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Sample records for basic leucine zipper

  1. Thermodynamic analysis of the heterodimerization of leucine zippers of Jun and Fos transcription factors

    International Nuclear Information System (INIS)

    Seldeen, Kenneth L.; McDonald, Caleb B.; Deegan, Brian J.; Farooq, Amjad

    2008-01-01

    Jun and Fos are components of the AP1 family of transcription factors and bind to the promoters of a diverse multitude of genes involved in critical cellular responses such as cell growth and proliferation, cell cycle regulation, embryonic development and cancer. Here, using the powerful technique of isothermal titration calorimetry, we characterize the thermodynamics of heterodimerization of leucine zippers of Jun and Fos. Our data suggest that the heterodimerization of leucine zippers is driven by enthalpic forces with unfavorable entropy change at physiological temperatures. Furthermore, the basic regions appear to modulate the heterodimerization of leucine zippers and may undergo at least partial folding upon heterodimerization. Large negative heat capacity changes accompanying the heterodimerization of leucine zippers are consistent with the view that leucine zippers do not retain α-helical conformations in isolation and that the formation of the native coiled-coil α-helical dimer is attained through a coupled folding-dimerization mechanism

  2. 3D Printing of Protein Models in an Undergraduate Laboratory: Leucine Zippers

    Science.gov (United States)

    Meyer, Scott C.

    2015-01-01

    An upper-division undergraduate laboratory experiment is described that explores the structure/function relationship of protein domains, namely leucine zippers, through a molecular graphics computer program and physical models fabricated by 3D printing. By generating solvent accessible surfaces and color-coding hydrophobic, basic, and acidic amino…

  3. Evolutionary and Expression Analyses of the Apple Basic Leucine Zipper Transcription Factor Family

    Science.gov (United States)

    Zhao, Jiao; Guo, Rongrong; Guo, Chunlei; Hou, Hongmin; Wang, Xiping; Gao, Hua

    2016-01-01

    Transcription factors (TFs) play essential roles in the regulatory networks controlling many developmental processes in plants. Members of the basic leucine (Leu) zipper (bZIP) TF family, which is unique to eukaryotes, are involved in regulating diverse processes, including flower and vascular development, seed maturation, stress signaling, and defense responses to pathogens. The bZIP proteins have a characteristic bZIP domain composed of a DNA-binding basic region and a Leu zipper dimerization region. In this study, we identified 112 apple (Malus domestica Borkh) bZIP TF-encoding genes, termed MdbZIP genes. Synteny analysis indicated that segmental and tandem duplication events, as well as whole genome duplication, have contributed to the expansion of the apple bZIP family. The family could be divided into 11 groups based on structural features of the encoded proteins, as well as on the phylogenetic relationship of the apple bZIP proteins to those of the model plant Arabidopsis thaliana (AtbZIP genes). Synteny analysis revealed that several paired MdbZIP genes and AtbZIP gene homologs were located in syntenic genomic regions. Furthermore, expression analyses of group A MdbZIP genes showed distinct expression levels in 10 different organs. Moreover, changes in these expression profiles in response to abiotic stress conditions and various hormone treatments identified MdbZIP genes that were responsive to high salinity and drought, as well as to different phytohormones. PMID:27066030

  4. Evolutionary and Expression Analyses of the Apple Basic Leucine Zipper Transcription Factor Family

    Directory of Open Access Journals (Sweden)

    Jiao eZhao

    2016-03-01

    Full Text Available Transcription factors (TFs play essential roles in the regulatory networks controlling many developmental processes in plants. Members of the basic leucine (Leu zipper (bZIP TF family, which is unique to eukaryotes, are involved in regulating diverse processes, including flower and vascular development, seed maturation, stress signaling and defense responses to pathogens. The bZIP proteins have a characteristic bZIP domain composed of a DNA-binding basic region and a Leu zipper dimerization region. In this study, we identified 112 apple (Malus domestica Borkh bZIP TF-encoding genes, termed MdbZIP genes. Synteny analysis indicated that segmental and tandem duplication events, as well as whole genome duplication, have contributed to the expansion of the apple bZIP family. The family could be divided into 11 groups based on structural features of the encoded proteins, as well as on the phylogenetic relationship of the apple bZIP proteins to those of the model plant Arabidopsis thaliana (AtbZIP genes. Synteny analysis revealed that several paired MdbZIP genes and AtbZIP gene homologs were located in syntenic genomic regions. Furthermore, expression analyses of group A MdbZIP genes showed distinct expression levels in ten different organs. Moreover, changes in these expression profiles in response to abiotic stress conditions and various hormone treatments identified MdbZIP genes that were responsive to high salinity and drought, as well as to different phytohormones.

  5. The retro-GCN4 leucine zipper sequence forms a stable three-dimensional structure

    Science.gov (United States)

    Mittl, Peer R. E.; Deillon, Christine; Sargent, David; Liu, Niankun; Klauser, Stephan; Thomas, Richard M.; Gutte, Bernd; Grütter, Markus G.

    2000-01-01

    The question of whether a protein whose natural sequence is inverted adopts a stable fold is still under debate. We have determined the 2.1-Å crystal structure of the retro-GCN4 leucine zipper. In contrast to the two-stranded helical coiled-coil GCN4 leucine zipper, the retro-leucine zipper formed a very stable, parallel four-helix bundle, which now lends itself to further structural and functional studies. PMID:10716989

  6. An engineered allosteric switch in leucine-zipper oligomerization.

    Science.gov (United States)

    Gonzalez, L; Plecs, J J; Alber, T

    1996-06-01

    Controversy remains about the role of core side-chain packing in specifying protein structure. To investigate the influence of core packing on the oligomeric structure of a coiled coil, we engineered a GCN4 leucine zipper mutant that switches from two to three strands upon binding the hydrophobic ligands cyclohexane and benzene. In solution these ligands increased the apparent thermal stability and the oligomerization order of the mutant leucine zipper. The crystal structure of the peptide-benzene complex shows a single benzene molecule bound at the engineered site in the core of the trimer. These results indicate that coiled coils are well-suited to function as molecular switches and emphasize that core packing is an important determinant of oligomerization specificity.

  7. The leucine zipper domains of the transcription factors GCN4 and c-Jun have ribonuclease activity.

    Directory of Open Access Journals (Sweden)

    Yaroslav Nikolaev

    Full Text Available Basic-region leucine zipper (bZIP proteins are one of the largest transcription factor families that regulate a wide range of cellular functions. Owing to the stability of their coiled coil structure leucine zipper (LZ domains of bZIP factors are widely employed as dimerization motifs in protein engineering studies. In the course of one such study, the X-ray structure of the retro-version of the LZ moiety of yeast transcriptional activator GCN4 suggested that this retro-LZ may have ribonuclease activity. Here we show that not only the retro-LZ but also the authentic LZ of GCN4 has weak but distinct ribonuclease activity. The observed cleavage of RNA is unspecific, it is not suppressed by the ribonuclease A inhibitor RNasin and involves the breakage of 3',5'-phosphodiester bonds with formation of 2',3'-cyclic phosphates as the final products as demonstrated by HPLC/electrospray ionization mass spectrometry. Several mutants of the GCN4 leucine zipper are catalytically inactive, providing important negative controls and unequivocally associating the enzymatic activity with the peptide under study. The leucine zipper moiety of the human factor c-Jun as well as the entire c-Jun protein are also shown to catalyze degradation of RNA. The presented data, which was obtained in the test-tube experiments, adds GCN4 and c-Jun to the pool of proteins with multiple functions (also known as moonlighting proteins. If expressed in vivo, the endoribonuclease activity of these bZIP-containing factors may represent a direct coupling between transcription activation and controlled RNA turnover. As an additional result of this work, the retro-leucine zipper of GCN4 can be added to the list of functional retro-peptides.

  8. Arabidopsis basic leucine zipper transcription factors involved in an abscisic acid-dependent signal transduction pathway under drought and high-salinity conditions

    OpenAIRE

    Uno, Yuichi; Furihata, Takashi; Abe, Hiroshi; Yoshida, Riichiro; Shinozaki, Kazuo; Yamaguchi-Shinozaki, Kazuko

    2000-01-01

    The induction of the dehydration-responsive Arabidopsis gene, rd29B, is mediated mainly by abscisic acid (ABA). Promoter analysis of rd29B indicated that two ABA-responsive elements (ABREs) are required for the dehydration-responsive expression of rd29B as cis-acting elements. Three cDNAs encoding basic leucine zipper (bZIP)-type ABRE-binding proteins were isolated by using the yeast one-hybrid system and were designated AREB1, AREB2, and AREB3 (ABA-responsive ...

  9. SdAb heterodimer formation using leucine zippers

    Science.gov (United States)

    Goldman, Ellen R.; Anderson, George P.; Brozozog-Lee, P. Audrey; Zabetakis, Dan

    2013-05-01

    Single domain antibodies (sdAb) are variable domains cloned from camel, llama, or shark heavy chain only antibodies, and are among the smallest known naturally derived antigen binding fragments. SdAb derived from immunized llamas are able to bind antigens with high affinity, and most are capable of refolding after heat or chemical denaturation to bind antigen again. We hypothesized that the ability to produce heterodimeric sdAb would enable reagents with the robust characteristics of component sdAb, but with dramatically improved overall affinity through increased avidity. Previously we had constructed multimeric sdAb by genetically linking sdAb that bind non-overlapping epitopes on the toxin, ricin. In this work we explored a more flexible approach; the construction of multivalent binding reagents using multimerization domains. We expressed anti-ricin sdAb that recognize different epitopes on the toxin as fusions with differently charged leucine zippers. When the initially produced homodimers are mixed the leucine zipper domains will pair to produce heterodimers. We used fluorescence resonance energy transfer to confirm heterodimer formation. Surface plasmon resonance, circular dichroism, enzyme linked immunosorbent assays, and fluid array assays were used to characterize the multimer constructs, and evaluate their utility in toxin detection.

  10. 'Zipbody' leucine zipper-fused Fab in E. coli in vitro and in vivo expression systems.

    Science.gov (United States)

    Ojima-Kato, Teruyo; Fukui, Kansuke; Yamamoto, Hiroaki; Hashimura, Dai; Miyake, Shiro; Hirakawa, Yuki; Yamasaki, Tomomi; Kojima, Takaaki; Nakano, Hideo

    2016-04-01

    A small antibody fragment, fragment of antigen binding (Fab), is favorable for various immunological assays. However, production efficiency of active Fab in microorganisms depends considerably on the clones. In this study, leucine zipper-peptide pairs that dimerize in parallel (ACID-p1 (LZA)/BASE-p1 (LZB) or c-Jun/c-Fos) were fused to the C-terminus of heavy chain (Hc, VH-CH1) and light chain (Lc, VL-CL), respectively, to accelerate the association of Hc and Lc to form Fab in Escherichia coli in vivo and in vitro expression systems. The leucine zipper-fused Fab named 'Zipbody' was constructed using anti-E. coli O157 monoclonal antibody obtained from mouse hybridoma and produced in both in vitro and in vivo expression systems in an active form, whereas Fab without the leucine zipper fusion was not. Similarly, Zipbody of rabbit monoclonal antibody produced in in vitro expression showed significant activity. The purified, mouse Zipbody produced in the E. coli strain Shuffle T7 Express had specificity toward the antigen; in bio-layer interferometry analysis, the KD value was measured to be 1.5-2.0 × 10(-8) M. These results indicate that leucine zipper fusion to Fab C-termini markedly enhances active Fab formation in E. coli. © The Author 2016. Published by Oxford University Press. All rights reserved. For Permissions, please e-mail: journals.permissions@oup.com.

  11. The N-terminal leucine-zipper motif in PTRF/cavin-1 is essential and sufficient for its caveolae-association

    Energy Technology Data Exchange (ETDEWEB)

    Wei, Zhuang [State Key Laboratory of Cell Biology, Institute of Biochemistry and Cell Biology, Shanghai Institutes for Biological Sciences, Chinese Academy of Sciences, Shanghai 200031 (China); Laboratory of System Biology, Institute of Biochemistry and Cell Biology, Shanghai Institutes for Biological Sciences, Chinese Academy of Sciences, Shanghai 200031 (China); Zou, Xinle [State Key Laboratory of Cell Biology, Institute of Biochemistry and Cell Biology, Shanghai Institutes for Biological Sciences, Chinese Academy of Sciences, Shanghai 200031 (China); Wang, Hongzhong; Lei, Jigang; Wu, Yuan [State Key Laboratory of Cell Biology, Institute of Biochemistry and Cell Biology, Shanghai Institutes for Biological Sciences, Chinese Academy of Sciences, Shanghai 200031 (China); Laboratory of System Biology, Institute of Biochemistry and Cell Biology, Shanghai Institutes for Biological Sciences, Chinese Academy of Sciences, Shanghai 200031 (China); Liao, Kan, E-mail: kliao@sibs.ac.cn [State Key Laboratory of Cell Biology, Institute of Biochemistry and Cell Biology, Shanghai Institutes for Biological Sciences, Chinese Academy of Sciences, Shanghai 200031 (China); Laboratory of System Biology, Institute of Biochemistry and Cell Biology, Shanghai Institutes for Biological Sciences, Chinese Academy of Sciences, Shanghai 200031 (China)

    2015-01-16

    Highlight: • The N-terminal leucine-zipper motif in PTRF/cavin-1 determines caveolar association. • Different cellular localization of PTRF/cavin-1 influences its serine 389 and 391 phosphorylation state. • PTRF/cavin-1 regulates cell motility via its caveolar association. - Abstract: PTRF/cavin-1 is a protein of two lives. Its reported functions in ribosomal RNA synthesis and in caveolae formation happen in two different cellular locations: nucleus vs. plasma membrane. Here, we identified that the N-terminal leucine-zipper motif in PTRF/cavin-1 was essential for the protein to be associated with caveolae in plasma membrane. It could counteract the effect of nuclear localization sequence in the molecule (AA 235–251). Deletion of this leucine-zipper motif from PTRF/cavin-1 caused the mutant to be exclusively localized in nuclei. The fusion of this leucine-zipper motif with histone 2A, which is a nuclear protein, could induce the fusion protein to be exported from nucleus. Cell migration was greatly inhibited in PTRF/cavin-1{sup −/−} mouse embryonic fibroblasts (MEFs). The inhibited cell motility could only be rescued by exogenous cavin-1 but not the leucine-zipper motif deleted cavin-1 mutant. Plasma membrane dynamics is an important factor in cell motility control. Our results suggested that the membrane dynamics in cell migration is affected by caveolae associated PTRF/cavin-1.

  12. The N-terminal leucine-zipper motif in PTRF/cavin-1 is essential and sufficient for its caveolae-association

    International Nuclear Information System (INIS)

    Wei, Zhuang; Zou, Xinle; Wang, Hongzhong; Lei, Jigang; Wu, Yuan; Liao, Kan

    2015-01-01

    Highlight: • The N-terminal leucine-zipper motif in PTRF/cavin-1 determines caveolar association. • Different cellular localization of PTRF/cavin-1 influences its serine 389 and 391 phosphorylation state. • PTRF/cavin-1 regulates cell motility via its caveolar association. - Abstract: PTRF/cavin-1 is a protein of two lives. Its reported functions in ribosomal RNA synthesis and in caveolae formation happen in two different cellular locations: nucleus vs. plasma membrane. Here, we identified that the N-terminal leucine-zipper motif in PTRF/cavin-1 was essential for the protein to be associated with caveolae in plasma membrane. It could counteract the effect of nuclear localization sequence in the molecule (AA 235–251). Deletion of this leucine-zipper motif from PTRF/cavin-1 caused the mutant to be exclusively localized in nuclei. The fusion of this leucine-zipper motif with histone 2A, which is a nuclear protein, could induce the fusion protein to be exported from nucleus. Cell migration was greatly inhibited in PTRF/cavin-1 −/− mouse embryonic fibroblasts (MEFs). The inhibited cell motility could only be rescued by exogenous cavin-1 but not the leucine-zipper motif deleted cavin-1 mutant. Plasma membrane dynamics is an important factor in cell motility control. Our results suggested that the membrane dynamics in cell migration is affected by caveolae associated PTRF/cavin-1

  13. Structure and function of homodomain-leucine zipper (HD-Zip) proteins.

    Science.gov (United States)

    Elhiti, Mohamed; Stasolla, Claudio

    2009-02-01

    Homeodomain-leucine zipper (HD-Zip) proteins are transcription factors unique to plants and are encoded by more than 25 genes in Arabidopsis thaliana. Based on sequence analyses these proteins have been classified into four distinct groups: HD-Zip I-IV. HD-Zip proteins are characterized by the presence of two functional domains; a homeodomain (HD) responsible for DNA binding and a leucine zipper domain (Zip) located immediately C-terminal to the homeodomain and involved in protein-protein interaction. Despite sequence similarities HD-ZIP proteins participate in a variety of processes during plant growth and development. HD-Zip I proteins are generally involved in responses related to abiotic stress, abscisic acid (ABA), blue light, de-etiolation and embryogenesis. HD-Zip II proteins participate in light response, shade avoidance and auxin signalling. Members of the third group (HD-Zip III) control embryogenesis, leaf polarity, lateral organ initiation and meristem function. HD-Zip IV proteins play significant roles during anthocyanin accumulation, differentiation of epidermal cells, trichome formation and root development.

  14. Growth hormone (GH)-independent dimerization of GH receptor by a leucine zipper results in constitutive activation

    DEFF Research Database (Denmark)

    Behncken, S N; Billestrup, Nils; Brown, R

    2000-01-01

    Growth hormone initiates signaling by inducing homodimerization of two GH receptors. Here, we have sought to determine whether constitutively active receptor can be created in the absence of the extracellular domain by substituting it with high affinity leucine zippers to create dimers of the gro......Growth hormone initiates signaling by inducing homodimerization of two GH receptors. Here, we have sought to determine whether constitutively active receptor can be created in the absence of the extracellular domain by substituting it with high affinity leucine zippers to create dimers...

  15. SREBP-1 dimerization specificity maps to both the helix-loop-helix and leucine zipper domains: use of a dominant negative

    DEFF Research Database (Denmark)

    Rishi, Vikas; Gal, Jozsef; Krylov, Dmitry

    2004-01-01

    -HLH-ZIP proteins MAX, USF, or MITF, even at 100 molar eq. Chimeric proteins containing the HLH domain of SREBP-1 and the leucine zipper from either MAX, USF, or MITF indicate that both the HLH and leucine zipper regions of SREBP-1 contribute to its dimerization specificity. Transient co-transfection studies...

  16. The role of basic leucine zipper transcription factor E4BP4 in the immune system and immune-mediated diseases.

    Science.gov (United States)

    Yin, Jinghua; Zhang, Jian; Lu, Qianjin

    2017-07-01

    Basic leucine zipper transcription factor E4BP4 (also known as NFIL3) has been implicated in the molecular and cellular mechanisms of functions and activities in mammals. The interactions between E4BP4 and major regulators of cellular processes have triggered significant interest in the roles of E4BP4 in the pathogenesis of certain chronic diseases. Indeed, novel discoveries have been emerging to illustrate the involvement of E4BP4 in multiple disorders. It is recognized that E4BP4 is extensively involved in some immune-mediated diseases, but the mechanisms of E4BP4 involvement in these complex diseases remain poorly defined. Here we review the regulatory mechanisms of E4BP4 engaging in not only the biological function but also the development of immune-mediated diseases, paving the way for future therapies. Copyright © 2017. Published by Elsevier Inc.

  17. Introduction of a leucine half-zipper engenders multiple high-quality crystals of a recalcitrant tRNA synthetase

    International Nuclear Information System (INIS)

    Guo, Min; Shapiro, Ryan; Schimmel, Paul; Yang, Xiang-Lei

    2010-01-01

    E. coli alanyl-tRNA synthetase is recalcitrant to crystallization. A group of leucine substitutions has transformed the protein. Although Escherichia coli alanyl-tRNA synthetase was among the first tRNA synthetases to be sequenced and extensively studied by functional analysis, it has proved to be recalcitrant to crystallization. This challenge remained even for crystallization of the catalytic fragment. By mutationally introducing three stacked leucines onto the solvent-exposed side of an α-helix, an engineered catalytic fragment of the synthetase was obtained that yielded multiple high-quality crystals and cocrystals with different ligands. The engineered α-helix did not form a leucine zipper that interlocked with the same α-helix from another molecule. Instead, using the created hydrophobic spine, it interacted with other surfaces of the protein as a leucine half-zipper (LHZ) to enhance the crystal lattice interactions. The LHZ made crystal lattice contacts in all crystals of different space groups. These results illustrate the power of introducing an LHZ into helices to facilitate crystallization. The authors propose that the method can be unified with surface-entropy reduction and can be broadly used for protein-surface optimization in crystallization

  18. Leucine zipper, down regulated in cancer-1 gene expression in prostate cancer

    OpenAIRE

    Salemi, Michele; Barone, Nunziata; La Vignera, Sandro; Condorelli, Rosita A.; Recupero, Domenico; Galia, Antonio; Fraggetta, Filippo; Aiello, Anna Maria; Pepe, Pietro; Castiglione, Roberto; Vicari, Enzo; Calogero, Aldo E.

    2016-01-01

    Numerous genetic alterations have been implicated in the development of prostate cancer (PCa). DNA and protein microarrays have enabled the identification of genes associated with apoptosis, which is important in PCa development. Despite the molecular mechanisms are not entirely understood, inhibition of apoptosis is a critical pathophysiological factor that contributes to the onset and progression of PCa. Leucine zipper, down-regulated in cancer 1 (LDOC-1) is a known regulator of the nuclear...

  19. Abscisic-acid-dependent basic leucine zipper (bZIP) transcription factors in plant abiotic stress.

    Science.gov (United States)

    Banerjee, Aditya; Roychoudhury, Aryadeep

    2017-01-01

    One of the major causes of significant crop loss throughout the world is the myriad of environmental stresses including drought, salinity, cold, heavy metal toxicity, and ultraviolet-B (UV-B) rays. Plants as sessile organisms have evolved various effective mechanism which enable them to withstand this plethora of stresses. Most of such regulatory mechanisms usually follow the abscisic-acid (ABA)-dependent pathway. In this review, we have primarily focussed on the basic leucine zipper (bZIP) transcription factors (TFs) activated by the ABA-mediated signalosome. Upon perception of ABA by specialized receptors, the signal is transduced via various groups of Ser/Thr kinases, which phosphorylate the bZIP TFs. Following such post-translational modification of TFs, they are activated so that they bind to specific cis-acting sequences called abscisic-acid-responsive elements (ABREs) or GC-rich coupling elements (CE), thereby influencing the expression of their target downstream genes. Several in silico techniques have been adopted so far to predict the structural features, recognize the regulatory modification sites, undergo phylogenetic analyses, and facilitate genome-wide survey of TF under multiple stresses. Current investigations on the epigenetic regulation that controls greater accessibility of the inducible regions of DNA of the target gene to the bZIP TFs exclusively under stress situations, along with the evolved stress memory responses via genomic imprinting mechanism, have been highlighted. The potentiality of overexpression of bZIP TFs, either in a homologous or in a heterologous background, in generating transgenic plants tolerant to various abiotic stressors have also been addressed by various groups. The present review will provide a coherent documentation on the functional characterization and regulation of bZIP TFs under multiple environmental stresses, with the major goal of generating multiple-stress-tolerant plant cultivars in near future.

  20. Divergent effects of endogenous and exogenous glucocorticoid-induced leucine zipper in animal models of inflammation and arthritis

    NARCIS (Netherlands)

    Ngo, Devi; Beaulieu, Elaine; Gu, Ran; Leaney, Alexandra; Santos, Leilani; Fan, Huapeng; Yang, Yuanhang; Kao, Wenping; Xu, Jiake; Escriou, Virginie; Loiler, Scott; Vervoordeldonk, Margriet J.; Morand, Eric F.

    2013-01-01

    Glucocorticoid-induced leucine zipper (GILZ) has effects on inflammatory pathways that suggest it to be a key inhibitory regulator of the immune system, and its expression is exquisitely sensitive to induction by glucocorticoids. We undertook this study to test our hypothesis that GILZ deficiency

  1. Preliminary structural studies on the leucine-zipper homology region of the human protein Bap31

    International Nuclear Information System (INIS)

    Mukasa, Takashi; Santelli, Eugenio; Reed, John C.; Pascual, Jaime

    2007-01-01

    A leucine-zipper with properties as apoptotic regulator in the ER has been crystallized. X-ray data to 2.5 Å resolution were collected, molecular replacement solutions were identified and refinement has been started. B-cell receptor-associated protein 31 (Bap31) is an integral membrane protein located in the endoplasmic reticulum (ER) that participates in the transport and quality control of membrane proteins and plays a role in determining cell sensitivity to ER stress and apoptosis. Its cytoplasmic region contains two target sites for caspase cleavage in certain apoptotic pathways. Here, the subcloning, expression, purification and crystallization of the Homo sapiens Bap31 leucine-zipper C-terminal fragment, which spans residues Gly160–Glu246, are reported. An N-terminally His-tagged protein was overexpressed in Escherichia coli and purified by chromatographic methods. X-ray diffraction data were collected in-house to 2.5 Å resolution. Crystals belong to space group P6 1 22/P6 5 22, with unit-cell parameters a = b = 70.7, c = 80.6 Å. Data analysis indicates the presence of one molecule per asymmetric unit

  2. Preliminary structural studies on the leucine-zipper homology region of the human protein Bap31

    Energy Technology Data Exchange (ETDEWEB)

    Mukasa, Takashi; Santelli, Eugenio [Program on Infectious Diseases, Center for Inflammation and Infectious Diseases, The Burnham Institute for Medical Research, 10901 North Torrey Pines Road, La Jolla, CA 92037 (United States); Reed, John C. [Program on Apoptosis, Cancer Center, The Burnham Institute for Medical Research, 10901 North Torrey Pines Road, La Jolla, CA 92037 (United States); Pascual, Jaime, E-mail: pascual@burnham.org [Program on Infectious Diseases, Center for Inflammation and Infectious Diseases, The Burnham Institute for Medical Research, 10901 North Torrey Pines Road, La Jolla, CA 92037 (United States)

    2007-04-01

    A leucine-zipper with properties as apoptotic regulator in the ER has been crystallized. X-ray data to 2.5 Å resolution were collected, molecular replacement solutions were identified and refinement has been started. B-cell receptor-associated protein 31 (Bap31) is an integral membrane protein located in the endoplasmic reticulum (ER) that participates in the transport and quality control of membrane proteins and plays a role in determining cell sensitivity to ER stress and apoptosis. Its cytoplasmic region contains two target sites for caspase cleavage in certain apoptotic pathways. Here, the subcloning, expression, purification and crystallization of the Homo sapiens Bap31 leucine-zipper C-terminal fragment, which spans residues Gly160–Glu246, are reported. An N-terminally His-tagged protein was overexpressed in Escherichia coli and purified by chromatographic methods. X-ray diffraction data were collected in-house to 2.5 Å resolution. Crystals belong to space group P6{sub 1}22/P6{sub 5}22, with unit-cell parameters a = b = 70.7, c = 80.6 Å. Data analysis indicates the presence of one molecule per asymmetric unit.

  3. Basic leucine zipper protein Cnc-C is a substrate and transcriptional regulator of the Drosophila 26S proteasome.

    Science.gov (United States)

    Grimberg, Kristian Björk; Beskow, Anne; Lundin, Daniel; Davis, Monica M; Young, Patrick

    2011-02-01

    While the 26S proteasome is a key proteolytic complex, little is known about how proteasome levels are maintained in higher eukaryotic cells. Here we describe an RNA interference (RNAi) screen of Drosophila melanogaster that was used to identify transcription factors that may play a role in maintaining levels of the 26S proteasome. We used an RNAi library against 993 Drosophila transcription factor genes to identify genes whose suppression in Schneider 2 cells stabilized a ubiquitin-green fluorescent protein reporter protein. This screen identified Cnc (cap 'n' collar [CNC]; basic region leucine zipper) as a candidate transcriptional regulator of proteasome component expression. In fact, 20S proteasome activity was reduced in cells depleted of cnc. Immunoblot assays against proteasome components revealed a general decline in both 19S regulatory complex and 20S proteasome subunits after RNAi depletion of this transcription factor. Transcript-specific silencing revealed that the longest of the seven transcripts for the cnc gene, cnc-C, was needed for proteasome and p97 ATPase production. Quantitative reverse transcription-PCR confirmed the role of Cnc-C in activation of transcription of genes encoding proteasome components. Expression of a V5-His-tagged form of Cnc-C revealed that the transcription factor is itself a proteasome substrate that is stabilized when the proteasome is inhibited. We propose that this single cnc gene in Drosophila resembles the ancestral gene family of mammalian nuclear factor erythroid-derived 2-related transcription factors, which are essential in regulating oxidative stress and proteolysis.

  4. Arabidopsis basic leucine zipper transcription factors involved in an abscisic acid-dependent signal transduction pathway under drought and high-salinity conditions.

    Science.gov (United States)

    Uno, Y; Furihata, T; Abe, H; Yoshida, R; Shinozaki, K; Yamaguchi-Shinozaki, K

    2000-10-10

    The induction of the dehydration-responsive Arabidopsis gene, rd29B, is mediated mainly by abscisic acid (ABA). Promoter analysis of rd29B indicated that two ABA-responsive elements (ABREs) are required for the dehydration-responsive expression of rd29B as cis-acting elements. Three cDNAs encoding basic leucine zipper (bZIP)-type ABRE-binding proteins were isolated by using the yeast one-hybrid system and were designated AREB1, AREB2, and AREB3 (ABA-responsive element binding protein). Transcription of the AREB1 and AREB2 genes is up-regulated by drought, NaCl, and ABA treatment in vegetative tissues. In a transient transactivation experiment using Arabidopsis leaf protoplasts, both the AREB1 and AREB2 proteins activated transcription of a reporter gene driven by ABRE. AREB1 and AREB2 required ABA for their activation, because their transactivation activities were repressed in aba2 and abi1 mutants and enhanced in an era1 mutant. Activation of AREBs by ABA was suppressed by protein kinase inhibitors. These results suggest that both AREB1 and AREB2 function as transcriptional activators in the ABA-inducible expression of rd29B, and further that ABA-dependent posttranscriptional activation of AREB1 and AREB2, probably by phosphorylation, is necessary for their maximum activation by ABA. Using cultured Arabidopsis cells, we demonstrated that a specific ABA-activated protein kinase of 42-kDa phosphorylated conserved N-terminal regions in the AREB proteins.

  5. Maternal Embryonic Leucine Zipper Kinase (MELK: A Novel Regulator in Cell Cycle Control, Embryonic Development, and Cancer

    Directory of Open Access Journals (Sweden)

    Pengfei Jiang

    2013-10-01

    Full Text Available Maternal embryonic leucine zipper kinase (MELK functions as a modulator of intracellular signaling and affects various cellular and biological processes, including cell cycle, cell proliferation, apoptosis, spliceosome assembly, gene expression, embryonic development, hematopoiesis, and oncogenesis. In these cellular processes, MELK functions by binding to numerous proteins. In general, the effects of multiple protein interactions with MELK are oncogenic in nature, and the overexpression of MELK in kinds of cancer provides some evidence that it may be involved in tumorigenic process. In this review, our current knowledge of MELK function and recent discoveries in MELK signaling pathway were discussed. The regulation of MELK in cancers and its potential as a therapeutic target were also described.

  6. Regulation of MIR165/166 by class II and class III homeodomain leucine zipper proteins establishes leaf polarity

    DEFF Research Database (Denmark)

    Merelo, Paz; Ram, Hathi; Caggiano, Monica Pia

    2016-01-01

    A defining feature of plant leaves is their flattened shape. This shape depends on an antagonism between the genes that specify adaxial (top) and abaxial (bottom) tissue identity; however, the molecular nature of this antagonism remains poorly understood. Class III homeodomain leucine zipper (HD-...... show that class III and class II HD-ZIP proteins act together to repress MIR165/166 via a conserved cis-element in their promoters. Organ morphology and tissue patterning in plants, therefore, depend on a bidirectional repressive circuit involving a set of miRNAs and its targets....

  7. Glucocorticoid Induced Leucine Zipper inhibits apoptosis of cardiomyocytes by doxorubicin

    International Nuclear Information System (INIS)

    Aguilar, David; Strom, Joshua; Chen, Qin M.

    2014-01-01

    Doxorubicin (Dox) is an indispensable chemotherapeutic agent for the treatment of various forms of neoplasia such as lung, breast, ovarian, and bladder cancers. Cardiotoxicity is a major concern for patients receiving Dox therapy. Previous work from our laboratory indicated that glucocorticoids (GCs) alleviate Dox-induced apoptosis in cardiomyocytes. Here we have found Glucocorticoid-Induced Leucine Zipper (GILZ) to be a mediator of GC-induced cytoprotection. GILZ was found to be induced in cardiomyocytes by GC treatment. Knocking down of GILZ using siRNA resulted in cancelation of GC-induced cytoprotection against apoptosis by Dox treatment. Overexpressing GILZ by transfection was able to protect cells from apoptosis induced by Dox as measured by caspase activation, Annexin V binding and morphologic changes. Western blot analyses indicate that GILZ overexpression prevented cytochrome c release from mitochondria and cleavage of caspase-3. When bcl-2 family proteins were examined, we found that GILZ overexpression causes induction of the pro-survival protein Bcl-xL. Since siRNA against Bcl-xL reverses GC induced cytoprotection, Bcl-xL induction represents an important event in GILZ-induced cytoprotection. Our data suggest that GILZ functions as a cytoprotective gene in cardiomyocytes. - Highlights: • Corticosteroids act as a cytoprotective agent in cardiomyocytes • Corticosteroids induce GILZ expression in cardiomyocytes • Elevated GILZ results in resistance against apoptosis induced by doxorubicin • GILZ induces Bcl-xL protein without inducing Bcl-xL mRNA

  8. A Petunia Homeodomain-Leucine Zipper Protein, PhHD-Zip, Plays an Important Role in Flower Senescence

    Science.gov (United States)

    Chang, Xiaoxiao; Donnelly, Linda; Sun, Daoyang; Rao, Jingping; Reid, Michael S.; Jiang, Cai-Zhong

    2014-01-01

    Flower senescence is initiated by developmental and environmental signals, and regulated by gene transcription. A homeodomain-leucine zipper transcription factor, PhHD-Zip, is up-regulated during petunia flower senescence. Virus-induced gene silencing of PhHD-Zip extended flower life by 20% both in unpollinated and pollinated flowers. Silencing PhHD-Zip also dramatically reduced ethylene production and the abundance of transcripts of genes involved in ethylene (ACS, ACO), and ABA (NCED) biosynthesis. Abundance of transcripts of senescence-related genes (SAG12, SAG29) was also dramatically reduced in the silenced flowers. Over-expression of PhHD-Zip accelerated petunia flower senescence. Furthermore, PhHD-Zip transcript abundance in petunia flowers was increased by application of hormones (ethylene, ABA) and abiotic stresses (dehydration, NaCl and cold). Our results suggest that PhHD-Zip plays an important role in regulating petunia flower senescence. PMID:24551088

  9. A Petunia homeodomain-leucine zipper protein, PhHD-Zip, plays an important role in flower senescence.

    Science.gov (United States)

    Chang, Xiaoxiao; Donnelly, Linda; Sun, Daoyang; Rao, Jingping; Reid, Michael S; Jiang, Cai-Zhong

    2014-01-01

    Flower senescence is initiated by developmental and environmental signals, and regulated by gene transcription. A homeodomain-leucine zipper transcription factor, PhHD-Zip, is up-regulated during petunia flower senescence. Virus-induced gene silencing of PhHD-Zip extended flower life by 20% both in unpollinated and pollinated flowers. Silencing PhHD-Zip also dramatically reduced ethylene production and the abundance of transcripts of genes involved in ethylene (ACS, ACO), and ABA (NCED) biosynthesis. Abundance of transcripts of senescence-related genes (SAG12, SAG29) was also dramatically reduced in the silenced flowers. Over-expression of PhHD-Zip accelerated petunia flower senescence. Furthermore, PhHD-Zip transcript abundance in petunia flowers was increased by application of hormones (ethylene, ABA) and abiotic stresses (dehydration, NaCl and cold). Our results suggest that PhHD-Zip plays an important role in regulating petunia flower senescence.

  10. A Petunia homeodomain-leucine zipper protein, PhHD-Zip, plays an important role in flower senescence.

    Directory of Open Access Journals (Sweden)

    Xiaoxiao Chang

    Full Text Available Flower senescence is initiated by developmental and environmental signals, and regulated by gene transcription. A homeodomain-leucine zipper transcription factor, PhHD-Zip, is up-regulated during petunia flower senescence. Virus-induced gene silencing of PhHD-Zip extended flower life by 20% both in unpollinated and pollinated flowers. Silencing PhHD-Zip also dramatically reduced ethylene production and the abundance of transcripts of genes involved in ethylene (ACS, ACO, and ABA (NCED biosynthesis. Abundance of transcripts of senescence-related genes (SAG12, SAG29 was also dramatically reduced in the silenced flowers. Over-expression of PhHD-Zip accelerated petunia flower senescence. Furthermore, PhHD-Zip transcript abundance in petunia flowers was increased by application of hormones (ethylene, ABA and abiotic stresses (dehydration, NaCl and cold. Our results suggest that PhHD-Zip plays an important role in regulating petunia flower senescence.

  11. Genome-wide analysis of the homeodomain-leucine zipper (HD-ZIP) gene family in peach (Prunus persica).

    Science.gov (United States)

    Zhang, C H; Ma, R J; Shen, Z J; Sun, X; Korir, N K; Yu, M L

    2014-04-08

    In this study, 33 homeodomain-leucine zipper (HD-ZIP) genes were identified in peach using the HD-ZIP amino acid sequences of Arabidopsis thaliana as a probe. Based on the phylogenetic analysis and the individual gene or protein characteristics, the HD-ZIP gene family in peach can be classified into 4 subfamilies, HD-ZIP I, II, III, and IV, containing 14, 7, 4, and 8 members, respectively. The most closely related peach HD-ZIP members within the same subfamilies shared very similar gene structure in terms of either intron/exon numbers or lengths. Almost all members of the same subfamily shared common motif compositions, thereby implying that the HD-ZIP proteins within the same subfamily may have functional similarity. The 33 peach HD-ZIP genes were distributed across scaffolds 1 to 7. Although the primary structure varied among HD-ZIP family proteins, their tertiary structures were similar. The results from this study will be useful in selecting candidate genes from specific subfamilies for functional analysis.

  12. Leucine zipper, down regulated in cancer-1 gene expression in prostate cancer

    Science.gov (United States)

    Salemi, Michele; Barone, Nunziata; La Vignera, Sandro; Condorelli, Rosita A.; Recupero, Domenico; Galia, Antonio; Fraggetta, Filippo; Aiello, Anna Maria; Pepe, Pietro; Castiglione, Roberto; Vicari, Enzo; Calogero, Aldo E.

    2016-01-01

    Numerous genetic alterations have been implicated in the development of prostate cancer (PCa). DNA and protein microarrays have enabled the identification of genes associated with apoptosis, which is important in PCa development. Despite the molecular mechanisms are not entirely understood, inhibition of apoptosis is a critical pathophysiological factor that contributes to the onset and progression of PCa. Leucine zipper, down-regulated in cancer 1 (LDOC-1) is a known regulator of the nuclear factor (NF)-mediated pathway of apoptosis through the inhibition of NF-κB. The present study investigated the expression of the LDOC-1 gene in LNCaP, PC-3, PNT1A and PNT2 prostate cell lines by reverse transcription-quantitative polymerase chain reaction. In addition LDOC-1 protein expression in normal prostate tissues and PCa was studied by immunohistochemistry. LDOC-1 messenger RNA resulted overexpressed in LNCaP and PC-3 PCa cell lines compared with the two normal prostate cell lines PNT1A and PNT2. The results of immunohistochemistry demonstrated a positive cytoplasmic LDOC-1 staining in all PCa and normal prostate samples, whereas no nuclear staining was observed in any sample. Furthermore, a more intense signal was evidenced in PCa samples. LDOC-1 gene overexpression in PCa suggests an activity of LDOC-1 in PCa cell lines. PMID:27698860

  13. Small leucine zipper protein functions as a negative regulator of estrogen receptor α in breast cancer.

    Directory of Open Access Journals (Sweden)

    Juyeon Jeong

    Full Text Available The nuclear transcription factor estrogen receptor α (ERα plays a critical role in breast cancer progression. ERα acts as an important growth stimulatory protein in breast cancer and the expression level of ERα is tightly related to the prognosis and treatment of patients. Small leucine zipper protein (sLZIP functions as a transcriptional cofactor by binding to various nuclear receptors, including glucocorticoid receptor, androgen receptor, and peroxisome proliferator-activated receptor γ. However, the role of sLZIP in the regulation of ERα and its involvement in breast cancer progression is unknown. We found that sLZIP binds to ERα and represses the transcriptional activity of ERα in ERα-positive breast cancer cells. sLZIP also suppressed the expression of ERα target genes. sLZIP disrupted the binding of ERα to the estrogen response element of the target gene promoter, resulting in suppression of cell proliferation. sLZIP is a novel co-repressor of ERα, and plays a negative role in ERα-mediated cell proliferation in breast cancer.

  14. Hepatocyte growth factor limits autoimmune neuroinflammation via glucocorticoid-induced leucine zipper expression in dendritic cells.

    Science.gov (United States)

    Benkhoucha, Mahdia; Molnarfi, Nicolas; Dunand-Sauthier, Isabelle; Merkler, Doron; Schneiter, Gregory; Bruscoli, Stefano; Riccardi, Carlo; Tabata, Yasuhiko; Funakoshi, Hiroshi; Nakamura, Toshikazu; Reith, Walter; Santiago-Raber, Marie-Laure; Lalive, Patrice H

    2014-09-15

    Autoimmune neuroinflammation, including multiple sclerosis and its animal model, experimental autoimmune encephalomyelitis (EAE), a prototype for T cell-mediated autoimmunity, is believed to result from immune tolerance dysfunction leading to demyelination and substantial neurodegeneration. We previously showed that CNS-restricted expression of hepatocyte growth factor (HGF), a potent neuroprotective factor, reduced CNS inflammation and clinical deficits associated with EAE. In this study, we demonstrate that systemic HGF treatment ameliorates EAE through the development of tolerogenic dendritic cells (DCs) with high expression levels of glucocorticoid-induced leucine zipper (GILZ), a transcriptional repressor of gene expression and a key endogenous regulator of the inflammatory response. RNA interference-directed neutralization of GILZ expression by DCs suppressed the induction of tolerance caused by HGF. Finally, adoptive transfer of HGF-treated DCs from wild-type but not GILZ gene-deficient mice potently mediated functional recovery in recipient mice with established EAE through effective modulation of autoaggressive T cell responses. Altogether, these results show that by inducing GILZ in DCs, HGF reproduces the mechanism of immune regulation induced by potent immunomodulatory factors such as IL-10, TGF-β1, and glucocorticoids and therefore that HGF therapy may have potential in the treatment of autoimmune dysfunctions. Copyright © 2014 by The American Association of Immunologists, Inc.

  15. Thermodynamic analysis of cavity creating mutations in an engineered leucine zipper and energetics of glycerol-induced coiled coil stabilization.

    Science.gov (United States)

    Dürr, E; Jelesarov, I

    2000-04-18

    Protein stability in vitro can be influenced either by introduction of mutations or by changes in the chemical composition of the solvent. Recently, we have characterized the thermodynamic stability and the rate of folding of the engineered dimeric leucine zipper A(2), which has a strengthened hydrophobic core [Dürr, E., Jelesarov, I., and Bosshard, H. R. (1999) Biochemistry 38, 870-880]. Here we report on the energetic consequences of a cavity introduced by Leu/Ala substitution at the tightly packed dimeric interface and how addition of 30% glycerol affects the folding thermodynamics of A(2) and the cavity mutants. Folding could be described by a two-state transition from two unfolded monomers to a coiled coil dimer. Removal of six methylene groups by Leu/Ala substitutions destabilized the dimeric coiled coil by 25 kJ mol(-1) at pH 3.5 and 25 degrees C in aqueous buffer. Destabilization was purely entropic at around room temperature and became increasingly enthalpic at elevated temperatures. Mutations were accompanied by a decrease of the unfolding heat capacity by 0.5 kJ K(-1) mol(-1). Addition of 30% glycerol increased the free energy of folding of A(2) and the cavity mutants by 5-10 kJ mol(-1) and lowered the unfolding heat capacity by 25% for A(2) and by 50% for the Leu/Ala mutants. The origin of the stabilizing effect of glycerol varied with temperature. Stabilization of the parent leucine zipper A(2) was enthalpic with an unfavorable entropic component between 0 and 100 degrees C. In the case of cavity mutants, glycerol induced enthalpic stabilization below 50 degrees C and entropic stabilization above 50 degrees C. The effect of glycerol could not be accounted for solely by the enthalpy and entropy of transfer or protein surface from water to glycerol/water mixture. We propose that in the presence of glycerol the folded coiled coil dimer is better packed and displays less intramolecular fluctuations, leading to enhanced enthalpic interactions and to an

  16. Expression and regulation of glucocorticoid-induced leucine zipper in the developing anterior pituitary gland.

    Science.gov (United States)

    Ellestad, Laura E; Malkiewicz, Stefanie A; Guthrie, H David; Welch, Glenn R; Porter, Tom E

    2009-02-01

    The expression profile of glucocorticoid-induced leucine zipper (GILZ) in the anterior pituitary during the second half of embryonic development in the chick is consistent with in vivo regulation by circulating corticosteroids. However, nothing else has been reported about the presence of GILZ in the neuroendocrine system. We sought to characterize expression and regulation of GILZ in the chicken embryonic pituitary gland and determine the effect of GILZ overexpression on anterior pituitary hormone levels. Pituitary GILZ mRNA levels increased during embryogenesis to a maximum on the day of hatch, and decreased through the first week after hatch. GILZ expression was rapidly upregulated by corticosterone in embryonic pituitary cells. To determine whether GILZ regulates hormone gene expression in the developing anterior pituitary, we overexpressed GILZ in embryonic pituitary cells and measured mRNA for the major pituitary hormones. Exogenous GILZ increased prolactin mRNA above basal levels, but not as high as that in corticosterone-treated cells, indicating that GILZ may play a small role in lactotroph differentiation. The largest effect we observed was a twofold increase in FSH beta subunit in cells transfected with GILZ but not treated with corticosterone, suggesting that GILZ may positively regulate gonadotroph development in a manner not involving glucocorticoids. In conclusion, this is the first report to characterize avian GILZ and examine its regulation in the developing neuroendocrine system. We have shown that GILZ is upregulated by glucocorticoids in the embryonic pituitary gland and may regulate expression of several pituitary hormones.

  17. Alanine Zipper-Like Coiled-Coil Domains Are Necessary for Homotypic Dimerization of Plant GAGA-Factors in the Nucleus and Nucleolus

    Science.gov (United States)

    Bloss, Ulrich; Hecker, Andreas; Elgass, Kirstin; Hummel, Sabine; Hahn, Achim; Caesar, Katharina; Schleifenbaum, Frank; Harter, Klaus; Berendzen, Kenneth W.

    2011-01-01

    GAGA-motif binding proteins control transcriptional activation or repression of homeotic genes. Interestingly, there are no sequence similarities between animal and plant proteins. Plant BBR/BPC-proteins can be classified into two distinct groups: Previous studies have elaborated on group I members only and so little is known about group II proteins. Here, we focused on the initial characterization of AtBPC6, a group II protein from Arabidopsis thaliana. Comparison of orthologous BBR/BPC sequences disclosed two conserved signatures besides the DNA binding domain. A first peptide signature is essential and sufficient to target AtBPC6-GFP to the nucleus and nucleolus. A second domain is predicted to form a zipper-like coiled-coil structure. This novel type of domain is similar to Leucine zippers, but contains invariant alanine residues with a heptad spacing of 7 amino acids. By yeast-2-hybrid and BiFC-assays we could show that this Alanine zipper domain is essential for homotypic dimerization of group II proteins in vivo. Interhelical salt bridges and charge-stabilized hydrogen bonds between acidic and basic residues of the two monomers are predicted to form an interaction domain, which does not follow the classical knobs-into-holes zipper model. FRET-FLIM analysis of GFP/RFP-hybrid fusion proteins validates the formation of parallel dimers in planta. Sequence comparison uncovered that this type of domain is not restricted to BBR/BPC proteins, but is found in all kingdoms. PMID:21347358

  18. Identification of glucocorticoid-induced leucine zipper as a key regulator of tumor cell proliferation in epithelial ovarian cancer

    Directory of Open Access Journals (Sweden)

    Fernandez Hervé

    2009-10-01

    Full Text Available Abstract Background Little is known about the molecules that contribute to tumor progression of epithelial ovarian cancer (EOC, currently a leading cause of mortality from gynecological malignancies. Glucocorticoid-Induced Leucine Zipper (GILZ, an intracellular protein widely expressed in immune tissues, has been reported in epithelial tissues and controls some of key signaling pathways involved in tumorigenesis. However, there has been no report on GILZ in EOC up to now. The objectives of the current study were to examine the expression of GILZ in EOC and its effect on tumor cell proliferation. Results GILZ expression was measured by immunohistochemical staining in tissue sections from 3 normal ovaries, 7 benign EOC and 50 invasive EOC. GILZ was not detected on the surface epithelium of normal ovaries and benign tumors. In contrast, it was expressed in the cytoplasm of tumor cells in 80% EOC specimens. GILZ immunostaining scores correlated positively to the proliferation marker Ki-67 (Spearman test in univariate analysis, P P Conclusion The present study is the first to identify GILZ as a molecule produced by ovarian cancer cells that promotes cell cycle progression and proliferation. Our findings clearly indicate that GILZ activates AKT, a crucial signaling molecule in tumorigenesis. GILZ thus appears as a potential key molecule in EOC.

  19. Glucocorticoid-induced leucine zipper expression is associated with response to treatment and immunoregulation in systemic lupus erythematosus.

    Science.gov (United States)

    Mohammadi, Saeed; Ebadpour, Mohammad Reza; Sedighi, Sima; Saeedi, Mohsen; Memarian, Ali

    2017-08-01

    Systemic lupus erythematosus (SLE) is an autoimmune disorder in which cytokine balance is disturbed. Glucocorticoids (GCs) are shown to balance immune response by transcriptional regulation of glucocorticoid receptor target genes such as Glucocorticoid-induced leucine zipper (GILZ) which has been introduced as an endogenous anti-inflammatory mediator. In the present study, we assessed the expression of GILZ in association with interferon-γ (IFN-γ), interleukine-10 (IL-10), and B lymphocyte stimulator (BLyS) plasma levels in SLE patients. A total of 40 female patients (18 under treatment and 22 newly diagnosed) were recruited in this study. Real-time RT PCR was conducted to quantify the mRNA expression of GILZ. The plasma levels of IFN-γ, IL-10, and BLyS were evaluated using ELISA method. GILZ was overexpressed among under treatment SLE patients. The mRNA expression of GILZ was significantly correlated with Systemic Lupus Erythematosus Disease Activity Index (SLEDAI) score. IFN-γ and BLyS were downregulated in response to therapies with negative correlations to GILZ. Moreover, IL-10 was upregulated among treated patients. The levels of IFN-γ and BLyS were correlated with the severity of disease, while IL-10 was negatively correlated with SLEDAI score. GILZ could be introduced as one of the acting molecules in mediating the regulatory effects of GCs on producing pro- and anti-inflammatory cytokines in SLE.

  20. A genome-wide survey of homeodomain-leucine zipper genes and analysis of cold-responsive HD-Zip I members' expression in tomato.

    Science.gov (United States)

    Zhang, Zhenzhu; Chen, Xiuling; Guan, Xin; Liu, Yang; Chen, Hongyu; Wang, Tingting; Mouekouba, Liana Dalcantara Ongouya; Li, Jingfu; Wang, Aoxue

    2014-01-01

    Homeodomain-leucine zipper (HD-Zip) proteins are a kind of transcriptional factors that play a vital role in plant growth and development. However, no detailed information of HD-Zip family in tomato has been reported till now. In this study, 51 HD-Zip genes (SlHZ01-51) in this family were identified and categorized into 4 classes by exon-intron and protein structure in tomato (Solanum lycopersicum) genome. The synthetical phylogenetic tree of tomato, Arabidopsis and rice HD-Zip genes were established for an insight into their evolutionary relationships and putative functions. The results showed that the contribution of segmental duplication was larger than that of tandem duplication for expansion and evolution of genes in this family of tomato. The expression profile results under abiotic stress suggested that all SlHZ I genes were responsive to cold stress. This study will provide a clue for the further investigation of functional identification and the role of tomato HD-Zip I subfamily in plant cold stress responses and developmental events.

  1. Engineering of a novel zipFv using leucine zipper motif against rabies virus glycoprotein G with improved protection potency in vivo.

    Science.gov (United States)

    Xi, Hualong; Zhang, Kaixin; Yin, Yanchun; Gu, Tiejun; Sun, Qing; Li, Zhuang; Cheng, Yue; Jiang, Chunlai; Kong, Wei; Wu, Yongge

    2017-06-01

    Rabies is an acute zoonotic infectious disease with a high fatality rate but is preventable with vaccination and rabies immunoglobulin (RIG). The single-chain Fv fragment (scFv), a small engineered antigen-binding protein derived from antibody variable heavy (V H ) and light (V L ) chains connected by a peptide linker, can potentially be used to replace RIG. Here, we produced two peptides V H -JUN-HIS and V L -FOS-HA separately in Escherichia coli and assembled them to form zipFv successfully in vitro. The new zipFv utilizes FOS and JUN leucine zippers to form an antibody structure similar to the IgG counterpart with two free N-terminal ends of V H and V L . The zipFv protein showed notable improvement in binding ability and affinity over its corresponding scFv. The zipFv also demonstrated greater stability in serum and the same protective rate as RIG against challenge with a standard rabies virus (CVS-24) in mice. Our results indicated zipFv as a novel and efficient antibody form with enhanced neutralizing potency. Copyright © 2017. Published by Elsevier B.V.

  2. GMP production and characterization of leucine zipper-tagged tumor necrosis factor-related apoptosis-inducing ligand (LZ-TRAIL) for phase I clinical trial.

    Science.gov (United States)

    Jiang, Jing; Liu, Xiaobin; Deng, Leixiu; Zhang, Peipei; Wang, Guangjun; Wang, Shifu; Liu, Honghao; Su, Yunpeng

    2014-10-05

    Tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) exhibits potent antitumor activity in a wide range of cancers without deleterious side effects on normal tissues. Several TRAIL derivatives have been developed to improve its pharmacokinetics and therapeutic effects through strategies such as adding a leucine zipper to increase the circulation half-life. To obtain clinical grade LZ-TRAIL for phase I clinical trial, a single batch of 30 L bioreactor culture was performed using the Escherichia coli BL21 (DE3) strain expressing the recombinant LZ-TRAIL. A robust LZ-TRAIL production fermentation process was developed, which could be scaled up from 5L to 50 L, and had a titer of approximately 1.4 g/l. A four-step purification strategy was carried out to obtain a final product with over 95% purity and 45% yield. The final material was filter sterilized, aseptically vialed, and stored at 4°C, and comprehensively characterized using multiple assays (vialed product was sterile, purity was 95%, aggregates were production of phase I clinical trial material. These preclinical investigations warrant further clinical development of this product for cancer therapy. Copyright © 2014. Published by Elsevier B.V.

  3. Application of HB17, an Arabidopsis class II homeodomain-leucine zipper transcription factor, to regulate chloroplast number and photosynthetic capacity.

    Science.gov (United States)

    Hymus, Graham J; Cai, Suqin; Kohl, Elizabeth A; Holtan, Hans E; Marion, Colleen M; Tiwari, Shiv; Maszle, Don R; Lundgren, Marjorie R; Hong, Melissa C; Channa, Namitha; Loida, Paul; Thompson, Rebecca; Taylor, J Philip; Rice, Elena; Repetti, Peter P; Ratcliffe, Oliver J; Reuber, T Lynne; Creelman, Robert A

    2013-11-01

    Transcription factors are proposed as suitable targets for the control of traits such as yield or food quality in plants. This study reports the results of a functional genomics research effort that identified ATHB17, a transcription factor from the homeodomain-leucine zipper class II family, as a novel target for the enhancement of photosynthetic capacity. It was shown that ATHB17 is expressed natively in the root quiescent centre (QC) from Arabidopsis embryos and seedlings. Analysis of the functional composition of genes differentially expressed in the QC from a knockout mutant (athb17-1) compared with its wild-type sibling revealed the over-representation of genes involved in auxin stimulus, embryo development, axis polarity specification, and plastid-related processes. While no other phenotypes were observed in athb17-1 plants, overexpression of ATHB17 produced a number of phenotypes in Arabidopsis including enhanced chlorophyll content. Image analysis of isolated mesophyll cells of 35S::ATHB17 lines revealed an increase in the number of chloroplasts per unit cell size, which is probably due to an increase in the number of proplastids per meristematic cell. Leaf physiological measurements provided evidence of improved photosynthetic capacity in 35S::ATHB17 lines on a per unit leaf area basis. Estimates of the capacity for ribulose-1,5-bisphosphate-saturated and -limited photosynthesis were significantly higher in 35S::ATHB17 lines.

  4. A leucine repeat motif in AbiA is required for resistance of Lactococcus lactis to phages representing three species.

    Science.gov (United States)

    Dinsmore, P K; O'Sullivan, D J; Klaenhammer, T R

    1998-05-28

    The abiA gene encodes an abortive bacteriophage infection mechanism that can protect Lactococcus species from infection by a variety of bacteriophages including three unrelated phage species. Five heptad leucine repeats suggestive of a leucine zipper motif were identified between residues 232 and 266 in the predicted amino acid sequence of the AbiA protein. The biological role of residues in the repeats was investigated by incorporating amino acid substitutions via site-directed mutagenesis. Each mutant was tested for phage resistance against three phages, phi 31, sk1, and c2, belonging to species P335, 936, and c2, respectively. The five residues that comprise the heptad repeats were designated L234, L242, A249, L256, and L263. Three single conservative mutations of leucine to valine in positions L235, L242, and L263 and a double mutation of two leucines (L235 and L242) to valines did not affect AbiA activity on any phages tested. Non-conservative single substitutions of charged amino acids for three of the leucines (L235, L242, and L256) virtually eliminated AbiA activity on all phages tested. Substitution of the alanine residue in the third repeat (A249) with a charged residue did not affect AbiA activity. Replacement of L242 with an alanine elimination phage resistance against phi 31, but partial resistance to sk1 and c2 remained. Two single proline substitutions for leucines L242 and L263 virtually eliminated AbiA activity against all phages, indicating that the predicted alpha-helical structure of this region is important. Mutations in an adjacent region of basic amino acids had various effects on phage resistance, suggesting that these basic residues are also important for AbiA activity. This directed mutagenesis analysis of AbiA indicated that the leucine repeat structure is essential for conferring phage resistance against three species of lactococcal bacteriophages.

  5. Unigenic Evolution: A Novel Genetic Method Localizes a Putative Leucine Zipper That Mediates Dimerization of the Saccharomyces Cerevisiae Regulator Gcr1p

    Science.gov (United States)

    Deminoff, S. J.; Tornow, J.; Santangelo, G. M.

    1995-01-01

    The GCR1 gene of Saccharomyces cerevisiae encodes a transcriptional activator that complexes with Rap1p and, through UAS(RPG) elements (Rap1p DNA binding sites), stimulates efficient expression of glycolytic and translational component genes. To map the functionally important domains in Gcr1p, we combined multiple rounds of random mutagenesis in vitro with in vivo selection of functional genes to locate conserved, or hypomutable, regions. We name this method unigenic evolution, a statistical analysis of mutations in evolutionary variants of a single gene in an otherwise isogenic background. Examination of the distribution of 315 mutations in 24 variant alleles allowed the localization of four hypomutable regions in GCR1 (A, B, C, and D). Dispensable N-terminal (intronic) and C-terminal portions of the evolved region of GCR1 were included in the analysis as controls and were, as expected, not hypomutable. The analysis of several insertion, deletion, and point mutations, combined with a comparison of the hypomutability and hydrophobicity plots of Gcr1p, suggested that some of the hypomutable regions may individually or in combination correspond to functionally important surface domains. In particular, we determined that region D contains a putative leucine zipper and is necessary and sufficient for Gcr1p homodimerization. PMID:8601472

  6. Glucocorticoid-Induced Leucine Zipper Protects the Retina From Light-Induced Retinal Degeneration by Inducing Bcl-xL in Rats.

    Science.gov (United States)

    Gu, Ruiping; Tang, Wenyi; Lei, Boya; Ding, Xinyi; Jiang, Cheng; Xu, Gezhi

    2017-07-01

    The aim of the present study was to investigate the neuroprotective effects of glucocorticoid-induced leucine zipper (GILZ) in a light-induced retinal degeneration model and to explore the underlying mechanisms. Intravitreal injection of recombinant GILZ-overexpressing lentivirus (OE-GILZ-rLV) and short hairpin RNA targeting GILZ recombinant lentivirus (shRNA-GILZ-rLV) was performed to up- and downregulate retinal GILZ, respectively. Three days after stable transduction, rats were exposed to continuous bright light (5000 lux) for 2 days. Retinal function was assessed by full-field electroretinography (ERG), and the retinal structure was examined for photoreceptor survival and death in rats kept under a 12-hour light:2-hour dark cycle following light exposure. The expression levels of retinal Bcl-xL, caspase-9, and caspase-3 were examined by Western blotting or real-time PCR at 1, 3, 5, and 7 days after light exposure. Exposure to bright light downregulated retinal GILZ in parallel with the downregulation of Bcl-xL and the upregulation of active caspase-3. Overexpression of retinal GILZ attenuated the decrease of Bcl-xL and the activation of caspase-9 and caspase-3 at 1, 3, 5, and 7 days after bright light exposure, respectively. GILZ silencing aggravated the downregulation of Bcl-xL induced by bright light exposure. Bright light exposure reduced the amplitude of ERG, increased the number of apoptotic photoreceptor cells, and decreased retinal thickness; and GILZ overexpression could attenuate all these effects. Overexpression of GILZ by OE-GILZ-rLV transduction protected the retina from light-induced cellular damage by activating antiapoptotic pathways.

  7. Adaptor protein containing PH domain, PTB domain and leucine zipper (APPL1) regulates the protein level of EGFR by modulating its trafficking

    International Nuclear Information System (INIS)

    Lee, Jae-Rin; Hahn, Hwa-Sun; Kim, Young-Hoon; Nguyen, Hong-Hoa; Yang, Jun-Mo; Kang, Jong-Sun; Hahn, Myong-Joon

    2011-01-01

    Highlights: ► APPL1 regulates the protein level of EGFR in response to EGF stimulation. ► Depletion of APPL1 accelerates the movement of EGF/EGFR from the cell surface to the perinuclear region in response to EGF. ► Knockdown of APPL1 enhances the activity of Rab5. -- Abstract: The EGFR-mediated signaling pathway regulates multiple biological processes such as cell proliferation, survival and differentiation. Previously APPL1 (adaptor protein containing PH domain, PTB domain and leucine zipper 1) has been reported to function as a downstream effector of EGF-initiated signaling. Here we demonstrate that APPL1 regulates EGFR protein levels in response to EGF stimulation. Overexpression of APPL1 enhances EGFR stabilization while APPL1 depletion by siRNA reduces EGFR protein levels. APPL1 depletion accelerates EGFR internalization and movement of EGF/EGFR from cell surface to the perinuclear region in response to EGF treatment. Conversely, overexpression of APPL1 decelerates EGFR internalization and translocation of EGF/EGFR to the perinuclear region. Furthermore, APPL1 depletion enhances the activity of Rab5 which is involved in internalization and trafficking of EGFR and inhibition of Rab5 in APPL1-depleted cells restored EGFR levels. Consistently, APPL1 depletion reduced activation of Akt, the downstream signaling effector of EGFR and this is restored by inhibition of Rab5. These findings suggest that APPL1 is required for EGFR signaling by regulation of EGFR stabilities through inhibition of Rab5.

  8. Surface properties of aqueous amino acid solutions II. Leucine-leucine hydrochloride and leucine-sodium leucinate mixtures.

    Science.gov (United States)

    Matubayasi, Norihiro; Matsuyama, Shohei; Akizuki, Ryosuke

    2005-08-15

    To understand the distinction between the effects of zwitterionic, anionic, and cationic l-leucine upon adsorption and lateral interactions at air/water surface, the surface tensions of aqueous solutions of l-leucine-l-leucine hydrochloride and l-leucine-sodium l-leucinate mixtures were measured as a function of concentration and composition at 25 degrees C. The surface activity decreases in the order l-leucine >l-leucine hydrochloride > sodium l-leucinate. Both l-leucine hydrochloride and sodium l-leucinate form gaseous adsorbed films through the experimentally accessible concentration range, while the adsorbed film of zwitterionic l-leucine shows a transition between gaseous and expanded film.

  9. CONTRIBUTIONS TO CLASSIFICATION ZIPPERS USED IN INDUSTRY FOOTWEAR AND LEATHER GOODS

    Directory of Open Access Journals (Sweden)

    MALCOCI Marina

    2016-05-01

    Full Text Available Now customary accessory companies of all leather garments, zipper began to enjoy popularity only after 80 years of its invention. The first is considered the inventor of the zipper Elias Howe. Essentially involves fashion, change, innovation, originality, creativity and is defined as a succession of trends or fads, short. Create fashionable leather confections from home means accepting a contract with the producer and / or consumer, showing a profit motivating all at the right time. Continuous which require the exercise involves creative skills of fashion design, leading to a wide range of products. Current zipper is composed of: slider, teeth, strips shooter stops. Currently there is possibility to customize shooters to customer requirements, and even to form their own zippers. The present work presents the classification criteria zippers. They are after construction fastener after destination zippers, after the role they fulfill zippers, after the presentation at the procurement zippers, after finishing module of the metallic elements, as visibility zipper in the product, by type of teeth, by nature material strip zipper, after the type of materials they are made pullers, after the nature of the materials they are made of sliders and stops after the oxidation of metal components after finishing module teeth after shaped zipper, after slider type, by mode of ornamentation zippers. Knowing appearance zippers, elements of which it is composed, and their classification criteria allow us to correlate the shape of the product and destination.

  10. Kinematics analysis of a robotic zipper prototype for miniaturisation

    NARCIS (Netherlands)

    Baharom, M.Z.; Delbressine, F.L.M.; Feijs, L.M.G.

    2016-01-01

    This paper presents the mechanism and kinematics analysis of a robotic zipper prototype. This generic version of the robotic zipper is dedicated to automatically open and close the zipper which could be beneficial to elderly, people with physical disability, upper limb reduction deficiencies (ULRD),

  11. Expression analysis of OsbZIP transcription factors in resistance ...

    African Journals Online (AJOL)

    zino

    2013-08-21

    Aug 21, 2013 ... Plant basic leucine zipper (bZIP) proteins play an essential role in the genes expression and regulation in higher plants. They have been shown to regulate diverse plant specific phenomena, including germination, floral induction and development, seed maturation, photomorphogenesis, biotic and.

  12. A p300 and SIRT1 Regulated Acetylation Switch of C/EBPα Controls Mitochondrial Function

    NARCIS (Netherlands)

    Zaini, Mohamad A; Müller, Christine; de Jong, Tristan V; Ackermann, Tobias; Hartleben, Götz; Kortman, Gertrud; Gührs, Karl-Heinz; Fusetti, Fabrizia; Krämer, Oliver H; Guryev, Victor; Calkhoven, Cornelis F

    2018-01-01

    Cellular metabolism is a tightly controlled process in which the cell adapts fluxes through metabolic pathways in response to changes in nutrient supply. Among the transcription factors that regulate gene expression and thereby cause changes in cellular metabolism is the basic leucine-zipper (bZIP)

  13. Model of how plants sense zinc deficiency

    DEFF Research Database (Denmark)

    Assuncao, Ana G.L.; Persson, Daniel Olof; Husted, Søren

    2013-01-01

    to develop plant-based solutions addressing nutrient-use-efficiency and adaptation to nutrient-limited or -toxic soils. Recently two transcription factors of the bZIP family (basic-region leucine zipper) have been identified in Arabidopsis and shown to be pivotal in the adaptation response to zinc deficiency...

