein Mfn2 was also abolished by sh-PPAR-β, resulting in abnormal mitochondrial Ca2+ ([Ca2+]M) transients as well as impaired mitochondrial bioenergetics. In conclusion, we demonstrated that by modulating mitochondrial energy metabolism through Mfn2 and mitochondrial Ca2+, PPAR-β took an important role in neuronal differentiation induced by flavonoid compound 4a. PMID:27315062

  3. Effects of continuous-wave, pulsed, and sinusoidal-amplitude-modulated microwaves on brain energy metabolism.

    PubMed

    Sanders, A P; Joines, W T; Allis, J W

    1985-01-01

    A comparison of the effects of continuous-wave, sinusoidal-amplitude-modulated, and pulsed square-wave-modulated 591-MHz microwave exposures on brain energy metabolism was made in male Sprague-Dawley rats (175-225 g). Brain NADH fluorescence, adenosine triphosphate (ATP) concentration, and creatine phosphate (CP) concentration were determined as a function of modulation frequency. Brain temperatures of animals were maintained between -0.1 and -0.4 degrees C from the preexposure temperature when subjected to as much as 20 mW/cm2 (average power) CW, pulsed, or sinusoidal-amplitude modulated 591-MHz radiation for 5 min. Sinusoidal-amplitude-modulated exposures at 16-24 Hz showed a trend toward preferential modulation frequency response in inducing an increase in brain NADH fluorescence. The pulse-modulated and sinusoidal-amplitude-modulated (16 Hz) microwaves were not significantly different from CW exposures in inducing increased brain NADH fluorescence and decreased ATP and CP concentrations. When the pulse-modulation frequency was decreased from 500 to 250 pulses per second the average incident power density threshold for inducing an increase in brain NADH fluorescence increased by a factor of 4--ie, from about 0.45 to about 1.85 mW/cm2. Since brain temperature did not increase, the microwave-induced increase in brain NADH and decrease in ATP and CP concentrations was not due to hyperthermia. This suggests a direct interaction mechanism and is consistent with the hypothesis of microwave inhibition of mitochondrial electron transport chain function of ATP production.

  4. Role of forkhead box protein A3 in age-associated metabolic decline

    PubMed Central

    Ma, Xinran; Xu, Lingyan; Gavrilova, Oksana; Mueller, Elisabetta

    2014-01-01

    Aging is associated with increased adiposity and diminished thermogenesis, but the critical transcription factors influencing these metabolic changes late in life are poorly understood. We recently demonstrated that the winged helix factor forkhead box protein A3 (Foxa3) regulates the expansion of visceral adipose tissue in high-fat diet regimens; however, whether Foxa3 also contributes to the increase in adiposity and the decrease in brown fat activity observed during the normal aging process is currently unknown. Here we report that during aging, levels of Foxa3 are significantly and selectively up-regulated in brown and inguinal white fat depots, and that midage Foxa3-null mice have increased white fat browning and thermogenic capacity, decreased adipose tissue expansion, improved insulin sensitivity, and increased longevity. Foxa3 gain-of-function and loss-of-function studies in inguinal adipose depots demonstrated a cell-autonomous function for Foxa3 in white fat tissue browning. Furthermore, our analysis revealed that the mechanisms of Foxa3 modulation of brown fat gene programs involve the suppression of peroxisome proliferator activated receptor γ coactivtor 1 α (PGC1α) levels through interference with cAMP responsive element binding protein 1-mediated transcriptional regulation of the PGC1α promoter. Overall, our data demonstrate a role for Foxa3 in energy expenditure and in age-associated metabolic disorders. PMID:25225406

  5. Quantitative Proteomics Analysis Reveals the Min System of Escherichia coli Modulates Reversible Protein Association with the Inner Membrane.

    PubMed

    Lee, Hsiao-Lin; Chiang, I-Chen; Liang, Suh-Yuen; Lee, Der-Yen; Chang, Geen-Dong; Wang, Kwan-Yu; Lin, Shu-Yu; Shih, Yu-Ling

    2016-05-01

    The Min system of Escherichia coli mediates placement of the division septum at the midcell. It oscillates from pole to pole to establish a concentration gradient of the division inhibition that is high at the poles but low at the midcell; the cell middle thereby becomes the most favorable site for division. Although Min oscillation is well studied from molecular and biophysical perspectives, it is still an enigma as to whether such a continuous, energy-consuming, and organized movement of the Min proteins would affect cellular processes other than the division site selection. To tackle this question, we compared the inner membrane proteome of the wild-type and Δmin strains using a quantitative approach. Forty proteins that showed differential abundance on the inner membrane of the mutant cells were identified and defined as proteins of interest (POIs). More than half of the POIs were peripheral membrane proteins, suggesting that the Min system affects mainly reversible protein association with the inner membrane. In addition, 6 out of 10 selected POIs directly interacted with at least one of the Min proteins, confirming the correlation between POIs and the Min system.Further analysis revealed a functional relationship between metabolism and the Min system. Metabolic enzymes accounted for 45% of the POIs, and there was a change of metabolites in the related reactions. We hypothesize that the Min system could alter the membrane location of proteins to modulate their enzymatic activity. Thus, the metabolic modulation in the Δmin mutant is likely an adaptive phenotype in cells of abnormal size and chromosome number due to an imbalanced abundance of proteins on the inner membrane. Taken together, the current work reports novel interactions of the Min system and reveals a global physiological impact of the Min system in addition to the division site placement.

  6. Modules of co-regulated metabolites in turmeric (Curcuma longa) rhizome suggest the existence of biosynthetic modules in plant specialized metabolism.

    PubMed

    Xie, Zhengzhi; Ma, Xiaoqiang; Gang, David R

    2009-01-01

    Turmeric is an excellent example of a plant that produces large numbers of metabolites from diverse metabolic pathways or networks. It is hypothesized that these metabolic pathways or networks contain biosynthetic modules, which lead to the formation of metabolite modules-groups of metabolites whose production is co-regulated and biosynthetically linked. To test whether such co-regulated metabolite modules do exist in this plant, metabolic profiling analysis was performed on turmeric rhizome samples that were collected from 16 different growth and development treatments, which had significant impacts on the levels of 249 volatile and non-volatile metabolites that were detected. Importantly, one of the many co-regulated metabolite modules that were indeed readily detected in this analysis contained the three major curcuminoids, whereas many other structurally related diarylheptanoids belonged to separate metabolite modules, as did groups of terpenoids. The existence of these co-regulated metabolite modules supported the hypothesis that the 3-methoxyl groups on the aromatic rings of the curcuminoids are formed before the formation of the heptanoid backbone during the biosynthesis of curcumin and also suggested the involvement of multiple polyketide synthases with different substrate selectivities in the formation of the array of diarylheptanoids detected in turmeric. Similar conclusions about terpenoid biosynthesis could also be made. Thus, discovery and analysis of metabolite modules can be a powerful predictive tool in efforts to understand metabolism in plants.

  7. Capacitance-modulated transistor detects odorant binding protein chiral interactions.

    PubMed

    Mulla, Mohammad Yusuf; Tuccori, Elena; Magliulo, Maria; Lattanzi, Gianluca; Palazzo, Gerardo; Persaud, Krishna; Torsi, Luisa

    2015-01-01

    Peripheral events in olfaction involve odorant binding proteins (OBPs) whose role in the recognition of different volatile chemicals is yet unclear. Here we report on the sensitive and quantitative measurement of the weak interactions associated with neutral enantiomers differentially binding to OBPs immobilized through a self-assembled monolayer to the gate of an organic bio-electronic transistor. The transduction is remarkably sensitive as the transistor output current is governed by the small capacitance of the protein layer undergoing minute changes as the ligand-protein complex is formed. Accurate determination of the free-energy balances and of the capacitance changes associated with the binding process allows derivation of the free-energy components as well as of the occurrence of conformational events associated with OBP ligand binding. Capacitance-modulated transistors open a new pathway for the study of ultra-weak molecular interactions in surface-bound protein-ligand complexes through an approach that combines bio-chemical and electronic thermodynamic parameters. PMID:25591754

  8. Capacitance-modulated transistor detects odorant binding protein chiral interactions

    NASA Astrophysics Data System (ADS)

    Mulla, Mohammad Yusuf; Tuccori, Elena; Magliulo, Maria; Lattanzi, Gianluca; Palazzo, Gerardo; Persaud, Krishna; Torsi, Luisa

    2015-01-01

    Peripheral events in olfaction involve odorant binding proteins (OBPs) whose role in the recognition of different volatile chemicals is yet unclear. Here we report on the sensitive and quantitative measurement of the weak interactions associated with neutral enantiomers differentially binding to OBPs immobilized through a self-assembled monolayer to the gate of an organic bio-electronic transistor. The transduction is remarkably sensitive as the transistor output current is governed by the small capacitance of the protein layer undergoing minute changes as the ligand-protein complex is formed. Accurate determination of the free-energy balances and of the capacitance changes associated with the binding process allows derivation of the free-energy components as well as of the occurrence of conformational events associated with OBP ligand binding. Capacitance-modulated transistors open a new pathway for the study of ultra-weak molecular interactions in surface-bound protein-ligand complexes through an approach that combines bio-chemical and electronic thermodynamic parameters.

  9. Contact sensitizers modulate the arachidonic acid metabolism of PMA-differentiated U-937 monocytic cells activated by LPS

    SciTech Connect

    Del Bufalo, Aurelia; Bernad, Jose; Dardenne, Christophe; Verda, Denis; Meunier, Jean Roch; Rousset, Francoise; Martinozzi-Teissier, Silvia; Pipy, Bernard

    2011-10-01

    For the effective induction of a hapten-specific T cell immune response toward contact sensitizers, in addition to covalent-modification of skin proteins, the redox and inflammatory statuses of activated dendritic cells are crucial. The aim of this study was to better understand how sensitizers modulate an inflammatory response through cytokines production and COX metabolism cascade. To address this purpose, we used the human monocytic-like U-937 cell line differentiated by phorbol myristate acetate (PMA) and investigated the effect of 6 contact sensitizers (DNCB, PPD, hydroquinone, propyl gallate, cinnamaldehyde and eugenol) and 3 non sensitizers (lactic acid, glycerol and tween 20) on the production of pro-inflammatory cytokines (IL-1{beta} and TNF-{alpha}) and on the arachidonic acid metabolic profile after bacterial lipopolysaccharide (LPS) stimulation. Our results showed that among the tested molecules, all sensitizers specifically prevent the production of PMA/LPS-induced COX-2 metabolites (PGE{sub 2,} TxB{sub 2} and PGD{sub 2}), eugenol and cinnamaldehyde inhibiting also the production of IL-1{beta} and TNF-{alpha}. We further demonstrated that there is no unique PGE{sub 2} inhibition mechanism: while the release of arachidonic acid (AA) from membrane phospholipids does not appear do be a target of modulation, COX-2 expression and/or COX-2 enzymatic activity are the major steps of prostaglandin synthesis that are inhibited by sensitizers. Altogether these results add a new insight into the multiple biochemical effects described for sensitizers. - Highlights: > We investigated how contact sensitizers modulate an inflammatory response. > We used macrophage-differentiated cell line, U-937 treated with PMA/LPS. > Sensitizers specifically inhibit the production of COX metabolites (PGE2, TxB2). > Several mechanisms of inhibition: COX-2 expression/enzymatic activity, isomerases. > New insight in the biochemical properties of sensitizers.

  10. A new level of regulation in gluconeogenesis: metabolic state modulates the intracellular localization of aldolase B and its interaction with liver fructose-1,6-bisphosphatase.

    PubMed

    Droppelmann, Cristian A; Sáez, Doris E; Asenjo, Joel L; Yáñez, Alejandro J; García-Rocha, Mar; Concha, Ilona I; Grez, Manuel; Guinovart, Joan J; Slebe, Juan C

    2015-12-01

    Understanding how glucose metabolism is finely regulated at molecular and cellular levels in the liver is critical for knowing its relationship to related pathologies, such as diabetes. In order to gain insight into the regulation of glucose metabolism, we studied the liver-expressed isoforms aldolase B and fructose-1,6-bisphosphatase-1 (FBPase-1), key enzymes in gluconeogenesis, analysing their cellular localization in hepatocytes under different metabolic conditions and their protein-protein interaction in vitro and in vivo. We observed that glucose, insulin, glucagon and adrenaline differentially modulate the intracellular distribution of aldolase B and FBPase-1. Interestingly, the in vitro protein-protein interaction analysis between aldolase B and FBPase-1 showed a specific and regulable interaction between them, whereas aldolase A (muscle isozyme) and FBPase-1 showed no interaction. The affinity of the aldolase B and FBPase-1 complex was modulated by intermediate metabolites, but only in the presence of K(+). We observed a decreased association constant in the presence of adenosine monophosphate, fructose-2,6-bisphosphate, fructose-6-phosphate and inhibitory concentrations of fructose-1,6-bisphosphate. Conversely, the association constant of the complex increased in the presence of dihydroxyacetone phosphate (DHAP) and non-inhibitory concentrations of fructose-1,6-bisphosphate. Notably, in vivo FRET studies confirmed the interaction between aldolase B and FBPase-1. Also, the co-expression of aldolase B and FBPase-1 in cultured cells suggested that FBPase-1 guides the cellular localization of aldolase B. Our results provide further evidence that metabolic conditions modulate aldolase B and FBPase-1 activity at the cellular level through the regulation of their interaction, suggesting that their association confers a catalytic advantage for both enzymes. PMID:26417114

  11. A new level of regulation in gluconeogenesis: metabolic state modulates the intracellular localization of aldolase B and its interaction with liver fructose-1,6-bisphosphatase.

    PubMed

    Droppelmann, Cristian A; Sáez, Doris E; Asenjo, Joel L; Yáñez, Alejandro J; García-Rocha, Mar; Concha, Ilona I; Grez, Manuel; Guinovart, Joan J; Slebe, Juan C

    2015-12-01

    Understanding how glucose metabolism is finely regulated at molecular and cellular levels in the liver is critical for knowing its relationship to related pathologies, such as diabetes. In order to gain insight into the regulation of glucose metabolism, we studied the liver-expressed isoforms aldolase B and fructose-1,6-bisphosphatase-1 (FBPase-1), key enzymes in gluconeogenesis, analysing their cellular localization in hepatocytes under different metabolic conditions and their protein-protein interaction in vitro and in vivo. We observed that glucose, insulin, glucagon and adrenaline differentially modulate the intracellular distribution of aldolase B and FBPase-1. Interestingly, the in vitro protein-protein interaction analysis between aldolase B and FBPase-1 showed a specific and regulable interaction between them, whereas aldolase A (muscle isozyme) and FBPase-1 showed no interaction. The affinity of the aldolase B and FBPase-1 complex was modulated by intermediate metabolites, but only in the presence of K(+). We observed a decreased association constant in the presence of adenosine monophosphate, fructose-2,6-bisphosphate, fructose-6-phosphate and inhibitory concentrations of fructose-1,6-bisphosphate. Conversely, the association constant of the complex increased in the presence of dihydroxyacetone phosphate (DHAP) and non-inhibitory concentrations of fructose-1,6-bisphosphate. Notably, in vivo FRET studies confirmed the interaction between aldolase B and FBPase-1. Also, the co-expression of aldolase B and FBPase-1 in cultured cells suggested that FBPase-1 guides the cellular localization of aldolase B. Our results provide further evidence that metabolic conditions modulate aldolase B and FBPase-1 activity at the cellular level through the regulation of their interaction, suggesting that their association confers a catalytic advantage for both enzymes.

  12. miR-182 Regulates Metabolic Homeostasis by Modulating Glucose Utilization in Muscle.

    PubMed

    Zhang, Duo; Li, Yan; Yao, Xuan; Wang, Hui; Zhao, Lei; Jiang, Haowen; Yao, Xiaohan; Zhang, Shengjie; Ye, Cheng; Liu, Wei; Cao, Hongchao; Yu, Shuxian; Wang, Yu-Cheng; Li, Qiong; Jiang, Jingjing; Liu, Yi; Zhang, Ling; Liu, Yun; Iwai, Naoharu; Wang, Hui; Li, Jingya; Li, Jia; Li, Xihua; Jin, Zi-Bing; Ying, Hao

    2016-07-19

    Understanding the fiber-type specification and metabolic switch in skeletal muscle provides insights into energy metabolism in physiology and diseases. Here, we show that miR-182 is highly expressed in fast-twitch muscle and negatively correlates with blood glucose level. miR-182 knockout mice display muscle loss, fast-to-slow fiber-type switching, and impaired glucose metabolism. Mechanistic studies reveal that miR-182 modulates glucose utilization in muscle by targeting FoxO1 and PDK4, which control fuel selection via the pyruvate dehydrogenase complex (PDHC). Short-term high-fat diet (HFD) feeding reduces muscle miR-182 levels by tumor necrosis factor α (TNFα), which contributes to the upregulation of FoxO1/PDK4. Restoration of miR-182 expression in HFD-fed mice induces a faster muscle phenotype, decreases muscle FoxO1/PDK4 levels, and improves glucose metabolism. Together, our work establishes miR-182 as a critical regulator that confers robust and precise controls on fuel usage and glucose homeostasis. Our study suggests that a metabolic shift toward a faster and more glycolytic phenotype is beneficial for glucose control. PMID:27396327

  13. Cell-wall invertases, key enzymes in the modulation of plant metabolism during defence responses.

    PubMed

    Proels, Reinhard Korbinian; Hückelhoven, Ralph

    2014-10-01

    Most plant-pathogen interactions do not result in pathogenesis because of pre-formed defensive plant barriers or pathogen-triggered activation of effective plant immune responses. The mounting of defence reactions is accompanied by a profound modulation of plant metabolism. Common metabolic changes are the repression of photosynthesis, the increase in heterotrophic metabolism and the synthesis of secondary metabolites. This enhanced metabolic activity is accompanied by the reduced export of sucrose or enhanced import of hexoses at the site of infection, which is mediated by an induced activity of cell-wall invertase (Cw-Inv). Cw-Inv cleaves sucrose, the major transport sugar in plants, irreversibly yielding glucose and fructose, which can be taken up by plant cells via hexose transporters. These hexose sugars not only function in metabolism, but also act as signalling molecules. The picture of Cw-Inv regulation in plant-pathogen interactions has recently been broadened and is discussed in this review. An interesting emerging feature is the link between Cw-Inv and the circadian clock and new modes of Cw-Inv regulation at the post-translational level.

  14. Testosterone Replacement Modulates Cardiac Metabolic Remodeling after Myocardial Infarction by Upregulating PPARα

    PubMed Central

    Yang, Jing

    2016-01-01

    Despite the importance of testosterone as a metabolic hormone, its effects on myocardial metabolism in the ischemic heart remain unclear. Myocardial ischemia leads to metabolic remodeling, ultimately resulting in ATP deficiency and cardiac dysfunction. In the present study, the effects of testosterone replacement on the ischemic heart were assessed in a castrated rat myocardial infarction model established by ligating the left anterior descending coronary artery 2 weeks after castration. The results of real-time PCR and Western blot analyses showed that peroxisome proliferator-activated receptor α (PPARα) decreased in the ischemic myocardium of castrated rats, compared with the sham-castration group, and the mRNA expression of genes involved in fatty acid metabolism (the fatty acid translocase CD36, carnitine palmitoyltransferase I, and medium-chain acyl-CoA dehydrogenase) and glucose transporter-4 also decreased. A decline in ATP levels in the castrated rats was accompanied by increased cardiomyocyte apoptosis and fibrosis and impaired cardiac function, compared with the sham-castration group, and these detrimental effects were reversed by testosterone replacement. Taken together, our findings suggest that testosterone can modulate myocardial metabolic remodeling by upregulating PPARα after myocardial infarction, exerting a protective effect on cardiac function. PMID:27413362

  15. Modulation of P1798 lymphosarcoma proliferation by protein phosphorylation

    SciTech Connect

    Michnoff, C.A.H.

    1983-01-01

    The role of protein kinases in modulating cell proliferation was examined. Studies characterized the regulation of cell proliferation by adenosine 3':5'-monophosphate-dependent protein kinase (cA-Pk). Calcium/calmodulin-dependent myosin light chain kinase (MLCK) was isolated and examined as a potential substrate regulated by cA-PK in the rapidly proliferating P1798 lymphosarcoma. Modulation of cell proliferation by cA-PK was characterized by quantitating cell division by (methyl-/sup 3/H) thymidine ((/sup 3/H)-dT) incorporation into DNA, cAMP accumulations, and activation of cA-PK using P1798 lymphosarcoma cells. Epinephrine and prostaglandin E/sub 1/ (PGE/sub 1/) were demonstrated to suppress (/sup 3/H)-dT incorporation into DNA, to stimulate cAMP accumulation, and to activate cA-PK with dose-dependency. Calcium/calmodulin-dependent MLCK was partially purified from P1798 lymphosarcoma. P1798 MLCK phosphorylated myosin regulatory light chains (P-LC) from thymus, cardiac and skeletal muscles. One mol (/sup 32/Pi) was transferred into one mol cardiac or skeletal P-LC by P1798 MLCK. Apparent Km values of 65 ..mu..M and 51 ..mu..M were determined for ATP and cardiac P-LC, respectively. The apparent molecular weight of P1798 MLCK was 135,000. P1798 MLCK was phosphorylated by cA-PK. Phosphorylated MLCK showed a 41% decrease in calcium-dependent activity. Two additional protein kinases from P1798 lymphosarcoma phosphorylated cardiac and skeletal light chains (MLC).

  16. Starch Granule Re-Structuring by Starch Branching Enzyme and Glucan Water Dikinase Modulation Affects Caryopsis Physiology and Metabolism.

    PubMed

    Shaik, Shahnoor S; Obata, Toshihiro; Hebelstrup, Kim H; Schwahn, Kevin; Fernie, Alisdair R; Mateiu, Ramona V; Blennow, Andreas

    2016-01-01

    Starch is of fundamental importance for plant development and reproduction and its optimized molecular assembly is potentially necessary for correct starch metabolism. Re-structuring of starch granules in-planta can therefore potentially affect plant metabolism. Modulation of granule micro-structure was achieved by decreasing starch branching and increasing starch-bound phosphate content in the barley caryopsis starch by RNAi suppression of all three Starch Branching Enzyme (SBE) isoforms or overexpression of potato Glucan Water Dikinase (GWD). The resulting lines displayed Amylose-Only (AO) and Hyper-Phosphorylated (HP) starch chemotypes, respectively. We studied the influence of these alterations on primary metabolism, grain composition, starch structural features and starch granule morphology over caryopsis development at 10, 20 and 30 days after pollination (DAP) and at grain maturity. While HP showed relatively little effect, AO showed significant reduction in starch accumulation with re-direction to protein and β-glucan (BG) accumulation. Metabolite profiling indicated significantly higher sugar accumulation in AO, with re-partitioning of carbon to accumulate amino acids, and interestingly it also had high levels of some important stress-related metabolites and potentially protective metabolites, possibly to elude deleterious effects. Investigations on starch molecular structure revealed significant increase in starch phosphate and amylose content in HP and AO respectively with obvious differences in starch granule morphology at maturity. The results demonstrate that decreasing the storage starch branching resulted in metabolic adjustments and re-directions, tuning to evade deleterious effects on caryopsis physiology and plant performance while only little effect was evident by increasing starch-bound phosphate as a result of overexpressing GWD.

  17. Starch Granule Re-Structuring by Starch Branching Enzyme and Glucan Water Dikinase Modulation Affects Caryopsis Physiology and Metabolism.

    PubMed

    Shaik, Shahnoor S; Obata, Toshihiro; Hebelstrup, Kim H; Schwahn, Kevin; Fernie, Alisdair R; Mateiu, Ramona V; Blennow, Andreas

    2016-01-01

    Starch is of fundamental importance for plant development and reproduction and its optimized molecular assembly is potentially necessary for correct starch metabolism. Re-structuring of starch granules in-planta can therefore potentially affect plant metabolism. Modulation of granule micro-structure was achieved by decreasing starch branching and increasing starch-bound phosphate content in the barley caryopsis starch by RNAi suppression of all three Starch Branching Enzyme (SBE) isoforms or overexpression of potato Glucan Water Dikinase (GWD). The resulting lines displayed Amylose-Only (AO) and Hyper-Phosphorylated (HP) starch chemotypes, respectively. We studied the influence of these alterations on primary metabolism, grain composition, starch structural features and starch granule morphology over caryopsis development at 10, 20 and 30 days after pollination (DAP) and at grain maturity. While HP showed relatively little effect, AO showed significant reduction in starch accumulation with re-direction to protein and β-glucan (BG) accumulation. Metabolite profiling indicated significantly higher sugar accumulation in AO, with re-partitioning of carbon to accumulate amino acids, and interestingly it also had high levels of some important stress-related metabolites and potentially protective metabolites, possibly to elude deleterious effects. Investigations on starch molecular structure revealed significant increase in starch phosphate and amylose content in HP and AO respectively with obvious differences in starch granule morphology at maturity. The results demonstrate that decreasing the storage starch branching resulted in metabolic adjustments and re-directions, tuning to evade deleterious effects on caryopsis physiology and plant performance while only little effect was evident by increasing starch-bound phosphate as a result of overexpressing GWD. PMID:26891365

  18. Starch Granule Re-Structuring by Starch Branching Enzyme and Glucan Water Dikinase Modulation Affects Caryopsis Physiology and Metabolism

    PubMed Central

    Shaik, Shahnoor S.; Obata, Toshihiro; Hebelstrup, Kim H.; Schwahn, Kevin; Fernie, Alisdair R.; Mateiu, Ramona V.; Blennow, Andreas

    2016-01-01

    Starch is of fundamental importance for plant development and reproduction and its optimized molecular assembly is potentially necessary for correct starch metabolism. Re-structuring of starch granules in-planta can therefore potentially affect plant metabolism. Modulation of granule micro-structure was achieved by decreasing starch branching and increasing starch-bound phosphate content in the barley caryopsis starch by RNAi suppression of all three Starch Branching Enzyme (SBE) isoforms or overexpression of potato Glucan Water Dikinase (GWD). The resulting lines displayed Amylose-Only (AO) and Hyper-Phosphorylated (HP) starch chemotypes, respectively. We studied the influence of these alterations on primary metabolism, grain composition, starch structural features and starch granule morphology over caryopsis development at 10, 20 and 30 days after pollination (DAP) and at grain maturity. While HP showed relatively little effect, AO showed significant reduction in starch accumulation with re-direction to protein and β-glucan (BG) accumulation. Metabolite profiling indicated significantly higher sugar accumulation in AO, with re-partitioning of carbon to accumulate amino acids, and interestingly it also had high levels of some important stress-related metabolites and potentially protective metabolites, possibly to elude deleterious effects. Investigations on starch molecular structure revealed significant increase in starch phosphate and amylose content in HP and AO respectively with obvious differences in starch granule morphology at maturity. The results demonstrate that decreasing the storage starch branching resulted in metabolic adjustments and re-directions, tuning to evade deleterious effects on caryopsis physiology and plant performance while only little effect was evident by increasing starch-bound phosphate as a result of overexpressing GWD. PMID:26891365

  19. Defining meal requirements for protein to optimize metabolic roles of amino acids

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Dietary protein provides essential amino acids (EAAs) for the synthesis of new proteins plus an array of other metabolic functions; many of these functions are sensitive to postprandial plasma and intracellular amino acid concentrations. Recent research has focused on amino acids as metabolic signal...

  20. Defining meal requirements for protein to optimize metabolic roles of amino acids

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Dietary protein provides essential amino acids (EAA) for the synthesis of new proteins plus an array of other metabolic functions; many of these functions are sensitive to post-prandial plasma and intracellular amino acid concentrations. Recent research has focused on amino acids as metabolic signal...

  1. Nur77 modulates hepatic lipid metabolism through suppression of SREBP1c activity

    SciTech Connect

    Pols, Thijs W.H.; Ottenhoff, Roelof; Vos, Mariska; Levels, Johannes H.M.; Quax, Paul H.A.; Meijers, Joost C.M.; Pannekoek, Hans; Groen, Albert K.; Vries, Carlie J.M. de

    2008-02-22

    NR4A nuclear receptors are induced in the liver upon fasting and regulate hepatic gluconeogenesis. Here, we studied the role of nuclear receptor Nur77 (NR4A1) in hepatic lipid metabolism. We generated mice expressing hepatic Nur77 using adenoviral vectors, and demonstrate that these mice exhibit a modulation of the plasma lipid profile and a reduction in hepatic triglyceride. Expression analysis of >25 key genes involved in lipid metabolism revealed that Nur77 inhibits SREBP1c expression. This results in decreased SREBP1c activity as is illustrated by reduced expression of its target genes stearoyl-coA desaturase-1, mitochondrial glycerol-3-phosphate acyltransferase, fatty acid synthase and the LDL receptor, and provides a mechanism for the physiological changes observed in response to Nur77. Expression of LXR target genes Abcg5 and Abcg8 is reduced by Nur77, and may suggest involvement of LXR in the inhibitory action of Nur77 on SREBP1c expression. Taken together, our study demonstrates that Nur77 modulates hepatic lipid metabolism through suppression of SREBP1c activity.

  2. System-wide assembly of pathways and modules hierarchically reveal metabolic mechanism of cerebral ischemia

    PubMed Central

    Zhu, Yan; Guo, Zhili; Zhang, Liangxiao; Zhang, Yingying; Chen, Yinying; Nan, Jingyi; Zhao, Buchang; Xiao, Hongbin; Wang, Zhong; Wang, Yongyan

    2015-01-01

    The relationship between cerebral ischemia and metabolic disorders is poorly understood, which is partly due to the lack of comparative fusing data for larger complete systems and to the complexity of metabolic cascade reactions. Based on the fusing maps of comprehensive serum metabolome, fatty acid and amino acid profiling, we identified 35 potential metabolic biomarkers for ischemic stroke. Our analyses revealed 8 significantly altered pathways by MetPA (Metabolomics Pathway Analysis, impact score >0.10) and 15 significantly rewired modules in a complex ischemic network using the Markov clustering (MCL) method; all of these pathways became more homologous as the number of overlapping nodes was increased. We then detected 24 extensive pathways based on the total modular nodes from the network analysis, 12 of which were new discovery pathways. We provided a new perspective from the viewpoint of abnormal metabolites for the overall study of ischemic stroke as well as a new method to simplify the network analysis by selecting the more closely connected edges and nodes to build a module map of stroke. PMID:26621314

  3. Modulation of glucose transporter protein by dietary flavonoids in type 2 diabetes mellitus.

    PubMed

    Hajiaghaalipour, Fatemeh; Khalilpourfarshbafi, Manizheh; Arya, Aditya

    2015-01-01

    Diabetes mellitus (DM) is a metabolic diseases characterized by hyperglycemia due to insufficient or inefficient insulin secretory response. This chronic disease is a global problem and there is a need for greater emphasis on therapeutic strategies in the health system. Phytochemicals such as flavonoids have recently attracted attention as source materials for the development of new antidiabetic drugs or alternative therapy for the management of diabetes and its related complications. The antidiabetic potential of flavonoids are mainly through their modulatory effects on glucose transporter by enhancing GLUT-2 expression in pancreatic β cells and increasing expression and promoting translocation of GLUT-4 via PI3K/AKT, CAP/Cb1/TC10 and AMPK pathways. This review highlights the recent findings on beneficial effects of flavonoids in the management of diabetes with particular emphasis on the investigations that explore the role of these compounds in modulating glucose transporter proteins at cellular and molecular level.

  4. Modulation of Glucose Transporter Protein by Dietary Flavonoids in Type 2 Diabetes Mellitus

    PubMed Central

    Hajiaghaalipour, Fatemeh; Khalilpourfarshbafi, Manizheh; Arya, Aditya

    2015-01-01

    Diabetes mellitus (DM) is a metabolic diseases characterized by hyperglycemia due to insufficient or inefficient insulin secretory response. This chronic disease is a global problem and there is a need for greater emphasis on therapeutic strategies in the health system. Phytochemicals such as flavonoids have recently attracted attention as source materials for the development of new antidiabetic drugs or alternative therapy for the management of diabetes and its related complications. The antidiabetic potential of flavonoids are mainly through their modulatory effects on glucose transporter by enhancing GLUT-2 expression in pancreatic β cells and increasing expression and promoting translocation of GLUT-4 via PI3K/AKT, CAP/Cb1/TC10 and AMPK pathways. This review highlights the recent findings on beneficial effects of flavonoids in the management of diabetes with particular emphasis on the investigations that explore the role of these compounds in modulating glucose transporter proteins at cellular and molecular level. PMID:25892959

  5. Trace levels of innate immune response modulating impurities (IIRMIs) synergize to break tolerance to therapeutic proteins.

    PubMed

    Verthelyi, Daniela; Wang, Vivian

    2010-12-22

    Therapeutic proteins such as monoclonal antibodies, replacement enzymes and toxins have significantly improved the therapeutic options for multiple diseases, including cancer and inflammatory diseases as well as enzyme deficiencies and inborn errors of metabolism. However, immune responses to these products are frequent and can seriously impact their safety and efficacy. Of the many factors that can impact protein immunogenicity, this study focuses on the role of innate immune response modulating impurities (IIRMIs) that could be present despite product purification and whether these impurities can synergize to facilitate an immunogenic response to therapeutic proteins. Using lipopolysaccharide (LPS) and CpG ODN as IIRMIs we showed that trace levels of these impurities synergized to induce IgM, IFNγ, TNFα and IL-6 expression. In vivo, trace levels of these impurities synergized to increase antigen-specific IgG antibodies to ovalbumin. Further, whereas mice treated with human erythropoietin showed a transient increase in hematocrit, those that received human erythropoietin containing low levels of IIRMIs had reduced response to erythropoietin after the 1(st) dose and developed long-lasting anemia following subsequent doses. This suggests that the presence of IIRMIs facilitated a breach in tolerance to the endogenous mouse erythropoietin. Overall, these studies indicate that the risk of enhancing immunogenicity should be considered when establishing acceptance limits of IIRMIs for therapeutic proteins.

  6. Effects of Tamarindus indica fruit pulp extract on abundance of HepG2 cell lysate proteins and their possible consequential impact on metabolism and inflammation.

    PubMed

    Chong, Ursula R W; Abdul-Rahman, Puteri S; Abdul-Aziz, Azlina; Hashim, Onn H; Mat-Junit, Sarni

    2013-01-01

    The fruit pulp extract of Tamarindus indica has been reported for its antioxidant and hypolipidemic properties. In this study, the methanol extract of T. indica fruit pulp was investigated for its effects on the abundance of HepG2 cell lysate proteins. Cell lysate was extracted from HepG2 cells grown in the absence and presence of the methanol extract of T. indica fruit pulp. Approximately 2500 spots were resolved using two-dimensional gel electrophoresis and the abundance of 20 cellular proteins was found to be significantly reduced. Among the proteins of reduced abundance, fourteen, including six proteins involved in metabolism (including ethanolamine phosphate cytidylyltransferase), four mitochondrial proteins (including prohibitin and respiratory chain proteins), and four proteins involved in translation and splicing, were positively identified by mass spectrometry and database search. The identified HepG2 altered abundance proteins, when taken together and analyzed by Ingenuity Pathways Analysis (IPA) software, are suggestive of the effects of T. indica fruit pulp extract on metabolism and inflammation, which are modulated by LXR/RXR. In conclusion, the methanol fruit pulp extract of T. indica was shown to cause reduced abundance of HepG2 mitochondrial, metabolic, and regulatory proteins involved in oxidative phosphorylation, protein synthesis, and cellular metabolism.

  7. Effects of Tamarindus indica Fruit Pulp Extract on Abundance of HepG2 Cell Lysate Proteins and Their Possible Consequential Impact on Metabolism and Inflammation

    PubMed Central

    Chong, Ursula R. W.; Abdul-Rahman, Puteri S.; Abdul-Aziz, Azlina; Hashim, Onn H.; Mat-Junit, Sarni

    2013-01-01

    The fruit pulp extract of Tamarindus indica has been reported for its antioxidant and hypolipidemic properties. In this study, the methanol extract of T. indica fruit pulp was investigated for its effects on the abundance of HepG2 cell lysate proteins. Cell lysate was extracted from HepG2 cells grown in the absence and presence of the methanol extract of T. indica fruit pulp. Approximately 2500 spots were resolved using two-dimensional gel electrophoresis and the abundance of 20 cellular proteins was found to be significantly reduced. Among the proteins of reduced abundance, fourteen, including six proteins involved in metabolism (including ethanolamine phosphate cytidylyltransferase), four mitochondrial proteins (including prohibitin and respiratory chain proteins), and four proteins involved in translation and splicing, were positively identified by mass spectrometry and database search. The identified HepG2 altered abundance proteins, when taken together and analyzed by Ingenuity Pathways Analysis (IPA) software, are suggestive of the effects of T. indica fruit pulp extract on metabolism and inflammation, which are modulated by LXR/RXR. In conclusion, the methanol fruit pulp extract of T. indica was shown to cause reduced abundance of HepG2 mitochondrial, metabolic, and regulatory proteins involved in oxidative phosphorylation, protein synthesis, and cellular metabolism. PMID:24455694

  8. Capacitance-modulated transistor detects odorant binding protein chiral interactions

    PubMed Central

    Mulla, Mohammad Yusuf; Tuccori, Elena; Magliulo, Maria; Lattanzi, Gianluca; Palazzo, Gerardo; Persaud, Krishna; Torsi, Luisa

    2015-01-01

    Peripheral events in olfaction involve odorant binding proteins (OBPs) whose role in the recognition of different volatile chemicals is yet unclear. Here we report on the sensitive and quantitative measurement of the weak interactions associated with neutral enantiomers differentially binding to OBPs immobilized through a self-assembled monolayer to the gate of an organic bio-electronic transistor. The transduction is remarkably sensitive as the transistor output current is governed by the small capacitance of the protein layer undergoing minute changes as the ligand–protein complex is formed. Accurate determination of the free-energy balances and of the capacitance changes associated with the binding process allows derivation of the free-energy components as well as of the occurrence of conformational events associated with OBP ligand binding. Capacitance-modulated transistors open a new pathway for the study of ultra-weak molecular interactions in surface-bound protein–ligand complexes through an approach that combines bio-chemical and electronic thermodynamic parameters. PMID:25591754

  9. Flexible Linker Modulates Glycosaminoglycan Affinity of Decorin Binding Protein A.

    PubMed

    Morgan, Ashli; Sepuru, Krishna Mohan; Feng, Wei; Rajarathnam, Krishna; Wang, Xu

    2015-08-18

    Decorin binding protein A (DBPA) is a glycosaminoglycan (GAG)-binding adhesin found on the surface of the bacterium Borrelia burgdorferi (B. burgdorferi), the causative agent of Lyme disease. DBPA facilitates bacterial adherence to extracellular matrices of human tissues and is crucial during the early stage of the infection process. Interestingly, DBPA from different strains (B31, N40, and PBr) show significant differences in GAG affinities, but the structural basis for the differences is not clear. In this study, we show that GAG affinity of N40 DBPA is modulated in part by flexible segments that control access to the GAG binding site, such that shortening of the linker leads to higher GAG affinity when analyzed using ELISA, gel mobility shift assay, solution NMR, and isothermal titration calorimetry. Our observation that GAG affinity differences among different B. burgdorferi strains can be attributed to a flexible linker domain regulating access to the GAG-binding domain is novel. It also provides a rare example of how neutral amino acids and dynamic segments in GAG binding proteins can have a large influence on GAG affinity and provides insights into why the number of basic amino acids in the GAG-binding site may not be the only factor determining GAG affinity of proteins. PMID:26223367

  10. Sensitivity of Xanthoria parietina to UV-A: role of metabolic modulators.

    PubMed

    Kováčik, Jozef; Klejdus, Bořivoj; Stork, František; Malčovská, Silvia

    2011-06-01

    Effects of methyl jasmonate (MeJA), salicylic acid (SA) or 2-aminoindane-2-phosphonic acid (AIP) pre-treatments on the sensitivity of Xanthoria parietina exposed to UV-A were studied. UV decreased chlorophylls and stimulated increase in hydrogen peroxide and superoxide level. Accumulation of soluble phenols and flavonoids increased in response to UV treatment. Metabolic modulators had negligible impact on these UV-induced changes. Within free amino acids, AIP (-UV variant) and SA and MeJA (+UV variants) altered their accumulation. AIP had no effect on the amount of phenylalanine. Benzoic and cinnamic acids were elevated by UV and mainly MeJA influenced their accumulations. Among lichen specific metabolites, vulpinic acid and ergosterol increased while usnic acid and atranorin decreased after exposure to UV; accumulation of parietin was not affected. Applied modulators showed a different effect on these lichen metabolites but biosynthetic pathway-specific trend of alteration was visible. Overall, MeJA showed the most pronounced effect among studied parameters. Accumulation of selected phenolics in response to UV-A seems to be an important feature of Xanthoria tolerance. Present finding in the context of phenolic metabolism in non-vascular plants and with respect to limited data about effect of studied modulators on non-vascular plants are discussed. PMID:21531571

  11. Sensitivity of Xanthoria parietina to UV-A: role of metabolic modulators.

    PubMed

    Kováčik, Jozef; Klejdus, Bořivoj; Stork, František; Malčovská, Silvia

    2011-06-01

    Effects of methyl jasmonate (MeJA), salicylic acid (SA) or 2-aminoindane-2-phosphonic acid (AIP) pre-treatments on the sensitivity of Xanthoria parietina exposed to UV-A were studied. UV decreased chlorophylls and stimulated increase in hydrogen peroxide and superoxide level. Accumulation of soluble phenols and flavonoids increased in response to UV treatment. Metabolic modulators had negligible impact on these UV-induced changes. Within free amino acids, AIP (-UV variant) and SA and MeJA (+UV variants) altered their accumulation. AIP had no effect on the amount of phenylalanine. Benzoic and cinnamic acids were elevated by UV and mainly MeJA influenced their accumulations. Among lichen specific metabolites, vulpinic acid and ergosterol increased while usnic acid and atranorin decreased after exposure to UV; accumulation of parietin was not affected. Applied modulators showed a different effect on these lichen metabolites but biosynthetic pathway-specific trend of alteration was visible. Overall, MeJA showed the most pronounced effect among studied parameters. Accumulation of selected phenolics in response to UV-A seems to be an important feature of Xanthoria tolerance. Present finding in the context of phenolic metabolism in non-vascular plants and with respect to limited data about effect of studied modulators on non-vascular plants are discussed.

  12. Modulation of the Arginase Pathway in the Context of Microbial Pathogenesis: A Metabolic Enzyme Moonlighting as an Immune Modulator

    PubMed Central

    Lahiri, Ayan; Chakravortty, Dipshikha

    2010-01-01

    Arginine is a crucial amino acid that serves to modulate the cellular immune response during infection. Arginine is also a common substrate for both inducible nitric oxide synthase (iNOS) and arginase. The generation of nitric oxide from arginine is responsible for efficient immune response and cytotoxicity of host cells to kill the invading pathogens. On the other hand, the conversion of arginine to ornithine and urea via the arginase pathway can support the growth of bacterial and parasitic pathogens. The competition between iNOS and arginase for arginine can thus contribute to the outcome of several parasitic and bacterial infections. There are two isoforms of vertebrate arginase, both of which catalyze the conversion of arginine to ornithine and urea, but they differ with regard to tissue distribution and subcellular localization. In the case of infection with Mycobacterium, Leishmania, Trypanosoma, Helicobacter, Schistosoma, and Salmonella spp., arginase isoforms have been shown to modulate the pathology of infection by various means. Despite the existence of a considerable body of evidence about mammalian arginine metabolism and its role in immunology, the critical choice to divert the host arginine pool by pathogenic organisms as a survival strategy is still a mystery in infection biology. PMID:20585552

  13. Impact of Dietary Carbohydrate and Protein Levels on Carbohydrate Metabolism

    ERIC Educational Resources Information Center

    Lasker, Denise Ann

    2009-01-01

    The goal of this dissertation was to investigate the impact of changing dietary carbohydrate (CARB) intakes within recommended dietary guidelines on metabolic outcomes specifically associated with glycemic regulations and carbohydrate metabolism. This research utilized both human and animal studies to examine changes in metabolism across a wide…

  14. DELLA proteins modulate Arabidopsis defences induced in response to caterpillar herbivory.

    PubMed

    Lan, Zhiyi; Krosse, Sebastian; Achard, Patrick; van Dam, Nicole M; Bede, Jacqueline C

    2014-02-01

    Upon insect herbivory, many plant species change the direction of metabolic flux from growth into defence. Two key pathways modulating these processes are the gibberellin (GA)/DELLA pathway and the jasmonate pathway. In this study, the effect of caterpillar herbivory on plant-induced responses was compared between wild-type Arabidopsis thaliana (L.) Heynh. and quad-della mutants that have constitutively elevated GA responses. The labial saliva (LS) of caterpillars of the beet armyworm, Spodoptera exigua, is known to influence induced plant defence responses. To determine the role of this herbivore cue in determining metabolic shifts, plants were subject to herbivory by caterpillars with intact or impaired LS secretions. In both wild-type and quad-della plants, a jasmonate burst is an early response to caterpillar herbivory. Negative growth regulator DELLA proteins are required for the LS-mediated suppression of hormone levels. Jasmonate-dependent marker genes are induced in response to herbivory independently of LS, with the exception of AtPDF1.2 that showed LS-dependent expression in the quad-della mutant. Early expression of the salicylic acid (SA)-marker gene, AtPR1, was not affected by herbivory which also reflected SA hormone levels; however, this gene showed LS-dependent expression in the quad-della mutant. DELLA proteins may positively regulate glucosinolate levels and suppress laccase-like multicopper oxidase activity in response to herbivory. The present results show a link between DELLA proteins and early, induced plant defences in response to insect herbivory; in particular, these proteins are necessary for caterpillar LS-associated attenuation of defence hormones. PMID:24399173

  15. Insulin sensitivity of muscle protein metabolism is altered in patients with chronic kidney disease and metabolic acidosis

    PubMed Central

    Garibotto, Giacomo; Sofia, Antonella; Russo, Rodolfo; Paoletti, Ernesto; Bonanni, Alice; Parodi, Emanuele L; Viazzi, Francesca; Verzola, Daniela

    2015-01-01

    An emergent hypothesis is that a resistance to the anabolic drive by insulin may contribute to loss of strength and muscle mass in patients with chronic kidney disease (CKD). We tested whether insulin resistance extends to protein metabolism using the forearm perfusion method with arterial insulin infusion in 7 patients with CKD and metabolic acidosis (bicarbonate 19 mmol/l) and 7 control individuals. Forearm glucose balance and protein turnover (2H-phenylalanine kinetics) were measured basally and in response to insulin infused at different rates for 2 h to increase local forearm plasma insulin concentration by approximately 20 and 50 μU/ml. In response to insulin, forearm glucose uptake was significantly increased to a lesser extent (−40%) in patients with CKD than controls. In addition, whereas in the controls net muscle protein balance and protein degradation were decreased by both insulin infusion rates, in patients with CKD net protein balance and protein degradation were sensitive to the high (0.035 mU/kg per min) but not the low (0.01 mU/kg per min) insulin infusion. Besides blunting muscle glucose uptake, CKD and acidosis interfere with the normal suppression of protein degradation in response to a moderate rise in plasma insulin. Thus, alteration of protein metabolism by insulin may lead to changes in body tissue composition which may become clinically evident in conditions characterized by low insulinemia. PMID:26308671

  16. Insulin Is a Key Modulator of Fetoplacental Endothelium Metabolic Disturbances in Gestational Diabetes Mellitus

    PubMed Central

    Sobrevia, Luis; Salsoso, Rocío; Fuenzalida, Bárbara; Barros, Eric; Toledo, Lilian; Silva, Luis; Pizarro, Carolina; Subiabre, Mario; Villalobos, Roberto; Araos, Joaquín; Toledo, Fernando; González, Marcelo; Gutiérrez, Jaime; Farías, Marcelo; Chiarello, Delia I.; Pardo, Fabián; Leiva, Andrea

    2016-01-01

    Gestational diabetes mellitus (GDM) is a disease of the mother that associates with altered fetoplacental vascular function. GDM-associated maternal hyperglycaemia result in fetal hyperglycaemia, a condition that leads to fetal hyperinsulinemia and altered L-arginine transport and synthesis of nitric oxide, i.e., endothelial dysfunction. These alterations in the fetoplacental endothelial function are present in women with GDM that were under diet or insulin therapy. Since these women and their newborn show normal glycaemia at term, other factors or conditions could be altered and/or not resolved by restoring normal level of circulating D-glucose. GDM associates with metabolic disturbances, such as abnormal handling of the locally released vasodilator adenosine, and biosynthesis and metabolism of cholesterol lipoproteins, or metabolic diseases resulting in endoplasmic reticulum stress and altered angiogenesis. Insulin acts as a potent modulator of all these phenomena under normal conditions as reported in primary cultures of cells obtained from the human placenta; however, GDM and the role of insulin regarding these alterations in this disease are poorly understood. This review focuses on the potential link between insulin and endoplasmic reticulum stress, hypercholesterolemia, and angiogenesis in GDM in the human fetoplacental vasculature. Based in reports in primary culture placental endothelium we propose that insulin is a factor restoring endothelial function in GDM by reversing ERS, hypercholesterolaemia and angiogenesis to a physiological state involving insulin activation of insulin receptor isoforms and adenosine receptors and metabolism in the human placenta from GDM pregnancies. PMID:27065887

  17. Synergistic effects between catalase inhibitors and modulators of nitric oxide metabolism on tumor cell apoptosis.

    PubMed

    Scheit, Katrin; Bauer, Georg

    2014-10-01

    Inhibitors of catalase (such as ascorbate, methyldopa, salicylic acid and neutralizing antibodies) synergize with modulators of nitric oxide (NO) metabolism (such as arginine, arginase inhibitor, NO synthase-inducing interferons and NO dioxygenase inhibitors) in the singlet oxygen-mediated inactivation of tumor cell protective catalase. This is followed by reactive oxygen species (ROS)-dependent apoptosis induction. TGF-beta, NADPH oxidase-1, NO synthase, dual oxidase-1 and caspase-9 are characterized as essential catalysts in this process. The FAS receptor and caspase-8 are required for amplification of ROS signaling triggered by individual compounds, but are dispensable when the synergistic effect is established. Our findings explain the antitumor effects of catalase inhibitors and of compounds that target NO metabolism, as well as their synergy. These data may have an impact on epidemiological studies related to secondary plant compounds and open new perspectives for the establishment of novel antitumor drugs and for the improvement of established chemotherapeutics.

  18. Modulation of the chromatin phosphoproteome by the Haspin protein kinase.

    PubMed

    Maiolica, Alessio; de Medina-Redondo, Maria; Schoof, Erwin M; Chaikuad, Apirat; Villa, Fabrizio; Gatti, Marco; Jeganathan, Siva; Lou, Hua Jane; Novy, Karel; Hauri, Simon; Toprak, Umut H; Herzog, Franz; Meraldi, Patrick; Penengo, Lorenza; Turk, Benjamin E; Knapp, Stefan; Linding, Rune; Aebersold, Ruedi

    2014-07-01

    Recent discoveries have highlighted the importance of Haspin kinase activity for the correct positioning of the kinase Aurora B at the centromere. Haspin phosphorylates Thr(3) of the histone H3 (H3), which provides a signal for Aurora B to localize to the centromere of mitotic chromosomes. To date, histone H3 is the only confirmed Haspin substrate. We used a combination of biochemical, pharmacological, and mass spectrometric approaches to study the consequences of Haspin inhibition in mitotic cells. We quantified 3964 phosphorylation sites on chromatin-associated proteins and identified a Haspin protein-protein interaction network. We determined the Haspin consensus motif and the co-crystal structure of the kinase with the histone H3 tail. The structure revealed a unique bent substrate binding mode positioning the histone H3 residues Arg(2) and Lys(4) adjacent to the Haspin phosphorylated threonine into acidic binding pockets. This unique conformation of the kinase-substrate complex explains the reported modulation of Haspin activity by methylation of Lys(4) of the histone H3. In addition, the identification of the structural basis of substrate recognition and the amino acid sequence preferences of Haspin aided the identification of novel candidate Haspin substrates. In particular, we validated the phosphorylation of Ser(137) of the histone variant macroH2A as a target of Haspin kinase activity. MacroH2A Ser(137) resides in a basic stretch of about 40 amino acids that is required to stabilize extranucleosomal DNA, suggesting that phosphorylation of Ser(137) might regulate the interactions of macroH2A and DNA. Overall, our data suggest that Haspin activity affects the phosphorylation state of proteins involved in gene expression regulation and splicing.

  19. Protein Stability and Dynamics Modulation: The Case of Human Frataxin

    PubMed Central

    Gallo, Mariana; Salvay, Andres G.; Ferreiro, Diego U.; Santos, Javier

    2012-01-01

    Frataxin (FXN) is an α/β protein that plays an essential role in iron homeostasis. Apparently, the function of human FXN (hFXN) depends on the cooperative formation of crucial interactions between helix α1, helix α2, and the C-terminal region (CTR) of the protein. In this work we quantitatively explore these relationships using a purified recombinant fragment hFXN90–195. This variant shows the hydrodynamic behavior expected for a monomeric globular domain. Circular dichroism, fluorescence, and NMR spectroscopies show that hFXN90–195 presents native-like secondary and tertiary structure. However, chemical and temperature induced denaturation show that CTR truncation significantly destabilizes the overall hFXN fold. Accordingly, limited proteolysis experiments suggest that the native-state dynamics of hFXN90–195 and hFXN90–210 are indeed different, being the former form much more sensitive to the protease at specific sites. The overall folding dynamics of hFXN fold was further explored with structure-based protein folding simulations. These suggest that the native ensemble of hFXN can be decomposed in at least two substates, one with consolidation of the CTR and the other without consolidation of the CTR. Explicit-solvent all atom simulations identify some of the proteolytic target sites as flexible regions of the protein. We propose that the local unfolding of CTR may be a critical step for the global unfolding of hFXN, and that modulation of the CTR interactions may strongly affect hFXN physiological function. PMID:23049850

  20. Patterns of indirect protein interactions suggest a spatial organization to metabolism.

    PubMed

    Pérez-Bercoff, Åsa; McLysaght, Aoife; Conant, Gavin C

    2011-11-01

    It has long been believed that cells organize their cytoplasm so as to efficiently channel metabolites between sequential enzymes. This metabolic channeling has the potential to yield higher metabolic fluxes as well as better regulatory control over metabolism. One mechanism for achieving such channeling is to ensure that sequential enzymes in a pathway are physically close to each other in the cell. We present evidence that indirect protein interactions between related enzymes represent a global mechanism for achieving metabolic channeling; the intuition being that protein interactions between enzymes and non-enzymatic mediator proteins are a powerful means of physically associating enzymes in a modular fashion. By analyzing the metabolic and protein-protein interactions networks of Escherichia coli, yeast and humans, we are able to show that all three species have many more indirect protein interactions linking enzymes that share metabolites than would be expected by chance. Moreover, these interactions are distributed non-randomly in the metabolic network. Our analyses in yeast and E. coli show that reactions possessing such interactions also show higher flux than do those lacking them. On the basis of these observations, we suggest that an important role of protein interactions with mediator proteins is to contribute to the spatial organization of the cell. This hypothesis is supported by the fact that these mediator proteins are also enriched with annotations related to signal transduction, a system where scaffolding proteins are known to limit cross-talk by controlling spatial localization.

  1. Metabolic syndrome and C-reactive protein in bank employees

    PubMed Central

    Cattafesta, Monica; Bissoli, Nazaré Souza; Salaroli, Luciane Bresciani

    2016-01-01

    Background The ultrasensitive C-reactive protein (us-CRP) is used for the diagnosis of cardiovascular disease, but it is not well described as a marker for the diagnosis of metabolic syndrome (MS). Methods An observational and transversal study of bank employees evaluated anthropometric, hemodynamic, and biochemical data. CRP values were determined using commercial kits from Roche Diagnostics Ltd, and MS criteria were analyzed according to National Cholesterol Education Program’s – Adult Treatment Panel III (NCEP/ATP III). Results A total of 88 individuals had MS, and 77.3% (n=68) of these showed alterations of us-CRP (P=0.0001, confidence interval [CI] 0.11–0.34). Individuals with MS had higher mean values of us-CRP in global measures (P=0.0001) and stratified by sex (P=0.004) than individuals without the syndrome. This marker exhibited significant differences with varying criteria for MS, such as waist circumference (P=0.0001), triglycerides (P=0.002), and diastolic blood pressure (P=0.007), and the highest levels of us-CRP were found in individuals with more MS criteria. Conclusion us-CRP was strongly associated with the presence of MS and MS criteria in this group of workers. us-CRP is a useful and effective marker for identifying the development of MS and may be used as a reference in routine care. PMID:27274294

  2. Effect of SO/sub 2/ on light modulation of plant metabolism. Progress report

    SciTech Connect

    Anderson, L.E.

    1985-01-01

    This progress report briefly notes conclusions of work done on SO/sub 2/ effect on light modulation of plant metabolism. Conclusions include: effect of light activation on kinetic parameters of fructosebisphosphatase - for this enzyme K/sub m/ decreases and V/sub max/ increases as a result of light activation; and the effect of sulfite and arsenite on light activation in 2 Pisum cultivars - the differences in sensitivity to SO/sub 2/ is directly reflected in differences in a thylakoid bound factor (LEM) to SO/sub 2/.

  3. Assessing the Metabolic Diversity of Streptococcus from a Protein Domain Point of View

    PubMed Central

    Koehorst, Jasper J.; Martins dos Santos, Vitor A. P.; Schaap, Peter J.

    2015-01-01

    Understanding the diversity and robustness of the metabolism of bacteria is fundamental for understanding how bacteria evolve and adapt to different environments. In this study, we characterised 121 Streptococcus strains and studied metabolic diversity from a protein domain perspective. Metabolic pathways were described in terms of the promiscuity of domains participating in metabolic pathways that were inferred to be functional. Promiscuity was defined by adapting existing measures based on domain abundance and versatility. The approach proved to be successful in capturing bacterial metabolic flexibility and species diversity, indicating that it can be described in terms of reuse and sharing functional domains in different proteins involved in metabolic activity. Additionally, we showed striking differences among metabolic organisation of the pathogenic serotype 2 Streptococcus suis and other strains. PMID:26366735

  4. The modulation of the symbiont/host interaction between Wolbachia pipientis and Aedes fluviatilis embryos by glycogen metabolism.

    PubMed

    da Rocha Fernandes, Mariana; Martins, Renato; Pessoa Costa, Evenilton; Pacidônio, Etiene Casagrande; Araujo de Abreu, Leonardo; da Silva Vaz, Itabajara; Moreira, Luciano A; da Fonseca, Rodrigo Nunes; Logullo, Carlos

    2014-01-01

    Wolbachia pipientis, a maternally transmitted bacterium that colonizes arthropods, may affect the general aspects of insect physiology, particularly reproduction. Wolbachia is a natural endosymbiont of Aedes fluviatilis, whose effects in embryogenesis and reproduction have not been addressed so far. In this context, we investigated the correlation between glucose metabolism and morphological alterations during A. fluviatilis embryo development in Wolbachia-positive (W+) and Wolbachia-negative (W-) mosquito strains. While both strains do not display significant morphological and larval hatching differences, larger differences were observed in hexokinase activity and glycogen contents during early and mid-stages of embryogenesis, respectively. To investigate if glycogen would be required for parasite-host interaction, we reduced Glycogen Synthase Kinase-3 (GSK-3) levels in adult females and their eggs by RNAi. GSK-3 knock-down leads to embryonic lethality, lower levels of glycogen and total protein and Wolbachia reduction. Therefore, our results suggest that the relationship between A. fluviatilis and Wolbachia may be modulated by glycogen metabolism.

  5. The Modulation of the Symbiont/Host Interaction between Wolbachia pipientis and Aedes fluviatilis Embryos by Glycogen Metabolism

    PubMed Central

    da Rocha Fernandes, Mariana; Martins, Renato; Pessoa Costa, Evenilton; Casagrande Pacidônio, Etiene; Araujo de Abreu, Leonardo; da Silva Vaz, Itabajara; Moreira, Luciano A.; da Fonseca, Rodrigo Nunes; Logullo, Carlos

    2014-01-01

    Wolbachia pipientis, a maternally transmitted bacterium that colonizes arthropods, may affect the general aspects of insect physiology, particularly reproduction. Wolbachia is a natural endosymbiont of Aedes fluviatilis, whose effects in embryogenesis and reproduction have not been addressed so far. In this context, we investigated the correlation between glucose metabolism and morphological alterations during A. fluviatilis embryo development in Wolbachia-positive (W+) and Wolbachia-negative (W−) mosquito strains. While both strains do not display significant morphological and larval hatching differences, larger differences were observed in hexokinase activity and glycogen contents during early and mid-stages of embryogenesis, respectively. To investigate if glycogen would be required for parasite-host interaction, we reduced Glycogen Synthase Kinase-3 (GSK-3) levels in adult females and their eggs by RNAi. GSK-3 knock-down leads to embryonic lethality, lower levels of glycogen and total protein and Wolbachia reduction. Therefore, our results suggest that the relationship between A. fluviatilis and Wolbachia may be modulated by glycogen metabolism. PMID:24926801

  6. Acetylation of glucokinase regulatory protein decreases glucose metabolism by suppressing glucokinase activity

    PubMed Central

    Park, Joo-Man; Kim, Tae-Hyun; Jo, Seong-Ho; Kim, Mi-Young; Ahn, Yong-Ho

    2015-01-01

    Glucokinase (GK), mainly expressed in the liver and pancreatic β-cells, is critical for maintaining glucose homeostasis. GK expression and kinase activity, respectively, are both modulated at the transcriptional and post-translational levels. Post-translationally, GK is regulated by binding the glucokinase regulatory protein (GKRP), resulting in GK retention in the nucleus and its inability to participate in cytosolic glycolysis. Although hepatic GKRP is known to be regulated by allosteric mechanisms, the precise details of modulation of GKRP activity, by post-translational modification, are not well known. Here, we demonstrate that GKRP is acetylated at Lys5 by the acetyltransferase p300. Acetylated GKRP is resistant to degradation by the ubiquitin-dependent proteasome pathway, suggesting that acetylation increases GKRP stability and binding to GK, further inhibiting GK nuclear export. Deacetylation of GKRP is effected by the NAD+-dependent, class III histone deacetylase SIRT2, which is inhibited by nicotinamide. Moreover, the livers of db/db obese, diabetic mice also show elevated GKRP acetylation, suggesting a broader, critical role in regulating blood glucose. Given that acetylated GKRP may affiliate with type-2 diabetes mellitus (T2DM), understanding the mechanism of GKRP acetylation in the liver could reveal novel targets within the GK-GKRP pathway, for treating T2DM and other metabolic pathologies. PMID:26620281

  7. Three Peptides from Soy Glycinin Modulate Glucose Metabolism in Human Hepatic HepG2 Cells.

    PubMed

    Lammi, Carmen; Zanoni, Chiara; Arnoldi, Anna

    2015-11-16

    Ile-Ala-Val-Pro-Gly-Glu-Val-Ala (IAVPGEVA), Ile-Ala-Val-Pro-Thr-Gly-Val-Ala (IAVPTGVA) and Leu-Pro-Tyr-Pro (LPYP), three peptides deriving from soy glycinin hydrolysis, are known to regulate cholesterol metabolism in human hepatic HepG2 cells. We have recently demonstrated that the mechanism of action involves the activation of adenosine monophosphate-activated protein kinase (AMPK). This fact suggested a potential activity of the same peptides on glucose metabolism that prompted us to also investigate this aspect in the same cells. After treatment with IAVPGEVA, IAVPTGVA and LPYP, HepG2 cells were analyzed using a combination of molecular techniques, including western blot analysis, glucose uptake experiments and fluorescence microscopy evaluation. The results showed that these peptides are indeed able to enhance the capacity of HepG2 cells to uptake glucose, via glucose transporter 1 GLUT1 and glucose transporter 4 GLUT4 activation, through the stimulation of protein kinase B Akt and adenosine monophosphate-activated protein kinase AMPK pathways, both involved in glucose metabolism.

  8. Ankyrin-repeat proteins from sponge symbionts modulate amoebal phagocytosis.

    PubMed

    Nguyen, Mary T H D; Liu, Michael; Thomas, Torsten

    2014-03-01

    Bacteria-eukaryote symbiosis occurs in all stages of evolution, from simple amoebae to mammals, and from facultative to obligate associations. Sponges are ancient metazoans that form intimate symbiotic interactions with complex communities of bacteria. The basic nutritional requirements of the sponge are in part satisfied by the phagocytosis of bacterial food particles from the surrounding water. How bacterial symbionts, which are permanently associated with the sponge, survive in the presence of phagocytic cells is largely unknown. Here, we present the discovery of a genomic fragment from an uncultured gamma-proteobacterial sponge symbiont that encodes for four proteins, whose closest known relatives are found in a sponge genome. Through recombinant approaches, we show that these four eukaryotic-like, ankyrin-repeat proteins (ARP) when expressed in Eschericha coli can modulate phagocytosis of amoebal cells and lead to accumulation of bacteria in the phagosome. Mechanistically, two ARPs appear to interfere with phagosome development in a similar way to reduced vacuole acidification, by blocking the fusion of the early phagosome with the lysosome and its digestive enzymes. Our results show that ARP from sponge symbionts can function to interfere with phagocytosis, and we postulate that this might be one mechanism by which symbionts can escape digestion in a sponge host.

  9. Metabolic Enzymes Enjoying New Partnerships as RNA-Binding Proteins.

    PubMed

    Castello, Alfredo; Hentze, Matthias W; Preiss, Thomas

    2015-12-01

    In the past century, few areas of biology advanced as much as our understanding of the pathways of intermediary metabolism. Initially considered unimportant in terms of gene regulation, crucial cellular fate changes, cell differentiation, or malignant transformation are now known to involve 'metabolic remodeling' with profound changes in the expression of many metabolic enzyme genes. This review focuses on the recent identification of RNA-binding activity of numerous metabolic enzymes. We discuss possible roles of this unexpected second activity in feedback gene regulation ('moonlighting') and/or in the control of enzymatic function. We also consider how metabolism-driven post-translational modifications could regulate enzyme-RNA interactions. Thus, RNA emerges as a new partner of metabolic enzymes with far-reaching possible consequences to be unraveled in the future.

  10. Metabolic Enzymes Enjoying New Partnerships as RNA-Binding Proteins

    PubMed Central

    Castello, Alfredo; Hentze, Matthias W.; Preiss, Thomas

    2015-01-01

    In the past century, few areas of biology advanced as much as our understanding of the pathways of intermediary metabolism. Initially considered unimportant in terms of gene regulation, crucial cellular fate changes, cell differentiation, or malignant transformation are now known to involve ‘metabolic remodeling’ with profound changes in the expression of many metabolic enzyme genes. This review focuses on the recent identification of RNA-binding activity of numerous metabolic enzymes. We discuss possible roles of this unexpected second activity in feedback gene regulation (‘moonlighting’) and/or in the control of enzymatic function. We also consider how metabolism-driven post-translational modifications could regulate enzyme–RNA interactions. Thus, RNA emerges as a new partner of metabolic enzymes with far-reaching possible consequences to be unraveled in the future. PMID:26520658

  11. Simulation of modulated protein crystal structure and diffraction data in a supercell and in superspace.

    PubMed

    Lovelace, Jeffrey J; Simone, Peter D; Petříček, Václav; Borgstahl, Gloria E O

    2013-06-01

    The toolbox for computational protein crystallography is full of easy-to-use applications for the routine solution and refinement of periodic diffraction data sets and protein structures. There is a gap in the available software when it comes to aperiodic crystallographic data. Current protein crystallography software cannot handle modulated data, and small-molecule software for aperiodic crystallography cannot work with protein structures. To adapt software for modulated protein data requires training data to test and debug the changed software. Thus, a comprehensive training data set consisting of atomic positions with associated modulation functions and the modulated structure factors packaged as both a three-dimensional supercell and as a modulated structure in (3+1)D superspace has been created. The (3+1)D data were imported into Jana2006; this is the first time that this has been performed for protein data.

  12. Metabolic Turnover of Synaptic Proteins: Kinetics, Interdependencies and Implications for Synaptic Maintenance

    PubMed Central

    Cohen, Laurie D.; Zuchman, Rina; Sorokina, Oksana; Müller, Anke; Dieterich, Daniela C.; Armstrong, J. Douglas; Ziv, Tamar; Ziv, Noam E.

    2013-01-01

    Chemical synapses contain multitudes of proteins, which in common with all proteins, have finite lifetimes and therefore need to be continuously replaced. Given the huge numbers of synaptic connections typical neurons form, the demand to maintain the protein contents of these connections might be expected to place considerable metabolic demands on each neuron. Moreover, synaptic proteostasis might differ according to distance from global protein synthesis sites, the availability of distributed protein synthesis facilities, trafficking rates and synaptic protein dynamics. To date, the turnover kinetics of synaptic proteins have not been studied or analyzed systematically, and thus metabolic demands or the aforementioned relationships remain largely unknown. In the current study we used dynamic Stable Isotope Labeling with Amino acids in Cell culture (SILAC), mass spectrometry (MS), Fluorescent Non–Canonical Amino acid Tagging (FUNCAT), quantitative immunohistochemistry and bioinformatics to systematically measure the metabolic half-lives of hundreds of synaptic proteins, examine how these depend on their pre/postsynaptic affiliation or their association with particular molecular complexes, and assess the metabolic load of synaptic proteostasis. We found that nearly all synaptic proteins identified here exhibited half-lifetimes in the range of 2–5 days. Unexpectedly, metabolic turnover rates were not significantly different for presynaptic and postsynaptic proteins, or for proteins for which mRNAs are consistently found in dendrites. Some functionally or structurally related proteins exhibited very similar turnover rates, indicating that their biogenesis and degradation might be coupled, a possibility further supported by bioinformatics-based analyses. The relatively low turnover rates measured here (∼0.7% of synaptic protein content per hour) are in good agreement with imaging-based studies of synaptic protein trafficking, yet indicate that the metabolic load

  13. PIPE: a protein-protein interaction passage extraction module for BioCreative challenge.

    PubMed

    Chang, Yung-Chun; Chu, Chun-Han; Su, Yu-Chen; Chen, Chien Chin; Hsu, Wen-Lian

    2016-01-01

    Identifying the interactions between proteins mentioned in biomedical literatures is one of the frequently discussed topics of text mining in the life science field. In this article, we propose PIPE, an interaction pattern generation module used in the Collaborative Biocurator Assistant Task at BioCreative V (http://www.biocreative.org/) to capture frequent protein-protein interaction (PPI) patterns within text. We also present an interaction pattern tree (IPT) kernel method that integrates the PPI patterns with convolution tree kernel (CTK) to extract PPIs. Methods were evaluated on LLL, IEPA, HPRD50, AIMed and BioInfer corpora using cross-validation, cross-learning and cross-corpus evaluation. Empirical evaluations demonstrate that our method is effective and outperforms several well-known PPI extraction methods. DATABASE URL. PMID:27524807

  14. DNA methylase and demethylase activities are modulated by one-carbon metabolism in Alzheimer's disease models.

    PubMed

    Fuso, Andrea; Nicolia, Vincenzina; Cavallaro, Rosaria A; Scarpa, Sigfrido

    2011-03-01

    Late-onset Alzheimer's disease seems to be a multi-factorial disease with both genetic and non-genetic, environmental, possible causes. Recently, epigenomics is achieving a major role in Alzheimer's research due to its involvement in different molecular pathways leading to neurodegeneration. Among the different epigenetic modifications, DNA methylation is one of the most relevant to the disease. We previously demonstrated that presenilin1 (PSEN1), a gene involved in amyloidogenesis, is modulated by DNA methylation in neuroblastoma cells and Alzheimer's mice in an experimental model of nutritionally altered one-carbon metabolism. This alteration, obtained by nutritional deficiency of B vitamins (folate, B12 and B6) hampered S-adenosylmethionine (SAM)-dependent methylation reactions. The aim of the present paper was to investigate the regulation of DNA methylation machinery in response to hypomethylating (B vitamin deficiency) and hypermethylating (SAM supplementation) alterations of the one-carbon metabolism. We found that DNA methylases (DNMT1, 3a and 3b) and a putative demethylase (MBD2) were differently modulated, in line with the previously observed changes of PSEN1 methylation pattern in the same experimental conditions.

  15. Structure-based design of small-molecule protein-protein interaction modulators: the story so far.

    PubMed

    Falchi, Federico; Caporuscio, Fabiana; Recanatini, Maurizio

    2014-03-01

    As the pivotal role of protein-protein interactions in cell growth, transcriptional activity, intracellular trafficking, signal transduction and pathological conditions has been assessed, experimental and in silico strategies have been developed to design protein-protein interaction modulators. State-of-the-art structure-based design methods, mainly pharmacophore modeling and docking, which have succeeded in the identification of enzyme inhibitors, receptor agonists and antagonists, and new tools specifically conceived to target protein-protein interfaces (e.g., hot-spot and druggable pocket prediction methods) have been applied in the search for small-molecule protein-protein interaction modulators. Many successful applications of structure-based design approaches that, despite the challenge of targeting protein-protein interfaces with small molecules, have led to the identification of micromolar and submicromolar hits are reviewed here.

  16. Protein engineering for metabolic engineering: Current and next-generation tools

    SciTech Connect

    Marcheschi, RJ; Gronenberg, LS; Liao, JC

    2013-04-16

    Protein engineering in the context of metabolic engineering is increasingly important to the field of industrial biotechnology. As the demand for biologically produced food, fuels, chemicals, food additives, and pharmaceuticals continues to grow, the ability to design and modify proteins to accomplish new functions will be required to meet the high productivity demands for the metabolism of engineered organisms. We review advances in selecting, modeling, and engineering proteins to improve or alter their activity. Some of the methods have only recently been developed for general use and are just beginning to find greater application in the metabolic engineering community. We also discuss methods of generating random and targeted diversity in proteins to generate mutant libraries for analysis. Recent uses of these techniques to alter cofactor use; produce non-natural amino acids, alcohols, and carboxylic acids; and alter organism phenotypes are presented and discussed as examples of the successful engineering of proteins for metabolic engineering purposes.

  17. Protein engineering for metabolic engineering: current and next-generation tools

    PubMed Central

    Marcheschi, Ryan J.; Gronenberg, Luisa S.; Liao, James C.

    2014-01-01

    Protein engineering in the context of metabolic engineering is increasingly important to the field of industrial biotechnology. As the demand for biologically-produced food, fuels, chemicals, food additives, and pharmaceuticals continues to grow, the ability to design and modify proteins to accomplish new functions will be required to meet the high productivity demands for the metabolism of engineered organisms. This article reviews advances of selecting, modeling, and engineering proteins to improve or alter their activity. Some of the methods have only recently been developed for general use and are just beginning to find greater application in the metabolic engineering community. We also discuss methods of generating random and targeted diversity in proteins to generate mutant libraries for analysis. Recent uses of these techniques to alter cofactor use, produce non-natural amino acids, alcohols, and carboxylic acids, and alter organism phenotypes are presented and discussed as examples of the successful engineering of proteins for metabolic engineering purposes. PMID:23589443

  18. Deep proteomics of mouse skeletal muscle enables quantitation of protein isoforms, metabolic pathways, and transcription factors.

    PubMed

    Deshmukh, Atul S; Murgia, Marta; Nagaraj, Nagarjuna; Treebak, Jonas T; Cox, Jürgen; Mann, Matthias

    2015-04-01

    Skeletal muscle constitutes 40% of individual body mass and plays vital roles in locomotion and whole-body metabolism. Proteomics of skeletal muscle is challenging because of highly abundant contractile proteins that interfere with detection of regulatory proteins. Using a state-of-the art MS workflow and a strategy to map identifications from the C2C12 cell line model to tissues, we identified a total of 10,218 proteins, including skeletal muscle specific transcription factors like myod1 and myogenin and circadian clock proteins. We obtain absolute abundances for proteins expressed in a muscle cell line and skeletal muscle, which should serve as a valuable resource. Quantitation of protein isoforms of glucose uptake signaling pathways and in glucose and lipid metabolic pathways provides a detailed metabolic map of the cell line compared with tissue. This revealed unexpectedly complex regulation of AMP-activated protein kinase and insulin signaling in muscle tissue at the level of enzyme isoforms.

  19. Deep Proteomics of Mouse Skeletal Muscle Enables Quantitation of Protein Isoforms, Metabolic Pathways, and Transcription Factors*

    PubMed Central

    Deshmukh, Atul S.; Murgia, Marta; Nagaraj, Nagarjuna; Treebak, Jonas T.; Cox, Jürgen; Mann, Matthias

    2015-01-01

    Skeletal muscle constitutes 40% of individual body mass and plays vital roles in locomotion and whole-body metabolism. Proteomics of skeletal muscle is challenging because of highly abundant contractile proteins that interfere with detection of regulatory proteins. Using a state-of-the art MS workflow and a strategy to map identifications from the C2C12 cell line model to tissues, we identified a total of 10,218 proteins, including skeletal muscle specific transcription factors like myod1 and myogenin and circadian clock proteins. We obtain absolute abundances for proteins expressed in a muscle cell line and skeletal muscle, which should serve as a valuable resource. Quantitation of protein isoforms of glucose uptake signaling pathways and in glucose and lipid metabolic pathways provides a detailed metabolic map of the cell line compared with tissue. This revealed unexpectedly complex regulation of AMP-activated protein kinase and insulin signaling in muscle tissue at the level of enzyme isoforms. PMID:25616865

  20. Gut epithelial inducible heat-shock proteins and their modulation by diet and the microbiota.

    PubMed

    Arnal, Marie-Edith; Lallès, Jean-Paul

    2016-03-01

    The epidemic of metabolic diseases has raised questions about the interplay between the human diet and the gut and its microbiota. The gut has two vital roles: nutrient absorption and intestinal barrier function. Gut barrier defects are involved in many diseases. Excess energy intake disturbs the gut microbiota and favors body entry of microbial compounds that stimulate chronic metabolic inflammation. In this context, the natural defense mechanisms of gut epithelial cells and the potential to boost them nutritionally warrant further study. One such important defense system is the activation of inducible heat-shock proteins (iHSPs) which protect the gut epithelium against oxidative stress and inflammation. Importantly, various microbial components can induce the expression of iHSPs. This review examines gut epithelial iHSPs as the main targets of microbial signals and nutrients and presents data on diseases involving disturbances of gut epithelial iHSPs. In addition, a broad literature analysis of dietary modulation of gut epithelial iHSPs is provided. Future research aims should include the identification of gut microbes that can optimize gut-protective iHSPs and the evaluation of iHSP-mediated health benefits of nutrients and food components.

  1. A computational analysis of protein interactions in metabolic networks reveals novel enzyme pairs potentially involved in metabolic channeling.

    PubMed

    Huthmacher, Carola; Gille, Christoph; Holzhütter, Hermann-Georg

    2008-06-01

    Protein-protein interactions are operative at almost every level of cell structure and function as, for example, formation of sub-cellular organelles, packaging of chromatin, muscle contraction, signal transduction, and regulation of gene expression. Public databases of reported protein-protein interactions comprise hundreds of thousands interactions, and this number is steadily growing. Elucidating the implications of protein-protein interactions for the regulation of the underlying cellular or extra-cellular reaction network remains a great challenge for computational biochemistry. In this work, we have undertaken a systematic and comprehensive computational analysis of reported enzyme-enzyme interactions in the metabolic networks of the model organisms Escherichia coli and Saccharomyces cerevisiae. We grouped all enzyme pairs according to the topological distance that the catalyzed reactions have in the metabolic network and performed a statistical analysis of reported enzyme-enzyme interactions within these groups. We found a higher frequency of reported enzyme-enzyme interactions within the group of enzymes catalyzing reactions that are adjacent in the network, i.e. sharing at least one metabolite. As some of these interacting enzymes have already been implicated in metabolic channeling our analysis may provide a useful screening for candidates of this phenomenon. To check for a possible regulatory role of interactions between enzymes catalyzing non-neighboring reactions, we determined potentially regulatory enzymes using connectivity in the network and absolute change of Gibbs free energy. Indeed a higher portion of reported interactions pertain to such potentially regulatory enzymes.

  2. Mammalian Mss51 is a skeletal muscle-specific gene modulating cellular metabolism

    PubMed Central

    Moyer, Adam L.; Wagner, Kathryn R.

    2015-01-01

    Background The transforming growth factor β (TGF-β) signaling pathways modulate skeletal muscle growth, regeneration, and cellular metabolism. Several recent gene expression studies have shown that inhibition of myostatin and TGF-β1 signaling consistently leads to a significant reduction in expression of Mss51, also named Zmynd17. The function of mammalian Mss51 is unknown although a putative homolog in yeast is a mitochondrial translational activator. Objective The objective of this work was to characterize mammalian Mss51. Methods Quantitative RT-PCR and immunoblot of subcellular fractionation were used to determine expression patterns and localization of Mss51. The CRISPR/Cas9 system was used to reduce expression of Mss51 in C2C12 myoblasts and the function of Mss51 was evaluated in assays of proliferation, differentiation and cellular metabolism. Results Mss51 was predominantly expressed in skeletal muscle and in those muscles dominated by fast-twitch fibers. In vitro, its expression was upregulated upon differentiation of C2C12 myoblasts into myotubes. Expression of Mss51 was modulated in response to altered TGF-β family signaling. In human muscle, Mss51 localized to the mitochondria. Its genetic disruption resulted in increased levels of cellular ATP, β-oxidation, glycolysis, and oxidative phosphorylation. Conclusions Mss51 is a novel, skeletal muscle-specific gene and a key target of myostatin and TGF-β1 signaling. Unlike myostatin, TGF-β1 and IGF-1, Mss51 does not regulate myoblast proliferation or differentiation. Rather, Mss51 appears to be one of the effectors of these growth factors on metabolic processes including fatty acid oxidation, glycolysis and oxidative phosphorylation. PMID:26634192

  3. Relevance of drug metabolizing enzyme activity modulation by tea polyphenols in the inhibition of esophageal tumorigenesis.

    PubMed

    Maliakal, Pius; Sankpal, Umesh T; Basha, Riyaz; Maliakal, Cima; Ledford, Andrea; Wanwimolruk, Sompon

    2011-09-01

    Tea is a popular, socially accepted, drink that is enjoyed by millions of people. A growing body of evidence suggests that moderate consumption of tea may protect against several forms of cancer. It is also known that bioactivation of precarcinogens and detoxification of ultimate carcinogens is carried out mainly by drug metabolizing enzymes such as cytochrome P450 (CYP). The present study investigates the effect of tea consumption on modulating CYP and phase II conjugating enzymes, and their association in the chemopreventive effect against esophageal tumorigenesis using both in vitro and in vivo techniques. Female Wistar rats were given aqueous solutions (2% w/v) of six different teas, standard green tea extract (GTE) (0.5% w/v), and dandelion tea (2% w/v) as the sole source of fluid for two weeks prior to and during the entire period of tumour induction (12 weeks). Animals were gavaged with 0.5 mg/kg N-nitrosomethylbenzylamine (NMBA) twice weekly for 12 weeks for esophageal tumor induction and the activities of different CYP isoforms and phase II enzymes were determined in the liver microsomes or cytosols. GTE, green tea and Dandelion tea caused decrease in tumour multiplication, tumour size and tumour volume; however, none of these tea preparations altered tumour incidence. No appreciable changes in drug metabolizing enzyme activity were observed in the treatment groups. Thus, the modulations in the activities of CYP 1A1/ 1A2 and CYP2E enzymes, by pre-treatment with green and dandelion teas, observed in our earlier experiments, seem to be compensated by the tumor inducing agent, NMBA. The balance between phase I carcinogen-activating enzymes and phase II detoxifying enzymes could be important in determining the risk of developing chemically-induced cancer and the present study in conjunction with the previous observations suggest a possible role of drug metabolizing enzymes in the anticancer effect of tea.

  4. Salmonella Modulates Metabolism During Growth under Conditions that Induce Expression of Virulence Genes

    SciTech Connect

    Kim, Young-Mo; Schmidt, Brian; Kidwai, Afshan S.; Jones, Marcus B.; Deatherage, Brooke L.; Brewer, Heather M.; Mitchell, Hugh D.; Palsson, Bernhard O.; McDermott, Jason E.; Heffron, Fred; Smith, Richard D.; Peterson, Scott N.; Ansong, Charles; Hyduke, Daniel R.; Metz, Thomas O.; Adkins, Joshua N.

    2013-04-05

    Salmonella enterica serovar Typhimurium (S. Typhimurium) is a facultative pathogen that uses complex mechanisms to invade and proliferate within mammalian host cells. To investigate possible contributions of metabolic processes in S. Typhimurium grown under conditions known to induce expression of virulence genes, we used a metabolomics-driven systems biology approach coupled with genome scale modeling. First, we identified distinct metabolite profiles associated with bacteria grown in either rich or virulence-inducing media and report the most comprehensive coverage of the S. Typhimurium metabolome to date. Second, we applied an omics-informed genome scale modeling analysis of the functional consequences of adaptive alterations in S. Typhimurium metabolism during growth under our conditions. Excitingly, we observed possible sequestration of metabolites recently suggested to have immune modulating roles. Modeling efforts highlighted a decreased cellular capability to both produce and utilize intracellular amino acids during stationary phase culture in virulence conditions, despite significant abundance increases for these molecules as observed by our metabolomics measurements. Model-guided analysis suggested that alterations in metabolism prioritized other activities necessary for pathogenesis instead, such as lipopolysaccharide biosynthesis.

  5. Dolomite supplementation improves bone metabolism through modulation of calcium-regulating hormone secretion in ovariectomized rats.

    PubMed

    Mizoguchi, Toshihide; Nagasawa, Sakae; Takahashi, Naoyuki; Yagasaki, Hiroshi; Ito, Michio

    2005-01-01

    Dolomite, a mineral composed of calcium magnesium carbonate (CaMg (CO3)2), is used as a food supplement that supplies calcium and magnesium. However, the effect of magnesium supplementation on bone metabolism in patients with osteoporosis is a matter of controversy. We examined the effects of daily supplementation with dolomite on calcium metabolism in ovariectomized (OVX) rats. Dolomite was administered daily to OVX rats for 9 weeks. The same amount of magnesium chloride as that supplied by the dolomite was given to OVX rats as a positive control. Histological examination revealed that ovariectomy decreased trabecular bone and increased adipose tissues in the femoral metaphysis. Dolomite or magnesium supplementation failed to improve these bone histological features. Calcium content in the femora was decreased in OVX rats. Neither calcium nor magnesium content in the femora in OVX rats was significantly increased by dolomite or magnesium administration. Urinary deoxypyridinoline excretion was significantly increased in OVX rats, and was not affected by the magnesium supplementation. Serum concentrations of magnesium were increased, and those of calcium were decreased, in OVX rats supplemented with dolomite or magnesium. However, there was a tendency toward decreased parathyroid hormone secretion and increased calcitonin secretion in OVX rats supplemented with dolomite or magnesium. Serum 1,25-dihydroxyvitamin D(3) and osteocalcin levels were significantly increased in the supplemented OVX rats. These results suggest that increased magnesium intake improves calcium metabolism in favor of increasing bone formation, through the modulation of calcium-regulating hormone secretion.

  6. Dolomite supplementation improves bone metabolism through modulation of calcium-regulating hormone secretion in ovariectomized rats.

    PubMed

    Mizoguchi, Toshihide; Nagasawa, Sakae; Takahashi, Naoyuki; Yagasaki, Hiroshi; Ito, Michio

    2005-01-01

    Dolomite, a mineral composed of calcium magnesium carbonate (CaMg (CO3)2), is used as a food supplement that supplies calcium and magnesium. However, the effect of magnesium supplementation on bone metabolism in patients with osteoporosis is a matter of controversy. We examined the effects of daily supplementation with dolomite on calcium metabolism in ovariectomized (OVX) rats. Dolomite was administered daily to OVX rats for 9 weeks. The same amount of magnesium chloride as that supplied by the dolomite was given to OVX rats as a positive control. Histological examination revealed that ovariectomy decreased trabecular bone and increased adipose tissues in the femoral metaphysis. Dolomite or magnesium supplementation failed to improve these bone histological features. Calcium content in the femora was decreased in OVX rats. Neither calcium nor magnesium content in the femora in OVX rats was significantly increased by dolomite or magnesium administration. Urinary deoxypyridinoline excretion was significantly increased in OVX rats, and was not affected by the magnesium supplementation. Serum concentrations of magnesium were increased, and those of calcium were decreased, in OVX rats supplemented with dolomite or magnesium. However, there was a tendency toward decreased parathyroid hormone secretion and increased calcitonin secretion in OVX rats supplemented with dolomite or magnesium. Serum 1,25-dihydroxyvitamin D(3) and osteocalcin levels were significantly increased in the supplemented OVX rats. These results suggest that increased magnesium intake improves calcium metabolism in favor of increasing bone formation, through the modulation of calcium-regulating hormone secretion. PMID:15750692

  7. Overcoming the metabolic burden of protein secretion in Schizosaccharomyces pombe--a quantitative approach using 13C-based metabolic flux analysis.