  14. In Planta Single-Molecule Pull-Down Reveals Tetrameric Stoichiometry of HD-ZIPIII:LITTLE ZIPPER Complexes.

    Science.gov (United States)

    Husbands, Aman Y; Aggarwal, Vasudha; Ha, Taekjip; Timmermans, Marja C P

    2016-08-01

    Deciphering complex biological processes markedly benefits from approaches that directly assess the underlying biomolecular interactions. Most commonly used approaches to monitor protein-protein interactions typically provide nonquantitative readouts that lack statistical power and do not yield information on the heterogeneity or stoichiometry of protein complexes. Single-molecule pull-down (SiMPull) uses single-molecule fluorescence detection to mitigate these disadvantages and can quantitatively interrogate interactions between proteins and other compounds, such as nucleic acids, small molecule ligands, and lipids. Here, we establish SiMPull in plants using the HOMEODOMAIN LEUCINE ZIPPER III (HD-ZIPIII) and LITTLE ZIPPER (ZPR) interaction as proof-of-principle. Colocalization analysis of fluorophore-tagged HD-ZIPIII and ZPR proteins provides strong statistical evidence of complex formation. In addition, we use SiMPull to directly quantify YFP and mCherry maturation probabilities, showing these differ substantially from values obtained in mammalian systems. Leveraging these probabilities, in conjunction with fluorophore photobleaching assays on over 2000 individual complexes, we determined HD-ZIPIII:ZPR stoichiometry. Intriguingly, these complexes appear as heterotetramers, comprising two HD-ZIPIII and two ZPR molecules, rather than heterodimers as described in the current model. This surprising result raises new questions about the regulation of these key developmental factors and is illustrative of the unique contribution SiMPull is poised to make to in planta protein interaction studies. © 2016 American Society of Plant Biologists. All rights reserved.

  15. Salt effects on hydrophobic interaction and charge screening in the folding of a negatively charged peptide to a coiled coil (leucine zipper).

    Science.gov (United States)

    Jelesarov, I; Dürr, E; Thomas, R M; Bosshard, H R

    1998-05-19

    The stability of a coiled coil or leucine zipper is controlled by hydrophobic interactions and electrostatic forces between the constituent helices. We have designed a 30-residue peptide with the repeating seven-residue pattern of a coiled coil, (abcdefg)n, and with Glu in positions e and g of each heptad. The glutamate side chains prevented folding at pH values above 6 because of electrostatic repulsion across the helix dimer interface as well as within the individual helices. Protonation of the carboxylates changed the conformation from a random coil monomer to a coiled coil dimer. Folding at alkaline pH where the peptide had a net charge of -7e was promoted by the addition of salts. The nature of the charge screening cation was less important than that of the anion. The high salt concentrations (>1 M) necessary to induce folding indicated that the salt-induced folding resulted from alterations in the protein-water interaction. Folding was promoted by the kosmotropic anions sulfate and fluoride and to a lesser extent by the weak kosmotrope formate, whereas chloride and the strong chaotrope perchlorate were ineffective. Kosmotropes are excluded from the protein surface, which is preferentially hydrated, and this promotes folding by strengthening hydrophobic interactions at the coiled coil interface. Although charge neutralization also contributed to folding, it was effective only when the screening cation was partnered by a good kosmotropic anion. Folding conformed to a two-state transition from random coil monomer to coiled coil dimer and was enthalpy driven and characterized by a change in the heat capacity of unfolding of 3.9 +/- 1.2 kJ mol-1 K-1. The rate of folding was analyzed by fluorescence stopped-flow measurements. Folding occurred in a biphasic reaction in which the rapid formation of an initial dimer (kf = 2 x 10(7) M-1 s-1) was followed by an equally rapid concentration-independent rearrangement to the folded dimer (k > 100 s-1).

  16. Novel and recurrent non-truncating mutations of the MITF basic domain: genotypic and phenotypic variations in Waardenburg and Tietz syndromes

    Science.gov (United States)

    Léger, Sandy; Balguerie, Xavier; Goldenberg, Alice; Drouin-Garraud, Valérie; Cabot, Annick; Amstutz-Montadert, Isabelle; Young, Paul; Joly, Pascal; Bodereau, Virginie; Holder-Espinasse, Muriel; Jamieson, Robyn V; Krause, Amanda; Chen, Hongsheng; Baumann, Clarisse; Nunes, Luis; Dollfus, Hélène; Goossens, Michel; Pingault, Véronique

    2012-01-01

    The microphthalmia-associated transcription factor (MITF) is a basic helix-loop-helix leucine zipper transcription factor, which regulates melanocyte development and the biosynthetic melanin pathway. A notable relationship has been described between non-truncating mutations of its basic domain and Tietz syndrome, which is characterized by albinoid-like hypopigmentation of the skin and hair, rather than the patchy depigmentation seen in Waardenburg syndrome, and severe hearing loss. Twelve patients with new or recurrent non-truncating mutations of the MITF basic domain from six families were enrolled in this study. We observed a wide range of phenotypes and some unexpected features. All the patients had blue irides and pigmentation abnormalities that ranged from diffuse hypopigmentation to Waardenburg-like patches. In addition, they showed congenital complete hearing loss, diffuse hypopigmentation of the skin, freckling and ocular abnormalities, more frequently than patients with MITF mutations outside the basic domain. In conclusion, the non-truncating mutations of the basic domain do not always lead to Tietz syndrome but rather to a large range of phenotypes. Sun-exposed freckles are interestingly observed more frequently in Asian populations. This variability argues for the possible interaction with modifier loci. PMID:22258527

  17. The homeodomain-leucine zipper (HD-Zip) class I transcription factors ATHB7 and ATHB12 modulate abscisic acid signalling by regulating protein phosphatase 2C and abscisic acid receptor gene activities.

    Science.gov (United States)

    Valdés, Ana Elisa; Overnäs, Elin; Johansson, Henrik; Rada-Iglesias, Alvaro; Engström, Peter

    2012-11-01

    Plants perceiving drought activate multiple responses to improve survival, including large-scale alterations in gene expression. This article reports on the roles in the drought response of two Arabidopsis thaliana homeodomain-leucine zipper class I genes; ATHB7 and ATHB12, both strongly induced by water-deficit and abscisic acid (ABA). ABA-mediated transcriptional regulation of both genes is shown to depend on the activity of protein phosphatases type 2C (PP2C). ATHB7 and ATHB12 are, thus, targets of the ABA signalling mechanism defined by the PP2Cs and the PYR/PYL family of ABA receptors, with which the PP2C proteins interact. Our results from chromatin immunoprecipitation and gene expression analyses demonstrate that ATHB7 and ATHB12 act as positive transcriptional regulators of PP2C genes, and thereby as negative regulators of abscisic acid signalling. In support of this notion, our results also show that ATHB7 and ATHB12 act to repress the transcription of genes encoding the ABA receptors PYL5 and PYL8 in response to an ABA stimulus. In summary, we demonstrate that ATHB7 and ATHB12 have essential functions in the primary response to drought, as mediators of a negative feedback effect on ABA signalling in the plant response to water deficit.

  18. SNARE zippering is hindered by polyphenols in the neuron

    Energy Technology Data Exchange (ETDEWEB)

    Yang, Yoosoo [Department of Genetic Engineering and Center for Human Interface Nanotechnology, Sungkyunkwan University, Suwon 440-746 (Korea, Republic of); Biomedical Research Institute, Korea Institute of Science and Technology, Seoul 136-791 (Korea, Republic of); Kim, Se-Hyun; Heo, Paul; Kong, Byoungjae; Shin, Jonghyeok; Jung, Young-Hun; Yoon, Keejung; Chung, Woo-Jae [Department of Genetic Engineering and Center for Human Interface Nanotechnology, Sungkyunkwan University, Suwon 440-746 (Korea, Republic of); Shin, Yeon-Kyun [Biomedical Research Institute, Korea Institute of Science and Technology, Seoul 136-791 (Korea, Republic of); Department of Biochemistry, Biophysics, and Molecular Biology, Iowa State University, Ames, IA 50011 (United States); Kweon, Dae-Hyuk, E-mail: dhkweon@skku.edu [Department of Genetic Engineering and Center for Human Interface Nanotechnology, Sungkyunkwan University, Suwon 440-746 (Korea, Republic of)

    2014-07-18

    Highlights: • Membrane fusion driven by SNARE complex is hindered by several polyphenols. • Distinctive inhibitory effect of each polyphenol on SNARE zippering in neuron was examined. • FRET between fluorescence protein-tagged SNAREs probed well SNARE zippering in PC12 cells. • Delphinidin and cyanidin inhibit N-terminal SNARE nucleation in Ca{sup 2+}-independent manner. • Myricetin inhibits Ca{sup 2+}-dependent transmembrane association of SNARE complex. - Abstract: Fusion of synaptic vesicles with the presynaptic plasma membrane in the neuron is mediated by soluble N-ethylmaleimide-sensitive fusion protein-attachment protein receptor (SNARE) proteins. SNARE complex formation is a zippering-like process which initiates at the N-terminus and proceeds to the C-terminal membrane-proximal region. Previously, we showed that this zippering-like process is regulated by several polyphenols, leading to the arrest of membrane fusion and the inhibition of neuroexocytosis. In vitro studies using purified SNARE proteins reconstituted in liposomes revealed that each polyphenol uniquely regulates SNARE zippering. However, the unique regulatory effect of each polyphenol in cells has not yet been examined. In the present study, we observed SNARE zippering in neuronal PC12 cells by measuring the fluorescence resonance energy transfer (FRET) changes of a cyan fluorescence protein (CFP) and a yellow fluorescence protein (YFP) fused to the N-termini or C-termini of SNARE proteins. We show that delphinidin and cyanidin inhibit the initial N-terminal nucleation of SNARE complex formation in a Ca{sup 2+}-independent manner, while myricetin inhibits Ca{sup 2+}-dependent transmembrane domain association of the SNARE complex in the cell. This result explains how polyphenols exhibit botulinum neurotoxin-like activity in vivo.

  19. A novel variable antibody fragment dimerized by leucine zippers with enhanced neutralizing potency against rabies virus G protein compared to its corresponding single-chain variable antibody fragment.

    Science.gov (United States)

    Li, Zhuang; Cheng, Yue; Xi, Hualong; Gu, Tiejun; Yuan, Ruosen; Chen, Xiaoxu; Jiang, Chunlai; Kong, Wei; Wu, Yongge

    2015-12-01

    Fatal rabies can be prevented effectively by post-exposure prophylactic (PEP) with rabies immunoglobulin (RIG). Single-chain variable fragments (scFv), which are composed of a variable heavy chain (VH) and a variable light chain (VL) connected by a peptide linker, can potentially be used to replace RIG. However, in our previous study, a scFv (scFV57S) specific for the rabies virus (RV) G protein showed a lower neutralizing potency than that of its parent IgG due to lower stability and altered peptide assembly pattern. In monoclonal antibodies, the VH and VL interact non-covalently, while in scFvs the VH is connected covalently with the VL by the artificial linker. In this study, we constructed and expressed two peptides 57VL-JUN-HIS and 57VH-FOS-HA in Escherichia coli. The well-known Fos and Jun leucine zippers were utilized to dimerize VH and VL similarly to the IgG counterpart. The two peptides assembled to form zipFv57S in vitro. Due to the greater similarity in structure with IgG, the zipFv57S protein showed a higher binding ability and affinity resulting in notable improvement of in vitro neutralizing activity over its corresponding scFv. The zipFv57S protein was also found to be more stable and showed similar protective rate as RIG in mice challenged with a lethal dose of RV. Our results not only indicated zipFv57S as an ideal alternative for RIG in PEP but also offered a novel and efficient hetero-dimerization pattern of VH and VL leading to enhanced neutralizing potency. Copyright © 2015. Published by Elsevier Ltd.

  20. Cliff : the automatized zipper

    NARCIS (Netherlands)

    Baharom, M.Z.; Toeters, M.J.; Delbressine, F.L.M.; Bangaru, C.; Feijs, L.M.G.

    2016-01-01

    It is our strong believe that fashion - more specifically apparel - can support us so much more in our daily life than it currently does. The Cliff project takes the opportunity to create a generic automatized zipper. It is a response to the struggle by elderly, people with physical disability, and

  1. Changes in leucine kinetics during meal absorption: effects of dietary leucine availability

    International Nuclear Information System (INIS)

    Nissen, S.; Haymond, M.W.

    1986-01-01

    Whole-body leucine and α/-ketoisocaproate (KIC) metabolism were estimated in mature dogs fed a complete meal, a meal devoid of branched-chain amino acids, and a meal devoid of all amino acids. Using a constant infusion of [4,5- 3 H]leucine and α-[1- 14 C]ketoisocaproate (KIC), combined with dietary [5,5,5- 2 H 3 ]leucine, the rate of whole-body proteolysis, protein synthesis, leucine oxidation, and interconversion leucine and KIC were estimated along with the rate of leucine absorption. Digestion of the complete meal resulted in a decrease in the rate of endogenous proteolysis, a small increase in the estimated rate of leucine entering protein, and a twofold increase in the rate of leucine oxidation. Ingestion of either the meal devoid of branched-chain amino acids or devoid of all amino acids resulted in a decrease in estimates of whole-body rates of proteolysis and protein synthesis, decreased leucine oxidation, and a decrease in the interconversion of leucine and KIC. The decrease in whole-body proteolysis was closely associated with the rise in plasma insulin concentrations following meal ingestion. Together these data suggest that the transition from tissue metabolism to anabolism is the result, at least in part, of decreased whole-body proteolysis. This meal-related decrease in proteolysis is independent of the dietary amino acid composition or content. In contrast, the rate of protein synthesis was sustained only when the meal complete in all amino acids was provided, indicating an overriding control of protein synthesis by amino acid availability

  2. CONTRIBUTIONS TO CLASSIFICATION ZIPPERS USED IN INDUSTRY FOOTWEAR AND LEATHER GOODS

    OpenAIRE

    MALCOCI Marina; PASCARI Ioana

    2016-01-01

    Now customary accessory companies of all leather garments, zipper began to enjoy popularity only after 80 years of its invention. The first is considered the inventor of the zipper Elias Howe. Essentially involves fashion, change, innovation, originality, creativity and is defined as a succession of trends or fads, short. Create fashionable leather confections from home means accepting a contract with the producer and / or consumer, showing a profit motivating all at the right time. Continuou...

  3. GenBank blastx search result: AK119754 [KOME

    Lifescience Database Archive (English)

    Full Text Available linkage group 20 Contains the gene for a novel protein similar to vertebrate C-Fos protein family, the gene ...for a novel protein similar to vertebrate jun dimerization protein 2, the gene for a novel protein similar to vertebrate... FOS-like antigen 2 protein (FOSL2), the gene for a novel protein similar to vertebrate... basic leucine zipper transcription factor ATF-like, the gene for a novel protein similar to vertebrate

  4. ATF5 polymorphisms influence ATF function and response to treatment in children with childhood acute lymphoblastic leukemia

    OpenAIRE

    Rousseau, Julie; Gagné, Vincent; Labuda, Malgorzata; Beaubois, Cyrielle; Sinnett, Daniel; Laverdière, Caroline; Moghrabi, Albert; Sallan, Stephen E.; Silverman, Lewis B.; Neuberg, Donna; Kutok, Jeffery L.; Krajinovic, Maja

    2011-01-01

    Asparaginase is a standard and critical component in the therapy of childhood acute lymphoblastic leukemia. Asparagine synthetase (ASNS) and the basic region leucine zipper activating transcription factor 5 (ATF5) and arginosuccinate synthase 1 (ASS1) have been shown to mediate the antileukemic effect of asparaginase and to display variable expression between leukemia cells that are resistant and sensitive to treatment. Fourteen polymorphisms in the regulatory and coding regions of these gene...

  5. Leucine aminopeptidase blood test

    Science.gov (United States)

    ... medlineplus.gov/ency/article/003559.htm Leucine aminopeptidase blood test To use the sharing features on this page, ... Alternative Names Serum leucine aminopeptidase; LAP - serum Images Blood test References Chernecky CC, Berger BJ. Leucine aminopeptidase (LAP) - ...

  6. Zipper plot: visualizing transcriptional activity of genomic regions.

    Science.gov (United States)

    Avila Cobos, Francisco; Anckaert, Jasper; Volders, Pieter-Jan; Everaert, Celine; Rombaut, Dries; Vandesompele, Jo; De Preter, Katleen; Mestdagh, Pieter

    2017-05-02

    Reconstructing transcript models from RNA-sequencing (RNA-seq) data and establishing these as independent transcriptional units can be a challenging task. Current state-of-the-art tools for long non-coding RNA (lncRNA) annotation are mainly based on evolutionary constraints, which may result in false negatives due to the overall limited conservation of lncRNAs. To tackle this problem we have developed the Zipper plot, a novel visualization and analysis method that enables users to simultaneously interrogate thousands of human putative transcription start sites (TSSs) in relation to various features that are indicative for transcriptional activity. These include publicly available CAGE-sequencing, ChIP-sequencing and DNase-sequencing datasets. Our method only requires three tab-separated fields (chromosome, genomic coordinate of the TSS and strand) as input and generates a report that includes a detailed summary table, a Zipper plot and several statistics derived from this plot. Using the Zipper plot, we found evidence of transcription for a set of well-characterized lncRNAs and observed that fewer mono-exonic lncRNAs have CAGE peaks overlapping with their TSSs compared to multi-exonic lncRNAs. Using publicly available RNA-seq data, we found more than one hundred cases where junction reads connected protein-coding gene exons with a downstream mono-exonic lncRNA, revealing the need for a careful evaluation of lncRNA 5'-boundaries. Our method is implemented using the statistical programming language R and is freely available as a webtool.

  7. Transcriptional Activation Domains of the Candida albicans Gcn4p and Gal4p Homologs▿ †

    OpenAIRE

    Martchenko, Mikhail; Levitin, Anastasia; Whiteway, Malcolm

    2006-01-01

    Many putative transcription factors in the pathogenic fungus Candida albicans contain sequence similarity to well-defined transcriptional regulators in the budding yeast Saccharomyces cerevisiae, but this sequence similarity is often limited to the DNA binding domains of the molecules. The Gcn4p and Gal4p proteins of Saccharomyces cerevisiae are highly studied and well-understood eukaryotic transcription factors of the basic leucine zipper (Gcn4p) and C6 zinc cluster (Gal4p) families; C. albi...

  8. The leucine-rich repeat structure.

    Science.gov (United States)

    Bella, J; Hindle, K L; McEwan, P A; Lovell, S C

    2008-08-01

    The leucine-rich repeat is a widespread structural motif of 20-30 amino acids with a characteristic repetitive sequence pattern rich in leucines. Leucine-rich repeat domains are built from tandems of two or more repeats and form curved solenoid structures that are particularly suitable for protein-protein interactions. Thousands of protein sequences containing leucine-rich repeats have been identified by automatic annotation methods. Three-dimensional structures of leucine-rich repeat domains determined to date reveal a degree of structural variability that translates into the considerable functional versatility of this protein superfamily. As the essential structural principles become well established, the leucine-rich repeat architecture is emerging as an attractive framework for structural prediction and protein engineering. This review presents an update of the current understanding of leucine-rich repeat structure at the primary, secondary, tertiary and quaternary levels and discusses specific examples from recently determined three-dimensional structures.

  9. Molecular characterisation of two novel maize LRR receptor-like kinases, which belong to the SERK gene family

    NARCIS (Netherlands)

    Baudino, S.; Hansen, S.; Brettschneider, R.; Hecht, V.F.G.; Dresselhaus, T.; Lörz, H.; Dumas, C.; Rogowsky, P.M.

    2001-01-01

    Genes encoding two novel members of the leucine-rich repeat receptor-like kinase (LRR-RLK) superfamily have been isolated from maize (Zea mays L.). These genes have been named ZmSERK1 and ZmSERK2 since features such as a putative leucine zipper (ZIP) and five leucine rich repeats in the

  10. Isolation and expression analysis of EcbZIP17 from different finger millet genotypes shows conserved nature of the gene.

    Science.gov (United States)

    Chopperla, Ramakrishna; Singh, Sonam; Mohanty, Sasmita; Reddy, Nanja; Padaria, Jasdeep C; Solanke, Amolkumar U

    2017-10-01

    Basic leucine zipper (bZIP) transcription factors comprise one of the largest gene families in plants. They play a key role in almost every aspect of plant growth and development and also in biotic and abiotic stress tolerance. In this study, we report isolation and characterization of EcbZIP17 , a group B bZIP transcription factor from a climate smart cereal, finger millet ( Eleusine coracana L.). The genomic sequence of EcbZIP17 is 2662 bp long encompassing two exons and one intron with ORF of 1722 bp and peptide length of 573 aa. This gene is homologous to AtbZIP17 ( Arabidopsis ), ZmbZIP17 (maize) and OsbZIP60 (rice) which play a key role in endoplasmic reticulum (ER) stress pathway. In silico analysis confirmed the presence of basic leucine zipper (bZIP) and transmembrane (TM) domains in the EcbZIP17 protein. Allele mining of this gene in 16 different genotypes by Sanger sequencing revealed no variation in nucleotide sequence, including the 618 bp long intron. Expression analysis of EcbZIP17 under heat stress exhibited similar pattern of expression in all the genotypes across time intervals with highest upregulation after 4 h. The present study established the conserved nature of EcbZIP17 at nucleotide and expression level.

  11. ABI-like transcription factor gene TaABL1 from wheat improves multiple abiotic stress tolerances in transgenic plants.

    Science.gov (United States)

    Xu, Dong-Bei; Gao, Shi-Qing; Ma, You-Zhi; Xu, Zhao-Shi; Zhao, Chang-Ping; Tang, Yi-Miao; Li, Xue-Yin; Li, Lian-Cheng; Chen, Yao-Feng; Chen, Ming

    2014-12-01

    The phytohormone abscisic acid (ABA) plays crucial roles in adaptive responses of plants to abiotic stresses. ABA-responsive element binding proteins (AREBs) are basic leucine zipper transcription factors that regulate the expression of downstream genes containing ABA-responsive elements (ABREs) in promoter regions. A novel ABI-like (ABA-insensitive) transcription factor gene, named TaABL1, containing a conserved basic leucine zipper (bZIP) domain was cloned from wheat. Southern blotting showed that three copies were present in the wheat genome. Phylogenetic analyses indicated that TaABL1 belonged to the AREB subfamily of the bZIP transcription factor family and was most closely related to ZmABI5 in maize and OsAREB2 in rice. Expression of TaABL1 was highly induced in wheat roots, stems, and leaves by ABA, drought, high salt, and low temperature stresses. TaABL1 was localized inside the nuclei of transformed wheat mesophyll protoplast. Overexpression of TaABL1 enhanced responses of transgenic plants to ABA and hastened stomatal closure under stress, thereby improving tolerance to multiple abiotic stresses. Furthermore, overexpression of TaABL1 upregulated or downregulated the expression of some stress-related genes controlling stomatal closure in transgenic plants under ABA and drought stress conditions, suggesting that TaABL1 might be a valuable genetic resource for transgenic molecular breeding.

  12. Temporary abdominal closure with zipper-mesh device for management of intra-abdominal sepsis

    Directory of Open Access Journals (Sweden)

    Edivaldo Massazo Utiyama

    Full Text Available OBJECTIVE: to present our experience with scheduled reoperations in 15 patients with intra-abdominal sepsis. METHODS: we have applied a more effective technique consisting of temporary abdominal closure with a nylon mesh sheet containing a zipper. We performed reoperations in the operating room under general anesthesia at an average interval of 84 hours. The revision consisted of debridement of necrotic material and vigorous lavage of the involved peritoneal area. The mean age of patients was 38.7 years (range, 15 to 72 years; 11 patients were male, and four were female. RESULTS: forty percent of infections were due to necrotizing pancreatitis. Sixty percent were due to perforation of the intestinal viscus secondary to inflammation, vascular occlusion or trauma. We performed a total of 48 reoperations, an average of 3.2 surgeries per patient. The mesh-zipper device was left in place for an average of 13 days. An intestinal ostomy was present adjacent to the zipper in four patients and did not present a problem for patient management. Mortality was 26.6%. No fistulas resulted from this technique. When intra-abdominal disease was under control, the mesh-zipper device was removed, and the fascia was closed in all patients. In three patients, the wound was closed primarily, and in 12 it was allowed to close by secondary intent. Two patients developed hernia; one was incisional and one was in the drain incision. CONCLUSION: the planned reoperation for manual lavage and debridement of the abdomen through a nylon mesh-zipper combination was rapid, simple, and well-tolerated. It permitted effective management of severe septic peritonitis, easy wound care and primary closure of the abdominal wall.

  13. Syntheses and Self-assembling Behaviors of Pentagonal Conjugates of Tryptophane Zipper-Forming Peptide

    Directory of Open Access Journals (Sweden)

    Nobuo Kimizuka

    2011-08-01

    Full Text Available Pentagonal conjugates of tryptophane zipper-forming peptide (CKTWTWTE with a pentaazacyclopentadecane core (Pentagonal-Gly-Trpzip and Pentagonal-Ala-Trpzip were synthesized and their self-assembling behaviors were investigated in water. Pentagonal-Gly-Trpzip self-assembled into nanofibers with the width of about 5 nm in neutral water (pH 7 via formation of tryptophane zipper, which irreversibly converted to nanoribbons by heating. In contrast, Pentagonal-Ala-Trpzip formed irregular aggregates in water.

  14. The HBZ gene, a key player in HTLV-1 pathogenesis

    Directory of Open Access Journals (Sweden)

    Green Patrick L

    2009-08-01

    Full Text Available Abstract Human T-cell leukemia virus type 1 (HTLV-1 causes adult T-cell leukemia/lymphoma (ATL and is also associated with a variety of lymphocyte-mediated diseases. The HTLV-1 basic leucine zipper (HBZ gene, found to be consistently expressed in ATL, has recently been the subject of intensive research efforts. In this review, we summarize recent findings about HBZ and discuss its roles and functions not only in the virus life cycle, but also in HTLV-1 disease pathogenesis.

  15. Hybrids of the bHLH and bZIP protein motifs display different DNA-binding activities in vivo vs. in vitro.

    Directory of Open Access Journals (Sweden)

    Hiu-Kwan Chow

    Full Text Available Minimalist hybrids comprising the DNA-binding domain of bHLH/PAS (basic-helix-loop-helix/Per-Arnt-Sim protein Arnt fused to the leucine zipper (LZ dimerization domain from bZIP (basic region-leucine zipper protein C/EBP were designed to bind the E-box DNA site, CACGTG, targeted by bHLHZ (basic-helix-loop-helix-zipper proteins Myc and Max, as well as the Arnt homodimer. The bHLHZ-like structure of ArntbHLH-C/EBP comprises the Arnt bHLH domain fused to the C/EBP LZ: i.e. swap of the 330 aa PAS domain for the 29 aa LZ. In the yeast one-hybrid assay (Y1H, transcriptional activation from the E-box was strong by ArntbHLH-C/EBP, and undetectable for the truncated ArntbHLH (PAS removed, as detected via readout from the HIS3 and lacZ reporters. In contrast, fluorescence anisotropy titrations showed affinities for the E-box with ArntbHLH-C/EBP and ArntbHLH comparable to other transcription factors (K(d 148.9 nM and 40.2 nM, respectively, but only under select conditions that maintained folded protein. Although in vivo yeast results and in vitro spectroscopic studies for ArntbHLH-C/EBP targeting the E-box correlate well, the same does not hold for ArntbHLH. As circular dichroism confirms that ArntbHLH-C/EBP is a much more strongly alpha-helical structure than ArntbHLH, we conclude that the nonfunctional ArntbHLH in the Y1H must be due to misfolding, leading to the false negative that this protein is incapable of targeting the E-box. Many experiments, including protein design and selections from large libraries, depend on protein domains remaining well-behaved in the nonnative experimental environment, especially small motifs like the bHLH (60-70 aa. Interestingly, a short helical LZ can serve as a folding- and/or solubility-enhancing tag, an important device given the focus of current research on exploration of vast networks of biomolecular interactions.

  16. Syntheses of DL-[2-13C]leucine and its use in the preparation of [3-DL-[2-13C]leucine]oxytocin and [8-DL-[2-13C]leucine]oxytocin

    International Nuclear Information System (INIS)

    Viswanatha, V.; Larsen, B.; Hruby, V.J.

    1979-01-01

    DL-[2- 13 C]Leucine was prepared by condensing the sodium salt of ethyl acetamido-[2- 13 C]cyanoacetate with isobutylbromide in hexamethylphosphoroustriamide followed by acid hydrolysis. N-BOC-DL-[2- 13 C]Leucine was prepared and incorporated into [8-DL-[2- 13 C]leucine]oxytocin by total synthesis. The 13 C-labeled hormone derivative [8-[2- 13 C]leucine]oxytocin was separated from its 8-position diastereoisomer by partition chromatography. The specifically 13 C-labeled peptide hormone diastereoisomeric analog [3-DL-[2- 13 C]leucine]oxytocin also was prepared by solid phase peptide synthesis. No suitable solvent system for partition chromatography separation of the latter diastereoisomeric peptide mixture could be found. However an excellent preparative separation of the diastereoisomers could be obtained by reverse phase high pressure liquid chromatography on a partisil 10 M9 ODS column using the solvent system 0.05 M ammonium acetate (pH 4.0), acetonitrile (81:19, v/v) to give pure [3-[2- 13 C]leucine]oxytocin and [3-D-[2- 13 C]leucine]oxytocin. An excellent separation of [8-[2- 13 C]leucine]oxytocin and the corresponding delata-D-leucine diastereoisomer derivative could also be accomplished by high pressure liquid chromatography. (author)

  17. Complexity of CNC transcription factors as revealed by gene targeting of the Nrf3 locus.

    Science.gov (United States)

    Derjuga, Anna; Gourley, Tania S; Holm, Teresa M; Heng, Henry H Q; Shivdasani, Ramesh A; Ahmed, Rafi; Andrews, Nancy C; Blank, Volker

    2004-04-01

    Cap'n'collar (CNC) family basic leucine zipper transcription factors play crucial roles in the regulation of mammalian gene expression and development. To determine the in vivo function of the CNC protein Nrf3 (NF-E2-related factor 3), we generated mice deficient in this transcription factor. We performed targeted disruption of two Nrf3 exons coding for CNC homology, basic DNA-binding, and leucine zipper dimerization domains. Nrf3 null mice developed normally and revealed no obvious phenotypic differences compared to wild-type animals. Nrf3(-/-) mice were fertile, and gross anatomy as well as behavior appeared normal. The mice showed normal age progression and did not show any apparent additional phenotype during their life span. We observed no differences in various blood parameters and chemistry values. We infected wild-type and Nrf3(-/-) mice with acute lymphocytic choriomeningitis virus and found no differences in these animals with respect to their number of virus-specific CD8 and CD4 T cells as well as their B-lymphocyte response. To determine whether the mild phenotype of Nrf3 null animals is due to functional redundancy, we generated mice deficient in multiple CNC factors. Contrary to our expectations, an absence of Nrf3 does not seem to cause additional lethality in compound Nrf3(-/-)/Nrf2(-/-) and Nrf3(-/-)/p45(-/-) mice. We hypothesize that the role of Nrf3 in vivo may become apparent only after appropriate challenge to the mice.

  18. OsbZIP58, a basic leucine zipper transcription factor, regulates starch biosynthesis in rice endosperm.

    Science.gov (United States)

    Wang, Jie-Chen; Xu, Heng; Zhu, Ying; Liu, Qiao-Quan; Cai, Xiu-Ling

    2013-08-01

    Starch composition and the amount in endosperm, both of which contribute dramatically to seed yield, cooking quality, and taste in cereals, are determined by a series of complex biochemical reactions. However, the mechanism regulating starch biosynthesis in cereal seeds is not well understood. This study showed that OsbZIP58, a bZIP transcription factor, is a key transcriptional regulator controlling starch synthesis in rice endosperm. OsbZIP58 was expressed mainly in endosperm during active starch synthesis. osbzip58 null mutants displayed abnormal seed morphology with altered starch accumulation in the white belly region and decreased amounts of total starch and amylose. Moreover, osbzip58 had a higher proportion of short chains and a lower proportion of intermediate chains of amylopectin. Furthermore, OsbZIP58 was shown to bind directly to the promoters of six starch-synthesizing genes, OsAGPL3, Wx, OsSSIIa, SBE1, OsBEIIb, and ISA2, and to regulate their expression. These findings indicate that OsbZIP58 functions as a key regulator of starch synthesis in rice seeds and provide new insights into seed quality control.

  19. Genome wide identification and expression analysis of Homeodomain leucine zipper subfamily IV (HDZ IV gene family from Musa accuminata

    Directory of Open Access Journals (Sweden)

    Ashutosh ePandey

    2016-02-01

    Full Text Available The homedodomain zipper family (HD-ZIP of transcription factors is present only in plants and plays important role in the regulation of plant-specific processes. The subfamily IV of HDZ transcription factors (HD-ZIP IV has primarily been implicated in the regulation of epidermal structure development. Though this gene family is present in all lineages of land plants, members of this gene family have not been identified in banana, which is one of the major staple fruit crops. In the present work, we identified 21 HDZIV genes in banana by the computational analysis of banana genome resource. Our analysis suggested that these genes putatively encode proteins having all the characteristic domains of HDZIV transcription factors. The phylogenetic analysis of the banana HDZIV family genes further confirmed that after separation from a common ancestor, the banana and poales lineages might have followed distinct evolutionary paths. Further, we conclude that segmental duplication played a major role in the evolution of banana HDZIV genes. All the identified banana HDZIV genes expresses in different banana tissue, however at varying levels. The transcript levels of some of the banana HDZIV genes were also detected in banana fruit pulp, suggesting their putative role in fruit attributes. A large number of genes of this family showed modulated expression under drought and salinity stress. Taken together, the present work lays a foundation for elucidation of functional aspects of the banana HDZIV genes and for their possible use in the banana improvement programs.

  20. The Single Needle Lockstitch Machine. [Setting Zippers.] Module 8.

    Science.gov (United States)

    South Carolina State Dept. of Education, Columbia. Office of Vocational Education.

    This module on setting zippers, one in a series on the single needle lockstitch sewing machine for student self-study, contains five sections. Each section includes the following parts: an introduction, directions, an objective, learning activities, student information, student self-check, check-out activities, and an instructor's final checklist.…

  1. Resistance of Type 5 chemical protective clothing against nanometric airborne particles: Behavior of seams and zipper.

    Science.gov (United States)

    Vinches, Ludwig; Hallé, Stéphane

    2017-12-01

    In the field of dermal protection, the use of chemical protective clothing (CPC) (including coveralls) are considered as the last barrier against airborne engineered nanomaterials (ENM). In the majority of cases, Type 5 CPC, used against solid particles (ISO 13982-1), perform well against ENM. But in a recent study, a penetration level (PL) of up to 8.5% of polydisperse sodium chloride airborne nanoparticles has been measured. Moreover, in all the previous studies, tests were performed on a sample of protective clothing material without seams or zippers. Thus, the potential for permeation through a zipper or seams has not yet been determined, even though these areas would be privileged entry points for airborne ENM. This work was designed to evaluate the PL of airborne ENM through coveralls and specifically the PL through the seams on different parts of the CPC and the zipper. Eight current models of CPC (Type 5) were selected. The samples were taken from places with and without seams and with a zipper. In some cases, a cover strip can be added to the zipper to enhance its sealing. Polydisperse nanoparticles were generated by nebulization of a sodium chloride solution. A penetration cell was developed to expose the sample to airborne nanometric particles. The NaCl particle concentration in number was measured with an ultrafine particle counter and the PL was defined as the downstream concentration divided by the upstream concentration. The results obtained show that the PL increased significantly in the presence of seams and could reach up to 90% depending on the seam's design. Moreover, this study classifies the different types of seams by their resistance against airborne ENM. As for the penetration of airborne NaCl particles through the zipper, the PL was greatly attenuated by the presence of a cover strip, but only for certain models of coveralls. Finally, the values of the pressure drop were directly linked to the type of seam. All of these conclusions provide

  2. [3H]Leucine incorporation method as a tool to measure secondary production by periphytic bacteria associated to the roots of floating aquatic macrophyte.

    Science.gov (United States)

    Miranda, M R; Guimarães, J R D; Coelho-Souza, A S

    2007-10-01

    The present study assessed the application of [(3)H]Leucine incorporation into protein by periphytic bacteria associated with the roots of the floating aquatic macrophyte Eichornia crassipes. Basic assumptions underlying the method, such as linearity of leucine incorporation, saturation level of incorporation rates, incorporation into other macromolecules, specificity of incorporation for bacterial assemblages and [(3)H]Leucine degradation during samples storage were tested, and two procedures for extracting the incorporated leucine were compared. Both methods gave the same results, however, the hot TCA extraction method was less time consuming than the alkaline extraction method. Incorporation of [(3)H]Leucine was linear for up to 40 min. Saturation concentration of [(3)H]Leucine incorporation into protein was 1500 nM. An experiment with prokaryotic and eukaryotic inhibitors showed no significant [(3)H]Leucine incorporation into eukaryotes even in high leucine concentrations. No significant amounts of radiolabel were incorporated into other macromolecules. The maximum time of sample storage after the incubation is 15 days. The leucine incorporation method can be a reliable tool to measure bacterial production in the periphyton root-associated bacteria.

  3. N-terminal segments modulate the α-helical propensities of the intrinsically disordered basic regions of bZIP proteins.

    Science.gov (United States)

    Das, Rahul K; Crick, Scott L; Pappu, Rohit V

    2012-02-17

    Basic region leucine zippers (bZIPs) are modular transcription factors that play key roles in eukaryotic gene regulation. The basic regions of bZIPs (bZIP-bRs) are necessary and sufficient for DNA binding and specificity. Bioinformatic predictions and spectroscopic studies suggest that unbound monomeric bZIP-bRs are uniformly disordered as isolated domains. Here, we test this assumption through a comparative characterization of conformational ensembles for 15 different bZIP-bRs using a combination of atomistic simulations and circular dichroism measurements. We find that bZIP-bRs have quantifiable preferences for α-helical conformations in their unbound monomeric forms. This helicity varies from one bZIP-bR to another despite a significant sequence similarity of the DNA binding motifs (DBMs). Our analysis reveals that intramolecular interactions between DBMs and eight-residue segments directly N-terminal to DBMs are the primary modulators of bZIP-bR helicities. We test the accuracy of this inference by designing chimeras of bZIP-bRs to have either increased or decreased overall helicities. Our results yield quantitative insights regarding the relationship between sequence and the degree of intrinsic disorder within bZIP-bRs, and might have general implications for other intrinsically disordered proteins. Understanding how natural sequence variations lead to modulation of disorder is likely to be important for understanding the evolution of specificity in molecular recognition through intrinsically disordered regions (IDRs). Copyright © 2011 Elsevier Ltd. All rights reserved.

  4. MicroRNA-27a Is Induced by Leucine and Contributes to Leucine-Induced Proliferation Promotion in C2C12 Cells

    Directory of Open Access Journals (Sweden)

    Guangmang Liu

    2013-07-01

    Full Text Available Leucine, a branched chain amino acid, is well known to stimulate protein synthesis in skeletal muscle. However, the role of leucine in myoblast proliferation remains unclear. In this study, we found that leucine could promote proliferation of C2C12 cells. Moreover, expressions of miR-27a and myostatin (a bona fide target of miR-27a were upregulated and downregulated, respectively, following leucine treatment. We also found that miR-27a loss-of-function by transfection of a miR-27a inhibitor suppressed the promotion of myoblast proliferation caused by leucine. Our results suggest that miR-27a is induced by leucine and contributes to leucine-induced proliferation promotion of myoblast.

  5. Leucine metabolism in patients with Hepatic Encephalopathy

    International Nuclear Information System (INIS)

    McGhee, A.S.; Kassouny, M.E.; Matthews, D.E.; Millikan, W.

    1986-01-01

    A primed continuous infusion of [ 15 N, 1- 13 C]leucine was used to determine whether increased oxidation and/or protein synthesis of leucine occurs in patients with cirrhosis. Five controls and patients were equilibrated on a metabolic balance diet [0.6 g protein per kg ideal body weight (IBW)]. An additional four patients were equilibrated in the same manner with the same type of diet with a protein level of 0.75 g per kg IBW. Plasma leucine and breath CO 2 enrichments were measured by mass spectrometry. Protein synthesis and leucine metabolism were identical in controls and patients when both were fed a diet with 0.6 g protein/kg IBW. Results indicate that systemic derangements of leucine metabolism are not the cause of Hepatic Encephalopathy

  6. Relationship between plasma and tissue parameters of leucine metabolism

    International Nuclear Information System (INIS)

    Vazquez, J.A.; Paul, H.S.; Adibi, S.A.

    1986-01-01

    Using a primed continuous infusion of [1- 14 C] leucine, the authors investigated parameters of leucine metabolism in plasma, expired air, and tissues of fed and 48-hour starved rats. The ratios of muscle/plasma specific activity of α-ketoisocaproate (KIC) in fed and starved rats, respectively were not significantly different from one (1.07 +/- 0.14 and 0.97 +/- 0.10, mean +/- SE, 7 rats). The ratio of muscle/plasma specific activity of leucine was also not significantly different from one (0.86 +/- 0.11) in fed rats, but was significantly lower than one (0.80 +/- 0.07) in starved rats. The rate of leucine oxidation was approximately 32% higher when calculated based on plasma KIC rather than leucine specific activity. However, starvation significantly increased the rate of leucine oxidation with either specific activity. The rate of leucine incorporation into whole body protein was unaffected by starvation (32.7 +/- 3.5 vs 36.1 +/- 2.5 μmol/100 g/h), but the incorporation into total protein of liver (1350 +/- 140 vs 780 +/- 33 nmol) and of skeletal muscle (1940 +/- 220 vs 820 +/- 60 nmol) was significantly decreased. The authors conclude that a) leucine or KIC specific activity in muscle is better predicted by plasma KIC than leucine specific activity, and b) the tracer infusion technique is valid for the study of leucine oxidation but not for leucine incorporation into whole body protein

  7. Protein and leucine metabolism in maple syrup urine disease

    International Nuclear Information System (INIS)

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

    1990-01-01

    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

  8. Comparative analysis of pharmacological treatments with N-acetyl-DL-leucine (Tanganil) and its two isomers (N-acetyl-L-leucine and N-acetyl-D-leucine) on vestibular compensation: Behavioral investigation in the cat.

    Science.gov (United States)

    Tighilet, Brahim; Leonard, Jacques; Bernard-Demanze, Laurence; Lacour, Michel

    2015-12-15

    Head roll tilt, postural imbalance and spontaneous nystagmus are the main static vestibular deficits observed after an acute unilateral vestibular loss (UVL). In the UVL cat model, these deficits are fully compensated over 6 weeks as the result of central vestibular compensation. N-Acetyl-dl-leucine is a drug prescribed in clinical practice for the symptomatic treatment of acute UVL patients. The present study investigated the effects of N-acetyl-dl-leucine on the behavioral recovery after unilateral vestibular neurectomy (UVN) in the cat, and compared the effects of each of its two isomers N-acetyl-L-leucine and N-acetyl-D-leucine. Efficacy of these three drug treatments has been evaluated with respect to a placebo group (UVN+saline water) on the global sensorimotor activity (observation grids), the posture control (support surface measurement), the locomotor balance (maximum performance at the rotating beam test), and the spontaneous vestibular nystagmus (recorded in the light). Whatever the parameters tested, the behavioral recovery was strongly and significantly accelerated under pharmacological treatments with N-acetyl-dl-leucine and N-acetyl-L-leucine. In contrast, the N-acetyl-D-leucine isomer had no effect at all on the behavioral recovery, and animals of this group showed the same recovery profile as those receiving a placebo. It is concluded that the N-acetyl-L-leucine isomer is the active part of the racemate component since it induces a significant acceleration of the vestibular compensation process similar (and even better) to that observed under treatment with the racemate component only. Copyright © 2015 Elsevier B.V. All rights reserved.

  9. DNA hybridization kinetics: zippering, internal displacement and sequence dependence.

    Science.gov (United States)

    Ouldridge, Thomas E; Sulc, Petr; Romano, Flavio; Doye, Jonathan P K; Louis, Ard A

    2013-10-01

    Although the thermodynamics of DNA hybridization is generally well established, the kinetics of this classic transition is less well understood. Providing such understanding has new urgency because DNA nanotechnology often depends critically on binding rates. Here, we explore DNA oligomer hybridization kinetics using a coarse-grained model. Strand association proceeds through a complex set of intermediate states, with successful binding events initiated by a few metastable base-pairing interactions, followed by zippering of the remaining bonds. But despite reasonably strong interstrand interactions, initial contacts frequently dissociate because typical configurations in which they form differ from typical states of similar enthalpy in the double-stranded equilibrium ensemble. Initial contacts must be stabilized by two or three base pairs before full zippering is likely, resulting in negative effective activation enthalpies. Non-Arrhenius behavior arises because the number of base pairs required for nucleation increases with temperature. In addition, we observe two alternative pathways-pseudoknot and inchworm internal displacement-through which misaligned duplexes can rearrange to form duplexes. These pathways accelerate hybridization. Our results explain why experimentally observed association rates of GC-rich oligomers are higher than rates of AT- rich equivalents, and more generally demonstrate how association rates can be modulated by sequence choice.

  10. Structural and dynamic studies of the dimerization and DNA-binding domains of the transcription factors v-Myc and Max

    International Nuclear Information System (INIS)

    Fieber, W.

    2001-05-01

    In the present work, solution structural and dynamic properties of the dimerization and DNA binding domains of the transcription factors v-Myc and Max were characterized by NMR and CD spectroscopy. It could be demonstrated that v-Myc in the absence of its authentic binding partner Max does not homodimerize, but exists in a monomeric and prestructured form. Two separated α-helical regions in the leucine zipper region and in the basic-H1 region, respectively, could be identified, while the latter appeared to be less stable. Both helices lack stabilizing tertiary side chain interactions and represent exceptional examples for loosely coupled, structured segments in a native protein. The structure of v-Myc is dynamic and can be described as a distribution of conformational substates. Motion within the substates comprise fast (picosecond to nanosecond) local backbone fluctuations like helical fraying, whereas motion between the substates comprise the relative orientation of the two helices and occur at larger time scales (microsecond to millisecond). The preformation of the specific protein and DNA binding sites, leucine zipper and the basic region, presumably allows rapid and accurate recognition of the respective binding partners. v-Myc-Max and Max-Max protein preparations were shown to form stable dimers. Thermodynamic analysis of the dissociation reactions of v-Myc-Max revealed a significant higher stability of the heterodimer than of the Max-Max homodimer over the whole temperature range. It could be demonstrated that the restricted conformational space of the v-Myc bHLHZip domain reduces the entropy penalty associated with dimerization and contributes to the preference of Max to form heterodimers with v-Myc rather than homodimers. (author)

  11. Leucine stimulation of skeletal muscle protein synthesis

    International Nuclear Information System (INIS)

    Layman, D.K.; Grogan, C.K.

    1986-01-01

    Previous work in this laboratory has demonstrated a stimulatory effect of leucine on skeletal muscle protein synthesis measured in vitro during catabolic conditions. Studies in other laboratories have consistently found this effect in diaphragm muscle, however, studies examining effects on nitrogen balance or with in vivo protein synthesis in skeletal muscle are equivocal. This experiment was designed to determine the potential of leucine to stimulate skeletal muscle protein synthesis in vivo. Male Sprague-Dawley rats weighing 200 g were fasted for 12 hrs, anesthetized, a jugular cannula inserted, and protein synthesis measured using a primed continuous infusion of 14 C-tyrosine. A plateau in specific activity was reached after 30 to 60 min and maintained for 3 hrs. The leucine dose consisted of a 240 umole priming dose followed by a continuous infusion of 160 umoles/hr. Leucine infusion stimulated protein synthesis in the soleus muscle (28%) and in the red (28%) and white portions (12%) of the gastrocnemius muscle compared with controls infused with only tyrosine. The increased rates of protein synthesis were due to increased incorporation of tyrosine into protein and to decreased specific activity of the free tyrosine pool. These data indicate that infusion of leucine has the potential to stimulate in vivo protein synthesis in skeletal muscles

  12. Efficacy and Safety of Leucine Supplementation in the Elderly.

    Science.gov (United States)

    Borack, Michael S; Volpi, Elena

    2016-12-01

    Leucine supplementation has grown in popularity due to the discovery of its anabolic effects on cell signaling and protein synthesis in muscle. The current recommendation is a minimum intake of 55 mg ⋅ kg -1 . d -1 Leucine acutely stimulates skeletal muscle anabolism and can overcome the anabolic resistance of aging. The value of chronic leucine ingestion for muscle growth is still unclear. Most of the research into leucine consumption has focused on efficacy. To our knowledge, very few studies have sought to determine the maximum safe level of intake. Limited evidence suggests that intakes of ≤1250 mg ⋅ kg -1 . d -1 do not appear to have any health consequences other than short-term elevated plasma ammonia concentrations. Similarly, no adverse events have been reported for the leucine metabolite β-hydroxy-β-methylbutyrate (HMB), although no studies have tested HMB toxicity in humans. Therefore, future research is needed to evaluate leucine and HMB toxicity in the elderly and in specific health conditions. © 2016 American Society for Nutrition.

  13. Application of zipper-fracturing of horizontal cluster wells in the Changning shale gas pilot zone, Sichuan Basin

    Directory of Open Access Journals (Sweden)

    Bin Qian

    2015-03-01

    Full Text Available After several years of exploration practices in the Changning-Weiyuan national shale gas pilot zone, the industrial production has been achieved in a number of vertical and horizontal wells completed by SRV fracturing, and a series of independent shale gas reservoir stimulation technologies have come into being. Next, it is necessary to consider how to enhance the efficiency of fracturing by a factory-mode operation. This paper presents the deployment of Changning Well Pad A, the first cluster horizontal shale gas well group, and proposes the optimal design for the factory operation mode of this Pad according to the requirements of wellpad fracturing stimulation technologies and the mountainous landform in the Sichuan Basin. Accordingly, a zipper-fracturing mode was firstly adopted in the factory fracturing on wellpad. With the application of standardized field process, zipper operation, assembly line work, staggered placement of downhole fractures, and microseismic monitoring in real time, the speed of fracturing reached 3.16 stages a day on average, and the stimulated reservoir volume was maximized, which has fully revealed how the factory operation mode contributes to the large-scale SRV fracturing of horizontal shale gas cluster wells on wellpads in the aspect of speed and efficiency. Moreover, the fracturing process, operation mode, surface facilities and post-fracturing preliminary evaluation of the zipper-fracturing in the well group were examined comprehensively. It is concluded from the practice that the zipper-fracturing in the two wells enhanced the efficiency by 78% and stimulated reservoir volume by 50% compared with the single-well fracturing at the preliminary stage in this area.

  14. Leucine and valine requirements of the growing germfree chicks

    International Nuclear Information System (INIS)

    Ishibashi, Teru; Kametaka, Masao; Ozaki, Akira; Yamamoto, Tetsuzo; Mitsuoka, Tomotari.

    1977-01-01

    To compare the requirements of leucine and valine of the growing germfree and conventional chicks, 7-day-old chicks were fed a diet with graded levels of leucine or valine for 7 days. Daily gains of body weight of the germfree chicks were greater than those of the conventional chicks when the dietary leucine or valine level was higher than the requirement level. Adversely, daily gains of body weight of the germfree chicks were less than those of the conventional chicks when the dietary leucine or valine level was very low. Leucine and valine requirements for maximum growth, however, was the same and estimated to be 0.95 and 0.78%, repectively, of the diet for both groups. After the feeding test, the chicks were injected with L-leucine- and L-valine-U- 14 C and expired carbon dioxide was collected for 2 and 3 hours, respectively. From the percentage of recovery of 14 C in the expired carbon dioxide, the leucine and valine requirements were found to be 0.80 and 0.53% of the diet for both groups, respectively. Three hours after feeding test, the plasma free valine concentration was estimated in the chicks which was not injected the isotope. The requirement of valine was estimated to be 0.59% for the germfree chicks and 0.54% for the conventional chicks. From the above data, it may be concluded that no difference, or very small if any, is found in the requirements of leucine and valine between the germfree chicks and those of the conventional chicks. (auth.)

  15. Leucine kinetics from [2H3]- and [13C]leucine infused simultaneously by gut and vein

    International Nuclear Information System (INIS)

    Hoerr, R.A.; Matthews, D.E.; Bier, D.M.; Young, V.R.

    1991-01-01

    In amino acid tracer kinetic studies of the fed state, ingested amino acid may be taken up during its initial transit through splanchnic tissues and thus not enter the plasma compartment where tracer is infused. To investigate this possibility, adult human subjects received simultaneous intravenous (iv) and intragastric (ig) leucine tracer infusions, first during a postabsorptive (PA) 4-h primed continuous ig infusion of L-[1-13C]-leucine and L-[5,5,5-2H3]leucine iv, followed on a separate day by a fed infusion, in which an ig infusion of a liquid formula was started 2 h before the tracer infusion and continued throughout the tracer study. Subjects were accustomed to a constant experimental diet supplying 1.5 g protein.kg-1.day-1 and 41-45 kcal.kg-1.day-1 for 7 and 12 days before the PA and fed studies, respectively. For the PA study, plasma enrichment for the ig tracer was 3.34 +/- 0.27 (SE) mol + excess and for the iv tracer it was 4.18 +/- 0.10 (P less than 0.02). Enrichments of alpha-keto-isocaproic acid (KIC) were 3.24 +/- 0.16 (ig) and 3.02 +/- 0.14 (iv), respectively [not significant (NS)]. For the fed study, plasma leucine enrichment for the ig tracer was 2.15 +/- 0.14 and for the iv tracer was 2.84 +/- 0.09 (P less than 0.02). KIC enrichments were 2.02 +/- 0.08 (ig) and 2.24 +/- 0.08 (iv), respectively (NS). In the PA study, the ratio of the plasma leucine enrichments for the ig and iv tracers was 0.80 +/- 0.06 and in the fed experiment, 0.76 +/- 0.05, respectively

  16. L-leucine methyl ester stimulates insulin secretion and islet glutamate dehydrogenase

    DEFF Research Database (Denmark)

    Knudsen, P; Kofod, Hans; Lernmark, A

    1983-01-01

    Column perifusion of collagenase-isolated mouse pancreatic islets was used to study the dynamics of insulin release in experiments lasting for several hours. The methyl esters of L-leucine and L-arginine were synthesized. Whereas L-arginine methyl ester (L-arginine OMe) had no effect, L-leucine OMe...... stimulated the release of insulin. The effect of L-leucine OMe was maximal at 5 mmol/liter. Whereas the Km for glucose-stimulated insulin release was unaffected by 1 mmol/liter L-leucine OMe, the maximal release of D-glucose was increased by the amino acid derivative that appeared more effective than L......-leucine. L-Leucine OMe was also a potent stimulus of insulin release from the perfused mouse pancreas. In the presence of 10 mmol/liter L-glutamine, 1 mmol/liter L-leucine OMe induced a 50- to 75-fold increase in insulin release. A similar stimulatory effect was also observed in column-perifused RIN 5F cells...

  17. Circadian secretion patterns of ß-endorphin and leucine enkephalin

    Directory of Open Access Journals (Sweden)

    E. H. de Wet

    1992-07-01

    Full Text Available ß-endorphin and leucine enkephalin are neuropeptides with potent opioid activity. In a study to investigate the circadian secretion patterns of the above-mentioned, blood samples were collected hourly from 12 healthy males who were subjected to the experiment for 24 hours. Radioimmunoassays were used in the analysis of plasma samples for ß-endorphin and leucine enkephalin. Peak concentrations of ß-endorphin were demonstrated from 08:00-09:00, while peak concentrations of leucine enkephalin occured from 23:00-07:00. Trough concentrations of ß-endorphin occurred from 24:00-05:00, while trough con­centrations of leucine enkephalin were demonstrated from 09:00-12:00. The illustrated circadian secretion pattern for ß-endorphin simulates the well-known circadian rhythm of cortisol. The answer to this may be in the fact that ß-endorphin and corticotropin stem from the same precursor. The illustrated circadian secretion pattern for leucine enkephalin simulates that of melatonin. The reason for this is unclear.

  18. Comprehensive characterization and RNA-Seq profiling of the HD-Zip transcription factor family in soybean (Glycine max) during dehydration and salt stress

    Science.gov (United States)

    The homeodomain leucine zipper (HD-Zip) transcription factor family is one of the largest plant specific superfamilies, and includes genes with roles in modulation of plant growth and response to environmental stresses. Many HD-Zip genes are well characterized in Arabidopsis (Arabidopsis thaliana), ...

  19. Leucine supplementation protects from insulin resistance by regulating adiposity levels.

    Directory of Open Access Journals (Sweden)

    Elke Binder

    Full Text Available BACKGROUND: Leucine supplementation might have therapeutic potential in preventing diet-induced obesity and improving insulin sensitivity. However, the underlying mechanisms are at present unclear. Additionally, it is unclear whether leucine supplementation might be equally efficacious once obesity has developed. METHODOLOGY/PRINCIPAL FINDINGS: Male C57BL/6J mice were fed chow or a high-fat diet (HFD, supplemented or not with leucine for 17 weeks. Another group of HFD-fed mice (HFD-pairfat group was food restricted in order to reach an adiposity level comparable to that of HFD-Leu mice. Finally, a third group of mice was exposed to HFD for 12 weeks before being chronically supplemented with leucine. Leucine supplementation in HFD-fed mice decreased body weight and fat mass by increasing energy expenditure, fatty acid oxidation and locomotor activity in vivo. The decreased adiposity in HFD-Leu mice was associated with increased expression of uncoupling protein 3 (UCP-3 in the brown adipose tissue, better insulin sensitivity, increased intestinal gluconeogenesis and preservation of islets of Langerhans histomorphology and function. HFD-pairfat mice had a comparable improvement in insulin sensitivity, without changes in islets physiology or intestinal gluconeogenesis. Remarkably, both HFD-Leu and HFD-pairfat mice had decreased hepatic lipid content, which likely helped improve insulin sensitivity. In contrast, when leucine was supplemented to already obese animals, no changes in body weight, body composition or glucose metabolism were observed. CONCLUSIONS/SIGNIFICANCE: These findings suggest that leucine improves insulin sensitivity in HFD-fed mice by primarily decreasing adiposity, rather than directly acting on peripheral target organs. However, beneficial effects of leucine on intestinal gluconeogenesis and islets of Langerhans's physiology might help prevent type 2 diabetes development. Differently, metabolic benefit of leucine supplementation

  20. Leucine kinetics from (2H3)- and ( sup 13 C)leucine infused simultaneously by gut and vein

    Energy Technology Data Exchange (ETDEWEB)

    Hoerr, R.A.; Matthews, D.E.; Bier, D.M.; Young, V.R. (Massachusetts Institute of Technology, Cambridge (USA))

    1991-01-01

    In amino acid tracer kinetic studies of the fed state, ingested amino acid may be taken up during its initial transit through splanchnic tissues and thus not enter the plasma compartment where tracer is infused. To investigate this possibility, adult human subjects received simultaneous intravenous (iv) and intragastric (ig) leucine tracer infusions, first during a postabsorptive (PA) 4-h primed continuous ig infusion of L-(1-13C)-leucine and L-(5,5,5-2H3)leucine iv, followed on a separate day by a fed infusion, in which an ig infusion of a liquid formula was started 2 h before the tracer infusion and continued throughout the tracer study. Subjects were accustomed to a constant experimental diet supplying 1.5 g protein.kg-1.day-1 and 41-45 kcal.kg-1.day-1 for 7 and 12 days before the PA and fed studies, respectively. For the PA study, plasma enrichment for the ig tracer was 3.34 +/- 0.27 (SE) mol + excess and for the iv tracer it was 4.18 +/- 0.10 (P less than 0.02). Enrichments of alpha-keto-isocaproic acid (KIC) were 3.24 +/- 0.16 (ig) and 3.02 +/- 0.14 (iv), respectively (not significant (NS)). For the fed study, plasma leucine enrichment for the ig tracer was 2.15 +/- 0.14 and for the iv tracer was 2.84 +/- 0.09 (P less than 0.02). KIC enrichments were 2.02 +/- 0.08 (ig) and 2.24 +/- 0.08 (iv), respectively (NS). In the PA study, the ratio of the plasma leucine enrichments for the ig and iv tracers was 0.80 +/- 0.06 and in the fed experiment, 0.76 +/- 0.05, respectively.

  1. Small leucine-rich proteoglycans in the aging skeleton

    DEFF Research Database (Denmark)

    Young, M F; Bi, Y; Ameye, L

    2006-01-01

    Small Leucine-Rich Proteoglyans (SLRPs) are major skeletal extracellular matrix (ECM) components that comprise a family of 13 members containing repeats of a leucine-rich motif. To examine SLRP function, we generated mice deficient in one or more member and analyzed them at the tissue, cell...

  2. Sequence Classification: 889214 [

    Lifescience Database Archive (English)

    Full Text Available elix-leucine zipper (bHLH/Zip) transcription factor that forms a complex with another bHLH/Zip protein, Rtg1p, to activate the retrog...rade (RTG) and TOR pathways; Rtg3p || http://www.ncbi.nlm.nih.gov/protein/6319365 ...

  3. Leucine metabolism in cirrhotic patients with hepatic encephalopathy

    International Nuclear Information System (INIS)

    McGhee, A.S.

    1985-01-01

    The purpose of this study was to determine whether increased oxidation of or protein synthesis requiring leucine occurs in cirrhotic patients. Five control subjects and four subjects with cirrhosis were equilibrated on a baseline diet (0.6 g protein per kg ideal body weight [IBW]) with sufficient nonprotein calories to preclude negative nitrogen balance. An additional four patients were equilibrated on the same type of diet with a higher protein level (0.75 g per kg IBW). Control subjects and the patients were then studied during continuous infusion of L-[ 15 N, 1- 13 C] leucine in the fasted state and, in the fed state, with a Propac diet which had the same distribution of energy nutrients as the baseline diets. Plasma levels of L-[ 15 N, 1- 13 C], L-[1- 13 C] and L-[ 15 N] leucine were measured during isotopic steady state by gas chromatography-mass spectrometry and fractional excretion of 13 CO 2 in breath samples were analyzed by isotopic ratio mass spectrometry. During the fasted and fed states leucine metabolism was measured to quantitate rates of nitrogen flux (Q/sub N/), carbon flux (Q/sub c/) and oxidation to carbon dioxide and water (C). From these measured values, proteins breakdown (B), protein synthesis (S), deamination (X 0 ) and reamination (X/sub N/) were calculated. The results showed that protein synthesis and leucine metabolism were identical in controls and patients when both were fed a diet with 0.6 g protein/kg IBW and maintenance level of nonprotein calories. The data also showed that leucine metabolism can be quantitatively and reproducibly measured in subjects with cirrhosis

  4. Nutritional efficiency of alpha-ketoisocaproate relative to leucine, assessed isotopically

    International Nuclear Information System (INIS)

    Kang, C.W.; Walser, M.

    1985-01-01

    The efficiency of alpha-ketoisocaproate as a dietary substitute for leucine was assessed in rats by two techniques: first, the minimal dose of alpha-ketoisocaproate required, as a supplement to a leucine-free diet, to achieve a growth rate as great as animals receiving leucine was found to be between 2.2 and 4.4 times larger. Therefore the nutritional efficiency of alpha-ketoisocaproate lies between 0.23 and 0.46. Second, alpha-[1- 14 C]-ketoisocaproate and [ 3 H]leucine were administered orally and the ratio of 14 C/ 3 H incorporated into the leucine of whole-body protein and fibrin was measured. This ratio, divided by the ratio 14 C/ 3 H injected, was the same in fibrin as in whole-body protein and averaged 0.39. Thus both techniques yield the same value, within the error of measurement, for the relative nutritional efficiency of alpha-ketoisocaproate. The authors also found that alpha-ketoisocaproate feeding at varying dosage did not alter this ratio in whole-body protein, suggesting that neither wide variations in growth rate nor exposure for 10 days to alpha-ketoisocaproate alters the relative rates of utilization (or oxidation) of alpha-ketoisocaproate vs. leucine

  5. Nutritional efficiency of alpha-ketoisocaproate relative to leucine, assessed isotopically

    Energy Technology Data Exchange (ETDEWEB)

    Kang, C.W.; Walser, M.

    1985-10-01

    The efficiency of alpha-ketoisocaproate as a dietary substitute for leucine was assessed in rats by two techniques: first, the minimal dose of alpha-ketoisocaproate required, as a supplement to a leucine-free diet, to achieve a growth rate as great as animals receiving leucine was found to be between 2.2 and 4.4 times larger. Therefore the nutritional efficiency of alpha-ketoisocaproate lies between 0.23 and 0.46. Second, alpha-(1- UC)-ketoisocaproate and (TH)leucine were administered orally and the ratio of UC/TH incorporated into the leucine of whole-body protein and fibrin was measured. This ratio, divided by the ratio UC/TH injected, was the same in fibrin as in whole-body protein and averaged 0.39. Thus both techniques yield the same value, within the error of measurement, for the relative nutritional efficiency of alpha-ketoisocaproate. The authors also found that alpha-ketoisocaproate feeding at varying dosage did not alter this ratio in whole-body protein, suggesting that neither wide variations in growth rate nor exposure for 10 days to alpha-ketoisocaproate alters the relative rates of utilization (or oxidation) of alpha-ketoisocaproate vs. leucine.

  6. Meta-analysis of Arabidopsis KANADI1 direct target genes identifies basic growth-promoting module acting upstream of hormonal signaling pathways

    DEFF Research Database (Denmark)

    Xie, Yakun; Straub, Daniel; Eguen, Teinai Ebimienere

    2015-01-01

    An intricate network of antagonistically acting transcription factors mediates formation of a flat leaf lamina of Arabidopsis thaliana plants. In this context, members of the class III homeodomain leucine zipper (HD-ZIPIII) transcription factor family specify the adaxial domain (future upper side......) of the leaf, while antagonistically acting KANADI transcription factors determine the abaxial domain (future lower side). Here we used an mRNA-seq approach to identify genes regulated by KANADI1 (KAN1) and subsequently performed a meta-analysis approach combining our datasets with published genome......-wide datasets. Our analysis revealed that KAN1 acts upstream of several genes encoding auxin biosynthetic enzymes. When exposed to shade, we find three YUCCA genes, YUC2, YUC5 and YUC8 to be transcriptionally upregulated, which correlates with an increase in the levels of free auxin. When ectopically expressed...

  7. Leucine supplementation in the management of protein energy ...

    African Journals Online (AJOL)

    Objective: To establish if leucine could be used as a therapeutic agent in the treatment of protein energy malnutrition. Study selection: Based on defined key words a search was carried out on Pubmed to retrieve all publications on leucine supplementation and muscle protein synthesis. Only studies that met the search ...

  8. Differentiating leucine incorporation of

    NARCIS (Netherlands)

    Yokokawa, T.; Sintes, E.; de Corte, D.; Olbrich, K.; Herndl, G.J.

    2012-01-01

    The abundance (based on catalyzed reporter deposition-fluorescence in situ hybrid ization, CARD-FISH) and leucine incorporation rates of Archaea and Bacteria were determined throughout the water column in the eastern Atlantic. Bacteria dominated throughout the water column, although their

  9. Leucine Supplementation Accelerates Connective Tissue Repair of Injured Tibialis Anterior Muscle

    Directory of Open Access Journals (Sweden)

    Marcelo G. Pereira

    2014-09-01

    Full Text Available This study investigated the effect of leucine supplementation on the skeletal muscle regenerative process, focusing on the remodeling of connective tissue of the fast twitch muscle tibialis anterior (TA. Young male Wistar rats were supplemented with leucine (1.35 g/kg per day; then, TA muscles from the left hind limb were cryolesioned and examined after 10 days. Although leucine supplementation induced increased protein synthesis, it was not sufficient to promote an increase in the cross-sectional area (CSA of regenerating myofibers (p > 0.05 from TA muscles. However, leucine supplementation reduced the amount of collagen and the activation of phosphorylated transforming growth factor-β receptor type I (TβR-I and Smad2/3 in regenerating muscles (p < 0.05. Leucine also reduced neonatal myosin heavy chain (MyHC-n (p < 0.05, increased adult MyHC-II expression (p < 0.05 and prevented the decrease in maximum tetanic strength in regenerating TA muscles (p < 0.05. Our results suggest that leucine supplementation accelerates connective tissue repair and consequent function of regenerating TA through the attenuation of TβR-I and Smad2/3 activation. Therefore, future studies are warranted to investigate leucine supplementation as a nutritional strategy to prevent or attenuate muscle fibrosis in patients with several muscle diseases.