    PubMed

    Klein, Tobias; Lange, Sabrina; Wilhelm, Nadine; Bureik, Matthias; Yang, Tae-Hoon; Heinzle, Elmar; Schneider, Konstantin

    2014-01-01

    Protein secretion in yeast is generally associated with a burden to cellular metabolism. To investigate this metabolic burden in Schizosaccharomyces pombe, we constructed a set of strains secreting the model protein maltase in different amounts. We quantified the influence of protein secretion on the metabolism applying (13)C-based metabolic flux analysis in chemostat cultures. Analysis of the macromolecular biomass composition revealed an increase in cellular lipid content at elevated levels of protein secretion and we observed altered metabolic fluxes in the pentose phosphate pathway, the TCA cycle, and around the pyruvate node including mitochondrial NADPH supply. Supplementing acetate to glucose or glycerol minimal media was found to improve protein secretion, accompanied by an increased cellular lipid content and carbon flux through the TCA cycle as well as increased mitochondrial NADPH production. Thus, systematic metabolic analyses can assist in identifying factors limiting protein secretion and in deriving strategies to overcome these limitations.

  8. A Small Protein Associated with Fungal Energy Metabolism Affects the Virulence of Cryptococcus neoformans in Mammals.

    PubMed

    McClelland, Erin E; Ramagopal, Udupi A; Rivera, Johanna; Cox, James; Nakouzi, Antonio; Prabu, Moses M; Almo, Steven C; Casadevall, Arturo

    2016-09-01

    The pathogenic yeast Cryptococcus neoformans causes cryptococcosis, a life-threatening fungal disease. C. neoformans has multiple virulence mechanisms that are non-host specific, induce damage and interfere with immune clearance. Microarray analysis of C. neoformans strains serially passaged in mice associated a small gene (CNAG_02591) with virulence. This gene, hereafter identified as HVA1 (hypervirulence-associated protein 1), encodes a protein that has homologs of unknown function in plant and animal fungi, consistent with a conserved mechanism. Expression of HVA1 was negatively correlated with virulence and was reduced in vitro and in vivo in both mouse- and Galleria-passaged strains of C. neoformans. Phenotypic analysis in hva1Δ and hva1Δ+HVA1 strains revealed no significant differences in established virulence factors. Mice infected intravenously with the hva1Δ strain had higher fungal burden in the spleen and brain, but lower fungal burden in the lungs, and died faster than mice infected with H99W or the hva1Δ+HVA1 strain. Metabolomics analysis demonstrated a general increase in all amino acids measured in the disrupted strain and a block in the TCA cycle at isocitrate dehydrogenase, possibly due to alterations in the nicotinamide cofactor pool. Macrophage fungal burden experiments recapitulated the mouse hypervirulent phenotype of the hva1Δ strain only in the presence of exogenous NADPH. The crystal structure of the Hva1 protein was solved, and a comparison of structurally similar proteins correlated with the metabolomics data and potential interactions with NADPH. We report a new gene that modulates virulence through a mechanism associated with changes in fungal metabolism. PMID:27583447

  9. A Small Protein Associated with Fungal Energy Metabolism Affects the Virulence of Cryptococcus neoformans in Mammals

    PubMed Central

    Cox, James; Nakouzi, Antonio; Prabu, Moses M.; Almo, Steven C.

    2016-01-01

    The pathogenic yeast Cryptococcus neoformans causes cryptococcosis, a life-threatening fungal disease. C. neoformans has multiple virulence mechanisms that are non-host specific, induce damage and interfere with immune clearance. Microarray analysis of C. neoformans strains serially passaged in mice associated a small gene (CNAG_02591) with virulence. This gene, hereafter identified as HVA1 (hypervirulence-associated protein 1), encodes a protein that has homologs of unknown function in plant and animal fungi, consistent with a conserved mechanism. Expression of HVA1 was negatively correlated with virulence and was reduced in vitro and in vivo in both mouse- and Galleria-passaged strains of C. neoformans. Phenotypic analysis in hva1Δ and hva1Δ+HVA1 strains revealed no significant differences in established virulence factors. Mice infected intravenously with the hva1Δ strain had higher fungal burden in the spleen and brain, but lower fungal burden in the lungs, and died faster than mice infected with H99W or the hva1Δ+HVA1 strain. Metabolomics analysis demonstrated a general increase in all amino acids measured in the disrupted strain and a block in the TCA cycle at isocitrate dehydrogenase, possibly due to alterations in the nicotinamide cofactor pool. Macrophage fungal burden experiments recapitulated the mouse hypervirulent phenotype of the hva1Δ strain only in the presence of exogenous NADPH. The crystal structure of the Hva1 protein was solved, and a comparison of structurally similar proteins correlated with the metabolomics data and potential interactions with NADPH. We report a new gene that modulates virulence through a mechanism associated with changes in fungal metabolism. PMID:27583447

  10. The origin of modern metabolic networks inferred from phylogenomic analysis of protein architecture.

    PubMed

    Caetano-Anollés, Gustavo; Kim, Hee Shin; Mittenthal, Jay E

    2007-05-29

    Metabolism represents a complex collection of enzymatic reactions and transport processes that convert metabolites into molecules capable of supporting cellular life. Here we explore the origins and evolution of modern metabolism. Using phylogenomic information linked to the structure of metabolic enzymes, we sort out recruitment processes and discover that most enzymatic activities were associated with the nine most ancient and widely distributed protein fold architectures. An analysis of newly discovered functions showed enzymatic diversification occurred early, during the onset of the modern protein world. Most importantly, phylogenetic reconstruction exercises and other evidence suggest strongly that metabolism originated in enzymes with the P-loop hydrolase fold in nucleotide metabolism, probably in pathways linked to the purine metabolic subnetwork. Consequently, the first enzymatic takeover of an ancient biochemistry or prebiotic chemistry was related to the synthesis of nucleotides for the RNA world. PMID:17517598

  11. Modulation of nociceptive ion channels and receptors via protein-protein interactions: implications for pain relief

    PubMed Central

    Rouwette, Tom; Avenali, Luca; Sondermann, Julia; Narayanan, Pratibha; Gomez-Varela, David; Schmidt, Manuela

    2015-01-01

    In the last 2 decades biomedical research has provided great insights into the molecular signatures underlying painful conditions. However, chronic pain still imposes substantial challenges to researchers, clinicians and patients alike. Under pathological conditions, pain therapeutics often lack efficacy and exhibit only minimal safety profiles, which can be largely attributed to the targeting of molecules with key physiological functions throughout the body. In light of these difficulties, the identification of molecules and associated protein complexes specifically involved in chronic pain states is of paramount importance for designing selective interventions. Ion channels and receptors represent primary targets, as they critically shape nociceptive signaling from the periphery to the brain. Moreover, their function requires tight control, which is usually implemented by protein-protein interactions (PPIs). Indeed, manipulation of such PPIs entails the modulation of ion channel activity with widespread implications for influencing nociceptive signaling in a more specific way. In this review, we highlight recent advances in modulating ion channels and receptors via their PPI networks in the pursuit of relieving chronic pain. Moreover, we critically discuss the potential of targeting PPIs for developing novel pain therapies exhibiting higher efficacy and improved safety profiles. PMID:26039491

  12. Mitochondrial functions modulate neuroendocrine, metabolic, inflammatory, and transcriptional responses to acute psychological stress

    PubMed Central

    Picard, Martin; McManus, Meagan J.; Gray, Jason D.; Nasca, Carla; Moffat, Cynthia; Kopinski, Piotr K.; Seifert, Erin L.; McEwen, Bruce S.; Wallace, Douglas C.

    2015-01-01

    The experience of psychological stress triggers neuroendocrine, inflammatory, metabolic, and transcriptional perturbations that ultimately predispose to disease. However, the subcellular determinants of this integrated, multisystemic stress response have not been defined. Central to stress adaptation is cellular energetics, involving mitochondrial energy production and oxidative stress. We therefore hypothesized that abnormal mitochondrial functions would differentially modulate the organism’s multisystemic response to psychological stress. By mutating or deleting mitochondrial genes encoded in the mtDNA [NADH dehydrogenase 6 (ND6) and cytochrome c oxidase subunit I (COI)] or nuclear DNA [adenine nucleotide translocator 1 (ANT1) and nicotinamide nucleotide transhydrogenase (NNT)], we selectively impaired mitochondrial respiratory chain function, energy exchange, and mitochondrial redox balance in mice. The resulting impact on physiological reactivity and recovery from restraint stress were then characterized. We show that mitochondrial dysfunctions altered the hypothalamic–pituitary–adrenal axis, sympathetic adrenal–medullary activation and catecholamine levels, the inflammatory cytokine IL-6, circulating metabolites, and hippocampal gene expression responses to stress. Each mitochondrial defect generated a distinct whole-body stress-response signature. These results demonstrate the role of mitochondrial energetics and redox balance as modulators of key pathophysiological perturbations previously linked to disease. This work establishes mitochondria as stress-response modulators, with implications for understanding the mechanisms of stress pathophysiology and mitochondrial diseases. PMID:26627253

  13. Disposition and metabolism of LY2452473, a selective androgen receptor modulator, in humans.

    PubMed

    Yi, Ping; Rehmel, Jessica Fayer; Cassidy, Kenneth; Hadden, Chad; Campanale, Kristina; Patel, Nita; Johnson, Jason

    2012-12-01

    The disposition and metabolism of isopropyl N-[(2S)-7-cyano-4-(2-pyridylmethyl)-2,3-dihydro-1H-cyclopenta[b]indol-2-yl]carbamate (LY2452473; a selective androgen receptor modulator) in humans was characterized after a single 15-mg (100 μCi) oral dose of [¹⁴C]LY2452473 to six healthy male subjects. LY2452473 was absorbed rapidly (time to reach maximum plasma concentration for both LY2452473 and total radioactivity was 2-3 h) and cleared slowly (plasma terminal t(½) of 27 h for LY2452473 and 51 h for the total radioactivity). LY2452473 and metabolites S5 (acetylamine) and S12 (hydroxylation on the cyclopentene) were major circulating entities in plasma, accounting for approximately 42, 21, and 35% of the total radioactivity exposure, respectively, as calculated from relative area under the concentration versus time curves from zero to 48 h derived from the plasma radiochromatograms. The radioactive dose was almost completely recovered after 312 h with 47.9% of the dose eliminated in urine and 46.6% in feces. Minimal LY2452473 was detected in excreta, indicating that metabolic clearance was the main route of elimination. Multiple metabolic pathways were observed with no single metabolic pathway accounting for more than 30% of the dose in excreta. Metabolite S10 (a diol across the cyclopenta-indole linkage) was the largest excretory metabolite (approximately 14% of the dose). S10 displayed interesting chemical and chromatographic properties, undergoing conversion to the corresponding epoxide under acidic conditions and conversion back to the diol under neutral conditions. An in vitro phenotyping approach indicated that CYP3A4 was the largest contributor to LY2452473 depletion. PMID:22961682

  14. Modulation of PML protein expression regulates JCV infection

    SciTech Connect

    Gasparovic, Megan L.; Maginnis, Melissa S.; O'Hara, Bethany A.; Dugan, Aisling S.; Atwood, Walter J.

    2009-08-01

    JC virus (JCV) is a human polyomavirus that infects the majority of the human population worldwide. It is responsible for the fatal demyelinating disease Progressive Multifocal Leukoencephalopathy. JCV binds to cells using the serotonin receptor 5-HT{sub 2A}R and alpha(2-6)- or alpha(2-3)-linked sialic acid. It enters cells using clathrin-dependent endocytosis and traffics to the early endosome and possibly to the endoplasmic reticulum. Viral DNA is then delivered to the nucleus where transcription, replication, and assembly of progeny occur. We found that the early regulatory protein large T antigen accumulates in microdomains in the nucleus adjacent to ND-10 or PML domains. This observation prompted us to explore the role of these domains in JCV infection. We found that a reduction of nuclear PML enhanced virus infection and that an increase in nuclear PML reduced infection. Infection with JCV did not directly modulate nuclear levels of PML but our data indicate that a host response involving interferon beta is likely to restrict virus infection by increasing nuclear PML.

  15. Use of Designer G Protein-Coupled Receptors to Dissect Metabolic Pathways.

    PubMed

    Wess, Jürgen

    2016-09-01

    G protein-coupled receptors (GPCRs) regulate virtually all metabolic processes, including glucose and energy homeostasis. Recently, the use of designer GPCRs referred to as designer receptors exclusively activated by designer drug (DREADDs) has made it possible to dissect metabolically relevant GPCR signaling pathways in a temporally and spatially controlled fashion in vivo. PMID:27381463

  16. Modulation of a human lymphoblastoid B cell line by cyclic AMP. Ig secretion and phosphatidylcholine metabolism

    SciTech Connect

    Shearer, W.T.; Patke, C.L.; Gilliam, E.B.; Rosenblatt, H.M.; Barron, K.S.; Orson, F.M.

    1988-09-01

    A transformed human B cell line, LA350, was found to be sensitive to cAMP-elevating agents by responding with rapid (0 to 2 h) severalfold elevations of intracellular cAMP to treatment with cholera toxin, isobutylmethylxanthine (IBMX), forskolin, and dibutyryl cAMP (all p less than 0.001). These cAMP-elevating agents also produced significant inhibitions of subsequent (48 to 72 h) Ig secretion by the same B cells as measured by a reverse hemolytic plaque assay and an enzyme-linked immunoadsorbent assay for IgM (both p less than 0.001). PMA- and IBMX-treated cells were particularly responsive to the effects of cholera toxin, showing a doubling of cAMP content and profound decrease in Ig production (p less than 0.001). Because our previous studies had correlated activation of the metabolic turnover of the phosphatidylcholine (PC) fraction of membrane phospholipids with enhanced Ig secretion, we examined the sensitivity of PC metabolism to cAMP in control and PMA-stimulated cells. Formation of PC was found to be inhibited by forskolin and IBMX (both p less than 0.002) but breakdown of PC was stimulated (p less than 0.001). These findings imply that as the enzymatic products of PC, choline phosphate and diacylglycerol, are depleted due to the combined effects of cAMP upon synthesis and turnover of PC, there is a decrease in Ig secretion. Since diacylglycerol activates protein kinase C, it appears reasonable that Ig secretion is at least partially regulated by cAMP-responsive alterations in PC metabolism produced by protein kinase C-induced phosphorylation. We conclude that the early cAMP-sensitive changes in PC metabolism in this activated B cell line may signal for subsequent alterations in Ig secretion.

  17. Study of Stationary Phase Metabolism Via Isotopomer Analysis of Amino Acids from an Isolated Protein

    SciTech Connect

    Shaikh, AfshanS.; Tang, YinjieJ.; Mukhopadhyay, Aindrila; Martin, Hector Garcia; Gin, Jennifer; Benke, Peter; Keasling, Jay D.

    2009-09-14

    Microbial production of many commercially important secondary metabolites occurs during stationary phase, and methods to measure metabolic flux during this growth phase would be valuable. Metabolic flux analysis is often based on isotopomer information from proteinogenic amino acids. As such, flux analysis primarily reflects the metabolism pertinent to the growth phase during which most proteins are synthesized. To investigate central metabolism and amino acids synthesis activity during stationary phase, addition of fully 13C-labeled glucose followed by induction of green fluorescent protein (GFP) expression during stationary phase was used. Our results indicate that Escherichia coli was able to produce new proteins (i.e., GFP) in the stationary phase, and the amino acids in GFP were mostly from degraded proteins synthesized during the exponential growth phase. Among amino acid biosynthetic pathways, only those for serine, alanine, glutamate/glutamine, and aspartate/asparagine had significant activity during the stationary phase.

  18. Altered Arginine Metabolism in Cells Transfected with Human Wild-Type Beta Amyloid Precursor Protein (βAPP).

    PubMed

    Jęśko, Henryk; Wilkaniec, Anna; Cieślik, Magdalena; Hilgier, Wojciech; Gąssowska, Magdalena; Lukiw, Walter J; Adamczyk, Agata

    2016-01-01

    Alterations of enzymes linked to arginine metabolism have been recently implicated in Alzheimer's disease (AD). Despite strong association of arginine changes with nitric oxide (NO) pathway, the impact of amyloid β (Aβ) peptides on arginine degradation and re-synthesis is unknown. In the present study we compared expression levels of arginases (ARG1, ARG2), neuronal, endothelial and inducible NO synthase isoforms (NNOS, ENOS, INOS), enzymes that metabolize arginine or resynthesize it from citrulline and the levels of corresponding amino acids in rat pheochromocytoma (PC12) cells overexpressing human Aβ precursor protein (APPwt cells). Moreover, we investigated the changes in miRNAs responsible for modulation of arginine metabolism in AD brains. Real-time PCR analysis revealed in APPwt cells significant decreases of ARG1 and ARG2 which are responsible for lysing arginine into ornithine and urea; this reduction was followed by significantly lower enzyme activity. NNOS and ENOS mRNAs were elevated in APPwt cells while iNOS was undetectable in both cell lines. The expression of argininosuccinate synthase (ASS) that metabolizes citrulline was down-regulated without changes in argininosuccinate lyase (ASL). Ornithine decarboxylase (ODC), which decarboxylates ornithine to form putrescine was also reduced. Arginine, the substrate for both arginases and NOS, was unchanged in APPwt cells. However, citrulline concentration was significantly higher. Elevated miRNA-9 and miRNA-128a found in AD brain tissues might modulate the expression of ASS and NOS, respectively. Our results indicate that Aβ affects arginine metabolism and this influence might have important role in the pathomechanism of AD.

  19. Altered Arginine Metabolism in Cells Transfected with Human Wild-Type Beta Amyloid Precursor Protein (βAPP).

    PubMed

    Jęśko, Henryk; Wilkaniec, Anna; Cieślik, Magdalena; Hilgier, Wojciech; Gąssowska, Magdalena; Lukiw, Walter J; Adamczyk, Agata

    2016-01-01

    Alterations of enzymes linked to arginine metabolism have been recently implicated in Alzheimer's disease (AD). Despite strong association of arginine changes with nitric oxide (NO) pathway, the impact of amyloid β (Aβ) peptides on arginine degradation and re-synthesis is unknown. In the present study we compared expression levels of arginases (ARG1, ARG2), neuronal, endothelial and inducible NO synthase isoforms (NNOS, ENOS, INOS), enzymes that metabolize arginine or resynthesize it from citrulline and the levels of corresponding amino acids in rat pheochromocytoma (PC12) cells overexpressing human Aβ precursor protein (APPwt cells). Moreover, we investigated the changes in miRNAs responsible for modulation of arginine metabolism in AD brains. Real-time PCR analysis revealed in APPwt cells significant decreases of ARG1 and ARG2 which are responsible for lysing arginine into ornithine and urea; this reduction was followed by significantly lower enzyme activity. NNOS and ENOS mRNAs were elevated in APPwt cells while iNOS was undetectable in both cell lines. The expression of argininosuccinate synthase (ASS) that metabolizes citrulline was down-regulated without changes in argininosuccinate lyase (ASL). Ornithine decarboxylase (ODC), which decarboxylates ornithine to form putrescine was also reduced. Arginine, the substrate for both arginases and NOS, was unchanged in APPwt cells. However, citrulline concentration was significantly higher. Elevated miRNA-9 and miRNA-128a found in AD brain tissues might modulate the expression of ASS and NOS, respectively. Our results indicate that Aβ affects arginine metabolism and this influence might have important role in the pathomechanism of AD. PMID:26971935

  20. HIV-1 Vpr Modulates Macrophage Metabolic Pathways: A SILAC-Based Quantitative Analysis

    PubMed Central

    Barrero, Carlos A.; Datta, Prasun K.; Sen, Satarupa; Deshmane, Satish; Amini, Shohreh; Khalili, Kamel; Merali, Salim

    2013-01-01

    Human immunodeficiency virus type 1 encoded viral protein Vpr is essential for infection of macrophages by HIV-1. Furthermore, these macrophages are resistant to cell death and are viral reservoir. However, the impact of Vpr on the macrophage proteome is yet to be comprehended. The goal of the present study was to use a stable-isotope labeling by amino acids in cell culture (SILAC) coupled with mass spectrometry-based proteomics approach to characterize the Vpr response in macrophages. Cultured human monocytic cells, U937, were differentiated into macrophages and transduced with adenovirus construct harboring the Vpr gene. More than 600 proteins were quantified in SILAC coupled with LC-MS/MS approach, among which 136 were significantly altered upon Vpr overexpression in macrophages. Quantified proteins were selected and clustered by biological functions, pathway and network analysis using Ingenuity computational pathway analysis. The proteomic data illustrating increase in abundance of enzymes in the glycolytic pathway (pentose phosphate and pyruvate metabolism) was further validated by western blot analysis. In addition, the proteomic data demonstrate down regulation of some key mitochondrial enzymes such as glutamate dehydrogenase 2 (GLUD2), adenylate kinase 2 (AK2) and transketolase (TKT). Based on these observations we postulate that HIV-1 hijacks the macrophage glucose metabolism pathway via the Vpr-hypoxia inducible factor 1 alpha (HIF-1 alpha) axis to induce expression of hexokinase (HK), glucose-6-phosphate dehyrogenase (G6PD) and pyruvate kinase muscle type 2 (PKM2) that facilitates viral replication and biogenesis, and long-term survival of macrophages. Furthermore, dysregulation of mitochondrial glutamate metabolism in macrophages can contribute to neurodegeneration via neuroexcitotoxic mechanisms in the context of NeuroAIDS. PMID:23874603

  1. Changes in gravity affect gene expression, protein modulation and metabolite pools of arabidopsis

    NASA Astrophysics Data System (ADS)

    Hampp, R.; Martzivanou, M.; Maier, R. M.; Magel, E.

    ), we investigated samples from sounding rocket experiments (5 min μ g) and show increased transcript levels for signalling proteins. By means of 2-dimensional SDS polyacrylamide gelelectrophoresis, coupled to spot identification after tryptic digest (MALDI-TOF), we further show that metabolic short-term responses can be adjusted by protein phosphorylation/dephosphorylation. Changes in gene expression / protein modulation are mirrored by respective alterations in metabolite pools. (Supported by a grant from the Deutsches Zentrum für Luft- und Raumfahrt (DLR, 50WB0143)).

  2. Hepatic autophagy contributes to the metabolic response to dietary protein restriction.

    PubMed

    Henagan, Tara M; Laeger, Thomas; Navard, Alexandra M; Albarado, Diana; Noland, Robert C; Stadler, Krisztian; Elks, Carrie M; Burk, David; Morrison, Christopher D

    2016-06-01

    Autophagy is an essential cellular response which acts to release stored cellular substrates during nutrient restriction, and particularly plays a key role in the cellular response to amino acid restriction. However, there has been limited work testing whether the induction of autophagy is required for adaptive metabolic responses to dietary protein restriction in the whole animal. Here, we found that moderate dietary protein restriction led to a series of metabolic changes in rats, including increases in food intake and energy expenditure, the downregulation of hepatic fatty acid synthesis gene expression and reduced markers of hepatic mitochondrial number. Importantly, these effects were also associated with an induction of hepatic autophagy. To determine if the induction of autophagy contributes to these metabolic effects, we tested the metabolic response to dietary protein restriction in BCL2-AAA mice, which bear a genetic mutation that impairs autophagy induction. Interestingly, BCL2-AAA mice exhibit exaggerated responses in terms of both food intake and energy expenditure, whereas the effects of protein restriction on hepatic metabolism were significantly blunted. These data demonstrate that restriction of dietary protein is sufficient to trigger hepatic autophagy, and that disruption of autophagy significantly alters both hepatic and whole animal metabolic response to dietary protein restriction. PMID:27173459

  3. Lipid-mediated Protein-protein Interactions Modulate Respiration-driven ATP Synthesis

    PubMed Central

    Nilsson, Tobias; Lundin, Camilla Rydström; Nordlund, Gustav; Ädelroth, Pia; von Ballmoos, Christoph; Brzezinski, Peter

    2016-01-01

    Energy conversion in biological systems is underpinned by membrane-bound proton transporters that generate and maintain a proton electrochemical gradient across the membrane which used, e.g. for generation of ATP by the ATP synthase. Here, we have co-reconstituted the proton pump cytochrome bo3 (ubiquinol oxidase) together with ATP synthase in liposomes and studied the effect of changing the lipid composition on the ATP synthesis activity driven by proton pumping. We found that for 100 nm liposomes, containing 5 of each proteins, the ATP synthesis rates decreased significantly with increasing fractions of DOPA, DOPE, DOPG or cardiolipin added to liposomes made of DOPC; with e.g. 5% DOPG, we observed an almost 50% decrease in the ATP synthesis rate. However, upon increasing the average distance between the proton pumps and ATP synthases, the ATP synthesis rate dropped and the lipid dependence of this activity vanished. The data indicate that protons are transferred along the membrane, between cytochrome bo3 and the ATP synthase, but only at sufficiently high protein densities. We also argue that the local protein density may be modulated by lipid-dependent changes in interactions between the two proteins complexes, which points to a mechanism by which the cell may regulate the overall activity of the respiratory chain. PMID:27063297

  4. iPPI-DB: an online database of modulators of protein-protein interactions.

    PubMed

    Labbé, Céline M; Kuenemann, Mélaine A; Zarzycka, Barbara; Vriend, Gert; Nicolaes, Gerry A F; Lagorce, David; Miteva, Maria A; Villoutreix, Bruno O; Sperandio, Olivier

    2016-01-01

    In order to boost the identification of low-molecular-weight drugs on protein-protein interactions (PPI), it is essential to properly collect and annotate experimental data about successful examples. This provides the scientific community with the necessary information to derive trends about privileged physicochemical properties and chemotypes that maximize the likelihood of promoting a given chemical probe to the most advanced stages of development. To this end we have developed iPPI-DB (freely accessible at http://www.ippidb.cdithem.fr), a database that contains the structure, some physicochemical characteristics, the pharmacological data and the profile of the PPI targets of several hundreds modulators of protein-protein interactions. iPPI-DB is accessible through a web application and can be queried according to two general approaches: using physicochemical/pharmacological criteria; or by chemical similarity to a user-defined structure input. In both cases the results are displayed as a sortable and exportable datasheet with links to external databases such as Uniprot, PubMed. Furthermore each compound in the table has a link to an individual ID card that contains its physicochemical and pharmacological profile derived from iPPI-DB data. This includes information about its binding data, ligand and lipophilic efficiencies, location in the PPI chemical space, and importantly similarity with known drugs, and links to external databases like PubChem, and ChEMBL.

  5. Network analysis reveals common host protein/s modulating pathogenesis of neurotropic viruses

    PubMed Central

    Ghosh, Sourish; Mukherjee, Sriparna; Sengupta, Nabonita; Roy, Arunava; Dey, Dhritiman; Chakraborty, Surajit; Chattopadhyay, Dhrubajyoti; Banerjee, Arpan; Basu, Anirban

    2016-01-01

    Network analysis through graph theory provides a quantitative approach to characterize specific proteins and their constituent assemblies that underlie host-pathogen interactions. In the present study, graph theory was used to analyze the interactome designed out of 50 differentially expressing proteins from proteomic analysis of Chandipura Virus (CHPV, Family: Rhabdoviridae) infected mouse brain tissue to identify the primary candidates for intervention. Using the measure of degree centrality, that quantifies the connectedness of a single protein within a milieu of several other interacting proteins, DJ-1 was selected for further molecular validation. To elucidate the generality of DJ-1’s role in propagating infection its role was also monitored in another RNA virus, Japanese Encephalitis Virus (JEV, Family: Flaviviridae) infection. Concurrently, DJ-1 got over-expressed in response to reactive oxygen species (ROS) generation following viral infection which in the early phase of infection migrated to mitochondria to remove dysfunctional mitochondria through the process of mitophagy. DJ-1 was also observed to modulate the viral replication and interferon responses along with low-density lipoprotein (LDL) receptor expression in neurons. Collectively these evidences reveal a comprehensive role for DJ-1 in neurotropic virus infection in the brain. PMID:27581498

  6. iPPI-DB: an online database of modulators of protein-protein interactions.

    PubMed

    Labbé, Céline M; Kuenemann, Mélaine A; Zarzycka, Barbara; Vriend, Gert; Nicolaes, Gerry A F; Lagorce, David; Miteva, Maria A; Villoutreix, Bruno O; Sperandio, Olivier

    2016-01-01

    In order to boost the identification of low-molecular-weight drugs on protein-protein interactions (PPI), it is essential to properly collect and annotate experimental data about successful examples. This provides the scientific community with the necessary information to derive trends about privileged physicochemical properties and chemotypes that maximize the likelihood of promoting a given chemical probe to the most advanced stages of development. To this end we have developed iPPI-DB (freely accessible at http://www.ippidb.cdithem.fr), a database that contains the structure, some physicochemical characteristics, the pharmacological data and the profile of the PPI targets of several hundreds modulators of protein-protein interactions. iPPI-DB is accessible through a web application and can be queried according to two general approaches: using physicochemical/pharmacological criteria; or by chemical similarity to a user-defined structure input. In both cases the results are displayed as a sortable and exportable datasheet with links to external databases such as Uniprot, PubMed. Furthermore each compound in the table has a link to an individual ID card that contains its physicochemical and pharmacological profile derived from iPPI-DB data. This includes information about its binding data, ligand and lipophilic efficiencies, location in the PPI chemical space, and importantly similarity with known drugs, and links to external databases like PubChem, and ChEMBL. PMID:26432833

  7. Network analysis reveals common host protein/s modulating pathogenesis of neurotropic viruses.

    PubMed

    Ghosh, Sourish; Mukherjee, Sriparna; Sengupta, Nabonita; Roy, Arunava; Dey, Dhritiman; Chakraborty, Surajit; Chattopadhyay, Dhrubajyoti; Banerjee, Arpan; Basu, Anirban

    2016-01-01

    Network analysis through graph theory provides a quantitative approach to characterize specific proteins and their constituent assemblies that underlie host-pathogen interactions. In the present study, graph theory was used to analyze the interactome designed out of 50 differentially expressing proteins from proteomic analysis of Chandipura Virus (CHPV, Family: Rhabdoviridae) infected mouse brain tissue to identify the primary candidates for intervention. Using the measure of degree centrality, that quantifies the connectedness of a single protein within a milieu of several other interacting proteins, DJ-1 was selected for further molecular validation. To elucidate the generality of DJ-1's role in propagating infection its role was also monitored in another RNA virus, Japanese Encephalitis Virus (JEV, Family: Flaviviridae) infection. Concurrently, DJ-1 got over-expressed in response to reactive oxygen species (ROS) generation following viral infection which in the early phase of infection migrated to mitochondria to remove dysfunctional mitochondria through the process of mitophagy. DJ-1 was also observed to modulate the viral replication and interferon responses along with low-density lipoprotein (LDL) receptor expression in neurons. Collectively these evidences reveal a comprehensive role for DJ-1 in neurotropic virus infection in the brain. PMID:27581498

  8. Lipid-mediated Protein-protein Interactions Modulate Respiration-driven ATP Synthesis.

    PubMed

    Nilsson, Tobias; Lundin, Camilla Rydström; Nordlund, Gustav; Ädelroth, Pia; von Ballmoos, Christoph; Brzezinski, Peter

    2016-04-11

    Energy conversion in biological systems is underpinned by membrane-bound proton transporters that generate and maintain a proton electrochemical gradient across the membrane which used, e.g. for generation of ATP by the ATP synthase. Here, we have co-reconstituted the proton pump cytochrome bo3 (ubiquinol oxidase) together with ATP synthase in liposomes and studied the effect of changing the lipid composition on the ATP synthesis activity driven by proton pumping. We found that for 100 nm liposomes, containing 5 of each proteins, the ATP synthesis rates decreased significantly with increasing fractions of DOPA, DOPE, DOPG or cardiolipin added to liposomes made of DOPC; with e.g. 5% DOPG, we observed an almost 50% decrease in the ATP synthesis rate. However, upon increasing the average distance between the proton pumps and ATP synthases, the ATP synthesis rate dropped and the lipid dependence of this activity vanished. The data indicate that protons are transferred along the membrane, between cytochrome bo3 and the ATP synthase, but only at sufficiently high protein densities. We also argue that the local protein density may be modulated by lipid-dependent changes in interactions between the two proteins complexes, which points to a mechanism by which the cell may regulate the overall activity of the respiratory chain.

  9. Dichotomous Metabolism of Enterococcus faecalis Induced by Hematin Starvation Modulates Colonic Gene Expression

    PubMed Central

    Allen, Toby D.; Moore, Danny R.; Wang, Xingmin; Casu, Viviana; May, Randal; Lerner, Megan R.; Houchen, Courtney; Brackett, Daniel J.; Huycke, Mark M.

    2009-01-01

    Summary Enterococcus faecalis is an intestinal commensal that cannot synthesize porphyrins and only expresses a functional respiratory chain when provided exogenous hematin. In the absence of hematin, E. faecalis reverts to fermentative metabolism and produces extracellular superoxide that can damage epithelial cell DNA. The acute response of the colonic mucosa to hematin-starved E. faecalis was identified by gene array. E. faecalis was inoculated into murine colons using a surgical ligation model that preserved tissue architecture and homeostasis. The mucosa was exposed to hematin-starved E. faecalis and compared to a control consisting of the same strain grown with hematin. At 1 hour post-inoculation six mucosal genes were differentially regulated and this increased to 42 genes at 6 hours. At 6 hours a highly significant biological interaction network was identified with functions that included NF-κB signaling, apoptosis, and cell cycle regulation. Colon biopsies showed no histological abnormalities by hematoxylin and eosin staining. Immunohistochemical staining, however, detected NF-κB activation in tissue macrophages using antibodies to the nuclear localization sequence for p65 and the F4/80 marker for murine macrophages. Similarly, hematin-starved E. faecalis strongly activated NF-κB in murine macrophages in vitro. Furthermore, primary and transformed colonic epithelial cells activated the G2/M checkpoint in vitro following exposure to hematin-starved E. faecalis. Modulation of this cell cycle checkpoint was due to extracellular superoxide produced as a result of the respiratory block in hematin-starved E. faecalis. These results demonstrate that the uniquely dichotomous metabolism of E. faecalis can significantly modulate gene expression in the colonic mucosa for pathways associated with inflammation, apoptosis, and cell cycle regulation. PMID:18809545

  10. The therapeutic potential of nuclear receptor modulators for treatment of metabolic disorders: PPARγ, RORs, and Rev-erbs.

    PubMed

    Marciano, David P; Chang, Mi Ra; Corzo, Cesar A; Goswami, Devrishi; Lam, Vinh Q; Pascal, Bruce D; Griffin, Patrick R

    2014-02-01

    Nuclear receptors (NRs) play central roles in metabolic syndrome, making them attractive drug targets despite the challenge of achieving functional selectivity. For instance, members of the thiazolidinedione class of insulin sensitizers offer robust efficacy but have been limited due to adverse effects linked to activation of genes not involved in insulin sensitization. Studies reviewed here provide strategies for targeting subsets of PPARγ target genes, enabling development of next-generation modulators with improved therapeutic index. Additionally, emerging evidence suggests that targeting the NRs ROR and Rev-erb holds promise for treating metabolic syndrome based on their involvement in circadian rhythm and metabolism.

  11. Interplay of Breast Cancer Resistance Protein (BCRP) and Metabolizing Enzymes.

    PubMed

    Tian, Ye; Bian, Yicong; Jiang, Yan; Qian, Sainan; Yu, Aiming; Zeng, Su

    2015-01-01

    The recent identification of the interplay between metabolizing enzymes and BCRP has drawn more and more attention from people. BCRP, a transporter belonging to ATP-binding cassette (ABC) family, has been hypothesized to play roles in many aspects including protecting the human body against therapeutics because it is expressed in the tissues that function as barriers in vivo. Efficient coupling of BCRP and metabolizing enzymes enables rapid elimination of foreign compounds from the body because BCRP could facilitate the excretion of metabolites catalyzed by phase I and II enzymes into bile, urine and feces. Without BCRP coupling, pass through the cell membrane may be difficult for them by passive diffusion because of the increment of the molecular weight and water solubility. Thus the metabolism-efflux alliance has extraordinary importance to drug metabolism, distribution, pharmacological effect, toxicity and elimination. In this manuscript, a brief discussion about the interplays of BCRP and metabolizing enzymes in liver, intestine, kidney, lung and other organs were presented and summarized. Many endogenous and exogenous compounds belong to different chemical groups, for instance, the dietary flavonoids and the steroidal hormones were involved. Clarifying the cooperation mechanisms of BCRP and enzymes could lead to a better prediction of drug clearance in vitro.

  12. Interplay of Breast Cancer Resistance Protein (BCRP) and Metabolizing Enzymes.

    PubMed

    Tian, Ye; Bian, Yicong; Jiang, Yan; Qian, Sainan; Yu, Aiming; Zeng, Su

    2015-01-01

    The recent identification of the interplay between metabolizing enzymes and BCRP has drawn more and more attention from people. BCRP, a transporter belonging to ATP-binding cassette (ABC) family, has been hypothesized to play roles in many aspects including protecting the human body against therapeutics because it is expressed in the tissues that function as barriers in vivo. Efficient coupling of BCRP and metabolizing enzymes enables rapid elimination of foreign compounds from the body because BCRP could facilitate the excretion of metabolites catalyzed by phase I and II enzymes into bile, urine and feces. Without BCRP coupling, pass through the cell membrane may be difficult for them by passive diffusion because of the increment of the molecular weight and water solubility. Thus the metabolism-efflux alliance has extraordinary importance to drug metabolism, distribution, pharmacological effect, toxicity and elimination. In this manuscript, a brief discussion about the interplays of BCRP and metabolizing enzymes in liver, intestine, kidney, lung and other organs were presented and summarized. Many endogenous and exogenous compounds belong to different chemical groups, for instance, the dietary flavonoids and the steroidal hormones were involved. Clarifying the cooperation mechanisms of BCRP and enzymes could lead to a better prediction of drug clearance in vitro. PMID:26652256

  13. Metformin revisited: Does this regulator of AMP-activated protein kinase secondarily affect bone metabolism and prevent diabetic osteopathy.

    PubMed

    McCarthy, Antonio Desmond; Cortizo, Ana María; Sedlinsky, Claudia

    2016-03-25

    Patients with long-term type 1 and type 2 diabetes mellitus (DM) can develop skeletal complications or "diabetic osteopathy". These include osteopenia, osteoporosis and an increased incidence of low-stress fractures. In this context, it is important to evaluate whether current anti-diabetic treatments can secondarily affect bone metabolism. Adenosine monophosphate-activated protein kinase (AMPK) modulates multiple metabolic pathways and acts as a sensor of the cellular energy status; recent evidence suggests a critical role for AMPK in bone homeostasis. In addition, AMPK activation is believed to mediate most clinical effects of the insulin-sensitizer metformin. Over the past decade, several research groups have investigated the effects of metformin on bone, providing a considerable body of pre-clinical (in vitro, ex vivo and in vivo) as well as clinical evidence for an anabolic action of metformin on bone. However, two caveats should be kept in mind when considering metformin treatment for a patient with type 2 DM at risk for diabetic osteopathy. In the first place, metformin should probably not be considered an anti-osteoporotic drug; it is an insulin sensitizer with proven macrovascular benefits that can secondarily improve bone metabolism in the context of DM. Secondly, we are still awaiting the results of randomized placebo-controlled studies in humans that evaluate the effects of metformin on bone metabolism as a primary endpoint.

  14. Metformin revisited: Does this regulator of AMP-activated protein kinase secondarily affect bone metabolism and prevent diabetic osteopathy

    PubMed Central

    McCarthy, Antonio Desmond; Cortizo, Ana María; Sedlinsky, Claudia

    2016-01-01

    Patients with long-term type 1 and type 2 diabetes mellitus (DM) can develop skeletal complications or “diabetic osteopathy”. These include osteopenia, osteoporosis and an increased incidence of low-stress fractures. In this context, it is important to evaluate whether current anti-diabetic treatments can secondarily affect bone metabolism. Adenosine monophosphate-activated protein kinase (AMPK) modulates multiple metabolic pathways and acts as a sensor of the cellular energy status; recent evidence suggests a critical role for AMPK in bone homeostasis. In addition, AMPK activation is believed to mediate most clinical effects of the insulin-sensitizer metformin. Over the past decade, several research groups have investigated the effects of metformin on bone, providing a considerable body of pre-clinical (in vitro, ex vivo and in vivo) as well as clinical evidence for an anabolic action of metformin on bone. However, two caveats should be kept in mind when considering metformin treatment for a patient with type 2 DM at risk for diabetic osteopathy. In the first place, metformin should probably not be considered an anti-osteoporotic drug; it is an insulin sensitizer with proven macrovascular benefits that can secondarily improve bone metabolism in the context of DM. Secondly, we are still awaiting the results of randomized placebo-controlled studies in humans that evaluate the effects of metformin on bone metabolism as a primary endpoint. PMID:27022443

  15. Bioenergetic cues shift FXR splicing towards FXRα2 to modulate hepatic lipolysis and fatty acid metabolism

    PubMed Central

    Correia, Jorge C.; Massart, Julie; de Boer, Jan Freark; Porsmyr-Palmertz, Margareta; Martínez-Redondo, Vicente; Agudelo, Leandro Z.; Sinha, Indranil; Meierhofer, David; Ribeiro, Vera; Björnholm, Marie; Sauer, Sascha; Dahlman-Wright, Karin; Zierath, Juleen R.; Groen, Albert K.; Ruas, Jorge L.

    2015-01-01

    Objective Farnesoid X receptor (FXR) plays a prominent role in hepatic lipid metabolism. The FXR gene encodes four proteins with structural differences suggestive of discrete biological functions about which little is known. Methods We expressed each FXR variant in primary hepatocytes and evaluated global gene expression, lipid profile, and metabolic fluxes. Gene delivery of FXR variants to Fxr−/− mouse liver was performed to evaluate their role in vivo. The effects of fasting and physical exercise on hepatic Fxr splicing were determined. Results We show that FXR splice isoforms regulate largely different gene sets and have specific effects on hepatic metabolism. FXRα2 (but not α1) activates a broad transcriptional program in hepatocytes conducive to lipolysis, fatty acid oxidation, and ketogenesis. Consequently, FXRα2 decreases cellular lipid accumulation and improves cellular insulin signaling to AKT. FXRα2 expression in Fxr−/− mouse liver activates a similar gene program and robustly decreases hepatic triglyceride levels. On the other hand, FXRα1 reduces hepatic triglyceride content to a lesser extent and does so through regulation of lipogenic gene expression. Bioenergetic cues, such as fasting and exercise, dynamically regulate Fxr splicing in mouse liver to increase Fxrα2 expression. Conclusions Our results show that the main FXR variants in human liver (α1 and α2) reduce hepatic lipid accumulation through distinct mechanisms and to different degrees. Taking this novel mechanism into account could greatly improve the pharmacological targeting and therapeutic efficacy of FXR agonists. PMID:26909306

  16. Amyloid precursor protein modulates β-catenin degradation

    PubMed Central

    Chen, Yuzhi; Bodles, Angela M

    2007-01-01

    Background The amyloid precursor protein (APP) is genetically associated with Alzheimer's disease (AD). Elucidating the function of APP should help understand AD pathogenesis and provide insights into therapeutic designs against this devastating neurodegenerative disease. Results We demonstrate that APP expression in primary neurons induces β-catenin phosphorylation at Ser33, Ser37, and Thr41 (S33/37/T41) residues, which is a prerequisite for β-catenin ubiquitinylation and proteasomal degradation. APP-induced phosphorylation of β-catenin resulted in the reduction of total β-catenin levels, suggesting that APP expression promotes β-catenin degradation. In contrast, treatment of neurons with APP siRNAs increased total β-catenin levels and decreased β-catenin phosphorylation at residues S33/37/T41. Further, β-catenin was dramatically increased in hippocampal CA1 pyramidal cells from APP knockout animals. Acute expression of wild type APP or of familial AD APP mutants in primary neurons downregulated β-catenin in membrane and cytosolic fractions, and did not appear to affect nuclear β-catenin or β-catenin-dependent transcription. Conversely, in APP knockout CA1 pyramidal cells, accumulation of β-catenin was associated with the upregulation of cyclin D1, a downstream target of β-catenin signaling. Together, these data establish that APP downregulates β-catenin and suggest a role for APP in sustaining neuronal function by preventing cell cycle reactivation and maintaining synaptic integrity. Conclusion We have provided strong evidence that APP modulates β-catenin degradation in vitro and in vivo. Future studies may investigate whether APP processing is necessary for β-catenin downregulation, and determine if excessive APP expression contributes to AD pathogenesis through abnormal β-catenin downregulation. PMID:18070361

  17. Distinct role of Arabidopsis mitochondrial P-type pentatricopeptide repeat protein-modulating editing protein, PPME, in nad1 RNA editing

    PubMed Central

    Leu, Kuan-Chieh; Hsieh, Ming-Hsiun; Wang, Huei-Jing; Hsieh, Hsu-Liang

    2016-01-01

    ABSTRACT The mitochondrion is an important power generator in most eukaryotic cells. To preserve its function, many essential nuclear-encoded factors play specific roles in mitochondrial RNA metabolic processes, including RNA editing. RNA editing consists of post-transcriptional deamination, which alters specific nucleotides in transcripts to mediate gene expression. In plant cells, many pentatricopeptide repeat proteins (PPRs) participate in diverse organellar RNA metabolic processes, but only PLS-type PPRs are involved in RNA editing. Here, we report a P-type PPR protein from Arabidopsis thaliana, P-type PPR-Modulating Editing (PPME), which has a distinct role in mitochondrial nad1 RNA editing via RNA binding activity. In the homozygous ppme mutant, cytosine (C)-to-uracil (U) conversions at both the nad1-898 and 937 sites were abolished, disrupting Arg300-to-Trp300 and Pro313-to-Ser313 amino acid changes in the mitochondrial NAD1 protein. NAD1 is a critical component of mitochondrial respiration complex I; its activity is severely reduced in the homozygous ppme mutant, resulting in significantly altered growth and development. Both abolished RNA editing and defective complex I activity were completely rescued by CaMV 35S promoter- and PPME native promoter-driven PPME genomic fragments tagged with GFP in a homozygous ppme background. Our experimental results demonstrate a distinct role of a P-type PPR protein, PPME, in RNA editing in plant organelles. PMID:27149614

  18. Integrating the protein and metabolic engineering toolkits for next-generation chemical biosynthesis.

    PubMed

    Pirie, Christopher M; De Mey, Marjan; Jones Prather, Kristala L; Ajikumar, Parayil Kumaran

    2013-04-19

    Through microbial engineering, biosynthesis has the potential to produce thousands of chemicals used in everyday life. Metabolic engineering and synthetic biology are fields driven by the manipulation of genes, genetic regulatory systems, and enzymatic pathways for developing highly productive microbial strains. Fundamentally, it is the biochemical characteristics of the enzymes themselves that dictate flux through a biosynthetic pathway toward the product of interest. As metabolic engineers target sophisticated secondary metabolites, there has been little recognition of the reduced catalytic activity and increased substrate/product promiscuity of the corresponding enzymes compared to those of central metabolism. Thus, fine-tuning these enzymatic characteristics through protein engineering is paramount for developing high-productivity microbial strains for secondary metabolites. Here, we describe the importance of protein engineering for advancing metabolic engineering of secondary metabolism pathways. This pathway integrated enzyme optimization can enhance the collective toolkit of microbial engineering to shape the future of chemical manufacturing.