  10. Increased Dietary Leucine Reduces Doxorubicin-Associated Cardiac Dysfunction in Rats

    Directory of Open Access Journals (Sweden)

    Thiago M. Fidale

    2018-01-01

    Full Text Available Cardiotoxicity is one of the most significant adverse effects of the oncologic treatment with doxorubicin, which is responsible for a substantial morbid and mortality. The occurrence of heart failure with ventricular dysfunction may lead to severe cardiomyopathy and ultimately to death. Studies have focused on the effects of leucine supplementation as a strategy to minimize or revert the clinical condition of induced proteolysis by several clinical onsets. However, the impact of leucine supplementation in heart failure induced by doxorubicin is unknown. Therefore, the objective of this work is to evaluate the effects of leucine supplementation on the cardiotoxicity in the heart of rats treated with doxorubicin. Rats treated with a 7.5 mg/kg cumulative dose of doxorubicin for 14 days presented a dilatation of the left ventricle (LV, and a reduction of the ejection fraction (FE. The 5% supplementation of leucine in the rats' food prevented the malfunctioning of the LV when administered with doxorubicin. Some alterations in the extracellular matrix remodeling were confirmed by the increase of collagen fibers in the doxorubicin group, which did not increase when the treatment was associated with leucine supplementation. Leucine attenuates heart failure in this experimental model with doxorubicin. Such protection is followed by the maintenance of interstitial collagen fibers.

  11. The structural determinants responsible for c-Fos protein proteasomal degradation differ according to the conditions of expression.

    Science.gov (United States)

    Ferrara, Patrizia; Andermarcher, Elisabetta; Bossis, Guillaume; Acquaviva, Claire; Brockly, Frédérique; Jariel-Encontre, Isabelle; Piechaczyk, Marc

    2003-03-13

    basic domain/leucine zipper-dependent mechanisms. Taken together, our data show that multiple degradation mechanisms, differing according to the conditions of expression, may operate on c-Fos to ensure a proper level and/or timing of expression. Moreover, they also indicate that the half-life of c-Fos during the G0/G1 phase transition is determined by a delicate balance between opposing stabilizing and destabilizing mechanisms operating at the same time.

  12. Effects of leucine supplemented diet on intestinal absorption in tumor bearing pregnant rats

    Directory of Open Access Journals (Sweden)

    de Mello Maria

    2002-04-01

    Full Text Available Abstract Background It is known that amino acid oxidation is increased in tumor-bearing rat muscles and that leucine is an important ketogenic amino acid that provides energy to the skeletal muscle. Methods To evaluate the effects of a leucine supplemented diet on the intestinal absorption alterations produced by Walker 256, growing pregnant rats were distributed into six groups. Three pregnant groups received a normal protein diet (18% protein: pregnant (N, tumor-bearing (WN, pair-fed rats (Np. Three other pregnant groups were fed a diet supplemented with 3% leucine (15% protein plus 3% leucine: leucine (L, tumor-bearing (WL and pair-fed with leucine (Lp. Non pregnant rats (C, which received a normal protein diet, were used as a control group. After 20 days, the animals were submitted to intestinal perfusion to measure leucine, methionine and glucose absorption. Results Tumor-bearing pregnant rats showed impairment in food intake, body weight gain and muscle protein content, which were less accentuated in WL than in WN rats. These metabolic changes led to reduction in both fetal and tumor development. Leucine absorption slightly increased in WN group. In spite of having a significant decrease in leucine and methionine absorption compared to L, the WL group has shown a higher absorption rate of methionine than WN group, probably due to the ingestion of the leucine supplemented diet inducing this amino acid uptake. Glucose absorption was reduced in both tumor-bearing groups. Conclusions Leucine supplementation during pregnancy in tumor-bearing rats promoted high leucine absorption, increasing the availability of the amino acid for neoplasic cells and, mainly, for fetus and host utilization. This may have contributed to the better preservation of body weight gain, food intake and muscle protein observed in the supplemented rats in relation to the non-supplemented ones.

  13. Effects of leucine supplemented diet on intestinal absorption in tumor bearing pregnant rats

    International Nuclear Information System (INIS)

    Ventrucci, Gislaine; Mello, Maria Alice Roston de; Gomes-Marcondes, Maria Cristina Cintra

    2002-01-01

    It is known that amino acid oxidation is increased in tumor-bearing rat muscles and that leucine is an important ketogenic amino acid that provides energy to the skeletal muscle. To evaluate the effects of a leucine supplemented diet on the intestinal absorption alterations produced by Walker 256, growing pregnant rats were distributed into six groups. Three pregnant groups received a normal protein diet (18% protein): pregnant (N), tumor-bearing (WN), pair-fed rats (Np). Three other pregnant groups were fed a diet supplemented with 3% leucine (15% protein plus 3% leucine): leucine (L), tumor-bearing (WL) and pair-fed with leucine (Lp). Non pregnant rats (C), which received a normal protein diet, were used as a control group. After 20 days, the animals were submitted to intestinal perfusion to measure leucine, methionine and glucose absorption. Tumor-bearing pregnant rats showed impairment in food intake, body weight gain and muscle protein content, which were less accentuated in WL than in WN rats. These metabolic changes led to reduction in both fetal and tumor development. Leucine absorption slightly increased in WN group. In spite of having a significant decrease in leucine and methionine absorption compared to L, the WL group has shown a higher absorption rate of methionine than WN group, probably due to the ingestion of the leucine supplemented diet inducing this amino acid uptake. Glucose absorption was reduced in both tumor-bearing groups. Leucine supplementation during pregnancy in tumor-bearing rats promoted high leucine absorption, increasing the availability of the amino acid for neoplasic cells and, mainly, for fetus and host utilization. This may have contributed to the better preservation of body weight gain, food intake and muscle protein observed in the supplemented rats in relation to the non-supplemented ones

  14. Prostate Cancer Cells in Different Androgen Receptor Status Employ Different Leucine Transporters.

    Science.gov (United States)

    Otsuki, Hideo; Kimura, Toru; Yamaga, Takashi; Kosaka, Takeo; Suehiro, Jun-Ichi; Sakurai, Hiroyuki

    2017-02-01

    Leucine stimulates cancer cell proliferation through the mTOR pathway, therefore, inhibiting leucine transporters may be a novel therapeutic target for cancer. L-type amino acid transporter (LAT) 1, a Na + -independent amino acid transporter, is highly expressed in many tumor cells. However, leucine transporter(s) in different stages of prostate cancer, particularly in the stages of castration resistance with androgen receptor (AR) expression, is unclear. LNCaP and DU145 and PC-3 cell lines were used as a model of androgen dependent, and metastatic prostate cancer. A new "LN-cr" cell line was established after culturing LNCaP cells for 6 months under androgen-free conditions, which is considered a model of castration resistant prostate cancer (CRPC) with androgen AR expression. The expression of leucine transporters was investigated with quantitative PCR and immunofluorescence. Uptake of 14 C Leucine was examined in the presence or absence of BCH (a pan-LAT inhibitor), JPH203 (an LAT1-specific inhibitor), or Na + . Cell growth was assessed with MTT assay. siRNA studies were performed to evaluate the indispensability of y + LAT2 on leucine uptake and cell viability in LN-cr. Cell viability showed a 90% decrease in the absence of leucine in all four cell lines. LNCaP cells principally expressed LAT3, and their leucine uptake was more than 90% Na + -independent. BCH, but not JPH203, inhibited leucine uptake, and cell proliferation (IC 50BCH :15 mM). DU145 and PC-3 cells predominantly expressed LAT1. Leucine uptake and cell growth were suppressed by BCH or JPH203 in a dose-dependent manner (IC 50BCH : ∼20 mM, IC 50JPH203 : ∼5 µM). In LN-cr cells, Na + -dependent uptake of leucine was 3.8 pmol/mgprotein/min, while, Na + -independent uptake was only 0.52 (P prostate cancer. Prostate 77:222-233, 2017. © 2016 Wiley Periodicals, Inc. © 2016 Wiley Periodicals, Inc.

  15. Function of the HD-Zip I gene Oshox22 in ABA-mediated drought and salt tolerances in rice

    NARCIS (Netherlands)

    Zhang, S.; Kohlen, W.; Jiang, L.; Bouwmeester, H.J.; Meijer, A.H.; Schluepmann, H.; Liu, C.M.; Ouwerkerk, P.B.F.

    2012-01-01

    Oshox22 belongs to the homeodomain-leucine zipper (HD-Zip) family I of transcription factors, most of which have unknown functions. Here we show that the expression of Oshox22 is strongly induced by salt stress, abscisic acid (ABA), and polyethylene glycol treatment (PEG), and weakly by cold stress.

  16. The Effect of Oral Leucine on Protein Metabolism in Adolescents with Type 1 Diabetes Mellitus

    Directory of Open Access Journals (Sweden)

    Wilson ThomasA

    2010-11-01

    Full Text Available Lack of insulin results in a catabolic state in subjects with insulin-dependent diabetes mellitus which is reversed by insulin treatment. Amino acid supply, especially branched chain amino acids such as leucine, enhances protein synthesis in both animal and human studies. This small study was undertaken to assess the acute effect of supplemental leucine on protein metabolism in adolescents with type 1 diabetes. L-[1-13C] Leucine was used to assess whole-body protein metabolism in six adolescent females (16–18 yrs with type 1 diabetes during consumption of a basal diet (containing 58 μmoles leucine/kg/h and the basal diet with supplemental leucine (232 μmoles leucine/kg/h. Net leucine balance was significantly higher with supplemental leucine ( μmoles leucine/kg body weight/hr than with the basal diet (, due to reduced protein degradation ( μmoles leucine/kg body weight/hr compared to the basal diet (, .

  17. Conformational composition of neutral leucine. Matrix isolation infrared and ab initio study

    International Nuclear Information System (INIS)

    Stepanian, Stepan G.; Ivanov, Alexander Yu.; Adamowicz, Ludwik

    2013-01-01

    Highlights: • FTIR spectra of leucine isolated in argon, neon and xenon matrices are obtained. • UV irradiation is used to separate bands of the leucine conformers. • Populations of the leucine conformers is determined. - Abstract: Low-temperature matrix-isolation FTIR spectroscopy and ab initio calculations are employed to determine conformational composition of neutral leucine. The presence of three leucine conformers in the matrices is revealed. This is in agreement with the results of a detailed study of the potential energy surface of leucine which demonstrates that only five out of 105 possible conformers should have populations in the matrices larger than 2% and only three conformers, which are the ones detected in the experiment, should have populations larger than 10%. UV irradiation of the matrix samples are used to separate bands of the different conformers. We also show that the populations of the leucine conformers in the gas phase at 440 K are significantly different from the ones in matrices. The population of the lowest energy conformer in the gas phase being approximately 23% in the gas phase increases to over 64% in matrices

  18. A zipper network model of the failure mechanics of extracellular matrices.

    Science.gov (United States)

    Ritter, Michael C; Jesudason, Rajiv; Majumdar, Arnab; Stamenovic, Dimitrije; Buczek-Thomas, Jo Ann; Stone, Phillip J; Nugent, Matthew A; Suki, Béla

    2009-01-27

    Mechanical failure of soft tissues is characteristic of life-threatening diseases, including capillary stress failure, pulmonary emphysema, and vessel wall aneurysms. Failure occurs when mechanical forces are sufficiently high to rupture the enzymatically weakened extracellular matrix (ECM). Elastin, an important structural ECM protein, is known to stretch beyond 200% strain before failing. However, ECM constructs and native vessel walls composed primarily of elastin and proteoglycans (PGs) have been found to fail at much lower strains. In this study, we hypothesized that PGs significantly contribute to tissue failure. To test this, we developed a zipper network model (ZNM), in which springs representing elastin are organized into long wavy fibers in a zipper-like formation and placed within a network of springs mimicking PGs. Elastin and PG springs possessed distinct mechanical and failure properties. Simulations using the ZNM showed that the failure of PGs alone reduces the global failure strain of the ECM well below that of elastin, and hence, digestion of elastin does not influence the failure strain. Network analysis suggested that whereas PGs drive the failure process and define the failure strain, elastin determines the peak and failure stresses. Predictions of the ZNM were experimentally confirmed by measuring the failure properties of engineered elastin-rich ECM constructs before and after digestion with trypsin, which cleaves the core protein of PGs without affecting elastin. This study reveals a role for PGs in the failure properties of engineered and native ECM with implications for the design of engineered tissues.

  19. Acetone Formation in the Vibrio Family: a New Pathway for Bacterial Leucine Catabolism

    Science.gov (United States)

    Nemecek-Marshall, Michele; Wojciechowski, Cheryl; Wagner, William P.; Fall, Ray

    1999-01-01

    There is current interest in biological sources of acetone, a volatile organic compound that impacts atmospheric chemistry. Here, we determined that leucine-dependent acetone formation is widespread in the Vibrionaceae. Sixteen Vibrio isolates, two Listonella species, and two Photobacterium angustum isolates produced acetone in the presence of l-leucine. Shewanella isolates produced much less acetone. Growth of Vibrio splendidus and P. angustum in a fermentor with controlled aeration revealed that acetone was produced after a lag in late logarithmic or stationary phase of growth, depending on the medium, and was not derived from acetoacetate by nonenzymatic decarboxylation in the medium. l-Leucine, but not d-leucine, was converted to acetone with a stoichiometry of approximately 0.61 mol of acetone per mol of l-leucine. Testing various potential leucine catabolites as precursors of acetone showed that only α-ketoisocaproate was efficiently converted by whole cells to acetone. Acetone production was blocked by a nitrogen atmosphere but not by electron transport inhibitors, suggesting that an oxygen-dependent reaction is required for leucine catabolism. Metabolic labeling with deuterated (isopropyl-d7)-l-leucine revealed that the isopropyl carbons give rise to acetone with full retention of deuterium in each methyl group. These results suggest the operation of a new catabolic pathway for leucine in vibrios that is distinct from the 3-hydroxy-3-methylglutaryl-coenzyme A pathway seen in pseudomonads. PMID:10601206

  20. Leucine zipper transcription factor-like 1 expression in gastric ...

    African Journals Online (AJOL)

    the gastric cancer tissues, and its relationship to cellular adhesion protein. ... related to cellular adhesion protein, but not to mRNA level. ... malignant tumors, and is correlated with gastric, ... The present study was aimed at investigating the.

  1. Rapid sensing of l-leucine by human and murine hypothalamic neurons: Neurochemical and mechanistic insights.

    Science.gov (United States)

    Heeley, Nicholas; Kirwan, Peter; Darwish, Tamana; Arnaud, Marion; Evans, Mark L; Merkle, Florian T; Reimann, Frank; Gribble, Fiona M; Blouet, Clemence

    2018-04-01

    Dietary proteins are sensed by hypothalamic neurons and strongly influence multiple aspects of metabolic health, including appetite, weight gain, and adiposity. However, little is known about the mechanisms by which hypothalamic neural circuits controlling behavior and metabolism sense protein availability. The aim of this study is to characterize how neurons from the mediobasal hypothalamus respond to a signal of protein availability: the amino acid l-leucine. We used primary cultures of post-weaning murine mediobasal hypothalamic neurons, hypothalamic neurons derived from human induced pluripotent stem cells, and calcium imaging to characterize rapid neuronal responses to physiological changes in extracellular l-Leucine concentration. A neurochemically diverse subset of both mouse and human hypothalamic neurons responded rapidly to l-leucine. Consistent with l-leucine's anorexigenic role, we found that 25% of mouse MBH POMC neurons were activated by l-leucine. 10% of MBH NPY neurons were inhibited by l-leucine, and leucine rapidly reduced AGRP secretion, providing a mechanism for the rapid leucine-induced inhibition of foraging behavior in rodents. Surprisingly, none of the candidate mechanisms previously implicated in hypothalamic leucine sensing (K ATP channels, mTORC1 signaling, amino-acid decarboxylation) were involved in the acute activity changes produced by l-leucine. Instead, our data indicate that leucine-induced neuronal activation involves a plasma membrane Ca 2+ channel, whereas leucine-induced neuronal inhibition is mediated by inhibition of a store-operated Ca 2+ current. A subset of neurons in the mediobasal hypothalamus rapidly respond to physiological changes in extracellular leucine concentration. Leucine can produce both increases and decreases in neuronal Ca 2+ concentrations in a neurochemically-diverse group of neurons, including some POMC and NPY/AGRP neurons. Our data reveal that leucine can signal through novel mechanisms to rapidly

  2. Journal of Biosciences | Indian Academy of Sciences

    Indian Academy of Sciences (India)

    ... identity to homeobox-leucine zipper protein HAHB-4. The analysis of proteome changes induced by N+ implantation could provid a new clue to studing mutation mechanism of ion implantation. To our knowledge, this is the first report about the analysis of proteome changes induced by N+ implantation in sunflower seeds.

  3. Interactions between the metabolism of L-leucine and D-glucose in the pancreatic. beta. -cells

    Energy Technology Data Exchange (ETDEWEB)

    Gylfe, E; Sehlin, J [Umeaa Univ. (Sweden). Dept. of Histology

    1976-01-01

    ..beta..-cell-rich pancreatic islets microdissected from obese-hyperglycemic mice were used to study interactions between the metabolism of L-leucine and D-glucose. L-leucine reduced the islet content of aspartic acid whereas D-glucose, when added to L-leucine-incubated islets, increased the contents of aspartic acid and ..gamma..-aminobutyric acid (GABA). D-glucose also increased the incorporation of L-leucine carbon into aspartic acid, GABA and glutamic acid, suggesting stimulation of a malate shuttle mechanism. When expressed per mole of the individual amino acids, the incorporation of L-leucine carbon into GABA was 2.5 - 4 times higher than into glutamic acid indicating intracellular compartmentation of the latter amino acid. Both L-leucine and D-leucine stimulated /sup 14/CO/sub 2/ production from /sup 14/C-labelled D-glucose. L-leucine did not affect /sup 3/H/sub 2/O production from tritiated D-glucose. The present data do not indicate a role of other amino acids or D-glucose in L-leucine-stimulated insulin release.

  4. Leucine signaling in the pathogenesis of type 2 diabetes and obesity.

    Science.gov (United States)

    Melnik, Bodo C

    2012-03-15

    Epidemiological evidence points to increased dairy and meat consumption, staples of the Western diet, as major risk factors for the development of type 2 diabetes (T2D). This paper presents a new concept and comprehensive review of leucine-mediated cell signaling explaining the pathogenesis of T2D and obesity by leucine-induced over-stimulation of mammalian target of rapamycin complex 1 (mTORC1). mTORC1, a pivotal nutrient-sensitive kinase, promotes growth and cell proliferation in response to glucose, energy, growth factors and amino acids. Dairy proteins and meat stimulate insulin/insulin-like growth factor 1 signaling and provide high amounts of leucine, a primary and independent stimulator for mTORC1 activation. The downstream target of mTORC1, the kinase S6K1, induces insulin resistance by phosphorylation of insulin receptor substrate-1, thereby increasing the metabolic burden of β-cells. Moreover, leucine-mediated mTORC1-S6K1-signaling plays an important role in adipogenesis, thus increasing the risk of obesity-mediated insulin resistance. High consumption of leucine-rich proteins explains exaggerated mTORC1-dependent insulin secretion, increased β-cell growth and β-cell proliferation promoting an early onset of replicative β-cell senescence with subsequent β-cell apoptosis. Disturbances of β-cell mass regulation with increased β-cell proliferation and apoptosis as well as insulin resistance are hallmarks of T2D, which are all associated with hyperactivation of mTORC1. In contrast, the anti-diabetic drug metformin antagonizes leucine-mediated mTORC1 signaling. Plant-derived polyphenols and flavonoids are identified as natural inhibitors of mTORC1 and exert anti-diabetic and anti-obesity effects. Furthermore, bariatric surgery in obesity reduces increased plasma levels of leucine and other branched-chain amino acids. Attenuation of leucine-mediated mTORC1 signaling by defining appropriate upper limits of the daily intake of leucine-rich animal and dairy

  5. Reaction of some selected soybean varieties ( Glycine max (L) Merril)

    African Journals Online (AJOL)

    In nematode endemic ecological zones, TGX-1985 – 8F is therefore recommended as it proved to contain some specialized genes that conferred a higher level of tolerance against root- knot nematode, Meloidogyne incognita. Key Words: Glycine max, root – knot nematode, Dominant loci, Mi – 1.2, leucine zipper and R ...

  6. Leucine metabolism in regulation of insulin secretion from pancreatic beta cells

    OpenAIRE

    Yang, Jichun; Chi, Yujing; Burkhardt, Brant R.; Guan, Youfei; Wolf, Bryan A

    2010-01-01

    Leucine, a the branched-chain amino acids that must be supplied in daily diet, plays an important role in controlling protein synthesis and regulating cell metabolism in various cell types. In pancreatic β cells, leucine acutely stimulates insulin secretion by serving as both metabolic fuel and allosteric activator of glutamate dehydrogenase to enhance glutaminolysis. Leucine has also been shown to regulate gene transcription and protein synthesis in pancreatic islet β cells via both mTOR-dep...

  7. Metabolic Mechanism for l-Leucine-Induced Metabolome To Eliminate Streptococcus iniae.

    Science.gov (United States)

    Du, Chao-Chao; Yang, Man-Jun; Li, Min-Yi; Yang, Jun; Peng, Bo; Li, Hui; Peng, Xuan-Xian

    2017-05-05

    Crucial metabolites that modulate hosts' metabolome to eliminate bacterial pathogens have been documented, but the metabolic mechanisms are largely unknown. The present study explores the metabolic mechanism for l-leucine-induced metabolome to eliminate Streptococcus iniae in tilapia. GC-MS-based metabolomics was used to investigate the tilapia liver metabolic profile in the presence of exogenous l-leucine. Thirty-seven metabolites of differential abundance were determined, and 11 metabolic pathways were enriched. Pattern recognition analysis identified serine and proline as crucial metabolites, which are the two metabolites identified in survived tilapias during S. iniae infection, suggesting that the two metabolites play crucial roles in l-leucine-induced elimination of the pathogen by the host. Exogenous l-serine reduces the mortality of tilapias infected by S. iniae, providing a robust proof supporting the conclusion. Furthermore, exogenous l-serine elevates expression of genes IL-1β and IL-8 in tilapia spleen, but not TNFα, CXCR4 and Mx, suggesting that the metabolite promotes a phagocytosis role of macrophages, which is consistent with the finding that l-leucine promotes macrophages to kill both Gram-positive and Gram-negative bacterial pathogens. Therefore, the ability of phagocytosis enhanced by exogenous l-leucine is partly attributed to elevation of l-serine. These results demonstrate a metabolic mechanism by which exogenous l-leucine modulates tilapias' metabolome to enhance innate immunity and eliminate pathogens.

  8. Comparison of the fates of ingested leucine and ingested 2-ketoisocaproate in rats

    Energy Technology Data Exchange (ETDEWEB)

    Imura, K.; Walser, M. (Johns Hopkins Univ. School of Medicine, Baltimore, MD (USA))

    1990-05-01

    We previously reported that the ratio, R, of 14C to 3H in the leucine of whole body protein, measured 6 h after ingestion of (3H)leucine and (1-14C)2-ketoisocaproate is equal to ratio of the dose of leucine to the dose of 2-ketoisocaproate (KIC) (on a leucine-free diet) required to achieve the same rate of growth. To determine whether R is dependent on the interval between injection and sampling, R was measured at intervals in purified whole body protein after oral injection of these isotopes in groups of rats; it was constant from 1 h onward for 1 wk, averaging 0.64 +/- 0.01 (means +/- SEM). Thus, the extent of incorporation into the leucine of whole body protein of ingested KIC remains close to 64% of the incorporation of ingested leucine administered as such simultaneously, from 1 h onward for at least 1 wk.

  9. Leucine supplementation attenuates macrophage foam-cell formation: Studies in humans, mice, and cultured macrophages.

    Science.gov (United States)

    Grajeda-Iglesias, Claudia; Rom, Oren; Hamoud, Shadi; Volkova, Nina; Hayek, Tony; Abu-Saleh, Niroz; Aviram, Michael

    2018-02-05

    Whereas atherogenicity of dietary lipids has been largely studied, relatively little is known about the possible contribution of dietary amino acids to macrophage foam-cell formation, a hallmark of early atherogenesis. Recently, we showed that leucine has antiatherogenic properties in the macrophage model system. In this study, an in-depth investigation of the role of leucine in macrophage lipid metabolism was conducted by supplementing humans, mice, or cultured macrophages with leucine. Macrophage incubation with serum obtained from healthy adults supplemented with leucine (5 g/d, 3 weeks) significantly decreased cellular cholesterol mass by inhibiting the rate of cholesterol biosynthesis and increasing cholesterol efflux from macrophages. Similarly, leucine supplementation to C57BL/6 mice (8 weeks) resulted in decreased cholesterol content in their harvested peritoneal macrophages (MPM) in relation with reduced cholesterol biosynthesis rate. Studies in J774A.1 murine macrophages revealed that leucine dose-dependently decreased cellular cholesterol and triglyceride mass. Macrophages treated with leucine (0.2 mM) showed attenuated uptake of very low-density lipoproteins and triglyceride biosynthesis rate, with a concurrent down-regulation of diacylglycerol acyltransferase-1, a key enzyme catalyzing triglyceride biosynthesis in macrophages. Similar effects were observed when macrophages were treated with α-ketoisocaproate, a key leucine metabolite. Finally, both in vivo and in vitro leucine supplementation significantly improved macrophage mitochondrial respiration and ATP production. The above studies, conducted in human, mice, and cultured macrophages, highlight a protective role for leucine attenuating macrophage foam-cell formation by mechanisms related to the metabolism of cholesterol, triglycerides, and energy production. © 2018 BioFactors, 2018. © 2018 International Union of Biochemistry and Molecular Biology.

  10. Determination of Optimum Condition of Leucine Content in Beef Protein Hydrolysate using Response Surface Methodology

    International Nuclear Information System (INIS)

    Siti Roha Ab Mutalib; Zainal Samicho; Noriham Abdullah

    2016-01-01

    The aim of this study is to determine the optimum condition of leucine content in beef hydrolysate. Beef hydrolysate was prepared by enzymatic hydrolysis using bromelain enzyme produced from pineapple peel. Parameter conditions such as concentration of bromelain, hydrolysis temperature and hydrolysis time were assessed to obtain the optimum leucine content of beef hydrolysate according to experimental design which was recommended by response surface methodology (RSM). Leucine content in beef hydrolysate was determined using AccQ. Tag amino acid analysis method using high performance liquid chromatography (HPLC). The condition of optimum leucine content was at bromelain concentration of 1.38 %, hydrolysis temperature of 42.5 degree Celcius and hydrolysis time of 31.59 hours with the predicted leucine content of 26.57 %. The optimum condition was verified with the leucine value obtained was 26.25 %. Since there was no significant difference (p>0.05) between the predicted and verified leucine values, thus it indicates that the predicted optimum condition by RSM can be accepted to predict the optimum leucine content in beef hydrolysate. (author)

  11. Genome-wide systematic characterization of the bZIP transcriptional factor family in tomato (Solanum lycopersicum L.).

    Science.gov (United States)

    Li, Dayong; Fu, Fuyou; Zhang, Huijuan; Song, Fengming

    2015-10-12

    Transcription factors of the basic leucine zipper (bZIP) family represent exclusively in eukaryotes and have been shown to regulate diverse biological processes in plant growth and development as well as in abiotic and biotic stress responses. However, little is known about the bZIP family in tomato (Solanum lycopersicum L.). The SlbZIP genes were identified using local BLAST and hidden Markov model profile searches. The phylogenetic trees, conserved motifs and gene structures were generated by MEGA6.06, MEME tool and gene Structure Display Server, respectively. The syntenic block diagrams were generated by the Circos software. The transcriptional gene expression profiles were obtained using Genevestigator tool and quantitative RT-PCR. In the present study, we carried out a genome-wide identification and systematic analyses of 69 SlbZIP genes that distributes unevenly on the tomato chromosomes. This family can be divided into 9 groups according to the phylogenetic relationship among the SlbZIP proteins. Six kinds of intron patterns (a-f) within the basic and hinge regions are defined. The additional conserved motifs and their presence of the group specificity were also identified. Further, we predicted the DNA-binding patterns and the dimerization property on the basis of the characteristic features in the basic and hinge regions and the leucine zipper, respectively, which supports our classification greatly and helps to classify 24 distinct subfamilies. Within the SlbZIP family, a total of 40 SlbZIP genes are located in the segmental duplicate regions in the tomato genome, suggesting that the segment chromosomal duplications contribute greatly to the expansion of the tomato SlbZIP family. Expression profiling analyses of 59 SlbZIP genes using quantitative RT-PCR and publicly available microarray data indicate that the tomato SlbZIP genes have distinct and diverse expression patterns in different tissues and developmental stages and many of the tomato bZIP genes

  12. Sugar sensing by ChREBP/Mondo-Mlx-new insight into downstream regulatory networks and integration of nutrient-derived signals.

    Science.gov (United States)

    Havula, Essi; Hietakangas, Ville

    2018-04-01

    Animals regulate their physiology with respect to nutrient status, which requires nutrient sensing pathways. Simple carbohydrates, sugars, are sensed by the basic-helix-loop-helix leucine zipper transcription factors ChREBP/Mondo, together with their heterodimerization partner Mlx, which are well-established activators of sugar-induced lipogenesis. Loss of ChREBP/Mondo-Mlx in mouse and Drosophila leads to sugar intolerance, that is, inability to survive on sugar containing diet. Recent evidence has revealed that ChREBP/Mondo-Mlx responds to sugar and fatty acid-derived metabolites through several mechanisms and cross-connects with other nutrient sensing pathways. ChREBP/Mondo-Mlx controls several downstream transcription factors and hormones, which mediate not only readjustment of metabolic pathways, but also control feeding behavior, intestinal digestion, and circadian rhythm. Copyright © 2017. Published by Elsevier Ltd.

  13. BACH transcription factors in innate and adaptive immunity.

    Science.gov (United States)

    Igarashi, Kazuhiko; Kurosaki, Tomohiro; Roychoudhuri, Rahul

    2017-07-01

    BTB and CNC homology (BACH) proteins are transcriptional repressors of the basic region leucine zipper (bZIP) transcription factor family. Recent studies indicate widespread roles of BACH proteins in controlling the development and function of the innate and adaptive immune systems, including the differentiation of effector and memory cells of the B and T cell lineages, CD4 + regulatory T cells and macrophages. Here, we emphasize similarities at a molecular level in the cell-type-specific activities of BACH factors, proposing that competitive interactions of BACH proteins with transcriptional activators of the bZIP family form a common mechanistic theme underlying their diverse actions. The findings contribute to a general understanding of how transcriptional repressors shape lineage commitment and cell-type-specific functions through repression of alternative lineage programmes.

  14. Leucine and HMB differentially modulate proteasome system in skeletal muscle under different sarcopenic conditions.

    Science.gov (United States)

    Baptista, Igor L; Silva, Willian J; Artioli, Guilherme G; Guilherme, Joao Paulo L F; Leal, Marcelo L; Aoki, Marcelo S; Miyabara, Elen H; Moriscot, Anselmo S

    2013-01-01

    In the present study we have compared the effects of leucine supplementation and its metabolite β-hydroxy-β-methyl butyrate (HMB) on the ubiquitin-proteasome system and the PI3K/Akt pathway during two distinct atrophic conditions, hindlimb immobilization and dexamethasone treatment. Leucine supplementation was able to minimize the reduction in rat soleus mass driven by immobilization. On the other hand, leucine supplementation was unable to provide protection against soleus mass loss in dexamethasone treated rats. Interestingly, HMB supplementation was unable to provide protection against mass loss in all treatments. While solely fiber type I cross sectional area (CSA) was protected in immobilized soleus of leucine-supplemented rats, none of the fiber types were protected by leucine supplementation in rats under dexamethasone treatment. In addition and in line with muscle mass results, HMB treatment did not attenuate CSA decrease in all fiber types against either immobilization or dexamethasone treatment. While leucine supplementation was able to minimize increased expression of both Mafbx/Atrogin and MuRF1 in immobilized rats, leucine was only able to minimize Mafbx/Atrogin in dexamethasone treated rats. In contrast, HMB was unable to restrain the increase in those atrogenes in immobilized rats, but in dexamethasone treated rats, HMB minimized increased expression of Mafbx/Atrogin. The amount of ubiquitinated proteins, as expected, was increased in immobilized and dexamethasone treated rats and only leucine was able to block this increase in immobilized rats but not in dexamethasone treated rats. Leucine supplementation maintained soleus tetanic peak force in immobilized rats at normal level. On the other hand, HMB treatment failed to maintain tetanic peak force regardless of treatment. The present data suggested that the anti-atrophic effects of leucine are not mediated by its metabolite HMB.

  15. Leucine and HMB differentially modulate proteasome system in skeletal muscle under different sarcopenic conditions.

    Directory of Open Access Journals (Sweden)

    Igor L Baptista

    Full Text Available In the present study we have compared the effects of leucine supplementation and its metabolite β-hydroxy-β-methyl butyrate (HMB on the ubiquitin-proteasome system and the PI3K/Akt pathway during two distinct atrophic conditions, hindlimb immobilization and dexamethasone treatment. Leucine supplementation was able to minimize the reduction in rat soleus mass driven by immobilization. On the other hand, leucine supplementation was unable to provide protection against soleus mass loss in dexamethasone treated rats. Interestingly, HMB supplementation was unable to provide protection against mass loss in all treatments. While solely fiber type I cross sectional area (CSA was protected in immobilized soleus of leucine-supplemented rats, none of the fiber types were protected by leucine supplementation in rats under dexamethasone treatment. In addition and in line with muscle mass results, HMB treatment did not attenuate CSA decrease in all fiber types against either immobilization or dexamethasone treatment. While leucine supplementation was able to minimize increased expression of both Mafbx/Atrogin and MuRF1 in immobilized rats, leucine was only able to minimize Mafbx/Atrogin in dexamethasone treated rats. In contrast, HMB was unable to restrain the increase in those atrogenes in immobilized rats, but in dexamethasone treated rats, HMB minimized increased expression of Mafbx/Atrogin. The amount of ubiquitinated proteins, as expected, was increased in immobilized and dexamethasone treated rats and only leucine was able to block this increase in immobilized rats but not in dexamethasone treated rats. Leucine supplementation maintained soleus tetanic peak force in immobilized rats at normal level. On the other hand, HMB treatment failed to maintain tetanic peak force regardless of treatment. The present data suggested that the anti-atrophic effects of leucine are not mediated by its metabolite HMB.

  16. Incorporation of [h]leucine and [h]valine into protein of freshwater bacteria: field applications.

    Science.gov (United States)

    Jørgensen, N O

    1992-11-01

    Incorporation of leucine and valine into proteins of freshwater bacteria as a measure of bacterial production was tested in two eutrophic Danish lakes and was related to bacterial production measured by thymidine incorporation. In a depth profile (0 to 8 m) in Frederiksborg Castle Lake, incorporation of 100 nM leucine and valine gave similar rates of protein production. In terms of carbon, this production was about 50% lower than incorporation of 10 nM thymidine. In another depth profile in the same lake, incorporations of 10 nM valine and 100 nM leucine were identical, but differed from incorporations of 10 nM leucine and 100 nM valine. Bacterial carbon production calculated from incorporations of 10 nM thymidine and 10 nM leucine was similar, whereas 10 nM valine and 100 nM leucine and valine indicated an up to 2.4-fold-higher rate of carbon production. In a diel study in Lake Bagsvaerd, incorporation of 100 nM leucine and valine indicated a similar protein production, but the calculated carbon production was about 1.9-fold higher than the production based on uptake of 10 nM thymidine. Different diel changes in incorporation of the two amino acids and in incorporation of thymidine were observed. In both lakes, concentrations of naturally occurring leucine and valine were activity of a H isotope added at a concentration of 100 nM usually was diluted a maximum of 5%. Net assimilation of natural free amino acids in the lakes sustained 8 to 69% of the net bacterial carbon requirement, estimated from incorporation of leucine, valine, or thymidine. The present results indicate that incorporation of leucine and valine permits realistic measurements of bacterial production in freshwater environments.

  17. Concentration-Dependent Patterns of Leucine Incorporation by Coastal Picoplankton

    Science.gov (United States)

    Alonso, Cecilia; Pernthaler, Jakob

    2006-01-01

    Coastal pelagic environments are believed to feature concentration gradients of dissolved organic carbon at a microscale, and they are characterized by pronounced seasonal differences in substrate availability for the heterotrophic picoplankton. Microbial taxa that coexist in such habitats might thus differ in their ability to incorporate substrates at various concentrations. We investigated the incorporation patterns of leucine in four microbial lineages from the coastal North Sea at concentrations between 0.1 and 100 nM before and during a spring phytoplankton bloom. Community bulk incorporation rates and the fraction of leucine-incorporating cells in the different populations were analyzed. Significantly fewer bacterial cells incorporated the amino acid before (13 to 35%) than during (23 to 47%) the bloom at all but the highest concentration. The incorporation rate per active cell in the prebloom situation was constant above 0.1 nM added leucine, whereas it increased steeply with substrate concentration during the bloom. At both time points, a high proportion of members of the Roseobacter clade incorporated leucine at all concentrations (55 to 80% and 86 to 94%, respectively). In contrast, the fractions of leucine-incorporating cells increased substantially with substrate availability in bacteria from the SAR86 clade (8 to 31%) and from DE cluster 2 of the Flavobacteria-Sphingobacteria (14 to 33%). The incorporation patterns of marine Euryarchaeota were between these extremes (30 to 56% and 48 to 70%, respectively). Our results suggest that the contribution of microbial taxa to the turnover of particular substrates may be concentration dependent. This may help us to understand the specific niches of coexisting populations that appear to compete for the same resources. PMID:16517664

  18. The Populus Class III HD ZIP, popREVOLUTA, influences cambium initiation and patterning of woody stems

    Science.gov (United States)

    Marcel Robischon; Juan Du; Eriko Miura; Andrew Groover

    2011-01-01

    The secondary growth of a woody stem requires the formation of a vascular cambium at an appropriate position and proper patterning of the vascular tissues derived from the cambium. Class III homeodomain-leucine zipper (HD ZIP) transcription factors have been implicated in polarity determination and patterning in lateral organs and primary vascular tissues and in the...

  19. Defect-mediated relaxation in the random tiling phase of a binary mixture: Birth, death and mobility of an atomic zipper

    Energy Technology Data Exchange (ETDEWEB)

    Tondl, Elisabeth; Ramsay, Malcolm; Harrowell, Peter; Widmer-Cooper, Asaph [School of Chemistry, University of Sydney, Sydney, NSW 2006 (Australia)

    2014-03-14

    This paper describes the mechanism of defect-mediated relaxation in a dodecagonal square-triangle random tiling phase exhibited by a simulated binary mixture of soft discs in 2D. We examine the internal transitions within the elementary mobile defect (christened the “zipper”) that allow it to move, as well as the mechanisms by which the zipper is created and annihilated. The structural relaxation of the random tiling phase is quantified and we show that this relaxation is well described by a model based on the distribution of waiting times for each atom to be visited by the diffusing zipper. This system, representing one of the few instances where a well defined mobile defect is capable of structural relaxation, can provide a valuable test case for general theories of relaxation in complex and disordered materials.

  20. Defect-mediated relaxation in the random tiling phase of a binary mixture: Birth, death and mobility of an atomic zipper

    International Nuclear Information System (INIS)

    Tondl, Elisabeth; Ramsay, Malcolm; Harrowell, Peter; Widmer-Cooper, Asaph

    2014-01-01

    This paper describes the mechanism of defect-mediated relaxation in a dodecagonal square-triangle random tiling phase exhibited by a simulated binary mixture of soft discs in 2D. We examine the internal transitions within the elementary mobile defect (christened the “zipper”) that allow it to move, as well as the mechanisms by which the zipper is created and annihilated. The structural relaxation of the random tiling phase is quantified and we show that this relaxation is well described by a model based on the distribution of waiting times for each atom to be visited by the diffusing zipper. This system, representing one of the few instances where a well defined mobile defect is capable of structural relaxation, can provide a valuable test case for general theories of relaxation in complex and disordered materials

  1. Results of a prospective randomised study comparing a non-invasive surgical zipper versus intracutaneous sutures for wound closure

    NARCIS (Netherlands)

    Roolker, W.; Kraaneveld, E.; Been, H. D.; Marti, R. K.

    2002-01-01

    A prospective randomised study was undertaken to investigate the advantages and disadvantages of a non-invasive surgical zipper (Medizip) vs intracutaneous sutures skin closure in orthopaedic surgery. The study group consisted of 120 consecutive patients, 45 men and 75 women with a mean age of 47

  2. The ontogeny of seizures induced by leucine-enkephalin and beta-endorphin.

    Science.gov (United States)

    Snead, O C; Stephens, H

    1984-06-01

    Rats ranging in postnatal age from 6 hours to 28 days were implanted with cortical and depth electrodes as well as an indwelling cannula in the lateral ventricle. We then administered varying amounts of the opiate peptides leucine-enkephalin and beta-endorphin intracerebroventricularly with continuous electroencephalographic monitoring. Leucine-enkephalin produced electrical seizure activity in rats as young as 2 days. beta-Endorphin administration was associated with seizures at the fifth postnatal day, with a high incidence of apnea resulting in death in animals as young as 6 hours. An adult seizure response to beta-endorphin and leucine-enkephalin was seen at 15 and 28 days of age, respectively. Naloxone blocked the seizure produced by these opiate peptides in all age groups. The data indicate that the opiate peptides are potent epileptogenic compounds in developing brain, that seizures induced by leucine-enkephalin differ from those caused by beta-endorphin, and that petit mal-like seizure activity can be an adult response in the rodent.

  3. Effect of burn and first-pass splanchnic leucine extraction on protein kinetics in rats

    International Nuclear Information System (INIS)

    Karlstad, M.D.; DeMichele, S.J.; Istfan, N.; Blackburn, G.L.; Bistrian, B.R.

    1988-01-01

    The effects of burn and first-pass splanchnic leucine extraction (FPE) on protein kinetics and energy expenditure were assessed by measuring O 2 consumption, CO 2 production, nitrogen balance, leucine kinetics, and tissue fractional protein synthetic rates (FSR-%/day) in enterally fed rats. Anesthetized male rats (200 g) were scalded on their dorsum with boiling water (25-30% body surface area) and enterally fed isovolemic diets that provided 60 kcal/day and 2.4 g of amino acids/day for 3 days. Controls were not burned. An intravenous or intragastric infusion of L-[1- 14 C]leucine was used to assess protein kinetics on day 3. FPE was taken as the ratio of intragastric to intravenous plasma leucine specific activity. There was a 69% reduction in cumulative nitrogen balance (P less than 0.001) and a 17-19% increase in leucine oxidation (P less than 0.05) and total energy expenditure (P less than 0.01) in burned rats. A 15% decrease in plasma leucine clearance (P less than 0.05) was accompanied by a 20% increase in plasma [leucine] (P less than 0.01) in burned rats. Burn decreased rectus muscle FSR from 5.0 +/- 0.4 to 3.5 +/- 0.5 (P less than 0.05) and increased liver FSR from 19.0 +/- 0.5 to 39.2 +/- 3.4 (P less than 0.01). First pass extraction of dietary leucine by the splanchnic bed was 8% in controls and 26% in burned rats. Leucine kinetics corrected for FPE showed increased protein degradation with burn that was not evident without FPE correction. This hypermetabolic burn model can be useful in the design of enteral diets that optimize rates of protein synthesis and degradation

  4. Application of Key Events Dose Response Framework to defining the upper intake level of leucine in young men.

    Science.gov (United States)

    Pencharz, Paul B; Russell, Robert M

    2012-12-01

    Leucine is sold in large doses in health food stores and is ingested by weight-training athletes. The safety of ingestion of large doses of leucine is unknown. Before designing chronic high-dose leucine supplementation experiments, we decided to determine the effect of graded doses of leucine in healthy participants. The Key Events Dose Response Framework is an organizational and analytical framework that dissects the various biologic steps (key events) that occur between exposure to a substance and an eventual adverse effect. Each biologic event is looked at for its unique dose-response characteristics. For nutrients, there are a number of biologic homeostatic mechanisms that work to keep circulating/tissue levels in a safe, nontoxic range. If a response mechanism at a particular key event is especially vulnerable and easily overwhelmed, this is known as a determining event, because this event drives the overall slope or shape of the dose-response relationship. In this paper, the Key Events Dose Framework has been applied to the problem of leucine toxicity and leucine's tolerable upper level. After analyzing the experimental data vis a vis key events for leucine leading to toxicity, it became evident that the rate of leucine oxidation was the determining event. A dose-response study has been conducted to graded intakes of leucine in healthy human adult male volunteers. All participants were started at the mean requirement level of leucine [50 mg/(kg · d)] and the highest leucine intake was 1250 mg/( kg · d), which is 25 times the mean requirement. No gut intolerance was seen. Blood glucose fell progressively but remained within normal values without any changes in plasma insulin. Maximal leucine oxidation levels occurred at an intake of 550 mg leucine/( kg · d), after which plasma leucine progressively increased and plasma ammonia also increased in response to leucine intakes >500 mg/( kg · d). Thus, the "key determining event" appears to be when the

  5. Measurement of L-[1-14C]leucine kinetics in splanchnic and leg tissues in humans. Effect of amino acid infusion

    International Nuclear Information System (INIS)

    Gelfand, R.A.; Glickman, M.G.; Castellino, P.; Louard, R.J.; DeFronzo, R.A.

    1988-01-01

    Although whole-body leucine flux is widely measured to study body protein turnover in humans, the contribution of specific tissues to the total-body measurement remains unknown. By combining the organ-balance technique with the systemic infusion of L-[1-14C]leucine, we quantitated leucine production and disposal by splanchnic and leg tissues and by the whole body, simultaneously, in six normal men before and during amino acid infusion. At steady state, disposal of arterial leucine by splanchnic and leg tissues was calculated from the percent extraction (E) of L-[1-14C]leucine counts: uptake = E x [Leu]a x flow. Tissue release of cold leucine (from protein turnover) into vein was calculated as the difference between leucine uptake and the net tissue leucine balance. In the postabsorptive state, despite substantial (P less than .01) extraction of L-[1-14C]leucine by splanchnic (23 +/- 1%) and leg (18 +/- 2%) tissues, net leucine balance across both tissue beds was small, indicating active simultaneous disposal and production of leucine at nearly equivalent rates. Splanchnic tissues accounted for approximately 50% of the measured total-body leucine flux. During amino acid infusion, the net leucine balance across splanchnic and leg tissues became positive, reflecting not only an increase in leucine uptake but also a marked suppression (by approximately 50%, P less than .02) of cold leucine release. This reduction in splanchnic and leg leucine release was indicated by a sharp decline in whole-body endogenous leucine flux

  6. Multimerization of the p12 domain is necessary for Mason–Pfizer monkey virus Gag assembly in vitro

    Czech Academy of Sciences Publication Activity Database

    Knejzlík, Z.; Smékalová, Z.; Ruml, Tomáš; Sakalian, M.

    2007-01-01

    Roč. 365, č. 2 (2007), s. 260-270 ISSN 0042-6822 R&D Projects: GA MŠk(CZ) 1M0520; GA ČR GESCO/06/E001 Grant - others:NIH(US) AI43230 Institutional research plan: CEZ:AV0Z40550506 Keywords : M-PMV * p12 protein * oligomerization * leucine zipper Subject RIV: CE - Biochemistry Impact factor: 3.765, year: 2007

  7. Nutritional leucine supplementation attenuates cardiac failure in tumour-bearing cachectic animals.

    Science.gov (United States)

    Toneto, Aline Tatiane; Ferreira Ramos, Luiz Alberto; Salomão, Emilianne Miguel; Tomasin, Rebeka; Aereas, Miguel Arcanjo; Gomes-Marcondes, Maria Cristina Cintra

    2016-12-01

    The condition known as cachexia presents in most patients with malignant tumours, leading to a poor quality of life and premature death. Although the cancer-cachexia state primarily affects skeletal muscle, possible damage in the cardiac muscle remains to be better characterized and elucidated. Leucine, which is a branched chain amino acid, is very useful for preserving lean body mass. Thus, this amino acid has been studied as a coadjuvant therapy in cachectic cancer patients, but whether this treatment attenuates the effects of cachexia and improves cardiac function remains poorly understood. Therefore, using an experimental cancer-cachexia model, we evaluated whether leucine supplementation ameliorates cachexia in the heart. Male Wistar rats were fed either a leucine-rich or a normoprotein diet and implanted or not with subcutaneous Walker-256 carcinoma. During the cachectic stage (approximately 21 days after tumour implantation), when the tumour mass was greater than 10% of body weight, the rats were subjected to an electrocardiogram analysis to evaluate the heart rate, QT-c, and T wave amplitude. The myocardial tissues were assayed for proteolytic enzymes (chymotrypsin, alkaline phosphatase, cathepsin, and calpain), cardiomyopathy biomarkers (myeloperoxidase, tissue inhibitor of metalloproteinases, and total plasminogen activator inhibitor 1), and caspase-8, -9, -3, and -7 activity. Both groups of tumour-bearing rats, especially the untreated group, had electrocardiography alterations that were suggestive of ischemia, dilated cardiomyopathy, and sudden death risk. Additionally, the rats in the untreated tumour-bearing group but not their leucine-supplemented littermates exhibited remarkable increases in chymotrypsin activity and all three heart failure biomarkers analysed, including an increase in caspase-3 and -7 activity. Our data suggest that a leucine-rich diet could modulate heart damage, cardiomyocyte proteolysis, and apoptosis driven by cancer

  8. Radiobiological half-lives for carbon-14 and hydrogen-3 leucine in man

    International Nuclear Information System (INIS)

    Classic, K.L.; Schwenk, W.F.; Haymond, M.W.

    1986-01-01

    In vivo estimates of protein metabolism in many are often made by oral or intravenous administration of leucine or its ∼-ketoacid, ∼-ketoisocaproate, labeled with 14 C or 3 H. Previous estimates of radiation dose from such tracers have been based on the measurement of 14 CO 2 in breath. Using measurements of the decay of 3 H or 14 C leucine from plasma proteins, longer biological half-lives for these compounds were obtained. The estimated total-body radiation absorbed dose is 0.97 mrad/uCi for [1- 14 C]KIC (or [1- 14 C]leucine) and 0.11 mrad/ + Ci for ]4,5- 3 H]leucine (or [ 3 H]KIC). Assuming administered doses of 100 μCi each, the total-body radiation absorbed dose is still well within the limits set by the FDA for Radioactive Drug Research Committees. 12 references, 3 figures, 3 tables

  9. Infusion and sampling site effects on two-pool model estimates of leucine metabolism

    International Nuclear Information System (INIS)

    Helland, S.J.; Grisdale-Helland, B.; Nissen, S.

    1988-01-01

    To assess the effect of site of isotope infusion on estimates of leucine metabolism infusions of alpha-[4,5-3H]ketoisocaproate (KIC) and [U- 14 C]leucine were made into the left or right ventricles of sheep and pigs. Blood was sampled from the opposite ventricle. In both species, left ventricular infusions resulted in significantly lower specific radioactivities (SA) of [ 14 C]leucine and [ 3 H]KIC. [ 14 C]KIC SA was found to be insensitive to infusion and sampling sites. [ 14 C]KIC was in addition found to be equal to the SA of [ 14 C]leucine only during the left heart infusions. Therefore, [ 14 C]KIC SA was used as the only estimate for [ 14 C]SA in the equations for the two-pool model. This model eliminated the influence of site of infusion and blood sampling on the estimates for leucine entry and reduced the impact on the estimates for proteolysis and oxidation. This two-pool model could not compensate for the underestimation of transamination reactions occurring during the traditional venous isotope infusion and arterial blood sampling

  10. Methodological investigation on the use of 14C-leucine and 15N-leucine for studying the absorption of amino acids in the experimental rat

    International Nuclear Information System (INIS)

    Bergner, H.; Bergner, U.; Adam, K.

    1980-01-01

    After nine days of adaptation to a whole-egg diet, albino rats were given 14 C-U-L-leucine and 15 N-L-leucine in addition by the oral route. Each rat received the labelled leucine via a pellet made from the whole-egg diet after food deprivation for 15 h. Thereafter, the experimental animals consumed the unlabelled experimental diet ad libitum. Four times, 30 min, and 1, 2, 4 and 8 h after ingestion of the labelled food, four experimental rats were sacrificed. The contents of the digestive tract and tissue samples were examined for 14 C and 15 N. The halftime of disappearance of the 14 C activity and of the 15 N excess from the TCA-soluble fraction of the gastric contents lay between 1.9 and 2.2 h. Up to the fourth hour of experiment, the 15 N level of the TCA-soluble fraction of the gastric contents was high. The free leucine is obviously absorbed in the stomach and is used for the synthesis of enzyme protein and mucoproteides. In the TCA-soluble fraction of the total contents of the small intestine, the following values (expressed as percentages of the total amounts ingested at the times of measurement) were found: 14 C = 2.0; 6.5; 9.6; 7.4 and 1.5; 15 N excess = 0.8; 1.2; 1.6; 1.6 and 1.2 Were these values regarded as non-absorbed leucine, the 14 C values obtained during the one-to-four hour period of experiment would unequivocally be too high. Presumably, they are simulated by other 14 C-metabolites which originate from the leucine catabolism and reach the intestinal lumen through the intestinal wall. Amino acids labelled with 15 N should be preferred in studies on the absorption of amino acids because, in case of catabolization, the 15 N-amino group is excreted mainly in the form of urea. 14 C-amino acids can be recommended for such studies only if the specific 14 C activity of the amino acid used is also measured. (author)

  11. Leucine Modulates Mitochondrial Biogenesis and SIRT1-AMPK Signaling in C2C12 Myotubes

    Directory of Open Access Journals (Sweden)

    Chunzi Liang

    2014-01-01

    Full Text Available Previous studies from this laboratory demonstrate that dietary leucine protects against high fat diet-induced mitochondrial impairments and stimulates mitochondrial biogenesis and energy partitioning from adipocytes to muscle cells through SIRT1-mediated mechanisms. Moreover, β-hydroxy-β-methyl butyrate (HMB, a metabolite of leucine, has been reported to activate AMPK synergistically with resveratrol in C2C12 myotubes. Therefore, we hypothesize that leucine-induced activation of SIRT1 and AMPK is the central event that links the upregulated mitochondrial biogenesis and fatty acid oxidation in skeletal muscle. Thus, C2C12 myotubes were treated with leucine (0.5 mM, alanine (0.5 mM, valine (0.5 mM, EX527 (SIRT1 inhibitor, 25 μM, and Compound C (AMPK inhibitor, 25 μM alone or in combination to determine the roles of AMPK and SIRT1 in leucine-modulation of energy metabolism. Leucine significantly increased mitochondrial content, mitochondrial biogenesis-related genes expression, fatty acid oxidation, SIRT1 activity and gene expression, and AMPK phosphorylation in C2C12 myotubes compared to the controls, while EX527 and Compound C markedly attenuated these effects. Furthermore, leucine treatment for 24 hours resulted in time-dependent increases in cellular NAD+, SIRT1 activity, and p-AMPK level, with SIRT1 activation preceding that of AMPK, indicating that leucine activation of SIRT1, rather than AMPK, is the primary event.

  12. Expression, purification, crystallization and preliminary X-ray analysis of a C-terminal fragment of the Epstein–Barr virus ZEBRA protein

    Energy Technology Data Exchange (ETDEWEB)

    Morand, Patrice [European Molecular Biology Laboratory, Grenoble Outstation, BP 181, 38042 Grenoble CEDEX 9 (France); Laboratoire de Virologie Moléculaire et Structurale, EA 2939, Université Joseph Fourier, Grenoble (France); Budayova-Spano, Monika [European Molecular Biology Laboratory, Grenoble Outstation, BP 181, 38042 Grenoble CEDEX 9 (France); Perrissin, Monique [Laboratoire de Virologie Moléculaire et Structurale, EA 2939, Université Joseph Fourier, Grenoble (France); Müller, Christoph W., E-mail: mueller@embl-grenoble.fr; Petosa, Carlo [European Molecular Biology Laboratory, Grenoble Outstation, BP 181, 38042 Grenoble CEDEX 9 (France)

    2006-03-01

    A C-terminal fragment of the Epstein–Barr virus lytic switch protein ZEBRA has been crystallized in complex with DNA. A C-terminal fragment of the Epstein–Barr virus immediate-early transcription factor ZEBRA has been expressed as a recombinant protein in Escherichia coli and purified to homogeneity. The fragment behaves as a dimer in solution, consistent with the presence of a basic region leucine-zipper (bZIP) domain. Crystals of the fragment in complex with a DNA duplex were grown by the hanging-drop vapour-diffusion technique using polyethylene glycol 4000 and magnesium acetate as crystallization agents. Crystals diffract to better than 2.5 Å resolution using synchrotron radiation (λ = 0.976 Å). Crystals belong to space group C2, with unit-cell parameters a = 94.2, b = 26.5, c = 98.1 Å, β = 103.9°.

  13. Reviewing the Effects of l-Leucine Supplementation in the Regulation of Food Intake, Energy Balance, and Glucose Homeostasis

    Directory of Open Access Journals (Sweden)

    João A.B. Pedroso

    2015-05-01

    Full Text Available Leucine is a well-known activator of the mammalian target of rapamycin (mTOR. Because mTOR signaling regulates several aspects of metabolism, the potential of leucine as a dietary supplement for treating obesity and diabetes mellitus has been investigated. The objective of the present review was to summarize and discuss the available evidence regarding the mechanisms and the effects of leucine supplementation on the regulation of food intake, energy balance, and glucose homeostasis. Based on the available evidence, we conclude that although central leucine injection decreases food intake, this effect is not well reproduced when leucine is provided as a dietary supplement. Consequently, no robust evidence indicates that oral leucine supplementation significantly affects food intake, although several studies have shown that leucine supplementation may help to decrease body adiposity in specific conditions. However, more studies are necessary to assess the effects of leucine supplementation in already-obese subjects. Finally, although several studies have found that leucine supplementation improves glucose homeostasis, the underlying mechanisms involved in these potential beneficial effects remain unknown and may be partially dependent on weight loss.

  14. Investigation on the absorption of 14C-leucine and 15N-leucine in rats after feeding a fish meal diet in comparison with a gelatine diet

    International Nuclear Information System (INIS)

    Bergner, U.; Adam, K.; Bergner, H.

    1981-01-01

    Albino rats received after nine days of adaptation to a fish meal diet in comparison with a gelatine diet 14 C-U-L-leucine and 15 N-L-leucine via a pellet made from the specific diet after food deprivation for 15 h. Thereafter, the animals consumed the non-labelled experimental diet ad libitum. 30 min, and 1, 2, 4 and 8 h, resp., after intake of the labelled food, four rats at a time were sacrificed. The contents of the digestive tract and tissue samples were examined for 14 C and 15 N and their percentages in the TCA-soluble fraction determined. If these values are regarded as non-absorbed leucine, the 14 C values obtained up to the four hour period of the experiment would be too high. Presumably, they are in the case of both diets simulated by other 14 C metabolites which originate from the leucine catabolism and reach the intestinal lumen. Amino acids labelled with 15 N should be preferred in studies on the absorption of amino acids because, in case of catabolization, the 15 N aminogroup is excreted mainly as urea via urine. (author)

  15. Differential effects of leucine and leucine-enriched whey protein on skeletal muscle protein synthesis in aged mice

    NARCIS (Netherlands)

    Dijk, Francina J.; Dijk, van Miriam; Walrand, Stéphane; Loon, van Luc J.C.; Norren, van Klaske; Luiking, Yvette C.

    2018-01-01

    Background & aims: It has been suggested that anabolic resistance, or a blunted protein synthetic response to anabolic stimuli, contributes to the failure of muscle mass maintenance in older adults. The amino acid leucine is one of the most prominent food-related anabolic stimuli. However, data

  16. Leucine-based receptor sorting motifs are dependent on the spacing relative to the plasma membrane

    DEFF Research Database (Denmark)

    Geisler, C; Dietrich, J; Nielsen, B L

    1998-01-01

    Many integral membrane proteins contain leucine-based motifs within their cytoplasmic domains that mediate internalization and intracellular sorting. Two types of leucine-based motifs have been identified. One type is dependent on phosphorylation, whereas the other type, which includes an acidic...... amino acid, is constitutively active. In this study, we have investigated how the spacing relative to the plasma membrane affects the function of both types of leucine-based motifs. For phosphorylation-dependent leucine-based motifs, a minimal spacing of 7 residues between the plasma membrane...... and the phospho-acceptor was required for phosphorylation and thereby activation of the motifs. For constitutively active leucine-based motifs, a minimal spacing of 6 residues between the plasma membrane and the acidic residue was required for optimal activity of the motifs. In addition, we found that the acidic...

  17. Radiation-induced tetramer-to-dimer transition of Escherichia coli lactose repressor

    International Nuclear Information System (INIS)

    Goffinont, S.; Davidkova, M.; Spotheim-Maurizot, M.

    2009-01-01

    The wild type lactose repressor of Escherichia coli is a tetrameric protein formed by two identical dimers. They are associated via a C-terminal 4-helix bundle (called tetramerization domain) whose stability is ensured by the interaction of leucine zipper motifs. Upon in vitro γ-irradiation the repressor losses its ability to bind the operator DNA sequence due to damage of its DNA-binding domains. Using an engineered dimeric repressor for comparison, we show here that irradiation induces also the change of repressor oligomerisation state from tetramer to dimer. The splitting of the tetramer into dimers can result from the oxidation of the leucine residues of the tetramerization domain.

  18. The perennial ryegrass GenomeZipper: targeted use of genome resources for comparative grass genomics.

    Science.gov (United States)

    Pfeifer, Matthias; Martis, Mihaela; Asp, Torben; Mayer, Klaus F X; Lübberstedt, Thomas; Byrne, Stephen; Frei, Ursula; Studer, Bruno

    2013-02-01

    Whole-genome sequences established for model and major crop species constitute a key resource for advanced genomic research. For outbreeding forage and turf grass species like ryegrasses (Lolium spp.), such resources have yet to be developed. Here, we present a model of the perennial ryegrass (Lolium perenne) genome on the basis of conserved synteny to barley (Hordeum vulgare) and the model grass genome Brachypodium (Brachypodium distachyon) as well as rice (Oryza sativa) and sorghum (Sorghum bicolor). A transcriptome-based genetic linkage map of perennial ryegrass served as a scaffold to establish the chromosomal arrangement of syntenic genes from model grass species. This scaffold revealed a high degree of synteny and macrocollinearity and was then utilized to anchor a collection of perennial ryegrass genes in silico to their predicted genome positions. This resulted in the unambiguous assignment of 3,315 out of 8,876 previously unmapped genes to the respective chromosomes. In total, the GenomeZipper incorporates 4,035 conserved grass gene loci, which were used for the first genome-wide sequence divergence analysis between perennial ryegrass, barley, Brachypodium, rice, and sorghum. The perennial ryegrass GenomeZipper is an ordered, information-rich genome scaffold, facilitating map-based cloning and genome assembly in perennial ryegrass and closely related Poaceae species. It also represents a milestone in describing synteny between perennial ryegrass and fully sequenced model grass genomes, thereby increasing our understanding of genome organization and evolution in the most important temperate forage and turf grass species.

  19. Leucine elicits myotube hypertrophy and enhances maximal contractile force in tissue engineered skeletal muscle in vitro.

    Science.gov (United States)

    Martin, Neil R W; Turner, Mark C; Farrington, Robert; Player, Darren J; Lewis, Mark P

    2017-10-01

    The amino acid leucine is thought to be important for skeletal muscle growth by virtue of its ability to acutely activate mTORC1 and enhance muscle protein synthesis, yet little data exist regarding its impact on skeletal muscle size and its ability to produce force. We utilized a tissue engineering approach in order to test whether supplementing culture medium with leucine could enhance mTORC1 signaling, myotube growth, and muscle function. Phosphorylation of the mTORC1 target proteins 4EBP-1 and rpS6 and myotube hypertrophy appeared to occur in a dose dependent manner, with 5 and 20 mM of leucine inducing similar effects, which were greater than those seen with 1 mM. Maximal contractile force was also elevated with leucine supplementation; however, although this did not appear to be enhanced with increasing leucine doses, this effect was completely ablated by co-incubation with the mTOR inhibitor rapamycin, showing that the augmented force production in the presence of leucine was mTOR sensitive. Finally, by using electrical stimulation to induce chronic (24 hr) contraction of engineered skeletal muscle constructs, we were able to show that the effects of leucine and muscle contraction are additive, since the two stimuli had cumulative effects on maximal contractile force production. These results extend our current knowledge of the efficacy of leucine as an anabolic nutritional aid showing for the first time that leucine supplementation may augment skeletal muscle functional capacity, and furthermore validates the use of engineered skeletal muscle for highly-controlled investigations into nutritional regulation of muscle physiology. © 2017 The Authors. Journal of Cellular Physiology Published by wiley periodicals, Inc.

  20. The actions of exogenous leucine on mTOR signalling and amino acid transporters in human myotubes

    Directory of Open Access Journals (Sweden)

    Cameron-Smith David

    2011-06-01

    Full Text Available Abstract Background The branched-chain amino acid (BCAA leucine has been identified to be a key regulator of skeletal muscle anabolism. Activation of anabolic signalling occurs via the mammalian target of rapamycin (mTOR through an undefined mechanism. System A and L solute carriers transport essential amino acids across plasma membranes; however it remains unknown whether an exogenous supply of leucine regulates their gene expression. The aim of the present study was to investigate the effects of acute and chronic leucine stimulation of anabolic signalling and specific amino acid transporters, using cultured primary human skeletal muscle cells. Results Human myotubes were treated with leucine, insulin or co-treated with leucine and insulin for 30 min, 3 h or 24 h. Activation of mTOR signalling kinases were examined, together with putative nutrient sensor human vacuolar protein sorting 34 (hVps34 and gene expression of selected amino acid transporters. Phosphorylation of mTOR and p70S6K was transiently increased following leucine exposure, independently to insulin. hVps34 protein expression was also significantly increased. However, genes encoding amino acid transporters were differentially regulated by insulin and not leucine. Conclusions mTOR signalling is transiently activated by leucine within human myotubes independently of insulin stimulation. While this occurred in the absence of changes in gene expression of amino acid transporters, protein expression of hVps34 increased.