  19. Design, synthesis, and biological characterization of metabolically stable selective androgen receptor modulators.

    PubMed

    Marhefka, Craig A; Gao, Wenqing; Chung, Kiwon; Kim, Juhyun; He, Yali; Yin, Donghua; Bohl, Casey; Dalton, James T; Miller, Duane D

    2004-02-12

    A series of nonsteroidal ligands were synthesized as second-generation agonists for the androgen receptor (AR). These ligands were designed to eliminate metabolic sites identified in one of our first-generation AR agonists, which was inactive in vivo due to its rapid metabolism to inactive constituents. The binding affinity of these compounds was evaluated using AR isolated from rat ventral prostate. These second-generation compounds bound the AR in a high affinity and stereoselective manner, with K(i) values ranging from about 4 to 130 nM. The ability of these ligands to stimulate AR-mediated transcriptional activation was examined in cells transfected with the human AR and a hormone-dependent luciferase reporter gene. Although some compounds were unable to stimulate AR-mediated transcription, several demonstrated activity similar to that of dihydrotestosterone (DHT, an endogenous steroidal ligand for the AR). We also evaluated the in vivo pharmacologic activity of selected compounds in castrated male rats. Three compounds were identified as selective androgen receptor modulators (SARMs), exhibiting significant anabolic activity while having only moderate to minimal androgenic activity in vivo.

  20. Sneaker Male Squid Produce Long-lived Spermatozoa by Modulating Their Energy Metabolism.

    PubMed

    Hirohashi, Noritaka; Tamura-Nakano, Miwa; Nakaya, Fumio; Iida, Tomohiro; Iwata, Yoko

    2016-09-01

    Spermatozoa released by males should remain viable until fertilization. Hence, sperm longevity is governed by intrinsic and environmental factors in accordance with the male mating strategy. However, whether intraspecific variation of insemination modes can impact sperm longevity remains to be elucidated. In the squid Heterololigo bleekeri, male dimorphism (consort and sneaker) is linked to two discontinuous insemination modes that differ in place and time. Notably, only sneaker male spermatozoa inseminated long before egg spawning can be stored in the seminal receptacle. We found that sneaker spermatozoa exhibited greater persistence in fertilization competence and flagellar motility than consort ones because of a larger amount of flagellar glycogen. Sneaker spermatozoa also showed higher capacities in glucose uptake and lactate efflux. Lactic acidosis was considered to stabilize CO2-triggered self-clustering of sneaker spermatozoa, thus establishing hypoxia-induced metabolic changes and sperm survival. These results, together with comparative omics analyses, suggest that postcopulatory reproductive contexts define sperm longevity by modulating the inherent energy levels and metabolic pathways. PMID:27385589

  1. Low level light in combination with metabolic modulators for effective therapy

    NASA Astrophysics Data System (ADS)

    Dong, Tingting; Zhang, Qi; Hamblin, Michael R.; Wu, Mei X.

    2015-03-01

    Vascular damage occurs frequently at the injured brain causing hypoxia and is associated with poor outcomes in the clinics. We found high levels of glycolysis, reduced ATP generation, and increased formation of reactive oxygen species (ROS) and apoptosis in neurons under hypoxia. Strikingly, these adverse events were reversed significantly by noninvasive exposure of injured brain to low-level light (LLL). LLL illumination sustained the mitochondrial membrane potential, constrained cytochrome C leakage in hypoxic cells, and protected them from apoptosis, underscoring a unique property of LLL. The effect of LLL was further bolstered by combination with metabolic substrates such as pyruvate or lactate both in vivo and in vitro. The combinational treatment retained memory and learning activities of injured mice to a normal level, whereas those treated with LLL or pyruvate alone, or sham light displayed partial or severe deficiency in these cognitive functions. In accordance with well-protected learning and memory function, the hippocampal region primarily responsible for learning and memory was completely protected by a combination of LLL and pyruvate, in marked contrast to the severe loss of hippocampal tissue due to secondary damage in control mice. These data clearly suggest that energy metabolic modulators can additively or synergistically enhance the therapeutic effect of LLL in energy-producing insufficient tissues like injured brain. Keywords:

  2. Low-level light in combination with metabolic modulators for effective therapy of injured brain

    PubMed Central

    Dong, Tingting; Zhang, Qi; Hamblin, Michael R; Wu, Mei X

    2015-01-01

    Vascular damage occurs frequently at the injured brain causing hypoxia and is associated with poor outcomes in the clinics. We found high levels of glycolysis, reduced adenosine triphosphate generation, and increased formation of reactive oxygen species and apoptosis in neurons under hypoxia. Strikingly, these adverse events were reversed significantly by noninvasive exposure of injured brain to low-level light (LLL). Low-level light illumination sustained the mitochondrial membrane potential, constrained cytochrome c leakage in hypoxic cells, and protected them from apoptosis, underscoring a unique property of LLL. The effect of LLL was further bolstered by combination with metabolic substrates such as pyruvate or lactate both in vivo and in vitro. The combinational treatment retained memory and learning activities of injured mice to a normal level, whereas other treatment displayed partial or severe deficiency in these cognitive functions. In accordance with well-protected learning and memory function, the hippocampal region primarily responsible for learning and memory was completely protected by combination treatment, in marked contrast to the severe loss of hippocampal tissue because of secondary damage in control mice. These data clearly suggest that energy metabolic modulators can additively or synergistically enhance the therapeutic effect of LLL in energy-producing insufficient tissue–like injured brain. PMID:25966949

  3. Neuronal Control of Metabolism through Nutrient-Dependent Modulation of Tracheal Branching

    PubMed Central

    Linneweber, Gerit A.; Jacobson, Jake; Busch, Karl Emanuel; Hudry, Bruno; Christov, Christo P.; Dormann, Dirk; Yuan, Michaela; Otani, Tomoki; Knust, Elisabeth; de Bono, Mario; Miguel-Aliaga, Irene

    2014-01-01

    Summary During adaptive angiogenesis, a key process in the etiology and treatment of cancer and obesity, the vasculature changes to meet the metabolic needs of its target tissues. Although the cues governing vascular remodeling are not fully understood, target-derived signals are generally believed to underlie this process. Here, we identify an alternative mechanism by characterizing the previously unrecognized nutrient-dependent plasticity of the Drosophila tracheal system: a network of oxygen-delivering tubules developmentally akin to mammalian blood vessels. We find that this plasticity, particularly prominent in the intestine, drives—rather than responds to—metabolic change. Mechanistically, it is regulated by distinct populations of nutrient- and oxygen-responsive neurons that, through delivery of both local and systemic insulin- and VIP-like neuropeptides, sculpt the growth of specific tracheal subsets. Thus, we describe a novel mechanism by which nutritional cues modulate neuronal activity to give rise to organ-specific, long-lasting changes in vascular architecture. PMID:24439370

  4. Emergence of Complexity in Protein Functions and Metabolic Networks

    NASA Technical Reports Server (NTRS)

    Pohorille, Andzej

    2009-01-01

    In modern organisms proteins perform a majority of cellular functions, such as chemical catalysis, energy transduction and transport of material across cell walls. Although great strides have been made towards understanding protein evolution, a meaningful extrapolation from contemporary proteins to their earliest ancestors is virtually impossible. In an alternative approach, the origin of water-soluble proteins was probed through the synthesis of very large libraries of random amino acid sequences and subsequently subjecting them to in vitro evolution. In combination with computer modeling and simulations, these experiments allow us to address a number of fundamental questions about the origins of proteins. Can functionality emerge from random sequences of proteins? How did the initial repertoire of functional proteins diversify to facilitate new functions? Did this diversification proceed primarily through drawing novel functionalities from random sequences or through evolution of already existing proto-enzymes? Did protein evolution start from a pool of proteins defined by a frozen accident and other collections of proteins could start a different evolutionary pathway? Although we do not have definitive answers to these questions, important clues have been uncovered. Considerable progress has been also achieved in understanding the origins of membrane proteins. We will address this issue in the example of ion channels - proteins that mediate transport of ions across cell walls. Remarkably, despite overall complexity of these proteins in contemporary cells, their structural motifs are quite simple, with -helices being most common. By combining results of experimental and computer simulation studies on synthetic models and simple, natural channels, I will show that, even though architectures of membrane proteins are not nearly as diverse as those of water-soluble proteins, they are sufficiently flexible to adapt readily to the functional demands arising during

  5. Milk protein for improved metabolic health: a review of the evidence

    PubMed Central

    2013-01-01

    Epidemiological evidence shows that consumption of dairy products is associated with decreased prevalence of metabolic related disorders, whilst evidence from experimental studies points towards dairy protein as a dietary component which may aid prevention of type 2 diabetes (T2DM). Poor metabolic health is a common characteristic of overweight, obesity and aging, and is the forerunner of T2DM and cardiovascular disease (CVD), and an ever increasing global health issue. Progressive loss of metabolic control is evident from a blunting of carbohydrate, fat and protein metabolism, which is commonly manifested through decreased insulin sensitivity, inadequate glucose and lipid control, accompanied by a pro-inflammatory environment and hypertension. Adverse physiological changes such as excess visceral adipose tissue deposition and expansion, lipid overspill and infiltration into liver, muscle and other organs, and sarcopaenia or degenerative loss of skeletal muscle mass and function all underpin this adverse profile. ‘Sarcobesity’ and sarcopaenic diabetes are rapidly growing health issues. As well as through direct mechanisms, dairy protein may indirectly improve metabolic health by aiding loss of body weight and fat mass through enhanced satiety, whilst promoting skeletal muscle growth and function through anabolic effects of dairy protein-derived branch chain amino acids (BCAAs). BCAAs enhance muscle protein synthesis, lean body mass and skeletal muscle metabolic function. The composition and processing of dairy protein has an impact on digestion, absorption, BCAA kinetics and function, hence the optimisation of dairy protein composition through selection and combination of specific protein components in milk may provide a way to maximize benefits for metabolic health. PMID:23822206

  6. Metabolic and trophic interactions modulate methane production by Arctic peat microbiota in response to warming.

    PubMed

    Tveit, Alexander Tøsdal; Urich, Tim; Frenzel, Peter; Svenning, Mette Marianne

    2015-05-12

    Arctic permafrost soils store large amounts of soil organic carbon (SOC) that could be released into the atmosphere as methane (CH4) in a future warmer climate. How warming affects the complex microbial network decomposing SOC is not understood. We studied CH4 production of Arctic peat soil microbiota in anoxic microcosms over a temperature gradient from 1 to 30 °C, combining metatranscriptomic, metagenomic, and targeted metabolic profiling. The CH4 production rate at 4 °C was 25% of that at 25 °C and increased rapidly with temperature, driven by fast adaptations of microbial community structure, metabolic network of SOC decomposition, and trophic interactions. Below 7 °C, syntrophic propionate oxidation was the rate-limiting step for CH4 production; above this threshold temperature, polysaccharide hydrolysis became rate limiting. This change was associated with a shift within the functional guild for syntrophic propionate oxidation, with Firmicutes being replaced by Bacteroidetes. Correspondingly, there was a shift from the formate- and H2-using Methanobacteriales to Methanomicrobiales and from the acetotrophic Methanosarcinaceae to Methanosaetaceae. Methanogenesis from methylamines, probably stemming from degradation of bacterial cells, became more important with increasing temperature and corresponded with an increased relative abundance of predatory protists of the phylum Cercozoa. We concluded that Arctic peat microbiota responds rapidly to increased temperatures by modulating metabolic and trophic interactions so that CH4 is always highly produced: The microbial community adapts through taxonomic shifts, and cascade effects of substrate availability cause replacement of functional guilds and functional changes within taxa.

  7. Metabolic and trophic interactions modulate methane production by Arctic peat microbiota in response to warming

    PubMed Central

    Tveit, Alexander Tøsdal; Urich, Tim; Frenzel, Peter; Svenning, Mette Marianne

    2015-01-01

    Arctic permafrost soils store large amounts of soil organic carbon (SOC) that could be released into the atmosphere as methane (CH4) in a future warmer climate. How warming affects the complex microbial network decomposing SOC is not understood. We studied CH4 production of Arctic peat soil microbiota in anoxic microcosms over a temperature gradient from 1 to 30 °C, combining metatranscriptomic, metagenomic, and targeted metabolic profiling. The CH4 production rate at 4 °C was 25% of that at 25 °C and increased rapidly with temperature, driven by fast adaptations of microbial community structure, metabolic network of SOC decomposition, and trophic interactions. Below 7 °C, syntrophic propionate oxidation was the rate-limiting step for CH4 production; above this threshold temperature, polysaccharide hydrolysis became rate limiting. This change was associated with a shift within the functional guild for syntrophic propionate oxidation, with Firmicutes being replaced by Bacteroidetes. Correspondingly, there was a shift from the formate- and H2-using Methanobacteriales to Methanomicrobiales and from the acetotrophic Methanosarcinaceae to Methanosaetaceae. Methanogenesis from methylamines, probably stemming from degradation of bacterial cells, became more important with increasing temperature and corresponded with an increased relative abundance of predatory protists of the phylum Cercozoa. We concluded that Arctic peat microbiota responds rapidly to increased temperatures by modulating metabolic and trophic interactions so that CH4 is always highly produced: The microbial community adapts through taxonomic shifts, and cascade effects of substrate availability cause replacement of functional guilds and functional changes within taxa. PMID:25918393

  8. Resveratrol modulates drug- and carcinogen-metabolizing enzymes in a healthy volunteer study.

    PubMed

    Chow, H-H Sherry; Garland, Linda L; Hsu, Chiu-Hsieh; Vining, Donna R; Chew, Wade M; Miller, Jessica A; Perloff, Marjorie; Crowell, James A; Alberts, David S

    2010-09-01

    Resveratrol has been shown to exhibit cancer-preventive activities in preclinical studies. We conducted a clinical study to determine the effect of pharmacologic doses of resveratrol on drug- and carcinogen-metabolizing enzymes. Forty-two healthy volunteers underwent baseline assessment of cytochrome P450 (CYP) and phase II detoxification enzymes. CYP1A2, CYP2D6, CYP2C9, and CYP3A4 enzyme activities were measured by the metabolism of caffeine, dextromethorphan, losartan, and buspirone, respectively. Blood lymphocyte glutathione S-transferase (GST) activity and GST-pi level and serum total and direct bilirubin, a surrogate for UDP-glucuronosyl transferase (UGT) 1A1 activity, were measured to assess phase II enzymes. After the baseline evaluation, study participants took 1 g of resveratrol once daily for 4 weeks. Enzyme assessment was repeated upon intervention completion. Resveratrol intervention was found to inhibit the phenotypic indices of CYP3A4, CYP2D6, and CYP2C9 and to induce the phenotypic index of 1A2. Overall, GST and UGT1A1 activities were minimally affected by the intervention, although an induction of GST-pi level and UGT1A1 activity was observed in individuals with low baseline enzyme level/activity. We conclude that resveratrol can modulate enzyme systems involved in carcinogen activation and detoxification, which may be one mechanism by which resveratrol inhibits carcinogenesis. However, pharmacologic doses of resveratrol could potentially lead to increased adverse drug reactions or altered drug efficacy due to inhibition or induction of certain CYPs. Further clinical development of resveratrol for cancer prevention should consider evaluation of lower doses of resveratrol to minimize adverse metabolic drug interactions. PMID:20716633

  9. Siderophore Biosynthesis Governs the Virulence of Uropathogenic Escherichia coli by Coordinately Modulating the Differential Metabolism.

    PubMed

    Su, Qiao; Guan, Tianbing; He, Yan; Lv, Haitao

    2016-04-01

    Urinary tract infections impose substantial health burdens on women worldwide. Urinary tract infections often incur a high risk of recurrence and antibiotic resistance, and uropathogenic E. coli accounts for approximately 80% of clinically acquired cases. The diagnosis of, treatment of, and drug development for urinary tract infections remain substantial challenges due to the complex pathogenesis of this condition. The clinically isolated UPEC 83972 strain was found to produce four siderophores: yersiniabactin, aerobactin, salmochelin, and enterobactin. The biosyntheses of some of these siderophores implies that the virulence of UPEC is mediated via the targeting of primary metabolism. However, the differential modulatory roles of siderophore biosyntheses on the differential metabolomes of UPEC and non-UPEC strains remain incompletely understood. In the present study, we sought to investigate how the differential metabolomes can be used to distinguish UPEC from non-UPEC strains and to determine the associated regulatory roles of siderophore biosynthesis. Our results are the first to demonstrate that the identified differential metabolomes strongly differentiated UPEC from non-UPEC strains. Furthermore, we performed metabolome assays of mutants with different patterns of siderophore deletions; the data revealed that the mutations of all four siderophores exerted a stronger modulatory role on the differential metabolomes of the UPEC and non-UPEC strains relative to the mutation of any single siderophore and that this modulatory role primarily involved amino acid metabolism, oxidative phosphorylation in the carbon fixation pathway, and purine and pyrimidine metabolism. Surprisingly, the modulatory roles were strongly dependent on the type and number of mutated siderophores. Taken together, these results demonstrated that siderophore biosynthesis coordinately modulated the differential metabolomes and thus may indicate novel targets for virulence-based diagnosis

  10. Coxiella burnetii effector CvpB modulates phosphoinositide metabolism for optimal vacuole development.

    PubMed

    Martinez, Eric; Allombert, Julie; Cantet, Franck; Lakhani, Anissa; Yandrapalli, Naresh; Neyret, Aymeric; Norville, Isobel H; Favard, Cyril; Muriaux, Delphine; Bonazzi, Matteo

    2016-06-01

    The Q fever bacterium Coxiella burnetii replicates inside host cells within a large Coxiella-containing vacuole (CCV) whose biogenesis relies on the Dot/Icm-dependent secretion of bacterial effectors. Several membrane trafficking pathways contribute membranes, proteins, and lipids for CCV biogenesis. These include the endocytic and autophagy pathways, which are characterized by phosphatidylinositol 3-phosphate [PI(3)P]-positive membranes. Here we show that the C. burnetii secreted effector Coxiella vacuolar protein B (CvpB) binds PI(3)P and phosphatidylserine (PS) on CCVs and early endosomal compartments and perturbs the activity of the phosphatidylinositol 5-kinase PIKfyve to manipulate PI(3)P metabolism. CvpB association to early endosome triggers vacuolation and clustering, leading to the channeling of large PI(3)P-positive membranes to CCVs for vacuole expansion. At CCVs, CvpB binding to early endosome- and autophagy-derived PI(3)P and the concomitant inhibition of PIKfyve favor the association of the autophagosomal machinery to CCVs for optimal homotypic fusion of the Coxiella-containing compartments. The importance of manipulating PI(3)P metabolism is highlighted by mutations in cvpB resulting in a multivacuolar phenotype, rescuable by gene complementation, indicative of a defect in CCV biogenesis. Using the insect model Galleria mellonella, we demonstrate the in vivo relevance of defective CCV biogenesis by highlighting an attenuated virulence phenotype associated with cvpB mutations. PMID:27226300

  11. In vivo versus in vitro protein abundance analysis of Shigella dysenteriae type 1 reveals changes in the expression of proteins involved in virulence, stress and energy metabolism

    PubMed Central

    2011-01-01

    Background Shigella dysenteriae serotype 1 (SD1) causes the most severe form of epidemic bacillary dysentery. Quantitative proteome profiling of Shigella dysenteriae serotype 1 (SD1) in vitro (derived from LB cell cultures) and in vivo (derived from gnotobiotic piglets) was performed by 2D-LC-MS/MS and APEX, a label-free computationally modified spectral counting methodology. Results Overall, 1761 proteins were quantitated at a 5% FDR (false discovery rate), including 1480 and 1505 from in vitro and in vivo samples, respectively. Identification of 350 cytoplasmic membrane and outer membrane (OM) proteins (38% of in silico predicted SD1 membrane proteome) contributed to the most extensive survey of the Shigella membrane proteome reported so far. Differential protein abundance analysis using statistical tests revealed that SD1 cells switched to an anaerobic energy metabolism under in vivo conditions, resulting in an increase in fermentative, propanoate, butanoate and nitrate metabolism. Abundance increases of transcription activators FNR and Nar supported the notion of a switch from aerobic to anaerobic respiration in the host gut environment. High in vivo abundances of proteins involved in acid resistance (GadB, AdiA) and mixed acid fermentation (PflA/PflB) indicated bacterial survival responses to acid stress, while increased abundance of oxidative stress proteins (YfiD/YfiF/SodB) implied that defense mechanisms against oxygen radicals were mobilized. Proteins involved in peptidoglycan turnover (MurB) were increased, while β-barrel OM proteins (OmpA), OM lipoproteins (NlpD), chaperones involved in OM protein folding pathways (YraP, NlpB) and lipopolysaccharide biosynthesis (Imp) were decreased, suggesting unexpected modulations of the outer membrane/peptidoglycan layers in vivo. Several virulence proteins of the Mxi-Spa type III secretion system and invasion plasmid antigens (Ipa proteins) required for invasion of colonic epithelial cells, and release of bacteria

  12. PPARα inhibition modulates multiple reprogrammed metabolic pathways in kidney cancer and attenuates tumor growth.

    PubMed

    Abu Aboud, Omran; Donohoe, Dallas; Bultman, Scott; Fitch, Mark; Riiff, Tim; Hellerstein, Marc; Weiss, Robert H

    2015-06-01

    Kidney cancer [renal cell carcinoma (RCC)] is the sixth-most-common cancer in the United States, and its incidence is increasing. The current progression-free survival for patients with advanced RCC rarely extends beyond 1-2 yr due to the development of therapeutic resistance. We previously identified peroxisome proliferator-activating receptor-α (PPARα) as a potential therapeutic target for this disease and showed that a specific PPARα antagonist, GW6471, induced apoptosis and cell cycle arrest at G0/G1 in RCC cell lines associated with attenuation of cell cycle regulatory proteins. We now extend that work and show that PPARα inhibition attenuates components of RCC metabolic reprogramming, capitalizing on the Warburg effect. The specific PPARα inhibitor GW6471, as well as a siRNA specific to PPARα, attenuates the enhanced fatty acid oxidation and oxidative phosphorylation associated with glycolysis inhibition, and PPARα antagonism also blocks the enhanced glycolysis that has been observed in RCC cells; this effect did not occur in normal human kidney epithelial cells. Such cell type-specific inhibition of glycolysis corresponds with changes in protein levels of the oncogene c-Myc and has promising clinical implications. Furthermore, we show that treatment with GW6471 results in RCC tumor growth attenuation in a xenograft mouse model, with minimal obvious toxicity, a finding associated with the expected on-target effects on c-Myc. These studies demonstrate that several pivotal cancer-relevant metabolic pathways are inhibited by PPARα antagonism. Our data support the concept that targeting PPARα, with or without concurrent inhibition of glycolysis, is a potential novel and effective therapeutic approach for RCC that targets metabolic reprogramming in this tumor.

  13. Approaches to Optimizing Animal Cell Culture Process: Substrate Metabolism Regulation and Protein Expression Improvement

    NASA Astrophysics Data System (ADS)

    Zhang, Yuanxing

    Some high value proteins and vaccines for medical and veterinary applications by animal cell culture have an increasing market in China. In order to meet the demands of large-scale productions of proteins and vaccines, animal cell culture technology has been widely developed. In general, an animal cell culture process can be divided into two stages in a batch culture. In cell growth stage a high specific growth rate is expected to achieve a high cell density. In production stage a high specific production rate is stressed for the expression and secretion of qualified protein or replication of virus. It is always critical to maintain high cell viability in fed-batch and perfusion cultures. More concern has been focused on two points by the researchers in China. First, the cell metabolism of substrates is analyzed and the accumulation of toxic by-products is decreased through regulating cell metabolism in the culture process. Second, some important factors effecting protein expression are understood at the molecular level and the production ability of protein is improved. In pace with the rapid development of large-scale cell culture for the production of vaccines, antibodies and other recombinant proteins in China, the medium design and process optimization based on cell metabolism regulation and protein expression improvement will play an important role. The chapter outlines the main advances in metabolic regulation of cell and expression improvement of protein in animal cell culture in recent years.

  14. Conjugated Linoleic Acid Supplementation under a High-Fat Diet Modulates Stomach Protein Expression and Intestinal Microbiota in Adult Mice

    PubMed Central

    Chaplin, Alice; Parra, Pilar; Serra, Francisca; Palou, Andreu

    2015-01-01

    The gastrointestinal tract constitutes a physiological interface integrating nutrient and microbiota-host metabolism. Conjugated linoleic acids (CLA) have been reported to contribute to decreased body weight and fat accretion. The modulation by dietary CLA of stomach proteins related to energy homeostasis or microbiota may be involved, although this has not been previously analysed. This is examined in the present study, which aims to underline the potential mechanisms of CLA which contribute to body weight regulation. Adult mice were fed either a normal fat (NF, 12% kJ content as fat) or a high-fat (HF, 43% kJ content as fat) diet. In the latter case, half of the animals received daily oral supplementation of CLA. Expression and content of stomach proteins and specific bacterial populations from caecum were analysed. CLA supplementation was associated with an increase in stomach protein expression, and exerted a prebiotic action on both Bacteroidetes/Prevotella and Akkermansia muciniphila. However, CLA supplementation was not able to override the negative effects of HF diet on Bifidobacterium spp., which was decreased in both HF and HF+CLA groups. Our data show that CLA are able to modulate stomach protein expression and exert a prebiotic effect on specific gut bacterial species. PMID:25915857

  15. Conjugated Linoleic Acid Supplementation under a High-Fat Diet Modulates Stomach Protein Expression and Intestinal Microbiota in Adult Mice.

    PubMed

    Chaplin, Alice; Parra, Pilar; Serra, Francisca; Palou, Andreu

    2015-01-01

    The gastrointestinal tract constitutes a physiological interface integrating nutrient and microbiota-host metabolism. Conjugated linoleic acids (CLA) have been reported to contribute to decreased body weight and fat accretion. The modulation by dietary CLA of stomach proteins related to energy homeostasis or microbiota may be involved, although this has not been previously analysed. This is examined in the present study, which aims to underline the potential mechanisms of CLA which contribute to body weight regulation. Adult mice were fed either a normal fat (NF, 12% kJ content as fat) or a high-fat (HF, 43% kJ content as fat) diet. In the latter case, half of the animals received daily oral supplementation of CLA. Expression and content of stomach proteins and specific bacterial populations from caecum were analysed. CLA supplementation was associated with an increase in stomach protein expression, and exerted a prebiotic action on both Bacteroidetes/Prevotella and Akkermansia muciniphila. However, CLA supplementation was not able to override the negative effects of HF diet on Bifidobacterium spp., which was decreased in both HF and HF+CLA groups. Our data show that CLA are able to modulate stomach protein expression and exert a prebiotic effect on specific gut bacterial species.

  16. Regulating the ethylene response of a plant by modulation of F-box proteins

    SciTech Connect

    Guo, Hongwei; Ecker, Joseph R

    2014-01-07

    The relationship between F-box proteins and proteins invovled in the ethylene response in plants is described. In particular, F-box proteins may bind to proteins involved in the ethylene response and target them for degradation by the ubiquitin/proteasome pathway. The transcription factor EIN3 is a key transcription factor mediating ethylne-regulated gene expression and morphological responses. EIN3 is degraded through a ubiquitin/proteasome pathway mediated by F-box proteins EBF1 and EBF2. The link between F-box proteins and the ethylene response is a key step in modulating or regulating the response of a plant to ethylene. Described herein are transgenic plants having an altered sensitivity to ethylene, and methods for making transgenic plant haing an althered sensitivity to ethylene by modulating the level of activity of F-box proteins. Methods of altering the ethylene response in a plant by modulating the activity or expression of an F-box protein are described. Also described are methods of identifying compounds that modulate the ethylene response in plants by modulating the level of F-box protein expression or activity.

  17. A pair of mouse KRAB zinc finger proteins modulates multiple indicators of female reproduction.

    PubMed

    Krebs, Christopher J; Robins, Diane M

    2010-04-01

    Krüppel-associated box-zinc finger proteins (KRAB-ZFPs) are the largest class of transcriptional regulators in mammals, yet few have been assigned biological roles. Cloning the genes underlying the regulator of sex-limitation (rsl) phenotype, in which the normally male-specific sex-limited protein (SLP) is expressed in female mice, identified two KRAB-ZFPs, Rsl1 and Rsl2, as influencing sexually dimorphic liver gene expression. Combined absence of both repressors in rsl mice leads to increased expression in female liver of major urinary proteins (MUPs) and certain enzymes of steroid metabolism, as well as SLP. We hypothesized that this altered gene expression might affect reproductive physiology in rsl females. Urinary MUP (uMUP) concentration varied with the estrous cycle in both wt and rsl females but was consistently higher in rsl urine. A behavioral odor test revealed that wild-type (wt) males preferred rsl to wt females, possibly due to elevated uMUPs providing greater pheromone presentation. To ascribe activity to Rsl1, Rsl2, or both, the genes were individually expressed as liver-specific transgenes. RSL2 overexpression accentuated uMUP fluctuations across the estrous cycle, whereas RSL1 overexpression did not. In addition, puberty onset, as indicated by vaginal opening (VO), occurred 2 days earlier in rsl females, and excess RSL2, but not RSL1, restored VO timing to wt. Hence, transcriptional repression by RSL in liver modifies female mouse reproduction via targets that likely impact both hormonal and pheromonal cues. The large and rapidly diversifying KRAB-ZFP family may modulate biological processes, including reproduction, to confer individual differences that may isolate populations and ultimately lead to speciation.

  18. Keratin 8 absence down-regulates colonocyte HMGCS2 and modulates colonic ketogenesis and energy metabolism

    PubMed Central

    Helenius, Terhi O.; Misiorek, Julia O.; Nyström, Joel H.; Fortelius, Lina E.; Habtezion, Aida; Liao, Jian; Asghar, M. Nadeem; Zhang, Haiyan; Azhar, Salman; Omary, M. Bishr; Toivola, Diana M.

    2015-01-01

    Simple-type epithelial keratins are intermediate filament proteins important for mechanical stability and stress protection. Keratin mutations predispose to human liver disorders, whereas their roles in intestinal diseases are unclear. Absence of keratin 8 (K8) in mice leads to colitis, decreased Na/Cl uptake, protein mistargeting, and longer crypts, suggesting that keratins contribute to intestinal homeostasis. We describe the rate-limiting enzyme of the ketogenic energy metabolism pathway, mitochondrial 3-hydroxy-3-methylglutaryl-CoA synthase 2 (HMGCS2), as a major down-regulated protein in the K8-knockout (K8−/−) colon. K8 absence leads to decreased quantity and activity of HMGCS2, and the down-regulation is not dependent on the inflammatory state, since HMGCS2 is not decreased in dextran sulfate sodium-induced colitis. Peroxisome proliferator–activated receptor α, a transcriptional activator of HMGCS2, is similarly down-regulated. Ketogenic conditions—starvation or ketogenic diet—increase K8+/+ HMGCS2, whereas this response is blunted in the K8−/− colon. Microbiota-produced short-chain fatty acids (SCFAs), substrates in the colonic ketone body pathway, are increased in stool, which correlates with decreased levels of their main transporter, monocarboxylate transporter 1 (MCT1). Microbial populations, including the main SCFA-butyrate producers in the colon, were not altered in the K8−/−. In summary, the regulation of the SCFA-MCT1-HMGCS2 axis is disrupted in K8−/− colonocytes, suggesting a role for keratins in colonocyte energy metabolism and homeostasis. PMID:25904331

  19. Doxorubicin Induces Inflammatory Modulation and Metabolic Dysregulation in Diabetic Skeletal Muscle.

    PubMed

    Supriya, Rashmi; Tam, Bjorn T; Pei, Xiao M; Lai, Christopher W; Chan, Lawrence W; Yung, Benjamin Y; Siu, Parco M

    2016-01-01

    Anti-cancer agent doxorubicin (DOX) has been demonstrated to worsen insulin signaling, engender muscle atrophy, trigger pro-inflammation, and induce a shift to anaerobic glycolytic metabolism in skeletal muscle. The myotoxicity of DOX in diabetic skeletal muscle remains largely unclear. This study examined the effects of DOX on insulin signaling, muscle atrophy, pro-/anti-inflammatory microenvironment, and glycolysis metabolic regulation in skeletal muscle of db/db diabetic and db/+ non-diabetic mice. Non-diabetic db/+ mice and diabetic db/db mice were randomly assigned to the following groups: db/+CON, db/+DOX, db/dbCON, and db/dbDOX. Mice in db/+DOX and db/dbDOX groups were intraperitoneally injected with DOX at a dose of 15 mg per kg body weight whereas mice in db/+CON and db/dbCON groups were injected with the same volume of saline instead of DOX. Gastrocnemius was immediately harvested, weighed, washed with cold phosphate buffered saline, frozen in liquid nitrogen, and stored at -80°C for later analysis. The effects of DOX on diabetic muscle were neither seen in insulin signaling markers (Glut4, pIRS1Ser(636∕639), and pAktSer(473)) nor muscle atrophy markers (muscle mass, MuRF1 and MAFbx). However, DOX exposure resulted in enhancement of pro-inflammatory favoring microenvironment (as indicated by TNF-α, HIFα and pNFκBp65) accompanied by diminution of anti-inflammatory favoring microenvironment (as indicated by IL15, PGC1α and pAMPKβ1Ser108). Metabolism of diabetic muscle was shifted to anaerobic glycolysis after DOX exposure as demonstrated by our analyses of PDK4, LDH and pACCSer(79). Our results demonstrated that there might be a link between inflammatory modulation and the dysregulation of aerobic glycolytic metabolism in DOX-injured diabetic skeletal muscle. These findings help to understand the pathogenesis of DOX-induced myotoxicity in diabetic muscle. PMID:27512375

  20. Doxorubicin Induces Inflammatory Modulation and Metabolic Dysregulation in Diabetic Skeletal Muscle

    PubMed Central

    Supriya, Rashmi; Tam, Bjorn T.; Pei, Xiao M.; Lai, Christopher W.; Chan, Lawrence W.; Yung, Benjamin Y.; Siu, Parco M.

    2016-01-01

    Anti-cancer agent doxorubicin (DOX) has been demonstrated to worsen insulin signaling, engender muscle atrophy, trigger pro-inflammation, and induce a shift to anaerobic glycolytic metabolism in skeletal muscle. The myotoxicity of DOX in diabetic skeletal muscle remains largely unclear. This study examined the effects of DOX on insulin signaling, muscle atrophy, pro-/anti-inflammatory microenvironment, and glycolysis metabolic regulation in skeletal muscle of db/db diabetic and db/+ non-diabetic mice. Non-diabetic db/+ mice and diabetic db/db mice were randomly assigned to the following groups: db/+CON, db/+DOX, db/dbCON, and db/dbDOX. Mice in db/+DOX and db/dbDOX groups were intraperitoneally injected with DOX at a dose of 15 mg per kg body weight whereas mice in db/+CON and db/dbCON groups were injected with the same volume of saline instead of DOX. Gastrocnemius was immediately harvested, weighed, washed with cold phosphate buffered saline, frozen in liquid nitrogen, and stored at −80°C for later analysis. The effects of DOX on diabetic muscle were neither seen in insulin signaling markers (Glut4, pIRS1Ser636∕639, and pAktSer473) nor muscle atrophy markers (muscle mass, MuRF1 and MAFbx). However, DOX exposure resulted in enhancement of pro-inflammatory favoring microenvironment (as indicated by TNF-α, HIFα and pNFκBp65) accompanied by diminution of anti-inflammatory favoring microenvironment (as indicated by IL15, PGC1α and pAMPKβ1Ser108). Metabolism of diabetic muscle was shifted to anaerobic glycolysis after DOX exposure as demonstrated by our analyses of PDK4, LDH and pACCSer79. Our results demonstrated that there might be a link between inflammatory modulation and the dysregulation of aerobic glycolytic metabolism in DOX-injured diabetic skeletal muscle. These findings help to understand the pathogenesis of DOX-induced myotoxicity in diabetic muscle. PMID:27512375

  1. Dietary Proteins as Determinants of Metabolic and Physiologic Functions of the Gastrointestinal Tract

    PubMed Central

    Jahan-Mihan, Alireza; Luhovyy, Bohdan L.; Khoury, Dalia El; Anderson, G. Harvey

    2011-01-01

    Dietary proteins elicit a wide range of nutritional and biological functions. Beyond their nutritional role as the source of amino acids for protein synthesis, they are instrumental in the regulation of food intake, glucose and lipid metabolism, blood pressure, bone metabolism and immune function. The interaction of dietary proteins and their products of digestion with the regulatory functions of the gastrointestinal (GI) tract plays a dominant role in determining the physiological properties of proteins. The site of interaction is widespread, from the oral cavity to the colon. The characteristics of proteins that influence their interaction with the GI tract in a source-dependent manner include their physico-chemical properties, their amino acid composition and sequence, their bioactive peptides, their digestion kinetics and also the non-protein bioactive components conjugated with them. Within the GI tract, these products affect several regulatory functions by interacting with receptors releasing hormones, affecting stomach emptying and GI transport and absorption, transmitting neural signals to the brain, and modifying the microflora. This review discusses the interaction of dietary proteins during digestion and absorption with the physiological and metabolic functions of the GI tract, and illustrates the importance of this interaction in the regulation of amino acid, glucose, lipid metabolism, and food intake. PMID:22254112

  2. Modulation of Glycosaminoglycans Affects PrPSc Metabolism but Does Not Block PrPSc Uptake

    PubMed Central

    Wolf, Hanna; Graßmann, Andrea; Bester, Romina; Hossinger, André; Möhl, Christoph; Paulsen, Lydia; Groschup, Martin H.; Schätzl, Hermann

    2015-01-01

    ABSTRACT Mammalian prions are unconventional infectious agents composed primarily of the misfolded aggregated host prion protein PrP, termed PrPSc. Prions propagate by the recruitment and conformational conversion of cellular prion protein into abnormal prion aggregates on the cell surface or along the endocytic pathway. Cellular glycosaminoglycans have been implicated as the first attachment sites for prions and cofactors for cellular prion replication. Glycosaminoglycan mimetics and obstruction of glycosaminoglycan sulfation affect prion replication, but the inhibitory effects on different strains and different stages of the cell infection have not been thoroughly addressed. We examined the effects of a glycosaminoglycan mimetic and undersulfation on cellular prion protein metabolism, prion uptake, and the establishment of productive infections in L929 cells by two mouse-adapted prion strains. Surprisingly, both treatments reduced endogenous sulfated glycosaminoglycans but had divergent effects on cellular PrP levels. Chemical or genetic manipulation of glycosaminoglycans did not prevent PrPSc uptake, arguing against their roles as essential prion attachment sites. However, both treatments effectively antagonized de novo prion infection independently of the prion strain and reduced PrPSc formation in chronically infected cells. Our results demonstrate that sulfated glycosaminoglycans are dispensable for prion internalization but play a pivotal role in persistently maintained PrPSc formation independent of the prion strain. IMPORTANCE Recently, glycosaminoglycans (GAGs) became the focus of neurodegenerative disease research as general attachment sites for cell invasion by pathogenic protein aggregates. GAGs influence amyloid formation in vitro. GAGs are also found in intra- and extracellular amyloid deposits. In light of the essential role GAGs play in proteinopathies, understanding the effects of GAGs on protein aggregation and aggregate dissemination is crucial

  3. Decrease in age-related tau hyperphosphorylation and cognitive improvement following vitamin D supplementation are associated with modulation of brain energy metabolism and redox state.

    PubMed

    Briones, T L; Darwish, H

    2014-03-14

    In the present study we examined whether vitamin D supplementation can reduce age-related tau hyperphosphorylation and cognitive impairment by enhancing brain energy homeostasis and protein phosphatase 2A (PP2A) activity, and modulating the redox state. Male F344 rats aged 20 months (aged) and 6 months (young) were randomly assigned to either vitamin D supplementation or no supplementation (control). Rats were housed in pairs and the supplementation group (n=10 young and n=10 aged) received subcutaneous injections of vitamin D (1, α25-dihydroxyvitamin D3) for 21 days. Control animals (n=10 young and n=10 aged) received equal volume of normal saline and behavioral testing in the water maze started on day 14 after the initiation of vitamin D supplementation. Tau phosphorylation, markers of brain energy metabolism (ADP/ATP ratio and adenosine monophosphate-activated protein kinase) and redox state (levels of reactive oxygen species, activity of superoxide dismutase, and glutathione levels) as well as PP2A activity were measured in hippocampal tissues. Our results extended previous findings that: (1) tau phosphorylation significantly increased during aging; (2) markers of brain energy metabolism and redox state are significantly decreased in aging; and (3) aged rats demonstrated significant learning and memory impairment. More importantly, we found that age-related changes in brain energy metabolism, redox state, and cognitive function were attenuated by vitamin D supplementation. No significant differences were seen in tau hyperphosphorylation, markers of energy metabolism and redox state in the young animal groups. Our data suggest that vitamin D ameliorated the age-related tau hyperphosphorylation and cognitive decline by enhancing brain energy metabolism, redox state, and PP2A activity making it a potentially useful therapeutic option to alleviate the effects of aging.

  4. Avian acute phase protein ovotransferrin modulates phagocyte function

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Acute phase proteins (APP) are serum proteins elevated in response to a variety of physiological injuries including infection and inflammation. These pathogen nonspecific proteins are predominantly synthesized in the liver and serve as a humoral component of innate immunity by way of recognizing and...