  1. Impaired intracortical transmission in G2019S leucine rich-repeat kinase Parkinson patients.

    Science.gov (United States)

    Ponzo, Viviana; Di Lorenzo, Francesco; Brusa, Livia; Schirinzi, Tommaso; Battistini, Stefania; Ricci, Claudia; Sambucci, Manolo; Caltagirone, Carlo; Koch, Giacomo

    2017-05-01

    A mutation in leucine-rich repeat kinase 2 is the most common cause of hereditary Parkinson's disease (PD), yet the neural mechanisms and the circuitry potentially involved are poorly understood. We used different transcranial magnetic stimulation protocols to explore in the primary motor cortex the activity of intracortical circuits and cortical plasticity (long-term potentiation) in patients with the G2019S leucine-rich repeat kinase 2 gene mutation when compared with idiopathic PD patients and age-matched healthy subjects. Paired pulse transcranial magnetic stimulation was used to investigate short intracortical inhibition and facilitation and short afferent inhibition. Intermittent theta burst stimulation, a form of repetitive transcranial magnetic stimulation, was used to test long-term potentiation-like cortical plasticity. Leucine-rich repeat kinase 2 and idiopathic PD were tested both in ON and in OFF l-dopa therapy. When compared with idiopathic PD and healthy subjects, leucine-rich repeat kinase 2 PD patients showed a remarkable reduction of short intracortical inhibition in both ON and in OFF l-dopa therapy. This reduction was paralleled by an increase of intracortical facilitation in OFF l-dopa therapy. Leucine-rich repeat kinase 2 PD showed abnormal long-term potentiation-like cortical plasticity in ON l-dopa therapy. The motor cortex in leucine-rich repeat kinase 2 mutated PD patients is strongly disinhibited and hyperexcitable. These abnormalities could be a result of an impairment of inhibitory (gamma-Aminobutyric acid) transmission eventually related to altered neurotransmitter release. © 2017 International Parkinson and Movement Disorder Society. © 2017 International Parkinson and Movement Disorder Society.

  2. Use of the [(14)C]leucine incorporation technique to measure bacterial production in river sediments and the epiphyton.

    Science.gov (United States)

    Fischer, H; Pusch, M

    1999-10-01

    Bacterial production is a key parameter for the understanding of carbon cycling in aquatic ecosystems, yet it remains difficult to measure in many aquatic habitats. We therefore tested the applicability of the [(14)C]leucine incorporation technique for the measurement of bulk bacterial production in various habitats of a lowland river ecosystem. To evaluate the method, we determined (i) extraction efficiencies of bacterial protein from the sediments, (ii) substrate saturation of leucine in sediments, the biofilms on aquatic plants (epiphyton), and the pelagic zone, (iii) bacterial activities at different leucine concentrations, (iv) specificity of leucine uptake by bacteria, and (v) the effect of the incubation technique (perfused-core incubation versus slurry incubation) on leucine incorporation into protein. Bacterial protein was best extracted from sediments and precipitated by hot trichloroacetic acid treatment following ultrasonication. For epiphyton, an alkaline-extraction procedure was most efficient. Leucine incorporation saturation occurred at 1 microM in epiphyton and 100 nM in the pelagic zone. Saturation curves in sediments were difficult to model but showed the first level of leucine saturation at 50 microM. Increased uptake at higher leucine concentrations could be partly attributed to eukaryotes. Addition of micromolar concentrations of leucine did not enhance bacterial electron transport activity or DNA replication activity. Similar rates of leucine incorporation into protein calculated for whole sediment cores were observed after slurry and perfused-core incubations, but the rates exhibited strong vertical gradients after the core incubation. We conclude that the leucine incorporation method can measure bacterial production in a wide range of aquatic habitats, including fluvial sediments, if substrate saturation and isotope dilution are determined.

  3. Use of the [14C]Leucine Incorporation Technique To Measure Bacterial Production in River Sediments and the Epiphyton

    Science.gov (United States)

    Fischer, Helmut; Pusch, Martin

    1999-01-01

    Bacterial production is a key parameter for the understanding of carbon cycling in aquatic ecosystems, yet it remains difficult to measure in many aquatic habitats. We therefore tested the applicability of the [14C]leucine incorporation technique for the measurement of bulk bacterial production in various habitats of a lowland river ecosystem. To evaluate the method, we determined (i) extraction efficiencies of bacterial protein from the sediments, (ii) substrate saturation of leucine in sediments, the biofilms on aquatic plants (epiphyton), and the pelagic zone, (iii) bacterial activities at different leucine concentrations, (iv) specificity of leucine uptake by bacteria, and (v) the effect of the incubation technique (perfused-core incubation versus slurry incubation) on leucine incorporation into protein. Bacterial protein was best extracted from sediments and precipitated by hot trichloroacetic acid treatment following ultrasonication. For epiphyton, an alkaline-extraction procedure was most efficient. Leucine incorporation saturation occurred at 1 μM in epiphyton and 100 nM in the pelagic zone. Saturation curves in sediments were difficult to model but showed the first level of leucine saturation at 50 μM. Increased uptake at higher leucine concentrations could be partly attributed to eukaryotes. Addition of micromolar concentrations of leucine did not enhance bacterial electron transport activity or DNA replication activity. Similar rates of leucine incorporation into protein calculated for whole sediment cores were observed after slurry and perfused-core incubations, but the rates exhibited strong vertical gradients after the core incubation. We conclude that the leucine incorporation method can measure bacterial production in a wide range of aquatic habitats, including fluvial sediments, if substrate saturation and isotope dilution are determined. PMID:10508068

  4. Chronic leucine supplementation improves glycemic control in etiologically distinct mouse models of obesity and diabetes mellitus

    Directory of Open Access Journals (Sweden)

    Hou Jue

    2010-07-01

    Full Text Available Abstract Background Leucine may function as a signaling molecule to regulate metabolism. We have previously shown that dietary leucine supplementation significantly improves glucose and energy metabolism in diet-induced obese mice, suggesting that leucine supplementation could potentially be a useful adjuvant therapy for obesity and type 2 diabetes. Since the underlying cause for obesity and type 2 diabetes is multifold, we further investigated metabolic effects of leucine supplementation in obese/diabetes mouse models with different etiologies, and explored the underlying molecular mechanisms. Methods Leucine supplementation was carried out in NONcNZO10/LtJ (RCS10 - a polygenic model predisposed to beta cell failure and type 2 diabetes, and in B6.Cg-Ay/J (Ay - a monogenic model for impaired central melanocortin receptor signaling, obesity, and severe insulin resistance. Mice in the treatment group received the drinking water containing 1.5% leucine for up to 8 months; control mice received the tap water. Body weight, body composition, blood HbA1c levels, and plasma glucose and insulin levels were monitored throughout and/or at the end of the study period. Indirect calorimetry, skeletal muscle gene expression, and adipose tissue inflammation were also assessed in Ay mice. Results Leucine supplementation significantly reduced HbA1c levels throughout the study period in both RCS10 and Ay mice. However, the treatment had no long term effect on body weight or adiposity. The improvement in glycemic control was associated with an increased insulin response to food challenge in RCS10 mice and decreased plasma insulin levels in Ay mice. In leucine-treated Ay mice, energy expenditure was increased by ~10% (p y mice whereas the expression levels of MCP-1 and TNF-alpha and macrophage infiltration in adipose tissue were significantly reduced. Conclusions Chronic leucine supplementation significantly improves glycemic control in multiple mouse models of

  5. Investigation on the absorption of /sup 14/C-leucine and /sup 15/N-leucine in rats after feeding a fish meal diet in comparison with a gelatine diet

    Energy Technology Data Exchange (ETDEWEB)

    Bergner, U; Adam, K; Bergner, H [Humboldt-Universitaet, Berlin (German Democratic Republic). Sektion Tierproduktion und Veterinaermedizin

    1981-01-01

    Albino rats received after nine days of adaptation to a fish meal diet in comparison with a gelatine diet /sup 14/C-U-L-leucine and /sup 15/N-L-leucine via a pellet made from the specific diet after food deprivation for 15 h. Thereafter, the animals consumed the non-labelled experimental diet ad libitum. 30 min, and 1, 2, 4 and 8 h, resp., after intake of the labelled food, four rats at a time were sacrificed. The contents of the digestive tract and tissue samples were examined for /sup 14/C and /sup 15/N and their percentages in the TCA-soluble fraction determined. If these values are regarded as non-absorbed leucine, the /sup 14/C values obtained up to the four hour period of the experiment would be too high. Presumably, they are in the case of both diets simulated by other /sup 14/C metabolites which originate from the leucine catabolism and reach the intestinal lumen. Amino acids labelled with /sup 15/N should be preferred in studies on the absorption of amino acids because, in case of catabolization, the /sup 15/N aminogroup is excreted mainly as urea via urine.

  6. Leucine incorporation into mixed skeletal muscle protein in humans

    International Nuclear Information System (INIS)

    Nair, K.S.; Halliday, D.; Griggs, R.C.

    1988-01-01

    Fractional mixed skeletal muscle protein synthesis (FMPS) was estimated in 10 postabsorptive healthy men by determining the increment in the abundance of [ 13 C]-leucine in quadriceps muscle protein during an intravenous infusion of L-[1- 13 C]leucine. Whole-body muscle protein synthesis (MPS) was calculated based on the estimation of muscle mass from creatinine excretion and compared with whole-body protein synthesis (WBPS) calculated from the nonoxidative portion of leucine flux. A significant correlation was found between MPS. The contribution of MPS to WBPS was 27 ± 1%, which is comparable to the reports in other species. Morphometric analyses of adjacent muscle samples in eight subjects demonstrated that the biopsy specimens consisted of 86.5 ± 2% muscular as opposed to other tissues. Because fiber type composition varies between biopsies, the authors examined the relationship between proportions of each fiber type and FMPS. Variation in the composition of biopsies and in fiber-type proportion did not affect the estimation of muscle protein synthesis rate. They conclude that stable isotope techniques using serial needle biopsies permit the direct measurement of FMPS in humans and that this estimation is correlated with an indirect estimation of WBPS

  7. Increased dependence of leucine in posttraumatic sepsis: leucine/tyrosine clearance ratio as an indicator of hepatic impairment in septic multiple organ failure syndrome.

    Science.gov (United States)

    Pittiruti, M; Siegel, J H; Sganga, G; Coleman, B; Wiles, C E; Belzberg, H; Wedel, S; Placko, R

    1985-09-01

    The body clearance of 10 plasma amino acids (AA) was determined from the rate of compared muscle-released AA and AA administered by infusion of total parenteral nutrition (TPN) compared to their estimated extracellular (ECW) pool in patients with multiple trauma with (n = 10) or without (n = 16) sepsis at 8-hour intervals. In both nonseptic and septic trauma, increasing TPN increased the mean clearance rate of all infused AA. When the individual AA clearance rates were normalized by the total AA infusion rate, regression-covariance analysis revealed that patients with sepsis had relatively impaired clearances of alanine (p less than 0.01) and methionine, proline, phenylalanine, and tyrosine p less than 0.05 for all). In contrast, the clearances of branched-chain AA (BCAA) valine and isoleucine were maintained, and the clearance of leucine was higher (p less than 0.05) in trauma patients with sepsis than in those without. At any AA infusion rate, compared with surviving patients with sepsis (p less than 0.05), patients who developed fatal multiple organ failure syndrome (MOFS) showed increased clearances of all BCAA with further impaired clearance of tyrosine. The clearance ratio of leucine/tyrosine was increased in MOFS at any AA infusion rate (p less than 0.0001), was an indicator of severity, and, if persistent, was a manifestation of a fatal outcome. Because tyrosine metabolism occurs almost entirely in the liver while leucine can be utilized by viscera and muscle, these data suggest early and progressive septic impairment of the pattern of hepatic uptake and oxidation of AA with a greater body dependence on BCAA, especially leucine, as septic MOFS develops.

  8. Leucine deprivation stimulates fat loss via increasing CRH expression in the hypothalamus and activating the sympathetic nervous system.

    Science.gov (United States)

    Cheng, Ying; Zhang, Qian; Meng, Qingshu; Xia, Tingting; Huang, Zhiying; Wang, Chunxia; Liu, Bin; Chen, Shanghai; Xiao, Fei; Du, Ying; Guo, Feifan

    2011-09-01

    We previously showed that leucine deprivation decreases abdominal fat mass largely by increasing energy expenditure, as demonstrated by increased lipolysis in white adipose tissue (WAT) and uncoupling protein 1 (UCP1) expression in brown adipose tissue (BAT). The goal of the present study was to investigate the possible involvement of central nervous system (CNS) in this regulation and elucidate underlying molecular mechanisms. For this purpose, levels of genes and proteins related to lipolysis in WAT and UCP1 expression in BAT were analyzed in wild-type mice after intracerebroventricular administration of leucine or corticotrophin-releasing hormone antibodies, or in mice deleted for three β-adrenergic receptors, after being maintained on a leucine-deficient diet for 7 d. Here, we show that intracerebroventricular administration of leucine significantly attenuates abdominal fat loss and blocks activation of hormone sensitive lipase in WAT and induction of UCP1 in BAT in leucine-deprived mice. Furthermore, we provide evidence that leucine deprivation stimulates fat loss by increasing expression of corticotrophin-releasing hormone in the hypothalamus via activation of stimulatory G protein/cAMP/protein kinase A/cAMP response element-binding protein pathway. Finally, we show that the effect of leucine deprivation on fat loss is mediated by activation of the sympathetic nervous system. These results suggest that CNS plays an important role in regulating fat loss under leucine deprivation and thereby provide novel and important insights concerning the importance of CNS leucine in the regulation of energy homeostasis.

  9. Biophysical properties of regions flanking the bHLH-Zip motif in the p22 Max protein

    International Nuclear Information System (INIS)

    Pursglove, Sharon E.; Fladvad, Malin; Bellanda, Massimo; Moshref, Ahmad; Henriksson, Marie; Carey, Jannette; Sunnerhagen, Maria

    2004-01-01

    The Max protein is the central dimerization partner in the Myc-Max-Mad network of transcriptional regulators, and a founding structural member of the family of basic-helix-loop-helix (bHLH)-leucine zipper (Zip) proteins. Biologically important regions flanking its bHLH-Zip motif have been disordered or absent in crystal structures. The present study shows that these regions are resistant to proteolysis in both the presence and absence of DNA, and that Max dimers containing both flanking regions have significantly higher helix content as measured by circular dichroism than that predicted from the crystal structures. Nuclear magnetic resonance measurements in the absence of DNA also support the inferred structural order. Deletion of both flanking regions is required to achieve maximal DNA affinity as measured by EMSA. Thus, the previously observed functionalities of these Max regions in DNA binding, phosphorylation, and apoptosis are suggested to be linked to structural properties

  10. The Role of Nrf2: Adipocyte Differentiation, Obesity, and Insulin Resistance

    Directory of Open Access Journals (Sweden)

    Hyun-Ae Seo

    2013-01-01

    Full Text Available Metabolic diseases, such as type 2 diabetes and obesity, are increasing globally, and much work has been performed to elucidate the regulatory mechanisms of these diseases. Nuclear factor E2-related factor 2 (Nrf2 is a basic leucine zipper transcription factor that serves as a primary cellular defense against the cytotoxic effects of oxidative stress. Recent studies have proposed a close relationship between oxidative stress and energy metabolism-associated disease. The Nrf2 pathway, as a master regulator of cellular defense against oxidative stress, has emerged as a critical target of energy metabolism; however, its effects are controversial. This review examines the current state of research on the role of Nrf2 on energy metabolism, specifically with respect to its participation in adipocyte differentiation, obesity, and insulin resistance, and discusses the possibility of using Nrf2 as a therapeutic target in the clinic.

  11. Plant transducers of the endoplasmic reticulum unfolded protein response

    KAUST Repository

    Iwata, Yuji; Koizumi, Nozomu

    2012-01-01

    The unfolded protein response (UPR) activates a set of genes to overcome accumulation of unfolded proteins in the endoplasmic reticulum (ER), a condition termed ER stress, and constitutes an essential part of ER protein quality control that ensures efficient maturation of secretory and membrane proteins in eukaryotes. Recent studies on Arabidopsis and rice identified the signaling pathway in which the ER membrane-localized ribonuclease IRE1 (inositol-requiring enzyme 1) catalyzes unconventional cytoplasmic splicing of mRNA, thereby producing the active transcription factor Arabidopsis bZIP60 (basic leucine zipper 60) and its ortholog in rice. Here we review recent findings identifying the molecular components of the plant UPR, including IRE1/bZIP60 and the membrane-bound transcription factors bZIP17 and bZIP28, and implicating its importance in several physiological phenomena such as pathogen response. © 2012 Elsevier Ltd.

  12. Plant transducers of the endoplasmic reticulum unfolded protein response

    KAUST Repository

    Iwata, Yuji

    2012-12-01

    The unfolded protein response (UPR) activates a set of genes to overcome accumulation of unfolded proteins in the endoplasmic reticulum (ER), a condition termed ER stress, and constitutes an essential part of ER protein quality control that ensures efficient maturation of secretory and membrane proteins in eukaryotes. Recent studies on Arabidopsis and rice identified the signaling pathway in which the ER membrane-localized ribonuclease IRE1 (inositol-requiring enzyme 1) catalyzes unconventional cytoplasmic splicing of mRNA, thereby producing the active transcription factor Arabidopsis bZIP60 (basic leucine zipper 60) and its ortholog in rice. Here we review recent findings identifying the molecular components of the plant UPR, including IRE1/bZIP60 and the membrane-bound transcription factors bZIP17 and bZIP28, and implicating its importance in several physiological phenomena such as pathogen response. © 2012 Elsevier Ltd.

  13. Phylogenetic analysis of F-bZIP transcription factors indicates conservation of the zinc deficiency response across land plants

    DEFF Research Database (Denmark)

    Castro, Pedro Humberto Araújo R F; Lilay, Grmay Hailu; Muñoz-Mérida, Antonio

    2017-01-01

    Basic leucine zipper (bZIP) transcription factors control important developmental and physiological processes in plants. In Arabidopsis thaliana, the three gene F-bZIP subfamily has been associated with zinc deficiency and salt stress response. Benefiting from the present abundance of plant genomic...... data, we performed an evolutionary and structural characterization of plant F-bZIPs. We observed divergence during seed plant evolution, into two groups and inferred different selective pressures for each. Group 1 contains AtbZIP19 and AtbZIP23 and appears more conserved, whereas Group 2, containing...... of AtZIP4. A survey of AtZIP4 orthologs promoters across different plant taxa revealed an enrichment of the Zinc Deficiency Response Element (ZDRE) to which both AtbZIP19/23 bind. Overall, our results indicate that while the AtbZIP24 function in the regulation of the salt stress response may...

  14. N-Acetyl-L-Leucine Accelerates Vestibular Compensation after Unilateral Labyrinthectomy by Action in the Cerebellum and Thalamus

    Science.gov (United States)

    Xiong, Guoming; Potschka, Heidrun; Jahn, Klaus; Bartenstein, Peter; Brandt, Thomas; Dutia, Mayank; Dieterich, Marianne; Strupp, Michael; la Fougère, Christian; Zwergal, Andreas

    2015-01-01

    An acute unilateral vestibular lesion leads to a vestibular tone imbalance with nystagmus, head roll tilt and postural imbalance. These deficits gradually decrease over days to weeks due to central vestibular compensation (VC). This study investigated the effects of i.v. N-acetyl-DL-leucine, N-acetyl-L-leucine and N-acetyl-D-leucine on VC using behavioural testing and serial [18F]-Fluoro-desoxyglucose ([18F]-FDG)-μPET in a rat model of unilateral chemical labyrinthectomy (UL). Vestibular behavioural testing included measurements of nystagmus, head roll tilt and postural imbalance as well as sequential whole-brain [18F]-FDG-μPET was done before and on days 1,3,7 and 15 after UL. A significant reduction of postural imbalance scores was identified on day 7 in the N-acetyl-DL-leucine (p metabolism (rCGM) by means of μPET revealed that only N-acetyl-L-leucine but not N-acetyl-D-leucine caused a significant increase of rCGM in the vestibulocerebellum and a decrease in the posterolateral thalamus and subthalamic region on days 3 and 7. A similar pattern was found when comparing the effect of N-acetyl-L-leucine on rCGM in an UL-group and a sham UL-group without vestibular damage. In conclusion, N-acetyl-L-leucine improves compensation of postural symptoms after UL in a dose-dependent and specific manner, most likely by activating the vestibulocerebellum and deactivating the posterolateral thalamus. PMID:25803613

  15. The Effect of Oral Leucine on Protein Metabolism in Adolescents with Type 1 Diabetes Mellitus

    OpenAIRE

    Vardhini Desikan; Izolda Mileva; Jeremy Garlick; Andrew H. Lane; Thomas A. Wilson; Margaret A. McNurlan

    2010-01-01

    Lack of insulin results in a catabolic state in subjects with insulin-dependent diabetes mellitus which is reversed by insulin treatment. Amino acid supply, especially branched chain amino acids such as leucine, enhances protein synthesis in both animal and human studies. This small study was undertaken to assess the acute effect of supplemental leucine on protein metabolism in adolescents with type 1 diabetes. L-[1-13C] Leucine was used to assess whole-body protein metabolism in six adolesc...

  16. An .I.ex vivo ./I.model to study v-Myb-induced leukemogenicity

    Czech Academy of Sciences Publication Activity Database

    Dvořáková, Marta; Králová, Jarmila; Karafiát, Vít; Bartůněk, Petr; Dvořák, Michal

    2001-01-01

    Roč. 27, č. 2 (2001), s. 437-445 ISSN 1079-9796 R&D Projects: GA ČR GV301/98/K042; GA ČR GA204/00/0554; GA AV ČR IPP2052002 Grant - others:HHMI(US) 75195-540401 Institutional research plan: CEZ:AV0Z5052915 Keywords : v-Myb oncoprotein * PEST domain * leucine zipper Subject RIV: EB - Genetics ; Molecular Biology Impact factor: 1.703, year: 2001

  17. Leucine signaling in the pathogenesis of type 2 diabetes and obesity

    OpenAIRE

    Melnik, Bodo C

    2012-01-01

    Epidemiological evidence points to increased dairy and meat consumption, staples of the Western diet, as major risk factors for the development of type 2 diabetes (T2D). This paper presents a new concept and comprehensive review of leucine-mediated cell signaling explaining the pathogenesis of T2D and obesity by leucine-induced over-stimulation of mammalian target of rapamycin complex 1 (mTORC1). mTORC1, a pivotal nutrient-sensitive kinase, promotes growth and cell proliferation in response t...

  18. Isolation and characterization of awamori yeast mutants with L-leucine accumulation that overproduce isoamyl alcohol.

    Science.gov (United States)

    Takagi, Hiroshi; Hashida, Keisuke; Watanabe, Daisuke; Nasuno, Ryo; Ohashi, Masataka; Iha, Tomoya; Nezuo, Maiko; Tsukahara, Masatoshi

    2015-02-01

    Awamori shochu is a traditional distilled alcoholic beverage made from steamed rice in Okinawa, Japan. Although it has a unique aroma that is distinguishable from that of other types of shochu, no studies have been reported on the breeding of awamori yeasts. In yeast, isoamyl alcohol (i-AmOH), known as the key flavor of bread, is mainly produced from α-ketoisocaproate in the pathway of L-leucine biosynthesis, which is regulated by end-product inhibition of α-isopropylmalate synthase (IPMS). Here, we isolated mutants resistant to the L-leucine analog 5,5,5-trifluoro-DL-leucine (TFL) derived from diploid awamori yeast of Saccharomyces cerevisiae. Some of the mutants accumulated a greater amount of intracellular L-leucine, and among them, one mutant overproduced i-AmOH in awamori brewing. This mutant carried an allele of the LEU4 gene encoding the Ser542Phe/Ala551Val variant IPMS, which is less sensitive to feedback inhibition by L-leucine. Interestingly, we found that either of the constituent mutations (LEU4(S542F) and LEU4(A551V)) resulted in the TFL tolerance of yeast cells and desensitization to L-leucine feedback inhibition of IPMS, leading to intracellular L-leucine accumulation. Homology modeling also suggested that L-leucine binding was drastically inhibited in the Ser542Phe, Ala551Val, and Ser542Phe/Ala551Val variants due to steric hindrance in the cavity of IPMS. As we expected, awamori yeast cells expressing LEU4(S542F), LEU4(A551V), and LEU4(S542F/A551V) showed a prominent increase in extracellular i-AmOH production, compared with that of cells carrying the vector only. The approach described here could be a practical method for the breeding of novel awamori yeasts to expand the diversity of awamori taste and flavor. Copyright © 2014 The Society for Biotechnology, Japan. Published by Elsevier B.V. All rights reserved.

  19. Leucine stimulates protein synthesis in skeletal muscle of neonatal pigs by enhancing mTORC1 activation.

    Science.gov (United States)

    Suryawan, Agus; Jeyapalan, Asumthia S; Orellana, Renan A; Wilson, Fiona A; Nguyen, Hanh V; Davis, Teresa A

    2008-10-01

    Skeletal muscle in the neonate grows at a rapid rate due in part to an enhanced sensitivity to the postprandial rise in amino acids, particularly leucine. To elucidate the molecular mechanism by which leucine stimulates protein synthesis in neonatal muscle, overnight-fasted 7-day-old piglets were treated with rapamycin [an inhibitor of mammalian target of rapamycin (mTOR) complex (mTORC)1] for 1 h and then infused with leucine for 1 h. Fractional rates of protein synthesis and activation of signaling components that lead to mRNA translation were determined in skeletal muscle. Rapamycin completely blocked leucine-induced muscle protein synthesis. Rapamycin markedly reduced raptor-mTOR association, an indicator of mTORC1 activation. Rapamycin blocked the leucine-induced phosphorylation of mTOR, S6 kinase 1 (S6K1), and eukaryotic initiation factor (eIF)4E-binding protein-1 (4E-BP1) and formation of the eIF4E.eIF4G complex and increased eIF4E.4E-BP1 complex abundance. Rapamycin had no effect on the association of mTOR with rictor, a crucial component for mTORC2 activation, or G protein beta-subunit-like protein (GbetaL), a component of mTORC1 and mTORC2. Neither leucine nor rapamycin affected the phosphorylation of AMP-activated protein kinase (AMPK), PKB, or tuberous sclerosis complex (TSC)2, signaling components that reside upstream of mTOR. Eukaryotic elongation factor (eEF)2 phosphorylation was not affected by leucine or rapamycin, although current dogma indicates that eEF2 phosphorylation is mTOR dependent. Together, these in vivo data suggest that leucine stimulates muscle protein synthesis in neonates by enhancing mTORC1 activation and its downstream effectors.

  20. Role and metabolism of free leucine in skeletal muscle in protein sparing action of dietary carbohydrate and fat

    International Nuclear Information System (INIS)

    Nakano, Kiwao; Ishikawa, Tamotsu

    1977-01-01

    Feeding rats with either a carbohydrate meal or a fat meal to the previously fasted rats caused significant decrease in urinary output of urea and total nitrogen. The content of free leucine in skeletal muscle decreased in the rats fed either a carbohydrate meal or a fat meal. Feeding of either a carbohydrate meal or a fat meal stimulated incorporation of L-leucine-1- 14 C into protein fraction of skeletal muscle and reduced its oxidation to 14 CO 2 . These results suggest that the metabolism of leucine is under nutritional regulation and that the decrease in content of free leucine in skeletal muscle might be caused by enhanced reutilization of leucine into protein by the feeding of a carbohydrate meal or a fat meal. The role of free leucine in skeletal muscle as a regulator of protein turnover in the tissue are discussed in relation to the metabolism of this branched chain amino acid. (auth.)

  1. Catabolism of leucine to branched-chain fatty acids in Staphylococcus xylosus

    DEFF Research Database (Denmark)

    Beck, Hans Christian; Hansen, A M; Lauritsen, F R

    2004-01-01

    Staphylococcus xylosus is an important starter culture in the production of flavours from the branched-chain amino acids leucine, valine and isoleucine in fermented meat products. The sensorially most important flavour compounds are the branched-chain aldehydes and acids derived from the correspo......Staphylococcus xylosus is an important starter culture in the production of flavours from the branched-chain amino acids leucine, valine and isoleucine in fermented meat products. The sensorially most important flavour compounds are the branched-chain aldehydes and acids derived from...

  2. ATP-binding motifs play key roles in Krp1p, kinesin-related protein 1, function for bi-polar growth control in fission yeast

    International Nuclear Information System (INIS)

    Rhee, Dong Keun; Cho, Bon A; Kim, Hyong Bai

    2005-01-01

    Kinesin is a microtubule-based motor protein with various functions related to the cell growth and division. It has been reported that Krp1p, kinesin-related protein 1, which belongs to the kinesin heavy chain superfamily, localizes on microtubules and may play an important role in cytokinesis. However, the function of Krp1p has not been fully elucidated. In this study, we overexpressed an intact form and three different mutant forms of Krp1p in fission yeast constructed by site-directed mutagenesis in two ATP-binding motifs or by truncation of the leucine zipper-like motif (LZiP). We observed hyper-extended microtubules and the aberrant nuclear shape in Krp1p-overexpressed fission yeast. As a functional consequence, a point mutation of ATP-binding domain 1 (G89E) in Krp1p reversed the effect of Krp1p overexpression in fission yeast, whereas the specific mutation in ATP-binding domain 2 (G238E) resulted in the altered cell polarity. Additionally, truncation of the leucine zipper-like domain (LZiP) at the C-terminal of Krp1p showed a normal nuclear division. Taken together, we suggest that krp1p is involved in regulation of cell-polarized growth through ATP-binding motifs in fission yeast

  3. Overexpression and characterization of an extracellular leucine aminopeptidase from Aspergillus oryzae.

    Science.gov (United States)

    Matsushita-Morita, Mayumi; Tada, Sawaki; Suzuki, Satoshi; Hattori, Ryota; Marui, Junichiro; Furukawa, Ikuyo; Yamagata, Youhei; Amano, Hitoshi; Ishida, Hiroki; Takeuchi, Michio; Kashiwagi, Yutaka; Kusumoto, Ken-Ichi

    2011-02-01

    Leucine aminopeptidase (LAP), an enzyme used in the food industry, is an exopeptidase that removes an amino acid residue, primarily leucine (Leu), from the N-terminus of peptides and protein substrates. In this study, we focused on the leucine aminopeptidase A (lapA) gene from Aspergillus oryzae RIB40. To purify and characterize the LapA, lapA was overexpressed in A. oryzae RIB40 using the amyB promoter. LAP activity in the culture supernatant of one transformant harboring the lapA expression plasmid was 33 times that of the host strain. LapA was purified from the culture supernatant of this lapA-overexpressing strain by column chromatography. The purified recombinant LapA had a molecular mass of 33 kDa, and its N-terminal amino acid was the tyrosine at position 80 of the deduced amino acid sequence. Optimal enzyme activity was observed at 60°C and pH 8.5, and the enzyme was stable at temperatures up to 60°C and in the pH range 7.5-11. In transcriptional analysis, lapA was induced under alkaline conditions and expressed at a relatively low level under normal conditions. LapA showed maximum hydrolyzing activity for the substrate leucine para-nitroanilide (Leu-pNA), followed by substrates Phe-pNA (39% activity compared with Leu-pNA), Met-pNA, Lys-pNA, and Arg-pNA. In addition, LapA preferentially hydrolyzed peptides longer than tripeptides.

  4. The Influence of 8 Weeks of Whey-Protein and Leucine Supplementation on Physical and Cognitive Performance

    Science.gov (United States)

    2010-01-01

    Influence of 8 Weeks of Whey Protein and Leucine Supplementation on Physical and Cognitive Performance 5a. GONTRAGT NUMBER FA8650-04-D-6472 5b. GRANT NUMBER...investigate the ability of whey -protein and leucine supplementation to enhance physical and cognitive performance and body composition. Thirty moderately fit...composition before and after supplementing their daily diet for 8 wk with either 19.7 g of whey protein and 6.2 g leucine (WPL) or a calorie-equivalent placebo

  5. Functional mapping and implications of substrate specificity of the yeast high-affinity leucine permease Bap2.

    Science.gov (United States)

    Usami, Yuki; Uemura, Satsohi; Mochizuki, Takahiro; Morita, Asami; Shishido, Fumi; Inokuchi, Jin-ichi; Abe, Fumiyoshi

    2014-07-01

    Leucine is a major amino acid in nutrients and proteins and is also an important precursor of higher alcohols during brewing. In Saccharomyces cerevisiae, leucine uptake is mediated by multiple amino acid permeases, including the high-affinity leucine permease Bap2. Although BAP2 transcription has been extensively analyzed, the mechanisms by which a substrate is recognized and moves through the permease remain unknown. Recently, we determined 15 amino acid residues required for Tat2-mediated tryptophan import. Here we introduced homologous mutations into Bap2 amino acid residues and showed that 7 residues played a role in leucine import. Residues I109/G110/T111 and E305 were located within the putative α-helix break in TMD1 and TMD6, respectively, according to the structurally homologous Escherichia coli arginine/agmatine antiporter AdiC. Upon leucine binding, these α-helix breaks were assumed to mediate a conformational transition in Bap2 from an outward-open to a substrate-binding occluded state. Residues Y336 (TMD7) and Y181 (TMD3) were located near I109 and E305, respectively. Bap2-mediated leucine import was inhibited by some amino acids according to the following order of severity: phenylalanine, leucine>isoleucine>methionine, tyrosine>valine>tryptophan; histidine and asparagine had no effect. Moreover, this order of severity clearly coincided with the logP values (octanol-water partition coefficients) of all amino acids except tryptophan. This result suggests that the substrate partition efficiency to the buried Bap2 binding pocket is the primary determinant of substrate specificity rather than structural amino acid side chain recognition. Copyright © 2014 Elsevier B.V. All rights reserved.

  6. Leucine and its transporter provide protection against cigarette smoke-induced cell death: A potential therapy for emphysema

    Directory of Open Access Journals (Sweden)

    Bannhi Das

    2014-01-01

    Full Text Available Cigarette smoke (CS is a major risk factor for emphysematous changes in the lungs and the underlying mechanism involves CS-induced cell death. In the present study we investigated the ability of nutrients to rescue CS-induced cell death. We observed that pre-treatment with excess leucine can partially rescue CS extract-induced cell death in Saccharomyces cerevisiae and alveolar epithelial A549 cells. Excess dietary leucine was also effective in alleviating effects of CS in guinea pig lungs. Further investigation to understand the underlying mechanism showed that CS exposure causes downregulation of leucine transporter that results in inactivation of mTOR, which is a positive regulator of protein synthesis and cell proliferation. Notably, leucine supplemented diet ameliorated even existing CS-induced emphysematous changes in guinea pig lung, a condition hitherto thought to be irreversible. Thus the current study documents a new mechanism by which CS affects cellular physiology wherein leucine transporter is a key target.

  7. Hypertrophy-Promoting Effects of Leucine Supplementation and Moderate Intensity Aerobic Exercise in Pre-Senescent Mice

    Directory of Open Access Journals (Sweden)

    Zhi Xia

    2016-05-01

    Full Text Available Several studies have indicated a positive influence of leucine supplementation and aerobic training on the aging skeletal muscle signaling pathways that control muscle protein balance and muscle remodeling. However, the effect of a combined intervention requires further clarification. Thirteen month old CD-1® mice were subjected to moderate aerobic exercise (45 min swimming per day with 3% body weight workload and fed a chow diet with 5% leucine or 3.4% alanine for 8 weeks. Serum and plasma were prepared for glucose, urea nitrogen, insulin and amino acid profile analysis. The white gastrocnemius muscles were used for determination of muscle size and signaling proteins involved in protein synthesis and degradation. The results show that both 8 weeks of leucine supplementation and aerobic training elevated the activity of mTOR (mammalian target of rapamycin and its downstream target p70S6K and 4E-BP1, inhibited the ubiquitin-proteasome system, and increased fiber cross-sectional area (CSA in white gastrocnemius muscle. Moreover, leucine supplementation in combination with exercise demonstrated more significant effects, such as greater CSA, protein content and altered phosphorylation (suggestive of increased activity of protein synthesis signaling proteins, in addition to lower expression of proteins involved in protein degradation compared to leucine or exercise alone. The current study shows moderate aerobic training combined with 5% leucine supplementation has the potential to increase muscle size in fast-twitch skeletal muscle during aging, potentially through increased protein synthesis and decreased protein breakdown.

  8. Leucine-rich diet alters the eukaryotic translation initiation factors expression in skeletal muscle of tumour-bearing rats

    Directory of Open Access Journals (Sweden)

    Mello Maria

    2007-03-01

    Full Text Available Abstract Background Cancer-cachexia induces a variety of metabolic disorders on protein turnorver, decreasing protein synthesis and increasing protein degradation. Controversly, insulin, other hormones, and branched-chain amino acids, especially leucine, stimulate protein synthesis and modulate the activity of translation initiation factors involved in protein synthesis. Since the tumour effects are more pronounced when associated with pregnancy, ehancing muscle-wasting proteolysis, in this study, the influence of a leucine-rich diet on the protein synthesis caused by cancer were investigated. Methods Pregnant rats with or without Walker 256 tumour were distributed into six groups. During 20 days of experiment, three groups were fed with a control diet: C – pregnant control, W – tumour-bearing, and P – pair-fed, which received the same amount of food as ingested by the W group; three other groups of pregnant rats were fed a leucine-rich diet: L – pregnant leucine, WL – tumour-bearing, and PL – pair-fed, which received the same amount of food as ingested by the WL group. Results The gastrocnemius muscle of WL rats showed increased incorporation of leucine in protein compared to W rats; the leucine-rich diet also prevented the decrease in plasma insulin normally seen in W. The expression of translation initiation factors increased when tumour-bearing rats fed leucine-rich diet, with increase of ~35% for eIF2α and eIF5, ~17% for eIF4E and 20% for eIF4G; the expression of protein kinase S6K1 and protein kinase C was also highly enhanced. Conclusion The results suggest that a leucine-rich diet increased the protein synthesis in skeletal muscle in tumour-bearing rats possibly through the activation of eIF factors and/or the S6kinase pathway.

  9. Leucine-rich diet alters the eukaryotic translation initiation factors expression in skeletal muscle of tumour-bearing rats

    International Nuclear Information System (INIS)

    Ventrucci, Gislaine; Mello, Maria Alice R; Gomes-Marcondes, Maria Cristina C

    2007-01-01

    Cancer-cachexia induces a variety of metabolic disorders on protein turnorver, decreasing protein synthesis and increasing protein degradation. Controversly, insulin, other hormones, and branched-chain amino acids, especially leucine, stimulate protein synthesis and modulate the activity of translation initiation factors involved in protein synthesis. Since the tumour effects are more pronounced when associated with pregnancy, ehancing muscle-wasting proteolysis, in this study, the influence of a leucine-rich diet on the protein synthesis caused by cancer were investigated. Pregnant rats with or without Walker 256 tumour were distributed into six groups. During 20 days of experiment, three groups were fed with a control diet: C – pregnant control, W – tumour-bearing, and P – pair-fed, which received the same amount of food as ingested by the W group; three other groups of pregnant rats were fed a leucine-rich diet: L – pregnant leucine, WL – tumour-bearing, and PL – pair-fed, which received the same amount of food as ingested by the WL group. The gastrocnemius muscle of WL rats showed increased incorporation of leucine in protein compared to W rats; the leucine-rich diet also prevented the decrease in plasma insulin normally seen in W. The expression of translation initiation factors increased when tumour-bearing rats fed leucine-rich diet, with increase of ~35% for eIF2α and eIF5, ~17% for eIF4E and 20% for eIF4G; the expression of protein kinase S6K1 and protein kinase C was also highly enhanced. The results suggest that a leucine-rich diet increased the protein synthesis in skeletal muscle in tumour-bearing rats possibly through the activation of eIF factors and/or the S6kinase pathway

  10. Leucine-rich diet alters the eukaryotic translation initiation factors expression in skeletal muscle of tumour-bearing rats

    Energy Technology Data Exchange (ETDEWEB)

    Ventrucci, Gislaine [Laboratório de Nutrição e Câncer, Departamento de Fisiologia e Biofísica, Instituto de Biologia, Universidade Estadual de Campinas (UNICAMP), Campinas, 13083-970, São Paulo (Brazil); Mello, Maria Alice R [Departamento de Fisiologia e Biofísica, Instituto Biociências, Universidade Estadual de São Paulo, UNESP, Rio Claro, 13506-900, São Paulo (Brazil); Gomes-Marcondes, Maria Cristina C [Laboratório de Nutrição e Câncer, Departamento de Fisiologia e Biofísica, Instituto de Biologia, Universidade Estadual de Campinas (UNICAMP), Campinas, 13083-970, São Paulo (Brazil)

    2007-03-06

    Cancer-cachexia induces a variety of metabolic disorders on protein turnorver, decreasing protein synthesis and increasing protein degradation. Controversly, insulin, other hormones, and branched-chain amino acids, especially leucine, stimulate protein synthesis and modulate the activity of translation initiation factors involved in protein synthesis. Since the tumour effects are more pronounced when associated with pregnancy, ehancing muscle-wasting proteolysis, in this study, the influence of a leucine-rich diet on the protein synthesis caused by cancer were investigated. Pregnant rats with or without Walker 256 tumour were distributed into six groups. During 20 days of experiment, three groups were fed with a control diet: C – pregnant control, W – tumour-bearing, and P – pair-fed, which received the same amount of food as ingested by the W group; three other groups of pregnant rats were fed a leucine-rich diet: L – pregnant leucine, WL – tumour-bearing, and PL – pair-fed, which received the same amount of food as ingested by the WL group. The gastrocnemius muscle of WL rats showed increased incorporation of leucine in protein compared to W rats; the leucine-rich diet also prevented the decrease in plasma insulin normally seen in W. The expression of translation initiation factors increased when tumour-bearing rats fed leucine-rich diet, with increase of ~35% for eIF2α and eIF5, ~17% for eIF4E and 20% for eIF4G; the expression of protein kinase S6K1 and protein kinase C was also highly enhanced. The results suggest that a leucine-rich diet increased the protein synthesis in skeletal muscle in tumour-bearing rats possibly through the activation of eIF factors and/or the S6kinase pathway.

  11. Recognition of double-stranded DNA using energetically activated duplexes with interstrand zippers of 1-, 2-or 4-pyrenyl-functionalized O2 '-alkylated RNA monomers

    DEFF Research Database (Denmark)

    Karmakar, Saswata; Madsen, Andreas Stahl; Guenther, Dale C.

    2014-01-01

    '-alkylated uridine monomers X-Z by means of thermal denaturation experiments, optical spectroscopy, force-field simulations and recognition experiments using DNA hairpins as model targets. We demonstrate that Invaders with +1 interstrand zippers of X or Y monomers efficiently recognize mixed-sequence DNA...

  12. Leucine minimizes denervation-induced skeletal muscle atrophy of rats through akt/mtor signaling pathways

    Directory of Open Access Journals (Sweden)

    Carolina Barbosa Ribeiro

    2015-03-01

    Full Text Available The aim of the present study was to evaluate the effect of leucine treatment (0.30 mM on muscle weight and signaling of myoproteins related to synthesis and degradation pathways of soleus muscle following seven days of complete sciatic nerve lesion.Wistar rats (n=24 of 3 to 4 months of age (192 ± 23 g were used. The animals were randomly distributed into four experimental groups (n=6/group: control, treated with leucine (L, denervated (D and denervated treated with leucine (DL.Dependent measures were proteins levels of AKT, AMPK, mTOR, and ACC performed by Western blot. Leucine induced a reduction in the phosphorylation of AMPK (p<0.05 by 16% in the L and by 68% in the DL groups as compared with control group. Denervation increased AMPK by 24% in the D group as compared with the control group (p<0.05. AKT was also modulated by denervation and leucine treatment, highlighted by the elevation of AKT phosphorylation in the D (65%, L (98% and DL (146% groups as compared with the control group (p<0.05. AKT phosphorylation was 49% higher in the D group as compared with the DL group.Furthermore, denervation decreased mTOR phosphorylation by 29% in the D group as compared with the control group. However, leucine treatment induced an increase of 49% in the phosphorylation of mTOR in the L group as compared with the control group, and an increase of 154% in the DL as compared with the D group ( p<0.05. ACC phosphorylation was 20% greater in the D group than the control group. Furthermore, ACC in the soleus was 22% lower in the in the L group and 50% lower in the DL group than the respective control group (p<0.05.In conclusion, leucine treatment minimized the deleterious effects of denervation on rat soleus muscle by increasing anabolic (AKT and mTOR and decreasing catabolic (AMPK pathways. These results may be interesting for muscle recovery following acute denervation, which may contribute to musculoskeletal rehabilitation after denervation.

  13. Embryonic expression of zebrafish MiT family genes tfe3b, tfeb, and tfec.

    Science.gov (United States)

    Lister, James A; Lane, Brandon M; Nguyen, Anhthu; Lunney, Katherine

    2011-11-01

    The MiT family comprises four genes in mammals: Mitf, Tfe3, Tfeb, and Tfec, which encode transcription factors of the basic-helix-loop-helix/leucine zipper class. Mitf is well-known for its essential role in the development of melanocytes, however the functions of the other members of this family, and of interactions between them, are less well understood. We have now characterized the complete set of MiT genes from zebrafish, which totals six instead of four. The zebrafish genome contain two mitf (mitfa and mitfb), two tfe3 (tfe3a and tfe3b), and single tfeb and tfec genes; this distribution is shared with other teleosts. We present here the sequence and embryonic expression patterns for the zebrafish tfe3b, tfeb, and tfec genes, and identify a new isoform of tfe3a. These findings will assist in elucidating the roles of the MiT gene family over the course of vertebrate evolution. Copyright © 2011 Wiley-Liss, Inc.

  14. Discovery of a Regulatory Motif for Human Satellite DNA Transcription in Response to BATF2 Overexpression.

    Science.gov (United States)

    Bai, Xuejia; Huang, Wenqiu; Zhang, Chenguang; Niu, Jing; Ding, Wei

    2016-03-01

    One of the basic leucine zipper transcription factors, BATF2, has been found to suppress cancer growth and migration. However, little is known about the genes downstream of BATF2. HeLa cells were stably transfected with BATF2, then chromatin immunoprecipitation-sequencing was employed to identify the DNA motifs responsive to BATF2. Comprehensive bioinformatics analyses indicated that the most significant motif discovered as TTCCATT[CT]GATTCCATTC[AG]AT was primarily distributed among the chromosome centromere regions and mostly within human type II satellite DNA. Such motifs were able to prime the transcription of type II satellite DNA in a directional and asymmetrical manner. Consistently, satellite II transcription was up-regulated in BATF2-overexpressing cells. The present study provides insight into understanding the role of BATF2 in tumours and the importance of satellite DNA in the maintenance of genomic stability. Copyright© 2016 International Institute of Anticancer Research (Dr. John G. Delinassios), All rights reserved.

  15. Intragastric administration of leucine or isoleucine lowers the blood glucose response to a mixed-nutrient drink by different mechanisms in healthy, lean volunteers.

    Science.gov (United States)

    Ullrich, Sina S; Fitzgerald, Penelope Ce; Schober, Gudrun; Steinert, Robert E; Horowitz, Michael; Feinle-Bisset, Christine

    2016-11-01

    The branched-chain amino acids leucine and isoleucine lower blood glucose after oral glucose ingestion, and the intraduodenal infusion of leucine decreases energy intake in healthy, lean men. We investigated the effects of the intragastric administration of leucine and isoleucine on the gastric emptying of, and blood glucose responses to, a physiologic mixed-macronutrient drink and subsequent energy intake. In 2 separate studies, 12 healthy, lean subjects received on 3 separate occasions an intragastric infusion of 5 g leucine (leucine-5g) or an intragastric infusion of 10 g leucine (leucine-10g), an intragastric infusion of 5 g isoleucine (isoleucine-5g) or an intragastric infusion of 10 g isoleucine (isoleucine-10g), or a control. Fifteen minutes later, subjects consumed a mixed-nutrient drink (400 kcal, 56 g carbohydrates, 15 g protein, and 12 g fat), and gastric emptying ( 13 C-acetate breath test) and blood glucose, plasma insulin, C-peptide, glucagon, glucagon-like peptide-1 (GLP-1), glucose-dependent insulinotropic polypeptide (GIP), and cholecystokinin (leucine study only) were measured for 60 min. Immediately afterward, energy intake from a cold, buffet-style meal was assessed. Compared with the control, leucine-10g decreased the blood glucose area under the curve (AUC) (P blood glucose (P = 0.07), whereas effects of leucine-5g were NS. Leucine-10g, but not leucine-5g, increased plasma insulin and C-peptide AUCs (P blood glucose AUC and peak blood glucose (P blood glucose AUC. Isoleucine did not affect energy intake. In healthy subjects, both leucine and isoleucine reduced blood glucose in response to a mixed-nutrient drink but did not affect subsequent energy intake. The mechanisms underlying glucose lowering appear to differ; leucine stimulated insulin, whereas isoleucine acted insulin independently. These trials were registered at www.anzctr.org.au as 12613000899741 and 12614000837628. © 2016 American Society for Nutrition.

  16. Lean body mass change over 6 years is associated with dietary leucine intake in an older Danish population.

    Science.gov (United States)

    McDonald, Cameron Keith; Ankarfeldt, Mikkel Z; Capra, Sandra; Bauer, Judy; Raymond, Kyle; Heitmann, Berit Lilienthal

    2016-05-01

    Higher protein intake, and particularly higher leucine intake, is associated with attenuated loss of lean body mass (LBM) over time in older individuals. Dietary leucine is thought to be a key mediator of anabolism. This study aimed to assess this relationship over 6 years among younger and older adult Danes. Dietary leucine intake was assessed at baseline and after 6 years in men and women, aged 35-65 years, participating in the Danish cohort of the WHO-MONICA (Multinational MONItoring of trends and determinants in CArdiovascular disease) study (n 368). Changes in LBM over the 6 years were measured by bioelectrical impedance using equations developed for this Danish population. The association between leucine and LBM changes was examined using multivariate linear regression and ANCOVA analyses adjusted for potential confounders. After adjustment for baseline LBM, sex, age, energy intake and physical activity, leucine intake was associated with LBM change in those older than 65 years (n 79), with no effect seen in those younger than 65 years. Older participants in the highest quartile of leucine intake (7·1 g/d) experienced LBM maintenance, whereas lower intakes were associated with LBM loss over 6 years (for trend: β=0·434, P=0·03). Sensitivity analysis indicated no effect modification of sex or the presence of CVD. Greater leucine intake in conjunction with adequate total protein intake was associated with long-term LBM retention in a healthy older Danish population. This study corroborates findings from laboratory investigations in relation to protein and leucine intakes and LBM change. A more diverse and larger sample is needed for confirmation of these results.

  17. Leucine Biosynthesis Is Involved in Regulating High Lipid Accumulation in Yarrowia lipolytica

    DEFF Research Database (Denmark)

    Kerkhoven, Eduard J.; Kim, Young-Mo; Wei, Siwei

    2017-01-01

    correlation was observed between the responses on the transcript and protein levels. Combination of DGA1 overexpression with nitrogen limitation resulted in a high level of lipid accumulation accompanied by downregulation of several amino acid biosynthetic pathways, including that of leucine in particular......, and these changes were further correlated with a decrease in metabolic fluxes. This downregulation was supported by the measured decrease in the level of 2-isopropylmalate, an intermediate of leucine biosynthesis. Combining the multi-omics data with putative transcription factor binding motifs uncovered...

  18. Host epithelial cell invasion by Campylobacter jejuni: trigger or zipper mechanism?

    Directory of Open Access Journals (Sweden)

    Tadhg eÓ Cróinín

    2012-03-01

    Full Text Available Campylobacter jejuni, a spiral-shaped Gram-negative pathogen, is a highly frequent cause of gastrointestinal foodborne illness in humans worldwide. Clinical outcome of C. jejuni infections ranges from mild to severe diarrheal disease, and some other complications including reactive arthritis and Guillain–Barré syndrome. This review article highlights various C. jejuni pathogenicity factors, host cell determinants and proposed signaling mechanisms involved in human host cell invasion and their potential role in the development of C. jejuni-mediated disease. A model is presented which outlines the various important interactions of C. jejuni with the intestinal epithelium, and we discuss the pro’s and con’s for the zipper over the trigger mechanism of invasion. Future work should clarify the contradictory role of some previously identified factors, and should identify and characterize novel virulence determinants, which are crucial to provide fresh insights into the diversity of strategies employed by this pathogen to cause disease.

  19. The Perennial Ryegrass GenomeZipper: Targeted Use of Genome Resources for Comparative Grass Genomics1[C][W

    Science.gov (United States)

    Pfeifer, Matthias; Martis, Mihaela; Asp, Torben; Mayer, Klaus F.X.; Lübberstedt, Thomas; Byrne, Stephen; Frei, Ursula; Studer, Bruno

    2013-01-01

    Whole-genome sequences established for model and major crop species constitute a key resource for advanced genomic research. For outbreeding forage and turf grass species like ryegrasses (Lolium spp.), such resources have yet to be developed. Here, we present a model of the perennial ryegrass (Lolium perenne) genome on the basis of conserved synteny to barley (Hordeum vulgare) and the model grass genome Brachypodium (Brachypodium distachyon) as well as rice (Oryza sativa) and sorghum (Sorghum bicolor). A transcriptome-based genetic linkage map of perennial ryegrass served as a scaffold to establish the chromosomal arrangement of syntenic genes from model grass species. This scaffold revealed a high degree of synteny and macrocollinearity and was then utilized to anchor a collection of perennial ryegrass genes in silico to their predicted genome positions. This resulted in the unambiguous assignment of 3,315 out of 8,876 previously unmapped genes to the respective chromosomes. In total, the GenomeZipper incorporates 4,035 conserved grass gene loci, which were used for the first genome-wide sequence divergence analysis between perennial ryegrass, barley, Brachypodium, rice, and sorghum. The perennial ryegrass GenomeZipper is an ordered, information-rich genome scaffold, facilitating map-based cloning and genome assembly in perennial ryegrass and closely related Poaceae species. It also represents a milestone in describing synteny between perennial ryegrass and fully sequenced model grass genomes, thereby increasing our understanding of genome organization and evolution in the most important temperate forage and turf grass species. PMID:23184232

  20. Endocrine responses and acute mTOR pathway phosphorylation to resistance exercise with leucine and whey

    Directory of Open Access Journals (Sweden)

    MT Lane

    2017-02-01

    Full Text Available Leucine ingestion reportedly activates the mTOR pathway in skeletal muscle, contributing to a hypertrophy response. The purpose of the study was to compare the post-resistance exercise effects of leucine and whey protein supplementation on endocrine responses and muscle mTOR pathway phosphorylation. On visit 1, subjects (X±SD; n=20; age=27.8±2.8yrs provided baseline blood samples for analysis of cortisol, glucose and insulin; a muscle biopsy of the vastus lateralis muscle to assess mTOR signaling pathway phosphorylation; and were tested for maximum strength on the leg press and leg extension exercises. For visits 2 and 3, subjects were randomized in a double-blind crossover design to ingest either leucine and whey protein (10g+10g; supplement or a non-caloric placebo. During these visits, 5 sets of 10 repetitions were performed on both exercises, immediately followed by ingestion of the supplement or placebo. Blood was sampled 30 min post-, and a muscle biopsy 45 min post-exercise. Western blots quantified total and phosphorylated proteins. Insulin increased (α<.05 with supplementation with no change in glucose compared to placebo. Relative phosphorylation of AKT and rpS6 were greater with leucine and whey supplementation compared to placebo. Supplementation of leucine and whey protein immediately after heavy resistance exercise increases anabolic signaling in human skeletal muscle.

  1. l-Leucine Supplementation Worsens the Adiposity of Already Obese Rats by Promoting a Hypothalamic Pattern of Gene Expression that Favors Fat Accumulation

    Directory of Open Access Journals (Sweden)

    Thais T. Zampieri

    2014-04-01

    Full Text Available Several studies showed that l-leucine supplementation reduces adiposity when provided before the onset of obesity. We studied rats that were exposed to a high-fat diet (HFD for 10 weeks before they started to receive l-leucine supplementation. Fat mass was increased in l-leucine-supplemented rats consuming the HFD. Accordingly, l-leucine produced a hypothalamic pattern of gene expression that favors fat accumulation. In conclusion, l-leucine supplementation worsened the adiposity of rats previously exposed to HFD possibly by central mechanisms.

  2. Oral leucine supplementation is sensed by the brain but neither reduces food intake nor induces an anorectic pattern of gene expression in the hypothalamus.

    Directory of Open Access Journals (Sweden)

    Thais T Zampieri

    Full Text Available Leucine activates the intracellular mammalian target of the rapamycin (mTOR pathway, and hypothalamic mTOR signaling regulates food intake. Although central infusion of leucine reduces food intake, it is still uncertain whether oral leucine supplementation is able to affect the hypothalamic circuits that control energy balance. We observed increased phosphorylation of p70s6k in the mouse hypothalamus after an acute oral gavage of leucine. We then assessed whether acute oral gavage of leucine induces the activation of neurons in several hypothalamic nuclei and in the brainstem. Leucine did not induce the expression of Fos in hypothalamic nuclei, but it increased the number of Fos-immunoreactive neurons in the area postrema. In addition, oral gavage of leucine acutely increased the 24 h food intake of mice. Nonetheless, chronic leucine supplementation in the drinking water did not change the food intake and the weight gain of ob/ob mice and of wild-type mice consuming a low- or a high-fat diet. We assessed the hypothalamic gene expression and observed that leucine supplementation increased the expression of enzymes (BCAT1, BCAT2 and BCKDK that metabolize branched-chain amino acids. Despite these effects, leucine supplementation did not induce an anorectic pattern of gene expression in the hypothalamus. In conclusion, our data show that the brain is able to sense oral leucine intake. However, the food intake is not modified by chronic oral leucine supplementation. These results question the possible efficacy of leucine supplementation as an appetite suppressant to treat obesity.

  3. Isovalerianeacidæmi--en sjælden og alvorlig defekt i nedbrydningen af leucin

    DEFF Research Database (Denmark)

    Lund, Ann-Britt Kiholm; Lund, Allan

    2011-01-01

    Isovaleric acidaemia (IVA) is an organic acidemia caused by deficient metabolism of the essential amino acid leucine. We describe the biochemistry, diagnostics, and treatment of IVA, and present the known Danish patients.......Isovaleric acidaemia (IVA) is an organic acidemia caused by deficient metabolism of the essential amino acid leucine. We describe the biochemistry, diagnostics, and treatment of IVA, and present the known Danish patients....

  4. Amino acid transport across the tonoplast of vacuoles isolated from barley mesophyll protoplasts: Uptake of alanine, leucine, and glutamine

    International Nuclear Information System (INIS)

    Dietz, K.J.; Jaeger, R.; Kaiser, G.; Martinoia, E.

    1990-01-01

    Mesophyll protoplasts from leaves of well-fertilized barley (Hordeum vulgare L.) plants contained amino acids at concentrations as high as 120 millimoles per liter. With the exception of glutamic acid, which is predominantly localized in the cytoplasm, a major part of all other amino acids was contained inside the large central vacuole. Alanine, leucine, and glutamine are the dominant vacuolar amino acids in barley. Their transport into isolated vacuoles was studied using 14 C-labeled amino acids. Uptake was slow in the absence of ATP. A three- to sixfold stimulation of uptake was observed after addition of ATP or adenylyl imidodiphosphate an ATP analogue not being hydrolyzed by ATPases. Other nucleotides were ineffective in increasing the rate of uptake. ATP-Stimulated amino acid transport was not dependent on the transtonoplast pH or membrane potential. p-Chloromercuriphenylsulfonic acid and n-ethyl maleimide increased transport independently of ATP. Neutral amino acids such as valine or leucine effectively decreased the rate of alanine transport. Glutamine and glycine were less effective or not effective as competitive inhibitors of alanine transport. The results indicate the existence of a uniport translocator specific for neutral or basic amino acids that is under control of metabolic effectors

  5. Dietary leucine--an environmental modifier of insulin resistance acting on multiple levels of metabolism

    DEFF Research Database (Denmark)

    Macotela, Yazmin; Emanuelli, Brice; Bång, Anneli M

    2011-01-01

    homeostasis and insulin signaling. After 8 weeks on HFD, mice developed obesity, fatty liver, inflammatory changes in adipose tissue and insulin resistance at the level of IRS-1 phosphorylation, as well as alterations in metabolomic profile of amino acid metabolites, TCA cycle intermediates, glucose...... and cholesterol metabolites, and fatty acids in liver, muscle, fat and serum. Doubling dietary leucine reversed many of the metabolite abnormalities and caused a marked improvement in glucose tolerance and insulin signaling without altering food intake or weight gain. Increased dietary leucine was also associated......Environmental factors, such as the macronutrient composition of the diet, can have a profound impact on risk of diabetes and metabolic syndrome. In the present study we demonstrate how a single, simple dietary factor--leucine--can modify insulin resistance by acting on multiple tissues...

  6. Zippered release from polymer-gated carbon nanotubes

    KAUST Repository

    Mashat, Afnan

    2012-01-01

    A thermosensitive drug delivery system based on polymer-gated carbon nanotubes (CNTs) that are loaded with the anticancer drug doxorubicin (DOX) is herein reported. The development of carbon nanotubes for various biomedical applications is the research focus of many research groups and holds great promise. The major drawback of these materials is the toxicity that is associated with conjugated carbon systems. Functionalization of CNTs with polymers has proved very successful in lowering the toxicity and improving the pharmacokinetic profile. In this work, CNTs are coated with polyethylenimine (PEI) and polyvinyl alcohol (PVA) via the "zipper effect" that provides both support and control over drug release. PEI/PVA provides excellent support to increase DOX loading on the nanocarrier. The system is controlled by changes in temperature due to the complexation (low temperature) and decomplexation (high temperature) of PEI and PVA via hydrogen bonding. The release of DOX was tested in three cell lines (Lung fibroblast (LF), Breast Adenocarcinoma (BA), and HeLa). It was further tested in primary cell lines (Human Dermal Fibroblast adult (HDFa) and Human Dermal Fibroblast neonatal (HDFn)). When the bonds between PEI and PVA are decomplexed at high temperature (≥40 °C), drug release was observed as verified by fluorescence microscopy. There was no drug release at room temperature (25 °C) and a slow release at normal body temperature (37 °C). This system represents a promising method for incorporating stimuli triggered polymer-gated CNTs in future controlled release applications. © 2012 The Royal Society of Chemistry.

  7. Synergistic effects of leucine and resveratrol on insulin sensitivity and fat metabolism in adipocytes and mice

    Directory of Open Access Journals (Sweden)

    Bruckbauer Antje

    2012-08-01

    Full Text Available Abstract Background Sirtuins are important regulators of glucose and fat metabolism, and sirtuin activation has been proposed as a therapeutic target for insulin resistance and diabetes. We have shown leucine to increase mitochondrial biogenesis and fat oxidation via Sirt1 dependent pathways. Resveratrol is a widely recognized activator of Sirt; however, the biologically-effective high concentrations used in cell and animal studies are generally impractical or difficult to achieve in humans. Accordingly, we sought to determine whether leucine would exhibit synergy with low levels of resveratrol on sirtuin-dependent outcomes in adipocytes and in diet-induced obese (DIO mice. Methods 3T3-L1 mouse adipocytes were treated with Leucine (0.5 mM, β-hydroxy-β-methyl butyrate (HMB (5 μM or Resveratrol (200 nM alone or in combination. In addition, diet-induced obese mice were treated for 6-weeks with low (2 g/kg diet or high (10 g/kg diet dose HMB, Leucine (24 g/kg diet; 200% of normal level or low (12.5 mg/kg diet or high (225 mg/kg diet dose resveratrol, alone or as combination with leucine-resveratrol or HMB-resveratrol. Results Fatty acid oxidation, AMPK, Sirt1 and Sirt3 activity in 3T3-L1 adipocytes and in muscle cells, were significantly increased by the combinations compared to the individual treatments. Similarly, 6-week feeding of low-dose resveratrol combined with either leucine or its metabolite HMB to DIO mice increased adipose Sirt1 activity, muscle glucose and palmitate uptake (measured via PET/CT, insulin sensitivity (HOMAIR, improved inflammatory stress biomarkers (CRP, IL-6, MCP-1, adiponectin and reduced adiposity comparable to the effects of high dose resveratrol, while low-dose resveratrol exerted no independent effect. Conclusion These data demonstrate that either leucine or its metabolite HMB may be combined with a low concentration of resveratrol to exert synergistic effects on Sirt1-dependent outcomes; this may result in more

  8. SIRT4 Is a Lysine Deacylase that Controls Leucine Metabolism and Insulin Secretion

    DEFF Research Database (Denmark)

    Anderson, Kristin A; Huynh, Frank K; Fisher-Wellman, Kelsey

    2017-01-01

    in leucine oxidation, and we show a primary role for SIRT4 in controlling this pathway in mice. Furthermore, we find that dysregulated leucine metabolism in SIRT4KO mice leads to elevated basal and stimulated insulin secretion, which progressively develops into glucose intolerance and insulin resistance....... These findings identify a robust enzymatic activity for SIRT4, uncover a mechanism controlling branched-chain amino acid flux, and position SIRT4 as a crucial player maintaining insulin secretion and glucose homeostasis during aging....

  9. Leucine-rich diet alters the 1H-NMR based metabolomic profile without changing the Walker-256 tumour mass in rats.

    Science.gov (United States)

    Viana, Laís Rosa; Canevarolo, Rafael; Luiz, Anna Caroline Perina; Soares, Raquel Frias; Lubaczeuski, Camila; Zeri, Ana Carolina de Mattos; Gomes-Marcondes, Maria Cristina Cintra

    2016-10-03

    Cachexia is one of the most important causes of cancer-related death. Supplementation with branched-chain amino acids, particularly leucine, has been used to minimise loss of muscle tissue, although few studies have examined the effect of this type of nutritional supplementation on the metabolism of the tumour-bearing host. Therefore, the present study evaluated whether a leucine-rich diet affects metabolomic derangements in serum and tumour tissues in tumour-bearing Walker-256 rats (providing an experimental model of cachexia). After 21 days feeding Wistar female rats a leucine-rich diet, distributed in L-leucine and LW-leucine Walker-256 tumour-bearing groups, we examined the metabolomic profile of serum and tumour tissue samples and compared them with samples from tumour-bearing rats fed a normal protein diet (C - control; W - tumour-bearing groups). We utilised 1 H-NMR as a means to study the serum and tumour metabolomic profile, tumour proliferation and tumour protein synthesis pathway. Among the 58 serum metabolites examined, we found that 12 were altered in the tumour-bearing group, reflecting an increase in activity of some metabolic pathways related to energy production, which diverted many nutrients toward tumour growth. Despite displaying increased tumour cell activity (i.e., higher Ki-67 and mTOR expression), there were no differences in tumour mass associated with changes in 23 metabolites (resulting from valine, leucine and isoleucine synthesis and degradation, and from the synthesis and degradation of ketone bodies) in the leucine-tumour group. This result suggests that the majority of nutrients were used for host maintenance. A leucine rich-diet, largely used to prevent skeletal muscle loss, did not affect Walker 256 tumour growth and led to metabolomic alterations that may partially explain the positive effects of leucine for the whole tumour-bearing host.

  10. Lean body mass change over 6 years is associated with dietary leucine intake in an older Danish population

    DEFF Research Database (Denmark)

    McDonald, Cameron Keith; Ankarfeldt, Mikkel Z.; Capra, Sandra

    2016-01-01

    Higher protein intake, and particularly higher leucine intake, is associated with attenuated loss of lean body mass (LBM) over time in older individuals. Dietary leucine is thought to be a key mediator of anabolism. This study aimed to assess this relationship over 6 years among younger and older...... corroborates findings from laboratory investigations in relation to protein and leucine intakes and LBM change. A more diverse and larger sample is needed for confirmation of these results....

  11. Seizures induced by carbachol, morphine, and leucine-enkephalin: a comparison.

    Science.gov (United States)

    Snead, O C

    1983-04-01

    The electrical, behavioral, and pharmacological properties of seizures induced by morphine, leucine-enkephalin, and the muscarinic cholinergic agonist carbachol were examined and compared. Low-dose carbachol given intracerebroventricularly (ICV) produced seizures similar electrically to those produced by ICV morphine and leucine-enkephalin, although there was some difference in site of subcortical origin of onset. Carbachol and morphine were similar in that they had the same anticonvulsant profile, produced similar behavioral changes, caused generalized absence seizures in low doses and generalized convulsive seizures in high doses, and were capable of chemical kindling. However, opiate-induced seizures were not overcome by cholinergic antagonists, nor were carbachol seizures blocked by opiate antagonists. These data suggest that there may be a common noncholinergic, nonopiatergic system involved in mediating carbachol- and morphine-induced seizures but not enkephalin seizures.

  12. Estimating bacterial production in marine waters from the simultaneous incorporation of thymidine and leucine.

    Science.gov (United States)

    Chin-Leo, G; Kirchman, D L

    1988-08-01

    We examined the simultaneous incorporation of [H]thymidine and [C]leucine to obtain two independent indices of bacterial production (DNA and protein syntheses) in a single incubation. Incorporation rates of leucine estimated by the dual-label method were generally higher than those obtained by the single-label method, but the differences were small (dual/single = 1.1 +/- 0.2 [mean +/- standard deviation]) and were probably due to the presence of labeled leucyl-tRNA in the cold trichloroacetic acid-insoluble fraction. There were no significant differences in thymidine incorporation between dual- and single-label incubations (dual/ single = 1.03 +/- 0.13). Addition of the two substrates in relatively large amounts (25 nM) did not apparently increase bacterial activity during short incubations (leucine incorporation rates covaried over depth profiles of the Chesapeake Bay. Estimates of bacterial production based on thymidine and leucine differed by less than 25%. Although the need for appropriate conversion factors has not been eliminated, the dual-label approach can be used to examine the variation in bacterial production while ensuring that the observed variation in incorporation rates is due to real changes in bacterial production rather than changes in conversion factors or introduction of other artifacts.

  13. A Novel Antibiotic Mechanism of l-Cyclopropylalanine Blocking the Biosynthetic Pathway of Essential Amino Acid l-Leucine

    Directory of Open Access Journals (Sweden)

    Bingji Ma

    2017-12-01

    Full Text Available The unusual amino acid l-cyclopropylalanine was isolated from the mushroom Amanita virgineoides after detection in an anti-fungal screening test. l-Cyclopropylalanine was found to exhibit broad-spectrum inhibition against fungi and bacteria. The anti-fungal activity was found to be abolished in the presence of the amino acid l-leucine, but not any other amino acids, indicating that l-cyclopropylalanine may block the biosynthesis of the essential amino acid l-leucine, thereby inhibiting fungal and bacteria growth. Further biochemical studies found l-cyclopropylalanine indeed inhibits α-isopropylmalate synthase (α-IMPS, the enzyme that catalyzes the rate-limiting step in the biosynthetic pathway of l-leucine. Inhibition of essential l-leucine synthesis in fungal and bacteria organisms, a pathway absent in host organisms such as humans, may represent a novel antibiotic mechanism to counter the ever-increasing problem of drug resistance to existing antibiotics.

  14. Experiment list: SRX150478 [Chip-atlas[Archive

    Lifescience Database Archive (English)

    Full Text Available osis=Fibrocystic Disease 66690540,98.1,24.9,110111 GSM935398: Harvard ChipSeq MCF10A-Er-Src 4OHTAM 1uM 12hr c-Fos Harvard... Control source_name=MCF10A-Er-Src || biomaterial_provider=Struhl laboratory || lab=Harvard || ...lab description=Struhl - Harvard University || datatype=ChipSeq || datatype descr... is a leucine-zipper. || antibody vendorname=Santa Cruz Biotech || antibody vendorid=sc-7202 || control=Harvard..._Control || control description=input library was prepared at Harvard. || control=Harvard

  15. Leucine supplementation improves acquired growth hormone resistance in rats with protein-energy malnutrition.

    Science.gov (United States)

    Gao, Xuejin; Tian, Feng; Wang, Xinying; Zhao, Jie; Wan, Xiao; Zhang, Li; Wu, Chao; Li, Ning; Li, Jieshou

    2015-01-01

    Protein-energy malnutrition (PEM) can lead to growth hormone (GH) resistance. Leucine supplementation diets have been shown to increase protein synthesis in muscles. Our study aimed at investigating if long-term leucine supplementation could modulate GH-insulin-like growth factor (IGF)-1 system function and mammalian target of rapamycin (mTOR)-related signal transduction in skeletal muscles in a rat model of severe malnutrition. Male Sprague-Dawley rats (n = 50; weight, 302 ± 5 g) were divided into 5 treatment groups, including 2 control groups (a normal control group that was fed chow and ad libitum water [CON, n = 10] and a malnourished control group [MC, n = 10] that was fed a 50% chow diet). After undergoing a weight loss stage for 4 weeks, rats received either the chow diet (MC-CON, n = 10), the chow diet supplemented with low-dose leucine (MC-L, n = 10), or the chow diet supplemented with high-dose leucine (MC-H, n = 10) for 2 weeks. The muscle masses of the gastrocnemius, soleus, and extensor digitorum longus were significantly reduced in the MC group. Re-feeding increased muscle mass, especially in the MC-L and MC-H groups. In the MC group, serum IGF-1, IGF-binding protein (IGFBP)-3, and hepatic growth hormone receptor (GHR) levels were significantly decreased and phosphorylation of the downstream anabolic signaling effectors protein kinase B (Akt), mTOR, and ribosomal protein S6 kinase 1 (S6K1) were significantly lower than in other groups. However, serum IGF-1 and IGF binding protein (IGFBP)-3 concentrations and hepatic growth hormone receptor (GHR) levels were significantly higher in the MC-L and MC-H groups than in the MC-CON group, and serum IGFBP-1 levels was significantly reduced in the MC-L and MC-H groups. These changes were consistent with those observed for hepatic mRNA expression levels. Phosphorylation of the downstream anabolic signaling effectors Akt, mTOR, and S6K1 were also significantly higher in the MC-L and MC-H groups than in the MC

  16. Characterization of the Zebrafish Homolog of Zipper Interacting Protein Kinase

    Directory of Open Access Journals (Sweden)

    Brandon W. Carr

    2014-06-01

    Full Text Available Zipper-interacting protein kinase (ZIPK is a conserved vertebrate-specific regulator of actomyosin contractility in smooth muscle and non-muscle cells. Murine ZIPK has undergone an unusual divergence in sequence and regulation compared to other ZIPK orthologs. In humans, subcellular localization is controlled by phosphorylation of threonines 299 and 300. In contrast, ZIPK subcellular localization in mouse and rat is controlled by interaction with PAR-4. We carried out a comparative biochemical characterization of the regulation of the zebrafish ortholog of ZIPK. Like the human orthologs zebrafish ZIPK undergoes nucleocytoplasmic-shuttling and is abundant in the cytoplasm, unlike the primarily nuclear rat ZIPK. Rat ZIPK, but not human or zebrafish ZIPK, interacts with zebrafish PAR-4. Mutation of the conserved residues required for activation of the mammalian orthologs abrogated activity of the zebrafish ZIPK. In contrast to the human ortholog, mutation of threonine 299 and 300 in the zebrafish ZIPK has no effect on the activity or subcellular localization. Thus, we found that zebrafish ZIPK functions in a manner most similar to the human ZIPK and quite distinct from murine orthologs, yet the regulation of subcellular localization is not conserved.

  17. Dietary leucine--an environmental modifier of insulin resistance acting on multiple levels of metabolism.

    Directory of Open Access Journals (Sweden)

    Yazmin Macotela

    Full Text Available Environmental factors, such as the macronutrient composition of the diet, can have a profound impact on risk of diabetes and metabolic syndrome. In the present study we demonstrate how a single, simple dietary factor--leucine--can modify insulin resistance by acting on multiple tissues and at multiple levels of metabolism. Mice were placed on a normal or high fat diet (HFD. Dietary leucine was doubled by addition to the drinking water. mRNA, protein and complete metabolomic profiles were assessed in the major insulin sensitive tissues and serum, and correlated with changes in glucose homeostasis and insulin signaling. After 8 weeks on HFD, mice developed obesity, fatty liver, inflammatory changes in adipose tissue and insulin resistance at the level of IRS-1 phosphorylation, as well as alterations in metabolomic profile of amino acid metabolites, TCA cycle intermediates, glucose and cholesterol metabolites, and fatty acids in liver, muscle, fat and serum. Doubling dietary leucine reversed many of the metabolite abnormalities and caused a marked improvement in glucose tolerance and insulin signaling without altering food intake or weight gain. Increased dietary leucine was also associated with a decrease in hepatic steatosis and a decrease in inflammation in adipose tissue. These changes occurred despite an increase in insulin-stimulated phosphorylation of p70S6 kinase indicating enhanced activation of mTOR, a phenomenon normally associated with insulin resistance. These data indicate that modest changes in a single environmental/nutrient factor can modify multiple metabolic and signaling pathways and modify HFD induced metabolic syndrome by acting at a systemic level on multiple tissues. These data also suggest that increasing dietary leucine may provide an adjunct in the management of obesity-related insulin resistance.

  18. Additive insulinogenic action of Opuntia ficus-indica cladode and fruit skin extract and leucine after exercise in healthy males

    Science.gov (United States)

    2013-01-01

    Background Oral intake of a specific extract of Opuntia ficus-indica cladode and fruit skin (OpunDia™) (OFI) has been shown to increase serum insulin concentration while reducing blood glucose level for a given amount of glucose ingestion after an endurance exercise bout in healthy young volunteers. However, it is unknown whether OFI-induced insulin stimulation after exercise is of the same magnitude than the stimulation by other insulinogenic agents like leucine as well as whether OFI can interact with those agents. Therefore, the aims of the present study were: 1) to compare the degree of insulin stimulation by OFI with the effect of leucine administration; 2) to determine whether OFI and leucine have an additive action on insulin stimulation post-exercise. Methods Eleven subjects participated in a randomized double-blind cross-over study involving four experimental sessions. In each session the subjects successively underwent a 2-h oral glucose tolerance test (OGTT) after a 30-min cycling bout at ~70% VO2max. At t0 and t60 during the OGTT, subjects ingested 75 g glucose and capsules containing either 1) a placebo; 2) 1000 mg OFI; 3) 3 g leucine; 4) 1000 mg OFI + 3 g leucine. Blood samples were collected before and at 30-min intervals during the OGTT for determination of blood glucose and serum insulin. Results Whereas no effect of leucine was measured, OFI reduced blood glucose at t90 by ~7% and the area under the glucose curve by ~15% and increased serum insulin concentration at t90 by ~35% compared to placebo (P<0.05). From t60 to the end of the OGTT, serum insulin concentration was higher in OFI+leucine than in placebo which resulted in a higher area under the insulin curve (+40%, P<0.05). Conclusion Carbohydrate-induced insulin stimulation post-exercise can be further increased by the combination of OFI with leucine. OFI and leucine could be interesting ingredients to include together in recovery drinks to resynthesize muscle glycogen faster post

  19. Estimating Bacterial Production in Marine Waters from the Simultaneous Incorporation of Thymidine and Leucine

    OpenAIRE

    Chin-Leo, Gerardo; Kirchman, David L.

    1988-01-01

    We examined the simultaneous incorporation of [3H]thymidine and [14C]leucine to obtain two independent indices of bacterial production (DNA and protein syntheses) in a single incubation. Incorporation rates of leucine estimated by the dual-label method were generally higher than those obtained by the single-label method, but the differences were small (dual/single = 1.1 ± 0.2 [mean ± standard deviation]) and were probably due to the presence of labeled leucyl-tRNA in the cold trichloroacetic ...

  20. Agglomerated novel spray-dried lactose-leucine tailored as a carrier to enhance the aerosolization performance of salbutamol sulfate from DPI formulations.

    Science.gov (United States)

    Molina, Carlos; Kaialy, Waseem; Chen, Qiao; Commandeur, Daniel; Nokhodchi, Ali

    2017-12-19

    Spray-drying allows to modify the physicochemical/mechanical properties of particles along with their morphology. In the present study, L -leucine with varying concentrations (0.1, 0.5, 1, 5, and 10% w/v) were incorporated into lactose monohydrate solution for spray-drying to enhance the aerosolization performance of dry powder inhalers containing spray-dried lactose-leucine and salbutamol sulfate. The prepared spray-dried lactose-leucine carriers were analyzed using laser diffraction (particle size), differential scanning calorimetry (thermal behavior), scanning electron microscopy (morphology), powder X-ray diffraction (crystallinity), Fourier transform infrared spectroscopy (interaction at molecular level), and in vitro aerosolization performance (deposition). The results showed that the efficacy of salbutamol sulfate's aerosolization performance was, in part, due to the introduction of L -leucine in the carrier, prior to being spray-dried, accounting for an increase in the fine particle fraction (FPF) of salbutamol sulfate from spray-dried lactose-leucine (0.5% leucine) in comparison to all other carriers. It was shown that all of the spray-dried carriers were spherical in their morphology with some agglomerates and contained a mixture of amorphous, α-lactose, and β-lactose. It was also interesting to note that spray-dried lactose-leucine particles were agglomerated during the spray-drying process to make coarse particles (volume mean diameter of 79 to 87 μm) suitable as a carrier in DPI formulations.

  1. A leucine-supplemented diet improved protein content of skeletal muscle in young tumor-bearing rats

    Directory of Open Access Journals (Sweden)

    Gomes-Marcondes M.C.C.

    2003-01-01

    Full Text Available Cancer cachexia induces host protein wastage but the mechanisms are poorly understood. Branched-chain amino acids play a regulatory role in the modulation of both protein synthesis and degradation in host tissues. Leucine, an important amino acid in skeletal muscle, is higher oxidized in tumor-bearing animals. A leucine-supplemented diet was used to analyze the effects of Walker 256 tumor growth on body composition in young weanling Wistar rats divided into two main dietary groups: normal diet (N, 18% protein and leucine-rich diet (L, 15% protein plus 3% leucine, which were further subdivided into control (N or L or tumor-bearing (W or LW subgroups. After 12 days, the animals were sacrificed and their carcass analyzed. The tumor-bearing groups showed a decrease in body weight and fat content. Lean carcass mass was lower in the W and LW groups (W = 19.9 ± 0.6, LW = 23.1 ± 1.0 g vs N = 29.4 ± 1.3, L = 28.1 ± 1.9 g, P < 0.05. Tumor weight was similar in both tumor-bearing groups fed either diet. Western blot analysis showed that myosin protein content in gastrocnemius muscle was reduced in tumor-bearing animals (W = 0.234 ± 0.033 vs LW = 0.598 ± 0.036, N = 0.623 ± 0.062, L = 0.697 ± 0.065 arbitrary intensity, P < 0.05. Despite accelerated tumor growth, LW animals exhibited a smaller reduction in lean carcass mass and muscle myosin maintenance, suggesting that excess leucine in the diet could counteract, at least in part, the high host protein wasting in weanling tumor-bearing rats.

  2. Contrasting ability to take up leucine and thymidine among freshwater bacterial groups: implications for bacterial production measurements

    Science.gov (United States)

    Pérez, María Teresa; Hörtnagl, Paul; Sommaruga, Ruben

    2010-01-01

    We examined the ability of different freshwater bacterial groups to take up leucine and thymidine in two lakes. Utilization of both substrates by freshwater bacteria was examined at the community level by looking at bulk incorporation rates and at the single-cell level by combining fluorescent in situ hybridization and signal amplification by catalysed reporter deposition with microautoradiography. Our results showed that leucine was taken up by 70–80% of Bacteria-positive cells, whereas only 15–43% of Bacteria-positive cells were able to take up thymidine. When a saturating substrate concentration in combination with a short incubation was used, 80–90% of Betaproteobacteria and 67–79% of Actinobacteria were positive for leucine uptake, whereas thymidine was taken up by bacterial group. Bacterial abundance was a good predictor of the relative contribution of bacterial groups to leucine uptake, whereas when thymidine was used Actinobacteria represented the large majority (> 80%) of the cells taking up this substrate. Increasing the substrate concentration to 100 nM did not affect the percentage of R-BT cells taking up leucine (> 90% even at low concentrations), but moderately increased the fraction of thymidine-positive R-BT cells to a maximum of 35% of the hybridized cells. Our results show that even at very high concentrations, thymidine is not taken up by all, otherwise active, bacterial cells. PMID:19725866

  3. Experiment list: SRX150477 [Chip-atlas[Archive

    Lifescience Database Archive (English)

    Full Text Available osis=Fibrocystic Disease 70636654,97.7,24.3,106225 GSM935397: Harvard ChipSeq MCF10A-Er-Src 4OHTAM 1uM 4hr c-Fos Harvard... Control source_name=MCF10A-Er-Src || biomaterial_provider=Struhl laboratory || lab=Harvard || l... a leucine-zipper. || antibody vendorname=Santa Cruz Biotech || antibody vendorid=sc-7202 || control=Harvard..._Control || control description=input library was prepared at Harvard. || control=Harvard..._Control || control description=input library was prepared at Harvard. || controlid=wgEncodeEH002871

  4. Experiment list: SRX150476 [Chip-atlas[Archive

    Lifescience Database Archive (English)

    Full Text Available osis=Fibrocystic Disease 60050220,94.5,18.5,85444 GSM935396: Harvard ChipSeq MCF10A-Er-Src 4OHTAM 1uM 36hr c-Fos Harvard... Control source_name=MCF10A-Er-Src || biomaterial_provider=Struhl laboratory || lab=Harvard || l...is a leucine-zipper. || antibody vendorname=Santa Cruz Biotech || antibody vendorid=sc-7202 || control=Harvard..._Control || control description=input library was prepared at Harvard. || control=Harvard..._Control || control description=input library was prepared at Harvard. || controlid=wgEncodeEH0028

  5. MicroRNA-193b-3p acts as a tumor suppressor by targeting the MYB oncogene in T-cell acute lymphoblastic leukemia

    OpenAIRE

    Mets, E; Van der Meulen, J; Van Peer, G; Boice, M; Mestdagh, P; Van de Walle, I; Lammens, T; Goossens, S; De Moerloose, B; Benoit, Y; Van Roy, N; Clappier, E; Poppe, B; Vandesompele, J; Wendel, H-G

    2014-01-01

    The MYB oncogene is a leucine zipper transcription factor essential for normal and malignant hematopoiesis. In T-cell acute lymphoblastic leukemia (T-ALL), elevated MYB levels can arise directly through T-cell receptor-mediated MYB translocations, genomic MYB duplications or enhanced TAL1 complex binding at the MYB locus or indirectly through the TAL1/miR-223/FBXW7 regulatory axis. In this study, we used an unbiased MYB 3′untranslated region–microRNA (miRNA) library screen and identified 33 p...

  6. The effect of a dietary leucine excess on the immunoresponsiveness ...

    African Journals Online (AJOL)

    Karen J. Tinker and A.M. Gous. Departments of Biochemistry, and Animal Science and Poultry Science, University of Natal, Pietermaritzburg. The effect of a dietary leucine imbalance on chickens was investigated. The two criteria considered were growth and antibody production following immunization with sheep red.

  7. Autophagy and leucine promote chronological longevity and respiration proficiency during calorie restriction in yeast.

    Science.gov (United States)

    Aris, John P; Alvers, Ashley L; Ferraiuolo, Roy A; Fishwick, Laura K; Hanvivatpong, Amanda; Hu, Doreen; Kirlew, Christine; Leonard, Michael T; Losin, Kyle J; Marraffini, Michelle; Seo, Arnold Y; Swanberg, Veronica; Westcott, Jennifer L; Wood, Michael S; Leeuwenburgh, Christiaan; Dunn, William A

    2013-10-01

    We have previously shown that autophagy is required for chronological longevity in the budding yeast Saccharomyces cerevisiae. Here we examine the requirements for autophagy during extension of chronological life span (CLS) by calorie restriction (CR). We find that autophagy is upregulated by two CR interventions that extend CLS: water wash CR and low glucose CR. Autophagy is required for full extension of CLS during water wash CR under all growth conditions tested. In contrast, autophagy was not uniformly required for full extension of CLS during low glucose CR, depending on the atg allele and strain genetic background. Leucine status influenced CLS during CR. Eliminating the leucine requirement in yeast strains or adding supplemental leucine to growth media extended CLS during CR. In addition, we observed that both water wash and low glucose CR promote mitochondrial respiration proficiency during aging of autophagy-deficient yeast. In general, the extension of CLS by water wash or low glucose CR was inversely related to respiration deficiency in autophagy-deficient cells. Also, autophagy is required for full extension of CLS under non-CR conditions in buffered media, suggesting that extension of CLS during CR is not solely due to reduced medium acidity. Thus, our findings show that autophagy is: (1) induced by CR, (2) required for full extension of CLS by CR in most cases (depending on atg allele, strain, and leucine availability) and, (3) promotes mitochondrial respiration proficiency during aging under CR conditions. Copyright © 2013 Elsevier Inc. All rights reserved.

  8. Lipid-protein interactions. The leucine transport system of Lactococcus lactis.

    NARCIS (Netherlands)

    Veld, Geertruida Elisabeth in 't

    1992-01-01

    In summary, it is concluded, that a functionally reconstituted leucine transport system of L. lactis is affected by bilayer features in the following order of importance: lipid headgroup (H+-bonding) › acyl chain carbon number (thickness) › cholesterol (fluidity) › acyl chain unsaturation (indirect

  9. Leucine Modulation of the mTOR Pathway for Cognition Modulation: Kinetic and In Vitro Studies and Model Development

    Science.gov (United States)

    2015-09-30

    ultimately at regional levels. In other words , the arrival of free leucine at a tissue site and taken up by the cells would impart a signal for...immunohistochemical techniques and immortalized rat hippocampal cells. Figure 2. Schematic of the Impact of Leucine on the mTOR Protein Synthesis Pathway

  10. Interaction between leucine and phosphodiesterase 5 inhibition in modulating insulin sensitivity and lipid metabolism

    Directory of Open Access Journals (Sweden)

    Fu L

    2015-05-01

    Full Text Available Lizhi Fu,1 Fenfen Li,1 Antje Bruckbauer,2 Qiang Cao,1 Xin Cui,1 Rui Wu,1 Hang Shi,1 Bingzhong Xue,1 Michael B Zemel21Department of Biology, Center for Obesity Reversal, Georgia State University, Atlanta, GA, 2NuSirt Biopharma Inc., Nashville, TN, USA Purpose: Leucine activates SIRT1/AMP-activated protein kinase (AMPK signaling and markedly potentiates the effects of other sirtuin and AMPK activators on insulin signaling and lipid metabolism. Phosphodiesterase 5 inhibition increases nitric oxide–cGMP signaling, which in turn exhibits a positive feedback loop with both SIRT1 and AMPK, thus amplifying peroxisome proliferator-activated receptor γ co-activator α (PGC1α-mediated effects. Methods: We evaluated potential synergy between leucine and PDE5i on insulin sensitivity and lipid metabolism in vitro and in diet-induced obese (DIO mice. Results: Leucine (0.5 mM exhibited significant synergy with subtherapeutic doses (0.1–10 nM of PDE5-inhibitors (sildenafil and icariin on fat oxidation, nitric oxide production, and mitochondrial biogenesis in hepatocytes, adipocytes, and myotubes. Effects on insulin sensitivity, glycemic control, and lipid metabolism were then assessed in DIO-mice. DIO-mice exhibited fasting and postprandial hyperglycemia, insulin resistance, and hepatic steatosis, which were not affected by the addition of leucine (24 g/kg diet. However, the combination of leucine and a subtherapeutic dose of icariin (25 mg/kg diet for 6 weeks reduced fasting glucose (38%, P<0.002, insulin (37%, P<0.05, area under the glucose tolerance curve (20%, P<0.01, and fully restored glucose response to exogenous insulin challenge. The combination also inhibited hepatic lipogenesis, stimulated hepatic and muscle fatty acid oxidation, suppressed hepatic inflammation, and reversed high-fat diet-induced steatosis. Conclusion: These robust improvements in insulin sensitivity, glycemic control, and lipid metabolism indicate therapeutic potential for

  11. Excessive Leucine-mTORC1-Signalling of Cow Milk-Based Infant Formula: The Missing Link to Understand Early Childhood Obesity.

    Science.gov (United States)

    Melnik, Bodo C

    2012-01-01

    Increased protein supply by feeding cow-milk-based infant formula in comparison to lower protein content of human milk is a well-recognized major risk factor of childhood obesity. However, there is yet no conclusive biochemical concept explaining the mechanisms of formula-induced childhood obesity. It is the intention of this article to provide the biochemical link between leucine-mediated signalling of mammalian milk proteins and adipogenesis as well as early adipogenic programming. Leucine has been identified as the predominant signal transducer of mammalian milk, which stimulates the nutrient-sensitive kinase mammalian target of rapamycin complex 1 (mTORC1). Leucine thus functions as a maternal-neonatal relay for mTORC1-dependent neonatal β-cell proliferation and insulin secretion. The mTORC1 target S6K1 plays a pivotal role in stimulation of mesenchymal stem cells to differentiate into adipocytes and to induce insulin resistance. It is of most critical concern that infant formulas provide higher amounts of leucine in comparison to human milk. Exaggerated leucine-mediated mTORC1-S6K1 signalling induced by infant formulas may thus explain increased adipogenesis and generation of lifelong elevated adipocyte numbers. Attenuation of mTORC1 signalling of infant formula by leucine restriction to physiologic lower levels of human milk offers a great chance for the prevention of childhood obesity and obesity-related metabolic diseases.

  12. Leucine incorporation by aerobic anoxygenic phototrophic bacteria in the Delaware estuary

    Science.gov (United States)

    Stegman, Monica R; Cottrell, Matthew T; Kirchman, David L

    2014-01-01

    Aerobic anoxygenic phototrophic (AAP) bacteria are well known to be abundant in estuaries, coastal regions and in the open ocean, but little is known about their activity in any aquatic ecosystem. To explore the activity of AAP bacteria in the Delaware estuary and coastal waters, single-cell 3H-leucine incorporation by these bacteria was examined with a new approach that combines infrared epifluorescence microscopy and microautoradiography. The approach was used on samples from the Delaware coast from August through December and on transects through the Delaware estuary in August and November 2011. The percent of active AAP bacteria was up to twofold higher than the percentage of active cells in the rest of the bacterial community in the estuary. Likewise, the silver grain area around active AAP bacteria in microautoradiography preparations was larger than the area around cells in the rest of the bacterial community, indicating higher rates of leucine consumption by AAP bacteria. The cell size of AAP bacteria was 50% bigger than the size of other bacteria, about the same difference on average as measured for activity. The abundance of AAP bacteria was negatively correlated and their activity positively correlated with light availability in the water column, although light did not affect 3H-leucine incorporation in light–dark experiments. Our results suggest that AAP bacteria are bigger and more active than other bacteria, and likely contribute more to organic carbon fluxes than indicated by their abundance. PMID:24824666

  13. Leucine incorporation by aerobic anoxygenic phototrophic bacteria in the Delaware estuary.

    Science.gov (United States)

    Stegman, Monica R; Cottrell, Matthew T; Kirchman, David L

    2014-11-01

    Aerobic anoxygenic phototrophic (AAP) bacteria are well known to be abundant in estuaries, coastal regions and in the open ocean, but little is known about their activity in any aquatic ecosystem. To explore the activity of AAP bacteria in the Delaware estuary and coastal waters, single-cell (3)H-leucine incorporation by these bacteria was examined with a new approach that combines infrared epifluorescence microscopy and microautoradiography. The approach was used on samples from the Delaware coast from August through December and on transects through the Delaware estuary in August and November 2011. The percent of active AAP bacteria was up to twofold higher than the percentage of active cells in the rest of the bacterial community in the estuary. Likewise, the silver grain area around active AAP bacteria in microautoradiography preparations was larger than the area around cells in the rest of the bacterial community, indicating higher rates of leucine consumption by AAP bacteria. The cell size of AAP bacteria was 50% bigger than the size of other bacteria, about the same difference on average as measured for activity. The abundance of AAP bacteria was negatively correlated and their activity positively correlated with light availability in the water column, although light did not affect (3)H-leucine incorporation in light-dark experiments. Our results suggest that AAP bacteria are bigger and more active than other bacteria, and likely contribute more to organic carbon fluxes than indicated by their abundance.

  14. Identification of the Transcription Factor Znc1p, which Regulates the Yeast-to-Hypha Transition in the Dimorphic Yeast Yarrowia lipolytica

    Science.gov (United States)

    Martinez-Vazquez, Azul; Gonzalez-Hernandez, Angelica; Domínguez, Ángel; Rachubinski, Richard; Riquelme, Meritxell; Cuellar-Mata, Patricia; Guzman, Juan Carlos Torres

    2013-01-01

    The dimorphic yeast Yarrowia lipolytica is used as a model to study fungal differentiation because it grows as yeast-like cells or forms hyphal cells in response to changes in environmental conditions. Here, we report the isolation and characterization of a gene, ZNC1, involved in the dimorphic transition in Y. lipolytica. The ZNC1 gene encodes a 782 amino acid protein that contains a Zn(II)2C6 fungal-type zinc finger DNA-binding domain and a leucine zipper domain. ZNC1 transcription is elevated during yeast growth and decreases during the formation of mycelium. Cells in which ZNC1 has been deleted show increased hyphal cell formation. Znc1p-GFP localizes to the nucleus, but mutations within the leucine zipper domain of Znc1p, and to a lesser extent within the Zn(II)2C6 domain, result in a mislocalization of Znc1p to the cytoplasm. Microarrays comparing gene expression between znc1::URA3 and wild-type cells during both exponential growth and the induction of the yeast-to-hypha transition revealed 1,214 genes whose expression was changed by 2-fold or more under at least one of the conditions analyzed. Our results suggest that Znc1p acts as a transcription factor repressing hyphal cell formation and functions as part of a complex network regulating mycelial growth in Y. lipolytica. PMID:23826133

  15. Leucine-induced activation of translational initiation is partly regulated by the branched-chain α-keto acid dehydrogenase complex in C2C12 cells

    International Nuclear Information System (INIS)

    Nakai, Naoya; Shimomura, Yoshiharu; Tamura, Tomohiro; Tamura, Noriko; Hamada, Koichiro; Kawano, Fuminori; Ohira, Yoshinobu

    2006-01-01

    Branched-chain amino acid leucine has been shown to activate the translational regulators through the mammalian target of rapamycin. However, the leucine's effects are self-limiting because leucine promotes its own disposal by an oxidative pathway. The irreversible and rate-limiting step in the leucine oxidation pathway is catalyzed by the branched-chain α-keto acid dehydrogenase (BCKDH) complex. The complex contains E1 (α2β2), E2, and E3 subunits, and its activity is abolished by phosphorylation of the E1α subunit by BCKDH kinase. The relationship between the activity of BCKDH complex and leucine-mediated activation of the protein translation was investigated using the technique of RNA interference. The activity of BCKDH complex in C2C12 cell was modulated by transfection of small interfering RNA (siRNA) for BCKDH E2 subunit or BCKDH kinase. Transfection of siRNAs decreased the mRNA expression and protein amount of corresponding gene. Suppression of either E2 subunit or kinase produced opposite effects on the cell proliferation and the activation of translational regulators by leucine. Suppression of BCKDH kinase for 48 h resulted in decreasing cell proliferation. In contrast, E2 suppression led to increased amount of total cellular protein. The phosphorylation of p70 S6 kinase by leucine was increased in E2-siRNA transfected C2C12 cells, whereas the leucine's effect was diminished in kinase-siRNA transfected cells. These results suggest that the activation of the translational regulators by leucine was partly regulated by the activity of BCKDH complex

  16. The Akt/mTOR pathway: Data comparing young and aged mice with leucine supplementation at the onset of skeletal muscle regeneration

    Directory of Open Access Journals (Sweden)

    Richard A. Perry, Jr.

    2016-09-01

    Full Text Available The data described herein is related to the article “Differential Effects of Leucine Supplementation in Young and Aged Mice at the Onset of Skeletal Muscle Regeneration” [1]. Aging is associated with a decreased ability of skeletal muscle to regenerate following injury. Leucine supplementation has been extensively shown, in young subjects, to promote protein synthesis during regeneration; however, the effects of leucine supplementation on the Akt/mTOR pathway in aged mice at the onset of muscle regeneration are not fully elucidated. In this article, we present data on the Akt/mTOR protein synthesis pathway at the onset of muscle regeneration in young and aged C57BL/6J mice that are and are not receiving leucine supplementation. More specifically, protein content of total Akt, mTOR, p70S6K and 4EBP-1 are presented. Additionally, we provide relative (phosphorylated:total protein content comparisons of these targets as they present themselves in young and aged mice who have neither been injured nor received leucine supplementation. Lastly, markers of atrophy (FoxO1/O3, MuRF-1, Atrogin-1 are also reported in these young and aged control groups. Keywords: MTOR, Skeletal muscle, Regeneration, Leucine supplementation, Aging

  17. Parkinson's Disease: Leucine-Rich Repeat Kinase 2 and Autophagy, Intimate Enemies

    Directory of Open Access Journals (Sweden)

    José M. Bravo-San Pedro

    2012-01-01

    Full Text Available Parkinson's disease is the second common neurodegenerative disorder, after Alzheimer's disease. It is a clinical syndrome characterized by loss of dopamine-generating cells in the substancia nigra, a region of the midbrain. The etiology of Parkinson's disease has long been through to involve both genetic and environmental factors. Mutations in the leucine-rich repeat kinase 2 gene cause late-onset Parkinson's disease with a clinical appearance indistinguishable from Parkinson's disease idiopathic. Autophagy is an intracellular catabolic mechanism whereby a cell recycles or degrades damage proteins and cytoplasmic organelles. This degradative process has been associated with cellular dysfunction in neurodegenerative processes including Parkinson's disease. We discuss the role of leucine-rich repeat kinase 2 in autophagy, and how the deregulations of this degradative mechanism in cells can be implicated in the Parkinson's disease etiology.

  18. Enteral leucine and protein synthesis in skeletal and cardiac muscle

    Science.gov (United States)

    There are three members of the Branch Chain Amino Acids: leucine, isoleucine, and valine. As essential amino acids, these amino acids have important functions which include a primary role in protein structure and metabolism. It is intriguing that the requirement for BCAA in humans comprise about 40–...

  19. Leucine alters hepatic glucose/lipid homeostasis via the myostatin-AMP-activated protein kinase pathway - potential implications for nonalcoholic fatty liver disease

    OpenAIRE

    Zarfeshani, Aida; Ngo, Sherry; Sheppard, Allan M

    2014-01-01

    Background Elevated plasma levels of the branched-chain amino acid (BCAA) leucine are associated with obesity and insulin resistance (IR), and thus the propensity for type 2 diabetes mellitus development. However, other clinical studies suggest the contradictory view that leucine may in fact offer a degree of protection against metabolic syndrome. Aiming to resolve this apparent paradox, we assessed the effect of leucine supplementation on the metabolism of human hepatic HepG2 cells. Results ...

  20. Blue light-induced LOV domain dimerization enhances the affinity of Aureochrome 1a for its target DNA sequence

    Science.gov (United States)

    Heintz, Udo; Schlichting, Ilme

    2016-01-01

    The design of synthetic optogenetic tools that allow precise spatiotemporal control of biological processes previously inaccessible to optogenetic control has developed rapidly over the last years. Rational design of such tools requires detailed knowledge of allosteric light signaling in natural photoreceptors. To understand allosteric communication between sensor and effector domains, characterization of all relevant signaling states is required. Here, we describe the mechanism of light-dependent DNA binding of the light-oxygen-voltage (LOV) transcription factor Aureochrome 1a from Phaeodactylum tricornutum (PtAu1a) and present crystal structures of a dark state LOV monomer and a fully light-adapted LOV dimer. In combination with hydrogen/deuterium-exchange, solution scattering data and DNA-binding experiments, our studies reveal a light-sensitive interaction between the LOV and basic region leucine zipper DNA-binding domain that together with LOV dimerization results in modulation of the DNA affinity of PtAu1a. We discuss the implications of these results for the design of synthetic LOV-based photosensors with application in optogenetics. DOI: http://dx.doi.org/10.7554/eLife.11860.001 PMID:26754770

  1. Nuclear Import of the Parsley bZIP Transcription Factor CPRF2 Is Regulated by Phytochrome Photoreceptors

    Science.gov (United States)

    Kircher, Stefan; Wellmer, Frank; Nick, Peter; Rügner, Alexander; Schäfer, Eberhard; Harter, Klaus

    1999-01-01

    In plants, light perception by photoreceptors leads to differential expression of an enormous number of genes. An important step for differential gene expression is the regulation of transcription factor activities. To understand these processes in light signal transduction we analyzed the three well-known members of the common plant regulatory factor (CPRF) family from parsley (Petroselinum crispum). Here, we demonstrate that these CPRFs, which belong to the basic- region leucine-zipper (bZIP) domain-containing transcription factors, are differentially distributed within parsley cells, indicating different regulatory functions within the regulatory networks of the plant cell. In particular, we show by cell fractionation and immunolocalization approaches that CPRF2 is transported from the cytosol into the nucleus upon irradiation due to action of phytochrome photoreceptors. Two NH2-terminal domains responsible for cytoplasmic localization of CPRF2 in the dark were characterized by deletion analysis using a set of CPRF2-green fluorescent protein (GFP) gene fusion constructs transiently expressed in parsley protoplasts. We suggest that light-induced nuclear import of CPRF2 is an essential step in phytochrome signal transduction. PMID:9922448

  2. Overexpression of Arachis hypogaea AREB1 Gene Enhances Drought Tolerance by Modulating ROS Scavenging and Maintaining Endogenous ABA Content

    Directory of Open Access Journals (Sweden)

    Ling Li

    2013-06-01

    Full Text Available AhAREB1 (Arachis hypogaea Abscisic-acid Response Element Binding Protein 1 is a member of the basic domain leucine zipper (bZIP-type transcription factor in peanut. Previously, we found that expression of AhAREB1 was specifically induced by abscisic acid (ABA, dehydration and drought. To understand the drought defense mechanism regulated by AhAREB1, transgenic Arabidopsis overexpressing AhAREB1 was conducted in wild-type (WT, and a complementation experiment was employed to ABA non-sensitivity mutant abi5 (abscisic acid-insensitive 5. Constitutive expression of AhAREB1 confers water stress tolerance and is highly sensitive to exogenous ABA. Microarray and further real-time PCR analysis revealed that drought stress, reactive oxygen species (ROS scavenging, ABA synthesis/metabolism-related genes and others were regulated in transgenic Arabidopsis overexpressing AhAREB1. Accordingly, low level of ROS, but higher ABA content was detected in the transgenic Arabidopsis plants’ overexpression of AhAREB1. Taken together, it was concluded that AhAREB1 modulates ROS accumulation and endogenous ABA level to improve drought tolerance in transgenic Arabidopsis.

  3. SlbZIP38, a Tomato bZIP Family Gene Downregulated by Abscisic Acid, Is a Negative Regulator of Drought and Salt Stress Tolerance

    Science.gov (United States)

    Pan, Yanglu; Hu, Xin; Li, Chunyan; Xu, Xing; Su, Chenggang; Li, Jinhua; Song, Hongyuan; Zhang, Xingguo; Pan, Yu

    2017-01-01

    The basic leucine zipper (bZIP) transcription factors have crucial roles in plant stress responses. In this study, the bZIP family gene SlbZIP38 (GenBank accession No: XM004239373) was isolated from a tomato (Solanum lycopersicum cv. Ailsa Craig) mature leaf cDNA library. The DNA sequence of SlbZIP38 encodes a protein of 484 amino acids, including a highly conserved bZIP DNA-binding domain in the C-terminal region. We found that SlbZIP38 was differentially expressed in various organs of the tomato plant and was downregulated by drought, salt stress, and abscisic acid (ABA). However, overexpression of SlbZIP38 significantly decreased drought and salt stress tolerance in tomatoes (Ailsa Craig). The findings that SlbZIP38 overexpression reduced the chlorophyll and free proline content in leaves but increased the malondialdehyde content may explain the reduced drought and salt tolerance observed in these lines. These results suggest that SlbZIP38 is a negative regulator of drought and salt resistance that acts by modulating ABA signaling. PMID:29261143

  4. Divergence of the bZIP Gene Family in Strawberry, Peach, and Apple Suggests Multiple Modes of Gene Evolution after Duplication

    Directory of Open Access Journals (Sweden)

    Xiao-Long Wang

    2015-01-01

    Full Text Available The basic leucine zipper (bZIP transcription factors are the most diverse members of dimerizing transcription factors. In the present study, 50, 116, and 47 bZIP genes were identified in Malus domestica (apple, Prunus persica (peach, and Fragaria vesca (strawberry, respectively. Species-specific duplication was the main contributor to the large number of bZIPs observed in apple. After WGD in apple genome, orthologous bZIP genes corresponding to strawberry on duplicated regions in apple genome were retained. However, in peach ancestor, these syntenic regions were quickly lost or deleted. Maybe the positive selection contributed to the expansion of clade S to adapt to the development and environment stresses. In addition, purifying selection was mainly responsible for bZIP sequence-specific DNA binding. The analysis of orthologous pairs between chromosomes indicates that these orthologs derived from one gene duplication located on one of the nine ancient chromosomes in the Rosaceae. The comparative analysis of bZIP genes in three species provides information on the evolutionary fate of bZIP genes in apple and peach after they diverged from strawberry.

  5. The target landscape of clinical kinase drugs.

    Science.gov (United States)

    Klaeger, Susan; Heinzlmeir, Stephanie; Wilhelm, Mathias; Polzer, Harald; Vick, Binje; Koenig, Paul-Albert; Reinecke, Maria; Ruprecht, Benjamin; Petzoldt, Svenja; Meng, Chen; Zecha, Jana; Reiter, Katrin; Qiao, Huichao; Helm, Dominic; Koch, Heiner; Schoof, Melanie; Canevari, Giulia; Casale, Elena; Depaolini, Stefania Re; Feuchtinger, Annette; Wu, Zhixiang; Schmidt, Tobias; Rueckert, Lars; Becker, Wilhelm; Huenges, Jan; Garz, Anne-Kathrin; Gohlke, Bjoern-Oliver; Zolg, Daniel Paul; Kayser, Gian; Vooder, Tonu; Preissner, Robert; Hahne, Hannes; Tõnisson, Neeme; Kramer, Karl; Götze, Katharina; Bassermann, Florian; Schlegl, Judith; Ehrlich, Hans-Christian; Aiche, Stephan; Walch, Axel; Greif, Philipp A; Schneider, Sabine; Felder, Eduard Rudolf; Ruland, Juergen; Médard, Guillaume; Jeremias, Irmela; Spiekermann, Karsten; Kuster, Bernhard

    2017-12-01

    Kinase inhibitors are important cancer therapeutics. Polypharmacology is commonly observed, requiring thorough target deconvolution to understand drug mechanism of action. Using chemical proteomics, we analyzed the target spectrum of 243 clinically evaluated kinase drugs. The data revealed previously unknown targets for established drugs, offered a perspective on the "druggable" kinome, highlighted (non)kinase off-targets, and suggested potential therapeutic applications. Integration of phosphoproteomic data refined drug-affected pathways, identified response markers, and strengthened rationale for combination treatments. We exemplify translational value by discovering SIK2 (salt-inducible kinase 2) inhibitors that modulate cytokine production in primary cells, by identifying drugs against the lung cancer survival marker MELK (maternal embryonic leucine zipper kinase), and by repurposing cabozantinib to treat FLT3-ITD-positive acute myeloid leukemia. This resource, available via the ProteomicsDB database, should facilitate basic, clinical, and drug discovery research and aid clinical decision-making. Copyright © 2017 The Authors, some rights reserved; exclusive licensee American Association for the Advancement of Science. No claim to original U.S. Government Works.

  6. Bach2 is involved in neuronal differentiation of N1E-115 neuroblastoma cells

    International Nuclear Information System (INIS)

    Shim, Ki Shuk; Rosner, Margit; Freilinger, Angelika; Lubec, Gert; Hengstschlaeger, Markus

    2006-01-01

    Bach1 and Bach2 are evolutionarily related members of the BTB-basic region leucine zipper transcription factor family. We found that Bach2 downregulates cell proliferation of N1E-115 cells and negatively affects their potential to differentiate. Nuclear localization of the cyclin-dependent kinase inhibitor p21 is known to arrest cell cycle progression, and cytoplasmic p21 has been shown to promote neuronal differentiation of N1E-115 cells. We found that ectopic Bach2 causes upregulation of p21 expression in the nucleus and in the cytoplasm in undifferentiated N1E-115 cells. In differentiated cells, Bach2 specifically triggers upregulation of cytoplasmic p21. Our data suggest that Bach2 expression could represent a switch during the process of neuronal differentiation. Bach2 is not expressed in neuronal precursor cells. It would have negative effects on proliferation and differentiation of these cells. In differentiated neuronal cells Bach2 expression is upregulated, which could allow Bach2 to function as a gatekeeper of the differentiated status

  7. Leucine Affects α-Amylase Synthesis through PI3K/Akt-mTOR Signaling Pathways in Pancreatic Acinar Cells of Dairy Calves.

    Science.gov (United States)

    Guo, Long; Liang, Ziqi; Zheng, Chen; Liu, Baolong; Yin, Qingyan; Cao, Yangchun; Yao, Junhu

    2018-05-23

    Dietary nutrient utilization, particularly starch, is potentially limited by digestion in dairy cow small intestine because of shortage of α-amylase. Leucine acts as an effective signal molecular in the mTOR signaling pathway, which regulates a series of biological processes, especially protein synthesis. It has been reported that leucine could affect α-amylase synthesis and secretion in ruminant pancreas, but mechanisms have not been elaborated. In this study, pancreatic acinar (PA) cells were used as a model to determine the cellular signal of leucine influence on α-amylase synthesis. PA cells were isolated from newborn Holstein dairy bull calves and cultured in Dulbecco's modifed Eagle's medium/nutrient mixture F12 liquid media containing four leucine treatments (0, 0.23, 0.45, and 0.90 mM, respectively), following α-amylase activity, zymogen granule, and signal pathway factor expression detection. Rapamycin, a specific inhibitor of mTOR, was also applied to PA cells. Results showed that leucine increased ( p synthesis of α-amylase as well as phosphorylation of PI3K, Akt, mTOR, and S6K1 while reduced ( p synthesis. In addition, the extracellular leucine dosage significantly influenced intracellular metabolism of isoleucine ( p synthesis through promoting the PI3K/Akt-mTOR pathway and reducing the GCN2 pathway in PA cells of dairy calves. These pathways form the signaling network that controls the protein synthesis and metabolism. It would be of great interest in future studies to explore the function of leucine in ruminant nutrition.

  8. Changes in the transport of leucine-14C across the red cell membrane in children with acute lymphoblastic leukemia

    International Nuclear Information System (INIS)

    Stepniewski, M.; Cyklis, R.; Szafran, Z.; Armata, J.; Nawrocka-Kanska, B.

    1981-01-01

    Distribution of leucine- 14 C between intracellular water of red blood cells and incubation medium was significantly higher in 13 children with acute lymphoblastic leukemia than in 22 healthy children. The distribution ratio of leucine- 14 C was significantly lower when measured in the group of 6 children in the period of remission, as compared with children in the acute phase of the disease and only slightly higher than in the control group. The results of this study indicate the existence of structural changes in leukemic red cell membrane responsible for the observed disturbances of leucine transport. (author)

  9. Experiment list: SRX190296 [Chip-atlas[Archive

    Lifescience Database Archive (English)

    Full Text Available ex AP-1. As such, the FOS proteins have been implicated as regulators of cell prolifer...rming the transcription factor complex AP-1. As such, the FOS proteins have been implicated... against the N-terminus of Fra-2 of human origin. Recommended for detection of Fra-2 of m, r, ... rabbit polyclonal raised against the N-terminus of Fra-2 of human origin. Recommended for detection of F...s encode leucine zipper proteins that can dimerize with proteins of the JUN family, thereby forming the transcription factor compl

  10. Influence of haloperidol on the 3H-leucine incorporation in incretory organs of the mouse

    International Nuclear Information System (INIS)

    Lange, E.; Hackenberg, P.

    1978-01-01

    The 3 H-leucine incorporation in proteins of incretory organs of the mouse, the exocrine pancreas, and the renal tubuli was studied autoradiographically after administration of therapylike doses of the neuroleptic drug Haloperidol. With exception of the pancreas, a dosage dependent increase of the 3 H-leucine incorporation was observed in the treated animals. The results reveal an activation of the hypothalamus-hypophysis-adrenocortical system due to a 10-days administration of Haloperidol. These results are in conformity with former ones in brain and liver. (author)

  11. A route to anionic hydrophilic films of copolymers of l-leucine, l-aspartic acid and l-aspartic acid esters

    NARCIS (Netherlands)

    Sederel, W.L.; Bantjes, A.; Feijen, Jan

    1975-01-01

    A series of copolymers of l-leucine and β-benzyl-l-aspartate [Leu/Asp(OBz)] covering the range 30–70 mol % of l-leucine, was synthesized by the N-carboxyanhydride (NCA) method. The copolymers were characterized by elemental analysis, infra-red spectroscopy and viscometry. For all compositions high

  12. Sestrin regulation of TORC1: Is Sestrin a leucine sensor?

    Energy Technology Data Exchange (ETDEWEB)

    Lee, Jun Hee; Cho, Uhn-Soo; Karin, Michael (Michigan); (UCSD)

    2016-06-07

    Sestrins are highly conserved, stress-inducible proteins that inhibit target of rapamycin complex 1 (TORC1) signaling. After their transcriptional induction, both vertebrate and invertebrate Sestrins turn on the adenosine monophosphate (AMP)–activated protein kinase (AMPK), which activates the tuberous sclerosis complex (TSC), a key inhibitor of TORC1 activation. However, Sestrin overexpression, on occasion, can result in TORC1 inhibition even in AMPK-deficient cells. This effect has been attributed to Sestrin’s ability to bind the TORC1-regulating GATOR2 protein complex, which was postulated to control trafficking of TORC1 to lysosomes. How the binding of Sestrins to GATOR2 is regulated and how it contributes to TORC1 inhibition are unknown. New findings suggest that the amino acid leucine specifically disrupts the association of Sestrin2 with GATOR2, thus explaining how leucine and related amino acids stimulate TORC1 activity. We discuss whether and how these findings fit what has already been learned about Sestrin-mediated TORC1 inhibition from genetic studies conducted in fruit flies and mammals.

  13. Incorporation of [3H]Leucine and [3H]Valine into Protein of Freshwater Bacteria: Field Applications

    OpenAIRE

    Jørgensen, Niels O. G.

    1992-01-01

    Incorporation of leucine and valine into proteins of freshwater bacteria as a measure of bacterial production was tested in two eutrophic Danish lakes and was related to bacterial production measured by thymidine incorporation. In a depth profile (0 to 8 m) in Frederiksborg Castle Lake, incorporation of 100 nM leucine and valine gave similar rates of protein production. In terms of carbon, this production was about 50% lower than incorporation of 10 nM thymidine. In another depth profile in t...

  14. Methodological study on determining endogenous amino acid excretion of broiler chickens by single intravenous injection of 3H-leucine

    International Nuclear Information System (INIS)

    Yao Junhu; Wang Kangning; Yang Feng; Zhou Anguo; Cai Xuelin; Duanmu Dao

    1999-01-01

    Forty broiler chickens (1.5 kg of body weight, BW) were randomly divided into 20 groups. Every fifth group was force-fed a nitrogen-free diet (NFD) or a NFd + 3.20% enzyme hydrolysed casein (EHC) diet or diets with 5% and 20% crude protein (CP) in which soybean meal (sol.) was the sole nitrogen source. 30μCi 3 H-leucine/kg BW was intravenously injected into all birds just after the force-feeding. Venous blood samples were taken at 5 min, 4h, 24h, 36h and 48h after the injection, and the amount of excreta for the whole period of 48h was collected. The amino acids excreted after force-feeding NFD + 3.20% EHC of CP5% diet were theoretically endogenous. The ratios of specific radioactivity (SR) in excreta and the value of definite integral in free plasma from 0 to 48 h after injection of labelled leucine were not different (P > 0.05) when NFD, NFD + 3.20% EHC or CP5% diet was fed. From these results and theoretical analysis, it was suggested that for the birds with CP20% diet, the ratio of SR in endogenous leucine and value of definite integral in free plasma from 0 to 48 h after injection of labelled leucine would be the same as that of the birds with NFD diet, and thus endogenous losses of leucine and other amino acids, by the endogenous amino acid pattern measured with NFD diet, could be estimated for CP20% diet. The endogenous amino acid losses measured by this new technique was 120.50% of those measured by NFD method. It was suggested that single intravenous injection of 3 H-leucine first proposed would be more valuable for determining endogenous amino acid losses, especially when practical nitrogen-containing diet was fed

  15. Study on the interaction between leucine-enkephalin and hypothalamus-pituitary-thyroid axis

    International Nuclear Information System (INIS)

    Li Fengying; Chen Jialun; Chen Mingdao; Tang Jinfeng; Li Jiping

    2001-01-01

    Objective: To study the possible interaction between leucine enkephalin and hypothalamus-pituitary-thyroid axis. Methods: Mice models of hyperthyroidism and hypothyroidism were produced. Serum thyroid hormonal levels (T 3 , T 4 , TSH, TRH), the leucine enkephalin content of the whole brain and 5-HT of the hypothalamus were determined in the animals sacrificed on different days after the animal models were established. Results: In hyperthyroid rats, the levels of T 3 , T 4 increased progressively (P 3 , T 4 levels were persistent lower (P < 0.001) along with gradually increasing of serum TSH and TRH levels while pituitary TSh hypothalamus TRH content decreased gradually (P < 0.01), but rose back when reaching the nadir, Besides, LEK elevated and 5-HT decreased (P<0.01). Conclusion: The thyroid functional hormonal changes are not necessarily accompanied by a corresponding increase or decrease of brain LEK

  16. Synergistic effects of polyphenols and methylxanthines with Leucine on AMPK/Sirtuin-mediated metabolism in muscle cells and adipocytes.

    Directory of Open Access Journals (Sweden)

    Antje Bruckbauer

    Full Text Available The AMPK-Sirt1 pathway is an important regulator of energy metabolism and therefore a potential target for prevention and therapy of metabolic diseases. We recently demonstrated leucine and its metabolite β-hydroxy-β-methylbutyrate (HMB to synergize with low-dose resveratrol (200 nM to activate sirtuin signaling and stimulate energy metabolism. Here we show that leucine exerts a direct effect on Sirt1 kinetics, reducing its Km for NAD(+ by >50% and enabling low doses of resveratrol to further activate the enzyme (p = 0.012. To test which structure elements of resveratrol are necessary for synergy, we assessed potential synergy of structurally similar and dissimilar polyphenols as well as other compounds converging on the same pathways with leucine using fatty acid oxidation (FAO as screening tool. Dose-response curves for FAO were constructed and the highest non-effective dose (typically 1-10 nM was used with either leucine (0.5 mM or HMB (5 µM to treat adipocytes and myotubes for 24 h. Significant synergy was detected for stilbenes with FAO increase in adipocytes by 60-70% (p2000% (p1 µM and exhibited little or no synergy. Thus, the six-carbon ring structure bound to a carboxylic group seems to be a necessary element for leucine/HMB synergy with other stilbenes and hydroxycinnamic acids to stimulate AMPK/Sirt1 dependent FAO; these effects occur at concentrations that produce no independent effects and are readily achievable via oral administration.

  17. Influence of haloperidol on the /sup 3/H-leucine incorporation in incretory organs of the mouse

    Energy Technology Data Exchange (ETDEWEB)

    Lange, E; Hackenberg, P [Bezirksnervenklinik, Schwerin (German Democratic Republic)

    1978-01-01

    The /sup 3/H-leucine incorporation in proteins of incretory organs of the mouse, the exocrine pancreas, and the renal tubuli was studied autoradiographically after administration of therapylike doses of the neuroleptic drug Haloperidol. With exception of the pancreas, a dosage dependent increase of the /sup 3/H-leucine incorporation was observed in the treated animals. The results reveal an activation of the hypothalamus-hypophysis-adrenocortical system due to a 10-days administration of Haloperidol. These results are in conformity with former ones in brain and liver.

  18. Effect of dietary nutrients on ileal endogenous losses of threonine, cysteine, methionine, lysine, leucine and protein in broiler chicks.

    Science.gov (United States)

    Cerrate, S; Vignale, S K; Ekmay, R; England, J; Coon, C

    2018-04-01

    An isotope dose technique was utilized (i) to determine endogenous amino acid (AA) and protein losses and (ii) to propose adjusted values for AA requirements. The endogenous flow rate was calculated from the pool of enrichment in plasma AA, assuming similitude to enrichment of endogenous AA. In experiment 1, chicks were orally administered D4-lysine at 2% of estimated lysine intake from 16 to 24 days to find the isotopic steady state of the atom percent excess (APE) of lysine for plasma and jejunal and ileal digesta. The APE of D4-lysine in plasma, jejunal digesta and ileal digesta reached the isotopic steady state at 5.5, 3.4 and 2.0 days, respectively, by using the broken-line model. It was assumed that the isotopic steady state at 5 days identified for D4-lysine is also representative for the 15N-labeled AA. In experiment 2, chicks were fed diets from 1 to 21 days with increasing levels of fat (6%, 8%, 12%, 13% extract ether), protein (26%, 28.5%, 31% CP) or fiber (14%, 16%, 18% NDF) by adding poultry fat, soybean meal, blended animal protein or barley. Chicks were orally administered 15N-threonine, 15N-cysteine, 15N-methionine, 15N-lysine and 15N-leucine at 2% of estimated daily intake for 5 days from 17 to 21 days of age. Dietary nutrients influenced endogenous losses (EL), where dietary fat stimulated EL of lysine (P=0.06), leucine and protein (P=0.07); dietary protein enhanced EL of leucine and protein; and finally the dietary fiber increased EL of leucine. Dietary nutrients also affected apparent ileal digestibility (AID). Dietary fat increased AID of cysteine but decreased AID of lysine. Dietary protein reduced AID of protein, threonine, lysine and leucine, and similarly dietary fiber decreased AID of protein, threonine, methionine, lysine and leucine. In contrast, dietary fat or protein did not affect real ileal digestibility (RID) of protein and AA except threonine and leucine. The dietary fiber reduced the RID of protein, threonine and leucine. This

  19. L-leucine partially rescues translational and developmental defects associated with zebrafish models of Cornelia de Lange syndrome.

    Science.gov (United States)

    Xu, Baoshan; Sowa, Nenja; Cardenas, Maria E; Gerton, Jennifer L

    2015-03-15

    Cohesinopathies are human genetic disorders that include Cornelia de Lange syndrome (CdLS) and Roberts syndrome (RBS) and are characterized by defects in limb and craniofacial development as well as mental retardation. The developmental phenotypes of CdLS and other cohesinopathies suggest that mutations in the structure and regulation of the cohesin complex during embryogenesis interfere with gene regulation. In a previous project, we showed that RBS was associated with highly fragmented nucleoli and defects in both ribosome biogenesis and protein translation. l-leucine stimulation of the mTOR pathway partially rescued translation in human RBS cells and development in zebrafish models of RBS. In this study, we investigate protein translation in zebrafish models of CdLS. Our results show that phosphorylation of RPS6 as well as 4E-binding protein 1 (4EBP1) was reduced in nipbla/b, rad21 and smc3-morphant embryos, a pattern indicating reduced translation. Moreover, protein biosynthesis and rRNA production were decreased in the cohesin morphant embryo cells. l-leucine partly rescued protein synthesis and rRNA production in the cohesin morphants and partially restored phosphorylation of RPS6 and 4EBP1. Concomitantly, l-leucine treatment partially improved cohesinopathy embryo development including the formation of craniofacial cartilage. Interestingly, we observed that alpha-ketoisocaproate (α-KIC), which is a keto derivative of leucine, also partially rescued the development of rad21 and nipbla/b morphants by boosting mTOR-dependent translation. In summary, our results suggest that cohesinopathies are caused in part by defective protein synthesis, and stimulation of the mTOR pathway through l-leucine or its metabolite α-KIC can partially rescue development in zebrafish models for CdLS. © The Author 2014. Published by Oxford University Press.

  20. Gibberellic acid, amino acids (glycine and L-leucine), vitamin B 2 ...

    African Journals Online (AJOL)

    The combined effects of zinc, gibberellic acid, vitamin B2, amino acids (glycine and L-leucine) on pigment production were evaluated in a liquid culture of Monascus purpureus. In this study, response surface design was used to optimize each parameter. The data were analyzed using Minitab 14 software. Five parameters ...

  1. Leucine, starch and bicarbonate utilization by specific bacterial groups in surface shelf waters off Galicia (NW Spain).

    Science.gov (United States)

    Teira, E; Hernando-Morales, V; Guerrero-Feijóo, E; Varela, M M

    2017-06-01

    The capability of different bacterial populations to degrade abundant polymers, such as algal-derived polysaccharides, or to utilize preferentially polymers over monomers, remains largely unknown. In this study, microautoradiography was combined with fluorescence in situ hybridization (MAR-FISH) to evaluate the ability of Bacteroidetes, SAR11, Roseobacter spp., Gammaproteobacteria and SAR86 cells to use bicarbonate, leucine and starch under natural light conditions at two locations in shelf surface waters off NW Spain. The percentage of cells incorporating bicarbonate was relatively high (mean 32% ± 4%) and was positively correlated with the intensity of solar radiation. The proportion of cells using starch (mean 56% ± 4%) or leucine (mean 47% ± 4%) was significantly higher than that using bicarbonate. On average, SAR11, Roseobacter spp. and Gammaproteobacteria showed a similarly high percentage of cells using leucine (47%-65% of hybridized cells) than using starch (51%-64% of hybridized cells), while Bacteroidetes and SAR86 cells preferentially used starch (53% of hybridized cells) over leucine (34%-40% of hybridized cells). We suggest that the great percentage of bacteria using starch is related to a high ambient availability of polymers associated to algal cell lysis, which, in turn, weakens the short-term coupling between phytoplankton release and bacterial production. © 2017 Society for Applied Microbiology and John Wiley & Sons Ltd.

  2. Kinetics of free radical reactions in irradiated crystalline L-leucine

    International Nuclear Information System (INIS)

    Dole, M.; Mahdavi, M.

    1983-01-01

    Four aspects of the radiation chemistry of crystalline L-leucine are presented. They are as follows: the transformation of one type of free radical into another as observed by following the ESR spectrum of individual peaks in the initial stages of the free radical decay at room or higher temperatures after a gamma irradiation at 77K; the catalytic effect of hydrogen gas in significantly accelerating the free radical decay; the unexpected effect of argon gas in decreasing the rate of the free radical decay; and the accurate agreement of the decay data with the diffusion controlled second order Waite equation when the latter is expressed in the linear form. The major free radical which exists in irradiated L-leucine gives rise to a spectrum of 8 Doublets for a total of 16 lines. Numbering the peaks from the least intense to the strongest, peaks numbers 3, 4, 6, and 7, initially increase in intensity before decreasing while peak number 5 rapidly decreases. However, during this initial period the total free radical concentration as measured by the total moment of the ESR spectrum decreases. The kinetics of these effects are discussed. (author)

  3. Effect of biostimulator on the incorporation of DL(1-14C)leucine into skeletal muscle proteins of buffalo

    International Nuclear Information System (INIS)

    Nath, N.C.; Singh, L.N.

    1982-01-01

    Incorporation of 14 C-leucine into muscle proteins of buffalo calves was studied in vitro using muscle fibre preparations from biceps femoris. Biostimulator (a spleen tissue extract) stimulated the incorporation of 14 C-leucine into total proteins to some extent, but inhibited the synthesis of sarcoplasmic proteins. There was no significant difference in the relative proportion of the individual sarcoplasmic and myofibrillar proteins in the presence or absence of biostimulator. In one major sarcoplasmic protein there was higher specific activity in the presence of biostimulator. In all the remaining 4 proteins the incorporation was inhibited. Among the myofibrillar proteins, the incorporation into troponins, myosin light chains and tropomyosin was stimulated in the presence of biostimulator. Myosin heavy chain and acting did not show any change in incorporation of 14 C-leucine after addition of the biostimulator. (author)

  4. ADOBE ILLUSTRATOR AND GIMP - AN APPROACH TO GARMENT DESIGN

    Directory of Open Access Journals (Sweden)

    INDRIE Liliana

    2016-05-01

    Full Text Available Designing clothes has become easier with the use of clothing design software. One of the most popular and basic software made for garments design is Adobe Illustrator. It enables designers and small to medium businesses to create clothing designs with easy access to all assets, including images, colors, brushes, and type styles. While Illustrator has all the necessary elements and features that are ideal for creating designer clothing, GIMP provides basic set of tools for image editing. When used in combination with one another, these two programs provide just about all the tools an apparel designer needs to draw fashion design sketches, technical flat sketches, CAD presentations, graphic artworks, design embroideries etc. In this paper we show how to use the instruments of those two software in order to draw a woman jacket’s garments components and to fill with texture the jacket created. By utilizing Illustrator’s Symbol we created libraries of jacket components (puller zipper, label, then the brushes to design the zipper detail. Finally, by a drag and drop we added the zipper, the zipper puller and the label. GIMP gave us access to precise fabric textures which make our output so realistic that you can almost touch the fabric.

  5. Leucine - /sup 14/C transport through erythrocyte cell membrane in newborns with hypertrophic constriction of the pylorus

    Energy Technology Data Exchange (ETDEWEB)

    Stepniewski, M; Janik, A [Akademia Medyczna, Krakow (Poland)

    1980-01-01

    In 12 newborns with hypertrophic constriction of the pylorus the coefficient of the leucine - /sup 14/C distribution in the erythrocyte intracellular twice: the first time during day prior the operation corresponding to advanced malnutrition of the newborns, and the second time seven days after pylorotomy. During the second period the effects of hyponutrition were partially balanced. The obtained data were compared with that noted in 12 healthy newborns and additionally with data of examination done with samples of conserved blood. In newborns with hypertrophic constriction of the pylorus the coefficient of leucine distribution prior the operation was significantly lower than that in the control group and conserved blood. After seven days from operation a significant increase of above coefficient is compared with the control group and erythrocytes in conserved blood has been noted. Above results suggest that disturbances in penetration of leucine through cell membranes of erythrocytes are in association with malnutrition caused by constriction of the pylorus.

  6. Use of the [14C]Leucine Incorporation Technique To Measure Bacterial Production in River Sediments and the Epiphyton

    OpenAIRE

    Fischer, Helmut; Pusch, Martin

    1999-01-01

    Bacterial production is a key parameter for the understanding of carbon cycling in aquatic ecosystems, yet it remains difficult to measure in many aquatic habitats. We therefore tested the applicability of the [14C]leucine incorporation technique for the measurement of bulk bacterial production in various habitats of a lowland river ecosystem. To evaluate the method, we determined (i) extraction efficiencies of bacterial protein from the sediments, (ii) substrate saturation of leucine in sedi...

  7. Metabolism of leucine and alanine in growing rats fed the diets with various protein to energy ratios

    International Nuclear Information System (INIS)

    Tanaka, Hideyuki; Yamaguchi, Michio; Kametaka, Masao

    1975-01-01

    In order to clarify the nutritional significance of metabolism of the carbon skeleton of individual amino acids, the metabolic fates of L-leucine-U- 14 C and L-alanine-U- 14 C were investigated in growing rats fed the diets with various protein calories percents (PC%) at 410 kcal of metabolizable energy. The incorporation of 14 C into body protein in 12 hr after the injection of leucine- 14 C was about 73% of the dose in the 0 and 5 PC% groups, though it decreased with increasing the levels of dietary protein from 10 to 30 PC%. The value of 14 C recovery in body protein almost agreed with the net protein utilization (NPU) determined for the whole egg protein in a similar experimental condition. The 14 C recovery in expired CO 2 and body lipid suggested that the carbon skeleton of leucine is well utilized as an energy source when the dietary carbohydrate is extensively replaced by protein. While, the incorporation of 14 C into body protein from alanine- 14 C was less than about 11% of the dose in all the dietary groups, and the majority of 14 C was recovered in expired CO 2 and body lipid in a remarked contrast to leucine. A similar pattern in urinary excretion of 14 C was obtained for these amino acids, and the refracted rise of 14 C from 10 PC% may give an indication for minimum protein requirements. (auth.)

  8. Post-transcriptional gene silencing of ribosomal protein S6 kinase 1 restores insulin action in leucine-treated skeletal muscle

    DEFF Research Database (Denmark)

    Deshmukh, A; Salehzadeh, F; Metayer-Coustard, S

    2009-01-01

    Excessive nutrients, especially amino acids, impair insulin action on glucose metabolism in skeletal muscle. We tested the hypothesis that the branched-chain amino acid leucine reduces acute insulin action in primary myotubes via a negative feedback mechanism involving ribosomal protein S6 kinase 1...... to excessive leucine. In conclusion, S6K1 plays an important role in the regulation of insulin action on glucose metabolism in skeletal muscle....