  5. CELLULAR MECHANISMS OF PROTEIN METABOLISM IN THE NEPHRON

    PubMed Central

    Oliver, Jean; MacDowell, Muriel

    1958-01-01

    1. The exaggeration of "hyaline droplet" formation observed in the renal lesion of epidemic hemorrhagic fever when treated with the infusion of large amounts of human serum albumin and the histochemical characteristics of the droplets so formed afford evidence towards their identification with the protein absorption droplets of experimental procedures and with those that occur in other renal diseases. 2. Protein absorption droplets (hyaline droplets) are the visible aspect of pathological modifications of a physiological process; i.e., the continuing reabsorption of plasma proteins by the proximal convolutions. 3. The mitochondria of the renal cells are directly involved in both the physiological and the abnormal reabsorption and disposal of the plasma proteins; the absorption droplets are a complex of reabsorbed proteins and mitochondrial substances and enzymes; they result whenever disposal is at a rate insufficient to prevent accumulation. 4. Failure of intracellular disposal of reabsorbed protein is determined by (a) the quantitative and qualitative characteristics of the protein and (b) by the functional state of renal cell. 5. Various factors in renal disease that result in disturbances of reabsorption and of intracellular disposal, both with and without droplet formation, are described. PMID:13525582

  6. Selective Modulation of MicroRNA Expression with Protein Ingestion Following Concurrent Resistance and Endurance Exercise in Human Skeletal Muscle.

    PubMed

    Camera, Donny M; Ong, Jun N; Coffey, Vernon G; Hawley, John A

    2016-01-01

    We examined changes in the expression of 13 selected skeletal muscle microRNAs (miRNAs) implicated in exercise adaptation responses following a single bout of concurrent exercise. In a randomized cross-over design, seven healthy males undertook a single trial consisting of resistance exercise (8 × 5 leg extension, 80% 1 Repetition Maximum) followed by cycling (30 min at ~70% VO2peak) with either post-exercise protein (PRO: 25 g whey protein) or placebo (PLA) ingestion. Muscle biopsies (vastus lateralis) were obtained at rest and 4 h post-exercise. Detection of miRNA via quantitative Polymerase Chain Reaction (qPCR) revealed post-exercise increases in miR-23a-3p (~90%), miR-23b-3p (~39%), miR-133b (~80%), miR-181-5p (~50%), and miR-378-5p (~41%) at 4 h post-exercise with PRO that also resulted in higher abundance compared to PLA (P < 0.05). There was a post-exercise decrease in miR-494-3p abundance in PLA only (~88%, P < 0.05). There were no changes in the total abundance of target proteins post-exercise or between conditions. Protein ingestion following concurrent exercise can modulate the expression of miRNAs implicated in exercise adaptations compared to placebo. The selective modulation of miRNAs with target proteins that may prioritize myogenic compared to oxidative/metabolic adaptive responses indicate that miRNAs can play a regulatory role in the molecular machinery enhancing muscle protein synthesis responses with protein ingestion following concurrent exercise. PMID:27014087

  7. Selective Modulation of MicroRNA Expression with Protein Ingestion Following Concurrent Resistance and Endurance Exercise in Human Skeletal Muscle

    PubMed Central

    Camera, Donny M.; Ong, Jun N.; Coffey, Vernon G.; Hawley, John A.

    2016-01-01

    We examined changes in the expression of 13 selected skeletal muscle microRNAs (miRNAs) implicated in exercise adaptation responses following a single bout of concurrent exercise. In a randomized cross-over design, seven healthy males undertook a single trial consisting of resistance exercise (8 × 5 leg extension, 80% 1 Repetition Maximum) followed by cycling (30 min at ~70% VO2peak) with either post-exercise protein (PRO: 25 g whey protein) or placebo (PLA) ingestion. Muscle biopsies (vastus lateralis) were obtained at rest and 4 h post-exercise. Detection of miRNA via quantitative Polymerase Chain Reaction (qPCR) revealed post-exercise increases in miR-23a-3p (~90%), miR-23b-3p (~39%), miR-133b (~80%), miR-181-5p (~50%), and miR-378-5p (~41%) at 4 h post-exercise with PRO that also resulted in higher abundance compared to PLA (P < 0.05). There was a post-exercise decrease in miR-494-3p abundance in PLA only (~88%, P < 0.05). There were no changes in the total abundance of target proteins post-exercise or between conditions. Protein ingestion following concurrent exercise can modulate the expression of miRNAs implicated in exercise adaptations compared to placebo. The selective modulation of miRNAs with target proteins that may prioritize myogenic compared to oxidative/metabolic adaptive responses indicate that miRNAs can play a regulatory role in the molecular machinery enhancing muscle protein synthesis responses with protein ingestion following concurrent exercise. PMID:27014087

  8. Sulfur alleviates arsenic toxicity by reducing its accumulation and modulating proteome, amino acids and thiol metabolism in rice leaves

    NASA Astrophysics Data System (ADS)

    Dixit, Garima; Singh, Amit Pal; Kumar, Amit; Dwivedi, Sanjay; Deeba, Farah; Kumar, Smita; Suman, Shankar; Adhikari, Bijan; Shukla, Yogeshwar; Trivedi, Prabodh Kumar; Pandey, Vivek; Tripathi, Rudra Deo

    2015-11-01

    Arsenic (As) contamination of water is a global concern and rice consumption is the biggest dietary exposure to human posing carcinogenic risks, predominantly in Asia. Sulfur (S) is involved in di-sulfide linkage in many proteins and plays crucial role in As detoxification. Present study explores role of variable S supply on rice leaf proteome, its inclination towards amino acids (AA) profile and non protein thiols under arsenite exposure. Analysis of 282 detected proteins on 2-DE gel revealed 113 differentially expressed proteins, out of which 80 were identified by MALDI-TOF-TOF. The identified proteins were mostly involved in glycolysis, TCA cycle, AA biosynthesis, photosynthesis, protein metabolism, stress and energy metabolism. Among these, glycolytic enzymes play a major role in AA biosynthesis that leads to change in AAs profiling. Proteins of glycolytic pathway, photosynthesis and energy metabolism were also validated by western blot analysis. Conclusively S supplementation reduced the As accumulation in shoot positively skewed thiol metabolism and glycolysis towards AA accumulation under AsIII stress.

  9. Sulfur alleviates arsenic toxicity by reducing its accumulation and modulating proteome, amino acids and thiol metabolism in rice leaves.

    PubMed

    Dixit, Garima; Singh, Amit Pal; Kumar, Amit; Dwivedi, Sanjay; Deeba, Farah; Kumar, Smita; Suman, Shankar; Adhikari, Bijan; Shukla, Yogeshwar; Trivedi, Prabodh Kumar; Pandey, Vivek; Tripathi, Rudra Deo

    2015-01-01

    Arsenic (As) contamination of water is a global concern and rice consumption is the biggest dietary exposure to human posing carcinogenic risks, predominantly in Asia. Sulfur (S) is involved in di-sulfide linkage in many proteins and plays crucial role in As detoxification. Present study explores role of variable S supply on rice leaf proteome, its inclination towards amino acids (AA) profile and non protein thiols under arsenite exposure. Analysis of 282 detected proteins on 2-DE gel revealed 113 differentially expressed proteins, out of which 80 were identified by MALDI-TOF-TOF. The identified proteins were mostly involved in glycolysis, TCA cycle, AA biosynthesis, photosynthesis, protein metabolism, stress and energy metabolism. Among these, glycolytic enzymes play a major role in AA biosynthesis that leads to change in AAs profiling. Proteins of glycolytic pathway, photosynthesis and energy metabolism were also validated by western blot analysis. Conclusively S supplementation reduced the As accumulation in shoot positively skewed thiol metabolism and glycolysis towards AA accumulation under AsIII stress. PMID:26552588

  10. Sulfur alleviates arsenic toxicity by reducing its accumulation and modulating proteome, amino acids and thiol metabolism in rice leaves

    PubMed Central

    Dixit, Garima; Singh, Amit Pal; Kumar, Amit; Dwivedi, Sanjay; Deeba, Farah; Kumar, Smita; Suman, Shankar; Adhikari, Bijan; Shukla, Yogeshwar; Trivedi, Prabodh Kumar; Pandey, Vivek; Tripathi, Rudra Deo

    2015-01-01

    Arsenic (As) contamination of water is a global concern and rice consumption is the biggest dietary exposure to human posing carcinogenic risks, predominantly in Asia. Sulfur (S) is involved in di-sulfide linkage in many proteins and plays crucial role in As detoxification. Present study explores role of variable S supply on rice leaf proteome, its inclination towards amino acids (AA) profile and non protein thiols under arsenite exposure. Analysis of 282 detected proteins on 2-DE gel revealed 113 differentially expressed proteins, out of which 80 were identified by MALDI-TOF-TOF. The identified proteins were mostly involved in glycolysis, TCA cycle, AA biosynthesis, photosynthesis, protein metabolism, stress and energy metabolism. Among these, glycolytic enzymes play a major role in AA biosynthesis that leads to change in AAs profiling. Proteins of glycolytic pathway, photosynthesis and energy metabolism were also validated by western blot analysis. Conclusively S supplementation reduced the As accumulation in shoot positively skewed thiol metabolism and glycolysis towards AA accumulation under AsIII stress. PMID:26552588

  11. Labeling Cell Surface GPIs and GPI-Anchored Proteins through Metabolic Engineering with Artificial Inositol Derivatives.

    PubMed

    Lu, Lili; Gao, Jian; Guo, Zhongwu

    2015-08-10

    Glycosylphosphatidylinositol (GPI) anchoring of proteins to the cell surface is important for various biological processes, but GPI-anchored proteins are difficult to study. An effective strategy was developed for the metabolic engineering of cell-surface GPIs and GPI-anchored proteins by using inositol derivatives carrying an azido group. The azide-labeled GPIs and GPI-anchored proteins were then tagged with biotin on live cells through a click reaction, which allows further elaboration with streptavidin-conjugated dyes or other molecules. The strategy can be used to label GPI-anchored proteins with various tags for biological studies.

  12. Proteins involved in flor yeast carbon metabolism under biofilm formation conditions.

    PubMed

    Moreno-García, Jaime; García-Martínez, Teresa; Moreno, Juan; Mauricio, Juan Carlos

    2015-04-01

    A lack of sugars during the production of biologically aged wines after fermentation of grape must causes flor yeasts to metabolize other carbon molecules formed during fermentation (ethanol and glycerol, mainly). In this work, a proteome analysis involving OFFGEL fractionation prior to LC/MS detection was used to elucidate the carbon metabolism of a flor yeast strain under biofilm formation conditions (BFC). The results were compared with those obtained under non-biofilm formation conditions (NBFC). Proteins associated to processes such as non-fermentable carbon uptake, the glyoxylate and TCA cycles, cellular respiration and inositol metabolism were detected at higher concentrations under BFC than under the reference conditions (NBFC). This study constitutes the first attempt at identifying the flor yeast proteins responsible for the peculiar sensory profile of biologically aged wines. A better metabolic knowledge of flor yeasts might facilitate the development of effective strategies for improved production of these special wines.

  13. Proteins involved in flor yeast carbon metabolism under biofilm formation conditions.

    PubMed

    Moreno-García, Jaime; García-Martínez, Teresa; Moreno, Juan; Mauricio, Juan Carlos

    2015-04-01

    A lack of sugars during the production of biologically aged wines after fermentation of grape must causes flor yeasts to metabolize other carbon molecules formed during fermentation (ethanol and glycerol, mainly). In this work, a proteome analysis involving OFFGEL fractionation prior to LC/MS detection was used to elucidate the carbon metabolism of a flor yeast strain under biofilm formation conditions (BFC). The results were compared with those obtained under non-biofilm formation conditions (NBFC). Proteins associated to processes such as non-fermentable carbon uptake, the glyoxylate and TCA cycles, cellular respiration and inositol metabolism were detected at higher concentrations under BFC than under the reference conditions (NBFC). This study constitutes the first attempt at identifying the flor yeast proteins responsible for the peculiar sensory profile of biologically aged wines. A better metabolic knowledge of flor yeasts might facilitate the development of effective strategies for improved production of these special wines. PMID:25475262

  14. Myocardial oxidative metabolism and protein synthesis during mechanical circulatory support by extracorporeal membrane oxygenation.

    PubMed

    Priddy, Colleen M O'Kelly; Kajimoto, Masaki; Ledee, Dolena R; Bouchard, Bertrand; Isern, Nancy; Olson, Aaron K; Des Rosiers, Christine; Portman, Michael A

    2013-02-01

    Extracorporeal membrane oxygenation (ECMO) provides essential mechanical circulatory support necessary for survival in infants and children with acute cardiac decompensation. However, ECMO also causes metabolic disturbances, which contribute to total body wasting and protein loss. Cardiac stunning can also occur, which prevents ECMO weaning, and contributes to high mortality. The heart may specifically undergo metabolic impairments, which influence functional recovery. We tested the hypothesis that ECMO alters oxidative metabolism and protein synthesis. We focused on the amino acid leucine and integration with myocardial protein synthesis. We used a translational immature swine model in which we assessed in heart 1) the fractional contribution of leucine (FcLeucine) and pyruvate to mitochondrial acetyl-CoA formation by nuclear magnetic resonance and 2) global protein fractional synthesis (FSR) by gas chromatography-mass spectrometry. Immature mixed breed Yorkshire male piglets (n = 22) were divided into four groups based on loading status (8 h of normal circulation or ECMO) and intracoronary infusion [(13)C(6),(15)N]-L-leucine (3.7 mM) alone or with [2-(13)C]-pyruvate (7.4 mM). ECMO decreased pulse pressure and correspondingly lowered myocardial oxygen consumption (∼40%, n = 5), indicating decreased overall mitochondrial oxidative metabolism. However, FcLeucine was maintained and myocardial protein FSR was marginally increased. Pyruvate addition decreased tissue leucine enrichment, FcLeucine, and Fc for endogenous substrates as well as protein FSR. The heart under ECMO shows reduced oxidative metabolism of substrates, including amino acids, while maintaining 1) metabolic flexibility indicated by ability to respond to pyruvate and 2) a normal or increased capacity for global protein synthesis.

  15. Energizing eukaryotic cell-free protein synthesis with glucose metabolism.

    PubMed

    Anderson, Mark J; Stark, Jessica C; Hodgman, C Eric; Jewett, Michael C

    2015-07-01

    Eukaryotic cell-free protein synthesis (CFPS) is limited by the dependence on costly high-energy phosphate compounds and exogenous enzymes to power protein synthesis (e.g., creatine phosphate and creatine kinase, CrP/CrK). Here, we report the ability to use glucose as a secondary energy substrate to regenerate ATP in a Saccharomyces cerevisiae crude extract CFPS platform. We observed synthesis of 3.64±0.35 μg mL(-1) active luciferase in batch reactions with 16 mM glucose and 25 mM phosphate, resulting in a 16% increase in relative protein yield (μg protein/$ reagents) compared to the CrP/CrK system. Our demonstration provides the foundation for development of cost-effective eukaryotic CFPS platforms.

  16. Metabolic profiling of chickpea-Fusarium interaction identifies differential modulation of disease resistance pathways.

    PubMed

    Kumar, Yashwant; Dholakia, Bhushan B; Panigrahi, Priyabrata; Kadoo, Narendra Y; Giri, Ashok P; Gupta, Vidya S

    2015-08-01

    Chickpea is the third most widely grown legume in the world and mainly used as a vegetarian source of human dietary protein. Fusarium wilt, caused by Fusarium oxysporum f. sp. ciceri (Foc), is one of the major threats to global chickpea production. Host resistance is the best way to protect crops from diseases; however, in spite of using various approaches, the mechanism of Foc resistance in chickpea remains largely obscure. In the present study, non-targeted metabolic profiling at several time points of resistant and susceptible chickpea cultivars using high-resolution liquid chromatography-mass spectrometry was applied to better understand the mechanistic basis of wilt resistance or susceptibility. Multivariate analysis of the data (OPLS-DA) revealed discriminating metabolites in chickpea root tissue after Foc inoculation such as flavonoids, isoflavonoids, alkaloids, amino acids and sugars. Foc inoculated resistant plants had more flavonoids and isoflavonoids along with their malonyl conjugates. Many antifungal metabolites that were induced after Foc infection viz., aurantion-obstine β-glucosides and querecitin were elevated in resistant cultivar. Overall, diverse genetic and biochemical mechanisms were operational in the resistant cultivar for Foc defense as compared to the susceptible plant. The resistant chickpea plants employed the above-mentioned metabolic pathways as potential defense strategy against Foc.

  17. Antisense Mediated Splicing Modulation For Inherited Metabolic Diseases: Challenges for Delivery

    PubMed Central

    Pérez, Belen; Vilageliu, Lluisa; Grinberg, Daniel

    2014-01-01

    In the past few years, research in targeted mutation therapies has experienced significant advances, especially in the field of rare diseases. In particular, the efficacy of antisense therapy for suppression of normal, pathogenic, or cryptic splice sites has been demonstrated in cellular and animal models and has already reached the clinical trials phase for Duchenne muscular dystrophy. In different inherited metabolic diseases, splice switching oligonucleotides (SSOs) have been used with success in patients' cells to force pseudoexon skipping or to block cryptic splice sites, in both cases recovering normal transcript and protein and correcting the enzyme deficiency. However, future in vivo studies require individual approaches for delivery depending on the gene defect involved, given the different patterns of tissue and organ expression. Herein we review the state of the art of antisense therapy targeting RNA splicing in metabolic diseases, grouped according to their expression patterns—multisystemic, hepatic, or in central nervous system (CNS)—and summarize the recent progress achieved in the field of in vivo delivery of oligonucleotides to each organ or system. Successful body-wide distribution of SSOs and preferential distribution in the liver after systemic administration have been reported in murine models for different diseases, while for CNS limited data are available, although promising results with intratechal injections have been achieved. PMID:24506780

  18. Streptococcus mutans copes with heat stress by multiple transcriptional regulons modulating virulence and energy metabolism

    PubMed Central

    Liu, Chengcheng; Niu, Yulong; Zhou, Xuedong; Zheng, Xin; Wang, Shida; Guo, Qiang; Li, Yuqing; Li, Mingyun; Li, Jiyao; Yang, Yi; Ding, Yi; Lamont, Richard J.; Xu, Xin

    2015-01-01

    Dental caries is closely associated with the virulence of Streptococcus mutans. The virulence expression of S. mutans is linked to its stress adaptation to the changes in the oral environment. In this work we used whole-genome microarrays to profile the dynamic transcriptomic responses of S. mutans during physiological heat stress. In addition, we evaluated the phenotypic changes, including, eDNA release, initial biofilm formation, extracellular polysaccharides generation, acid production/acid tolerance, and ATP turnover of S. mutans during heat stress. There were distinct patterns observed in the way that S. mutans responded to heat stress that included 66 transcription factors for the expression of functional genes being differentially expressed. Especially, response regulators of two component systems (TCSs), the repressors of heat shock proteins and regulators involved in sugar transporting and metabolism co-ordinated to enhance the cell’s survival and energy generation against heat stress in S. mutans. PMID:26251057

  19. Hepatocellular carcinoma protein carbonylation in virus C and metabolic syndrome patients.

    PubMed

    Martin, Fernando Ariel; Mebarki, Mouniya; Paradis, Valérie; Friguet, Bertrand; Radman, Miroslav

    2014-10-01

    Metabolic syndrome (MS) is becoming the leading cause of chronic liver diseases worldwide. Hepatocellular carcinoma (HCC) development in MS is peculiar compared to other chronic liver diseases. Carbohydrate and lipid metabolic imbalance in MS increase reactive oxygen species damaging proteins. In the present work we study the difference in protein oxidative damage (carbonylation) in human HCC derived from virus C infection (VHC) and from MS (MS_HCC) as the only subjacent cause. We selected a patient cohort containing of 10 non-tumoral and 10 tumoral liver resections in each study group (virus C and MS HCC) based on clinical patient history and histological parameters. Protein samples were labeled to saturation using CF 647-hydrazide™ dye. This approach allows us to perform carbonyl detection alongside with a DIGE experiment. We detected a total of 1184 spots with 36 differentially expressed proteins and 47 spots differentially carbonylated between VHC and MS_HCC (fold change >1.5, p<0.05). VHC up-regulated proteins are involved in signaling pathways related to cancer development such as signaling by EGFR, Wnt, Cdc20 and cell cycle. Further, up-regulated proteins in MS HCC, are implicated in metabolism of carbohydrates and amino acids. Differential carbonylation analysis between VHC and MS_HCC showed protein damage in proteins such as glucose phosphate isomerase, isocitrate dehydrogenase, and 3-ketoacyl-CoA thiolase. Higher protein carbonylation in MS_HCC samples was observed in proteins involved in redox response and lipid metabolism. In conclusion, the observed difference in protein oxidative damage between MS and Virus C derived carcinoma could account for the different cancer development pathway. PMID:26461368

  20. Cardioselective Dominant-negative Thyroid Hormone Receptor (Δ337T) Modulates Myocardial Metabolism and Contractile Dfficiency

    SciTech Connect

    Hyyti, Outi M.; Olson, Aaron; Ge, Ming; Ning, Xue-Han; Buroker, Norman E.; Chung, Youngran; Jue, Thomas; Portman, Michael A.

    2008-06-03

    Dominant- negative thyroid hormone receptors (TRs) show elevated expression relative to ligand-binding TRs during cardiac hypertrophy. We tested the hypothesis that overexpression of a dominant-negative TR alters cardiac metabolism and contractile efficiency (CE). We used mice expressing the cardioselective dominant-negative TRβ1 mutation Δ337T. Isolated working Δ337T hearts and nontransgenic control (Con) hearts were perfused with 13C-labeled free fatty acids (FFA), acetoacetate (ACAC), lactate, and glucose at physiological concentrations for 30 min. 13C NMR spectroscopy and isotopomer analyses were used to determine substrate flux and fractional contributions (Fc) of acetyl-CoA to the citric acid cycle (CAC). Δ337T hearts exhibited rate depression but higher developed pressure and CE, defined as work per oxygen consumption (MV˙ O2). Unlabeled substrate Fc from endogenous sources was higher in Δ337T, but ACAC Fc was lower. Fluxes through CAC, lactate, ACAC, and FFA were reduced in Δ337T. CE and Fc differences were reversed by pacing Δ337T to Con rates, accompanied by an increase in FFA Fc. Δ337T hearts lacked the ability to increase MV˙ O2. Decreases in protein expression for glucose transporter-4 and hexokinase-2 and increases in pyruvate dehydrogenase kinase-2 and -4 suggest that these hearts are unable to increase carbohydrate oxidation in response to stress. These data show that Δ337T alters the metabolic phenotype in murine heart by reducing substrate flux for multiple pathways. Some of these changes are heart rate dependent, indicating that the substrate shift may represent an accommodation to altered contractile protein kinetics, which can be disrupted by pacing stress.

  1. Surface proteins of Bordetella pertussis: comparison of virulent and avirulent strains and effects of phenotypic modulation.

    PubMed Central

    Armstrong, S K; Parker, C D

    1986-01-01

    The surface proteins of several Bordetella strains and their modulated derivatives were examined by surface radioiodination, cell fractionation, and Western blotting. A surface protein with a high Mr, missing in a mutant lacking the filamentous hemagglutinin, was identified in virulent Bordetella pertussis and Bordetella parapertussis cells and was absent in avirulent B. pertussis strains. The electrophoretic profiles of lipopolysaccharide and the 40,000-Mr anion-selective porin were not determinants which correlated with phase variation or phenotypic modulation. At least three envelope proteins (91,000, 32,000, and 30,000 molecular weight) were found only in virulent B. pertussis strains and were absent or diminished in the avirulent phase and most phenotypically modulated strains. Two transposon-induced mutants unable to produce hemolysin, dermonecrotic toxin, pertussis toxin, and filamentous hemagglutinin also lacked these three envelope proteins, confirming that virulence-associated envelope proteins were genetically regulated with other virulence-associated traits. Images PMID:2876957

  2. 2P2Idb v2: update of a structural database dedicated to orthosteric modulation of protein-protein interactions.

    PubMed

    Basse, Marie-Jeanne; Betzi, Stéphane; Morelli, Xavier; Roche, Philippe

    2016-01-01

    2P2Idb is a hand-curated structural database dedicated to protein-protein interactions with known small molecule orthosteric modulators. It compiles the structural information related to orthosteric inhibitors and their target [i.e. related 3D structures available in the RCSB Protein Data Bank (PDB)] and provides links to other useful databases. 2P2Idb includes all interactions for which both the protein-protein and protein-inhibitor complexes have been structurally characterized. Since its first release in 2010, the database has grown constantly and the current version contains 27 protein-protein complexes and 274 protein-inhibitor complexes corresponding to 242 unique small molecule inhibitors which represent almost a 5-fold increase compared to the previous version. A number of new data have been added, including new protein-protein complexes, binding affinities, molecular descriptors, precalculated interface parameters and links to other webservers. A new query tool has been implemented to search for inhibitors within the database using standard molecular descriptors. A novel version of the 2P2I-inspector tool has been implemented to calculate a series of physical and chemical parameters of the protein interfaces. Several geometrical parameters including planarity, eccentricity and circularity have been added as well as customizable distance cutoffs. This tool has also been extended to protein-ligand interfaces. The 2P2I database thus represents a wealth of structural source of information for scientists interested in the properties of protein-protein interactions and the design of protein-protein interaction modulators. Database URL: http://2p2idb.cnrs-mrs.fr.

  3. Modulation by estrogen of synthesis of specific uterine proteins.

    PubMed

    Skipper, J K; Eakle, S D; Hamilton, T H

    1980-11-01

    The contemporary procedure for high resolution two dimensional gel electrophoresis was extended to include an initial nondenaturing dimension of electrophoresis. Use of the resulting three dimensional procedure revealed that the previously described single peak of estrogen-induced protein in the uterus of the rat contains at least three distinct proteins whose rates of synthesis are regulated by estrogen. These proteins were localized within partial protein maps, thereby providing definitive operational definitions for the detection and identification of each. It was unambiguously demonstrated that each of the three proteins is continuously synthesized in control uteri. These findings cast doubt on the simplistic hypothesis that estrogen induces a single key protein that triggers a "cascade" of sequential transcriptional events in the uterus. Our finding that the major uterine protein induced by estrogen is also synthesized in liver and muscle cells is significant in that it points to a more general cellular function for the protein, rather than a unique role within uterine cells. Finally, our procedure for three dimensional gel electrophoresis opens new avenues for the detection of minor proteins in heterogeneous protein mixtures, such as those from the tissues of higher animals. PMID:7428041

  4. Effect of Prolonged Simulated Microgravity on Metabolic Proteins in Rat Hippocampus: Steps toward Safe Space Travel.

    PubMed

    Wang, Yun; Javed, Iqbal; Liu, Yahui; Lu, Song; Peng, Guang; Zhang, Yongqian; Qing, Hong; Deng, Yulin

    2016-01-01

    Mitochondria are not only the main source of energy in cells but also produce reactive oxygen species (ROS), which result in oxidative stress when in space. This oxidative stress is responsible for energy imbalances and cellular damage. In this study, a rat tail suspension model was used in individual experiments for 7 and 21 days to explore the effect of simulated microgravity (SM) on metabolic proteins in the hippocampus, a vital brain region involved in learning, memory, and navigation. A comparative (18)O-labeled quantitative proteomic strategy was used to observe the differential expression of metabolic proteins. Forty-two and sixty-seven mitochondrial metabolic proteins were differentially expressed after 21 and 7 days of SM, respectively. Mitochondrial Complex I, III, and IV, isocitrate dehydrogenase and malate dehydrogenase were down-regulated. Moreover, DJ-1 and peroxiredoxin 6, which defend against oxidative damage, were up-regulated in the hippocampus. Western blot analysis of proteins DJ-1 and COX 5A confirmed the mass spectrometry results. Despite these changes in mitochondrial protein expression, no obvious cell apoptosis was observed after 21 days of SM. The results of this study indicate that the oxidative stress induced by SM has profound effects on metabolic proteins.

  5. Amino Acid Flux from Metabolic Network Benefits Protein Translation: the Role of Resource Availability

    PubMed Central

    Hu, Xiao-Pan; Yang, Yi; Ma, Bin-Guang

    2015-01-01

    Protein translation is a central step in gene expression and affected by many factors such as codon usage bias, mRNA folding energy and tRNA abundance. Despite intensive previous studies, how metabolic amino acid supply correlates with protein translation efficiency remains unknown. In this work, we estimated the amino acid flux from metabolic network for each protein in Escherichia coli and Saccharomyces cerevisiae by using Flux Balance Analysis. Integrated with the mRNA expression level, protein abundance and ribosome profiling data, we provided a detailed description of the role of amino acid supply in protein translation. Our results showed that amino acid supply positively correlates with translation efficiency and ribosome density. Moreover, with the rank-based regression model, we found that metabolic amino acid supply facilitates ribosome utilization. Based on the fact that the ribosome density change of well-amino-acid-supplied genes is smaller than poorly-amino-acid-supply genes under amino acid starvation, we reached the conclusion that amino acid supply may buffer ribosome density change against amino acid starvation and benefit maintaining a relatively stable translation environment. Our work provided new insights into the connection between metabolic amino acid supply and protein translation process by revealing a new regulation strategy that is dependent on resource availability. PMID:26056817

  6. Quantitative H2S-mediated protein sulfhydration reveals metabolic reprogramming during the integrated stress response.

    PubMed

    Gao, Xing-Huang; Krokowski, Dawid; Guan, Bo-Jhih; Bederman, Ilya; Majumder, Mithu; Parisien, Marc; Diatchenko, Luda; Kabil, Omer; Willard, Belinda; Banerjee, Ruma; Wang, Benlian; Bebek, Gurkan; Evans, Charles R; Fox, Paul L; Gerson, Stanton L; Hoppel, Charles L; Liu, Ming; Arvan, Peter; Hatzoglou, Maria

    2015-01-01

    The sulfhydration of cysteine residues in proteins is an important mechanism involved in diverse biological processes. We have developed a proteomics approach to quantitatively profile the changes of sulfhydrated cysteines in biological systems. Bioinformatics analysis revealed that sulfhydrated cysteines are part of a wide range of biological functions. In pancreatic β cells exposed to endoplasmic reticulum (ER) stress, elevated H2S promotes the sulfhydration of enzymes in energy metabolism and stimulates glycolytic flux. We propose that transcriptional and translational reprogramming by the integrated stress response (ISR) in pancreatic β cells is coupled to metabolic alternations triggered by sulfhydration of key enzymes in intermediary metabolism. PMID:26595448

  7. Mitochondrial Localization of Telomeric Protein TIN2 Links Telomere Regulation to Metabolic Control

    PubMed Central

    Chen, Liuh-Yow; Zhang, Yi; Zhang, Qinfen; Li, Hongzhi; Luo, Zhenhua; Fang, Hezhi; Kim, Sok Ho; Qin, Li; Yotnda, Patricia; Xu, Jianmin; Tu, Benjamin P.; Bai, Yidong; Songyang, Zhou

    2012-01-01

    Summary Both mitochondria, which are metabolic powerhouses, and telomeres, which help maintain genomic stability, have been implicated in cancer and aging. However, the signaling events that connect these two cellular structures remain poorly understood. Here we report that the canonical telomeric protein TIN2 is also a regulator of metabolism. TIN2 is recruited to telomeres and associates with multiple telomere regulators including TPP1. TPP1 interacts with TIN2 N-terminus, which contains overlapping mitochondrial and telomeric targeting sequences, and controls TIN2 localization. We have found that TIN2 is post-translationally processed in mitochondria, and regulates mitochondria oxidative phosphorylation. Reducing TIN2 expression by RNAi knockdown inhibited glycolysis and reactive oxygen species (ROS) and production, and enhanced ATP levels and oxygen consumption in cancer cells. These results suggest a link between telomeric proteins and metabolic control, providing an additional mechanism by which telomeric proteins regulate cancer and aging. PMID:22885005

  8. Protein deacetylation by SIRT1: an emerging key post-translational modification in metabolic regulation.

    PubMed

    Yu, Jiujiu; Auwerx, Johan

    2010-07-01

    The biological function of most proteins relies on reversible post-translational modifications, among which phosphorylation is most prominently studied and well recognized. Recently, a growing amount of evidence indicates that acetylation-deacetylation reactions, when applied to crucial mediators, can also robustly affect the function of target proteins and thereby have wide-ranging physiological impacts. Sirtuin 1 (SIRT1), which functions as a nicotinamide adenine dinucleotide (NAD(+))-dependent protein deacetylase, deacetylates a wide variety of metabolic molecules in response to the cellular energy and redox status and as such causes significant changes in metabolic homeostasis. This review surveys the evidence for the emerging role of SIRT1-mediated deacetylation in the control of metabolic homeostasis. PMID:20026274

  9. Exosome engineering for efficient intracellular delivery of soluble proteins using optically reversible protein–protein interaction module

    PubMed Central

    Yim, Nambin; Ryu, Seung-Wook; Choi, Kyungsun; Lee, Kwang Ryeol; Lee, Seunghee; Choi, Hojun; Kim, Jeongjin; Shaker, Mohammed R.; Sun, Woong; Park, Ji-Ho; Kim, Daesoo; Do Heo, Won; Choi, Chulhee

    2016-01-01

    Nanoparticle-mediated delivery of functional macromolecules is a promising method for treating a variety of human diseases. Among nanoparticles, cell-derived exosomes have recently been highlighted as a new therapeutic strategy for the in vivo delivery of nucleotides and chemical drugs. Here we describe a new tool for intracellular delivery of target proteins, named ‘exosomes for protein loading via optically reversible protein–protein interactions' (EXPLORs). By integrating a reversible protein–protein interaction module controlled by blue light with the endogenous process of exosome biogenesis, we are able to successfully load cargo proteins into newly generated exosomes. Treatment with protein-loaded EXPLORs is shown to significantly increase intracellular levels of cargo proteins and their function in recipient cells in vitro and in vivo. These results clearly indicate the potential of EXPLORs as a mechanism for the efficient intracellular transfer of protein-based therapeutics into recipient cells and tissues. PMID:27447450

  10. Astrocyte arachidonate and palmitate uptake and metabolism is differentially modulated by dibutyryl-cAMP treatment.

    PubMed

    Seeger, D R; Murphy, C C; Murphy, E J

    2016-07-01

    Astrocytes play a vital role in brain lipid metabolism; however the impact of the phenotypic shift in astrocytes to a reactive state on arachidonic acid metabolism is unknown. Therefore, we determined the impact of dibutyryl-cAMP (dBcAMP) treatment on radiolabeled arachidonic acid ([1-(14)C]20:4n-6) and palmitic acid ([1-(14)C]16:0) uptake and metabolism in primary cultured murine cortical astrocytes. In dBcAMP treated astrocytes, total [1-(14)C]20:4n-6 uptake was increased 1.9-fold compared to control, while total [1-(14)C]16:0 uptake was unaffected. Gene expression of long-chain acyl-CoA synthetases (Acsl), acyl-CoA hydrolase (Acot7), fatty acid binding protein(s) (Fabp) and alpha-synuclein (Snca) were determined using qRT-PCR. dBcAMP treatment increased expression of Acsl3 (4.8-fold) and Acsl4 (1.3-fold), which preferentially use [1-(14)C]20:4n-6 and are highly expressed in astrocytes, consistent with the increase in [1-(14)C]20:4n-6 uptake. However, expression of Fabp5 and Fabp7 were significantly reduced by 25% and 45%, respectively. Acot7 (20%) was also reduced, suggesting dBcAMP treatment favors acyl-CoA formation. dBcAMP treatment enhanced [1-(14)C]20:4n-6 (2.2-fold) and [1-(14)C]16:0 (1.6-fold) esterification into total phospholipids, but the greater esterification of [1-(14)C]20:4n-6 is consistent with the observed uptake through increased Acsl, but not Fabp expression. Although total [1-(14)C]16:0 uptake was not affected, there was a dramatic decrease in [1-(14)C]16:0 in the free fatty acid pool as esterification into the phospholipid pool was increased, which is consistent with the increase in Acsl3 and Acsl4 expression. In summary, our data demonstrates that dBcAMP treatment increases [1-(14)C]20:4n-6 uptake in astrocytes and this increase appears to be due to increased expression of Acsl3 and Acsl4 coupled with a reduction in Acot7 expression. PMID:27255639

  11. HMGA proteins as modulators of chromatin structure during transcriptional activation

    PubMed Central

    Ozturk, Nihan; Singh, Indrabahadur; Mehta, Aditi; Braun, Thomas; Barreto, Guillermo

    2013-01-01

    High mobility group (HMG) proteins are the most abundant non-histone chromatin associated proteins. HMG proteins bind to DNA and nucleosome and alter the structure of chromatin locally and globally. Accessibility to DNA within chromatin is a central factor that affects DNA-dependent nuclear processes, such as transcription, replication, recombination, and repair. HMG proteins associate with different multi-protein complexes to regulate these processes by mediating accessibility to DNA. HMG proteins can be subdivided into three families: HMGA, HMGB, and HMGN. In this review, we will focus on recent advances in understanding the function of HMGA family members, specifically their role in gene transcription regulation during development and cancer. PMID:25364713

  12. Small Molecule Control of Intracellular Protein Levels Through Modulation of the Ubiquitin Proteasome System

    PubMed Central

    Buckley, Dennis L.

    2015-01-01

    Traditionally, biological probes and drugs have targeted the activities of proteins (such as enzymes and receptors) that can be easily controlled by small molecules. The remaining majority of the proteome has been deemed “undruggable”. By using small molecule modulators of the ubiquitin proteasome, protein levels, rather than protein activities can be targeted instead, increasing the number of druggable targets. While targeting the proteasome itself can lead to a global increase in protein levels, targeting other components of the UPS (e.g., the hundreds of E3 ubiquitin ligases) can lead to an increase in protein levels in a more targeted fashion. Alternatively, multiple strategies for inducing protein degradation with small molecule probes are emerging. With the ability to induce and inhibit the degradation of targeted proteins, small molecule modulators of the UPS have the potential to significantly expand the druggable portion of the proteome beyond traditional targets such as enzymes and receptors. PMID:24459094

  13. Direct Modulation of Heterotrimeric G Protein-coupled Signaling by a Receptor Kinase Complex.

    PubMed

    Tunc-Ozdemir, Meral; Urano, Daisuke; Jaiswal, Dinesh Kumar; Clouse, Steven D; Jones, Alan M

    2016-07-01

    Plants and some protists have heterotrimeric G protein complexes that activate spontaneously without canonical G protein-coupled receptors (GPCRs). In Arabidopsis, the sole 7-transmembrane regulator of G protein signaling 1 (AtRGS1) modulates the G protein complex by keeping it in the resting state (GDP-bound). However, it remains unknown how a myriad of biological responses is achieved with a single G protein modulator. We propose that in complete contrast to G protein activation in animals, plant leucine-rich repeat receptor-like kinases (LRR RLKs), not GPCRs, provide this discrimination through phosphorylation of AtRGS1 in a ligand-dependent manner. G protein signaling is directly activated by the pathogen-associated molecular pattern flagellin peptide 22 through its LRR RLK, FLS2, and co-receptor BAK1.

  14. The TIM Barrel Architecture Facilitated the Early Evolution of Protein-Mediated Metabolism.

    PubMed

    Goldman, Aaron David; Beatty, Joshua T; Landweber, Laura F

    2016-01-01

    The triosephosphate isomerase (TIM) barrel protein fold is a structurally repetitive architecture that is present in approximately 10% of all enzymes. It is generally assumed that this ubiquity in modern proteomes reflects an essential historical role in early protein-mediated metabolism. Here, we provide quantitative and comparative analyses to support several hypotheses about the early importance of the TIM barrel architecture. An information theoretical analysis of protein structures supports the hypothesis that the TIM barrel architecture could arise more easily by duplication and recombination compared to other mixed α/β structures. We show that TIM barrel enzymes corresponding to the most taxonomically broad superfamilies also have the broadest range of functions, often aided by metal and nucleotide-derived cofactors that are thought to reflect an earlier stage of metabolic evolution. By comparison to other putatively ancient protein architectures, we find that the functional diversity of TIM barrel proteins cannot be explained simply by their antiquity. Instead, the breadth of TIM barrel functions can be explained, in part, by the incorporation of a broad range of cofactors, a trend that does not appear to be shared by proteins in general. These results support the hypothesis that the simple and functionally general TIM barrel architecture may have arisen early in the evolution of protein biosynthesis and provided an ideal scaffold to facilitate the metabolic transition from ribozymes, peptides, and geochemical catalysts to modern protein enzymes.

  15. Combined intervention of dietary soybean proteins and swim training: effects on bone metabolism in ovariectomized rats.

    PubMed

    Figard, Hélène; Mougin, Fabienne; Gaume, Vincent; Berthelot, Alain

    2006-01-01

    Soybean proteins, a rich source of isoflavones, taken immediately after an ovariectomy prevent bone loss in rats. Exercise-induced stimuli are essential for bone growth. Few studies exist about the combined effects of swim training and soybean protein supplementation on bone metabolism. So, the purpose of this study was to investigate, in 48 female Sprague-Dawley rats (12 weeks old) the effects of an 8-week swim-training regimen (1 h/day, 5 days/week) and dietary soybean proteins (200 g/kg diet) on bone metabolism. Rats were randomly assigned to four groups: (1) ovariectomized fed with a semisynthetic control diet; (2) ovariectomized fed with a soybean protein-enriched semisynthetic diet; (3) ovariectomized trained to exercise and fed with control diet; (4) ovariectomized trained to exercise and fed with a soybean protein diet. Following the treatment period, body weight gain was identical in the four groups. Soybean protein supplementation increased bone calcium content, and reduced plasma osteocalcin values, without significant modification of calcium balance and net calcium absorption. Swim training enhanced plasma and bone calcium content and calcium balance and net calcium absorption. It did not modify either plasma osteocalcin values or urinary deoxypyridinoline excretion. Both exercise and soybean protein intake increased plasma on bone calcium without modifying net calcium absorption or bone markers. In conclusion, we demonstrated, in ovariectomized rats, that swimming exercise and dietary supplementation with soy proteins do not have synergistic effects on calcium metabolism and bone markers.

  16. Disorder transitions and conformational diversity cooperatively modulate biological function in proteins.

    PubMed

    Zea, Diego Javier; Monzon, Alexander Miguel; Gonzalez, Claudia; Fornasari, María Silvina; Tosatto, Silvio C E; Parisi, Gustavo

    2016-06-01

    Structural differences between conformers sustain protein biological function. Here, we studied in a large dataset of 745 intrinsically disordered proteins, how ordered-disordered transitions modulate structural differences between conformers as derived from crystallographic data. We found that almost 50% of the proteins studied show no transitions and have low conformational diversity while the rest show transitions and a higher conformational diversity. In this last subset, 60% of the proteins become more ordered after ligand binding, while 40% more disordered. As protein conformational diversity is inherently connected with protein function our analysis suggests differences in structure-function relationships related to order-disorder transitions.

  17. Engineering protein-based machines to emulate key steps of metabolism (biological energy conversion)

    PubMed

    Urry; Peng; Hayes; McPherson; Xu; Woods; Gowda; Pattanaik

    1998-04-01

    Metabolism is the conversion of available energy sources to those energy forms required for sustaining and propagating living organisms; this is simply biological energy conversion. Proteins are the machines of metabolism; they are the engines of motility and the other machines that interconvert energy forms not involving motion. Accordingly, metabolic engineering becomes the use of natural protein-based machines for the good of society. In addition, metabolic engineering can utilize the principles, whereby proteins function, to design new protein-based machines to fulfill roles for society that proteins have never been called upon throughout evolution to fulfill. This article presents arguments for a universal mechanism whereby proteins perform their diverse energy conversions; it begins with background information, and then asserts a set of five axioms for protein folding, assembly, and function and for protein engineering. The key process is the hydrophobic folding and assembly transition exhibited by properly balanced amphiphilic protein sequences. The fundamental molecular process is the competition for hydration between hydrophobic and polar, e.g., charged, residues. This competition determines Tt, the onset temperature for the hydrophobic folding and assembly transition, Nhh, the numbers of waters of hydrophobic hydration, and the pKa of ionizable functions. Reported acid-base titrations and pH dependence of microwave dielectric relaxation data simultaneously demonstrate the interdependence of Tt, Nhh and the pKa using a series of microbially prepared protein-based poly(30mers) with one glutamic acid residue per 30mer and with an increasing number of more hydrophobic phenylalanine residues replacing valine residues. Also, reduction of nicotinamides and flavins is shown to lower Tt, i.e., to increase hydrophobicity. Furthermore, the argument is presented, and related to an extended Henderson-Hasselbalch equation, wherein reduction of nicotinamides represents

  18. The Role of Maternal Dietary Proteins in Development of Metabolic Syndrome in Offspring

    PubMed Central

    Jahan-Mihan, Alireza; Rodriguez, Judith; Christie, Catherine; Sadeghi, Marjan; Zerbe, Tara

    2015-01-01

    The prevalence of metabolic syndrome and obesity has been increasing. Pre-natal environment has been suggested as a factor influencing the risk of metabolic syndrome in adulthood. Both observational and experimental studies showed that maternal diet is a major modifier of the development of regulatory systems in the offspring in utero and post-natally. Both protein content and source in maternal diet influence pre- and early post-natal development. High and low protein dams’ diets have detrimental effect on body weight, blood pressure191 and metabolic and intake regulatory systems in the offspring. Moreover, the role of the source of protein in a nutritionally adequate maternal diet in programming of food intake regulatory system, body weight, glucose metabolism and blood pressure in offspring is studied. However, underlying mechanisms are still elusive. The purpose of this review is to examine the current literature related to the role of proteins in maternal diets in development of characteristics of the metabolic syndrome in offspring. PMID:26561832

  19. A liver stress-endocrine nexus promotes metabolic integrity during dietary protein dilution.

    PubMed

    Maida, Adriano; Zota, Annika; Sjøberg, Kim A; Schumacher, Jonas; Sijmonsma, Tjeerd P; Pfenninger, Anja; Christensen, Marie M; Gantert, Thomas; Fuhrmeister, Jessica; Rothermel, Ulrike; Schmoll, Dieter; Heikenwälder, Mathias; Iovanna, Juan L; Stemmer, Kerstin; Kiens, Bente; Herzig, Stephan; Rose, Adam J

    2016-09-01

    Dietary protein intake is linked to an increased incidence of type 2 diabetes (T2D). Although dietary protein dilution (DPD) can slow the progression of some aging-related disorders, whether this strategy affects the development and risk for obesity-associated metabolic disease such as T2D is unclear. Here, we determined that DPD in mice and humans increases serum markers of metabolic health. In lean mice, DPD promoted metabolic inefficiency by increasing carbohydrate and fat oxidation. In nutritional and polygenic murine models of obesity, DPD prevented and curtailed the development of impaired glucose homeostasis independently of obesity and food intake. DPD-mediated metabolic inefficiency and improvement of glucose homeostasis were independent of uncoupling protein 1 (UCP1), but required expression of liver-derived fibroblast growth factor 21 (FGF21) in both lean and obese mice. FGF21 expression and secretion as well as the associated metabolic remodeling induced by DPD also required induction of liver-integrated stress response-driven nuclear protein 1 (NUPR1). Insufficiency of select nonessential amino acids (NEAAs) was necessary and adequate for NUPR1 and subsequent FGF21 induction and secretion in hepatocytes in vitro and in vivo. Taken together, these data indicate that DPD promotes improved glucose homeostasis through an NEAA insufficiency-induced liver NUPR1/FGF21 axis. PMID:27548521

  20. Integrating gene and protein expression data with genome-scale metabolic networks to infer functional pathways

    PubMed Central

    2013-01-01

    Background The study of cellular metabolism in the context of high-throughput -omics data has allowed us to decipher novel mechanisms of importance in biotechnology and health. To continue with this progress, it is essential to efficiently integrate experimental data into metabolic modeling. Results We present here an in-silico framework to infer relevant metabolic pathways for a particular phenotype under study based on its gene/protein expression data. This framework is based on the Carbon Flux Path (CFP) approach, a mixed-integer linear program that expands classical path finding techniques by considering additional biophysical constraints. In particular, the objective function of the CFP approach is amended to account for gene/protein expression data and influence obtained paths. This approach is termed integrative Carbon Flux Path (iCFP). We show that gene/protein expression data also influences the stoichiometric balancing of CFPs, which provides a more accurate picture of active metabolic pathways. This is illustrated in both a theoretical and real scenario. Finally, we apply this approach to find novel pathways relevant in the regulation of acetate overflow metabolism in Escherichia coli. As a result, several targets which could be relevant for better understanding of the phenomenon leading to impaired acetate overflow are proposed. Conclusions A novel mathematical framework that determines functional pathways based on gene/protein expression data is presented and validated. We show that our approach is able to provide new insights into complex biological scenarios such as acetate overflow in Escherichia coli. PMID:24314206

  1. Acute metabolic acidosis decreases muscle protein synthesis but not albumin synthesis in humans.

    PubMed

    Kleger, G R; Turgay, M; Imoberdorf, R; McNurlan, M A; Garlick, P J; Ballmer, P E

    2001-12-01

    Chronic metabolic acidosis induces negative nitrogen balance by either increased protein breakdown or decreased protein synthesis. Few data exist regarding effects of acute metabolic acidosis on protein synthesis. We investigated fractional synthesis rates (FSRs) of muscle protein and albumin, plasma concentrations of insulin-like growth factor-I (IGF-I), thyroid-stimulating hormone (TSH), and thyroid hormones (free thyroxin [fT(4)] and triiodothyronine [fT(3)]) in seven healthy human volunteers after a stable controlled metabolic period of 5 days and again 48 hours later after inducing metabolic acidosis by oral ammonium chloride intake (4.2 mmol/kg/d divided in six daily doses). Muscle and albumin FSRs were obtained by the [(2)H(5)ring]phenylalanine flooding technique. Ammonium chloride induced a significant decrease in pH (7.43 +/- 0.02 versus 7.32 +/- 0.04; P < 0.0001) and bicarbonate concentration (24.6 +/- 1.6 versus 16.0 +/- 2.7 mmol/L; P < 0.0001) within 48 hours. Nitrogen balance decreased significantly on the second day of acidosis. The FSR of muscle protein decreased (1.94 +/- 0.25 versus 1.30 +/- 0.39; P < 0.02), whereas the FSR of albumin remained constant. TSH levels increased significantly (1.1 +/- 0.5 versus 1.9 +/- 1.1 mU/L; P = 0.03), whereas IGF-I, fT(4), and fT(3) levels showed no significant change. We conclude that acute metabolic acidosis for 48 hours in humans induces a decrease in muscle protein synthesis, which contributes substantially to a negative nitrogen balance. In contrast to prolonged metabolic acidosis of 7 days, a short period of acidosis in the present study did not downregulate albumin synthesis.

  2. Spaceflight and protein metabolism, with special reference to humans

    NASA Technical Reports Server (NTRS)

    Stein, T. P.; Gaprindashvili, T.

    1994-01-01

    Human space missions have shown that human spaceflight is associated with a loss of body protein. Specific changes include a loss of lean body mass, decreased muscle mass in the calves, decreased muscle strength, and changes in plasma proteins and amino acids. The major muscle loss is believed to be associated with the antigravity (postural) muscle. The most significant loss of protein appears to occur during the first month of flight. The etiology is believed to be multifactorial with contributions from disuse atrophy, undernutrition, and a stress type of response. This article reviews the results of American and Russian space missions to investigate this problem in humans, monkeys, and rats. The relationship of the flight results with ground-based models including bedrest for humans and hindlimb unweighting for rats is also discussed. The results suggest that humans adapt to spaceflight much better than either monkeys or rats.

  3. Role of acyl carrier protein isoforms in plant lipid metabolism

    SciTech Connect

    Not Available

    1990-01-01

    Although acyl carrier protein (ACP) is the best studied protein in plant fatty acid biosynthesis, the in vivo forms of ACPs and their steady state pools have not been examined previously in either seed or leaf. Information about the relative pool sizes of free ACP and its acyl-ACP intermediates is essential for understanding regulation of de novo fatty acid biosynthesis in plants. In this study we utilized antibodies directed against spinach ACP as a sensitive assay to analyze the acyl groups while they were still covalently attached to ACPs. 4 refs., 4 figs.

  4. Investigation into the role of catabolite control protein A in the metabolic regulation of Streptococcus suis serotype 2 using gene expression profile analysis

    PubMed Central

    LANG, XULONG; WAN, ZHONGHAI; PAN, YING; WANG, XIURAN; WANG, XIAOXU; BU, ZHAOYANG; QIAN, JING; ZENG, HUAZONG; WANG, XINGLONG

    2015-01-01

    Catabolite control protein A (CcpA) serves a key function in the catabolism of Streptococcus suis serotype 2 (S. suis 2) by affecting the biological function and metabolic regulatory mechanisms of this bacterium. The aim of the present study was to identify variations in CcpA expression in S. suis 2 using gene expression profile analysis. Using sequencing and functional analysis, CcpA was demonstrated to play a regulatory role in the expression and regulation of virulence genes, carbon metabolism and immunoregulation in S. suis 2. Gene Ontology and Kyto Encyclopedia of Genes and Genomes analyses indicated that CcpA in S. suis 2 is involved in the regulation of multiple metabolic processes. Furthermore, combined analysis of the transcriptome and metabolite data suggested that metabolites varied due to the modulation of gene expression levels under the influence of CcpA regulation. In addition, metabolic network analysis indicated that CcpA impacted carbon metabolism to a certain extent. Therefore, the present study has provided a more comprehensive analysis of the role of CcpA in the metabolic regulation of S. suis 2, which may facilitate future investigation into this mechanism. Furthermore, the results of the present study provide a foundation for further research into the regulatory function of CcpA and associated metabolic pathways in S. suis 2. PMID:26170923

  5. Modulating Composition and Metabolic Activity of the Gut Microbiota in IBD Patients

    PubMed Central

    Matijašić, Mario; Meštrović, Tomislav; Perić, Mihaela; Čipčić Paljetak, Hana; Panek, Marina; Vranešić Bender, Darija; Ljubas Kelečić, Dina; Krznarić, Željko; Verbanac, Donatella

    2016-01-01

    The healthy intestine represents a remarkable interface where sterile host tissues come in contact with gut microbiota, in a balanced state of homeostasis. The imbalance of gut homeostasis is associated with the onset of many severe pathological conditions, such as inflammatory bowel disease (IBD), a chronic gastrointestinal disorder increasing in incidence and severely influencing affected individuals. Despite the recent development of next generation sequencing and bioinformatics, the current scientific knowledge of specific triggers and diagnostic markers to improve interventional approaches in IBD is still scarce. In this review we present and discuss currently available and emerging therapeutic options in modulating composition and metabolic activity of gut microbiota in patients affected by IBD. Therapeutic approaches at the microbiota level, such as dietary interventions alone or with probiotics, prebiotics and synbiotics, administration of antibiotics, performing fecal microbiota transplantation (FMT) and the use of nematodes, all represent a promising opportunities towards establishing and maintaining of well-being as well as improving underlying IBD symptoms. PMID:27104515

  6. Potentiating the antitumour response of CD8(+) T cells by modulating cholesterol metabolism.

    PubMed

    Yang, Wei; Bai, Yibing; Xiong, Ying; Zhang, Jin; Chen, Shuokai; Zheng, Xiaojun; Meng, Xiangbo; Li, Lunyi; Wang, Jing; Xu, Chenguang; Yan, Chengsong; Wang, Lijuan; Chang, Catharine C Y; Chang, Ta-Yuan; Zhang, Ti; Zhou, Penghui; Song, Bao-Liang; Liu, Wanli; Sun, Shao-cong; Liu, Xiaolong; Li, Bo-liang; Xu, Chenqi

    2016-03-31

    CD8(+) T cells have a central role in antitumour immunity, but their activity is suppressed in the tumour microenvironment. Reactivating the cytotoxicity of CD8(+) T cells is of great clinical interest in cancer immunotherapy. Here we report a new mechanism by which the antitumour response of mouse CD8(+) T cells can be potentiated by modulating cholesterol metabolism. Inhibiting cholesterol esterification in T cells by genetic ablation or pharmacological inhibition of ACAT1, a key cholesterol esterification enzyme, led to potentiated effector function and enhanced proliferation of CD8(+) but not CD4(+) T cells. This is due to the increase in the plasma membrane cholesterol level of CD8(+) T cells, which causes enhanced T-cell receptor clustering and signalling as well as more efficient formation of the immunological synapse. ACAT1-deficient CD8(+) T cells were better than wild-type CD8(+) T cells at controlling melanoma growth and metastasis in mice. We used the ACAT inhibitor avasimibe, which was previously tested in clinical trials for treating atherosclerosis and showed a good human safety profile, to treat melanoma in mice and observed a good antitumour effect. A combined therapy of avasimibe plus an anti-PD-1 antibody showed better efficacy than monotherapies in controlling tumour progression. ACAT1, an established target for atherosclerosis, is therefore also a potential target for cancer immunotherapy. PMID:26982734

  7. Potentiating the antitumour response of CD8+ T cells by modulating cholesterol metabolism

    PubMed Central

    Yang, Wei; Bai, Yibing; Xiong, Ying; Zhang, Jin; Chen, Shuokai; Zheng, Xiaojun; Meng, Xiangbo; Li, Lunyi; Wang, Jing; Xu, Chenguang; Yan, Chengsong; Wang, Lijuan; Chang, Catharine C. Y.; Chang, Ta-Yuan; Zhang, Ti; Zhou, Penghui; Song, Bao-Liang; Liu, Wanli; Sun, Shao-cong; Liu, Xiaolong; Li, Bo-liang; Xu, Chenqi

    2016-01-01

    CD8+ T cells have a central role in antitumour immunity, but their activity is suppressed in the tumour microenvironment1–4. Reactivating the cytotoxicity of CD8+ T cells is of great clinical interest in cancer immunotherapy. Here we report a new mechanism by which the antitumour response of mouse CD8+ T cells can be potentiated by modulating cholesterol metabolism. Inhibiting cholesterol esterification in T cells by genetic ablation or pharmacological inhibition of ACAT1, a key cholesterol esterification enzyme5, led to potentiated effector function and enhanced proliferation of CD8+ but not CD4+ T cells. This is due to the increase in the plasma membrane cholesterol level of CD8+ T cells, which causes enhanced T-cell receptor clustering and signalling as well as more efficient formation of the immunological synapse. ACAT1-deficient CD8+ T cells were better than wild-type CD8+ T cells at controlling melanoma growth and metastasis in mice. We used the ACAT inhibitor avasimibe, which was previously tested in clinical trials for treating atherosclerosis and showed a good human safety profile6,7, to treat melanoma in mice and observed a good antitumour effect. A combined therapy of avasimibe plus an anti-PD-1 antibody showed better efficacy than monotherapies in controlling tumour progression. ACAT1, an established target for atherosclerosis, is therefore also a potential target for cancer immunotherapy. PMID:26982734

  8. Caffeine Impact on Metabolic Syndrome Components Is Modulated by a CYP1A2 Variant.

    PubMed

    Platt, Daniel E; Ghassibe-Sabbagh, Michella; Salameh, Pascale; Salloum, Angelique K; Haber, Marc; Mouzaya, Francis; Gauguier, Dominique; Al-Sarraj, Yasser; El-Shanti, Hatem; Zalloua, Pierre A; Abchee, Antoine B

    2016-01-01

    Cultural, dietary, and lifestyle factors are the main modulators of type 2 diabetes mellitus (T2DM) disease risk. Coffee is one of the most popular worldwide beverages, and recent epidemiological studies have showed that coffee consumption is associated with a lower risk of T2DM. This study investigates the impact of coffee intake on T2DM risk and assesses the effect of CYP variants with caffeine exposures on T2DM. Data from 7,607 study subjects were analyzed by logistic regression models, among whom 3,290 GWAS data were available for CYP variants association studies using Plink analysis. These data suggest a protective relationship for women, but not for men; however, the results were not statistically significant in this dataset and there is a significant interaction in favor of women regarding heavy coffee consumption. The interaction between male gender and heavy coffee consumption becomes significant, thereby tending to cancel the protective effect of coffee for males. CYP rs2470890 allele 'C' increases the odds of T2DM by a factor of around 1.2 but decreases the odds of caffeine boosting T2DM of 1.7 by a factor of 0.77. rs2470890 showed an association with T2DM only when the interaction with coffee was considered, thereby setting an example of genetic activation by dietary changes associating with metabolic syndrome. PMID:26588584

  9. Modulation of phosphate/pyrophosphate metabolism to regenerate the periodontium. A novel in vivo approach

    PubMed Central

    Rodrigues, Thaisângela L.; Nagatomo, Kanako J.; Foster, Brian L.; Nociti, Francisco H.; Somerman, Martha J.

    2013-01-01

    Background The developing periodontium is sensitive to local levels of phosphate (Pi) and pyrophosphate (PPi), as demonstrated by cementum phenotypes resulting from loss of function of protein regulators of Pi/PPi homeostasis. The progressive ankylosis protein (ANK) regulates transport of PPi, and Ank knock-out (KO) mice feature rapidly forming and thick cementum. We hypothesized that, besides affecting cementum formation, decreased extracellular PPi levels in Ank KO mice would also impact cementum regeneration. Methods Periodontal fenestration defects (2mm/1mm/0.5mm) were created on the buccal aspects of mandibular molars in Ank KO and wild-type (WT) mice. Mandibles were harvested at 15 and 30 days post-surgery for histology, histomorphometry, evaluation of in vivo fluorochrome labeling, and immunohistochemistry(IHC) for proteins including bone sialoprotein (BSP), osteopontin (OPN), dentin matrix protein 1 (DMP1), and ectonucleotide pyrophosphatase/phosphodiesterase 1 (NPP1). Results A greater amount of new cementum was observed for Ank KO mice at 15 and 30 days post-surgery (p<0.05), that was confirmed by fluorochrome labeling showing a higher new cementum appositional activity in the defect areas in Ank KO vs. controls. At days 15 and 30 during healing, regenerating cementum and associated cells in Ank KO recapitulated expression patterns mapped during development, including limited BSP and positive OPN and DMP1 in the cementum matrix, as well as elevated NPP1 in cementoblasts. Conclusions Within the limits of the study, these findings suggest that reduced local levels of PPi can promote increased cementum regeneration. Therefore, local modulation of Pi/PPi may be a potential therapeutic approach for achieving improved cementum regeneration. PMID:21488756

  10. RNAi screens reveal novel metabolic regulators: RIP140, MAP4k4 and the lipid droplet associated fat specific protein (FSP) 27.

    PubMed

    Puri, V; Virbasius, J V; Guilherme, A; Czech, M P

    2008-01-01

    Adipose tissue modulates whole body metabolism and insulin sensitivity by controlling circulating lipid levels and producing molecules that can regulate fatty acid metabolism in such tissues as muscle and liver. We have developed RNA interference (RNAi) screens to identify genes in cultured adipocytes that regulate insulin signalling and key metabolic pathways. These short interfering RNA (siRNA)-based screens identified the transcriptional corepressor receptor interacting protein 140 (RIP140) (J Clin Invest 116: 125, 2006) and the mitogen-activated protein kinase (MAP4k4) (Proc Natl Acad Sci USA 103: 2087, 2006) as negative regulators of insulin-responsive hexose uptake and oxidative metabolism. Gene expression profiling revealed that RIP140 depletion upregulates the expression of clusters of genes in the pathways of glucose uptake, glycolysis, tricarboxylic acid cycle, fatty acid oxidation, mitochondrial biogenesis and oxidative phosphorylation. RIP140-null mice resist weight gain on a high-fat diet and display enhanced glucose tolerance. MAP4k4 depletion in adipocytes increases many of the RIP140-sensitive genes, increases adipogenesis and mediates some actions of tumour necrosis factor-alpha (TNF-alpha). Remarkably, another hit in our RNAi screens was fat specific protein 27 (FSP27), a highly expressed isoform of Cidea. We discovered that FSP27 unexpectedly associates specifically with lipid droplets and regulates fat storage. We conclude that RIP140, MAP4k4 and the novel lipid droplet protein FSP27 are powerful regulators of adipose tissue metabolism and are potential therapeutic targets for controlling metabolic disease. The discovery of these novel proteins validates the power of RNAi screening for discovery of new therapeutic approaches to type 2 diabetes and obesity. PMID:18171433

  11. Pathway analysis of Pichia pastoris to elucidate methanol metabolism and its regulation for production of recombinant proteins.

    PubMed

    Unrean, Pornkamol

    2014-01-01

    This research rationally analyzes metabolic pathways of Pichia pastoris to study the metabolic flux responses of this yeast under methanol metabolism. A metabolic model of P. pastoris was constructed and analyzed by elementary mode analysis (EMA). EMA was used to comprehensively identify the cell's metabolic flux profiles and its underlying regulation mechanisms for the production of recombinant proteins from methanol. Change in phenotypes and flux profiles during methanol adaptation with varying feed mixture of glycerol and methanol was examined. EMA identified increasing and decreasing fluxes during the glycerol-methanol metabolic shift, which well agreed with experimental observations supporting the validity of the metabolic network model. Analysis of all the identified pathways also led to the determination of the metabolic capacities as well as the optimum metabolic pathways for recombinant protein synthesis during methanol induction. The network sensitivity analysis revealed that the production of proteins can be improved by manipulating the flux ratios at the pyruvate branch point. In addition, EMA suggested that protein synthesis is optimum under hypoxic culture conditions. The metabolic modeling and analysis presented in this study could potentially form a valuable knowledge base for future research on rational design and optimization of P. pastoris by determining target genes, pathways, and culture conditions for enhanced recombinant protein synthesis. The metabolic pathway analysis is also of considerable value for production of therapeutic proteins by P. pastoris in biopharmaceutical applications.

  12. A human protein containing multiple types of protease-inhibitory modules

    PubMed Central

    Trexler, Mária; Bányai, László; Patthy, László

    2001-01-01

    By using sensitive homology-search and gene-finding programs, we have found that a genomic region from the tip of the short arm of human chromosome 16 (16p13.3) encodes a putative secreted protein consisting of a domain related to the whey acidic protein (WAP) domain, a domain homologous with follistatin modules of the Kazal-domain family (FS module), an immunoglobulin-related domain (Ig domain), two tandem domains related to Kunitz-type protease inhibitor modules (KU domains), and a domain belonging to the recently defined NTR-module family (NTR domain). The gene encoding these WAP, FS, Ig, KU, and NTR modules (hereafter referred to as the WFIKKN gene) is intron-depleted—its single 1,157-bp intron splits the WAP module. The validity of our gene prediction was confirmed by sequencing a WFIKKN cDNA cloned from a lung cDNA library. Studies on the tissue-expression pattern of the WFIKKN gene have shown that the gene is expressed primarily in pancreas, kidney, liver, placenta, and lung. As to the function of the WFIKKN protein, it is noteworthy that it contains FS, WAP, and KU modules, i.e., three different module types homologous with domains frequently involved in inhibition of serine proteases. The protein also contains an NTR module, a domain type implicated in inhibition of zinc metalloproteinases of the metzincin family. On the basis of its intriguing homologies, we suggest that the WFIKKN protein is a multivalent protease inhibitor that may control the action of multiple types of serine proteases as well as metalloproteinase(s). PMID:11274388

  13. Fluoranthene metabolism and associated proteins in Mycobacterium sp. JS14.

    PubMed

    Lee, Sung-Eun; Seo, Jong-Su; Keum, Young-Soo; Lee, Kwang-Jun; Li, Qing X

    2007-06-01

    Fluoranthene is a polycyclic aromatic hydrocarbon (PAH) commonly present in PAH-contaminated soils. We studied fluoranthene catabolism and associated proteins in Mycobacterium sp. JS14, a bacterium isolated from a PAH-contaminated soil in Hilo (HI, USA). Fluoranthene degrades in at least three separated pathways via 1-indanone, 2',3'-dihydroxybiphenyl-2,3,-dicarboxylic acid, and naphthalene-1,8-dicarboxylic acid. Part of the diverse catabolism is converged into phthalate catabolism. An increased expression of 25 proteins related to fluoranthene catabolism is found with 1-D PAGE or 2-DE and nano-LC-MS/MS. Detection of fluoranthene catabolism associated proteins coincides well with its multiple degradation pathways that are mapped via metabolites identified. Among the up-regulated proteins, PAH ring-hydroxylating dioxygenase alpha-subunit and beta-subunit and 2,3-dihydroxybiphenyl 1,2-dioxygenase are notably induced. The up-regulation of trans-2-carboxybenzalpyruvate hydratase suggests that some of fluoranthene metabolites may be further degraded through aromatic dicarboxylic acid pathways. Catalase and superoxide dismutase were up-regulated to control unexpected oxidative stress during the fluoranthene catabolism. The up-regulation of chorismate synthase and nicotine-nucleotide phosphorylase may be necessary for sustaining shikimate pathway and pyrimidine biosynthesis, respectively. A fluoranthene degradation pathway for Mycobacterium sp. JS14 was proposed and confirmed by proteomic study by identifying almost all the enzymes required during the initial steps of fluoranthene degradation. PMID:17514677

  14. Effects of atorvastatin on human c reactive protein metabolism

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Statins are known to reduce plasma C-reactive protein (CRP) concentrations. Our goals were to define the mechanisms by which CRP was reduced by maximal dose atorvastatin. Eight subjects with combined hyperlipidemia (5 men and 3 postmenopausal women) were enrolled in a randomized, placebo-controlled...

  15. Modulating non-native aggregation and electrostatic protein-protein interactions with computationally designed single-point mutations.

    PubMed

    O'Brien, C J; Blanco, M A; Costanzo, J A; Enterline, M; Fernandez, E J; Robinson, A S; Roberts, C J

    2016-06-01

    Non-native protein aggregation is a ubiquitous challenge in the production, storage and administration of protein-based biotherapeutics. This study focuses on altering electrostatic protein-protein interactions as a strategy to modulate aggregation propensity in terms of temperature-dependent aggregation rates, using single-charge variants of human γ-D crystallin. Molecular models were combined to predict amino acid substitutions that would modulate protein-protein interactions with minimal effects on conformational stability. Experimental protein-protein interactions were quantified by the Kirkwood-Buff integrals (G22) from laser scattering, and G22 showed semi-quantitative agreement with model predictions. Experimental initial-rates for aggregation showed that increased (decreased) repulsive interactions led to significantly increased (decreased) aggregation resistance, even based solely on single-point mutations. However, in the case of a particular amino acid (E17), the aggregation mechanism was altered by substitution with R or K, and this greatly mitigated improvements in aggregation resistance. The results illustrate that predictions based on native protein-protein interactions can provide a useful design target for engineering aggregation resistance; however, this approach needs to be balanced with consideration of how mutations can impact aggregation mechanisms. PMID:27160179

  16. Metal ion modulated electron transfer in photosynthetic proteins.

    SciTech Connect

    Utschig, L. M.; Thurnauer, M. C.; Chemistry

    2004-07-01

    Photosynthetic purple bacterial reaction center (RC) proteins are ideal native systems for addressing basic questions regarding the nature of biological electron transfer because both the protein structure and the electron-transfer reactions are well-characterized. Metal ion binding to the RC can affect primary photochemistry and provides a probe for understanding the involvement of local protein environments in electron transfer. The RC has two distinct transition metal ion binding sites, the well-known non-heme Fe{sup 2+} site buried in the protein interior and a recently discovered Zn{sup 2+} site located on the surface of the protein. Fe{sup 2+} removal and Zn{sup 2+} binding systematically affect different electron-transfer steps in the RC. Factors involved in the metal ion alteration of RC electron transfer may provide a paradigm for other biological systems involved in electron transfer.

  17. Function and expression of a novel rat salt-tolerant protein: evidence of a role in cellular sodium metabolism.

    PubMed

    Tsuji, E; Tsuji, Y; Sasaguri, M; Arakawa, K

    1998-09-01

    Higher dietary salt intake in humans is associated with higher BP, but the BP response to NaCl, so-called salt sensitivity, is heterogeneous among individuals. It has been postulated that modifications in cellular cation metabolism may be related to salt sensitivity in mammalian hypertension. The authors have isolated a novel rat complementary DNA, called salt-tolerant protein (STP), that can functionally complement Saccharomyces cervisiae HAL1, which improves salt tolerance by modulating the cation transport system. On high-salt (8% NaCl) diets, both Dahl salt-sensitive and salt-resistant rats displayed an elevated BP and increased STP mRNA expression. Immunohistochemistry using an anti-rat STP antibody demonstrated the presence of STP immunoreactivity in the proximal tubules. In cells that transiently expressed STP, the intracellular [Na+]/[K+] ratio was higher than that in control cells. STP contains predicted coiled-coil and Src homology 3 domains, and shows a partially high degree of nucleotide identity to human thyroid-hormone receptor interacting protein. These results suggest that STP may play an important role in salt sensitivity through cellular sodium metabolism by mediating signal transduction and a hormone-dependent transcription mechanism. PMID:9727364

  18. Allosteric Modulation of protein oligomerization: an emerging approach to drug design

    NASA Astrophysics Data System (ADS)

    Gabizon, Ronen; Friedler, Assaf

    2014-03-01

    Many disease-related proteins are in equilibrium between different oligomeric forms. The regulation of this equilibrium plays a central role in maintaining the activity of these proteins in vitro and in vivo. Modulation of the oligomerization equilibrium of proteins by molecules that bind preferentially to a specific oligomeric state is emerging as a potential therapeutic strategy that can be applied to many biological systems such as cancer and viral infections. The target proteins for such compounds are diverse in structure and sequence, and may require different approaches for shifting their oligomerization equilibrium. The discovery of such oligomerization-modulating compounds is thus achieved based on existing structural knowledge about the specific target proteins, as well as on their interactions with partner proteins or with ligands. In silico design and combinatorial tools such as peptide arrays and phage display are also used for discovering compounds that modulate protein oligomerization. The current review highlights some of the recent developments in the design of compounds aimed at modulating the oligomerization equilibrium of proteins, including the "shiftides" approach developed in our lab.

  19. Myocardial Oxidative Metabolism and Protein Synthesis during Mechanical Circulatory Support by Extracorporeal Membrane Oxygenation

    SciTech Connect

    Priddy, MD, Colleen M.; Kajimoto, Masaki; Ledee, Dolena; Bouchard, Bertrand; Isern, Nancy G.; Olson, Aaron; Des Rosiers, Christine; Portman, Michael A.

    2013-02-01

    Extracorporeal membrane oxygenation (ECMO) provides mechanical circulatory support essential for survival in infants and children with acute cardiac decompensation. However, ECMO also causes metabolic disturbances, which contribute to total body wasting and protein loss. Cardiac stunning can also occur which prevents ECMO weaning, and contributes to high mortality. The heart may specifically undergo metabolic impairments, which influence functional recovery. We tested the hypothesis that ECMO alters oxidative. We focused on the amino acid leucine, and integration with myocardial protein synthesis. We used a translational immature swine model in which we assessed in heart (i) the fractional contribution of leucine (FcLeucine) and pyruvate (FCpyruvate) to mitochondrial acetyl-CoA formation by nuclear magnetic resonance and (ii) global protein fractional synthesis (FSR) by gas chromatography-mass spectrometry. Immature mixed breed Yorkshire male piglets (n = 22) were divided into four groups based on loading status (8 hours of normal circulation or ECMO) and intracoronary infusion [13C6,15N]-L-leucine (3.7 mM) alone or with [2-13C]-pyruvate (7.4 mM). ECMO decreased pulse pressure and correspondingly lowered myocardial oxygen consumption (~ 40%, n = 5), indicating decreased overall mitochondrial oxidative metabolism. However, FcLeucine was maintained and myocardial protein FSR was marginally increased. Pyruvate addition decreased tissue leucine enrichment, FcLeucine, and Fc for endogenous substrates as well as protein FSR. Conclusion: The heart under ECMO shows reduced oxidative metabolism of substrates, including amino acids, while maintaining (i) metabolic flexibility indicated by ability to respond to pyruvate, and (ii) a normal or increased capacity for global protein synthesis, suggesting an improved protein balance.

  20. Maternal protein restriction impairs the transcriptional metabolic flexibility of skeletal muscle in adult rat offspring.

    PubMed

    da Silva Aragão, Raquel; Guzmán-Quevedo, Omar; Pérez-García, Georgina; Manhães-de-Castro, Raul; Bolaños-Jiménez, Francisco

    2014-08-14

    Skeletal muscle exhibits a remarkable flexibility in the usage of fuel in response to the nutrient intake and energy demands of the organism. In fact, increased physical activity and fasting trigger a transcriptional programme in skeletal muscle cells leading to a switch from carbohydrate to lipid oxidation. Impaired metabolic flexibility has been reported to be associated with obesity and type 2 diabetes, but it is not known whether the disability to adapt to metabolic demands is a cause or a consequence of these pathological conditions. Inasmuch as a poor nutritional environment during early life is a predisposing factor for the development of metabolic diseases in adulthood, in the present study, we aimed to determine the long-term effects of maternal malnutrition on the metabolic flexibility of offspring skeletal muscle. To this end, the transcriptional responses of the soleus and extensor digitorum longus muscles to fasting were evaluated in adult rats born to dams fed a control (17 % protein) or a low-protein (8 % protein, protein restricted (PR)) diet throughout pregnancy and lactation. With the exception of reduced body weight and reduced plasma concentrations of TAG, PR rats exhibited a metabolic profile that was the same as that of the control rats. In the fed state, PR rats exhibited an enhanced expression of key regulatory genes of fatty acid oxidation including CPT1a, PGC-1α, UCP3 and PPARα and an impaired expression of genes that increase the capacity for fat oxidation in response to fasting. These results suggest that impaired metabolic inflexibility precedes and may contribute to the development of metabolic disorders associated with early malnutrition. PMID:24823946

  1. In Silico Studies of C3 Metabolic Pathway Proteins of Wheat (Triticum aestivum)

    PubMed Central

    Naeem, Muhammad Kashif; Rauf, Sobiah; Iqbal, Hina; Nawaz Shah, Muhammad Kausar; Mir, Asif

    2013-01-01

    Photosynthesis is essential for plant productivity and critical for plant growth. More than 90% of plants have a C3 metabolic pathway primarily for carbon assimilation. Improving crop yields for food and fuel is a major challenge for plant biology. To enhance the production of wheat there is need to adopt the strategies that can create the change in plants at the molecular level. During the study we have employed computational bioinformatics and interactomics analysis of C3 metabolic pathway proteins in wheat. The three-dimensional protein modeling provided insight into molecular mechanism and enhanced understanding of physiological processes and biological systems. Therefore in our study, initially we constructed models for nine proteins involving C3 metabolic pathway, as these are not determined through wet lab experiment (NMR, X-ray Crystallography) and not available in RCSB Protein Data Bank and UniProt KB. On the basis of docking interaction analysis, we proposed the schematic diagram of C3 metabolic pathway. Accordingly, there also exist vice versa interactions between 3PGK and Rbcl. Future site and directed mutagenesis experiments in C3 plants could be designed on the basis of our findings to confirm the predicted protein interactions. PMID:23484105

  2. Global Profiling of Protein Lysine Malonylation in Escherichia coli Reveals Its Role in Energy Metabolism.

    PubMed

    Qian, Lili; Nie, Litong; Chen, Ming; Liu, Ping; Zhu, Jun; Zhai, Linhui; Tao, Sheng-Ce; Cheng, Zhongyi; Zhao, Yingming; Tan, Minjia

    2016-06-01

    Protein lysine malonylation is a recently identified post-translational modification (PTM), which is evolutionarily conserved from bacteria to mammals. Although analysis of lysine malonylome in mammalians suggested that this modification was related to energy metabolism, the substrates and biological roles of malonylation in prokaryotes are still poorly understood. In this study, we performed qualitative and quantitative analyses to globally identify lysine malonylation substrates in Escherichia coli. We identified 1745 malonylation sites in 594 proteins in E. coli, representing the first and largest malonylome data set in prokaryotes up to date. Bioinformatic analyses showed that lysine malonylation was significantly enriched in protein translation, energy metabolism pathways and fatty acid biosynthesis, implying the potential roles of protein malonylation in bacterial physiology. Quantitative proteomics by fatty acid synthase inhibition in both auxotrophic and prototrophic E. coli strains revealed that lysine malonylation is closely associated with E. coli fatty acid metabolism. Protein structural analysis and mutagenesis experiment suggested malonylation could impact enzymatic activity of citrate synthase, a key enzyme in citric acid (TCA) cycle. Further comparative analysis among lysine malonylome, succinylome and acetylome data showed that these three modifications could participate in some similar enriched metabolism pathways, but they could also possibly play distinct roles such as in fatty acid synthesis. These data expanded our knowledge of lysine malonylation in prokaryotes, providing a resource for functional study of lysine malonylation in bacteria. PMID:27183143

  3. The RHOX Homeodomain Proteins Regulate the Expression of Insulin and Other Metabolic Regulators in the Testis*

    PubMed Central

    MacLean, James A.; Hu, Zhiying; Welborn, Joshua P.; Song, Hye-Won; Rao, Manjeet K.; Wayne, Chad M.; Wilkinson, Miles F.

    2013-01-01

    Defects in cellular metabolism have been widely implicated in causing male infertility, but there has been little progress in understanding the underlying mechanism. Here we report that several key metabolism genes are regulated in the testis by Rhox5, the founding member of a large X-linked homeobox gene cluster. Among these Rhox5-regulated genes are insulin 2 (Ins2), resistin (Retn), and adiponectin (Adipoq), all of which encode secreted proteins that have profound and wide-ranging effects on cellular metabolism. The ability of Rhox5 to regulate their levels in the testis has the potential to dictate metabolism locally in this organ, given the existence of the blood-testes barrier. We demonstrate that Ins2 is a direct target of Rhox5 in Sertoli cells, and we show that this regulation is physiologically significant, because Rhox5-null mice fail to up-regulate Ins2 expression during the first wave of spermatogenesis and have insulin-signaling defects. We identify other Rhox family members that induce Ins2 transcription, define protein domains and homeodomain amino acid residues crucial for this property, and demonstrate that this regulation is conserved. Rhox5-null mice also exhibit altered expression of other metabolism genes, including those encoding the master transcriptional regulators of metabolism, PPARG and PPARGC1A, as well as SCD1, the rate-limiting enzyme for fatty acid metabolism. These results, coupled with the known roles of RHOX5 and its target metabolism genes in spermatogenesis in vivo, lead us to propose a model in which RHOX5 is a central transcription factor that promotes the survival of male germ cells via its effects on cellular metabolism. PMID:24121513

  4. The RHOX homeodomain proteins regulate the expression of insulin and other metabolic regulators in the testis.

    PubMed

    MacLean, James A; Hu, Zhiying; Welborn, Joshua P; Song, Hye-Won; Rao, Manjeet K; Wayne, Chad M; Wilkinson, Miles F

    2013-11-29

    Defects in cellular metabolism have been widely implicated in causing male infertility, but there has been little progress in understanding the underlying mechanism. Here we report that several key metabolism genes are regulated in the testis by Rhox5, the founding member of a large X-linked homeobox gene cluster. Among these Rhox5-regulated genes are insulin 2 (Ins2), resistin (Retn), and adiponectin (Adipoq), all of which encode secreted proteins that have profound and wide-ranging effects on cellular metabolism. The ability of Rhox5 to regulate their levels in the testis has the potential to dictate metabolism locally in this organ, given the existence of the blood-testes barrier. We demonstrate that Ins2 is a direct target of Rhox5 in Sertoli cells, and we show that this regulation is physiologically significant, because Rhox5-null mice fail to up-regulate Ins2 expression during the first wave of spermatogenesis and have insulin-signaling defects. We identify other Rhox family members that induce Ins2 transcription, define protein domains and homeodomain amino acid residues crucial for this property, and demonstrate that this regulation is conserved. Rhox5-null mice also exhibit altered expression of other metabolism genes, including those encoding the master transcriptional regulators of metabolism, PPARG and PPARGC1A, as well as SCD1, the rate-limiting enzyme for fatty acid metabolism. These results, coupled with the known roles of RHOX5 and its target metabolism genes in spermatogenesis in vivo, lead us to propose a model in which RHOX5 is a central transcription factor that promotes the survival of male germ cells via its effects on cellular metabolism.

  5. Rubella virus capsid protein modulation of viral genomic and subgenomic RNA synthesis

    SciTech Connect

    Tzeng, W.-P.; Frey, Teryl K. . E-mail: tfrey@gsu.edu

    2005-07-05

    The ratio of the subgenomic (SG) to genome RNA synthesized by rubella virus (RUB) replicons expressing the green fluorescent protein reporter gene (RUBrep/GFP) is substantially higher than the ratio of these species synthesized by RUB (4.3 for RUBrep/GFP vs. 1.3-1.4 for RUB). It was hypothesized that this modulation of the viral RNA synthesis was by one of the virus structural protein genes and it was found that introduction of the capsid (C) protein gene into the replicons as an in-frame fusion with GFP resulted in an increase of genomic RNA production (reducing the SG/genome RNA ratio), confirming the hypothesis and showing that the C gene was the moiety responsible for the modulation effect. The N-terminal one-third of the C gene was required for the effect of be exhibited. A similar phenomenon was not observed with the replicons of Sindbis virus, a related Alphavirus. Interestingly, modulation was not observed when RUBrep/GFP was co-transfected with either other RUBrep or plasmid constructs expressing the C gene, demonstrating that modulation could occur only when the C gene was provided in cis. Mutations that prevented translation of the C protein failed to modulate RNA synthesis, indicating that the C protein was the moiety responsible for modulation; consistent with this conclusion, modulation of RNA synthesis was maintained when synonymous codon mutations were introduced at the 5' end of the C gene that changed the C gene sequence without altering the amino acid sequence of the C protein. These results indicate that C protein translated in proximity of viral replication complexes, possibly from newly synthesized SG RNA, participate in regulating the replication of viral RNA.

  6. Leishmania amazonensis: PKC-like protein kinase modulates the (Na++K+)ATPase activity.

    PubMed

    Almeida-Amaral, Elmo Eduardo de; Caruso-Neves, Celso; Lara, Lucienne Silva; Pinheiro, Carla Mônica; Meyer-Fernandes, José Roberto

    2007-08-01

    The present study aimed to identify the presence of protein kinase C-like (PKC-like) in Leishmania amazonensis and to elucidate its possible role in the modulation of the (Na(+)+K(+))ATPase activity. Immunoblotting experiments using antibody against a consensus sequence (Ac 543-549) of rabbit protein kinase C (PKC) revealed the presence of a protein kinase of 80 kDa in L. amazonensis. Measurements of protein kinase activity showed the presence of both (Ca(2+)-dependent) and (Ca(2+)-independent) protein kinase activity in plasma membrane and cytosol. Phorbol ester (PMA) activation of the Ca(2+)-dependent protein kinase stimulated the (Na(+)+K(+))ATPase activity, while activation of the Ca(2+)-independent protein kinase was inhibitory. Both effects of protein kinase on the (Na(+)+K(+))ATPase of the plasma membrane were lower than that observed in intact cells. PMA induced the translocation of protein kinase from cytosol to plasma membrane, indicating that the maximal effect of protein kinase on the (Na(+)+K(+))ATPase activity depends on the synergistic action of protein kinases from both plasma membrane and cytosol. This is the first demonstration of a protein kinase activated by PMA in L. amazonensis and the first evidence for a possible role in the regulation of the (Na(+)+K(+))ATPase activity in this trypanosomatid. Modulation of the (Na(+)+K(+))ATPase by protein kinase in a trypanosomatid opens up new possibilities to understand the regulation of ion homeostasis in this parasite. PMID:17475255

  7. Data set of interactomes and metabolic pathways of proteins differentially expressed in brains with Alzheimer׳s disease.

    PubMed

    Minjarez, Benito; Calderón-González, Karla Grisel; Valero Rustarazo, Ma Luz; Herrera-Aguirre, María Esther; Labra-Barrios, María Luisa; Rincon-Limas, Diego E; Sánchez Del Pino, Manuel M; Mena, Raul; Luna-Arias, Juan Pedro

    2016-06-01

    Alzheimer׳s disease is one of the main causes of dementia in the elderly and its frequency is on the rise worldwide. It is considered the result of complex interactions between genetic and environmental factors, being many of them unknown. Therefore, there is a dire necessity for the identification of novel molecular players for the understanding of this disease. In this data article we determined the protein expression profiles of whole protein extracts from cortex regions of brains from patients with Alzheimer׳s disease in comparison to a normal brain. We identified 721 iTRAQ-labeled polypeptides with more than 95% in confidence. We analyzed all proteins that changed in their expression level and located them in the KEGG metabolic pathways, as well as in the mitochondrial complexes of the electron transport chain and ATP synthase. In addition, we analyzed the over- and sub-expressed polypeptides through IPA software, specifically Core I and Biomarkers I modules. Data in this article is related to the research article "Identification of proteins that are differentially expressed in brains with Alzheimer's disease using iTRAQ labeling and tandem mass spectrometry" (Minjarez et al., 2016) [1]. PMID:27257613

  8. Data set of interactomes and metabolic pathways of proteins differentially expressed in brains with Alzheimer׳s disease.