  9. Identification of a novel missense mutation of MAF in a Japanese family with congenital cataract by whole exome sequencing: a clinical report and review of literature.

    Science.gov (United States)

    Narumi, Yoko; Nishina, Sachiko; Tokimitsu, Motoharu; Aoki, Yoko; Kosaki, Rika; Wakui, Keiko; Azuma, Noriyuki; Murata, Toshinori; Takada, Fumio; Fukushima, Yoshimitsu; Kosho, Tomoki

    2014-05-01

    Congenital cataracts are the most important cause of severe visual impairment in infants. Genetic factors contribute to the disease development and 29 genes are known to cause congenital cataracts. Identifying the genetic cause of congenital cataracts can be difficult because of genetic heterogeneity. V-maf avian musculoaponeurotic fibrosarcoma oncogene homolog (MAF) encodes a basic region/leucine zipper transcription factor that plays a key role as a regulator of embryonic lens fiber cell development. MAF mutations have been reported to cause juvenile-onset pulverulent cataract, microcornea, iris coloboma, and other anterior segment dysgenesis. We report on six patients in a family who have congenital cataracts were identified MAF mutation by whole exome sequencing (WES). The heterozygous MAF mutation Q303L detected in the present family occurs in a well conserved glutamine residue at the basic region of the DNA-binding domain. All affected members showed congenital cataracts. Three of the six members showed microcornea and one showed iris coloboma. Congenital cataracts with MAF mutation exhibited phenotypically variable cataracts within the family. Review of the patients with MAF mutations supports the notion that congenital cataracts caused by MAF mutations could be accompanied by microcornea and/or iris coloboma. WES is a useful tool for detecting disease-causing mutations in patients with genetically heterogeneous conditions. © 2014 Wiley Periodicals, Inc.

  10. Finding a Leucine in a Haystack: Searching the Proteome for ambigous Leucine-Aspartic Acid motifs

    KAUST Repository

    Arold, Stefan T.

    2016-01-25

    Leucine-aspartic acid (LD) motifs are short helical protein-protein interaction motifs involved in cell motility, survival and communication. LD motif interactions are also implicated in cancer metastasis and are targeted by several viruses. LD motifs are notoriously difficult to detect because sequence pattern searches lead to an excessively high number of false positives. Hence, despite 20 years of research, only six LD motif–containing proteins are known in humans, three of which are close homologues of the paxillin family. To enable the proteome-wide discovery of LD motifs, we developed LD Motif Finder (LDMF), a web tool based on machine learning that combines sequence information with structural predictions to detect LD motifs with high accuracy. LDMF predicted 13 new LD motifs in humans. Using biophysical assays, we experimentally confirmed in vitro interactions for four novel LD motif proteins. Thus, LDMF allows proteome-wide discovery of LD motifs, despite a highly ambiguous sequence pattern. Functional implications will be discussed.

  11. The 15N-leucine single-injection method allows for determining endogenous losses and true digestibility of amino acids in cecectomized roosters.

    Directory of Open Access Journals (Sweden)

    Rujiu Hu

    Full Text Available This study was conducted to assess the influence of dietary protein content in poultry when using the 15N-leucine single-injection method to determine endogenous amino acid losses (EAALs in poultry. Forty-eight cecectomized roosters (2.39 ± 0.23 kg were randomly allocated to eight dietary treatments containing protein levels of 0, 3%, 6%, 9%, 12%, 15%, 18% and 21%. Each bird was precisely fed an experimental diet of 25 g/kg of body weight. After feeding, all roosters were subcutaneously injected with a 15N-leucine solution at a dose of 20 mg/kg of body weight. Blood was sampled 23 h after the injection, and excreta samples were continuously collected during the course of the 48-h experiment. The ratio of 15N-enrichment of leucine in crude mucin to free leucine in plasma ranged from 0.664 to 0.763 and remained relatively consistent (P > 0.05 across all treatments. The amino acid (AA profiles of total endogenous AAs, except isoleucine, alanine, aspartic acid, cysteine, proline and serine, were not influenced (P > 0.05 by dietary protein contents. The predominant endogenous AAs in the excreta were glutamic acid, aspartic acid, threonine, serine and proline. The order of the relative proportions of these predominant AAs also remained relatively constant (P > 0.05. The endogenous losses of total AAs determined with the 15N-leucine single-injection method increased curvilinearly with the dietary protein contents. The true digestibility of most AAs and total AAs was independent of their respective dietary protein levels. Collectively, the 15N-leucine single-injection method is appropriate for determining EAALs and the true digestibility of AAs in poultry fed varying levels of protein-containing ingredients.

  12. Elaboration of biscuits with oatmeal and fat palm with added L-leucine and calcium for sarcopenia

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    Telma Elita Bertolin

    2013-06-01

    Full Text Available The objective of this study was to evaluate the effect of the addition of oatmeal and palm fat in the elaboration of biscuits with added L-leucine and calcium in order to develop a product for sarcopenia in the elderly. The biscuits, or cookies, were elaborated applying a central composite rotational design with surface response methodology, and the significant linear, quadratic and interaction terms were used in the second order mathematical model. Physical, physicochemical and sensory analyses were performed by a trained panel. Based on the best results obtained, three cookie formulations were selected for sensory evaluation by the target group and physicochemical determinations. The formulations with the highest sensory scores for appearance and texture and medium scores for color and expansion index were selected. The addition of calcium and leucine increased significantly the concentration of these components in the biscuits elaborated resulting in a cookie with more than 30% of DRI (Dietary Reference Intake for calcium and leucine. The formulations selected showed high acceptance by the target group; therefore, they can be included in the diet of elderly with sarcopenia as a functional food.

  13. Deuterium incorporation experiments from (3R)- and (3S)-[3-2H]leucine into characteristic isoprenoidal lipid-core of halophilic archaea suggests the involvement of isovaleryl-CoA dehydrogenase.

    Science.gov (United States)

    Yamauchi, Noriaki; Tanoue, Ryo

    2017-11-01

    The stereochemical reaction course for the two C-3 hydrogens of leucine to produce a characteristic isoprenoidal lipid in halophilic archaea was observed using incubation experiments with whole cell Halobacterium salinarum. Deuterium-labeled (3R)- and (3S)-[3- 2 H]leucine were freshly prepared as substrates from 2,3-epoxy-4-methyl-1-pentanol. Incorporation of deuterium from (3S)-[3- 2 H]leucine and loss of deuterium from (3R)-[3- 2 H]leucine in the lipid-core of H. salinarum was observed. Taken together with the results of our previous report, involving the incubation of chiral-labeled [5- 2 H]leucine, these results strongly suggested an involvement of isovaleryl-CoA dehydrogenase in leucine conversion to isoprenoid lipid in halophilic archaea. The stereochemical course of the reaction (anti-elimination) might have been the same as that previously reported for mammalian enzyme reactions. Thus, these results suggested that branched amino acids were metabolized to mevalonate in archaea in a manner similar to other organisms.

  14. Sulforaphane Protects against Cardiovascular Disease via Nrf2 Activation

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    Yang Bai

    2015-01-01

    Full Text Available Cardiovascular disease (CVD causes an unparalleled proportion of the global burden of disease and will remain the main cause of mortality for the near future. Oxidative stress plays a major role in the pathophysiology of cardiac disorders. Several studies have highlighted the cardinal role played by the overproduction of reactive oxygen or nitrogen species in the pathogenesis of ischemic myocardial damage and consequent cardiac dysfunction. Isothiocyanates (ITC are sulfur-containing compounds that are broadly distributed among cruciferous vegetables. Sulforaphane (SFN is an ITC shown to possess anticancer activities by both in vivo and epidemiological studies. Recent data have indicated that the beneficial effects of SFN in CVD are due to its antioxidant and anti-inflammatory properties. SFN activates NF-E2-related factor 2 (Nrf2, a basic leucine zipper transcription factor that serves as a defense mechanism against oxidative stress and electrophilic toxicants by inducing more than a hundred cytoprotective proteins, including antioxidants and phase II detoxifying enzymes. This review will summarize the evidence from clinical studies and animal experiments relating to the potential mechanisms by which SFN modulates Nrf2 activation and protects against CVD.

  15. Genome-Wide Identification and Structural Analysis of bZIP Transcription Factor Genes in Brassica napus.

    Science.gov (United States)

    Zhou, Yan; Xu, Daixiang; Jia, Ledong; Huang, Xiaohu; Ma, Guoqiang; Wang, Shuxian; Zhu, Meichen; Zhang, Aoxiang; Guan, Mingwei; Lu, Kun; Xu, Xinfu; Wang, Rui; Li, Jiana; Qu, Cunmin

    2017-10-24

    The basic region/leucine zipper motif (bZIP) transcription factor family is one of the largest families of transcriptional regulators in plants. bZIP genes have been systematically characterized in some plants, but not in rapeseed ( Brassica napus ). In this study, we identified 247 BnbZIP genes in the rapeseed genome, which we classified into 10 subfamilies based on phylogenetic analysis of their deduced protein sequences. The BnbZIP genes were grouped into functional clades with Arabidopsis genes with similar putative functions, indicating functional conservation. Genome mapping analysis revealed that the BnbZIPs are distributed unevenly across all 19 chromosomes, and that some of these genes arose through whole-genome duplication and dispersed duplication events. All expression profiles of 247 bZIP genes were extracted from RNA-sequencing data obtained from 17 different B . napus ZS11 tissues with 42 various developmental stages. These genes exhibited different expression patterns in various tissues, revealing that these genes are differentially regulated. Our results provide a valuable foundation for functional dissection of the different BnbZIP homologs in B . napus and its parental lines and for molecular breeding studies of bZIP genes in B . napus .

  16. Deregulation of sucrose-controlled translation of a bZIP-type transcription factor results in sucrose accumulation in leaves.

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    Sunil Kumar Thalor

    Full Text Available Sucrose is known to repress the translation of Arabidopsis thaliana AtbZIP11 transcript which encodes a protein belonging to the group of S (S--stands for small basic region-leucine zipper (bZIP-type transcription factor. This repression is called sucrose-induced repression of translation (SIRT. It is mediated through the sucrose-controlled upstream open reading frame (SC-uORF found in the AtbZIP11 transcript. The SIRT is reported for 4 other genes belonging to the group of S bZIP in Arabidopsis. Tobacco tbz17 is phylogenetically closely related to AtbZIP11 and carries a putative SC-uORF in its 5'-leader region. Here we demonstrate that tbz17 exhibits SIRT mediated by its SC-uORF in a manner similar to genes belonging to the S bZIP group of the Arabidopsis genus. Furthermore, constitutive transgenic expression of tbz17 lacking its 5'-leader region containing the SC-uORF leads to production of tobacco plants with thicker leaves composed of enlarged cells with 3-4 times higher sucrose content compared to wild type plants. Our finding provides a novel strategy to generate plants with high sucrose content.

  17. Deregulation of sucrose-controlled translation of a bZIP-type transcription factor results in sucrose accumulation in leaves.

    Science.gov (United States)

    Thalor, Sunil Kumar; Berberich, Thomas; Lee, Sung Shin; Yang, Seung Hwan; Zhu, Xujun; Imai, Ryozo; Takahashi, Yoshihiro; Kusano, Tomonobu

    2012-01-01

    Sucrose is known to repress the translation of Arabidopsis thaliana AtbZIP11 transcript which encodes a protein belonging to the group of S (S--stands for small) basic region-leucine zipper (bZIP)-type transcription factor. This repression is called sucrose-induced repression of translation (SIRT). It is mediated through the sucrose-controlled upstream open reading frame (SC-uORF) found in the AtbZIP11 transcript. The SIRT is reported for 4 other genes belonging to the group of S bZIP in Arabidopsis. Tobacco tbz17 is phylogenetically closely related to AtbZIP11 and carries a putative SC-uORF in its 5'-leader region. Here we demonstrate that tbz17 exhibits SIRT mediated by its SC-uORF in a manner similar to genes belonging to the S bZIP group of the Arabidopsis genus. Furthermore, constitutive transgenic expression of tbz17 lacking its 5'-leader region containing the SC-uORF leads to production of tobacco plants with thicker leaves composed of enlarged cells with 3-4 times higher sucrose content compared to wild type plants. Our finding provides a novel strategy to generate plants with high sucrose content.

  18. Elucidation of the Specific Formation of Homo- and Heterodimeric Forms of ThbZIP1 and Its Role in Stress

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    Xianguang Nie

    2014-06-01

    Full Text Available Protein–protein interactions are important for the molecular understanding of the biological processes of proteins. The dimerization of bZIPs (basic leucine zipper proteins is involved in modifying binding site specificities, altering dimer stability, and permitting a new set of specific protein-to-protein interactions to occur at the promoter. In the present study, we studied the whether ThbZIP1 form homo- and heterodimers using the yeast two-hybrid method. Five bZIP genes were cloned from Tamarix hispida to investigate their interaction with ThbZIP1. Our results showed that ThbZIP1 can form homodimers with itself, and three out of five bZIPs could interact with the ThbZIP1 protein to form heterodimers. Real-time RT-PCR results suggested that these ThbZIPs can all respond to abiotic stresses and abscisic acid (ABA, and shared very similar expression patterns in response to NaCl, ABA or PEG6000. Subcellular localization studies showed that all ThbZIPs are targeted to the nucleus. Our results showed that ThbZIP1 are dimeric proteins, which can form homo- or heterodimers.

  19. Crystallization and preliminary crystallographic studies of the W2 domain of Drosophila melanogaster eukaryotic translation initiation factor 5C domain-containing protein

    International Nuclear Information System (INIS)

    Zhao, Hui; Wang, Hong; Liu, Huihui; Teng, Maikun; Li, Xu

    2012-01-01

    The crystallization and preliminary crystallographic studies of the carboxy-terminal domain of D. melanogaster eukaryotic translation initiation factor 5C domain-containing protein are reported. The Drosophila melanogaster eukaryotic translation initiation factor 5C domain-containing protein (ECP) is composed of two independently folded domains which belong to the basic leucine-zipper and W2 domain-containing protein (BZW) family. Based on the sequence similarity between the C-terminal W2 domain of ECP and some eukaryotic translation initiation factors (such as eIF2B∊, eIF4γ, eIF5 etc.), ECP has been speculated to participate in the translation initiation process. Structural information on the C-terminal W2 domain of ECP would be helpful in understanding the specific cellular function of this protein. Here, the W2 domain of ECP was expressed and crystallized. Crystals grown by the hanging-drop vapour-diffusion method diffracted to 2.70 Å resolution and belonged to space group I4, with unit-cell parameters a = b = 81.05, c = 57.44 Å. The Matthews coefficient suggested that there was one molecule per asymmetric unit in the crystal

  20. Tfe3 expression is closely associated to macrophage terminal differentiation of human hematopoietic myeloid precursors

    International Nuclear Information System (INIS)

    Zanocco-Marani, Tommaso; Vignudelli, Tatiana; Gemelli, Claudia; Pirondi, Sara; Testa, Anna; Montanari, Monica; Parenti, Sandra; Tenedini, Elena; Grande, Alexis; Ferrari, Sergio

    2006-01-01

    The MItf-Tfe family of basic helix-loop-helix leucine zipper (bHLH-Zip) transcription factors encodes four family members: MItf, Tfe3, TfeB and TfeC. In vitro, each protein of the family binds DNA in a homo- or heterodimeric form with other family members. Tfe3 is involved in chromosomal translocations recurrent in different tumors and it has been demonstrated, by in vivo studies, that it plays, redundantly with MItf, an important role in the process of osteoclast formation, in particular during the transition from mono-nucleated to multi-nucleated osteoclasts. Since mono-nucleated osteoclasts derive from macrophages we investigated whether Tfe3 might play a role upstream during hematopoietic differentiation. Here we show that Tfe3 is able to induce mono-macrophagic differentiation of U937 cells, in association with a decrease of cell proliferation and an increase of apoptosis. We also show that Tfe3 does not act physiologically during commitment of CD34+ hematopoietic stem cells (HSCs), since it is not able to direct HSCs toward a specific lineage as observed by clonogenic assay, but is a strong actor of terminal differentiation since it allows human primary myeloblasts' maturation toward the macrophage lineage

  1. Ser298 of MITF, a mutation site in Waardenburg syndrome type 2, is a phosphorylation site with functional significance.

    Science.gov (United States)

    Takeda, K; Takemoto, C; Kobayashi, I; Watanabe, A; Nobukuni, Y; Fisher, D E; Tachibana, M

    2000-01-01

    MITF (microphthalmia-associated transcription factor) is a basic-helix-loop-helix-leucine zipper (bHLHZip) factor which regulates expression of tyrosinase and other melanocytic genes via a CATGTG promoter sequence, and is involved in melanocyte differentiation. Mutations of MITF in mice or humans with Waardenburg syndrome type 2 (WS2) often severely disrupt the bHLHZip domain, suggesting the importance of this structure. Here, we show that Ser298, which locates downstream of the bHLHZip and was previously found to be mutated in individuals with WS2, plays an important role in MITF function. Glycogen synthase kinase 3 (GSK3) was found to phosphorylate Ser298 in vitro, thereby enhancing the binding of MITF to the tyrosinase promoter. The same serine was found to be phosphorylated in vivo, and expression of dominant-negative GSK3beta selectively suppressed the ability of MITF to transactivate the tyrosinase promoter. Moreover, mutation of Ser298, as found in a WS2 family, disabled phos-phorylation of MITF by GSK3beta and impaired MITF function. These findings suggest that the Ser298 is important for MITF function and is phosphorylated probably by GSK3beta.

  2. Endoplasmic reticulum stress-responsive transcription factor ATF6α directs recruitment of the Mediator of RNA polymerase II transcription and multiple histone acetyltransferase complexes.

    Science.gov (United States)

    Sela, Dotan; Chen, Lu; Martin-Brown, Skylar; Washburn, Michael P; Florens, Laurence; Conaway, Joan Weliky; Conaway, Ronald C

    2012-06-29

    The basic leucine zipper transcription factor ATF6α functions as a master regulator of endoplasmic reticulum (ER) stress response genes. Previous studies have established that, in response to ER stress, ATF6α translocates to the nucleus and activates transcription of ER stress response genes upon binding sequence specifically to ER stress response enhancer elements in their promoters. In this study, we investigate the biochemical mechanism by which ATF6α activates transcription. By exploiting a combination of biochemical and multidimensional protein identification technology-based mass spectrometry approaches, we have obtained evidence that ATF6α functions at least in part by recruiting to the ER stress response enhancer elements of ER stress response genes a collection of RNA polymerase II coregulatory complexes, including the Mediator and multiple histone acetyltransferase complexes, among which are the Spt-Ada-Gcn5 acetyltransferase (SAGA) and Ada-Two-A-containing (ATAC) complexes. Our findings shed new light on the mechanism of action of ATF6α, and they outline a straightforward strategy for applying multidimensional protein identification technology mass spectrometry to determine which RNA polymerase II transcription factors and coregulators are recruited to promoters and other regulatory elements to control transcription.

  3. Cryptocephal, the Drosophila melanogaster ATF4, is a specific coactivator for ecdysone receptor isoform B2.

    Directory of Open Access Journals (Sweden)

    Sebastien A Gauthier

    Full Text Available The ecdysone receptor is a heterodimer of two nuclear receptors, the Ecdysone receptor (EcR and Ultraspiracle (USP. In Drosophila melanogaster, three EcR isoforms share common DNA and ligand-binding domains, but these proteins differ in their most N-terminal regions and, consequently, in the activation domains (AF1s contained therein. The transcriptional coactivators for these domains, which impart unique transcriptional regulatory properties to the EcR isoforms, are unknown. Activating transcription factor 4 (ATF4 is a basic-leucine zipper transcription factor that plays a central role in the stress response of mammals. Here we show that Cryptocephal (CRC, the Drosophila homolog of ATF4, is an ecdysone receptor coactivator that is specific for isoform B2. CRC interacts with EcR-B2 to promote ecdysone-dependent expression of ecdysis-triggering hormone (ETH, an essential regulator of insect molting behavior. We propose that this interaction explains some of the differences in transcriptional properties that are displayed by the EcR isoforms, and similar interactions may underlie the differential activities of other nuclear receptors with distinct AF1-coactivators.

  4. Deletion of Batf3-dependent antigen-presenting cells does not affect atherosclerotic lesion formation in mice.

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    Jesus Gil-Pulido

    Full Text Available Atherosclerosis is the main underlying cause for cardiovascular events such as myocardial infarction and stroke and its development might be influenced by immune cells. Dendritic cells (DCs bridge innate and adaptive immune responses by presenting antigens to T cells and releasing a variety of cytokines. Several subsets of DCs can be discriminated that engage specific transcriptional pathways for their development. Basic leucine zipper transcription factor ATF-like 3 (Batf3 is required for the development of classical CD8α+ and CD103+ DCs. By crossing mice deficient in Batf3 with atherosclerosis-prone low density lipoprotein receptor (Ldlr-/--deficient mice we here aimed to further address the contribution of Batf3-dependent CD8α+ and CD103+ antigen-presenting cells to atherosclerosis. We demonstrate that deficiency in Batf3 entailed mild effects on the immune response in the spleen but did not alter atherosclerotic lesion formation in the aorta or aortic root, nor affected plaque phenotype in low density lipoprotein receptor-deficient mice fed a high fat diet. We thus provide evidence that Batf3-dependent antigen-presenting cells do not have a prominent role in atherosclerosis.

  5. Redundant and distinct functions of the ABA response loci ABA-INSENSITIVE(ABI)5 and ABRE-BINDING FACTOR (ABF)3.

    Science.gov (United States)

    Finkelstein, Ruth; Gampala, Srinivas S L; Lynch, Tim J; Thomas, Terry L; Rock, Christopher D

    2005-09-01

    Abscisic acid-responsive gene expression is regulated by numerous transcription factors, including a subgroup of basic leucine zipper factors that bind to the conserved cis-acting sequences known as ABA-responsive elements. Although one of these factors, ABA-insensitive 5 (ABI5), was identified genetically, the paucity of genetic data for the other family members has left it unclear whether they perform unique functions or act redundantly to ABI5 or each other. To test for potential redundancy with ABI5, we identified the family members with most similar effects and interactions in transient expression systems (ABF3 and ABF1), then characterized loss-of-function lines for those loci. The abf1 and abf3 monogenic mutant lines had at most minimal effects on germination or seed-specific gene expression, but the enhanced ABA- and stress-resistance of abf3 abi5 double mutants revealed redundant action of these genes in multiple stress responses of seeds and seedlings. Although ABI5, ABF3, and ABF1 have some overlapping effects, they appear to antagonistically regulate each other's expression at specific stages. Consequently, loss of any one factor may be partially compensated by increased expression of other family members.

  6. Activation of mTORC1 by leucine is potentiated by branched-chain amino acids and even more so by essential amino acids following resistance exercise.

    Science.gov (United States)

    Moberg, Marcus; Apró, William; Ekblom, Björn; van Hall, Gerrit; Holmberg, Hans-Christer; Blomstrand, Eva

    2016-06-01

    Protein synthesis is stimulated by resistance exercise and intake of amino acids, in particular leucine. Moreover, activation of mammalian target of rapamycin complex 1 (mTORC1) signaling by leucine is potentiated by the presence of other essential amino acids (EAA). However, the contribution of the branched-chain amino acids (BCAA) to this effect is yet unknown. Here we compare the stimulatory role of leucine, BCAA, and EAA ingestion on anabolic signaling following exercise. Accordingly, eight trained volunteers completed four sessions of resistance exercise during which they ingested either placebo, leucine, BCAA, or EAA (including the BCAA) in random order. Muscle biopsies were taken at rest, immediately after exercise, and following 90 and 180 min of recovery. Following 90 min of recovery the activity of S6 kinase 1 (S6K1) was greater than at rest in all four trials (PlaceboBCAABCAA. However, after 180 min of recovery this difference between EAA and BCAA had disappeared, although with both these supplements the increases were still higher than with leucine (40%, P BCAA. Copyright © 2016 the American Physiological Society.

  7. Leucine pulses enhance skeletal muscle protein synthesis during continuous feeding in neonatal pigs

    Science.gov (United States)

    Infants unable to maintain oral feeding can be nourished by orogastric tube. We have shown that orogastric continuous feeding restricts muscle protein synthesis compared with intermittent bolus feeding in neonatal pigs. To determine whether leucine leu infusion can be used to enhance protein synthes...

  8. In vitro conditions for 14C-leucine incorporation into the protein of cultured ovaries of the silkworm, Bombyx mori

    International Nuclear Information System (INIS)

    Miyadai, Toshiaki; Yamashita, Okitsugu

    1980-01-01

    Vitellogenic ovaries of silkworm pupae were incubated in vitro in different media based on the Wyatt's medium to establish an adequate condition for culture of silkworm ovaries. Incorporation of 14 C-leucine into protein fraction was determined to assess the biochemical activity of the ovary. When ovaries were incubated in vitro for a short time by 6 hr, a saturation kinetics of incorporation of the labelled leucine was shown. Sequential substitution of K + ion to Na + ion in the medium had no effect on the incorporation of 14 C-leucine, but Mg 2+ ion appeared to stimulate synthetic activity at more than 10 mM. The activity was not affected at pH range 5.0-7.2. Neither different sugars, nor vitellogenin nor lipoprotein prepared from silkworm haemolymph affected the incorporation of 14 C-leucine, when added into the medium. The synthesis of protein depended upon the developmental stages of the cultured ovaries and was most active in 6-day-old ovary. Ovaries developing in pupal body showed comparable changes in synthetic activity. It is concluded that the chemical composition of the medium does not exert a strict effect on synthetic activity of protein in short-term cultures and the ovaries cultured in vitro maintain the activity comparable with those found in in situ condition. (author)

  9. Effects of a leucine and pyridoxine-containing nutraceutical on body weight and composition in obese subjects

    Directory of Open Access Journals (Sweden)

    Zemel MB

    2013-08-01

    Full Text Available Michael B Zemel,1,2 Antje Bruckbauer11NuSirt Sciences Inc, 2Nutrition Department, The University of Tennessee, Knoxville, TN, USABackground: We recently demonstrated leucine to modulate energy partitioning between adipose tissue and muscle. Further, leucine exhibits a synergy with B6, resulting in reduced adipocyte lipid storage coupled with increased muscle fat oxidation. Accordingly, a nutraceutical (NuShape™ containing 2.25 g leucine and 30 mg B6 increased fat oxidation by > 30 g/day in a 28-day randomized controlled trial. The present study evaluated the long-term efficacy of this combination in modulating body weight and composition.Methods: Two 24-week, placebo-controlled, randomized trials, one with weight maintenance (n = 20 and one hypocaloric (-500 kcal/day; n = 24, were conducted using the nutraceutical Nushape in obese subjects.Results: The supplement resulted in fat loss in the maintenance study (-1.12 ± 0.36 and -1.82 ± 0.70 kg at 12 and 24 weeks, P < 0.01 versus placebo while no change was found in the placebo group. In the hypocaloric study, the supplement group lost up to twice as much weight (6.18 ± 1.02 versus 3.40 ± 0.81 kg at 12 weeks and 8.15 ± 1.33 versus 5.25 ± 1.13 kg at 24 weeks, P < 0.01 and fat (4.96 ± 0.61 versus 2.31 ± 0.53 kg at 12 weeks and 7.00 ± 0.95 versus 4.22 ± 0.74 kg at 24 weeks, P < 0.01 than the placebo group.Conclusion: This nutraceutical combination results in significant fat loss in the absence of caloric restriction and markedly enhances weight and fat loss by 50%–80% over a 24-week period.Keywords: leucine, vitamin B6, pyridoxine, fat loss, weight loss, caloric restriction, adiposity, body composition

  10. Branched-chain amino acid interactions with reference to amino acid requirements in adult men: Valine metabolism at different leucine intakes

    International Nuclear Information System (INIS)

    Pelletier, V.; Marks, L.; Wagner, D.A.; Hoerr, R.A.; Young, V.R.

    1991-01-01

    The authors explored whether the oxidation of valine and by implication the physiological requirement for this amino acid are affected by changes in leucine intake over a physiological range. Six young adult men received, in random order, four L-amino acid-based diets for 5 d supplying either 20 or 10 mg valine.kg body wt-1.d-1, each in combination with 80 or 40 mg leucine.kg-1.d-1. On day 6 subjects were studied with an 8-h continuous intravenous infusion of [1-13C]valine (and [2H3]leucine) to determine valine oxidation in the fasted state (first 3 h) and fed state (last 5 h). Valine oxidation in the fasted state was similar among all diets but was lower (P less than 0.05) in the fed state for the 10 vs 20 mg valine.kg-1.d-1 intake. Leucine intake did not affect valine oxidation. Mean daily valine balance approximated +1.3 mg.kg-1.d-1 for the 20-mg intake and -1.6 mg.kg-1.d-1 for the 10-mg intake. These findings support our previously suggested mean valine requirement estimate of approximately 20 mg.kg-1.d-1

  11. Leucine insertion caused by a yeast amber suppressor

    Energy Technology Data Exchange (ETDEWEB)

    Liebman, S W [Univ. of Rochester School of Medicine and Dentistry, NY; Stewart, J W; Parker, J H; Sherman, F

    1977-01-01

    The amber suppressor SUP52 can cause the production of approximately 15 to 20% of the normal amount of iso-l-cytochrome c when coupled to the amber (UAG) mutant cyc1-76. The suppressed iso-l-cytochrome c contains a residue of leucine at the position corresponding to the site of the amber codon. SUP52 also supresses another amber allele cyc1-179, but only with a low efficiency of approximately 2%. It does not appear to act at all on ochre (UAA) mutants. SUP52 was found to be on the left arm of chromosome X closely linked to the centromere.

  12. Expression, purification and preliminary biochemical and structural characterization of the leucine rich repeat namesake domain of leucine rich repeat kinase 2.

    Science.gov (United States)

    Vancraenenbroeck, Renée; Lobbestael, Evy; Weeks, Stephen D; Strelkov, Sergei V; Baekelandt, Veerle; Taymans, Jean-Marc; De Maeyer, Marc

    2012-03-01

    Mutations in leucine-rich repeat kinase 2 (LRRK2) are the most common cause of familial Parkinson's disease. Much research effort has been directed towards the catalytic core region of LRRK2 composed of GTPase (ROC, Ras of complex proteins) and kinase domains and a connecting COR (C-terminus of ROC) domain. In contrast, the precise functions of the protein-protein interaction domains, such as the leucine-rich repeat (LRR) domain, are not known. In the present study, we modeled the LRRK2 LRR domain (LRR(LRRK2)) using a template assembly approach, revealing the presence of 14 LRRs. Next, we focused on the expression and purification of LRR(LRRK2) in Escherichia coli. Buffer optimization revealed that the protein requires the presence of a zwitterionic detergent, namely Empigen BB, during solubilization and the subsequent purification and characterization steps. This indicates that the detergent captures the hydrophobic surface patches of LRR(LRRK2) thereby suppressing its aggregation. Circular dichroism (CD) spectroscopy measured 18% α-helices and 21% β-sheets, consistent with predictions from the homology model. Size exclusion chromatography (SEC) and dynamic light scattering measurements showed the presence of a single species, with a Stokes radius corresponding to the model dimensions of a protein monomer. Furthermore, no obvious LRR(LRRK2) multimerization was detected via cross-linking studies. Finally, the LRR(LRRK2) clinical mutations did not influence LRR(LRRK2) secondary, tertiary or quaternary structure as determined via SEC and CD spectroscopy. We therefore conclude that these mutations are likely to affect putative LRR(LRRK2) inter- and intramolecular interactions. Copyright © 2011 Elsevier B.V. All rights reserved.

  13. Effects of tannic acid on trypsin and leucine aminopeptidase activities in gypsy moth larval midgut

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    Mrdaković Marija

    2013-01-01

    Full Text Available The effects of allelochemical stress on genetic variations in the specific activities of gypsy moth digestive enzymes (trypsin and leucine aminopeptidase and relative midgut mass (indirect measure of food consumption, as well as variability in their plasticity, were investigated in fifth instar gypsy moths originating from two populations with different trophic adaptations (oak and locust-tree forests. Thirty-two full-sib families from the Quercus population and twenty-six full-sib families from the Robinia population were reared on an artificial diet with or without supplementation with tannic acid. Between population differences were observed as higher average specific activity of trypsin and relative midgut mass in larvae from the Robinia population. Significant broad-sense heritabilities were observed for the specific activity of trypsin in the control state, and for specific activity of leucine aminopeptidase in a stressful environment. Significantly lower heritability for relative midgut mass was recorded in larvae from the Robinia population reared under stressful conditions. Significant variability of trypsin plasticity in larvae from both populations and significant variability of leucine aminopeptidase plasticity in larvae from the Robinia population point to the potential for the evolution of enzyme adaptive plastic responses to the presence of stressor. Non-significant across-environment genetic correlations do not represent a constraint for the evolution of enzyme plasticity. [Projekat Ministarstva nauke Republike Srbije, br. 173027

  14. Cloaked similarity between HIV-1 and SARS-CoV suggests an anti-SARS strategy

    Directory of Open Access Journals (Sweden)

    Kliger Yossef

    2003-09-01

    Full Text Available Abstract Background Severe acute respiratory syndrome (SARS is a febrile respiratory illness. The disease has been etiologically linked to a novel coronavirus that has been named the SARS-associated coronavirus (SARS-CoV, whose genome was recently sequenced. Since it is a member of the Coronaviridae, its spike protein (S2 is believed to play a central role in viral entry by facilitating fusion between the viral and host cell membranes. The protein responsible for viral-induced membrane fusion of HIV-1 (gp41 differs in length, and has no sequence homology with S2. Results Sequence analysis reveals that the two viral proteins share the sequence motifs that construct their active conformation. These include (1 an N-terminal leucine/isoleucine zipper-like sequence, and (2 a C-terminal heptad repeat located upstream of (3 an aromatic residue-rich region juxtaposed to the (4 transmembrane segment. Conclusions This study points to a similar mode of action for the two viral proteins, suggesting that anti-viral strategy that targets the viral-induced membrane fusion step can be adopted from HIV-1 to SARS-CoV. Recently the FDA approved Enfuvirtide, a synthetic peptide corresponding to the C-terminal heptad repeat of HIV-1 gp41, as an anti-AIDS agent. Enfuvirtide and C34, another anti HIV-1 peptide, exert their inhibitory activity by binding to a leucine/isoleucine zipper-like sequence in gp41, thus inhibiting a conformational change of gp41 required for its activation. We suggest that peptides corresponding to the C-terminal heptad repeat of the S2 protein may serve as inhibitors for SARS-CoV entry.

  15. Excessive Leucine-mTORC1-Signalling of Cow Milk-Based Infant Formula: The Missing Link to Understand Early Childhood Obesity

    OpenAIRE

    Melnik, Bodo C.

    2012-01-01

    Increased protein supply by feeding cow-milk-based infant formula in comparison to lower protein content of human milk is a well-recognized major risk factor of childhood obesity. However, there is yet no conclusive biochemical concept explaining the mechanisms of formula-induced childhood obesity. It is the intention of this article to provide the biochemical link between leucine-mediated signalling of mammalian milk proteins and adipogenesis as well as early adipogenic programming. Leucine ...

  16. Activation of mTORC1 by leucine is potentiated by branched-chain amino acids and even more so by essential amino acids following resistance exercise

    DEFF Research Database (Denmark)

    Moberg, Marcus; Apró, William; Ekblom, Björn

    2016-01-01

    Protein synthesis is stimulated by resistance exercise and intake of amino acids, in particular leucine. Moreover, activation of mammalian target of rapamycin complex 1 (mTORC1) signaling by leucine is potentiated by the presence of other essential amino acids (EAA). However, the contribution...... of the branched-chain amino acids (BCAA) to this effect is yet unknown. Here we compare the stimulatory role of leucine, BCAA, and EAA ingestion on anabolic signaling following exercise. Accordingly, eight trained volunteers completed four sessions of resistance exercise during which they ingested either placebo...

  17. Further observations on incorporation of the 14C-leucine into proteins by freshly secreted milk

    International Nuclear Information System (INIS)

    Singh, L.N.

    1976-01-01

    Using freshly secreted bovine milk, no incorporation of DL (1- 14 C)-leucine was observed in the total milk proteins and acid precipitated casein, when these protein fractions were isolated from skim milk. A significant portion of the radioactivity however, remained associated with the heat coagulable whey proteins and proteose-peptone fractions. This association was shown to be due to non enzymatic physical sequestering of the radioactive amino acid or its metabolites with these proteins. Most of the radioactivity was associated with the cream layer proteins and the cellular fraction. The results obtained using filtered milk, incubated milk and certain antibiotics also indicated that the incorporation of 14 C leucine into proteins by freshly secreted milk may be a purely microbial process and physical sequestering of an amino acids with milk proteins. (author)

  18. Compartmental Modeling and Dosimetry of in Vivo Metabolic Studies of Leucine and Three Secretory Proteins in Humans Using Radioactive Tracers

    Science.gov (United States)

    Venkatakrishnan, Vaidehi

    1995-01-01

    Physical and mathematical models provide a systematic means of looking at biological systems. Radioactive tracer kinetic studies open a unique window to study complex tracee systems such as protein metabolism in humans. This research deals with compartmental modeling of tracer kinetic data on leucine and apolipoprotein metabolism obtained using an endogenous tritiated leucine tracer administered as a bolus, and application of compartmental modeling techniques for dosimetric evaluation of metabolic studies of radioiodinated apolipoproteins. Dr. Waldo R. Fisher, Department of Medicine, was the coordinating research supervisor and the work was carried out in his laboratory. A compartmental model for leucine kinetics in humans has been developed that emphasizes its recycling pathways which were examined over two weeks. This model builds on a previously published model of Cobelli et al, that analyzed leucine kinetic data up to only eight hours. The proposed model includes different routes for re-entry of leucine from protein breakdown into plasma accounting for proteins which turn over at different rates. This new model successfully incorporates published models of three secretory proteins: albumin, apoA-I, and VLDL apoB, in toto thus increasing its validity and utility. The published model of apoA-I, based on an exogenous radioiodinated tracer, was examined with data obtained using an endogenous leucine tracer using compartmental techniques. The analysis concludes that the major portion of apoA-I enters plasma by a fast pathway but the major fraction of apoA-I in plasma resides with a second slow pathway; further the study is suggestive of a precursor-product relationship between the two plasma apoA-I pools. The possible relevance of the latter suggestion to the aberrant kinetics of apoA-I in Tangier disease is discussed. The analysis of apoA-II data resulted in similar conclusions. A methodology for evaluating the dosimetry of radioiodinated apolipoproteins by

  19. Effect of voluntary exercise and dietary protein levels on incorporation of 14C-leucine into protein by mice liver slices in vitro

    International Nuclear Information System (INIS)

    Yashiro, Masanori; Kimura, Shuichi

    1983-01-01

    The effect of voluntary exercise on incorporation of 14 C-leucine into protein by mice liver slices in vitro were examined with mice fed 4 %, 6 % and 20 % protein diets. The incorporation of 14 C-leucine increased as dietary protein levels decreased and was significantly higher in liver slices of exercise groups than in slices of non-exercise groups. (author)

  20. Impact of prolonged leucine supplementation on protein synthesis and lean growth in neonatal pigs

    Science.gov (United States)

    Most low-birth weight infants experience extrauterine growth failure due to reduced nutrient intake as a result of feeding intolerance. The objective of this study was to determine whether prolonged enteral leucine supplementation improves lean growth in neonatal pigs fed a restricted protein diet. ...

  1. Cloning and characterization of an aromatic amino acid and leucine permease of Penicillium chrysogenum

    NARCIS (Netherlands)

    Trip, Hein; Evers, Melchior E.; Konings, Wil N.; Driessen, Arnold J.M.

    2002-01-01

    The gene encoding the amino acid permease ArlP (Aromatic and leucine Permease) was isolated from the filamentous fungus Penicillium chrysogenum after PCR using degenerated oligonucleotides based on conserved regions of fungal amino acid permeases. The cDNA clone was used for expression of the

  2. Peptide synthesis by enzymatic catalysis: new application to the total radiosynthesis of the tritiated leucine-enkephalin hormone, using Y carboxypeptidase

    International Nuclear Information System (INIS)

    Hellio, F.

    1986-01-01

    A new method of enzymatic labelling of peptide hormones is described. The enzyme used, a protease, Y carboxypeptidase is able, in some conditions, to catalyze the formation of peptide bounds. This property has been used for the synthesis of a pentapeptide, the tritiated leucine-enkephalin, with the incorporation of every radioactive amino acid. The specific radioactivity of the labelled molecule is 139 Ci/mmole and its biological properties (receptor binding and immunoreactivity) are identical with native leucine-enkephalin properties [fr

  3. Fine-tuning of T-cell development by the CD3γ di-leucine-based TCR-sorting motif

    DEFF Research Database (Denmark)

    Lauritsen, Jens Peter Holst; Boding, Lasse; Buus, Terkild B

    2015-01-01

    The CD3γ di-leucine-based (diL) receptor-sorting motif plays a central role in TCR down-regulation and in clonal expansion of virus-specific T cells. However, the role of the CD3γ diL motif in T-cell development is not known. In this study, we show that protein kinase C-induced TCR down-regulatio......The CD3γ di-leucine-based (diL) receptor-sorting motif plays a central role in TCR down-regulation and in clonal expansion of virus-specific T cells. However, the role of the CD3γ diL motif in T-cell development is not known. In this study, we show that protein kinase C-induced TCR down...

  4. Postprandial leucine and insulin responses and toxicological effects of a novel whey protein hydrolysate-based supplement in rats

    Directory of Open Access Journals (Sweden)

    Toedebusch Ryan G

    2012-06-01

    Full Text Available Abstract The purpose of this study was: aim 1 compare insulin and leucine serum responses after feeding a novel hydrolyzed whey protein (WPH-based supplement versus a whey protein isolate (WPI in rats during the post-absorptive state, and aim 2 to perform a thorough toxicological analysis on rats that consume different doses of the novel WPH-based supplement over a 30-day period. In male Wistar rats (~250 g, n = 40, serum insulin and leucine concentrations were quantified up to 120 min after one human equivalent dose of a WPI or the WPH-based supplement. In a second cohort of rats (~250 g, n = 20, we examined serum/blood and liver/kidney histopathological markers after 30 days of feeding low (1human equivalent dose, medium (3 doses and high (6 doses amounts of the WPH-based supplement. In aim 1, higher leucine levels existed at 15 min after WPH vs. WPI ingestion (p = 0.04 followed by higher insulin concentrations at 60 min (p = 0.002. In aim 2, liver and kidney histopathology/toxicology markers were not different 30 days after feeding with low, medium, high dose WPH-based supplementation or water only. There were no between-condition differences in body fat or lean mass or circulating clinical chemistry markers following the 30-day feeding intervention in aim 2. In comparison to WPI, acute ingestion of a novel WPH-based supplement resulted in a higher transient leucine response with a sequential increase in insulin. Furthermore, chronic ingestion of the tested whey protein hydrolysate supplement appears safe.

  5. Molecular cloning and characterization of leucine aminopeptidase gene from Taenia pisiformis.

    Science.gov (United States)

    Zhang, Shaohua; Cai, Xuepeng; Luo, Xuenong; Wang, Shuai; Guo, Aijiang; Hou, Junling; Wu, Run

    2018-03-01

    Leucine aminopeptidase (LAP, EC: 3.4.11.1) is an important metalloexopeptidase that catalyze the hydrolysis of amino-terminal leucine residues from polypeptides and proteins. In this study, a full length of cDNA encoding leucine aminopeptidase of Taenia pisiformis (TpLAP) was cloned by rapid amplification of cDNA-ends using the polymerase chain reaction (RACE-PCR) method. The full-length cDNA of the TpLAP gene is 1823 bp and contains a 1569 bp ORF encoding 533 amino acids with a putative mass of 56.4 kDa. TpLAP contains two characteristic motifs of the M17LAP family in the C-terminal sequence: the metal binding site 265-[VGKG]-271 and the catalytic domain motif 351-[NTDAEGRL]-357. The soluble GST-TpLAP protein was expressed in Escherichia coli Transetta (DE3) and four specific anti-TpLAP monoclonal antibodies (mAbs) were prepared. In enzymatic assays, the optimal activity was observed at pH 9.5 at 45 °C. GST-TpLAP displayed a hydrolyzing activity for the Leu-pNA substrate with a maximum activity of 46 U/ml. The enzymatic activity was significantly enhanced by Mn 2+ and completely inhibited by 20 nM bestatin and 0.15 mM EDTA. The native TpLAP was detected specifically in ES components of adult T. pisiformis by western blotting using anti-TpLAP mAb as a probe. Quantitative real-time PCR revealed that the TpLAP gene was expressed at a high level in adult worm tissues, especially in the gravid proglottids (50.71-fold). Immunolocalization analysis showed that TpLAP was located primarily in the subtegumental parenchyma zone and the uterine wall of adult worms. Our results indicate that TpLAP is a new member of the M17LAP family and can be considered as a stage-differentially expressed protein. These findings might provide new insights into the study of the mechanisms of growth, development and survival of T. pisiformis in the final host and have potential value as an attractive target for drug therapy or vaccine intervention. Copyright © 2018 Elsevier Inc

  6. Lysine and Leucine Deficiencies Affect Myocytes Development and IGF Signaling in Gilthead Sea Bream (Sparus aurata.

    Directory of Open Access Journals (Sweden)

    Sheida Azizi

    Full Text Available Optimizing aquaculture production requires better knowledge of growth regulation and improvement in diet formulation. A great effort has been made to replace fish meal for plant protein sources in aquafeeds, making necessary the supplementation of such diets with crystalline amino acids (AA to cover the nutritional requirements of each species. Lysine and Leucine are limiting essential AA in fish, and it has been demonstrated that supplementation with them improves growth in different species. However, the specific effects of AA deficiencies in myogenesis are completely unknown and have only been studied at the level of hepatic metabolism. It is well-known that the TOR pathway integrates the nutritional and hormonal signals to regulate protein synthesis and cell proliferation, to finally control muscle growth, a process also coordinated by the expression of myogenic regulatory factors (MRFs. This study aimed to provide new information on the impact of Lysine and Leucine deficiencies in gilthead sea bream cultured myocytes examining their development and the response of insulin-like growth factors (IGFs, MRFs, as well as key molecules involved in muscle growth regulation like TOR. Leucine deficiency did not cause significant differences in most of the molecules analyzed, whereas Lysine deficiency appeared crucial in IGFs regulation, decreasing significantly IGF-I, IGF-II and IGF-IRb mRNA levels. This treatment also down-regulated the gene expression of different MRFs, including Myf5, Myogenin and MyoD2. These changes were also corroborated by a significant decrease in proliferation and differentiation markers in the Lysine-deficient treatment. Moreover, both Lysine and Leucine limitation induced a significant down-regulation in FOXO3 gene expression, which deserves further investigation. We believe that these results will be relevant for the production of a species as appreciated for human consumption as it is gilthead sea bream and demonstrates

  7. Leucine-rich repeat-containing G-protein-coupled receptors as markers of adult stem cells

    NARCIS (Netherlands)

    Barker, N.; Clevers, H.

    2010-01-01

    Molecular markers are used to characterize and track adult stem cells. Colon cancer research has led to the identification of 2 related receptors, leucine-rich repeat-containing, G-protein-coupled receptors (Lgr)5 and Lgr6, that are expressed by small populations of cells in a variety of adult

  8. Leucine incorporation by aerobic anoxygenic phototrophic bacteria in the Delaware estuary

    OpenAIRE

    Stegman, Monica R; Cottrell, Matthew T; Kirchman, David L

    2014-01-01

    Aerobic anoxygenic phototrophic (AAP) bacteria are well known to be abundant in estuaries, coastal regions and in the open ocean, but little is known about their activity in any aquatic ecosystem. To explore the activity of AAP bacteria in the Delaware estuary and coastal waters, single-cell 3H-leucine incorporation by these bacteria was examined with a new approach that combines infrared epifluorescence microscopy and microautoradiography. The approach was used on samples from the Delaware c...

  9. Effect of leucine supplementation on fat free mass with prolonged hypoxic exposure during a 13-day trek to Everest Base Camp: a double-blind randomized study.

    Science.gov (United States)

    Wing-Gaia, Stacie L; Gershenoff, Dana C; Drummond, Micah J; Askew, E Wayne

    2014-03-01

    Loss of body weight and fat-free mass (FFM) are commonly noted with prolonged exposure to hypobaric hypoxia. Recent evidence suggests protein supplementation, specifically leucine, may potentially attenuate loss of FFM in subcaloric conditions during normoxia. The purpose of this study was to determine if leucine supplementation would prevent the loss of FFM in subcaloric conditions during prolonged hypoxia. Eighteen physically active male (n = 10) and female (n = 8) trekkers completed a 13-day trek in Nepal to Everest Base Camp with a mean altitude of 4140 m (range 2810-5364 m). In this double-blind study, participants were randomized to ingest either leucine (LEU) (7 g leucine, 93 kcal, 14.5 g whey-based protein) or an isocaloric isonitrogenous control (CON) (0.3 g LEU, 93 kcal, 11.3 g collagen protein) twice daily prior to meals. Body weight, body composition, and circumferences of bicep, thigh, and calf were measured pre- and post-trek. There was a significant time effect for body weight (-2.2% ± 1.7%), FFM (-1.7% ± 1.5%), fat mass (-4.0% ± 6.9%), and circumferences (p FFM (CON -2.1 ± 1.5%; LEU -1.2 ± 1.6%), fat mass (CON -2.9% ± 5.9%; LEU -5.4% ± 8.1%), or circumferences. Although a significant loss of body weight, FFM, and fat mass was noted in 13 days of high altitude exposure, FFM loss was not attenuated by leucine. Future studies are needed to determine if leucine attenuates loss of FFM with longer duration high altitude exposure.

  10. A new method to measure muscle protein synthesis in humans by endogenously introduced d9-leucine and using blood for precursor enrichment determination

    Science.gov (United States)

    Tran, Lee; Masters, Haley; Roust, Lori R; Katsanos, Christos S

    2015-01-01

    Enrichment from the easily accessible blood amino acid pool is commonly used as precursor enrichment to calculate rates of muscle protein fractional synthesis in relevant human studies in lieu of the less accessible muscle fluid amino acid pool. However, the accuracy of this approach depends largely on the extent to which there is low discrepancy in free amino acid enrichment between blood and muscle. Steady-state gradient (i.e., ratio) of amino acid enrichment between blood and muscle fluid in the basal state and in response to amino acid infusion were determined in five healthy subjects, and in association with two separate tracers: d9-leucine, introduced endogenously by the metabolism of d10-leucine (i.e., l-[2,3,3,4,5,5,5,6,6,6-2H10]leucine) infused in blood, and 13C6-phenylalanine introduced/infused in blood. The blood-to-muscle fluid amino acid enrichment ratio was lower (P enrichment introduced endogenously by intravenous infusion of d10-leucine provides a closer estimate of the muscle fluid amino acid enrichment, and its associated changes, than blood phenylalanine enrichment to calculate rates of muscle protein synthesis in humans. PMID:26243214

  11. Alterations in protein and amino acid metabolism in rats fed a branched-chain amino acid- or leucine-enriched diet during postprandial and postabsorptive states.

    Science.gov (United States)

    Holecek, Milan; Siman, Pavel; Vodenicarovova, Melita; Kandar, Roman

    2016-01-01

    Many people believe in favourable effects of branched-chain amino acids (BCAAs; valine, leucine, and isoleucine), especially leucine, on muscle protein balance and consume BCAAs for many years. We determined the effects of the chronic intake of a BCAA- or leucine-enriched diet on protein and amino acid metabolism in fed and postabsorptive states. Rats were fed a standard diet, a diet with a high content of valine, leucine, and isoleucine (HVLID), or a high content of leucine (HLD) for 2 months. Half of the animals in each group were sacrificed in the fed state on the last day, and the other half were sacrificed after overnight fast. Protein synthesis was assessed using the flooding dose method (L-[3,4,5-(3)H]phenylalanine), proteolysis on the basis of chymotrypsin-like activity (CHTLA) of proteasome and cathepsin B and L activities. Chronic intake of HVLID or HLD enhanced plasma levels of urea, alanine and glutamine. HVLID also increased levels of all three BCAA and branched-chain keto acids (BCKA), HLD increased leucine, ketoisocaproate and alanine aminotransferase and decreased valine, ketovaline, isoleucine, ketoisoleucine, and LDL cholesterol. Tissue weight and protein content were lower in extensor digitorum longus muscles in the HLD group and higher in kidneys in the HVLID and HLD groups. Muscle protein synthesis in postprandial state was higher in the HVLID group, and CHTLA was lower in muscles of the HVLID and HLD groups compared to controls. Overnight starvation enhanced alanine aminotransferase activity in muscles, and decreased protein synthesis in gastrocnemius (in HVLID group) and extensor digitorum longus (in HLD group) muscles more than in controls. Effect of HVLID and HLD on CHTLA in muscles in postabsorptive state was insignificant. The results failed to demonstrate positive effects of the chronic consumption of a BCAA-enriched diet on protein balance in skeletal muscle and indicate rather negative effects from a leucine-enriched diet. The primary

  12. CD3 gamma contains a phosphoserine-dependent di-leucine motif involved in down-regulation of the T cell receptor

    DEFF Research Database (Denmark)

    Dietrich, J; Hou, X; Wegener, A M

    1994-01-01

    -regulation of the TCR. Furthermore, analysis of a series of CD3 gamma truncation mutants indicated that in addition to S126 phosphorylation a motif C-terminal of S126 was required for TCR down-regulation. Point mutation analyses confirmed this observation and demonstrated that a membrane-proximal di-leucine motif (L131......, indicating that the TCR was down-regulated by endocytosis via clathrin coated pits. Based on the present results and previously published observations on intracellular receptor sorting, a general model for intracellular sorting of receptors containing di-leucine- or tyrosine-based motifs is proposed....

  13. Central ions and lateral asparagine/glutamine zippers stabilize the post-fusion hairpin conformation of the SARS coronavirus spike glycoprotein

    International Nuclear Information System (INIS)

    Duquerroy, Stephane; Vigouroux, Armelle; Rottier, Peter J.M.; Rey, Felix A.; Jan Bosch, Berend

    2005-01-01

    The coronavirus spike glycoprotein is a class I membrane fusion protein with two characteristic heptad repeat regions (HR1 and HR2) in its ectodomain. Here, we report the X-ray structure of a previously characterized HR1/HR2 complex of the severe acute respiratory syndrome coronavirus spike protein. As expected, the HR1 and HR2 segments are organized in antiparallel orientations within a rod-like molecule. The HR1 helices form an exceptionally long (120 A) internal coiled coil stabilized by hydrophobic and polar interactions. A striking arrangement of conserved asparagine and glutamine residues of HR1 propagates from two central chloride ions, providing hydrogen-bonding 'zippers' that strongly constrain the path of the HR2 main chain, forcing it to adopt an extended conformation at either end of a short HR2 α-helix

  14. Quantitative analysis and prediction of curvature in leucine-rich repeat proteins.

    Science.gov (United States)

    Hindle, K Lauren; Bella, Jordi; Lovell, Simon C

    2009-11-01

    Leucine-rich repeat (LRR) proteins form a large and diverse family. They have a wide range of functions most of which involve the formation of protein-protein interactions. All known LRR structures form curved solenoids, although there is large variation in their curvature. It is this curvature that determines the shape and dimensions of the inner space available for ligand binding. Unfortunately, large-scale parameters such as the overall curvature of a protein domain are extremely difficult to predict. Here, we present a quantitative analysis of determinants of curvature of this family. Individual repeats typically range in length between 20 and 30 residues and have a variety of secondary structures on their convex side. The observed curvature of the LRR domains correlates poorly with the lengths of their individual repeats. We have, therefore, developed a scoring function based on the secondary structure of the convex side of the protein that allows prediction of the overall curvature with a high degree of accuracy. We also demonstrate the effectiveness of this method in selecting a suitable template for comparative modeling. We have developed an automated, quantitative protocol that can be used to predict accurately the curvature of leucine-rich repeat proteins of unknown structure from sequence alone. This protocol is available as an online resource at http://www.bioinf.manchester.ac.uk/curlrr/.

  15. Transcription factor Nrf1 is topologically repartitioned across membranes to enable target gene transactivation through its acidic glucose-responsive domains.

    Science.gov (United States)

    Zhang, Yiguo; Ren, Yonggang; Li, Shaojun; Hayes, John D

    2014-01-01

    The membrane-bound Nrf1 transcription factor regulates critical homeostatic and developmental genes. The conserved N-terminal homology box 1 (NHB1) sequence in Nrf1 targets the cap'n'collar (CNC) basic basic-region leucine zipper (bZIP) factor to the endoplasmic reticulum (ER), but it is unknown how its activity is controlled topologically within membranes. Herein, we report a hitherto unknown mechanism by which the transactivation activity of Nrf1 is controlled through its membrane-topology. Thus after Nrf1 is anchored within ER membranes, its acidic transactivation domains (TADs), including the Asn/Ser/Thr-rich (NST) glycodomain situated between acidic domain 1 (AD1) and AD2, are transiently translocated into the lumen of the ER, where NST is glycosylated in the presence of glucose to yield an inactive 120-kDa Nrf1 glycoprotein. Subsequently, portions of the TADs partially repartition across membranes into the cyto/nucleoplasmic compartments, whereupon an active 95-kDa form of Nrf1 accumulates, a process that is more obvious in glucose-deprived cells and may involve deglycosylation. The repartitioning of Nrf1 out of membranes is monitored within this protein by its acidic-hydrophobic amphipathic glucose-responsive domains, particularly the Neh5L subdomain within AD1. Therefore, the membrane-topological organization of Nrf1 dictates its post-translational modifications (i.e. glycosylation, the putative deglycosylation and selective proteolysis), which together control its ability to transactivate target genes.

  16. The LINKS motif zippers trans-acyltransferase polyketide synthase assembly lines into a biosynthetic megacomplex.

    Science.gov (United States)

    Gay, Darren C; Wagner, Drew T; Meinke, Jessica L; Zogzas, Charles E; Gay, Glen R; Keatinge-Clay, Adrian T

    2016-03-01

    Polyketides such as the clinically-valuable antibacterial agent mupirocin are constructed by architecturally-sophisticated assembly lines known as trans-acyltransferase polyketide synthases. Organelle-sized megacomplexes composed of several copies of trans-acyltransferase polyketide synthase assembly lines have been observed by others through transmission electron microscopy to be located at the Bacillus subtilis plasma membrane, where the synthesis and export of the antibacterial polyketide bacillaene takes place. In this work we analyze ten crystal structures of trans-acyltransferase polyketide synthases ketosynthase domains, seven of which are reported here for the first time, to characterize a motif capable of zippering assembly lines into a megacomplex. While each of the three-helix LINKS (Laterally-INteracting Ketosynthase Sequence) motifs is observed to similarly dock with a spatially-reversed copy of itself through hydrophobic and ionic interactions, the amino acid sequences of this motif are not conserved. Such a code is appropriate for mediating homotypic contacts between assembly lines to ensure the ordered self-assembly of a noncovalent, yet tightly-knit, enzymatic network. LINKS-mediated lateral interactions would also have the effect of bolstering the vertical association of the polypeptides that comprise a polyketide synthase assembly line. Copyright © 2015 Elsevier Inc. All rights reserved.

  17. Fast axonal transport of 3H-leucin-labelled proteins in the unhurt and isolated optical nerve of rats

    International Nuclear Information System (INIS)

    Wagner, H.E.

    1981-01-01

    The distribution of radioactivity of amino acid molecules incorporated in protein after injection of 3 H-Leucin into the right bulb was investigated and determined along optical nerve after 1, 2, and 4 h. A slightly increased radioactivity at the point of entrance of the optical nerves into the optical duct was found. A slightly reduced axon diameter was discussed as a possible cause. The radioactivity brought into the optical nerve via the vascular system was determined by measuring the contralateral optical nerve. In relation to the axonally transported activity, it was low. The speed of the fast axonal transport is 168 mm/d. If the processes ruling the amino acids in the perikaryon are taken into consideration, the transport speed is 240 mm/d. The application of the protein synthesis prohibitor, Cycloheximide, 5 minutes after the injection of Leucinin completely prevented the appearance of axonally transported labelled proteins. When cycloheximide was administered 2 h after Leucin, a significantly loner radioactivity than in the nerve could be determined after another 2 h; i.e. the incorporation of Leucin was not completed yet after 2 h. The profile of active compounds was the same as in the control group. In other experiments, the axonal transport of labelled proteins in isolated optical nerve fibres was tested. If the separation was carried out 2 h after the injection of Leucin an extreme reduction in activity could be determined after 1 or 2 h. The continued distribution of activity after cycloheximide treatment and removal of perikarya in comparison with the control indicate the continuation of the transport, also after separation of the axon from the perikaryon. This means that, during the time of the experiment, the mechanism of the fast axonal transport functions independently of the perikaryon. (orig./MG) [de

  18. Expression of a truncated ATHB17 protein in maize increases ear weight at silking.

    Directory of Open Access Journals (Sweden)

    Elena A Rice

    Full Text Available ATHB17 (AT2G01430 is an Arabidopsis gene encoding a member of the α-subclass of the homeodomain leucine zipper class II (HD-Zip II family of transcription factors. The ATHB17 monomer contains four domains common to all class II HD-Zip proteins: a putative repression domain adjacent to a homeodomain, leucine zipper, and carboxy terminal domain. However, it also possesses a unique N-terminus not present in other members of the family. In this study we demonstrate that the unique 73 amino acid N-terminus is involved in regulation of cellular localization of ATHB17. The ATHB17 protein is shown to function as a transcriptional repressor and an EAR-like motif is identified within the putative repression domain of ATHB17. Transformation of maize with an ATHB17 expression construct leads to the expression of ATHB17Δ113, a truncated protein lacking the first 113 amino acids which encodes a significant portion of the repression domain. Because ATHB17Δ113 lacks the repression domain, the protein cannot directly affect the transcription of its target genes. ATHB17Δ113 can homodimerize, form heterodimers with maize endogenous HD-Zip II proteins, and bind to target DNA sequences; thus, ATHB17Δ113 may interfere with HD-Zip II mediated transcriptional activity via a dominant negative mechanism. We provide evidence that maize HD-Zip II proteins function as transcriptional repressors and that ATHB17Δ113 relieves this HD-Zip II mediated transcriptional repression activity. Expression of ATHB17Δ113 in maize leads to increased ear size at silking and, therefore, may enhance sink potential. We hypothesize that this phenotype could be a result of modulation of endogenous HD-Zip II pathways in maize.

  19. Expression of a truncated ATHB17 protein in maize increases ear weight at silking.

    Science.gov (United States)

    Rice, Elena A; Khandelwal, Abha; Creelman, Robert A; Griffith, Cara; Ahrens, Jeffrey E; Taylor, J Philip; Murphy, Lesley R; Manjunath, Siva; Thompson, Rebecca L; Lingard, Matthew J; Back, Stephanie L; Larue, Huachun; Brayton, Bonnie R; Burek, Amanda J; Tiwari, Shiv; Adam, Luc; Morrell, James A; Caldo, Rico A; Huai, Qing; Kouadio, Jean-Louis K; Kuehn, Rosemarie; Sant, Anagha M; Wingbermuehle, William J; Sala, Rodrigo; Foster, Matt; Kinser, Josh D; Mohanty, Radha; Jiang, Dongming; Ziegler, Todd E; Huang, Mingya G; Kuriakose, Saritha V; Skottke, Kyle; Repetti, Peter P; Reuber, T Lynne; Ruff, Thomas G; Petracek, Marie E; Loida, Paul J

    2014-01-01

    ATHB17 (AT2G01430) is an Arabidopsis gene encoding a member of the α-subclass of the homeodomain leucine zipper class II (HD-Zip II) family of transcription factors. The ATHB17 monomer contains four domains common to all class II HD-Zip proteins: a putative repression domain adjacent to a homeodomain, leucine zipper, and carboxy terminal domain. However, it also possesses a unique N-terminus not present in other members of the family. In this study we demonstrate that the unique 73 amino acid N-terminus is involved in regulation of cellular localization of ATHB17. The ATHB17 protein is shown to function as a transcriptional repressor and an EAR-like motif is identified within the putative repression domain of ATHB17. Transformation of maize with an ATHB17 expression construct leads to the expression of ATHB17Δ113, a truncated protein lacking the first 113 amino acids which encodes a significant portion of the repression domain. Because ATHB17Δ113 lacks the repression domain, the protein cannot directly affect the transcription of its target genes. ATHB17Δ113 can homodimerize, form heterodimers with maize endogenous HD-Zip II proteins, and bind to target DNA sequences; thus, ATHB17Δ113 may interfere with HD-Zip II mediated transcriptional activity via a dominant negative mechanism. We provide evidence that maize HD-Zip II proteins function as transcriptional repressors and that ATHB17Δ113 relieves this HD-Zip II mediated transcriptional repression activity. Expression of ATHB17Δ113 in maize leads to increased ear size at silking and, therefore, may enhance sink potential. We hypothesize that this phenotype could be a result of modulation of endogenous HD-Zip II pathways in maize.

  20. Reversible pH-controlled DNA-binding peptide nanotweezers: An in-silico study

    Directory of Open Access Journals (Sweden)

    Gaurav Sharma

    2008-11-01

    Full Text Available Gaurav Sharma1, Kaushal Rege2,3, David E Budil4, Martin L Yarmush2,5, Constantinos Mavroidis11Department of Mechanical and Industrial Engineering; 4Department of Chemistry and Chemical Biology, Northeastern University, Boston, MA, USA; 2The Center for Engineering in Medicine (CEM, Massachusetts General Hospital and Harvard Medical School, Boston, MA, USA; 3Department of Chemical Engineering, Arizona State University, Tempe, AZ, USA; 5Department of Biomedical Engineering, Rutgers University, NJ, USAAbstract: We describe the molecular dynamics (MD-aided engineering design of mutant peptides based on the α-helical coiled-coil GCN4 leucine zipper peptide (GCN4-p1 in order to obtain environmentally-responsive nanotweezers. The actuation mechanism of the nanotweezers depends on the modification of electrostatic charges on the residues along the length of the coiled coil. Modulating the solution pH between neutral and acidic values results in the reversible movement of helices toward and away from each other and creates a complete closed-open-closed transition cycle between the helices. Our results indicate that the mutants show a reversible opening of up to 15 Å (1.5 nm; approximately 150% of the initial separation upon pH actuation. Investigation on the physicochemical phenomena that influence conformational properties, structural stability, and reversibility of the coiled-coil peptide-based nanotweezers revealed that a rationale- and design-based approach is needed to engineer stable peptide or macromolecules into stimuli-responsive devices. The efficacy of the mutant that demonstrated the most significant reversible actuation for environmentally responsive modulation of DNA-binding activity was also demonstrated. Our results have significant implications in bioseparations and in the engineering of novel transcription factors.Keywords: bionanotechnology, nanotweezers, coiled-coil, GCN4, leucine zipper, molecular dynamics, environmentally

  1. Expression of a Truncated ATHB17 Protein in Maize Increases Ear Weight at Silking

    Science.gov (United States)

    Creelman, Robert A.; Griffith, Cara; Ahrens, Jeffrey E.; Taylor, J. Philip; Murphy, Lesley R.; Manjunath, Siva; Thompson, Rebecca L.; Lingard, Matthew J.; Back, Stephanie L.; Larue, Huachun; Brayton, Bonnie R.; Burek, Amanda J.; Tiwari, Shiv; Adam, Luc; Morrell, James A.; Caldo, Rico A.; Huai, Qing; Kouadio, Jean-Louis K.; Kuehn, Rosemarie; Sant, Anagha M.; Wingbermuehle, William J.; Sala, Rodrigo; Foster, Matt; Kinser, Josh D.; Mohanty, Radha; Jiang, Dongming; Ziegler, Todd E.; Huang, Mingya G.; Kuriakose, Saritha V.; Skottke, Kyle; Repetti, Peter P.; Reuber, T. Lynne; Ruff, Thomas G.; Petracek, Marie E.; Loida, Paul J.