    PubMed

    Minjarez, Benito; Calderón-González, Karla Grisel; Valero Rustarazo, Ma Luz; Herrera-Aguirre, María Esther; Labra-Barrios, María Luisa; Rincon-Limas, Diego E; Sánchez Del Pino, Manuel M; Mena, Raul; Luna-Arias, Juan Pedro

    2016-06-01

    Alzheimer׳s disease is one of the main causes of dementia in the elderly and its frequency is on the rise worldwide. It is considered the result of complex interactions between genetic and environmental factors, being many of them unknown. Therefore, there is a dire necessity for the identification of novel molecular players for the understanding of this disease. In this data article we determined the protein expression profiles of whole protein extracts from cortex regions of brains from patients with Alzheimer׳s disease in comparison to a normal brain. We identified 721 iTRAQ-labeled polypeptides with more than 95% in confidence. We analyzed all proteins that changed in their expression level and located them in the KEGG metabolic pathways, as well as in the mitochondrial complexes of the electron transport chain and ATP synthase. In addition, we analyzed the over- and sub-expressed polypeptides through IPA software, specifically Core I and Biomarkers I modules. Data in this article is related to the research article "Identification of proteins that are differentially expressed in brains with Alzheimer's disease using iTRAQ labeling and tandem mass spectrometry" (Minjarez et al., 2016) [1].

  9. Pharmacokinetics, metabolism and distribution of PEGs and PEGylated proteins: quo vadis?

    PubMed

    Baumann, Andreas; Tuerck, Dietrich; Prabhu, Saileta; Dickmann, Leslie; Sims, Jennifer

    2014-10-01

    The pharmacokinetics (PK), metabolism and biodistribution of polyethylene glycol (PEG) in PEGylated proteins are important to understand the increased cellular vacuolation reported in various tissues in animals. The tissue distribution profile of PEGylated proteins and 'metabolic' PEG is guided largely by absolute PEG load, PEG molecular weight and, where applicable, receptor-mediated uptake via the protein moiety. High molecular weight PEGs show slow renal clearance, and consequently have a greater potential to accumulate within cells. The intracellular nonbiodegradable PEG can accumulate within the lysosome ultimately causing distension and vacuolation observed by standard histological examinations. Improved bioanalytical methodologies will contribute to the identification of specific PK parameters including distribution behavior to support development of PEGylated proteins as therapeutics.

  10. Role of Protein–Protein Interactions in Cytochrome P450-Mediated Drug Metabolism and Toxicity

    PubMed Central

    2015-01-01

    Through their unique oxidative chemistry, cytochrome P450 monooxygenases (CYPs) catalyze the elimination of most drugs and toxins from the human body. Protein–protein interactions play a critical role in this process. Historically, the study of CYP–protein interactions has focused on their electron transfer partners and allosteric mediators, cytochrome P450 reductase and cytochrome b5. However, CYPs can bind other proteins that also affect CYP function. Some examples include the progesterone receptor membrane component 1, damage resistance protein 1, human and bovine serum albumin, and intestinal fatty acid binding protein, in addition to other CYP isoforms. Furthermore, disruption of these interactions can lead to altered paths of metabolism and the production of toxic metabolites. In this review, we summarize the available evidence for CYP protein–protein interactions from the literature and offer a discussion of the potential impact of future studies aimed at characterizing noncanonical protein–protein interactions with CYP enzymes. PMID:25133307

  11. The effects of dietary protein and amino acids on skeletal metabolism.

    PubMed

    Bihuniak, Jessica D; Insogna, Karl L

    2015-07-15

    Dietary protein is required for optimal skeletal growth and maturation. Although Recommended Dietary Allowances (RDAs) exist for global dietary protein intake, the level and sources of dietary protein that are optimal for skeletal health over the life continuum have not been established. This is partly due to the difficulty in quantifying the effects of variable levels of a nutrient's intake over a lifetime as well as the complex nature of the relationships between dietary protein and calcium economy. Areas of current uncertainty include the precise source and amount of dietary protein required for optimal skeletal accretion and maintenance of skeletal mass, as well as the site-specific effects of dietary protein. The cellular and molecular mechanisms that underpin the actions of dietary protein on mineral metabolism and skeletal homeostasis remain unclear. This review attempts to summarize recent data bearing on these questions. PMID:25843057

  12. Amino acid metabolism, substrate availability and the control of protein dynamics in the human kidney.

    PubMed

    Garibotto, G; Tessari, P; Sacco, P; Deferrari, G

    1999-01-01

    The mechanisms controlling protein metabolism in the human kidney are not well understood. During adult life, kidney protein content and the size of the kidney remain fairly constant, indicating that protein synthesis and degradation within the kidney are tightly regulated. However, kidney protein turnover may change in response to stimuli such as alterations in substrate availability, hormones or growth factors, acid-base balance, renal work or renal injury with a progressive decrease in the number of nephrons. These factors have been evaluated mainly in animals, in vitro or in vivo. Amino acids, the kidneys substrates for protein synthesis, are provided by several routes. Like in other organs, amino acids can reach the kidney cells through the arterial blood flow. However, they may also come from the degradation of reabsorbed low-molecular weight proteins filtered by the glomerulus. The human kidney has high rates of protein turnover and leucine oxidation. The magnitude of the protein turnover across the human kidney suggests that the protein dynamics is partly determined by intrarenal protein catabolism. As evaluated by a steady-state leucine multiple compartment analysis, kidney protein synthesis is dependent to a similar extent on intrarenal generation of amino acids from protein breakdown and from amino acids taken up from the arterial blood. Kidney mass may therefore depend not only on the availability of free amino acids, but also on filtered proteins which are degraded within the kidney. Future studies could define the mechanisms, metabolic pathways and mediators influencing kidney protein turnover in humans, with a view to better comprehension of the mechanisms of disease. PMID:10493563

  13. Role of a liver fatty acid-binding protein gene in lipid metabolism in chicken hepatocytes.

    PubMed

    Gao, G L; Na, W; Wang, Y X; Zhang, H F; Li, H; Wang, Q G

    2015-01-01

    This study investigated the role of the chicken liver fatty acid-binding protein (L-FABP) gene in lipid metabolism in hepatocytes, and the regulatory relationships between L-FABP and genes related to lipid metabolism. The short hairpin RNA (shRNA) interference vector with L-FABP and an eukaryotic expression vector were used. Chicken hepatocytes were subjected to shRNA-mediated knockdown or L-FABP cDNA overexpression. Expression levels of lipid metabolism-related genes and biochemical parameters were detected 24, 36, 48, 60, and 72 h after transfection with the interference or overexpression plasmids for L-FABP, PPARα and L-BABP expression levels, and the total amount of cholesterol, were significantly affected by L-FABP expression. L-FABP may affect lipid metabolism by regulating PPARα and L-BABP in chicken hepatocytes. PMID:25966259

  14. Protein phosphorylation and prevention of cytochrome oxidase inhibition by ATP: coupled mechanisms of energy metabolism regulation.

    PubMed

    Acin-Perez, Rebeca; Gatti, Domenico L; Bai, Yidong; Manfredi, Giovanni

    2011-06-01

    Rapid regulation of oxidative phosphorylation is crucial for mitochondrial adaptation to swift changes in fuels availability and energy demands. An intramitochondrial signaling pathway regulates cytochrome oxidase (COX), the terminal enzyme of the respiratory chain, through reversible phosphorylation. We find that PKA-mediated phosphorylation of a COX subunit dictates mammalian mitochondrial energy fluxes and identify the specific residue (S58) of COX subunit IV-1 (COXIV-1) that is involved in this mechanism of metabolic regulation. Using protein mutagenesis, molecular dynamics simulations, and induced fit docking, we show that mitochondrial energy metabolism regulation by phosphorylation of COXIV-1 is coupled with prevention of COX allosteric inhibition by ATP. This regulatory mechanism is essential for efficient oxidative metabolism and cell survival. We propose that S58 COXIV-1 phosphorylation has evolved as a metabolic switch that allows mammalian mitochondria to rapidly toggle between energy utilization and energy storage.

  15. Role of a liver fatty acid-binding protein gene in lipid metabolism in chicken hepatocytes.

    PubMed

    Gao, G L; Na, W; Wang, Y X; Zhang, H F; Li, H; Wang, Q G

    2015-01-01

    This study investigated the role of the chicken liver fatty acid-binding protein (L-FABP) gene in lipid metabolism in hepatocytes, and the regulatory relationships between L-FABP and genes related to lipid metabolism. The short hairpin RNA (shRNA) interference vector with L-FABP and an eukaryotic expression vector were used. Chicken hepatocytes were subjected to shRNA-mediated knockdown or L-FABP cDNA overexpression. Expression levels of lipid metabolism-related genes and biochemical parameters were detected 24, 36, 48, 60, and 72 h after transfection with the interference or overexpression plasmids for L-FABP, PPARα and L-BABP expression levels, and the total amount of cholesterol, were significantly affected by L-FABP expression. L-FABP may affect lipid metabolism by regulating PPARα and L-BABP in chicken hepatocytes.

  16. Role of the mixed-lineage protein kinase pathway in the metabolic stress response to obesity

    PubMed Central

    Kant, Shashi; Barrett, Tamera; Vertii, Anastassiia; Noh, Yun Hee; Jung, Dae Young; Kim, Jason K.; Davis, Roger J.

    2013-01-01

    Summary Saturated free fatty acid (FFA) is implicated in the metabolic response to obesity. In vitro studies indicate that FFA signaling may be mediated by the mixed-lineage protein kinase (MLK) pathway that activates cJun NH2-terminal kinase (JNK). Here we examined the role of the MLK pathway in vivo using a mouse model of diet-induced obesity. The ubiquitously expressed MLK2 and MLK3 protein kinases have partially redundant functions. We therefore compared wild-type and compound mutant mice that lack expression of MLK2 plus MLK3. MLK-deficiency protected mice against high fat diet-induced insulin resistance and obesity. Reduced JNK activation and increased energy expenditure contribute to the metabolic effects of MLK-deficiency. These data confirm that the MLK pathway plays a critical role in the metabolic response to obesity. PMID:23954791

  17. Sarcocystis fusiformis: some protein metabolic enzymes in various fractions of sarcocysts of buffalo (Bubalus bubalis).

    PubMed

    Gupta, R S; Kushwah, H S; Kushwah, A

    1993-01-01

    An investigation on the relative presence of some protein metabolic enzymes, namely aspartate aminotransferase (AST), alanine aminotransferase (ALT), NAD+ and NADP+ dependent glutamate dehydrogenase (GLDH) and arginase in cyst wall (CW), cyst fluid (CF) and zoite (ZT) fractions of the sarcocysts of Sarcocystis fusiformis in the oesophageal muscles of Indian water buffalo was carried out. Both the transaminases were present in all the fractions of the cyst, although in variable amounts. There was a higher level of AST activity than of ALT activity. AST activity was the highest in ZT, whereas ALT activity was at a maximum in the CF fraction. The levels of activity of NAD+ and NADP+ dependent GLDH and arginase remained beyond detectable limits. The study revealed that the intermediates of carbohydrate metabolism are linked to protein metabolism by transaminases. The possibility of concomitant removal of ammonia and its subsequent incorporation into the urea cycle is ruled out in this parasitic protozoan.

  18. Regulatory mechanism of protein metabolic pathway during the differentiation process of chicken male germ cell.

    PubMed

    Li, Dong; Zuo, Qisheng; Lian, Chao; Zhang, Lei; Shi, Qingqing; Zhang, Zhentao; Wang, Yingjie; Ahmed, Mahmoud F; Tang, Beibei; Xiao, Tianrong; Zhang, Yani; Li, Bichun

    2015-08-01

    We explored the regulatory mechanism of protein metabolism during the differentiation process of chicken male germ cells and provide a basis for improving the induction system of embryonic stem cell differentiation to male germ cells in vitro. We sequenced the transcriptome of embryonic stem cells, primordial germ cells, and spermatogonial stem cells with RNA sequencing (RNA-Seq), bioinformatics analysis methods, and detection of the key genes by quantitative reverse transcription PCR (qRT-PCR). Finally, we found 16 amino acid metabolic pathways enriched in the biological metabolism during the differentiation process of embryonic stem cells to primordial germ cells and 15 amino acid metabolic pathways enriched in the differentiation stage of primordial germ cells to spermatogonial stem cells. We found three pathways, arginine-proline metabolic pathway, tyrosine metabolic pathway, and tryptophan metabolic pathway, significantly enriched in the whole differentiation process of embryonic stem cells to spermatogonial stem cells. Moreover, for these three pathways, we screened key genes such as NOS2, ADC, FAH, and IDO. qRT-PCR results showed that the expression trend of these genes were the same to RNA-Seq. Our findings showed that the three pathways and these key genes play an important role in the differentiation process of embryonic stem cells to male germ cells. These results provide basic information for improving the induction system of embryonic stem cell differentiation to male germ cells in vitro.

  19. [Protein metabolism in the cerebral hemispheres during the emotional-algesic stress].

    PubMed

    Yakushev, V S; Davydov, V V; Bushueva, V V; Skurygin, V P; Krisanova, N V

    1985-01-01

    Emotional-algesic stress causes essential changes in the protein metabolism of cerebral hemispheres. These changes may be of great importance for the functioning of the brain and cause the disturbances of the higher nervous activity when the organism is influenced by the emotional stress factors. PMID:4039861

  20. Modulation of protein quality control systems by food phytochemicals.

    PubMed

    Murakami, Akira

    2013-05-01

    There is compelling evidence showing that dietary phytochemicals have exhibited pronounced bioactivities in a number of experimental models. In addition, a variety of epidemiological surveys have demonstrated that frequent ingestion of vegetables and fruits, which contain abundant phytochemicals, lowers the risk of onset of some diseases. However, the action mechanisms by which dietary phytochemicals show bioactivity remain to be fully elucidated and a fundamental question is why this class of chemicals has great potential for regulating health. Meanwhile, maintenance and repair of biological proteins by molecular chaperones, such as heat shock proteins, and clearance of abnormal proteins by the ubiquitin-proteasome system and autophagy play central roles in health, some disease prevention, and longevity. Interestingly, several recent studies have revealed that phytochemicals, including curcumin (yellow pigment in turmeric), resveratrol (phytoalexin in grapes), quercetin (general flavonol in onions and others), and isothiocyanates (preferentially present in cruciferous vegetables, such as broccoli and cabbage), are remarkable regulators of protein quality control systems, suggesting that their physiological and biological functions are exerted, at least in part, through activation of such unique mechanisms. This review article highlights recent findings regarding the effects of representative phytochemicals on protein quality control systems and their possible molecular mechanisms.

  1. DNA and RNA aptamers as modulators of protein function.

    PubMed

    Ulrich, Henning

    2005-03-01

    The SELEX technique (systematic evolution of ligands by exponential enrichment) is a combinatorial library approach in which DNA or RNA molecules are selected by their ability to bind their protein targets with high affinity and specificity. The isolated molecules are referred to as aptamers (from aptus = Latin "to fit"). First, RNA and DNA aptamers were identified that bind to proteins naturally interacting with nucleic acids, or to small organic molecules such as ATP. In the following years, the use of the SELEX technique was extended to isolate oligonucleotide ligands for a wide range of proteins of importance for therapy, and diagnostics. Since these RNA and DNA molecules bind their targets with similar affinities as antibodies, and are able to distinguish between isotypes of an enzyme, aptamers have been also called synthetic antibodies. Recently, the use of in vitro selection methods to isolate protein inhibitors has been extended to complex targets, such as receptors that are only functional in their membrane-bound form, cells, and trypanosomes. RNA aptamers have been expressed in living cells where they inhibit a protein implicated in intracellular signal transduction. The utility of aptamers for in vivo experiments, and diagnostic and therapeutic purposes, is considerably enhanced by introducing chemical modifications into the oligonucleotides to provide resistance against enzymatic degradation in body fluids. Recently, such inhibitors have been evolved for a great variety of targets, including receptors, growth factors, and adhesion molecules implicated in disease. Furthermore, some results were already obtained in animal models and clinical trials. PMID:16787315

  2. Modulation of protein quality control systems by food phytochemicals

    PubMed Central

    Murakami, Akira

    2013-01-01

    There is compelling evidence showing that dietary phytochemicals have exhibited pronounced bioactivities in a number of experimental models. In addition, a variety of epidemiological surveys have demonstrated that frequent ingestion of vegetables and fruits, which contain abundant phytochemicals, lowers the risk of onset of some diseases. However, the action mechanisms by which dietary phytochemicals show bioactivity remain to be fully elucidated and a fundamental question is why this class of chemicals has great potential for regulating health. Meanwhile, maintenance and repair of biological proteins by molecular chaperones, such as heat shock proteins, and clearance of abnormal proteins by the ubiquitin-proteasome system and autophagy play central roles in health, some disease prevention, and longevity. Interestingly, several recent studies have revealed that phytochemicals, including curcumin (yellow pigment in turmeric), resveratrol (phytoalexin in grapes), quercetin (general flavonol in onions and others), and isothiocyanates (preferentially present in cruciferous vegetables, such as broccoli and cabbage), are remarkable regulators of protein quality control systems, suggesting that their physiological and biological functions are exerted, at least in part, through activation of such unique mechanisms. This review article highlights recent findings regarding the effects of representative phytochemicals on protein quality control systems and their possible molecular mechanisms. PMID:23704811

  3. Oestradiol modulation of serotonin reuptake transporter and serotonin metabolism in the brain of monkeys.

    PubMed

    Sánchez, M G; Morissette, M; Di Paolo, T

    2013-06-01

    Serotonin (5-hydroxytryptamine; 5-HT) is an important brain neurotransmitter that is implicated in mental and neurodegenerative diseases and is modulated by ovarian hormones. Nevertheless, the effect of oestrogens on 5-HT neurotransmission in the primate caudate nucleus, putamen and nucleus accumbens, which are major components of the basal ganglia, and the anterior cerebral cortex, mainly the frontal and cingulate gyrus, is not well documented. The present study evaluated 5-HT reuptake transporter (SERT) and 5-HT metabolism in these brain regions in response to 1-month treatment with 17β-oestradiol in short-term (1 month) ovariectomised (OVX) monkeys (Macaca fascicularis). SERT-specific binding was measured by autoradiography using the radioligand [³H]citalopram. Biogenic amine concentrations were quantified by high-performance liquid chromatography. 17β-Oestradiol increased SERT in the superior frontal cortex and in the anterior cingulate cortex, in the nucleus accumbens, and in subregions of the caudate nucleus of OVX monkeys. 17β-Oestradiol left [³H]citalopram-specific binding unchanged in the putamen, as well as the dorsal and medial raphe nucleus. 17β-Oestradiol treatment decreased striatal concentrations of the precursor of 5-HT, 5-hydroxytryptophan, and increased 5-HT, dopamine and 3-methoxytyramine concentrations in the nucleus accumbens, caudate nucleus and putamen, whereas the concentrations of the metabolites 5-hydroxyindoleacetic acid, 3,4-dihydroxyphenylacetic acid and homovanillic acid remained unchanged. No effect of 17β-oestradiol treatment was observed for biogenic amine concentrations in the cortical regions. A significant positive correlation was observed between [³H]citalopram-specific binding and 5-HT concentrations in the caudate nucleus, putamen and nucleus accumbens, suggesting their link. These results have translational value for women with low oestrogen, such as those in surgical menopause or perimenopause. PMID:23414342

  4. Protein-directed modulation of high-LET hyperthermic radiosensitization

    SciTech Connect

    Chang, P.Y.

    1991-01-01

    A pair of Chinese Hamster Ovary cell lines, the wild-type CHO-SC1, and its temperature-sensitive mutant (CHO-tsH1) was used to examine the importance of protein synthesis in the development of thermotolerance. The classical biphasic thermotolerant survival response to hyperthermia was observed in the SC1 cells after continuous heating at 41.5C to 42.5C, while tsH1 showed no thermotolerance. In separate experiments, each cell line was triggered and challenged at 45C. The heat doses were separated with graded incubaton periods at 35C or 40C for thermotolerance development. SC1 cells expressed thermoresistance, with the synthesis of heat shock proteins, under both incubation conditions. tsH1 cells expressed thermotolerance similar to that seen in the SC1 cells when incubated at 35C, but the survival response with the non-permissive 40C incubation was much reduced in the absence of protein synthesis. The combined effects of heavy-ion radiation and hyperthermia were examined using the same cell system. A mild heat dose of 41.5C was used in conjunction with Neon particle radiation of various high LET values. The cell killing effects were highly dependent on the sequence of application of heat and Neon radiation. Heat applied immediately after Neon irradiation was more cytotoxic to SC1 cells than when heat was applied prior to the irradiation. The ability of cells to synthesize new proteins plays a key role in this sequence-dependent thermal radiosensitization. In the absence of protein synthesis in the tsH1 cells, the high-LET thermal enhancement for cell-killing was unchanged regardless of the sequence. In the presence of protein synthetic activity in the SC1 cells, the thermal enhancement of radiation-induced cell killing was LET-dependent.

  5. Human Glycolipid Transfer Protein (GLTP) Expression Modulates Cell Shape

    PubMed Central

    Gao, Yongguang; Chung, Taeowan; Zou, Xianqiong; Pike, Helen M.; Brown, Rhoderick E.

    2011-01-01

    Glycolipid transfer protein (GLTP) accelerates glycosphingolipid (GSL) intermembrane transfer via a unique lipid transfer/binding fold (GLTP-fold) that defines the GLTP superfamily and is the prototype for GLTP-like domains in larger proteins, i.e. phosphoinositol 4-phosphate adaptor protein-2 (FAPP2). Although GLTP-folds are known to play roles in the nonvesicular intracellular trafficking of glycolipids, their ability to alter cell phenotype remains unexplored. In the present study, overexpression of human glycolipid transfer protein (GLTP) was found to dramatically alter cell phenotype, with cells becoming round between 24 and 48 h after transfection. By 48 h post transfection, ∼70% conversion to the markedly round shape was evident in HeLa and HEK-293 cells, but not in A549 cells. In contrast, overexpression of W96A-GLTP, a liganding-site point mutant with abrogated ability to transfer glycolipid, did not alter cell shape. The round adherent cells exhibited diminished motility in wound healing assays and an inability to endocytose cholera toxin but remained viable and showed little increase in apoptosis as assessed by poly(ADP-ribose) polymerase cleavage. A round cell phenotype also was induced by overexpression of FAPP2, which binds/transfers glycolipid via its C-terminal GLTP-like fold, but not by a plant GLTP ortholog (ACD11), which is incapable of glycolipid binding/transfer. Screening for human protein partners of GLTP by yeast two hybrid screening and by immuno-pulldown analyses revealed regulation of the GLTP-induced cell rounding response by interaction with δ-catenin. Remarkably, while δ-catenin overexpression alone induced dendritic outgrowths, coexpression of GLTP along with δ-catenin accelerated transition to the rounded phenotype. The findings represent the first known phenotypic changes triggered by GLTP overexpression and regulated by direct interaction with a p120-catenin protein family member. PMID:21625605

  6. A Mitochondrial ATP synthase Subunit Interacts with TOR Signaling to Modulate Protein Homeostasis and Lifespan in Drosophila

    PubMed Central

    Sun, Xiaoping; Wheeler, Charles T.; Yolitz, Jason; Laslo, Mara; Alberico, Thomas; Sun, Yaning; Song, Qisheng; Zou, Sige

    2014-01-01

    SUMMARY Diet composition is a critical determinant of lifespan and nutrient imbalance is detrimental health. However, how nutrients interact with genetic factors to modulate lifespan remains elusive. We investigated how diet composition influences mitochondrial ATP synthase subunit d (ATPsyn-d) in modulating lifespan in Drosophila. ATPsyn-d knockdown extended lifespan in females fed low carbohydrate-to-protein (C:P) diets, but not the high C:P ratio diet. This extension was associated with increased resistance to oxidative stress, transcriptional changes in metabolism, proteostasis and immune genes, reduced protein damage and aggregation, and reduced phosphorylation of S6K and ERK in TOR and MAPK signaling, respectively. ATPsyn-d knockdown did not extend lifespan in females with reduced TOR signaling induced genetically by Tsc2 overexpression or pharmacologically by rapamycin. Our data reveal a link among diet, mitochondria, MAPK and TOR signaling in aging and stresses the importance of considering genetic background and diet composition in implementing interventions for promoting healthy aging. PMID:25220459

  7. Estrogen Modulates Expression of Tight Junction Proteins in Rat Vagina

    PubMed Central

    Oh, Kyung-Jin; Ahn, Kyuyoun

    2016-01-01

    Background. The objectives of this study were to investigate the localization of tight junctions and the modulation of zonula occludens- (ZO-) 1, occludin and claudin-1 expression by estrogen in castrated female rat vagina. Female Sprague-Dawley rats (230–240 g, n = 45) were divided into three groups and subjected to a sham operation (control group, n = 15), bilateral ovariectomy (Ovx group, n = 15), or bilateral ovariectomy followed by daily subcutaneous injection of 17β-estradiol (50 μg/kg/day, Ovx + Est group, n = 15). The cellular localization and expression of ZO-1, occludin, and claudin-1 were determined in each group by immunohistochemistry and western blot. Results. Expression of ZO-1 was diffuse in all groups, with the highest intensity in the superficial epithelium in the control group. Occludin was localized in the intermediate and basal epithelium. Claudin-1 was most intense in the superficial layer of the vaginal epithelium in the control group. Expression of ZO-1, occludin, and claudin-1 was significantly decreased after ovariectomy and was restored to the level of the control after estrogen replacement. Conclusions. Tight junctions are distinctly localized in rat vagina, and estrogen modulates the expression of tight junctions. Further researches are needed to clarify the functional role of tight junctions in vaginal lubrication. PMID:27127786

  8. Structural basis for modulation of a G-protein-coupled receptor by allosteric drugs.

    PubMed

    Dror, Ron O; Green, Hillary F; Valant, Celine; Borhani, David W; Valcourt, James R; Pan, Albert C; Arlow, Daniel H; Canals, Meritxell; Lane, J Robert; Rahmani, Raphaël; Baell, Jonathan B; Sexton, Patrick M; Christopoulos, Arthur; Shaw, David E

    2013-11-14

    The design of G-protein-coupled receptor (GPCR) allosteric modulators, an active area of modern pharmaceutical research, has proved challenging because neither the binding modes nor the molecular mechanisms of such drugs are known. Here we determine binding sites, bound conformations and specific drug-receptor interactions for several allosteric modulators of the M2 muscarinic acetylcholine receptor (M2 receptor), a prototypical family A GPCR, using atomic-level simulations in which the modulators spontaneously associate with the receptor. Despite substantial structural diversity, all modulators form cation-π interactions with clusters of aromatic residues in the receptor extracellular vestibule, approximately 15 Å from the classical, 'orthosteric' ligand-binding site. We validate the observed modulator binding modes through radioligand binding experiments on receptor mutants designed, on the basis of our simulations, either to increase or to decrease modulator affinity. Simulations also revealed mechanisms that contribute to positive and negative allosteric modulation of classical ligand binding, including coupled conformational changes of the two binding sites and electrostatic interactions between ligands in these sites. These observations enabled the design of chemical modifications that substantially alter a modulator's allosteric effects. Our findings thus provide a structural basis for the rational design of allosteric modulators targeting muscarinic and possibly other GPCRs.

  9. Structural basis for modulation of a G-protein-coupled receptor by allosteric drugs

    NASA Astrophysics Data System (ADS)

    Dror, Ron O.; Green, Hillary F.; Valant, Celine; Borhani, David W.; Valcourt, James R.; Pan, Albert C.; Arlow, Daniel H.; Canals, Meritxell; Lane, J. Robert; Rahmani, Raphaël; Baell, Jonathan B.; Sexton, Patrick M.; Christopoulos, Arthur; Shaw, David E.

    2013-11-01

    The design of G-protein-coupled receptor (GPCR) allosteric modulators, an active area of modern pharmaceutical research, has proved challenging because neither the binding modes nor the molecular mechanisms of such drugs are known. Here we determine binding sites, bound conformations and specific drug-receptor interactions for several allosteric modulators of the M2 muscarinic acetylcholine receptor (M2 receptor), a prototypical family A GPCR, using atomic-level simulations in which the modulators spontaneously associate with the receptor. Despite substantial structural diversity, all modulators form cation-π interactions with clusters of aromatic residues in the receptor extracellular vestibule, approximately 15Å from the classical, `orthosteric' ligand-binding site. We validate the observed modulator binding modes through radioligand binding experiments on receptor mutants designed, on the basis of our simulations, either to increase or to decrease modulator affinity. Simulations also revealed mechanisms that contribute to positive and negative allosteric modulation of classical ligand binding, including coupled conformational changes of the two binding sites and electrostatic interactions between ligands in these sites. These observations enabled the design of chemical modifications that substantially alter a modulator's allosteric effects. Our findings thus provide a structural basis for the rational design of allosteric modulators targeting muscarinic and possibly other GPCRs.

  10. Recent Progress on Bile Acid Receptor Modulators for Treatment of Metabolic Diseases.

    PubMed

    Xu, Yanping

    2016-07-28

    Bile acids are steroid-derived molecules synthesized in the liver, secreted from hepatocytes into the bile canaliculi, and subsequently stored in the gall bladder. During the feeding, bile flows into the duodenum, where it contributes to the solubilization and digestion of lipid-soluble nutrients. After a meal, bile-acid levels increase in the intestine, liver, and also in the systemic circulation. Therefore, serum bile-acid levels serve as an important sensing mechanism for nutrient and energy. Recent studies have described bile acids as versatile signaling molecules endowed with systemic endocrine functions. Bile acids are ligands for G-protein coupled receptors (GPCRs) such as TGR5 (also known as GPBAR1, M-BAR, and BG37) and nuclear hormone receptors including farnesoid X receptor (FXR; also known as NR1H4). Acting through these diverse signaling pathways, bile acids regulate triglyceride, cholesterol, glucose homeostasis, and energy expenditure. These bile-acid-controlled signaling pathways have become the source of promising novel drug targets to treat common metabolic and hepatic diseases. PMID:26878262

  11. Modulation of HDL metabolism by the niacin receptor GPR109A in mouse hepatocytes.

    PubMed

    Li, Xiaoyu; Millar, John S; Brownell, Nicholas; Briand, François; Rader, Daniel J

    2010-11-01

    The niacin receptor GPR109A is a G(i)-protein-coupled receptor which mediates the effects of niacin on inhibiting intracellular triglyceride lipolysis in adipocytes. However, the role of GPR109A in mediating the effects of niacin on high density lipoprotein (HDL) metabolism is unclear. We found niacin has no effect on HDL-C in GPR109A knockout mice. Furthermore, niacin lowered intracellular cAMP in primary hepatocytes mediated by GPR109A. We used an adeno-associated viral (AAV) serotype 8 vector encoding GPR109A under the control of the hepatic-specific thyroxine-binding globulin promoter to specifically overexpress GPR109A in mouse liver. Plasma HDL-C, hepatic ABCA1 and the HDL cholesterol production rate were significantly reduced in mice overexpressing GPR109A. Overexpression of GPR109A reduced primary hepatocyte free cholesterol efflux to apoA-I; conversely, GPR109A deficient hepatocytes had increased ABCA1-mediated cholesterol efflux. These data support the concept that the HDL-C lowering effect of niacin in wild-type mice is mediated through stimulation of GPR109A in hepatocytes; such an effect then leads to reduced hepatocyte ABCA1 expression and activity, decreased cholesterol efflux to nascent apoA-I, and reduced HDL-C levels. These results indicate that niacin-mediated activation of GP109A in liver lowers ABCA1 expression leading to reduced hepatic cholesterol efflux to HDL.

  12. Intein Applications: From Protein Purification and Labeling to Metabolic Control Methods*

    PubMed Central

    Wood, David W.; Camarero, Julio A.

    2014-01-01

    The discovery of inteins in the early 1990s opened the door to a wide variety of new technologies. Early engineered inteins from various sources allowed the development of self-cleaving affinity tags and new methods for joining protein segments through expressed protein ligation. Some applications were developed around native and engineered split inteins, which allow protein segments expressed separately to be spliced together in vitro. More recently, these early applications have been expanded and optimized through the discovery of highly efficient trans-splicing and trans-cleaving inteins. These new inteins have enabled a wide variety of applications in metabolic engineering, protein labeling, biomaterials construction, protein cyclization, and protein purification. PMID:24700459

  13. Concentration dependent effects of tobacco particulates from different types of cigarettes on expression of drug metabolizing proteins, and benzo(a)pyrene metabolism in primary normal human oral epithelial cells

    PubMed Central

    Sacks, Peter G.; Zhao, Zhong-Lin; Kosinska, Wieslawa; Fleisher, Kenneth E.; Gordon, Terry; Guttenplan, Joseph B.

    2011-01-01

    The ability of tobacco smoke (TS) to modulate phase I and II enzymes and affect metabolism of tobacco carcinogens is likely an important factor in its carcinogenicity. For the first time several types of TS particulates (TSP) were compared in different primary cultured human oral epithelial cells (NOE) for their abilities to affect metabolism of the tobacco carcinogen, (BaP) to genotoxic products, and expression of drug metabolizing enzymes. TSP from, reference filtered (2RF4), mentholated (MS), reference unfiltered, (IR3), ultra low tar (UL), and cigarettes that primarily heat tobacco (ECL) were tested. Cells pretreated with TSP concentrations of 0.2 – 10 µg/ml generally showed increased rates of BaP metabolism; those treated with TSP concentrations above 10 µg/ml showed decreased rates. Effects of TSPs were similar when expressed on a weight basis. Weights of TSP/cigarette varied in the order: MS ≈ IR3 > 2RF4 > ECL > UL. All TSPs induced the phase I proteins, cytochrome P450 1A1 (CYP1A1) and 1B1 (CYP1B1), phase II proteins, NAD(P)H dehydrogenase quinone 1 (NQO1), and microsomal glutathione S-transferase 1 (MGST1), and additionally, hydroxysteroid (17-beta) dehydrogenase 2 (HSD17B2), as assessed by qRT-PCR. The pattern of gene induction at probable physiological levels favored activation over detoxification. PMID:21722697

  14. Temperature-dependent solvation modulates the dimensions of disordered proteins

    PubMed Central

    Wuttke, René; Hofmann, Hagen; Nettels, Daniel; Borgia, Madeleine B.; Mittal, Jeetain; Best, Robert B.; Schuler, Benjamin

    2014-01-01

    For disordered proteins, the dimensions of the chain are an important property that is sensitive to environmental conditions. We have used single-molecule Förster resonance energy transfer to probe the temperature-induced chain collapse of five unfolded or intrinsically disordered proteins. Because this behavior is sensitive to the details of intrachain and chain–solvent interactions, the collapse allows us to probe the physical interactions governing the dimensions of disordered proteins. We find that each of the proteins undergoes a collapse with increasing temperature, with the most hydrophobic one, λ-repressor, undergoing a reexpansion at the highest temperatures. Although such a collapse might be expected due to the temperature dependence of the classical “hydrophobic effect,” remarkably we find that the largest collapse occurs for the most hydrophilic, charged sequences. Using a combination of theory and simulation, we show that this result can be rationalized in terms of the temperature-dependent solvation free energies of the constituent amino acids, with the solvation properties of the most hydrophilic residues playing a large part in determining the collapse. PMID:24706910

  15. Temperature-dependent solvation modulates the dimensions of disordered proteins.

    PubMed

    Wuttke, René; Hofmann, Hagen; Nettels, Daniel; Borgia, Madeleine B; Mittal, Jeetain; Best, Robert B; Schuler, Benjamin

    2014-04-01

    For disordered proteins, the dimensions of the chain are an important property that is sensitive to environmental conditions. We have used single-molecule Förster resonance energy transfer to probe the temperature-induced chain collapse of five unfolded or intrinsically disordered proteins. Because this behavior is sensitive to the details of intrachain and chain-solvent interactions, the collapse allows us to probe the physical interactions governing the dimensions of disordered proteins. We find that each of the proteins undergoes a collapse with increasing temperature, with the most hydrophobic one, λ-repressor, undergoing a reexpansion at the highest temperatures. Although such a collapse might be expected due to the temperature dependence of the classical "hydrophobic effect," remarkably we find that the largest collapse occurs for the most hydrophilic, charged sequences. Using a combination of theory and simulation, we show that this result can be rationalized in terms of the temperature-dependent solvation free energies of the constituent amino acids, with the solvation properties of the most hydrophilic residues playing a large part in determining the collapse.

  16. Correction of defective protein kinesis of human P-glycoprotein mutants by substrates and modulators.

    PubMed

    Loo, T W; Clarke, D M

    1997-01-10

    There is growing evidence that abnormal protein folding or trafficking (protein kinesis) leads to diseases. We have used P-glycoprotein as a model protein to develop strategies to overcome defects in protein kinesis. Misprocessed mutants of the human P-glycoprotein are retained in the endoplasmic reticulum as core-glycosylated biosynthetic intermediates and rapidly degraded. Synthesis of the mutant proteins in the presence of drug substrates or modulators such as capsaicin, cyclosporin, vinblastine, or verapamil, however, resulted in the appearance of a fully glycosylated and functional protein at the cell surface. These effects were dose-dependent and occurred within a few hours after the addition of substrate. The ability to facilitate processing of the misfolded mutants appeared to be independent of the cell lines used and location of the mutation. P-glycoproteins with mutations in transmembrane segments, extracellular or cytoplasmic loops, the nucleotide-binding domains, or the linker region were processed to the fully mature form in the presence of these substrates. These drug substrates or modulators acted as specific chemical chaperones for P-glycoprotein because they were ineffective on the deltaF508 mutant of cystic fibrosis transmembrane conductance regulator. Therefore, one possible strategy to prevent protein misfolding is to carry out synthesis in the presence of specific substrates or modulators of the protein.

  17. Metabolism

    MedlinePlus

    Metabolism refers to all the physical and chemical processes in the body that convert or use energy, ... Tortora GJ, Derrickson BH. Metabolism. In: Tortora GJ, Derrickson BH. Principles of Anatomy and Physiology . 14th ed. Hoboken, NJ: John H Wiley and Sons; 2013: ...

  18. High Throughput Screening for Drugs that Modulate Intermediate Filament Proteins

    PubMed Central

    Sun, Jingyuan; Groppi, Vincent E.; Gui, Honglian; Chen, Lu; Xie, Qing; Liu, Li

    2016-01-01

    Intermediate filament (IF) proteins have unique and complex cell and tissue distribution. Importantly, IF gene mutations cause or predispose to more than 80 human tissue-specific diseases (IF-pathies), with the most severe disease phenotypes being due to mutations at conserved residues that result in a disrupted IF network. A critical need for the entire IF-pathy field is the identification of drugs that can ameliorate or cure these diseases, particularly since all current therapies target the IF-pathy complication, such as diabetes or cardiovascular disease, rather than the mutant IF protein or gene. We describe a high throughput approach to identify drugs that can normalize disrupted IF proteins. This approach utilizes transduction of lentivirus that expresses green-fluorescent-protein-tagged keratin 18 (K18) R90C in A549 cells. The readout is drug ‘hits’ that convert the dot-like keratin filament distribution, due to the R90C mutation, to a wildtype-like filamentous array. A similar strategy can be used to screen thousands of compounds and can be utilized for practically any IF protein with a filament-disrupting mutation, and could therefore potentially target many IF-pathies. ‘Hits’ of interest require validation in cell culture then using in vivo experimental models. Approaches to study the mechanism of mutant-IF normalization by potential drugs of interest are also described. The ultimate goal of this drug screening approach is to identify effective and safe compounds that can potentially be tested for clinical efficacy in patients. PMID:26795471

  19. Model based engineering of Pichia pastoris central metabolism enhances recombinant protein production

    PubMed Central

    Nocon, Justyna; Steiger, Matthias G.; Pfeffer, Martin; Sohn, Seung Bum; Kim, Tae Yong; Maurer, Michael; Rußmayer, Hannes; Pflügl, Stefan; Ask, Magnus; Haberhauer-Troyer, Christina; Ortmayr, Karin; Hann, Stephan; Koellensperger, Gunda; Gasser, Brigitte; Lee, Sang Yup; Mattanovich, Diethard

    2014-01-01

    The production of recombinant proteins is frequently enhanced at the levels of transcription, codon usage, protein folding and secretion. Overproduction of heterologous proteins, however, also directly affects the primary metabolism of the producing cells. By incorporation of the production of a heterologous protein into a genome scale metabolic model of the yeast Pichia pastoris, the effects of overproduction were simulated and gene targets for deletion or overexpression for enhanced productivity were predicted. Overexpression targets were localized in the pentose phosphate pathway and the TCA cycle, while knockout targets were found in several branch points of glycolysis. Five out of 9 tested targets led to an enhanced production of cytosolic human superoxide dismutase (hSOD). Expression of bacterial β-glucuronidase could be enhanced as well by most of the same genetic modifications. Beneficial mutations were mainly related to reduction of the NADP/H pool and the deletion of fermentative pathways. Overexpression of the hSOD gene itself had a strong impact on intracellular fluxes, most of which changed in the same direction as predicted by the model. In vivo fluxes changed in the same direction as predicted to improve hSOD production. Genome scale metabolic modeling is shown to predict overexpression and deletion mutants which enhance recombinant protein production with high accuracy. PMID:24853352

  20. Cytochromes and iron sulfur proteins in sulfur metabolism of phototrophic bacteria

    NASA Technical Reports Server (NTRS)

    Fischer, U.

    1985-01-01

    Dissimilatory sulfur metabolism in phototrophic sulfur bacteria provides the bacteria with electrons for photosynthetic electron transport chain and, with energy. Assimilatory sulfate reduction is necessary for the biosynthesis of sulfur-containing cell components. Sulfide, thiosulfate, and elemental sulfur are the sulfur compounds most commonly used by phototrophic bacteria as electron donors for anoxygenic photosynthesis. Cytochromes or other electron transfer proteins, like high-potential-iron-sulfur protein (HIPIP) function as electron acceptors or donors for most enzymatic steps during the oxidation pathways of sulfide or thiosulfate. Yet, heme- or siroheme-containing proteins themselves undergo enzymatic activities in sulfur metabolism. Sirohemes comprise a porphyrin-like prosthetic group of sulfate reductase. eenzymatic reactions involve electron transfer. Electron donors or acceptors are necessary for each reaction. Cytochromes and iron sulfur problems, are able to transfer electrons.

  1. Effector proteins that modulate plant--insect interactions.

    PubMed

    Hogenhout, Saskia A; Bos, Jorunn I B

    2011-08-01

    Insect herbivores have highly diverse life cycles and feeding behaviors. They establish close interactions with their plant hosts and suppress plant defenses. Chewing herbivores evoke characteristic defense responses distinguishable from general mechanical damage. In addition, piercing-sucking hemipteran insects display typical feeding behavior that suggests active suppression of plant defense responses. Effectors that modulate plant defenses have been identified in the saliva of these insects. Tools for high-throughput effector identification and functional characterization have been developed. In addition, in some insect species it is possible to silence gene expression by RNAi. Together, this technological progress has enabled the identification of insect herbivore effectors and their targets that will lead to the development of novel strategies for pest resistances in plants.

  2. Development of dimeric modulators for anti-apoptotic Bcl-2 proteins

    PubMed Central

    Wang, Liangyou; Kong, Fansen; Kokoski, Candis; Andrews, David W.; Xing, Chengguo

    2008-01-01

    Bcl-2 family proteins can be classified into two subfamilies – anti-apoptotic members and pro-apoptotic members. Mechanistically, these two subfamilies can antagonize each other through heterodimerization while homodimerization have been proposed for each subfamily to carry out their corresponding anti-apoptotic or pro-apoptotic functions. To date, many small-molecule antagonists against anti-apoptotic Bcl-2 proteins have been developed, which are monomeric modulators. In this study, a series of BH3I-1 based dimeric modulators were developed with structure-activity relationship explored. Dimeric modulators compared to the monomeric antagonists have enhanced binding activity against anti-apoptotic Bcl-2 proteins. In addition, the acidic functional group was demonstrated to be critical for the binding interaction of the small-molecule antagonists with anti-apoptotic Bcl-2 proteins. Finally, the representative dimeric modulator revealed enhanced activity in inducing cytochrome c release from mitochondria compared to its monomeric counterpart. Taken together, dimerization of monomeric modulators is one practical approach to enhance the bioactivity of Bcl-2 antagonists. PMID:18023349

  3. 17β Estradiol Modulates Perfusion Pressure and Expression of 5-LOX and CYP450 4A in the Isolated Kidney of Metabolic Syndrome Female Rats

    PubMed Central

    Zúñiga-Muñoz, A. M.; Guarner Lans, V.; Soria-Castro, E.; Diaz-Diaz, E.; Torrico-Lavayen, R.; Tena-Betancourt, E.; Pérez-Torres, I.

    2015-01-01

    Prevalence of metabolic syndrome and progression of nephropathy depend on sex. We examined a protective effect of estradiol against nephropathy in metabolic syndrome through the modulation of the arachidonic acid metabolism by activating the 5-lipoxygenase and cytochrome p450 4A pathways. 28 female Wistar rats were divided into four groups of seven animals each: control, intact metabolic syndrome, ovariectomized metabolic syndrome, and metabolic syndrome ovariectomized plus estradiol. Blood pressure, body weight, body fat, triglycerides, insulin, HOMA-index, albuminuria, and TNF-α were increased in ovariectomized metabolic syndrome rats (p < 0.001). The perfusion pressure in isolated kidneys of ovariectomized metabolic syndrome rats in presence of 4 μg of arachidonic acid was increased. The inhibitors of the arachidonic acid metabolism Baicalein, Miconazole, and Indomethacin in these rats decreased the perfusion pressure by 57.62%, 99.83%, and 108.5%, respectively and they decreased creatinine clearance and the arachidonic acid percentage. Phospholipase A2 expression in the kidney of ovariectomized metabolic syndrome rats was not modified. 5-lipoxygenase was increased in metabolic syndrome ovariectomized rats while cytochrome p450 4A was decreased. In conclusion, the loss of estradiol increases renal damage while the treatment with estradiol benefits renal function by modulating arachidonic acid metabolism through the 5-lipoxygenase and cytochrome p450 4A pathways. PMID:26491436

  4. Peanut protein structure, polyphenol content and immune response to peanut proteins in vivo are modulated by laccase.

    PubMed

    Mihajlovic, L; Radosavljevic, J; Nordlund, E; Krstic, M; Bohn, T; Smit, J; Buchert, J; Cirkovic Velickovic, T

    2016-05-18

    Food texture can be improved by enzyme-mediated covalent cross-linking of different food components, such as proteins and carbohydrates. Cross-linking changes the biological and immunological properties of proteins and may change the sensitizing potential of food allergens. In this study we applied a microbial polyphenol oxidase, laccase, to cross-link peanut proteins. The size and morphology of the obtained cross-linked proteins were analyzed by electrophoresis and electron microscopy. Structural changes in proteins were analyzed by CD spectroscopy and by using specific antibodies to major peanut allergens. The bioavailability of peanut proteins was analyzed using a Caco-2 epithelial cell model. The in vivo sensitizing potential of laccase-treated peanut proteins was analyzed using a mouse model of food allergy. Finally, peanut polyphenols were analyzed by UHPLC-MS/MS, before and after the enzymatic reaction with laccase. Laccase treatment of peanut proteins yielded a covalently cross-linked material, with the modified tertiary structure of peanut proteins, improved bioavailability of Ara h 2 (by 70 fold, p < 0.05) and modulated allergic immune response in vivo. The modulation of the immune response was related to the increased production of IgG2a antibodies 11 fold (p < 0.05) and reduced IL-13 secretion in in vitro cultured splenocytes 7 fold (p < 0.05). Analysis of the peanut polyphenol content and profile by HPLC-MS/MS revealed that laccase treatment depleted the peanut extract of polyphenol compounds leaving mostly isorhamnetin derivatives and procyanidin dimer B-type in detectable amounts. Treatment of complex food extracts rich in polyphenols with laccase results in both protein cross-linking and modification of polyphenol compounds. These extensively cross-linked proteins have unchanged potency to induce allergic sensitization in vivo, but certain immunomodulatory changes were observed.

  5. Peanut protein structure, polyphenol content and immune response to peanut proteins in vivo are modulated by laccase.

    PubMed

    Mihajlovic, L; Radosavljevic, J; Nordlund, E; Krstic, M; Bohn, T; Smit, J; Buchert, J; Cirkovic Velickovic, T

    2016-05-18

    Food texture can be improved by enzyme-mediated covalent cross-linking of different food components, such as proteins and carbohydrates. Cross-linking changes the biological and immunological properties of proteins and may change the sensitizing potential of food allergens. In this study we applied a microbial polyphenol oxidase, laccase, to cross-link peanut proteins. The size and morphology of the obtained cross-linked proteins were analyzed by electrophoresis and electron microscopy. Structural changes in proteins were analyzed by CD spectroscopy and by using specific antibodies to major peanut allergens. The bioavailability of peanut proteins was analyzed using a Caco-2 epithelial cell model. The in vivo sensitizing potential of laccase-treated peanut proteins was analyzed using a mouse model of food allergy. Finally, peanut polyphenols were analyzed by UHPLC-MS/MS, before and after the enzymatic reaction with laccase. Laccase treatment of peanut proteins yielded a covalently cross-linked material, with the modified tertiary structure of peanut proteins, improved bioavailability of Ara h 2 (by 70 fold, p < 0.05) and modulated allergic immune response in vivo. The modulation of the immune response was related to the increased production of IgG2a antibodies 11 fold (p < 0.05) and reduced IL-13 secretion in in vitro cultured splenocytes 7 fold (p < 0.05). Analysis of the peanut polyphenol content and profile by HPLC-MS/MS revealed that laccase treatment depleted the peanut extract of polyphenol compounds leaving mostly isorhamnetin derivatives and procyanidin dimer B-type in detectable amounts. Treatment of complex food extracts rich in polyphenols with laccase results in both protein cross-linking and modification of polyphenol compounds. These extensively cross-linked proteins have unchanged potency to induce allergic sensitization in vivo, but certain immunomodulatory changes were observed. PMID:27138276

  6. Effects of dairy protein and fat on the metabolic syndrome and type 2 diabetes.

    PubMed

    Bjørnshave, Ann; Hermansen, Kjeld

    2014-01-01

    The incidence of the metabolic syndrome (MetS) and type 2 diabetes (T2D) is increasing worldwide. Evidence supports a negative relationship between the consumption of dairy products and risk of MetS and T2D. Dairy proteins are known to have a directly beneficial effect on hypertension, dyslipidemia, and hyperglycemia, but a detailed understanding of the underlying mechanisms is missing. It has been confirmed by observations that the insulinotropic effect of dairy proteins is associated with the amino acid composition; in particular branched-chain amino acids (BCAA) seem to be of vital importance. Dairy protein-derived peptides may also contribute to the insulinotropic effect via dipeptidyl peptidase-4 (DPP-4) inhibitory activity, and may lower the blood pressure (BP). The lipid metabolism may be improved by whey protein (WP), which acts to reduce the postprandial triglyceride (TG) response. The effect of dairy fat is much more controversial because of the potentially harmful effect exerted by saturated fatty acid (SFA) on metabolic health. Recent observations suggest less adverse effects of SFA on metabolic health than previous assumed. However, little is known about dairy lipid fractions belonging to the groups of monounsaturated fatty acids (MUFA), polyunsaturated fatty acids (PUFA), and phospholipids (PL). Dairy fat seems to act differently depending on the dairy product and the composition of macronutrients in the meal. Therefore, for a better understanding of the mechanisms behind the dairy protein and fat effect on MetS, we suggest that more human studies should be carried out to clarify the interactions of dairy protein and fat with macronutrients in the meal and other dairy components, such as micronutrients and microorganisms from fermented products.

  7. Effects of Dairy Protein and Fat on the Metabolic Syndrome and Type 2 Diabetes

    PubMed Central

    Bjørnshave, Ann; Hermansen, Kjeld

    2014-01-01

    The incidence of the metabolic syndrome (MetS) and type 2 diabetes (T2D) is increasing worldwide. Evidence supports a negative relationship between the consumption of dairy products and risk of MetS and T2D. Dairy proteins are known to have a directly beneficial effect on hypertension, dyslipidemia, and hyperglycemia, but a detailed understanding of the underlying mechanisms is missing. It has been confirmed by observations that the insulinotropic effect of dairy proteins is associated with the amino acid composition; in particular branched-chain amino acids (BCAA) seem to be of vital importance. Dairy protein-derived peptides may also contribute to the insulinotropic effect via dipeptidyl peptidase-4 (DPP-4) inhibitory activity, and may lower the blood pressure (BP). The lipid metabolism may be improved by whey protein (WP), which acts to reduce the postprandial triglyceride (TG) response. The effect of dairy fat is much more controversial because of the potentially harmful effect exerted by saturated fatty acid (SFA) on metabolic health. Recent observations suggest less adverse effects of SFA on metabolic health than previous assumed. However, little is known about dairy lipid fractions belonging to the groups of monounsaturated fatty acids (MUFA), polyunsaturated fatty acids (PUFA), and phospholipids (PL). Dairy fat seems to act differently depending on the dairy product and the composition of macronutrients in the meal. Therefore, for a better understanding of the mechanisms behind the dairy protein and fat effect on MetS, we suggest that more human studies should be carried out to clarify the interactions of dairy protein and fat with macronutrients in the meal and other dairy components, such as micronutrients and microorganisms from fermented products. PMID:25396403

  8. The Role of the Renal Ammonia Transporter Rhcg in Metabolic Responses to Dietary Protein

    PubMed Central

    Bounoure, Lisa; Ruffoni, Davide; Müller, Ralph; Kuhn, Gisela Anna; Devuyst, Olivier

    2014-01-01

    High dietary protein imposes a metabolic acid load requiring excretion and buffering by the kidney. Impaired acid excretion in CKD, with potential metabolic acidosis, may contribute to the progression of CKD. Here, we investigated the renal adaptive response of acid excretory pathways in mice to high-protein diets containing normal or low amounts of acid-producing sulfur amino acids (SAA) and examined how this adaption requires the RhCG ammonia transporter. Diets rich in SAA stimulated expression of enzymes and transporters involved in mediating NH4+ reabsorption in the thick ascending limb of the loop of Henle. The SAA-rich diet increased diuresis paralleled by downregulation of aquaporin-2 (AQP2) water channels. The absence of Rhcg transiently reduced NH4+ excretion, stimulated the ammoniagenic pathway more strongly, and further enhanced diuresis by exacerbating the downregulation of the Na+/K+/2Cl− cotransporter (NKCC2) and AQP2, with less phosphorylation of AQP2 at serine 256. The high protein acid load affected bone turnover, as indicated by higher Ca2+ and deoxypyridinoline excretion, phenomena exaggerated in the absence of Rhcg. In animals receiving a high-protein diet with low SAA content, the kidney excreted alkaline urine, with low levels of NH4+ and no change in bone metabolism. Thus, the acid load associated with high-protein diets causes a concerted response of various nephron segments to excrete acid, mostly in the form of NH4+, that requires Rhcg. Furthermore, bone metabolism is altered by a high-protein acidogenic diet, presumably to buffer the acid load. PMID:24652796

  9. Modulation of Membrane Protein Lateral Mobility by Polyphosphates and Polyamines

    NASA Astrophysics Data System (ADS)

    Schindler, Melvin; Koppel, Dennis E.; Sheetz, Michael P.

    1980-03-01

    The lateral mobility of fluorescein-labeled membrane glycoproteins was measured in whole unlysed erythrocytes and erythrocyte ghosts by the technique of ``fluorescence redistribution after fusion.'' Measurements were made on polyethylene glycol-fused cell pairs in which only one member of the couplet was initially fluorescently labeled. Diffusion coefficients were estimated from the rate of fluorescence redistribution determined from successive scans with a focused laser beam across individual fused pairs. This technique allows for the analysis of diffusion within cell membranes without the possible damaging photochemical events caused by photobleaching. It was found that lateral mobility of erythrocyte proteins can be increased by the addition of polyphosphates (i.e., ATP and 2,3-diphosphoglycerate) and decreased by the addition of organic polyamines (i.e., neomycin and spermine). This control is exerted by these molecules only when they contact the cytoplasmic side of the membrane and is not dependent upon high-energy phosphates. Microviscosity experiments employing diphenylhexatriene demonstrated no changes in membrane lipid state as a function of these reagents. Our results, in conjunction with data on the physical interactions of cytoskeletal proteins, suggest that the diffusion effector molecules alter the lateral mobility of erythrocyte membrane proteins through modifications of interactions in the shell, which is composed of spectrin, actin, and component 4.1.

  10. Axons modulate the expression of proteolipid protein in the CNS.

    PubMed

    Scherer, S S; Vogelbacker, H H; Kamholz, J

    1992-06-01

    We examined the expression of mRNA encoding proteolipid protein (PLP), the major myelin protein in the CNS, in developing rat cerebrum, and in normal and degenerating optic nerves. PLP transcripts were initiated at two clusters of start sites that were separated by about 30 base pairs. During the peak of PLP mRNA expression in developing cerebrum, a higher proportion of PLP transcripts were initiated from the distal start site, furthest from the open reading frame, than in mature cerebrum. We enucleated one eye of immature rats to cause Wallerian degeneration in the optic nerve. In these degenerating optic nerves, the steady state levels of PLP mRNA fell markedly, and the proportion of distally initiated PLP transcripts declined to the same proportion found in normal adult nerves. Changes in myelin gene expression were not limited to PLP mRNA, as the steady-state levels of myelin basic protein (MBP) mRNA paralleled those of PLP mRNA in the developing cerebrum and in degenerating optic nerves. Thus, oligodendrocytes require axons to maintain their normal levels of PLP and MBP transcripts and the high proportion of distally initiated PLP transcripts that characterize early myelination.

  11. MicroRNA-194 Modulates Glucose Metabolism and Its Skeletal Muscle Expression Is Reduced in Diabetes

    PubMed Central

    Latouche, Celine; Natoli, Alaina; Reddy-Luthmoodoo, Medini; Heywood, Sarah E.; Armitage, James A.; Kingwell, Bronwyn A.

    2016-01-01

    Background The regulation of microRNAs (miRNAs) at different stages of the progression of type 2 diabetes mellitus (T2DM) and their role in glucose homeostasis was investigated. Methods Microarrays were used to assess miRNA expression in skeletal muscle biopsies taken from healthy individuals and patients with pre-diabetes or T2DM, and insulin resistant offspring of rat dams fed a high fat diet during pregnancy. Results Twenty-three miRNAs were differentially expressed in patients with T2DM, and 7 in the insulin resistant rat offspring compared to their controls. Among these, only one miRNA was similarly regulated: miR-194 expression was significantly reduced by 25 to 50% in both the rat model and in human with pre-diabetes and established diabetes. Knockdown of miR-194 in L6 skeletal muscle cells induced an increase in basal and insulin-stimulated glucose uptake and glycogen synthesis. This occurred in conjunction with an increased glycolysis, indicated by elevated lactate production. Moreover, oxidative capacity was also increased as we found an enhanced glucose oxidation in presence of the mitochondrial uncoupler FCCP. When miR-194 was down-regulated in vitro, western blot analysis showed an increased phosphorylation of AKT and GSK3β in response to insulin, and an increase in expression of proteins controlling mitochondrial oxidative phosphorylation. Conclusions Type 2 diabetes mellitus is associated with regulation of several miRNAs in skeletal muscle. Interestingly, miR-194 was a unique miRNA that appeared regulated across different stages of the disease progression, from the early stages of insulin resistance to the development of T2DM. We have shown miR-194 is involved in multiple aspects of skeletal muscle glucose metabolism from uptake, through to glycolysis, glycogenesis and glucose oxidation, potentially via mechanisms involving AKT, GSK3 and oxidative phosphorylation. MiR-194 could be down-regulated in patients with early features of diabetes as an

  12. Strategies for Selection from Protein Libraries Composed of de Novo Designed Secondary Structure Modules

    NASA Astrophysics Data System (ADS)

    Matsuura, Tomoaki; Plückthun, Andreas

    2004-02-01

    As more and more protein structures are determined, it has become clear that there is only a limited number of protein folds in nature. To explore whether the protein folds found in nature are the only solutions to the protein folding problem, or that a lack of evolutionary pressure causes the paucity of different protein folds found, we set out to construct protein libraries without any restriction on topology. We generated different libraries (all α-helix, all β-strand and α-helix plus β-strand) with an average length of 100 amino acid residues, composed of designed secondary structure modules (α-helix, β-strand and β-turn) in various proportions, based primarily on the patterning of polar and non-polar residues. From the analysis of proteins chosen randomly from the libraries, we found that a substantial portion of pure α-helical proteins show properties similar to native proteins. Using these libraries as a starting point, we aim to establish a selection system which allows us to enrich proteins with favorable folding properties (non-aggregating, compactly folded) from the libraries. We have developed such a method based on ribosome display. This selection is based on two concepts: (1) misfolded proteins are more sensitive to proteolysis, (2) misfolded and/or aggregated proteins are more hydrophobic. We show that by applying each of these selection criteria proteins that are compactly folded and soluble can be enriched over insoluble and random coil proteins.

  13. Identification of lncRNA functions in lung cancer based on associated protein-protein interaction modules

    PubMed Central

    Wu, Chih-Hsun; Hsu, Chia-Lang; Lu, Pei-Chun; Lin, Wen-Chang; Juan, Hsueh-Fen; Huang, Hsuan-Cheng

    2016-01-01

    Long non-coding RNAs (lncRNAs) have been found to play important roles in various biological processes; however, many of their functions remain unclear. In this study, we present a novel approach to identify the lncRNA-associated protein-protein interaction (PPI) modules and ascertain their functions in human lung squamous cell carcinoma. We collected lncRNA and mRNA expression profiles of lung squamous cell carcinoma from The Cancer Genome Atlas. To identify the lncRNA-associated PPI modules, lncRNA-mRNA co-expression networks were first constructed based on the mutual ranks of expression correlations. Next, we examined whether the co-expressed mRNAs of a specific lncRNA were closely connected by PPIs. For this, a significantly connected mRNA set was considered to be the lncRNA-associated PPI module. Finally, the prospective functions of a lncRNA was inferred using Gene Ontology enrichment analysis on the associated module. We found that lncRNA-associated PPI modules were subtype-dependent and each subtype had unique molecular mechanisms. In addition, antisense lncRNAs and sense genes tended to be functionally associated. Our results might provide new directions for understanding lncRNA regulations in lung cancer. The analysis pipeline was implemented in a web tool, available at http://lncin.ym.edu.tw/. PMID:27786280

  14. Biochemical and metabolic mechanisms by which dietary whey protein may combat obesity and Type 2 diabetes.

    PubMed

    Jakubowicz, Daniela; Froy, Oren

    2013-01-01

    Consumption of milk and dairy products has been associated with reduced risk of metabolic disorders and cardiovascular disease. Milk contains two primary sources of protein, casein (80%) and whey (20%). Recently, the beneficial physiological effects of whey protein on the control of food intake and glucose metabolism have been reported. Studies have shown an insulinotropic and glucose-lowering properties of whey protein in healthy and Type 2 diabetes subjects. Whey protein seems to induce these effects via bioactive peptides and amino acids generated during its gastrointestinal digestion. These amino acids and peptides stimulate the release of several gut hormones, such as cholecystokinin, peptide YY and the incretins gastric inhibitory peptide and glucagon-like peptide 1 that potentiate insulin secretion from β-cells and are associated with regulation of food intake. The bioactive peptides generated from whey protein may also serve as endogenous inhibitors of dipeptidyl peptidase-4 (DPP-4) in the proximal gut, preventing incretin degradation. Indeed, recently, DPP-4 inhibitors were identified in whey protein hydrolysates. This review will focus on the emerging properties of whey protein and its potential clinical application for obesity and Type 2 diabetes.

  15. Innovative strategies to treat protein misfolding in inborn errors of metabolism: pharmacological chaperones and proteostasis regulators.

    PubMed

    Muntau, Ania C; Leandro, João; Staudigl, Michael; Mayer, Felix; Gersting, Søren W

    2014-07-01

    To attain functionality, proteins must fold into their three-dimensional native state. The intracellular balance between protein synthesis, folding, and degradation is constantly challenged by genetic or environmental stress factors. In the last ten years, protein misfolding induced by missense mutations was demonstrated to be the seminal molecular mechanism in a constantly growing number of inborn errors of metabolism. In these cases, loss of protein function results from early degradation of missense-induced misfolded proteins. Increasing knowledge on the proteostasis network and the protein quality control system with distinct mechanisms in different compartments of the cell paved the way for the development of new treatment strategies for conformational diseases using small molecules. These comprise proteostasis regulators that enhance the capacity of the proteostasis network and pharmacological chaperones that specifically bind and rescue misfolded proteins by conformational stabilization. They can be used either alone or in combination, the latter to exploit synergistic effects. Many of these small molecule compounds currently undergo preclinical and clinical pharmaceutical development and two have been approved: saproterin dihydrochloride for the treatment of phenylketonuria and tafamidis for the treatment of transthyretin-related hereditary amyloidosis. Different technologies are exploited for the discovery of new small molecule compounds that belong to the still young class of pharmaceutical products discussed here. These compounds may in the near future improve existing treatment strategies or even offer a first-time treatment to patients suffering from nowadays-untreatable inborn errors of metabolism. PMID:24687294

  16. Acrylamide administration alters protein phosphorylation and phospholipid metabolism in rat sciatic nerve

    SciTech Connect

    Berti-Mattera, L.N.; Eichberg, J.; Schrama, L.; LoPachin, R.M. )

    1990-05-01

    The effects of ACR on protein phosphorylation and phospholipid metabolism were assessed in rat sciatic nerve. After 5 days of ACR administration (50 mg/kg/day) an increase in the incorporation of 32P into phosphatidylinositol-4,5-bisphosphate, phosphatidylinositol-4-phosphate, and phosphatidylcholine was detected in proximal sciatic nerve segments. In contrast, no changes in phospholipid metabolism were observed in distal segments. After 9 days of ACR treatment when neurotoxicological symptoms were clearly apparent, a generalized increase in radiolabel uptake into phospholipids was noted exclusively in proximal nerve regions. ACR-induced increases in phospholipid metabolism were toxicologically specific since comparable administration of MBA (108 mg/kg/day X 5 or 9 days) produced only minor changes. ACR intoxication was also associated with a rise in sciatic nerve protein phosphorylation. After 9 days of ACR treatment, phosphorylation of beta-tubulin, P0, and several unidentified proteins (38 and 180 kDa) was increased in distal segments. In contrast, chronic administration of MBA caused increases in phosphorylation of beta-tubulin and the major myelin proteins of proximal nerve segments. In cell free homogenates prepared from sciatic nerves of treated and control rats, MBA caused an increase in phosphorylation of major myelin proteins similar to its effect in intact proximal nerve segments. The most striking effect observed in nerve homogenates of ACR-treated rats was a marked decrease in phosphorylation of an 80-kDa protein. Addition of ACR (1 mM) to homogenates of normal nerve had no effect on protein phosphorylation. Our results indicate that changes in the phosphorylation of phospholipids and proteins in sciatic nerve might be a component of the neurotoxic mechanism of ACR.

  17. Role of the AMP-activated protein kinase in regulating fatty acid metabolism during exercise.

    PubMed

    Steinberg, Gregory R

    2009-06-01

    During moderate-intensity exercise, fatty acids are the predominant substrate for working skeletal muscle. The release of fatty acids from adipose tissue stores, combined with the ability of skeletal muscle to actively fine tune the gradient between fatty acid and carbohydrate metabolism, depending on substrate availability and energetic demands, requires a coordinated system of metabolic control. Over the past decade, since the discovery that AMP-activated protein kinase (AMPK) was increased in accordance with exercise intensity, there has been significant interest in the proposed role of this ancient stress-sensing kinase as a critical integrative switch controlling metabolic responses during exercise. In this review, studies examining the role of AMPK as a regulator of fatty acid metabolism in both adipose tissue and skeletal muscle during exercise will be discussed. Exercise induces activation of AMPK in adipocytes and regulates triglyceride hydrolysis and esterfication through phosphorylation of hormone sensitive lipase (HSL) and glycerol-3-phosphate acyl-transferase, respectively. In skeletal muscle, exercise-induced activation of AMPK is associated with increases in fatty acid uptake, phosphorylation of HSL, and increased fatty acid oxidation, which is thought to occur via the acetyl-CoA carboxylase-malony-CoA-CPT-1 signalling axis. Despite the importance of AMPK in regulating fatty acid metabolism under resting conditions, recent evidence from transgenic models of AMPK deficiency suggest that alternative signalling pathways may also be important for the control of fatty acid metabolism during exercise.

  18. Nicotinamide N-methyltransferase regulates hepatic nutrient metabolism through Sirt1 protein stabilization

    PubMed Central

    Hong, Shangyu; Moreno-Navarrete, Jose M; Wei, Xiaojing; Kikukawa, Yusuke; Tzameli, Iphigenia; Prasad, Deepthi; Lee, Yoonjin; Asara, John M; Fernandez-Real, Jose Manuel; Maratos-Flier, Eleftheria; Pissios, Pavlos

    2015-01-01

    Nicotinamide N-methyltransferase (Nnmt) methylates nicotinamide, a form of vitamin B3, to produce N1-methylnicotinamide (MNAM). Nnmt is an emerging metabolic regulator in adipocytes but its role in the liver, a tissue with the strongest Nnmt expression, is not known. In spite of its overall high expression, here we find that hepatic expression of Nnmt is highly variable and correlates with multiple metabolic parameters in mice and in humans. Further, we find that suppression of hepatic Nnmt expression in vivo alters glucose and cholesterol metabolism and that the metabolic effects of Nnmt in the liver are mediated by its product MNAM. Supplementation of high fat diet with MNAM decreases serum and liver cholesterol and liver triglycerides levels in mice. Mechanistically, increasing Nnmt expression or MNAM levels stabilizes sirtuin 1 protein, an effect, which is required for their metabolic benefits. In summary, we describe a novel regulatory pathway for vitamin B3 that could provide a new opportunity for metabolic disease therapy. PMID:26168293

  19. The unfolded protein response mediates reversible tau phosphorylation induced by metabolic stress

    PubMed Central

    van der Harg, J M; Nölle, A; Zwart, R; Boerema, A S; van Haastert, E S; Strijkstra, A M; Hoozemans, J JM; Scheper, W

    2014-01-01

    The unfolded protein response (UPR) is activated in neurodegenerative tauopathies such as Alzheimer's disease (AD) in close connection with early stages of tau pathology. Metabolic disturbances are strongly associated with increased risk for AD and are a potent inducer of the UPR. Here, we demonstrate that metabolic stress induces the phosphorylation of endogenous tau via activation of the UPR. Strikingly, upon restoration of the metabolic homeostasis, not only the levels of the UPR markers pPERK, pIRE1α and BiP, but also tau phosphorylation are reversed both in cell models as well as in torpor, a physiological hypometabolic model in vivo. Intervention in the UPR using the global UPR inhibitor TUDCA or a specific small-molecule inhibitor of the PERK signaling pathway, inhibits the metabolic stress-induced phosphorylation of tau. These data support a role for UPR-mediated tau phosphorylation as part of an adaptive response to metabolic stress. Failure to restore the metabolic homeostasis will lead to prolonged UPR activation and tau phosphorylation, and may thus contribute to AD pathogenesis. We demonstrate that the UPR is functionally involved in the early stages of tau pathology. Our data indicate that targeting of the UPR may be employed for early intervention in tau-related neurodegenerative diseases. PMID:25165879

  20. A DEK Domain-Containing Protein Modulates Chromatin Structure and Function in Arabidopsis[W][OPEN

    PubMed Central

    Waidmann, Sascha; Kusenda, Branislav; Mayerhofer, Juliane; Mechtler, Karl; Jonak, Claudia

    2014-01-01

    Chromatin is a major determinant in the regulation of virtually all DNA-dependent processes. Chromatin architectural proteins interact with nucleosomes to modulate chromatin accessibility and higher-order chromatin structure. The evolutionarily conserved DEK domain-containing protein is implicated in important chromatin-related processes in animals, but little is known about its DNA targets and protein interaction partners. In plants, the role of DEK has remained elusive. In this work, we identified DEK3 as a chromatin-associated protein in Arabidopsis thaliana. DEK3 specifically binds histones H3 and H4. Purification of other proteins associated with nuclear DEK3 also established DNA topoisomerase 1α and proteins of the cohesion complex as in vivo interaction partners. Genome-wide mapping of DEK3 binding sites by chromatin immunoprecipitation followed by deep sequencing revealed enrichment of DEK3 at protein-coding genes throughout the genome. Using DEK3 knockout and overexpressor lines, we show that DEK3 affects nucleosome occupancy and chromatin accessibility and modulates the expression of DEK3 target genes. Furthermore, functional levels of DEK3 are crucial for stress tolerance. Overall, data indicate that DEK3 contributes to modulation of Arabidopsis chromatin structure and function. PMID:25387881

  1. Macroautophagy and Cell Responses Related to Mitochondrial Dysfunction, Lipid Metabolism and Unconventional Secretion of Proteins

    PubMed Central

    Demine, Stéphane; Michel, Sébastien; Vannuvel, Kayleen; Wanet, Anaïs; Renard, Patricia; Arnould, Thierry

    2012-01-01

    Macroautophagy has important physiological roles and its cytoprotective or detrimental function is compromised in various diseases such as many cancers and metabolic diseases. However, the importance of autophagy for cell responses has also been demonstrated in many other physiological and pathological situations. In this review, we discuss some of the recently discovered mechanisms involved in specific and unspecific autophagy related to mitochondrial dysfunction and organelle degradation, lipid metabolism and lipophagy as well as recent findings and evidence that link autophagy to unconventional protein secretion. PMID:24710422

  2. Protein sorting gone wrong--VPS10P domain receptors in cardiovascular and metabolic diseases.

    PubMed

    Schmidt, Vanessa; Willnow, Thomas E

    2016-02-01

    VPS10P domain receptors are a unique class of sorting receptors that direct intracellular transport of target proteins in neurons and that play central roles in neurodegenerative processes. Surprisingly, genome-wide association studies now implicate the very same receptors in cardiovascular and metabolic disturbances. In this review, we discuss current findings that uncovered some of the molecular mechanisms whereby sorting receptors, such as SORLA, sortilin, and SORCS1 control homeostasis in cardiovascular and metabolic tissues, and how they promote hypercholesterolemia, atherosclerosis, obesity, and diabetes, when being altered. PMID:26724530

  3. Positive Lysosomal Modulation As a Unique Strategy to Treat Age-Related Protein Accumulation Diseases

    PubMed Central

    Wisniewski, Meagan L.; Butler, David

    2012-01-01

    Abstract Lysosomes are involved in degrading and recycling cellular ingredients, and their disruption with age may contribute to amyloidogenesis, paired helical filaments (PHFs), and α-synuclein and mutant huntingtin aggregation. Lysosomal cathepsins are upregulated by accumulating proteins and more so by the modulator Z-Phe-Ala-diazomethylketone (PADK). Such positive modulators of the lysosomal system have been studied in the well-characterized hippocampal slice model of protein accumulation that exhibits the pathogenic cascade of tau aggregation, tubulin breakdown, microtubule destabilization, transport failure, and synaptic decline. Active cathepsins were upregulated by PADK; Rab proteins were modified as well, indicating enhanced trafficking, whereas lysosome-associated membrane protein and proteasome markers were unchanged. Lysosomal modulation reduced the pre-existing PHF deposits, restored tubulin structure and transport, and recovered synaptic components. Further proof-of-principle studies used Alzheimer disease mouse models. It was recently reported that systemic PADK administration caused dramatic increases in cathepsin B protein and activity levels, whereas neprilysin, insulin-degrading enzyme, α-secretase, and β-secretase were unaffected by PADK. In the transgenic models, PADK treatment resulted in clearance of intracellular amyloid beta (Aβ) peptide and concomitant reduction of extracellular deposits. Production of the less pathogenic Aβ1–38 peptide corresponded with decreased levels of Aβ1–42, supporting the lysosome's antiamyloidogenic role through intracellular truncation. Amelioration of synaptic and behavioral deficits also indicates a neuroprotective function of the lysosomal system, identifying lysosomal modulation as an avenue for disease-modifying therapies. From the in vitro and in vivo findings, unique lysosomal modulators represent a minimally invasive, pharmacologically controlled strategy against protein accumulation disorders

  4. Expression of metabolism-related proteins in triple-negative breast cancer

    PubMed Central

    Kim, Min-Ju; Kim, Do-Hee; Jung, Woo-Hee; Koo, Ja-Seung

    2014-01-01

    To investigate the dominant metabolic type of triple-negative breast cancer (TNBC) and evaluate its clinical implication through analysis of protein expression related to glycolysis, glutaminolysis, and mitochondrial oxidative phosphorylation. Tissue samples from 129 patients with TNBC who underwent mastectomy due to invasive breast cancer from 2000 to 2005 were prepared for tissue microarray. By immunohistochemical staining of the tissue microarrays, the markers of glycolysis-related proteins (Glut-1, CAIX, MCT4), glutaminolysis-related proteins (GLS1, GDH, ASCT2), and mitochondrial enzymes (ATP synthase, SDHA and SDHB) were analyzed. Based on the results, the metabolic phenotypes were defined based on positivity for more than two of three markers for each phenotype as follows: glycolysis type (Glut-1, CAIX and MCT4), glutaminolysis type (GLS1, GDH and ASCT2) and mitochondrial type (ATP synthase, SDHA and SDHB). The percentages of samples with metabolic phenotypes of tumor and stroma of TNBC were as follows: for tumor, mitochondrial type (85.3%) > glutaminolysis type (67.4%) > glycolysis type (63.0%); and for stroma, glutaminolysis type (37.2%) > glycolysis type (16.3%) > mitochondrial type (14.0%). The most common metabolic phenotype of TNBC was glycolysis type for basal-like type and non-glycolysis type for non-basal-like type (p=0.047). The correlation between glutaminolysis and mitochondrial type was statistically significant in both tumor and stroma (p<0.001). In conclusion, tumor cells of TNBC express glycolysis and mitochondrial metabolism-related proteins. Glycolysis type is the most common phenotype of basal-like type, and reversely, non-glycolysis type is the most common phenotype of non basal-like type. PMID:24427351

  5. NFκB is a central regulator of protein quality control in response to protein aggregation stresses via autophagy modulation

    PubMed Central

    Nivon, Mathieu; Fort, Loïc; Muller, Pascale; Richet, Emma; Simon, Stéphanie; Guey, Baptiste; Fournier, Maëlenn; Arrigo, André-Patrick; Hetz, Claudio; Atkin, Julie D.; Kretz-Remy, Carole

    2016-01-01

    During cell life, proteins often misfold, depending on particular mutations or environmental changes, which may lead to protein aggregates that are toxic for the cell. Such protein aggregates are the root cause of numerous diseases called “protein conformational diseases,” such as myofibrillar myopathy and familial amyotrophic lateral sclerosis. To fight against aggregates, cells are equipped with protein quality control mechanisms. Here we report that NFκB transcription factor is activated by misincorporation of amino acid analogues into proteins, inhibition of proteasomal activity, expression of the R120G mutated form of HspB5 (associated with myofibrillar myopathy), or expression of the G985R and G93A mutated forms of superoxide dismutase 1 (linked to familial amyotrophic lateral sclerosis). This noncanonical stimulation of NFκB triggers the up-regulation of BAG3 and HspB8 expression, two activators of selective autophagy, which relocalize to protein aggregates. Then NFκB-dependent autophagy allows the clearance of protein aggregates. Thus NFκB appears as a central and major regulator of protein aggregate clearance by modulating autophagic activity. In this context, the pharmacological stimulation of this quality control pathway might represent a valuable strategy for therapies against protein conformational diseases. PMID:27075172

  6. The relation of high fat diet, metabolic disturbances and brain oxidative dysfunction: modulation by hydroxy citric acid

    PubMed Central

    2011-01-01

    Aims This study aimed to examine the effect of high fat diet (HFD) to modulate brain dysfunction, and understand the linkages between obesity, metabolic disturbances and the brain oxidative stress (BOS) dysfunction and modulation with hydroxyl citric acid of G. Cambogia. Methods Rats were divided into 3 groups; 1st control, maintained on standard normal rat chow diet, 2nd HFD, maintained on high fat diet along 12 week and 3rd HFD+G, administered G. Cambogia for 4 weeks and each group include 8 rats. Blood, brain and abdominal fat were collected for biochemical measurements. Results HFD group showed significant increase in energy intake, final BW and BW gain. Also significant increase in weight of abdominal fat in HFD group. HFD induce metabolic disturbance through increasing the lipid profile (LDL, TG, TC), γGT and α-amylase activity, uric acid level and hyperglycemia, while decreasing creatine kinase (CK) activity. These changes associated with lowering in brain nitric oxide (NO) level and rising in serum butyrylcholinesterase (BChE), brain catalase activity and MDA levels as oxidative stress markers. These alterations improved by G. Cambogia that decrease BOS and increased NO level. Conclusions Rats fed HFD showed, metabolic disturbances produce hyperglycemia, hypertriglyceridemia, hypercholesterolemia and increased LDL associated with increased BOS. Involvement of BuChE, NO and oxidative stress associated with metabolic disturbances in the pathophysiological progression in brain, suggesting association between obesity, metabolic disorders and brain alteration while, using G. Cambogia, ameliorate the damaging effects of the HFD via lowering feed intake and BOS. PMID:21569551

  7. Modulating protein activity using tethered ligands with mutually exclusive binding sites

    PubMed Central

    Schena, Alberto; Griss, Rudolf; Johnsson, Kai

    2015-01-01

    The possibility to design proteins whose activities can be switched on and off by unrelated effector molecules would enable applications in various research areas, ranging from biosensing to synthetic biology. We describe here a general method to modulate the activity of a protein in response to the concentration of a specific effector. The approach is based on synthetic ligands that possess two mutually exclusive binding sites, one for the protein of interest and one for the effector. Tethering such a ligand to the protein of interest results in an intramolecular ligand–protein interaction that can be disrupted through the presence of the effector. Specifically, we introduce a luciferase controlled by another protein, a human carbonic anhydrase whose activity can be controlled by proteins or small molecules in vitro and on living cells, and novel fluorescent and bioluminescent biosensors. PMID:26198003

  8. Cellular prion protein and NMDA receptor modulation: protecting against excitotoxicity

    PubMed Central

    Black, Stefanie A. G.; Stys, Peter K.; Zamponi, Gerald W.; Tsutsui, Shigeki

    2014-01-01

    Although it is well established that misfolding of the cellular prion protein (PrPC) into the β-sheet-rich, aggregated scrapie conformation (PrPSc) causes a variety of transmissible spongiform encephalopathies (TSEs), the physiological roles of PrPC are still incompletely understood. There is accumulating evidence describing the roles of PrPC in neurodegeneration and neuroinflammation. Recently, we identified a functional regulation of NMDA receptors by PrPC that involves formation of a physical protein complex between these proteins. Excessive NMDA receptor activity during conditions such as ischemia mediates enhanced Ca2+ entry into cells and contributes to excitotoxic neuronal death. In addition, NMDA receptors and/or PrPC play critical roles in neuroinflammation and glial cell toxicity. Inhibition of NMDA receptor activity protects against PrPSc-induced neuronal death. Moreover, in mice lacking PrPC, infarct size is increased after focal cerebral ischemia, and absence of PrPC increases susceptibility of neurons to NMDA receptor-dependent death. Recently, PrPC was found to be a receptor for oligomeric beta-amyloid (Aβ) peptides, suggesting a role for PrPC in Alzheimer's disease (AD). Our recent findings suggest that Aβ peptides enhance NMDA receptor current by perturbing the normal copper- and PrPC-dependent regulation of these receptors. Here, we review evidence highlighting a role for PrPC in preventing NMDA receptor-mediated excitotoxicity and inflammation. There is a need for more detailed molecular characterization of PrPC-mediated regulation of NMDA receptors, such as determining which NMDA receptor subunits mediate pathogenic effects upon loss of PrPC-mediated regulation and identifying PrPC binding site(s) on the receptor. This knowledge will allow development of novel therapeutic interventions for not only TSEs, but also for AD and other neurodegenerative disorders involving dysfunction of PrPC. PMID:25364752

  9. Putative drug and vaccine target protein identification using comparative genomic analysis of KEGG annotated metabolic pathways of Mycoplasma hyopneumoniae.

    PubMed

    Damte, Dereje; Suh, Joo-Won; Lee, Seung-Jin; Yohannes, Sileshi Belew; Hossain, Md Akil; Park, Seung-Chun

    2013-07-01

    In the present study, a computational comparative and subtractive genomic/proteomic analysis aimed at the identification of putative therapeutic target and vaccine candidate proteins from Kyoto Encyclopedia of Genes and Genomes (KEGG) annotated metabolic pathways of Mycoplasma hyopneumoniae was performed for drug design and vaccine production pipelines against M.hyopneumoniae. The employed comparative genomic and metabolic pathway analysis with a predefined computational systemic workflow extracted a total of 41 annotated metabolic pathways from KEGG among which five were unique to M. hyopneumoniae. A total of 234 proteins were identified to be involved in these metabolic pathways. Although 125 non homologous and predicted essential proteins were found from the total that could serve as potential drug targets and vaccine candidates, additional prioritizing parameters characterize 21 proteins as vaccine candidate while druggability of each of the identified proteins evaluated by the DrugBank database prioritized 42 proteins suitable for drug targets.

  10. Cellular factors modulating the mechanism of tau protein aggregation

    PubMed Central

    Fontaine, Sarah N.; Sabbagh, Jonathan J.; Baker, Jeremy; Martinez-Licha, Carlos R.; Darling, April

    2015-01-01

    Pathological accumulation of the microtubule-associated protein tau, in the form of neurofibrillary tangles, is a major hallmark of Alzheimer’s disease, the most prevalent neurodegenerative condition worldwide. In addition to Alzheimer’s disease, a number of neurodegenerative diseases, called tauopathies, are characterized by the accumulation of aggregated tau in a variety of brain regions. While tau normally plays an important role in stabilizing the microtubule network of the cytoskeleton, its dissociation from microtubules and eventual aggregation into pathological deposits is an area of intense focus for therapeutic development. Here we discuss the known cellular factors that affect tau aggregation, from post-translational modifications to molecular chaperones. PMID:25666877

  11. Effect of branched chain amino acid infusions on body protein metabolism in cirrhosis of liver.

    PubMed Central

    Wright, P D; Holdsworth, J D; Dionigi, P; Clague, M B; James, O F

    1986-01-01

    Thirty seven patients with established cirrhosis of the liver were subjected to measurement of body protein metabolism using L-(1-14C) labelled leucine as a tracer. The effects of disease severity and those of solutions containing 0%, 16%, 35%, 53%, and 100% branched chain amino acids were evaluated. Significant increases in protein synthesis were noted with solutions containing 35%, 53%, and 100% branched chain amino acids, but in patients receiving 100% branched chain amino acids without additional essential amino acid supplement the increase in synthesis was matched by a significant increase in protein breakdown. Protein balance was thus improved only in patients receiving 35% and 53% branched chain amino acids. It was concluded that the high increase in protein breakdown in patients receiving 100% branched chain amino acids was undesirable, and such a solution should not be recommended for clinical use. PMID:3539714

  12. The essential polysome-associated RNA-binding protein RBP42 targets mRNAs involved in Trypanosoma brucei energy metabolism.

    PubMed

    Das, Anish; Morales, Rachel; Banday, Mahrukh; Garcia, Stacey; Hao, Li; Cross, George A M; Estevez, Antonio M; Bellofatto, Vivian

    2012-11-01

    RNA-binding proteins that target mRNA coding regions are emerging as regulators of post-transcriptional processes in eukaryotes. Here we describe a newly identified RNA-binding protein, RBP42, which targets the coding region of mRNAs in the insect form of the African trypanosome, Trypanosoma brucei. RBP42 is an essential protein and associates with polysome-bound mRNAs in the cytoplasm. A global survey of RBP42-bound mRNAs was performed by applying HITS-CLIP technology, which captures protein-RNA interactions in vivo using UV light. Specific RBP42-mRNA interactions, as well as mRNA interactions with a known RNA-binding protein, were purified using specific antibodies. Target RNA sequences were identified and quantified using high-throughput RNA sequencing. Analysis revealed that RBP42 bound mainly within the coding region of mRNAs that encode proteins involved in cellular energy metabolism. Although the mechanism of RBP42's function is unclear at present, we speculate that RBP42 plays a critical role in modulating T. brucei energy metabolism.

  13. Protein metabolism in growing pigs fed corn or cassava peel based diets containing graded protein levels.

    PubMed

    Tewe, O O

    1985-05-01

    Sixty-four Large White cross Landrace weanling pigs were randomly allotted to eight treatments in a two by four factorial arrangement. The two dietary variables were cassava peel (0 and 40 per cent) and crude protein (20, 15, 10 and 5 per cent). Total serum protein concentration was significantly (P less than 0.01) reduced by protein deficiency and by its interaction with cassava peel. The multiple coefficient of determination (R2) showed that protein intake was the primary factor determining changes in serum protein. R2 values for cyanide intake (independent variable) on serum protein (dependent variable) increased from day 30 to 90 of the trial. Serum urea was increased on the 5 per cent protein diets on days 60 and 90 of the trial. The R2 values for cyanide and protein intake on serum urea concentration increased from day 30 to day 90 of the trial. Serum creatinine increased (P less than 0.05) on the 5 per cent protein diet on day 90 of the trial. The R2 value for the effects of protein intake on serum creatinine was higher than for cyanide intake on days 30 and 90. The results confirm the progressive and pronounced effects of long term cyanide intake on serum nitrogenous metabolites in pigs consuming between 110 and 120 ppm hydrocyanic acid, especially in diets containing 10 per cent or less protein. PMID:2989987