    2014-01-01

    ATHB17 (AT2G01430) is an Arabidopsis gene encoding a member of the α-subclass of the homeodomain leucine zipper class II (HD-Zip II) family of transcription factors. The ATHB17 monomer contains four domains common to all class II HD-Zip proteins: a putative repression domain adjacent to a homeodomain, leucine zipper, and carboxy terminal domain. However, it also possesses a unique N-terminus not present in other members of the family. In this study we demonstrate that the unique 73 amino acid N-terminus is involved in regulation of cellular localization of ATHB17. The ATHB17 protein is shown to function as a transcriptional repressor and an EAR-like motif is identified within the putative repression domain of ATHB17. Transformation of maize with an ATHB17 expression construct leads to the expression of ATHB17Δ113, a truncated protein lacking the first 113 amino acids which encodes a significant portion of the repression domain. Because ATHB17Δ113 lacks the repression domain, the protein cannot directly affect the transcription of its target genes. ATHB17Δ113 can homodimerize, form heterodimers with maize endogenous HD-Zip II proteins, and bind to target DNA sequences; thus, ATHB17Δ113 may interfere with HD-Zip II mediated transcriptional activity via a dominant negative mechanism. We provide evidence that maize HD-Zip II proteins function as transcriptional repressors and that ATHB17Δ113 relieves this HD-Zip II mediated transcriptional repression activity. Expression of ATHB17Δ113 in maize leads to increased ear size at silking and, therefore, may enhance sink potential. We hypothesize that this phenotype could be a result of modulation of endogenous HD-Zip II pathways in maize. PMID:24736658

  2. Functional characterization of a competitive peptide antagonist of p65 in human macrophage-like cells suggests therapeutic potential for chronic inflammation

    Directory of Open Access Journals (Sweden)

    Srinivasan M

    2014-12-01

    Full Text Available Mythily Srinivasan,1 Corinne Blackburn,1 Debomoy K Lahiri2,3 1Department of Oral Pathology, Medicine and Radiology, Indiana University School of Dentistry, 2Institute of Psychiatry Research, Department of Psychiatry, 3Department of Medical and Molecular Genetics, School of Medicine, Indiana University-Purdue University, Indianapolis, IN, USA Abstract: Glucocorticoid-induced leucine zipper (GILZ is a glucocorticoid responsive protein that links the nuclear factor-kappa B (NFκB and the glucocorticoid signaling pathways. Functional and binding studies suggest that the proline-rich region at the carboxy terminus of GILZ binds the p65 subunit of NFκB and suppresses the immunoinflammatory response. A widely-used strategy in the discovery of peptide drugs involves exploitation of the complementary surfaces of naturally occurring binding partners. Previously, we observed that a synthetic peptide (GILZ-P derived from the proline-rich region of GILZ bound activated p65 and ameliorated experimental encephalomyelitis. Here we characterize the secondary structure of GILZ-P by circular dichroic analysis. GILZ-P adopts an extended polyproline type II helical conformation consistent with the structural conformation commonly observed in interfaces of transient intermolecular interactions. To determine the potential application of GILZ-P in humans, we evaluated the toxicity and efficacy of the peptide drug in mature human macrophage-like THP-1 cells. Treatment with GILZ-P at a wide range of concentrations commonly used for peptide drugs was nontoxic as determined by cell viability and apoptosis assays. Functionally, GILZ-P suppressed proliferation and glutamate secretion by activated macrophages by inhibiting nuclear translocation of p65. Collectively, our data suggest that the GILZ-P has therapeutic potential in chronic CNS diseases where persistent inflammation leads to neurodegeneration such as multiple sclerosis and Alzheimer’s disease. Keywords

  3. Silencing of the glucocorticoid-induced leucine zipper improves the immunogenicity of clinical-grade dendritic cells

    DEFF Research Database (Denmark)

    Cathelin, Dominique; Met, Özcan; Svane, Inge Marie

    2013-01-01

    The maturation cocktail composed of interleukin (IL)-6, IL-1β, tumor necrosis factor-α and prostaglandin E2 is considered the "gold standard" for inducing the maturation of dendritic cells (DCs) for use in cancer immunotherapy. Nevertheless, although this maturation cocktail induces increased...

  4. Effect of partially purified components of zoospores and mycelia of phytophthora infestans on uptake of 3H-leucine by potato tuber disks

    International Nuclear Information System (INIS)

    Nishimura, Norio; Tomiyama, Kohei; Doke, Noriyuki

    1980-01-01

    The zoosporial component of Phytophthora infestans, which was previously reported to cause reduction of 3 H-leucine uptake by potato tuber disks, was partially purified. Precipitate (A-fraction) was obtained by homogenizing zoospores with acetate buffer at pH 4.5 and centrifuging at 20,000 x g, and the A-fraction was suspended in borate buffer at pH 8.8, boiled for 1 hr and then centrifuged at 20,000 x g, giving the precipitate (B-fraction) and supernatant (C-fraction). Ten ml of 10 mM tris-HCl buffer containing 1 mM CaCl 2 at pH 7.4 was used to suspend A and B-fraction. The buffer was used as a control. A, B and C fractions obtained from 5 - 6 x 10 6 zoosprores reducted uptake of 3 H-leucine by the tuber disks of potato cv. Rishiri, but the inhibition rates caused by these fractions differed markedly. However, very high correlation was found between inhibition rates of 3 H-leucine uptake and sugar contents of these fractions. There was no difference in the inhibition rates between the zoosporial components of incompatible and compatible races, when the activities were expressed in terms of the sugar contents. The mycelial components of P. infestans extracted by the modified method of Lisker and Kuc which was used to extract phytoalexin elicitor from that of P. infestans, also had the same effect as the zoosporial components (A, B, and C-fraction) on 3 H-leucine uptake by the disks. C-fraction containing 15 μg of sugar per ml sufficed to inhibit 3 H-leucine uptake at the maximum rate, and the maximum rate of inhibition was attained within 2 hr after the zoosporial component (C-fraction containing 30 μg sugar/ml) was administered to the disks. (author)

  5. Leucine and protein metabolism in obese Zucker rats.

    Directory of Open Access Journals (Sweden)

    Pengxiang She

    Full Text Available 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 protein

  6. MHJ_0461 is a multifunctional leucine aminopeptidase on the surface of Mycoplasma hyopneumoniae.

    Science.gov (United States)

    Jarocki, Veronica M; Santos, Jerran; Tacchi, Jessica L; Raymond, Benjamin B A; Deutscher, Ania T; Jenkins, Cheryl; Padula, Matthew P; Djordjevic, Steven P

    2015-01-01

    Aminopeptidases are part of the arsenal of virulence factors produced by bacterial pathogens that inactivate host immune peptides. Mycoplasma hyopneumoniae is a genome-reduced pathogen of swine that lacks the genetic repertoire to synthesize amino acids and relies on the host for availability of amino acids for growth. M. hyopneumoniae recruits plasmin(ogen) onto its cell surface via the P97 and P102 adhesins and the glutamyl aminopeptidase MHJ_0125. Plasmin plays an important role in regulating the inflammatory response in the lungs of pigs infected with M. hyopneumoniae. We show that recombinant MHJ_0461 (rMHJ_0461) functions as a leucine aminopeptidase (LAP) with broad substrate specificity for leucine, alanine, phenylalanine, methionine and arginine and that MHJ_0461 resides on the surface of M. hyopneumoniae. rMHJ_0461 also binds heparin, plasminogen and foreign DNA. Plasminogen bound to rMHJ_0461 was readily converted to plasmin in the presence of tPA. Computational modelling identified putative DNA and heparin-binding motifs on solvent-exposed sites around a large pore on the LAP hexamer. We conclude that MHJ_0461 is a LAP that moonlights as a multifunctional adhesin on the cell surface of M. hyopneumoniae.

  7. Accumulation of D- vs. L-isomers of alanine and leucine in rat prostatic adenocarcinoma

    International Nuclear Information System (INIS)

    Conti, P.S.; Schmall, B.; Bigler, R.E.; Zanzonico, P.B.; Kleinert, E.; Whitmore, W.F. Jr.

    1985-01-01

    It has been reported that tumor tissue may accumulate some D-amino acids preferentially over the L-isomers. In order to investigate the potential use of carbon-11 labeled amino acid isomers for in vivo tumor studies with positron emission tomography in patients, the tissue distributions of alanine and leucine, substrates for the A-type and L-type amino acid transport systems, respectively, were studied in Copenhagen rates bearing the Dunning R3327G prostatic adenocarcinoma. The authors have previously reported differences in the accumulation of A-type vs. L-type amino acids in rat prostatic adenocarcinoma and normal tissues. All compounds were labeled with C-14 in the carboxyl position with specific activities of 30.0-56.6 mCi/mmol. Higher levels of C-14 activity (Relative Concentration (RC)=dpm found per gm tissue + dpm inject per gm animal mass) were observed in tumor tissue using D-alanine (0.71) compared to L- (0.21) or DL-alanine (0.27) at 45 min post-injection. While tumor/prostate and tumor/liver ratios were above 2 for all three substrates, tumor/blood and tumor/muscle were above one for only the D-isomer. Comparisons made with D-, L-, and DL-leucine also demonstrated a higher level of RC in tumor tissue with the D-isomer (0.84) vs. the L-(0.66) and DL-leucine (0.63). In this case, however, tumor/blood, tumor/prostate, and tumor/muscle ratios were above one for all three substrates, while tumor/liver ratios were below one. These results support the observation of a preferential accumulation of D-amino acids in tumor tissue over the natural L-isomers. Observed differences in the accumulation of the isomers in normal tissues are discussed

  8. Participation of ATM in cellular response to DNA damage induced by ionizing radiation

    International Nuclear Information System (INIS)

    Meng Xiangbing; Song Yi; Mao Jianping; Gong Bo; Dong Yan; Liu Bin; Sun Zhixian

    2000-01-01

    Objective: To clone ATM full length cDNA and cDNA fragments containing some functional domains and to identify proteins that interact with ATM and mediate DNA damage signal transduction in cellular response to DNA damage. Methods: ATM cDNA was amplified from MarthomTM-Ready cDNA kit of human leukocytes by LD-PCR. ATM-interacting proteins were screened by yeast two hybrid system. Results: ATM full-length cDNA and cDNA fragments containing PI3K kinase domain, leucine zipper and proline rich region were amplified from human cDNAs. Several candidate clones that interacted with ATM PI3K domain were identified. Conclusion: ATM mediates DNA damage signal transduction by interacting with many proteins

  9. Masking of the CD3 gamma di-leucine-based motif by zeta is required for efficient T-cell receptor expression

    DEFF Research Database (Denmark)

    Lauritsen, Jens Peter H; Bonefeld, Charlotte Menné; von Essen, Marina

    2004-01-01

    containing the di-leucine-based endocytosis motif of the TCR subunit CD3 gamma have indicated that the zeta chain can mask this motif. In this study, we show that successive truncations of the cytoplasmic tail of zeta led to reduced surface expression levels of completely assembled TCR complexes. The reduced...... TCR expression levels were caused by an increase in the TCR endocytic rate constant in combination with an unaffected exocytic rate constant. Furthermore, the TCR degradation rate constant was increased in cells with truncated zeta. Introduction of a CD3 gamma chain with a disrupted di-leucine...

  10. Transcription Factor Functional Protein-Protein Interactions in Plant Defense Responses

    Directory of Open Access Journals (Sweden)

    Murilo S. Alves

    2014-03-01

    Full Text Available Responses to biotic stress in plants lead to dramatic reprogramming of gene expression, favoring stress responses at the expense of normal cellular functions. Transcription factors are master regulators of gene expression at the transcriptional level, and controlling the activity of these factors alters the transcriptome of the plant, leading to metabolic and phenotypic changes in response to stress. The functional analysis of interactions between transcription factors and other proteins is very important for elucidating the role of these transcriptional regulators in different signaling cascades. In this review, we present an overview of protein-protein interactions for the six major families of transcription factors involved in plant defense: basic leucine zipper containing domain proteins (bZIP, amino-acid sequence WRKYGQK (WRKY, myelocytomatosis related proteins (MYC, myeloblastosis related proteins (MYB, APETALA2/ ETHYLENE-RESPONSIVE ELEMENT BINDING FACTORS (AP2/EREBP and no apical meristem (NAM, Arabidopsis transcription activation factor (ATAF, and cup-shaped cotyledon (CUC (NAC. We describe the interaction partners of these transcription factors as molecular responses during pathogen attack and the key components of signal transduction pathways that take place during plant defense responses. These interactions determine the activation or repression of response pathways and are crucial to understanding the regulatory networks that modulate plant defense responses.

  11. Batf3-dependent CD8α+ Dendritic Cells Aggravates Atherosclerosis via Th1 Cell Induction and Enhanced CCL5 Expression in Plaque Macrophages.

    Science.gov (United States)

    Li, Yalin; Liu, Xueyan; Duan, Wei; Tian, Hua; Zhu, Guangming; He, Hao; Yao, Shutong; Yi, Shuying; Song, Wengang; Tang, Hua

    2017-04-01

    Dendritic cells (DCs) play an important role in controlling T cell-mediated adaptive immunity in atherogenesis. However, the role of the basic leucine zipper transcription factor, ATF-like 3 (Batf3)-dependent CD8α + DC subset in atherogenesis remains unclear. Here we show that Batf3 -/- Apoe -/- mice, lacking CD8α + DCs, exhibited a significant reduction in atherogenesis and T help 1 (Th1) cells compared with Apoe -/- controls. Then, we found that CD8α + DCs preferentially induce Th1 cells via secreting interleukin-12 (IL-12), and that the expression of interferon-gamma (IFN-γ)or chemokine (C-C motif) ligand 5 (CCL5) in aorta were significantly decreased in Batf3 -/- Apoe -/- mice. We further demonstrated that macrophages were the major CCL5-expressing cells in the plaque, which was significantly reduced in Batf3 -/- Apoe -/- mice. Furthermore, we found CCL5 expression in macrophages was promoted by IFN-γ. Finally, we showed that Batf3 -/- Apoe -/- mice displayed decreased infiltration of leukocytes in the plaque. Thus, CD8α + DCs aggravated atherosclerosis, likely by inducing Th1 cell response, which promoted CCL5 expression in macrophages and increased infiltration of leukocytes and lesion inflammation. Copyright © 2017 The Authors. Published by Elsevier B.V. All rights reserved.

  12. Quantitative phosphoproteomic analysis of early seed development in rice (Oryza sativa L.).

    Science.gov (United States)

    Qiu, Jiehua; Hou, Yuxuan; Tong, Xiaohong; Wang, Yifeng; Lin, Haiyan; Liu, Qing; Zhang, Wen; Li, Zhiyong; Nallamilli, Babi R; Zhang, Jian

    2016-02-01

    Rice (Oryza sativa L.) seed serves as a major food source for over half of the global population. Though it has been long recognized that phosphorylation plays an essential role in rice seed development, the phosphorylation events and dynamics in this process remain largely unknown so far. Here, we report the first large scale identification of rice seed phosphoproteins and phosphosites by using a quantitative phosphoproteomic approach. Thorough proteomic studies in pistils and seeds at 3, 7 days after pollination resulted in the successful identification of 3885, 4313 and 4135 phosphopeptides respectively. A total of 2487 proteins were differentially phosphorylated among the three stages, including Kip related protein 1, Rice basic leucine zipper factor 1, Rice prolamin box binding factor and numerous other master regulators of rice seed development. Moreover, differentially phosphorylated proteins may be extensively involved in the biosynthesis and signaling pathways of phytohormones such as auxin, gibberellin, abscisic acid and brassinosteroid. Our results strongly indicated that protein phosphorylation is a key mechanism regulating cell proliferation and enlargement, phytohormone biosynthesis and signaling, grain filling and grain quality during rice seed development. Overall, the current study enhanced our understanding of the rice phosphoproteome and shed novel insight into the regulatory mechanism of rice seed development.

  13. A novel bZIP gene from Tamarix hispida mediates physiological responses to salt stress in tobacco plants.

    Science.gov (United States)

    Wang, Yucheng; Gao, Caiqiu; Liang, Yenan; Wang, Chao; Yang, Chuanping; Liu, Guifeng

    2010-02-15

    Basic leucine zipper proteins (bZIPs) are transcription factors that bind abscisic acid (ABA)-responsive elements (ABREs) and enable plants to withstand adverse environmental conditions. In the present study, a novel bZIP gene, ThbZIP1 was cloned from Tamarix hispida. Expression studies in T. hispida showed differential regulation of ThbZIP1 in response to treatment with NaCl, polyethylene glycol (PEG) 6000, NaHCO(3), and CdCl(2), suggesting that ThbZIP1 is involved in abiotic stress responses. To identify the physiological responses mediated by ThbZIP1, transgenic tobacco plants overexpressing exogenous ThbZIP1 were generated. Various physiological parameters related to salt stress were measured and compared between transgenic and wild type (WT) plants. Our results indicate that overexpression of ThbZIP1 can enhance the activity of both peroxidase (POD) and superoxide dismutase (SOD), and increase the content of soluble sugars and soluble proteins under salt stress conditions. These results suggest that ThbZIP1 contributes to salt tolerance by mediating signaling through multiple physiological pathways. Furthermore, ThbZIP1 confers stress tolerance to plants by enhancing reactive oxygen species (ROS) scavenging, facilitating the accumulation of compatible osmolytes, and inducing and/or enhancing the biosynthesis of soluble proteins. Copyright 2009 Elsevier GmbH. All rights reserved.

  14. Gladiolus hybridus ABSCISIC ACID INSENSITIVE 5 (GhABI5) is an important transcription factor in ABA signaling that can enhance Gladiolus corm dormancy and Arabidopsis seed dormancy.

    Science.gov (United States)

    Wu, Jian; Seng, Shanshan; Sui, Juanjuan; Vonapartis, Eliana; Luo, Xian; Gong, Benhe; Liu, Chen; Wu, Chenyu; Liu, Chao; Zhang, Fengqin; He, Junna; Yi, Mingfang

    2015-01-01

    The phytohormone abscisic acid (ABA) regulates plant development and is crucial for abiotic stress response. In this study, cold storage contributes to reducing endogenous ABA content, resulting in dormancy breaking of Gladiolus. The ABA inhibitor fluridone also promotes germination, suggesting that ABA is an important hormone that regulates corm dormancy. Here, we report the identification and functional characterization of the Gladiolus ABI5 homolog (GhABI5), which is a basic leucine zipper motif transcriptional factor (TF). GhABI5 is expressed in dormant vegetative organs (corm, cormel, and stolon) as well as in reproductive organs (stamen), and it is up-regulated by ABA or drought. Complementation analysis reveals that GhABI5 rescues the ABA insensitivity of abi5-3 during seed germination and induces the expression of downstream ABA response genes in Arabidopsis thaliana (EM1, EM6, and RD29B). Down-regulation of GhABI5 in dormant cormels via virus induced gene silence promotes sprouting and reduces the expression of downstream genes (GhLEA and GhRD29B). The results of this study reveal that GhABI5 regulates bud dormancy (vegetative organ) in Gladiolus in addition to its well-studied function in Arabidopsis seeds (reproductive organ).

  15. Gladiolus hybridus ABSCISIC ACID INSENSITIVE 5 (GhABI5 is an important transcription factor in ABA signaling that can enhance Gladiolus corm dormancy and Arabidopsis seed dormancy.

    Directory of Open Access Journals (Sweden)

    Jian eWu

    2015-11-01

    Full Text Available The phytohormone abscisic acid (ABA regulates plant development and is crucial for abiotic stress response. In this study, cold storage contributes to reducing endogenous ABA content, resulting in dormancy breaking of Gladiolus. The ABA inhibitor fluridone also promotes germination, suggesting that ABA is an important hormone that regulates corm dormancy. Here, we report the identification and functional characterization of the Gladiolus ABI5 homolog (GhABI5, which is a basic leucine zipper motif transcriptional factor (TF. GhABI5 is expressed in dormant vegetative organs (corm, cormel and stolon as well as in reproductive organs (stamen, and it is up-regulated by ABA or drought. Complementation analysis reveals that GhABI5 rescues the ABA insensitivity of abi5-3 during seed germination and induces the expression of downstream ABA response genes in Arabidopsis thaliana (EM1, EM6 and RD29B. Down-regulation of GhABI5 in dormant cormels via Virus Induced Gene Silence (VIGS promotes sprouting and reduces the expression of downstream genes (GhLEA and GhRD29B. The results of this study reveal that GhABI5 regulates bud dormancy (vegetative organ in Gladiolus in addition to its well-studied function in Arabidopsis seeds (reproductive organ.

  16. Determination of proteolytic activity using L-[4,5-3H]leucine-labelled globin as a substrate

    International Nuclear Information System (INIS)

    Maliopoulou, T.B.; Dionyssiou-Asteriou, A.; Loucopoulos, D.

    1980-01-01

    A simple and sensitive method for the assay of proteolytic enzyme activity is described. This is based on the digestion of L-[4,5- 3 H]leucine globin by proteolytic enzymes and radioactivity measurement of the trichloroacetic acid soluble cleavage products. (Auth.)

  17. Biosynthesis of the leucine derived α-, β- and γ-hydroxynitrile glucosides in barley (Hordeum vulgare L.)

    DEFF Research Database (Denmark)

    Knoch, Eva; Motawie, Mohammed Saddik; Olsen, Carl Erik

    2016-01-01

    Barley (Hordeum vulgare L.) produces five leucine-derived hydroxynitrile glucosides (HNGs), of which only epiheterodendrin is a cyanogenic glucoside. The four non-cyanogenic HNGs are the β-HNG epidermin and the γ-HNGs osmaronin, dihydroosmaronin and sutherlandin. By analyzing 247 spring barley...

  18. Characteristic expression of HTLV-1 basic zipper factor (HBZ transcripts in HTLV-1 provirus-positive cells

    Directory of Open Access Journals (Sweden)

    Yamada Yasuaki

    2008-04-01

    Full Text Available Abstract Background HTLV-1 causes adult T-cell leukemia (ATL. Although there have been many studies on the oncogenesis of the viral protein Tax, the precise oncogenic mechanism remains to be elucidated. Recently, a new viral factor, HTLV-1 basic Zip factor (HBZ, encoded from the minus strand mRNA was discovered and the current models of Tax-centered ATL cell pathogenesis are in conflict with this discovery. HBZs consisting of non-spliced and spliced isoforms (HBZ-SI are thought to be implicated in viral replication and T-cell proliferation but there is little evidence on the HBZ expression profile on a large scale. Results To investigate the role of HBZ-SI in HTLV-1 provirus-positive cells, the HBZ-SI and Tax mRNA loads in samples with a mixture of infected and non-infected cells were measured and then adjusted by dividing by the HTLV-I proviral load. We show here that the HBZ-SI mRNA level is 4-fold higher than non-spliced HBZ and is expressed by almost all cells harboring HTLV-1 provirus with variable intensity. The proviral-adjusted HBZ-SI and Tax quantification revealed a characteristic imbalanced expression feature of high HBZ and low Tax expression levels in primary ATL cells or high HBZ and very high Tax levels in HTLV-1-related cell lines (cell lines compared with a standard expression profile of low HBZ and low Tax in infected cells. Interestingly, according to the mutual Tax and HBZ expression status, HTLV-1-related cell lines were subcategorized into two groups, an ATL cell type with high HBZ and low Tax levels and another type with high Tax and either high or low HBZ, which was closely related to its cell origin. Conclusion This is the first comprehensive study to evaluate the mutual expression profile of HBZ and Tax in provirus-positive cells, revealing that there are quantitative and relative characteristic features among infected cells, primary ATL cells, and cell lines.

  19. BACE1 protein endocytosis and trafficking are differentially regulated by ubiquitination at lysine 501 and the Di-leucine motif in the carboxyl terminus.

    Science.gov (United States)

    Kang, Eugene L; Biscaro, Barbara; Piazza, Fabrizio; Tesco, Giuseppina

    2012-12-14

    β-Site amyloid precursor protein-cleaving enzyme (BACE1) is a membrane-tethered member of the aspartyl proteases that has been identified as β-secretase. BACE1 is targeted through the secretory pathway to the plasma membrane and then is internalized to endosomes. Sorting of membrane proteins to the endosomes and lysosomes is regulated by the interaction of signals present in their carboxyl-terminal fragment with specific trafficking molecules. The BACE1 carboxyl-terminal fragment contains a di-leucine sorting signal ((495)DDISLL(500)) and a ubiquitination site at Lys-501. Here, we report that lack of ubiquitination at Lys-501 (BACE1K501R) does not affect the rate of endocytosis but produces BACE1 stabilization and accumulation of BACE1 in early and late endosomes/lysosomes as well as at the cell membrane. In contrast, the disruption of the di-leucine motif (BACE1LLAA) greatly impairs BACE1 endocytosis and produces a delayed retrograde transport of BACE1 to the trans-Golgi network (TGN) and a delayed delivery of BACE1 to the lysosomes, thus decreasing its degradation. Moreover, the combination of the lack of ubiquitination at Lys-501 and the disruption of the di-leucine motif (BACE1LLAA/KR) produces additive effects on BACE1 stabilization and defective internalization. Finally, BACE1LLAA/KR accumulates in the TGN, while its levels are decreased in EEA1-positive compartments indicating that both ubiquitination at Lys-501 and the di-leucine motif are necessary for the trafficking of BACE1 from the TGN to early endosomes. Our studies have elucidated a differential role for the di-leucine motif and ubiquitination at Lys-501 in BACE1 endocytosis, trafficking, and degradation and suggest the involvement of multiple adaptor molecules.

  20. BACE1 Protein Endocytosis and Trafficking Are Differentially Regulated by Ubiquitination at Lysine 501 and the Di-leucine Motif in the Carboxyl Terminus*

    Science.gov (United States)

    Kang, Eugene L.; Biscaro, Barbara; Piazza, Fabrizio; Tesco, Giuseppina

    2012-01-01

    β-Site amyloid precursor protein-cleaving enzyme (BACE1) is a membrane-tethered member of the aspartyl proteases that has been identified as β-secretase. BACE1 is targeted through the secretory pathway to the plasma membrane and then is internalized to endosomes. Sorting of membrane proteins to the endosomes and lysosomes is regulated by the interaction of signals present in their carboxyl-terminal fragment with specific trafficking molecules. The BACE1 carboxyl-terminal fragment contains a di-leucine sorting signal (495DDISLL500) and a ubiquitination site at Lys-501. Here, we report that lack of ubiquitination at Lys-501 (BACE1K501R) does not affect the rate of endocytosis but produces BACE1 stabilization and accumulation of BACE1 in early and late endosomes/lysosomes as well as at the cell membrane. In contrast, the disruption of the di-leucine motif (BACE1LLAA) greatly impairs BACE1 endocytosis and produces a delayed retrograde transport of BACE1 to the trans-Golgi network (TGN) and a delayed delivery of BACE1 to the lysosomes, thus decreasing its degradation. Moreover, the combination of the lack of ubiquitination at Lys-501 and the disruption of the di-leucine motif (BACE1LLAA/KR) produces additive effects on BACE1 stabilization and defective internalization. Finally, BACE1LLAA/KR accumulates in the TGN, while its levels are decreased in EEA1-positive compartments indicating that both ubiquitination at Lys-501 and the di-leucine motif are necessary for the trafficking of BACE1 from the TGN to early endosomes. Our studies have elucidated a differential role for the di-leucine motif and ubiquitination at Lys-501 in BACE1 endocytosis, trafficking, and degradation and suggest the involvement of multiple adaptor molecules. PMID:23109336

  1. The role of leucine in isoprenoid metabolism. Incorporation of [3-13C]leucine and of [2-3H,4-14C]-β,β-dimethyl-acrylic acid into phytosterols by tissue cultures of Andrographis paniculata

    International Nuclear Information System (INIS)

    Anastasis, P.; Freer, I.; Overton, K.; Rycroft, D.; Singh, S.B.

    1985-01-01

    [3- 13 C]Leucine is incorporated into phytosterols by tissue cultures of Andrographis paniculata by breakdown to acetyl-CoA and its subsequent incorporation via (3S)-3-hydroxy-3-methylglutaryl-CoA (HMG-CoA) and mevalonic acid; [2- 3 H,4- 14 C]-β,β-dimethylacrylic acid also is not incorporated intact. (author)

  2. Postprandial oxidative losses of dietary leucine depend on the time interval between consecutive meals

    NARCIS (Netherlands)

    Myszkowska-Ryciak, J.; Keller, J.S.; Bujko, J.; Stankiewicz-Ciupa, J.; Koopmanschap, R.E.; Schreurs, V.V.A.M.

    2015-01-01

    Postprandial oxidative losses of egg white-bound [1-13C]-leucine were studied as 13C recovery in the breath of rats in relation to different time intervals between two meals. Male Wistar rats (n = 48; 68.3 ±5.9 g) divided into 4 groups (n = 12) were fed two meals a day (9:00

  3. T cell receptor zeta allows stable expression of receptors containing the CD3gamma leucine-based receptor-sorting motif

    DEFF Research Database (Denmark)

    Dietrich, J; Geisler, C

    1998-01-01

    The leucine-based motif in the T cell receptor (TCR) subunit CD3gamma constitutes a strong internalization signal. In fully assembled TCR this motif is inactive unless phosphorylated. In contrast, the motif is constitutively active in CD4/CD3gamma and Tac/CD3gamma chimeras independently of phosph......The leucine-based motif in the T cell receptor (TCR) subunit CD3gamma constitutes a strong internalization signal. In fully assembled TCR this motif is inactive unless phosphorylated. In contrast, the motif is constitutively active in CD4/CD3gamma and Tac/CD3gamma chimeras independently...... of phosphorylation and leads to rapid internalization and sorting of these chimeras to lysosomal degradation. Because the TCRzeta chain rescues incomplete TCR complexes from lysosomal degradation and allows stable surface expression of fully assembled TCR, we addressed the question whether TCRzeta has the potential...... to mask the CD3gamma leucine-based motif. By studying CD4/CD3gamma and CD16/CD3gamma chimeras, we found that CD16/CD3gamma chimeras associated with TCRzeta. The CD16/CD3gamma-TCRzeta complexes were stably expressed at the cell surface and had a low spontaneous internalization rate, indicating...

  4. The vibrational spectrum of the hydrated alanine-leucine peptide in the amide region from IR experiments and first principles calculations

    Science.gov (United States)

    Hassan, Irtaza; Donati, Luca; Stensitzki, Till; Keller, Bettina G.; Heyne, Karsten; Imhof, Petra

    2018-04-01

    We have combined infrared (IR) experiments with molecular dynamics (MD) simulations in solution at finite temperature to analyse the vibrational signature of the small floppy peptide Alanine-Leucine. IR spectra computed from first-principles MD simulations exhibit no distinct differences between conformational clusters of α -helix or β -sheet-like folds with different orientations of the bulky leucine side chain. All computed spectra show two prominent bands, in good agreement with the experiment, that are assigned to the stretch vibrations of the carbonyl and carboxyl group, respectively. Variations in band widths and exact maxima are likely due to small fluctuations in the backbone torsion angles.

  5. Effect of glucagon on cyclic AMP, albumin metabolism and incorporation of 14C-leucine into proteins in isolated parenchymal rat liver cells

    DEFF Research Database (Denmark)

    Dich, J; Gluud, C N

    1976-01-01

    wet wt. This is about the rate found in the perfused liver, Glucagon (10(-8-10(-6) M) inhibited albumin secretion and the incorporation of 14C-leucine into albumin, into total proteins in the medium and into total proteins in the cell suspension. The effect of glucagon on albumin secretion...... is compatible with an effect on the rate of synthesis. A positive correlation existed between the maximal level of cyclic AMP after glucagon administration and the inhibition of both albumin secretion and the incorporation of 149leucine....

  6. Small leucine-rich repeat proteoglycans associated with mature insoluble elastin serve as binding sites for galectins.

    Science.gov (United States)

    Itoh, Aiko; Nonaka, Yasuhiro; Ogawa, Takashi; Nakamura, Takanori; Nishi, Nozomu

    2017-11-01

    We previously reported that galectin-9 (Gal-9), an immunomodulatory animal lectin, could bind to insoluble collagen preparations and exerted direct cytocidal effects on immune cells. In the present study, we found that mature insoluble elastin is capable of binding Gal-9 and other members of the human galectin family. Lectin blot analysis of a series of commercial water-soluble elastin preparations, PES-(A) ~ PES-(E), revealed that only PES-(E) contained substances recognized by Gal-9. Gal-9-interacting substances in PES-(E) were affinity-purified, digested with trypsin and then analyzed by reversed-phase HPLC. Peptide fragments derived from five members of the small leucine-rich repeat proteoglycan family, versican, lumican, osteoglycin/mimecan, prolargin, and fibromodulin, were identified by N-terminal amino acid sequence analysis. The results indicate that Gal-9 and possibly other galectins recognize glycans attached to small leucine-rich repeat proteoglycans associated with insoluble elastin and also indicate the possibility that mature insoluble elastin serves as an extracellular reservoir for galectins.

  7. Immobilazation of aerobic microorganisms on glassy sintered material, illustrated by the example of the production of L leucine using Corynebacterium glutamicum. Immobilisierung von aeroben Mikroorganismen an Glassintermaterial am Beispiel der L-Leucin-Produktion mit Corynebacterium glutamicum

    Energy Technology Data Exchange (ETDEWEB)

    Buechs, J.

    1988-12-01

    The aim of this study was to develop the carrier fixation of aerobic microorganisms on open-pore sintered glass material. The fermentative production of L-leucine from {alpha} cetonic isocaproic acid with Corynebacterium glutamicum was chosen as an example of a microbial process with a high demand of oxygen. (orig.).

  8. Polymorphisms in leucine-rich repeat genes are associated with autism spectrum disorder susceptibility in populations of European ancestry

    NARCIS (Netherlands)

    Sousa, Ines; Clark, Taane G.; Holt, Richard; Pagnamenta, Alistair T.; Mulder, Erik J.; Minderaa, Ruud B.; Bailey, Anthony J.; Battaglia, Agatino; Klauck, Sabine M.; Poustka, Fritz; Monaco, Anthony P.

    2010-01-01

    Background: Autism spectrum disorders (ASDs) are a group of highly heritable neurodevelopmental disorders which are characteristically comprised of impairments in social interaction, communication and restricted interests/behaviours. Several cell adhesion transmembrane leucine-rich repeat (LRR)

  9. Expression of small leucine-rich proteoglycans in rat anterior pituitary gland.

    Science.gov (United States)

    Horiguchi, Kotaro; Syaidah, Rahimi; Fujiwara, Ken; Tsukada, Takehiro; Ramadhani, Dini; Jindatip, Depicha; Kikuchi, Motoshi; Yashiro, Takashi

    2013-01-01

    Proteoglycans are components of the extracellular matrix and comprise a specific core protein substituted with covalently linked glycosaminoglycan chains. Small leucine-rich proteoglycans (SLRPs) are a major family of proteoglycans and have key roles as potent effectors in cellular signaling pathways. Research during the last two decades has shown that SLRPs regulate biological functions in many tissues such as skin, tendon, kidney, liver, and heart. However, little is known of the expression of SLRPs, or the characteristics of the cells that produce them, in the anterior pituitary gland. Therefore, we have determined whether SLRPs are present in rat anterior pituitary gland. We have used real-time reverse transcription with the polymerase chain reaction to analyze the expression of SLRP genes and have identified the cells that produce SLRPs by using in situ hybridization with a digoxigenin-labeled cRNA probe. We have clearly detected the mRNA expression of SLRP genes, and cells expressing decorin, biglycan, fibromodulin, lumican, proline/arginine-rich end leucine-rich repeat protein (PRELP), and osteoglycin are located in the anterior pituitary gland. We have also investigated the possible double-staining of SLRP mRNA and pituitary hormones, S100 protein (a marker of folliculostellate cells), desmin (a marker of capillary pericytes), and isolectin B4 (a marker of endothelial cells). Decorin, biglycan, fibromodulin, lumican, PRELP, and osteoglycin mRNA have been identified in S100-protein-positive and desmin-positive cells. Thus, we conclude that folliculostellate cells and pericytes produce SLRPs in rat anterior pituitary gland.

  10. In vivo effect of sex steroids on uptake of 3H-leucine by female mouse brain

    International Nuclear Information System (INIS)

    Seiki, Kanji; Haruki, Yasuo; Imanishi, Yoshio

    1978-01-01

    In vivo effects of the sex steroids estrogen and progesterone on 3 H-leucine uptake by the brain of mature ovariectomized mice were examined. Animals were divided into three groups: group 1, consisting of control animals treated with sesame oil, group 2, animals treated with estrogen, and group 3, animals first treated with estrogen and then with progesterone. Each group was given a single i.p. injection of 3 H-leucine 2 hr after the last hormonal treatment, and sacrificed 2 hr later. Intensity of the uptake of radiolabeled leucine was measured by counting the number of reduced silver grains over cells in various brain regions using an autoradiographic technique. Group 1 showed a relatively high uptake in the supraoptic nucleus, paraventricular nucleus (PV) and ventromedial nucleus (VM) when compared with that in the remaining hypothalamic nuclei examined. Group 2 showed a significant enhancement of the uptake in all hypothalamic regions except the preoptic periventricular nucleus (PPV) when compared with that in group 1. Group 3 showed enhancement of the uptake in all hypothalamic nuclei when compared with that in group 1. However, only the PV, PPV, VM and periventricular arcuate nucleus revealed a significantly higher uptake than the respective nuclei in group 2. The remaining nuclei showed no change in uptake. Uptake by cells in the ependymal cells and cerebral cortex remained unchanged after hormonal treatment. The present results suggest that in female mice estrogen and estrogen plus progesterone stimulate protein synthesis in most of the hypothalamic nuclei and that the progesterone effect on protein synthesis is greatly influenced by estrogen-priming. (auth.)

  11. Friedelin Synthase from Maytenus ilicifolia: Leucine 482 Plays an Essential Role in the Production of the Most Rearranged Pentacyclic Triterpene

    Science.gov (United States)

    Souza-Moreira, Tatiana M.; Alves, Thaís B.; Pinheiro, Karina A.; Felippe, Lidiane G.; de Lima, Gustavo M. A.; Watanabe, Tatiana F.; Barbosa, Cristina C.; Santos, Vânia A. F. F. M.; Lopes, Norberto P.; Valentini, Sandro R.; Guido, Rafael V. C.; Furlan, Maysa; Zanelli, Cleslei F.

    2016-11-01

    Among the biologically active triterpenes, friedelin has the most-rearranged structure produced by the oxidosqualene cyclases and is the only one containing a cetonic group. In this study, we cloned and functionally characterized friedelin synthase and one cycloartenol synthase from Maytenus ilicifolia (Celastraceae). The complete coding sequences of these 2 genes were cloned from leaf mRNA, and their functions were characterized by heterologous expression in yeast. The cycloartenol synthase sequence is very similar to other known OSCs of this type (approximately 80% identity), although the M. ilicifolia friedelin synthase amino acid sequence is more related to β-amyrin synthases (65-74% identity), which is similar to the friedelin synthase cloned from Kalanchoe daigremontiana. Multiple sequence alignments demonstrated the presence of a leucine residue two positions upstream of the friedelin synthase Asp-Cys-Thr-Ala-Glu (DCTAE) active site motif, while the vast majority of OSCs identified so far have a valine or isoleucine residue at the same position. The substitution of the leucine residue with valine, threonine or isoleucine in M. ilicifolia friedelin synthase interfered with substrate recognition and lead to the production of different pentacyclic triterpenes. Hence, our data indicate a key role for the leucine residue in the structure and function of this oxidosqualene cyclase.

  12. Total protein synthesis in elderly people; a comparison of results with (/sup 15/N)glycine and (/sup 14/C)leucine

    Energy Technology Data Exchange (ETDEWEB)

    Golden, M H.N.; Waterlow, J C [London School of Hygiene and Tropical Medicine (UK)

    1977-09-01

    Total body protein turnover was studied in six elderly patients. During the study they were fed by continuous infusion of a liquid formula through a nasogastric tube. L-(1-/sup 14/C)leucine and (/sup 15/N)-glycine were infused at a constant rate for 30 h. The labelled glycine was infused into the intragastric line; the labelled leucine was given either by this route or intravenously. The specific radioactivity of free leucine in plasma and the rate of output of /sup 14/CO/sub 2/ in expired air both reached a plateau at 10 h, and remained constant until the end of the infusion at 30 h. The /sup 15/N abundance in urinary urea and total N was very similar. In neither was a plateau reached by 30 h but in four out of the six patients the abundance in urinary NH/sub 4//sup +/ had attained a plateau by the end of the infusion. Flux rates and rates of protein synthesis were calculated in four ways and a comparison of methods was used to examine the validity of the assumptions on which the different methods depended. The results suggest that the rate of protein turnover is reduced in the elderly, compared with younger subjects.

  13. Improved muscle function and quality after diet intervention with leucine-enriched whey and antioxidants in antioxidant deficient aged mice.

    Science.gov (United States)

    van Dijk, Miriam; Dijk, Francina J; Bunschoten, Annelies; van Dartel, Dorien A M; van Norren, Klaske; Walrand, Stephane; Jourdan, Marion; Verlaan, Sjors; Luiking, Yvette

    2016-04-05

    Antioxidant (AOX) deficiencies are commonly observed in older adults and oxidative stress has been suggested to contribute to sarcopenia. Here we investigate if 1) low levels of dietary antioxidants had a negative impact on parameters of muscle mass, function and quality, and 2) to study if nutritional interventions with AOX and/or leucine-enriched whey protein could improve these muscle parameters in aged mice. 18-months-old mice were fed a casein-based antioxidant-deficient (lowox) diet or a casein-based control-diet (CTRL) for 7 months. During the last 3 months, lowox-mice were subjected to either: a) continued lowox, b) supplementation with vitamin A/E, Selenium and Zinc (AOX), c) substitution of casein with leucine-enriched whey protein (PROT) or d) a combination of both AOX and PROT (TOTAL). After 7 months lowox-mice displayed lower muscle strength and more muscle fatigue compared to CTRL. Compared to lowox-mice, PROT-mice showed improved muscle power, grip strength and less muscle fatigue. AOX-mice showed improved oxidative status, less muscle fatigue, improved grip strength and mitochondrial dynamics compared to lowox-mice. The TOTAL-mice showed the combined effects of both interventions compared to lowox-mice. In conclusion, nutritional intervention with AOX and/or leucine-enriched whey protein can play a role in improving muscle health in a AOX-deficient mouse model.

  14. LETM1, a novel gene encoding a putative EF-hand Ca(2+)-binding protein, flanks the Wolf-Hirschhorn syndrome (WHS) critical region and is deleted in most WHS patients.

    Science.gov (United States)

    Endele, S; Fuhry, M; Pak, S J; Zabel, B U; Winterpacht, A

    1999-09-01

    Deletions within human chromosome 4p16.3 cause Wolf-Hirschhorn syndrome (WHS), which is characterized by severe mental and developmental defects. It is thought that haploinsufficiency of more than one gene contributes to the complex phenotype. We have cloned and characterized a novel gene (LETM1) that is deleted in nearly all WHS patients. LETM1 encodes a putative member of the EF-hand family of Ca(2+)-binding proteins. The protein contains two EF-hands, a transmembrane domain, a leucine zipper, and several coiled-coil domains. On the basis of its possible Ca(2+)-binding property and involvement in Ca(2+) signaling and/or homeostasis, we propose that haploinsufficiency of LETM1 may contribute to the neuromuscular features of WHS patients. Copyright 1999 Academic Press.

  15. How to find a leucine in a haystack? Structure, ligand recognition and regulation of leucine-aspartic acid (LD) motifs

    KAUST Repository

    Alam, Tanvir

    2014-05-29

    LD motifs (leucine-aspartic acidmotifs) are short helical protein-protein interaction motifs that have emerged as key players in connecting cell adhesion with cell motility and survival. LD motifs are required for embryogenesis, wound healing and the evolution of multicellularity. LD motifs also play roles in disease, such as in cancer metastasis or viral infection. First described in the paxillin family of scaffolding proteins, LD motifs and similar acidic LXXLL interaction motifs have been discovered in several other proteins, whereas 16 proteins have been reported to contain LDBDs (LD motif-binding domains). Collectively, structural and functional analyses have revealed a surprising multivalency in LD motif interactions and a wide diversity in LDBD architectures. In the present review, we summarize the molecular basis for function, regulation and selectivity of LD motif interactions that has emerged from more than a decade of research. This overview highlights the intricate multi-level regulation and the inherently noisy and heterogeneous nature of signalling through short protein-protein interaction motifs. © 2014 Biochemical Society.

  16. How to find a leucine in a haystack? Structure, ligand recognition and regulation of leucine-aspartic acid (LD) motifs

    KAUST Repository

    Alam, Tanvir; Alazmi, Meshari; Gao, Xin; Arold, Stefan T.

    2014-01-01

    LD motifs (leucine-aspartic acidmotifs) are short helical protein-protein interaction motifs that have emerged as key players in connecting cell adhesion with cell motility and survival. LD motifs are required for embryogenesis, wound healing and the evolution of multicellularity. LD motifs also play roles in disease, such as in cancer metastasis or viral infection. First described in the paxillin family of scaffolding proteins, LD motifs and similar acidic LXXLL interaction motifs have been discovered in several other proteins, whereas 16 proteins have been reported to contain LDBDs (LD motif-binding domains). Collectively, structural and functional analyses have revealed a surprising multivalency in LD motif interactions and a wide diversity in LDBD architectures. In the present review, we summarize the molecular basis for function, regulation and selectivity of LD motif interactions that has emerged from more than a decade of research. This overview highlights the intricate multi-level regulation and the inherently noisy and heterogeneous nature of signalling through short protein-protein interaction motifs. © 2014 Biochemical Society.

  17. Differentiating leucine incorporation of Archaea and Bacteria throughout the water column of the eastern Atlantic using metabolic inhibitors

    NARCIS (Netherlands)

    Yokokawa, Taichi; Sintes, Eva; de Corte, Daniele; Olbrich, Kerstin; Herndl, Gerhard J.

    2012-01-01

    The abundance (based on catalyzed reporter deposition-fluorescence in situ hybrid ization, CARD-FISH) and leucine incorporation rates of Archaea and Bacteria were determined throughout the water column in the eastern Atlantic. Bacteria dominated throughout the water column, although their

  18. Role of leucine in isoprenoid metabolism. Incorporation of (3-/sup 13/C)leucine and of (2-/sup 3/H,4-/sup 14/C)-. beta. ,. beta. -dimethyl-acrylic acid into phytosterols by tissue cultures of Andrographis paniculata

    Energy Technology Data Exchange (ETDEWEB)

    Anastasis, P; Freer, I; Overton, K; Rycroft, D; Singh, S B [Glasgow Univ. (UK). Dept. of Chemistry

    1985-02-01

    (3-/sup 13/C)Leucine is incorporated into phytosterols by tissue cultures of Andrographis paniculata by breakdown to acetyl-CoA and its subsequent incorporation via (3S)-3-hydroxy-3-methylglutaryl-CoA (HMG-CoA) and mevalonic acid; (2-/sup 3/H,4-/sup 14/C)-..beta..,..beta..-dimethylacrylic acid also is not incorporated intact.

  19. Application of Raman spectroscopy in type 2 diabetes screening in blood using leucine and isoleucine amino-acids as biomarkers and in comparative anti-diabetic drugs efficacy studies.

    Science.gov (United States)

    Birech, Zephania; Mwangi, Peter Waweru; Bukachi, Fredrick; Mandela, Keith Makori

    2017-01-01

    Diabetes is an irreversible condition characterized by elevated blood glucose levels. Currently, there are no predictive biomarkers for this disease and the existing ones such as hemoglobin A1c and fasting blood glucose are used only when diabetes symptoms are noticed. The objective of this work was first to explore the potential of leucine and isoleucine amino acids as diabetes type 2 biomarkers using their Raman spectroscopic signatures. Secondly, we wanted to explore whether Raman spectroscopy can be applied in comparative efficacy studies between commercially available anti-diabetic drug pioglitazone and the locally used anti-diabetic herbal extract Momordica spinosa (Gilg.)Chiov. Sprague Dawley (SD) rat's blood was used and were pipetted onto Raman substrates prepared from conductive silver paste smeared glass slides. Prominent Raman bands associated with glucose (926, 1302, 1125 cm-1), leucine (1106, 1248, 1302, 1395 cm-1) and isolecucine (1108, 1248, 1437 and 1585 cm-1) were observed. The Raman bands centered at 1125 cm-1, 1395 cm-1 and 1437 cm-1 associated respectively to glucose, leucine and isoleucine were chosen as biomarker Raman peaks for diabetes type 2. These Raman bands displayed decreased intensities in blood from diabetic SD rats administered antidiabetic drugs pioglitazone and herbal extract Momordica spinosa (Gilg.)Chiov. The intensity decrease indicated reduced concentration levels of the respective biomarker molecules: glucose (1125 cm-1), leucine (1395 cm-1) and isoleucine (1437 cm-1) in blood. The results displayed the power and potential of Raman spectroscopy in rapid (10 seconds) diabetes and pre-diabetes screening in blood (human or rat's) with not only glucose acting as a biomarker but also leucine and isoleucine amino-acids where intensities of respectively assigned bands act as references. It also showed that using Raman spectroscopic signatures of the chosen biomarkers, the method can be an alternative for performing comparative

  20. Application of Raman spectroscopy in type 2 diabetes screening in blood using leucine and isoleucine amino-acids as biomarkers and in comparative anti-diabetic drugs efficacy studies.

    Directory of Open Access Journals (Sweden)

    Zephania Birech

    Full Text Available Diabetes is an irreversible condition characterized by elevated blood glucose levels. Currently, there are no predictive biomarkers for this disease and the existing ones such as hemoglobin A1c and fasting blood glucose are used only when diabetes symptoms are noticed. The objective of this work was first to explore the potential of leucine and isoleucine amino acids as diabetes type 2 biomarkers using their Raman spectroscopic signatures. Secondly, we wanted to explore whether Raman spectroscopy can be applied in comparative efficacy studies between commercially available anti-diabetic drug pioglitazone and the locally used anti-diabetic herbal extract Momordica spinosa (Gilg.Chiov. Sprague Dawley (SD rat's blood was used and were pipetted onto Raman substrates prepared from conductive silver paste smeared glass slides. Prominent Raman bands associated with glucose (926, 1302, 1125 cm-1, leucine (1106, 1248, 1302, 1395 cm-1 and isolecucine (1108, 1248, 1437 and 1585 cm-1 were observed. The Raman bands centered at 1125 cm-1, 1395 cm-1 and 1437 cm-1 associated respectively to glucose, leucine and isoleucine were chosen as biomarker Raman peaks for diabetes type 2. These Raman bands displayed decreased intensities in blood from diabetic SD rats administered antidiabetic drugs pioglitazone and herbal extract Momordica spinosa (Gilg.Chiov. The intensity decrease indicated reduced concentration levels of the respective biomarker molecules: glucose (1125 cm-1, leucine (1395 cm-1 and isoleucine (1437 cm-1 in blood. The results displayed the power and potential of Raman spectroscopy in rapid (10 seconds diabetes and pre-diabetes screening in blood (human or rat's with not only glucose acting as a biomarker but also leucine and isoleucine amino-acids where intensities of respectively assigned bands act as references. It also showed that using Raman spectroscopic signatures of the chosen biomarkers, the method can be an alternative for performing

  1. Proliferation related acidic leucine-rich protein PAL31 functions as a caspase-3 inhibitor

    International Nuclear Information System (INIS)

    Sun Weiyong; Kimura, Hiromichi; Hattori, Naka; Tanaka, Satoshi; Matsuyama, Shigemi; Shiota, Kunio

    2006-01-01

    Proliferation related acidic leucine-rich protein PAL31 (PAL31) is expressed in proliferating cells and consists of 272 amino acids with a tandem structure of leucine-rich repeats in the N-terminus and a highly acidic region with a putative nuclear localization signal in the C-terminus. We previously reported that PAL31 is required for cell cycle progression. In the present study, we found that the antisense oligonucleotide of PAL31 induced apoptosis to the transfected Nb2 cells. Stable transfectants, in which PAL31 was regulated by an inducible promoter, were generated to gain further insight into the signaling role of PAL31 in the regulation of apoptosis. Expression of PAL31 resulted in the marked rescue of Rat1 cells from etoposide and UV radiation-induced apoptosis and the cytoprotection was correlated with the levels of PAL31 protein. Thus, cytoprotection from apoptosis is a physiological function of PAL31. PAL31 can suppress caspase-3 activity but not cytochrome c release in vitro, indicating that PAL31 is a direct caspase-3 inhibitor. In conclusion, PAL31 is a multifunctional protein working as a cell cycle progression factor as well as a cell survival factor

  2. Identification of a novel Leucine-rich repeat protein and candidate PP1 regulatory subunit expressed in developing spermatids

    Directory of Open Access Journals (Sweden)

    Sperry Ann O

    2008-01-01

    Full Text Available Abstract Background Spermatogenesis is comprised of a series of highly regulated developmental changes that transform the precursor germ cell into a highly specialized spermatozoon. The last phase of spermatogenesis, termed spermiogenesis, involves dramatic morphological change including formation of the acrosome, elongation and condensation of the nucleus, formation of the flagella, and disposal of unnecessary cytoplasm. A prominent cytoskeletal component of the developing spermatid is the manchette, a unique microtubular structure that surrounds the nucleus of the developing spermatid and is thought to assist in both the reshaping of the nucleus and redistribution of spermatid cytoplasm. Although the molecular motor KIFC1 has been shown to associate with the manchette, its precise role in function of the manchette and the identity of its testis specific protein partners are unknown. The purpose of this study was to identify proteins in the testis that interact with KIFC1 using a yeast 2 hybrid screen of a testis cDNA library. Results Thirty percent of the interacting clones identified in our screen contain an identical cDNA encoding a 40 kD protein. This interacting protein has 4 leucine-rich repeats in its amino terminal half and is expressed primarily in the testis; therefore we have named this protein testis leucine-rich repeat protein or TLRR. TLRR was also found to associate tightly with the KIFC1 targeting domain using affinity chromatography. In addition to the leucine-rich repeats, TLRR contains a consensus-binding site for protein phosphatase-1 (PP1. Immunocytochemistry using a TLRR specific antibody demonstrates that this protein is found near the manchette of developing spermatids. Conclusion We have identified a previously uncharacterized leucine-rich repeat protein that is expressed abundantly in the testis and associates with the manchette of developing spermatids, possibly through its interaction with the KIFC1 molecular motor

  3. Effect of infection by Phytophthora infestans and treatment with its zoosporial components on uptake of 3H-leucine and protein synthesis potato tuber tissue

    International Nuclear Information System (INIS)

    Nishimura, Norio; Tomiyama, Kohei; Doke, Noriyuki

    1978-01-01

    Both sides of aged disks of potato tubers were inoculated with incompatible race 0 or compatible race 1 of P. infestans. The uptake of radioactivity from 3 H-leucine into the disks was greatly reduced by the infection with the incompatible race, but the ratio of incorporated radioactivity in the acid soluble fraction of the disks to total radioactivity in the 20,000 x g supernatant was increased by the infection with the incompatible race 0. In case of the compatible race 1, the infection had less effect on the incorporation ratio of the radioactivity into the acid insoluble fraction than the case of the race 0. The results suggested that the reduction in 3 H-leucine uptake may be due to alteration of the physiological activity of host plasma membrane, instead of the reduction of metabolic activity. The treatment of tuber disks with the insoluble fraction of the zoosporial homogenate of race 0 or race 1 greatly reduced the uptake of 3 H-leucine by the disks. The supernatant fraction of the zoosporial homogenate also reduced the uptake, but the effect was lost by dialysis. The results suggest that some substances in the insoluble components of the zoosporial homogenate of P. infestans may be at least one cause of the reduction in the uptake of 3 H-leucine which begins to occur at very early period of the inoculation with race 0 or race 1 while penetration has not yet occurred. (Kobatake, H.)

  4. Effect of ascorbate, nitrate and nitrite on the amount of flavour compounds produced from leucine by Staphylococcus xylosus and Staphylococcus carnosus

    DEFF Research Database (Denmark)

    Olesen, Pelle Thonning; Stahnke, Louise Heller; Talon, R.

    2004-01-01

    Resting cells of Staphylococcus xylosus and S. carnosus were incubated with ascorbate, nitrate and nitrite in defined reaction medium and their degradation of H-3-labelled leucine into methyl-branched catabolites were studied using HPLC/radiometric detection. The experiments were carried out...... of nitrite and nitrate for S. xylosus. Addition of alpha-ketoglutarate generally increased the concentration of 3-methylbutanoic acid for both S. xylosus and S. carnosus....... with and without addition of alpha-ketoglutarate. The main catabolic product of leucine degradation was 3-methylbutanoic acid but also small amounts of alpha-hydroxy isocaproic acid were produced. Nitrite addition lowered the concentration of 3-methylbutanoic acid for both Staphylococcus species and this effect...

  5. Effect of leucine-to-methionine substitutions on the diffraction quality of histone chaperone SET/TAF-Iβ/INHAT crystals

    International Nuclear Information System (INIS)

    Senda, Miki; Muto, Shinsuke; Horikoshi, Masami; Senda, Toshiya

    2008-01-01

    The combination of leucine-to-methionine substitutions and optimization of cryoconditions improved the resolution of histone chaperone SET/TAF-Iβ/INHAT crystals from around 5.5 to 2.3 Å without changing the crystallization conditions, allowing successful structure determination of SET/TAF-Iβ/INHAT by the multiwavelength anomalous diffraction method. One of the most frequent problems in crystallization is poor quality of the crystals. In order to overcome this obstacle several methods have been utilized, including amino-acid substitutions of the target protein. Here, an example is presented of crystal-quality improvement by leucine-to-methionine substitutions. A variant protein with three amino-acid substitutions enabled improvement of the crystal quality of the histone chaperone SET/TAF-Iβ/INHAT when combined with optimization of the cryoconditions. This procedure improved the resolution of the SET/TAF-Iβ/INHAT crystals from around 5.5 to 2.3 Å without changing the crystallization conditions

  6. Leucine supplementation stimulates protein synthesis and reduces degradation signal activation in muscle of newborn pigs during acute endotoxemia

    Science.gov (United States)

    Sepsis disrupts skeletal muscle proteostasis and mitigates the anabolic response to leucine (Leu) in muscle of mature animals. We have shown that Leu stimulates muscle protein synthesis (PS) in healthy neonatal piglets. To determine if supplemental Leu can stimulate PS and reduce protein degradation...

  7. Interactive Smart Fashion Using User-Oriented Visible Light Communication: The Case of Modular Strapped Cuffs and Zipper Slider Types

    Directory of Open Access Journals (Sweden)

    Jai-Eun Kim

    2017-01-01

    Full Text Available Because LEDs offer flexible expressions such as brightness, color control, and various patterns, they are popularly used in multidevice interactions. Moreover, LEDs have excellent physical characteristics. However, existing LED light-based wearable interactions are designed for interest and attention. So, LED can be used in fashion as it can give new look to our style and at the same time also as an interaction device. Therefore, in this paper, we present the design guideline for regulating the technical implementation, design strategies, and directions of interactive LED devices. The technology and design concepts are demonstrated through a case study (analysis of an existing LED light-based wearable interaction. We also design a scenario-based iterative collaborative design process model. Finally, we develop a smart fashion of modular strapped cuffs and zipper slider types that can be attached and detached according to the user’s preference as the interactive smart fashion using user-oriented visible light communication, ultimately pursuing a visual-MIMO (Multiple-Input Multiple-Output product through stepwise strategy.

  8. The zipper mechanism in phagocytosis: energetic requirements and variability in phagocytic cup shape

    Directory of Open Access Journals (Sweden)

    Dart Anna E

    2010-11-01

    Full Text Available Abstract Background Phagocytosis is the fundamental cellular process by which eukaryotic cells bind and engulf particles by their cell membrane. Particle engulfment involves particle recognition by cell-surface receptors, signaling and remodeling of the actin cytoskeleton to guide the membrane around the particle in a zipper-like fashion. Despite the signaling complexity, phagocytosis also depends strongly on biophysical parameters, such as particle shape, and the need for actin-driven force generation remains poorly understood. Results Here, we propose a novel, three-dimensional and stochastic biophysical model of phagocytosis, and study the engulfment of particles of various sizes and shapes, including spiral and rod-shaped particles reminiscent of bacteria. Highly curved shapes are not taken up, in line with recent experimental results. Furthermore, we surprisingly find that even without actin-driven force generation, engulfment proceeds in a large regime of parameter values, albeit more slowly and with highly variable phagocytic cups. We experimentally confirm these predictions using fibroblasts, transfected with immunoreceptor FcγRIIa for engulfment of immunoglobulin G-opsonized particles. Specifically, we compare the wild-type receptor with a mutant receptor, unable to signal to the actin cytoskeleton. Based on the reconstruction of phagocytic cups from imaging data, we indeed show that cells are able to engulf small particles even without support from biological actin-driven processes. Conclusions This suggests that biochemical pathways render the evolutionary ancient process of phagocytic highly robust, allowing cells to engulf even very large particles. The particle-shape dependence of phagocytosis makes a systematic investigation of host-pathogen interactions and an efficient design of a vehicle for drug delivery possible.

  9. Biodegradability and tissue reaction of random copolymers of L-leucine, L-aspartic acid, and L-aspartic acid esters

    NARCIS (Netherlands)

    Marck, K.W.; Wildevuur, Ch.R.H.; Sederel, W.L.; Bantjes, A.; Feijen, Jan

    1977-01-01

    A series of copoly(α-amino acids) with varying percentages of hydrophilic (l-aspartic acid) and hydrophobic monomers (l-leucine, ß-methyl-l-aspartate, and ß-benzyl-l-aspartate) were implanted subcutaneously in rats and the macroscopic degradation behavior was studied. Three groups of materials (A,

  10. The enzymatic degradation and transport of leucine-enkephalin and 4-imidazolidinone enkephalin prodrugs at the blood-brain barrier

    DEFF Research Database (Denmark)

    Lund, L.; Bak, A.; Friis, G.J.

    1998-01-01

    In this study, the stability in and transport across a cell culture model of the blood-brain barrier (BBB) is investigated for leucine-enkephalin (Leu-enkephalin) and four 4-imidazolidinone prodrugs of Leu-enkephalin. The results show that Leu-enkephalin is degraded in the cell culture model...

  11. Establishing a Mathematical Equations and Improving the Production of L-tert-Leucine by Uniform Design and Regression Analysis.

    Science.gov (United States)

    Jiang, Wei; Xu, Chao-Zhen; Jiang, Si-Zhi; Zhang, Tang-Duo; Wang, Shi-Zhen; Fang, Bai-Shan

    2017-04-01

    L-tert-Leucine (L-Tle) and its derivatives are extensively used as crucial building blocks for chiral auxiliaries, pharmaceutically active ingredients, and ligands. Combining with formate dehydrogenase (FDH) for regenerating the expensive coenzyme NADH, leucine dehydrogenase (LeuDH) is continually used for synthesizing L-Tle from α-keto acid. A multilevel factorial experimental design was executed for research of this system. In this work, an efficient optimization method for improving the productivity of L-Tle was developed. And the mathematical model between different fermentation conditions and L-Tle yield was also determined in the form of the equation by using uniform design and regression analysis. The multivariate regression equation was conveniently implemented in water, with a space time yield of 505.9 g L -1  day -1 and an enantiomeric excess value of >99 %. These results demonstrated that this method might become an ideal protocol for industrial production of chiral compounds and unnatural amino acids such as chiral drug intermediates.

  12. Bacterial incorporation of leucine into protein down to -20 degrees C with evidence for potential activity in sub-eutectic saline ice formations.

    Science.gov (United States)

    Junge, Karen; Eicken, Hajo; Swanson, Brian D; Deming, Jody W

    2006-06-01

    Direct evidence for metabolism in a variety of frozen environments has pushed temperature limits for bacterial activity to increasingly lower temperatures, so far to -20 degrees C. To date, the metabolic activities of marine psychrophilic bacteria, important components of sea-ice communities, have not been studied in laboratory culture, not in ice and not below -12 degrees C. We measured [3H]-leucine incorporation into macromolecules (further fractionated biochemically) by the marine psychrophilic bacterium Colwellia psychrerythraea strain 34H over a range of anticipated activity-permissive temperatures, from +13 to -20 degrees C, including expected negative controls at -80 and -196 degrees C. For incubation temperatures below -1 degrees C, the cell suspensions [all in artificial seawater (ASW)] were first quick-frozen in liquid nitrogen. We also examined the effect of added extracellular polymeric substances (EPS) on [3H]-leucine incorporation. Results showed that live cells of strain 34H incorporated substantial amounts of [3H]-leucine into TCA-precipitable material (primarily protein) down to -20 degrees C. At temperatures from -1 to -20 degrees C, rates were enhanced by EPS. No activity was detected in the killed controls for strain 34H (or in Escherichia coli controls), which included TCA-killed, heat-killed, and sodium azide- and chloramphenicol-treated samples. Surprisingly, evidence for low but significant rates of intracellular incorporation of [3H]-leucine into protein was observed for both ASW-only and EPS-amended (and live only) samples incubated at -80 and -196 degrees C. Mechanisms that could explain the latter results require further study, but the process of vitrification promoted by rapid freezing and the presence of salts and organic polymers may be relevant. Overall, distinguishing between intracellular and extracellular aspects of bacterial activity appears important to understanding behavior at sub-freezing temperatures.

  13. Leucine supplementation of a chronically restricted protein and energy diet enhances mTOR pathway activation but not muscle protein synthesis in neonatal pigs.

    Science.gov (United States)

    Manjarín, Rodrigo; Columbus, Daniel A; Suryawan, Agus; Nguyen, Hanh V; Hernandez-García, Adriana D; Hoang, Nguyet-Minh; Fiorotto, Marta L; Davis, Teresa

    2016-01-01

    Suboptimal nutrient intake represents a limiting factor for growth and long-term survival of low-birth weight infants. The objective of this study was to determine if in neonates who can consume only 70 % of their protein and energy requirements for 8 days, enteral leucine supplementation will upregulate the mammalian target of rapamycin (mTOR) pathway in skeletal muscle, leading to an increase in protein synthesis and muscle anabolism. Nineteen 4-day-old piglets were fed by gastric tube 1 of 3 diets, containing (kg body weight(-1) · day(-1)) 16 g protein and 190 kcal (CON), 10.9 g protein and 132 kcal (R), or 10.8 g protein + 0.2 % leucine and 136 kcal (RL) at 4-h intervals for 8 days. On day 8, plasma AA and insulin levels were measured during 6 post-feeding intervals, and muscle protein synthesis rate and mTOR signaling proteins were determined at 120 min post-feeding. At 120 min, leucine was highest in RL (P protein synthesis, phosphorylation of S6 kinase (p-S6K1) and 4E-binding protein (p-4EBP1), and activation of eukaryotic initiation factor 4 complex (eIF4E · eIF4G). RL increased (P ≤ 0.01) p-S6K1, p-4EBP1 and eIF4E · eIF4G compared to R. In conclusion, when protein and energy intakes are restricted for 8 days, leucine supplementation increases muscle mTOR activation, but does not improve body weight gain or enhance skeletal muscle protein synthesis in neonatal pigs.

  14. The putative bZIP transcription factor BzpN slows proliferation and functions in the regulation of cell density by autocrine signals in Dictyostelium.

    Directory of Open Access Journals (Sweden)

    Jonathan E Phillips

    Full Text Available The secreted proteins AprA and CfaD function as autocrine signals that inhibit cell proliferation in Dictyostelium discoideum, thereby regulating cell numbers by a negative feedback mechanism. We report here that the putative basic leucine zipper transcription factor BzpN plays a role in the inhibition of proliferation by AprA and CfaD. Cells lacking BzpN proliferate more rapidly than wild-type cells but do not reach a higher stationary density. Recombinant AprA inhibits wild-type cell proliferation but does not inhibit the proliferation of cells lacking BzpN. Recombinant CfaD also inhibits wild-type cell proliferation, but promotes the proliferation of cells lacking BzpN. Overexpression of BzpN results in a reduced cell density at stationary phase, and this phenotype requires AprA, CfaD, and the kinase QkgA. Conditioned media from high-density cells stops the proliferation of wild-type but not bzpN(- cells and induces a nuclear localization of a BzpN-GFP fusion protein, though this localization does not require AprA or CfaD. Together, the data suggest that BzpN is necessary for some but not all of the effects of AprA and CfaD, and that BzpN may function downstream of AprA and CfaD in a signal transduction pathway that inhibits proliferation.

  15. The Putative bZIP Transcripton Factor BzpN Slows Proliferation and Functions in the Regulation of Cell Density by Autocrine Signals in Dictyostelium

    Science.gov (United States)

    Phillips, Jonathan E.; Huang, Eryong; Shaulsky, Gad; Gomer, Richard H.

    2011-01-01

    The secreted proteins AprA and CfaD function as autocrine signals that inhibit cell proliferation in Dictyostelium discoideum, thereby regulating cell numbers by a negative feedback mechanism. We report here that the putative basic leucine zipper transcription factor BzpN plays a role in the inhibition of proliferation by AprA and CfaD. Cells lacking BzpN proliferate more rapidly than wild-type cells but do not reach a higher stationary density. Recombinant AprA inhibits wild-type cell proliferation but does not inhibit the proliferation of cells lacking BzpN. Recombinant CfaD also inhibits wild-type cell proliferation, but promotes the proliferation of cells lacking BzpN. Overexpression of BzpN results in a reduced cell density at stationary phase, and this phenotype requires AprA, CfaD, and the kinase QkgA. Conditioned media from high-density cells stops the proliferation of wild-type but not bzpN− cells and induces a nuclear localization of a BzpN-GFP fusion protein, though this localization does not require AprA or CfaD. Together, the data suggest that BzpN is necessary for some but not all of the effects of AprA and CfaD, and that BzpN may function downstream of AprA and CfaD in a signal transduction pathway that inhibits proliferation. PMID:21760904

  16. The putative bZIP transcription factor BzpN slows proliferation and functions in the regulation of cell density by autocrine signals in Dictyostelium.

    Science.gov (United States)

    Phillips, Jonathan E; Huang, Eryong; Shaulsky, Gad; Gomer, Richard H

    2011-01-01

    The secreted proteins AprA and CfaD function as autocrine signals that inhibit cell proliferation in Dictyostelium discoideum, thereby regulating cell numbers by a negative feedback mechanism. We report here that the putative basic leucine zipper transcription factor BzpN plays a role in the inhibition of proliferation by AprA and CfaD. Cells lacking BzpN proliferate more rapidly than wild-type cells but do not reach a higher stationary density. Recombinant AprA inhibits wild-type cell proliferation but does not inhibit the proliferation of cells lacking BzpN. Recombinant CfaD also inhibits wild-type cell proliferation, but promotes the proliferation of cells lacking BzpN. Overexpression of BzpN results in a reduced cell density at stationary phase, and this phenotype requires AprA, CfaD, and the kinase QkgA. Conditioned media from high-density cells stops the proliferation of wild-type but not bzpN(-) cells and induces a nuclear localization of a BzpN-GFP fusion protein, though this localization does not require AprA or CfaD. Together, the data suggest that BzpN is necessary for some but not all of the effects of AprA and CfaD, and that BzpN may function downstream of AprA and CfaD in a signal transduction pathway that inhibits proliferation.

  17. Genomic survey of bZIP transcription factor genes related to tanshinone biosynthesis in Salvia miltiorrhiza

    Directory of Open Access Journals (Sweden)

    Yu Zhang

    2018-03-01

    Full Text Available Tanshinones are a class of bioactive components in the traditional Chinese medicine Salvia miltiorrhiza, and their biosynthesis and regulation have been widely studied. Current studies show that basic leucine zipper (bZIP proteins regulate plant secondary metabolism, growth and developmental processes. However, the bZIP transcription factors involved in tanshinone biosynthesis are unknown. Here, we conducted the first genome-wide survey of the bZIP gene family and analyzed the phylogeny, gene structure, additional conserved motifs and alternative splicing events in S. miltiorrhiza. A total of 70 SmbZIP transcription factors were identified and categorized into 11 subgroups based on their phylogenetic relationships with those in Arabidopsis. Moreover, seventeen SmbZIP genes underwent alternative splicing events. According to the transcriptomic data, the SmbZIP genes that were highly expressed in the Danshen root and periderm were selected. Based on the prediction of bZIP binding sites in the promoters and the co-expression analysis and co-induction patterns in response to Ag+ treatment via quantitative real-time polymerase chain reaction (qRT-PCR, we concluded that SmbZIP7 and SmbZIP20 potentially participate in the regulation of tanshinone biosynthesis. These results provide a foundation for further functional characterization of the candidate SmbZIP genes, which have the potential to increase tanshinone production. KEY WORDS: bZIP genes, Salvia miltiorrhiza, Phylogenetic analysis, Expression pattern analysis, Tanshinone biosynthesis

  18. Variation Analysis of Physiological Traits in Betula platyphylla Overexpressing TaLEA-ThbZIP Gene under Salt Stress.

    Directory of Open Access Journals (Sweden)

    Xiyang Zhao

    Full Text Available The aim of this study was to determine whether transgenic birch (Betula platyphylla ectopic overexpressing a late embryogenesis abundant (LEA gene and a basic leucine zipper (bZIP gene from the salt-tolerant genus Tamarix (salt cedar show increased tolerance to salt (NaCl stress. Co-transfer of TaLEA and ThbZIP in birch under the control of two independent CaMV 35S promoters significantly enhanced salt stress. PCR and northern blot analyses indicated that the two genes were ectopically overexpressed in several dual-gene transgenic birch lines. We compared the effects of salt stress among three transgenic birch lines (L-4, L-5, and L-8 and wild type (WT. In all lines, the net photosynthesis values were higher before salt stress treatment than afterwards. After the salt stress treatment, the transgenic lines L-4 and L-8 showed higher values for photosynthetic traits, chlorophyll fluorescence, peroxidase and superoxide dismutase activities, and lower malondialdehyde and Na+ contents, compared with those in WT and L-5. These different responses to salt stress suggested that the transcriptional level of the TaLEA and ThbZIP genes differed among the transgenic lines, resulting in a variety of genetic and phenotypic effects. The results of this research can provide a theoretical basis for the genetic engineering of salt-tolerant trees.

  19. Distinct Plasma Profile of Polar Neutral Amino Acids, Leucine, and Glutamate in Children with Autism Spectrum Disorders

    Science.gov (United States)

    Tirouvanziam, Rabindra; Obukhanych, Tetyana V.; Laval, Julie; Aronov, Pavel A.; Libove, Robin; Banerjee, Arpita Goswami; Parker, Karen J.; O'Hara, Ruth; Herzenberg, Leonard A.; Herzenberg, Leonore A.; Hardan, Antonio Y.

    2012-01-01

    The goal of this investigation was to examine plasma amino acid (AA) levels in children with Autism Spectrum Disorders (ASD, N = 27) and neuro-typically developing controls (N = 20). We observed reduced plasma levels of most polar neutral AA and leucine in children with ASD. This AA profile conferred significant post hoc power for discriminating…

  20. Molecular characterization of the di-leucine-based internalization motif of the T cell receptor

    DEFF Research Database (Denmark)

    Dietrich, J; Hou, X; Wegener, A M

    1996-01-01

    Several cell surface receptors including the T cell receptor (TCR) are phosphorylated and down-regulated following activation of protein kinases. We have recently shown that both phosphorylation of Ser-126 and the presence of the di-leucine sequence Leu-131 and Leu-132 in CD3 gamma are required f...... are important. 2) Recognition of phosphorylated CD3 gamma by molecules involved in receptor internalization. In this process Ser(P)-126, Asp-127, Leu-131, and Leu-132 are important....

  1. A 21.7 kb DNA segment on the left arm of yeast chromosome XIV carries WHI3, GCR2, SPX18, SPX19, an homologue to the heat shock gene SSB1 and 8 new open reading frames of unknown function.

    Science.gov (United States)

    Jonniaux, J L; Coster, F; Purnelle, B; Goffeau, A

    1994-12-01

    We report the amino acid sequence of 13 open reading frames (ORF > 299 bp) located on a 21.7 kb DNA segment from the left arm of chromosome XIV of Saccharomyces cerevisiae. Five open reading frames had been entirely or partially sequenced previously: WHI3, GCR2, SPX19, SPX18 and a heat shock gene similar to SSB1. The products of 8 other ORFs are new putative proteins among which N1394 is probably a membrane protein. N1346 contains a leucine zipper pattern and the corresponding ORF presents an HAP (global regulator of respiratory genes) upstream activating sequence in the promoting region. N1386 shares homologies with the DNA structure-specific recognition protein family SSRPs and the corresponding ORF is preceded by an MCB (MluI cell cycle box) upstream activating factor.

  2. 14C leucine chloromethylketone interaction with sarcoma 37 cell plasma membrane components

    International Nuclear Information System (INIS)

    Matthews, R.H.; Milo, G.E.; McMichael, T.L.; Lewis, N.J.

    1982-01-01

    Leucine chloromethylketone labelling of viable S37 cells was preferential for the plasma membrane fraction. The pattern of radiolabelling of the plasma membrane proteins was time-dependent. After 5 min the radiolabel was localized with glutamyl transpeptidase, and subsequently, with other physiologically active proteins as a function of time after incubation. Labelling of proteins was temperature-dependent and incubation of viable S37 cells with the radiolabelled substrate at 0 0 C yielded little or no radioactivity localized in the plasma membrane. The molecular weight of one radiolabelled substratemembrane protein complex was estimated on sodium dodecyl sulfate polyacrylamide gel electrophoresis to be between 100,000-200,000. (author)

  3. Repression of branched-chain amino acid synthesis in Staphylococcus aureus is mediated by isoleucine via CodY, and by a leucine-rich attenuator peptide.

    Science.gov (United States)

    Kaiser, Julienne C; King, Alyssa N; Grigg, Jason C; Sheldon, Jessica R; Edgell, David R; Murphy, Michael E P; Brinsmade, Shaun R; Heinrichs, David E

    2018-01-01

    Staphylococcus aureus requires branched-chain amino acids (BCAAs; isoleucine, leucine, valine) for protein synthesis, branched-chain fatty acid synthesis, and environmental adaptation by responding to their availability via the global transcriptional regulator CodY. The importance of BCAAs for S. aureus physiology necessitates that it either synthesize them or scavenge them from the environment. Indeed S. aureus uses specialized transporters to scavenge BCAAs, however, its ability to synthesize them has remained conflicted by reports that it is auxotrophic for leucine and valine despite carrying an intact BCAA biosynthetic operon. In revisiting these findings, we have observed that S. aureus can engage in leucine and valine synthesis, but the level of BCAA synthesis is dependent on the BCAA it is deprived of, leading us to hypothesize that each BCAA differentially regulates the biosynthetic operon. Here we show that two mechanisms of transcriptional repression regulate the level of endogenous BCAA biosynthesis in response to specific BCAA availability. We identify a trans-acting mechanism involving isoleucine-dependent repression by the global transcriptional regulator CodY and a cis-acting leucine-responsive attenuator, uncovering how S. aureus regulates endogenous biosynthesis in response to exogenous BCAA availability. Moreover, given that isoleucine can dominate CodY-dependent regulation of BCAA biosynthesis, and that CodY is a global regulator of metabolism and virulence in S. aureus, we extend the importance of isoleucine availability for CodY-dependent regulation of other metabolic and virulence genes. These data resolve the previous conflicting observations regarding BCAA biosynthesis, and reveal the environmental signals that not only induce BCAA biosynthesis, but that could also have broader consequences on S. aureus environmental adaptation and virulence via CodY.

  4. Repression of branched-chain amino acid synthesis in Staphylococcus aureus is mediated by isoleucine via CodY, and by a leucine-rich attenuator peptide.

    Directory of Open Access Journals (Sweden)

    Julienne C Kaiser

    2018-01-01

    Full Text Available Staphylococcus aureus requires branched-chain amino acids (BCAAs; isoleucine, leucine, valine for protein synthesis, branched-chain fatty acid synthesis, and environmental adaptation by responding to their availability via the global transcriptional regulator CodY. The importance of BCAAs for S. aureus physiology necessitates that it either synthesize them or scavenge them from the environment. Indeed S. aureus uses specialized transporters to scavenge BCAAs, however, its ability to synthesize them has remained conflicted by reports that it is auxotrophic for leucine and valine despite carrying an intact BCAA biosynthetic operon. In revisiting these findings, we have observed that S. aureus can engage in leucine and valine synthesis, but the level of BCAA synthesis is dependent on the BCAA it is deprived of, leading us to hypothesize that each BCAA differentially regulates the biosynthetic operon. Here we show that two mechanisms of transcriptional repression regulate the level of endogenous BCAA biosynthesis in response to specific BCAA availability. We identify a trans-acting mechanism involving isoleucine-dependent repression by the global transcriptional regulator CodY and a cis-acting leucine-responsive attenuator, uncovering how S. aureus regulates endogenous biosynthesis in response to exogenous BCAA availability. Moreover, given that isoleucine can dominate CodY-dependent regulation of BCAA biosynthesis, and that CodY is a global regulator of metabolism and virulence in S. aureus, we extend the importance of isoleucine availability for CodY-dependent regulation of other metabolic and virulence genes. These data resolve the previous conflicting observations regarding BCAA biosynthesis, and reveal the environmental signals that not only induce BCAA biosynthesis, but that could also have broader consequences on S. aureus environmental adaptation and virulence via CodY.

  5. Anti-leucine rich glioma inactivated 1 protein and anti-N-methyl-D-aspartate receptor encephalitis show distinct patterns of brain glucose metabolism in 18F-fluoro-2-deoxy-d-glucose positron emission tomography.

    Science.gov (United States)

    Wegner, Florian; Wilke, Florian; Raab, Peter; Tayeb, Said Ben; Boeck, Anna-Lena; Haense, Cathleen; Trebst, Corinna; Voss, Elke; Schrader, Christoph; Logemann, Frank; Ahrens, Jörg; Leffler, Andreas; Rodriguez-Raecke, Rea; Dengler, Reinhard; Geworski, Lilli; Bengel, Frank M; Berding, Georg; Stangel, Martin; Nabavi, Elham

    2014-06-20

    Pathogenic autoantibodies targeting the recently identified leucine rich glioma inactivated 1 protein and the subunit 1 of the N-methyl-D-aspartate receptor induce autoimmune encephalitis. A comparison of brain metabolic patterns in 18F-fluoro-2-deoxy-d-glucose positron emission tomography of anti-leucine rich glioma inactivated 1 protein and anti-N-methyl-D-aspartate receptor encephalitis patients has not been performed yet and shall be helpful in differentiating these two most common forms of autoimmune encephalitis. The brain 18F-fluoro-2-deoxy-d-glucose uptake from whole-body positron emission tomography of six anti-N-methyl-D-aspartate receptor encephalitis patients and four patients with anti-leucine rich glioma inactivated 1 protein encephalitis admitted to Hannover Medical School between 2008 and 2012 was retrospectively analyzed and compared to matched controls. Group analysis of anti-N-methyl-D-aspartate encephalitis patients demonstrated regionally limited hypermetabolism in frontotemporal areas contrasting an extensive hypometabolism in parietal lobes, whereas the anti-leucine rich glioma inactivated 1 protein syndrome was characterized by hypermetabolism in cerebellar, basal ganglia, occipital and precentral areas and minor frontomesial hypometabolism. This retrospective 18F-fluoro-2-deoxy-d-glucose positron emission tomography study provides novel evidence for distinct brain metabolic patterns in patients with anti-leucine rich glioma inactivated 1 protein and anti-N-methyl-D-aspartate receptor encephalitis.

  6. The Effects of Leucine, Zinc, and Chromium Supplements on Inflammatory Events of the Respiratory System in Type 2 Diabetic Rats.

    Directory of Open Access Journals (Sweden)

    Saeed Kolahian

    Full Text Available Diabetes mellitus is a major cause of serious micro- and macrovascular diseases that affect nearly every system in the body, including the respiratory system. Non-enzymatic protein glycation due to hyperglycaemic stress has fundamental implications due to the large capillary network and amount of connective tissue in the lung. The current study was designed to determine whether leucine, zinc, and chromium supplementations influence the function and histological structure of the respiratory tract in a rat model of type 2 diabetes. Seventy-seven rats were divided into eleven groups, consisting of 7 animals each. One group served as negative control and insulin and glibenclamide were used as positive control drugs. Thus, eight groups received the nutritional supplements alone or in combination with each other. Nutritional supplements and glibenclamide were added to the drinking water and neutral protamine Hagedorn insulin was subcutaneously injected during the 4 weeks of treatment period. The induction of type 2 diabetes in the rats caused an infiltration of mononuclear cells and edema in the submucosa of the trachea and lung, severe fibrosis around the vessels and airways, and perivascular and peribronchial infiltration of inflammatory cells and fibrin. In the diabetic group, the total inflammation score and Reid index significantly increased. Diabetes induction significantly reduced the total antioxidant status and elevated the lipid peroxidation products in the serum, lung lavage and lung tissue of the diabetic animals. Treatment with nutritional supplements significantly decreased the histopathological changes and inflammatory indices in the diabetic animals. Supplementation of diabetic rats with leucine, zinc, and chromium, alone and in combination, significantly increased the total antioxidant status and lipid peroxidation level in the diabetic animals. The nutritional supplements improved the enzymatic antioxidant activity of catalase

  7. The Effects of Leucine, Zinc, and Chromium Supplements on Inflammatory Events of the Respiratory System in Type 2 Diabetic Rats

    Science.gov (United States)

    Kolahian, Saeed; Sadri, Hassan; Shahbazfar, Amir Ali; Amani, Morvarid; Mazadeh, Anis; Mirani, Mehdi

    2015-01-01

    Diabetes mellitus is a major cause of serious micro- and macrovascular diseases that affect nearly every system in the body, including the respiratory system. Non-enzymatic protein glycation due to hyperglycaemic stress has fundamental implications due to the large capillary network and amount of connective tissue in the lung. The current study was designed to determine whether leucine, zinc, and chromium supplementations influence the function and histological structure of the respiratory tract in a rat model of type 2 diabetes. Seventy-seven rats were divided into eleven groups, consisting of 7 animals each. One group served as negative control and insulin and glibenclamide were used as positive control drugs. Thus, eight groups received the nutritional supplements alone or in combination with each other. Nutritional supplements and glibenclamide were added to the drinking water and neutral protamine Hagedorn insulin was subcutaneously injected during the 4 weeks of treatment period. The induction of type 2 diabetes in the rats caused an infiltration of mononuclear cells and edema in the submucosa of the trachea and lung, severe fibrosis around the vessels and airways, and perivascular and peribronchial infiltration of inflammatory cells and fibrin. In the diabetic group, the total inflammation score and Reid index significantly increased. Diabetes induction significantly reduced the total antioxidant status and elevated the lipid peroxidation products in the serum, lung lavage and lung tissue of the diabetic animals. Treatment with nutritional supplements significantly decreased the histopathological changes and inflammatory indices in the diabetic animals. Supplementation of diabetic rats with leucine, zinc, and chromium, alone and in combination, significantly increased the total antioxidant status and lipid peroxidation level in the diabetic animals. The nutritional supplements improved the enzymatic antioxidant activity of catalase, glutathione peroxidase

  8. Effects of resource availability and bacterivory on leucine incorporation in different groups of freshwater bacterioplankton, assessed using microautoradiography

    Czech Academy of Sciences Publication Activity Database

    Horňák, Karel; Jezbera, Jan; Nedoma, Jiří; Gasol, J.M.; Šimek, Karel

    2006-01-01

    Roč. 45, č. 3 (2006), s. 277-289 ISSN 0948-3055 R&D Projects: GA ČR GA206/05/0007 Grant - others:FRVŠ(CZ) 1062/2004 Institutional research plan: CEZ:AV0Z60170517 Keywords : leucin e incorporation * bacterial structure * bacterial function Subject RIV: EH - Ecology, Behaviour Impact factor: 2.209, year: 2006

  9. Pulsatile delivery of a leucine supplement during long-term continuous enteral feeding enhances lean growth in term neonatal pigs

    Science.gov (United States)

    Neonatal pigs are used as a model to study and optimize the clinical treatment of infants who are unable to maintain oral feeding. Using this model, we have previously shown that pulsatile administration of leucine during continuous feeding over 24 h via orogastric tube enhanced protein synthesis in...

  10. Sample Dilution and Bacterial Community Composition Influence Empirical Leucine-to-Carbon Conversion Factors in Surface Waters of the World's Oceans

    KAUST Repository

    Teira, Eva; Hernando-Morales, Ví ctor; Cornejo-Castillo, Francisco M.; Alonso-Sá ez, Laura; Sarmento, Hugo; Valencia-Vila, Joaquí n; Serrano Catalá , Teresa; Herná ndez-Ruiz, Marta; Varela, Marta M.; Ferrera, Isabel; Moran, Xose Anxelu G.; Gasol, Josep M.

    2015-01-01

    The transformation of leucine incorporation rates to prokaryotic carbon production rates requires the use of either theoretical or empirically determined conversion factors. Empirical leucine-to-carbon conversion factors (eCFs) vary widely across environments, and little is known about their potential controlling factors. We conducted 10 surface seawater manipulation experiments across the world's oceans, where the growth of the natural prokaryotic assemblages was promoted by filtration (i.e., removal of grazers [F treatment]) or filtration combined with dilution (i.e., also relieving resource competition [FD treatment]). The impact of sunlight exposure was also evaluated in the FD treatments, and we did not find a significant effect on the eCFs. The eCFs varied from 0.09 to 1.47 kg C mol Leu−1 and were significantly lower in the FD than in the F samples. Also, changes in bacterial community composition during the incubations, as assessed by automated ribosomal intergenic spacer analysis (ARISA), were more pronounced in the FD than in the F treatments, compared to unmanipulated controls. Thus, we discourage the common procedure of diluting samples (in addition to filtration) for eCF determination. The eCFs in the filtered treatment were negatively correlated with the initial chlorophyll a concentration, picocyanobacterial abundance (mostly Prochlorococcus), and the percentage of heterotrophic prokaryotes with high nucleic acid content (%HNA). The latter two variables explained 80% of the eCF variability in the F treatment, supporting the view that both Prochlorococcus and HNA prokaryotes incorporate leucine in substantial amounts, although this results in relatively low carbon production rates in the oligotrophic ocean.

  11. Sample Dilution and Bacterial Community Composition Influence Empirical Leucine-to-Carbon Conversion Factors in Surface Waters of the World's Oceans

    KAUST Repository

    Teira, Eva

    2015-09-25

    The transformation of leucine incorporation rates to prokaryotic carbon production rates requires the use of either theoretical or empirically determined conversion factors. Empirical leucine-to-carbon conversion factors (eCFs) vary widely across environments, and little is known about their potential controlling factors. We conducted 10 surface seawater manipulation experiments across the world\\'s oceans, where the growth of the natural prokaryotic assemblages was promoted by filtration (i.e., removal of grazers [F treatment]) or filtration combined with dilution (i.e., also relieving resource competition [FD treatment]). The impact of sunlight exposure was also evaluated in the FD treatments, and we did not find a significant effect on the eCFs. The eCFs varied from 0.09 to 1.47 kg C mol Leu−1 and were significantly lower in the FD than in the F samples. Also, changes in bacterial community composition during the incubations, as assessed by automated ribosomal intergenic spacer analysis (ARISA), were more pronounced in the FD than in the F treatments, compared to unmanipulated controls. Thus, we discourage the common procedure of diluting samples (in addition to filtration) for eCF determination. The eCFs in the filtered treatment were negatively correlated with the initial chlorophyll a concentration, picocyanobacterial abundance (mostly Prochlorococcus), and the percentage of heterotrophic prokaryotes with high nucleic acid content (%HNA). The latter two variables explained 80% of the eCF variability in the F treatment, supporting the view that both Prochlorococcus and HNA prokaryotes incorporate leucine in substantial amounts, although this results in relatively low carbon production rates in the oligotrophic ocean.

  12. An innovative approach to enhance methane hydrate formation kinetics with leucine for energy storage application

    International Nuclear Information System (INIS)

    Veluswamy, Hari Prakash; Kumar, Asheesh; Kumar, Rajnish; Linga, Praveen

    2017-01-01

    Highlights: • Innovative combinatorial hybrid approach to reduce nucleation stochasticity and enhance hydrate growth. • Methane hydrate growth curves are similar in UTR and STR configurations in presence of leucine. • Amalgamation of stirred (STR) and unstirred (UTR) configuration is demonstrated. • Reliable method for scale up and commercial production of Solidified Natural Gas (SNG). - Abstract: Natural gas storage in clathrate hydrates or solidified natural gas (SNG) offers the safest, cleanest and the most compact mode of storage aided by the relative ease in natural gas (NG) recovery with minimal cost compared to known conventional methods of NG storage. The stochastic nature of hydrate nucleation and the slow kinetics of hydrate growth are major challenges that needs to be addressed on the SNG production side. A deterministic and fast nucleation coupled with rapid crystallization kinetics would empower this beneficial technology for commercial application. We propose a hybrid combinatorial approach of methane hydrate formation utilizing the beneficial aspect of environmentally benign amino acid (leucine) as a kinetic promoter by combining stirred and unstirred reactor operation. This hybrid approach is simple, can easily be implemented and scaled-up to develop an economical SNG technology for efficient storage of natural gas on a large scale. Added benefits include the minimal energy requirement during hydrate growth resulting in overall cost reduction for SNG technology.

  13. Solvation thermodynamics of L-cystine, L-tyrosine, and L-leucine in aqueous-electrolyte media

    Science.gov (United States)

    Roy, Sanjay; Guin, Partha Sarathi; Mahali, Kalachand; Dolui, Bijoy Krishna

    2017-12-01

    Solubilities of L-cystine, L-tyrosine, and L-leucine in aqueous NaCl media at 298.15 K have been studied. Indispensable and related solvent parameters such as molar mass, molar volume, etc., were also determined. The results are used to evaluate the standard transfer Gibbs free energy, cavity forming enthalpy of transfer, cavity forming transfer Gibbs free energy and dipole-dipole interaction effects during the course of solvation. Various weak interactions involving solute-solvent or solvent-solvent molecules were characterized in order to find their role on the solvation of these amino acids.

  14. A conserved gene family encodes transmembrane proteins with fibronectin, immunoglobulin and leucine-rich repeat domains (FIGLER

    Directory of Open Access Journals (Sweden)

    Haga Christopher L

    2007-09-01

    Full Text Available Abstract Background In mouse the cytokine interleukin-7 (IL-7 is required for generation of B lymphocytes, but human IL-7 does not appear to have this function. A bioinformatics approach was therefore used to identify IL-7 receptor related genes in the hope of identifying the elusive human cytokine. Results Our database search identified a family of nine gene candidates, which we have provisionally named fibronectin immunoglobulin leucine-rich repeat (FIGLER. The FIGLER 1–9 genes are predicted to encode type I transmembrane glycoproteins with 6–12 leucine-rich repeats (LRR, a C2 type Ig domain, a fibronectin type III domain, a hydrophobic transmembrane domain, and a cytoplasmic domain containing one to four tyrosine residues. Members of this multichromosomal gene family possess 20–47% overall amino acid identity and are differentially expressed in cell lines and primary hematopoietic lineage cells. Genes for FIGLER homologs were identified in macaque, orangutan, chimpanzee, mouse, rat, dog, chicken, toad, and puffer fish databases. The non-human FIGLER homologs share 38–99% overall amino acid identity with their human counterpart. Conclusion The extracellular domain structure and absence of recognizable cytoplasmic signaling motifs in members of the highly conserved FIGLER gene family suggest a trophic or cell adhesion function for these molecules.

  15. Temperature-driven adaptation of the bacterial community in peat measured by using thymidine and leucine incorporation

    OpenAIRE

    Ranneklev, Sissel Brit; Bååth, Erland

    2001-01-01

    The temperature-driven adaptation of the bacterial community in peat was studied, by altering temperature to simulate self-heating and a subsequent return to mesophilic conditions. The technique used consisted of extracting the bacterial community from peat using homogenization-centrifugation and measuring the rates of thymidine (TdR) or leucine (Leu) incorporation by the extracted bacterial community at different temperatures. Increasing the peat incubation temperature from 25°C to 35, 45, o...

  16. Origin and diversification of leucine-rich repeat receptor-like protein kinase (LRR-RLK) genes in plants

    OpenAIRE

    Liu, Ping-Li; Du, Liang; Huang, Yuan; Gao, Shu-Min; Yu, Meng

    2017-01-01

    Background Leucine-rich repeat receptor-like protein kinases (LRR-RLKs) are the largest group of receptor-like kinases in plants and play crucial roles in development and stress responses. The evolutionary relationships among LRR-RLK genes have been investigated in flowering plants; however, no comprehensive studies have been performed for these genes in more ancestral groups. The subfamily classification of LRR-RLK genes in plants, the evolutionary history and driving force for the evolution...

  17. Copolymers based on N-acryloyl-L-leucine and urea methacrylate with pyridine moieties

    Directory of Open Access Journals (Sweden)

    Buruiana Emil C.

    2016-01-01

    Full Text Available By using free radical polymerization of (N-methacryloyloxyethyl-N′-4-picolyl-urea (MAcPU and N-acryloyl-L-leucine (AcLeu, an optically active copolymer, poly[(N-methacryloyloxyethyl-N′-4-picolyl-urea-co-N-acryloyl-L-leucine], MAcPU-co-AcLeu (1.86:1 molar ratio was prepared and subsequently functionalized at the pyridine-N with (1R/S-(−/+-10-camphorsulfonic acid (R/S-CSA and at carboxyl group with (R-(+-α-ethylbenzylamine (R-EBA or trans-4-stilbene methanol (t-StM. The structures, chemical composition and chiroptical activity of the monomers and the copolymers were characterized by spectral analysis (FTIR, 1H (13C-NMR, 1H,1H-COSY, UV/vis, thermal methods (TGA, DSC, fluorescence spectroscopy, gel permeation chromatography and specific rotation measurements. Influence of the optical activity of monomer and modifier on modified copolymers suggested a good correlation between the experimental data obtained (23[α]589=+12.5° for AcLeu and MAcPU-co-AcLeu, 23[α]589=0°+27.5° for (MAcPU-co-AcLeu-R/S-CSA, 23[α]589=+25° for (MAcPU-co-AcLeu-R-EBA, and 23[α]589 = 0° for (MAcPU-co-AcLeu-St. In addition, the photobehavior of the stilbene copolymer (MAcPU-co-AcLeu-St in film was investigated by UV-vis spectroscopy. The fluorescence quenching of the stilbene species in the presence of aliphatic/aromatic amine in DMF solution was evaluated, more efficiently being 4,4′−dipyridyl (detection limit: 7.2 x 10-6 mol/L.

  18. Estimation of whole body protein turnover by L-[1-[sup 13]C]-leucine tracer experiment in post-viral hepatocirrhotic patients

    Energy Technology Data Exchange (ETDEWEB)

    Xinhua, Zhu; Zhenduo, Tang; Tengchang, Dai; Zongqin, Xia [Shanghai Second Medical Univ. (China)

    1993-08-01

    A constant infusion of L-[1-[sup 1]'3C]-leucine given to the subject and the plasma [sup 13]C-leucine enrichment and breath [sup 1]'3CO[sub 2] enrichment were measured with GC-MS and gas isotope ratio MS respectively. The plateau isotope enrichments reached were used to calculate the whole body protein synthesis and breakdown rates using a two pools model. 15 post-viral hepatocirrhotic patients and 6 normal adults were studied. The whole body protein synthesis and breakdown rates were both markedly increased in cirrhotic patients, with the breakdown rate higher than the synthesis rate and hence leading to a negative nitrogen balance. The changes of 7 decompensated cirrhotic patients were more severe than 8 compensated cirrhotic patients. The results suggest that the protein malnutrition and negative nitrogen balance of post-viral hepatocirrhotic patients are not simply due to low intake of protein and that the hyperactive turnover, especially the excessive breakdown of body protein might play an important role.

  19. Whey and casein labelled with L-[1-13C]-leucine and muscle protein synthesis: effect of resistance exercise and protein ingestion

    DEFF Research Database (Denmark)

    Reitelseder, Søren; Agergaard, Jakob; Doessing, Simon

    2011-01-01

    to a single bolus intake of whey or casein after performance of heavy resistance exercise. Young male individuals were randomly assigned to participate in two protein trials (n = 9) or one control trial (n = 8). Infusion of l-[1-(13)C]leucine was carried out, and either whey, casein (0.3 g/kg lean body mass......), or a noncaloric control drink was ingested immediately after exercise. l-[1-(13)C]leucine-labeled whey and casein were used while muscle protein synthesis (MPS) was assessed. Blood and muscle tissue samples were collected to measure systemic hormone and amino acid concentrations, tracer enrichments......, and myofibrillar protein synthesis. Western blots were used to investigate the Akt signaling pathway. Plasma insulin and branched-chain amino acid concentrations increased to a greater extent after ingestion of whey compared with casein. Myofibrillar protein synthesis was equally increased 1-6 h postexercise after...

  20. Sample dilution and bacterial community composition influence empirical leucine-to-carbon conversion factors in surface waters of the world's oceans.

    Science.gov (United States)

    Teira, Eva; Hernando-Morales, Víctor; Cornejo-Castillo, Francisco M; Alonso-Sáez, Laura; Sarmento, Hugo; Valencia-Vila, Joaquín; Serrano Catalá, Teresa; Hernández-Ruiz, Marta; Varela, Marta M; Ferrera, Isabel; Gutiérrez Morán, Xosé Anxelu; Gasol, Josep M

    2015-12-01

    The transformation of leucine incorporation rates to prokaryotic carbon production rates requires the use of either theoretical or empirically determined conversion factors. Empirical leucine-to-carbon conversion factors (eCFs) vary widely across environments, and little is known about their potential controlling factors. We conducted 10 surface seawater manipulation experiments across the world's oceans, where the growth of the natural prokaryotic assemblages was promoted by filtration (i.e., removal of grazers [F treatment]) or filtration combined with dilution (i.e., also relieving resource competition [FD treatment]). The impact of sunlight exposure was also evaluated in the FD treatments, and we did not find a significant effect on the eCFs. The eCFs varied from 0.09 to 1.47 kg C mol Leu(-1) and were significantly lower in the FD than in the F samples. Also, changes in bacterial community composition during the incubations, as assessed by automated ribosomal intergenic spacer analysis (ARISA), were more pronounced in the FD than in the F treatments, compared to unmanipulated controls. Thus, we discourage the common procedure of diluting samples (in addition to filtration) for eCF determination. The eCFs in the filtered treatment were negatively correlated with the initial chlorophyll a concentration, picocyanobacterial abundance (mostly Prochlorococcus), and the percentage of heterotrophic prokaryotes with high nucleic acid content (%HNA). The latter two variables explained 80% of the eCF variability in the F treatment, supporting the view that both Prochlorococcus and HNA prokaryotes incorporate leucine in substantial amounts, although this results in relatively low carbon production rates in the oligotrophic ocean. Copyright © 2015, American Society for Microbiology. All Rights Reserved.

  1. Medium-Chain Triglycerides in Combination with Leucine and Vitamin D Benefit Cognition in Frail Elderly Adults: A Randomized Controlled Trial.

    Science.gov (United States)

    Abe, Sakiko; Ezaki, Osamu; Suzuki, Motohisa

    2017-01-01

    The combined supplementation of medium-chain triglycerides (MCTs), L-leucine-rich amino acids, and cholecalciferol (vitamin D 3 ) increase muscle strength and function in frail elderly individuals. However, their effects on cognition are unknown. We enrolled 38 elderly nursing home residents (mean age±SD, 86.6±4.8 y) in a 3-mo randomized, controlled, parallel group trial. The participants were randomly allocated to 3 groups: the first group received a L-leucine (1.2 g)- and cholecalciferol (20 μg)-enriched supplement with 6 g of MCT (LD+MCT); the second group received the same supplement with 6 g of long-chain triglycerides (LD+LCT); and the third group did not receive any supplements (control). Cognition was assessed at baseline and after the 3-mo intervention. The difference in changes among the groups was assessed with ANCOVA, adjusting for age and the baseline value as covariates. After 3 mo, the Mini-Mental State Examination (MMSE) score in the LD+MCT group increased by 10.6% (from 16.6 to 18.4 points, p<0.05). After 3 mo, the Nishimura geriatric rating scale for mental status (NM scale) score in the LD+MCT group increased by 30.6% (from 24.6 to 32.2 points, p<0.001), whereas that in the LD+LCT and control groups decreased by 11.2% (from 31.2 to 27.7 points, p<0.05) and 26.1% (from 27.2 to 20.1 points, p<0.001), respectively. The combined supplementation of MCTs (6 g), L-leucine-rich amino acids, and cholecalciferol may improve cognitive function in frail elderly individuals.

  2. Zipper-like magnetic molecularly imprinted microspheres for on/off-switchable recognition and extraction of 17β-estradiol from food samples.

    Science.gov (United States)

    Zhu, Wenting; Peng, Hailong; Luo, Mei; Yu, Ningxiang; Xiong, Hua; Wang, Ronghui; Li, Yanbin

    2018-09-30

    Zipper-like on/off-switchable and magnetic molecularly imprinted microspheres (SM-MIMs) were constructed using acrylamide (AAm) and 2-acrylamide-2-methyl propanesulfonic acid (AMPS) as functional monomers for 17β-estradiol (17β-E 2 ) recognition and extraction. The imprinted polymer interactions between poly(AAm) (PAAm) and poly(AMPS) (PAMPS) with on/off-switchable property to temperature, exhibited dissociation at relatively higher temperatures (such as 30 °C) and helped 17β-E 2 enter into imprinted sites, leading to higher binding capability. Conversely, the interpolymer complexes between PAAm and PAMPS formed and blocked 17β-E 2 access to imprinted sites at lower temperature (such as 20 °C). SM-MIMs were used as dispersive solid phase extraction (SPE) adsorbent with HPLC for 17β-E 2 pretreatment and detection in food samples, and low limit detection (2.52 µg L -1 ) and quantification (10.76 µg L -1 ) with higher recovery were obtained. Therefore, SM-MIMs may be a promising adsorbent for 17β-E 2 pretreatment in food samples owing to its advantages of on/off-switchable recognition, eco-friendly elution, and efficient separation. Copyright © 2018. Published by Elsevier Ltd.

  3. Adding Fish Oil to Whey Protein, Leucine, and Carbohydrate Over a Six-Week Supplementation Period Attenuates Muscle Soreness Following Eccentric Exercise in Competitive Soccer Players.

    Science.gov (United States)

    Philpott, Jordan D; Donnelly, Chris; Walshe, Ian H; MacKinley, Elizabeth E; Dick, James; Galloway, Stuart D R; Tipton, Kevin D; Witard, Oliver C

    2018-01-01

    Soccer players often experience eccentric exercise-induced muscle damage given the physical demands of soccer match-play. Since long chain n-3 polyunsaturated fatty acids (n-3PUFA) enhance muscle sensitivity to protein supplementation, dietary supplementation with a combination of fish oil-derived n-3PUFA, protein, and carbohydrate may promote exercise recovery. This study examined the influence of adding n-3PUFA to a whey protein, leucine, and carbohydrate containing beverage over a six-week supplementation period on physiological markers of recovery measured over three days following eccentric exercise. Competitive soccer players were assigned to one of three conditions (2 × 200 mL): a fish oil supplement beverage (FO; n = 10) that contained n-3PUFA (1100 mg DHA/EPA-approximately 550 mg DHA, 550 mg EPA), whey protein (15 g), leucine (1.8 g), and carbohydrate (20 g); a protein supplement beverage (PRO; n = 10) that contained whey protein (15 g), leucine (1.8 g), and carbohydrate (20 g); and a carbohydrate supplement beverage (CHO; n = 10) that contained carbohydrate (24 g). Eccentric exercise consisted of unilateral knee extension/flexion contractions on both legs separately. Maximal force production was impaired by 22% during the 72-hour recovery period following eccentric exercise (p recovery, was less in FO (1948 ± 1091 mm × 72 h) than PRO (4640 ± 2654 mm × 72 h, p soccer performance, or blood c-reactive protein concentrations were observed between groups. In conclusion, the addition of n-3PUFA to a beverage containing whey protein, leucine, and carbohydrate ameliorates the increase in muscle soreness and blood concentrations of creatine kinase following eccentric exercise in competitive soccer players.

  4. Dendrite Injury Triggers DLK-Independent Regeneration

    Directory of Open Access Journals (Sweden)

    Michelle C. Stone

    2014-01-01

    Full Text Available Axon injury triggers regeneration through activation of a conserved kinase cascade, which includes the dual leucine zipper kinase (DLK. Although dendrites are damaged during stroke, traumatic brain injury, and seizure, it is not known whether mature neurons monitor dendrite injury and initiate regeneration. We probed the response to dendrite damage using model Drosophila neurons. Two larval neuron types regrew dendrites in distinct ways after all dendrites were removed. Dendrite regeneration was also triggered by injury in adults. Next, we tested whether dendrite injury was initiated with the same machinery as axon injury. Surprisingly, DLK, JNK, and fos were dispensable for dendrite regeneration. Moreover, this MAP kinase pathway was not activated by injury to dendrites. Thus, neurons respond to dendrite damage and initiate regeneration without using the conserved DLK cascade that triggers axon regeneration.

  5. Polymorphisms in leucine-rich repeat genes are associated with autism spectrum disorder susceptibility in populations of European ancestry

    Directory of Open Access Journals (Sweden)

    Sousa Inês

    2010-03-01

    Full Text Available Abstract Background Autism spectrum disorders (ASDs are a group of highly heritable neurodevelopmental disorders which are characteristically comprised of impairments in social interaction, communication and restricted interests/behaviours. Several cell adhesion transmembrane leucine-rich repeat (LRR proteins are highly expressed in the nervous system and are thought to be key regulators of its development. Here we present an association study analysing the roles of four promising candidate genes - LRRTM1 (2p, LRRTM3 (10q, LRRN1 (3p and LRRN3 (7q - in order to identify common genetic risk factors underlying ASDs. Methods In order to gain a better understanding of how the genetic variation within these four gene regions may influence susceptibility to ASDs, a family-based association study was undertaken in 661 families of European ancestry selected from four different ASD cohorts. In addition, a case-control study was undertaken across the four LRR genes, using logistic regression in probands with ASD of each population against 295 ECACC controls. Results Significant results were found for LRRN3 and LRRTM3 (P LRRTM3. Conclusions Overall, our findings implicate the neuronal leucine-rich genes LRRN3 and LRRTM3 in ASD susceptibility.

  6. Advances in basic and clinical immunology in 2016.

    Science.gov (United States)

    Chinen, Javier; Badran, Yousef R; Geha, Raif S; Chou, Janet S; Fried, Ari J

    2017-10-01

    Advances in basic immunology in 2016 included studies that further characterized the role of different proteins in the differentiation of effector T and B cells, including cytokines and proteins involved in the actin cytoskeleton. Regulation of granule formation and secretion in cytotoxic cells was also further described by examining patients with familial hemophagocytic lymphohistiocytosis. The role of prenylation in patients with mevalonate kinase deficiency leading to inflammation has been established. We reviewed advances in clinical immunology, as well as new approaches of whole-genome sequencing and genes newly reported to be associated with immunodeficiency, such as linker of activation of T cells (LAT); B-cell CLL/lymphoma 11B (BCL11B); RGD, leucine-rich repeat, tropomodulin domain, and proline-rich domain-containing protein (RLTPR); moesin; and Janus kinase 1 (JAK1). Trials of hematopoietic stem cell transplantation and gene therapy for primary immunodeficiency have had relative success; the use of autologous virus-specific cytotoxic T cells has proved effective as well. New medications are being explored, such as pioglitazone, which is under study for its role in enhancing the oxidative burst in patients with chronic granulomatous disease. Development of vaccines for HIV infection continues to provide insight into the immune response against a virus with an extraordinary mutation rate. Copyright © 2017 American Academy of Allergy, Asthma & Immunology. Published by Elsevier Inc. All rights reserved.

  7. Bistable forespore engulfment in Bacillus subtilis by a zipper mechanism in absence of the cell wall.

    Directory of Open Access Journals (Sweden)

    Nikola Ojkic

    2014-10-01

    Full Text Available To survive starvation, the bacterium Bacillus subtilis forms durable spores. The initial step of sporulation is asymmetric cell division, leading to a large mother-cell and a small forespore compartment. After division is completed and the dividing septum is thinned, the mother cell engulfs the forespore in a slow process based on cell-wall degradation and synthesis. However, recently a new cell-wall independent mechanism was shown to significantly contribute, which can even lead to fast engulfment in [Formula: see text] 60 [Formula: see text] of the cases when the cell wall is completely removed. In this backup mechanism, strong ligand-receptor binding between mother-cell protein SpoIIIAH and forespore-protein SpoIIQ leads to zipper-like engulfment, but quantitative understanding is missing. In our work, we combined fluorescence image analysis and stochastic Langevin simulations of the fluctuating membrane to investigate the origin of fast bistable engulfment in absence of the cell wall. Our cell morphologies compare favorably with experimental time-lapse microscopy, with engulfment sensitive to the number of SpoIIQ-SpoIIIAH bonds in a threshold-like manner. By systematic exploration of model parameters, we predict regions of osmotic pressure and membrane-surface tension that produce successful engulfment. Indeed, decreasing the medium osmolarity in experiments prevents engulfment in line with our predictions. Forespore engulfment may thus not only be an ideal model system to study decision-making in single cells, but its biophysical principles are likely applicable to engulfment in other cell types, e.g. during phagocytosis in eukaryotes.

  8. An oncogenic axis of STAT-mediated BATF3 upregulation causing MYC activity in classical Hodgkin lymphoma and anaplastic large cell lymphoma.

    Science.gov (United States)

    Lollies, A; Hartmann, S; Schneider, M; Bracht, T; Weiß, A L; Arnolds, J; Klein-Hitpass, L; Sitek, B; Hansmann, M-L; Küppers, R; Weniger, M A

    2018-01-01

    Classical Hodgkin lymphoma (cHL) and anaplastic large cell lymphoma (ALCL) feature high expression of activator protein-1 (AP-1) transcription factors, which regulate various physiological processes but also promote lymphomagenesis. The AP-1 factor basic leucine zipper transcription factor, ATF-like 3 (BATF3), is highly transcribed in cHL and ALCL; however, its functional importance in lymphomagenesis is unknown. Here we show that proto-typical CD30 + lymphomas, namely cHL (21/30) and primary mediastinal B-cell lymphoma (8/9), but also CD30 + diffuse large B-cell lymphoma (15/20) frequently express BATF3 protein. Mass spectrometry and co-immunoprecipitation established interactions of BATF3 with JUN and JUNB in cHL and ALCL lines. BATF3 knockdown using short hairpin RNAs was toxic for cHL and ALCL lines, reducing their proliferation and survival. We identified MYC as a critical BATF3 target and confirmed binding of BATF3 to the MYC promoter. JAK/STAT signaling regulated BATF3 expression, as chemical JAK2 inhibition reduced and interleukin 13 stimulation induced BATF3 expression in cHL lines. Chromatin immunoprecipitation substantiated a direct regulation of BATF3 by STAT proteins in cHL and ALCL lines. In conclusion, we identified STAT-mediated BATF3 expression that is essential for lymphoma cell survival and promoted MYC activity in cHL and ALCL, hence we recognized a new oncogenic axis in these lymphomas.

  9. Exploring the bZIP transcription factor regulatory network in Neurospora crassa.

    Science.gov (United States)

    Tian, Chaoguang; Li, Jingyi; Glass, N Louise

    2011-03-01

    Transcription factors (TFs) are key nodes of regulatory networks in eukaryotic organisms, including filamentous fungi such as Neurospora crassa. The 178 predicted DNA-binding TFs in N. crassa are distributed primarily among six gene families, which represent an ancient expansion in filamentous ascomycete genomes; 98 TF genes show detectable expression levels during vegetative growth of N. crassa, including 35 that show a significant difference in expression level between hyphae at the periphery versus hyphae in the interior of a colony. Regulatory networks within a species genome include paralogous TFs and their respective target genes (TF regulon). To investigate TF network evolution in N. crassa, we focused on the basic leucine zipper (bZIP) TF family, which contains nine members. We performed baseline transcriptional profiling during vegetative growth of the wild-type and seven isogenic, viable bZIP deletion mutants. We further characterized the regulatory network of one member of the bZIP family, NCU03905. NCU03905 encodes an Ap1-like protein (NcAp-1), which is involved in resistance to multiple stress responses, including oxidative and heavy metal stress. Relocalization of NcAp-1 from the cytoplasm to the nucleus was associated with exposure to stress. A comparison of the NcAp-1 regulon with Ap1-like regulons in Saccharomyces cerevisiae, Schizosaccharomyces pombe, Candida albicans and Aspergillus fumigatus showed both conservation and divergence. These data indicate how N. crassa responds to stress and provide information on pathway evolution.

  10. DNA Methylation Influences Chlorogenic Acid Biosynthesis in Lonicera japonica by Mediating LjbZIP8 to Regulate Phenylalanine Ammonia-Lyase 2 Expression

    Directory of Open Access Journals (Sweden)

    Liangping Zha

    2017-07-01

    Full Text Available The content of active compounds differ in buds and flowers of Lonicera japonica (FLJ and L. japonica var. chinensis (rFLJ. Chlorogenic acid (CGAs were major active compounds of L. japonica and regarded as measurements for quality evaluation. However, little is known concerning the formation of active compounds at the molecular level. We quantified the major CGAs in FLJ and rFLJ, and found the concentrations of CGAs were higher in the buds of rFLJ than those of FLJ. Further analysis of CpG methylation of CGAs biosynthesis genes showed differences between FLJ and rFLJ in the 5′-UTR of phenylalanine ammonia-lyase 2 (PAL2. We identified 11 LjbZIP proteins and 24 rLjbZIP proteins with conserved basic leucine zipper domains, subcellular localization, and electrophoretic mobility shift assay showed that the transcription factor LjbZIP8 is a nuclear-localized protein that specifically binds to the G-box element of the LjPAL2 5′-UTR. Additionally, a transactivation assay and LjbZIP8 overexpression in transgenic tobacco indicated that LjbZIP8 could function as a repressor of transcription. Finally, treatment with 5-azacytidine decreased the transcription level of LjPAL2 and CGAs content in FLJ leaves. These results raise the possibility that DNA methylation might influence the recruitment of LjbZIP8, regulating PAL2 expression level and CGAs content in L. japonica.

  11. Modulation of organic acids and sugar content in tomato fruits by an abscisic acid-regulated transcription factor.

    Science.gov (United States)

    Bastías, Adriana; López-Climent, María; Valcárcel, Mercedes; Rosello, Salvador; Gómez-Cadenas, Aurelio; Casaretto, José A

    2011-03-01

    Growing evidence suggests that the phytohormone abscisic acid (ABA) plays a role in fruit development. ABA signaling components of developmental programs and responses to stress conditions include the group of basic leucine zipper transcriptional activators known as ABA-response element binding factors (AREBs/ABFs). AREB transcription factors mediate ABA-regulated gene expression involved in desiccation tolerance and are expressed mainly in seeds and in vegetative tissues under stress; however, they are also expressed in some fruits such as tomato. In order to get an insight into the role of ABA signaling in fruit development, the expression of two AREB-like factors were investigated during different developmental stages. In addition, tomato transgenic lines that overexpress and downregulate one AREB-like transcription factor, SlAREB1, were used to determine its effect on the levels of some metabolites determining fruit quality. Higher levels of citric acid, malic acid, glutamic acid, glucose and fructose were observed in SlAREB1-overexpressing lines compared with those in antisense suppression lines in red mature fruit pericarp. The higher hexose concentration correlated with increased expression of genes encoding a vacuolar invertase (EC 3.2.1.26) and a sucrose synthase (EC 2.4.1.13). No significant changes were found in ethylene content which agrees with the normal ripening phenotype observed in transgenic fruits. These results suggest that an AREB-mediated ABA signal affects the metabolism of these compounds during the fruit developmental program. Copyright © Physiologia Plantarum 2010.

  12. A bHLH gene from Tamarix hispida improves abiotic stress tolerance by enhancing osmotic potential and decreasing reactive oxygen species accumulation.

    Science.gov (United States)

    Ji, Xiaoyu; Nie, Xianguang; Liu, Yujia; Zheng, Lei; Zhao, Huimin; Zhang, Bing; Huo, Lin; Wang, Yucheng

    2016-02-01

    Basic helix-loop-helix (bHLH) leucine-zipper transcription factors play important roles in abiotic stress responses. However, their specific roles in abiotic stress tolerance are not fully known. Here, we functionally characterized a bHLH gene, ThbHLH1, from Tamarix hispida in abiotic stress tolerance. ThbHLH1 specifically binds to G-box motif with the sequence of 'CACGTG'. Transiently transfected T. hispida plantlets with transiently overexpressed ThbHLH1 and RNAi-silenced ThbHLH1 were generated for gain- and loss-of-function analysis. Transgenic Arabidopsis thaliana lines overexpressing ThbHLH1 were generated to confirm the gain- and loss-of-function analysis. Overexpression of ThbHLH1 significantly elevates glycine betaine and proline levels, increases Ca(2+) concentration and enhances peroxidase (POD) and superoxide dismutase (SOD) activities to decrease reactive oxygen species (ROS) accumulation. Additionally, ThbHLH1 regulates the expression of the genes including P5CS, BADH, CaM, POD and SOD, to activate the above physiological changes, and also induces the expression of stress tolerance-related genes LEAs and HSPs. These data suggest that ThbHLH1 induces the expression of stress tolerance-related genes to improve abiotic stress tolerance by increasing osmotic potential, improving ROS scavenging capability and enhancing second messenger in stress signaling cascades. © The Author 2016. Published by Oxford University Press. All rights reserved. For Permissions, please email: journals.permissions@oup.com.

  13. A p300 and SIRT1 Regulated Acetylation Switch of C/EBPα Controls Mitochondrial Function

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    Mohamad A. Zaini

    2018-01-01

    Full Text Available Summary: Cellular metabolism is a tightly controlled process in which the cell adapts fluxes through metabolic pathways in response to changes in nutrient supply. Among the transcription factors that regulate gene expression and thereby cause changes in cellular metabolism is the basic leucine-zipper (bZIP transcription factor CCAAT/enhancer-binding protein alpha (C/EBPα. Protein lysine acetylation is a key post-translational modification (PTM that integrates cellular metabolic cues with other physiological processes. Here, we show that C/EBPα is acetylated by the lysine acetyl transferase (KAT p300 and deacetylated by the lysine deacetylase (KDAC sirtuin1 (SIRT1. SIRT1 is activated in times of energy demand by high levels of nicotinamide adenine dinucleotide (NAD+ and controls mitochondrial biogenesis and function. A hypoacetylated mutant of C/EBPα induces the transcription of mitochondrial genes and results in increased mitochondrial respiration. Our study identifies C/EBPα as a key mediator of SIRT1-controlled adaption of energy homeostasis to changes in nutrient supply. : Zaini et al. show that the transcription factor C/EBPα is acetylated by p300 and deacetylated by the lysine deacetylase SIRT1. Hypoacetylated C/EBPα induces the transcription of mitochondrial genes and results in increased mitochondrial respiration. C/EBPα is a key mediator of SIRT1-controlled adaption of energy homeostasis to changes in nutrient supply. Keywords: C/EBPα, SIRT1, p300, lysine acetylation, mitochondrial function, cellular metabolism, NAD+, gene regulation

  14. Knockdown of XBP1 by RNAi in Mouse Granulosa Cells Promotes Apoptosis, Inhibits Cell Cycle, and Decreases Estradiol Synthesis

    Directory of Open Access Journals (Sweden)

    Nan Wang

    2017-05-01

    Full Text Available Granulosa cells are crucial for follicular growth, development, and follicular atresia. X-box binding protein 1 (XBP1, a basic region-leucine zipper protein, is widely involved in cell differentiation, proliferation, apoptosis, cellular stress response, and other signaling pathways. In this study, RNA interference, flow cytometry, western blot, real-time PCR, Cell Counting Kit (CCK8, and ELISA were used to investigate the effect of XBP1 on steroidogenesis, apoptosis, cell cycle, and proliferation of mouse granulosa cells. ELISA analysis showed that XBP1 depletion significantly decreased the concentrations of estradiol (E2. Additionally, the expression of estrogen synthesis enzyme Cyp19a1 was sharply downregulated. Moreover, flow cytometry showed that knockdown of XBP1 increased the apoptosis rate and arrests the cell cycle in S-phase in granulosa cells (GCs. Further study confirmed these results. The expression of CCAAT-enhancer-binding protein homologous protein (CHOP, cysteinyl aspartate specific proteases-3 (caspase-3, cleaved caspase-3, and Cyclin E was upregulated, while that of Bcl-2, Cyclin A1, and Cyclin B1 was downregulated. Simultaneously, CCK8 analysis indicated that XBP1 disruption inhibited cell proliferation. In addition, XBP1 knockdown also alters the expression of Has2 and Ptgs2, two essential genes for folliculogenesis. Collectively, these data reveal a novel critical role of XBP1 in folliculogenesis by regulating the cell cycle, apoptosis, and steroid synthesis of mouse granulosa cells.

  15. Glycogen synthase kinase 3 regulates expression of nuclear factor-erythroid-2 related transcription factor-1 (Nrf1) and inhibits pro-survival function of Nrf1

    Energy Technology Data Exchange (ETDEWEB)

    Biswas, Madhurima; Kwong, Erick K.; Park, Eujean; Nagra, Parminder; Chan, Jefferson Y., E-mail: jchan@uci.edu

    2013-08-01

    Nuclear factor E2-related factor-1 (Nrf1) is a basic leucine zipper transcription factor that is known to regulate antioxidant and cytoprotective gene expression. It was recently shown that Nrf1 is regulated by SCF–Fbw7 ubiquitin ligase. However our knowledge of upstream signals that targets Nrf1 for degradation by the UPS is not known. We report here that Nrf1 expression is negatively regulated by glycogen synthase kinase 3 (GSK3) in Fbw7-dependent manner. We show that GSK3 interacts with Nrf1 and phosphorylates the Cdc4 phosphodegron domain (CPD) in Nrf1. Mutation of serine residue in the CPD of Nrf1 to alanine (S350A), blocks Nrf1 from phosphorylation by GSK3, and stabilizes Nrf1. Knockdown of Nrf1 and expression of a constitutively active form of GSK3 results in increased apoptosis in neuronal cells in response to ER stress, while expression of the GSK3 phosphorylation resistant S350A–Nrf1 attenuates apoptotic cell death. Together these data suggest that GSK3 regulates Nrf1 expression and cell survival function in response to stress activation. Highlights: • The effect of GSK3 on Nrf1 expression was examined. • GSK3 destabilizes Nrf1 protein via Fbw7 ubiquitin ligase. • GSK3 binds and phosphorylates Nrf1. • Protection from stress-induced apoptosis by Nrf1 is inhibited by GSK3.

  16. A 5' UTR-Overlapping LncRNA Activates the Male-Determining Gene doublesex1 in the Crustacean Daphnia magna.

    Science.gov (United States)

    Kato, Yasuhiko; Perez, Christelle Alexa G; Mohamad Ishak, Nur Syafiqah; Nong, Quang D; Sudo, Yuumi; Matsuura, Tomoaki; Wada, Tadashi; Watanabe, Hajime

    2018-06-04

    Long noncoding RNAs (lncRNAs) are pervasively transcribed in the eukaryotic genome [1] and are important for the control of master regulatory genes that are involved in cell differentiation and development [2, 3]. Here, we show that a 5' UTR-overlapping lncRNA regulates the male-specific expression of the DM-domain gene doublesex1 (dsx1) in the crustacean Daphnia magna, which produces males in response to environmental stimuli. This lncRNA, named doublesex1 alpha promoter-associated long RNA (DAPALR), is transcribed upstream the transcription start site (TSS) in a sense orientation and subjected to 5' end capping and 3' end processing at a stem-loop structure before the dsx1 coding exon. Similar to dsx1, its expression is only activated in males by the juvenile hormone (JH) and basic-leucine zipper (bZIP) transcription factor Vrille (Vri) and is maintained during embryogenesis. Knockdown of DAPALR in males silenced dsx1 and led to feminization, including egg production, whereas ectopic expression of DAPALR in dsx1-silenced females resulted in the de-repression of dsx1. We further demonstrate that the DAPALR transcript overlaps the dsx1 5'-UTR, and this overlapping region is required for dsx1 activation. Our results suggest that DAPALR can transactivate and possibly maintain dsx1 expression. This might be important for converting transient environmental signals into stable male development, controlled by the continuous expression of dsx1. Copyright © 2018 Elsevier Ltd. All rights reserved.

  17. Effects of Leucine Supplementation and Serum Withdrawal on Branched-Chain Amino Acid Pathway Gene and Protein Expression in Mouse Adipocytes

    Science.gov (United States)

    Vivar, Juan C.; Knight, Megan S.; Pointer, Mildred A.; Gwathmey, Judith K.; Ghosh, Sujoy

    2014-01-01

    The essential branched-chain amino acids (BCAA), leucine, valine and isoleucine, are traditionally associated with skeletal muscle growth and maintenance, energy production, and generation of neurotransmitter and gluconeogenic precursors. Recent evidence from human and animal model studies has established an additional link between BCAA levels and obesity. However, details of the mechanism of regulation of BCAA metabolism during adipogenesis are largely unknown. We interrogated whether the expression of genes and proteins involved in BCAA metabolism are sensitive to the adipocyte differentiation process, and responsive to nutrient stress from starvation or BCAA excess. Murine 3T3-L1 preadipocytes were differentiated to adipocytes under control conditions and under conditions of L-leucine supplementation or serum withdrawal. RNA and proteins were isolated at days 0, 4 and 10 of differentiation to represent pre-differentiation, early differentiation and late differentiation stages. Expression of 16 BCAA metabolism genes was quantified by quantitative real-time PCR. Expression of the protein levels of branched-chain amino acid transaminase 2 (Bcat2) and branched-chain alpha keto acid dehydrogenase (Bckdha) was quantified by immunoblotting. Under control conditions, all genes displayed induction of gene expression during early adipogenesis (Day 4) compared to Day 0. Leucine supplementation resulted in an induction of Bcat2 and Bckdha genes during early and late differentiation. Western blot analysis demonstrated condition-specific concordance between gene and protein expression. Serum withdrawal resulted in undetectable Bcat2 and Bckdha protein levels at all timepoints. These results demonstrate that the expression of genes related to BCAA metabolism are regulated during adipocyte differentiation and influenced by nutrient levels. These results provide additional insights on how BCAA metabolism is associated with adipose tissue function and extends our understanding of

  18. Effects of leucine supplementation and serum withdrawal on branched-chain amino acid pathway gene and protein expression in mouse adipocytes.

    Directory of Open Access Journals (Sweden)

    Abderrazak Kitsy

    Full Text Available The essential branched-chain amino acids (BCAA, leucine, valine and isoleucine, are traditionally associated with skeletal muscle growth and maintenance, energy production, and generation of neurotransmitter and gluconeogenic precursors. Recent evidence from human and animal model studies has established an additional link between BCAA levels and obesity. However, details of the mechanism of regulation of BCAA metabolism during adipogenesis are largely unknown. We interrogated whether the expression of genes and proteins involved in BCAA metabolism are sensitive to the adipocyte differentiation process, and responsive to nutrient stress from starvation or BCAA excess. Murine 3T3-L1 preadipocytes were differentiated to adipocytes under control conditions and under conditions of L-leucine supplementation or serum withdrawal. RNA and proteins were isolated at days 0, 4 and 10 of differentiation to represent pre-differentiation, early differentiation and late differentiation stages. Expression of 16 BCAA metabolism genes was quantified by quantitative real-time PCR. Expression of the protein levels of branched-chain amino acid transaminase 2 (Bcat2 and branched-chain alpha keto acid dehydrogenase (Bckdha was quantified by immunoblotting. Under control conditions, all genes displayed induction of gene expression during early adipogenesis (Day 4 compared to Day 0. Leucine supplementation resulted in an induction of Bcat2 and Bckdha genes during early and late differentiation. Western blot analysis demonstrated condition-specific concordance between gene and protein expression. Serum withdrawal resulted in undetectable Bcat2 and Bckdha protein levels at all timepoints. These results demonstrate that the expression of genes related to BCAA metabolism are regulated during adipocyte differentiation and influenced by nutrient levels. These results provide additional insights on how BCAA metabolism is associated with adipose tissue function and extends our

  19. Metabolism of L-leucine-U-14C in young rats fed excess glycine diets

    International Nuclear Information System (INIS)

    Takeuchi, Hisanao; Tadauchi, Nobuo; Muramatsu, Keiichiro

    1975-01-01

    As reported previously, while the growth-depressing effect of excess glycine was prevented by supplementing L-arginine and L-methionine, the degradation of glycine-U-(SUP 14)C into expired carbon dioxide was not accelerated by the supplement of both amino acids. However, it was found that the incorporation of the isotope into the lipids of livers and carcasses increased in the rats fed the excess glycine diet containing both amino acids. The lipid synthesis utilizing excess glycine may be accelerated by adding both amino acids to the 10% casein diet containing excess glycine. In the present experiment, the metabolic fate of L-leucine-U-(SUP 14)C was studied with the rats fed the excess glycine diet with or without L-arginine and L-methionine. 10% casein (10C), 10% casein diet containing 7% glycine (10C7G), or 10C7G Supplemented with 1.4% L-arginine-HCL and 0.9% L-methionine (10C7GArgMet) was fed to each rat, and the diet suspension containing 4 sup(μ)Ci of L-leucine-U-(SUP 14)C per 100 g of body weight was fed forcibly after 12 hr fast. The radioactivity in expired carbon dioxide, TCA soluble fraction, protein, glycogen, lipids and urine, and the concentration of free amino acids in blood plasma, livers and urine were measured. The body weight gain and food intake of the 10C7G group were much smaller than those of the other groups. The recovery of (SUP 14)C-radioactivity in expired carbon dioxide was much lower in the 10C7GArgMet group than that of the other groups. (Kako, I.)

  20. Co-ingestion of a protein hydrolysate with or without additional leucine effectively reduces postprandial blood glucose excursions in type 2 diabetic men

    NARCIS (Netherlands)

    Manders, R.J.; Koopman, R.; Sluijsmans, W.E.; Berg, R. van den; Verbeek, K.; Saris, W.H.; Wagenmakers, A.J.; Loon, L.J. van

    2006-01-01

    This study examined postprandial plasma insulin and glucose responses after co-ingestion of an insulinotropic protein (Pro) hydrolysate with and without additional free leucine with a single bolus of carbohydrate (Cho). Male patients with long-standing Type 2 diabetes (n = 10) and healthy controls

  1. TCR comodulation of nonengaged TCR takes place by a protein kinase C and CD3 gamma di-leucine-based motif-dependent mechanism

    DEFF Research Database (Denmark)

    Bonefeld, Charlotte Menné; Rasmussen, B. A.; Lauritsen, J P

    2003-01-01

    of comodulation. Like internalization of engaged TCR, comodulation was dependent on protein tyrosine kinase activity. Finally, we found that in contrast to internalization of engaged TCR, comodulation was highly dependent on protein kinase C activity and the CD3 gamma di-leucine-based motif. Based...

  2. Authentication of pure L-leucine products manufactured in China by discriminating between plant and animal sources using nitrogen stable isotope technique.

    Science.gov (United States)

    Huang, Jingyu; Nkrumah, Philip N; Appiah-Sefah, Gloria; Tang, Shijiang

    2013-03-01

     L-leucine products among other branched chain amino acid supplements are highly susceptible to economically motivated adulteration. Curbing this menace is critical and timely. Hence, the δ(15) N composition of the L-leucine derived from plants and animals sources was estimated. The trophic enrichment phenomenon of δ(15) N composition was utilized to elucidate the sources. We finally established the distinction between the respective sources. Samples of plant sources (maize and soybean) and that of animal sources (pig fur and duck feather) were analyzed for δ(15) N isotopic signatures. An elemental analyzer which was connected to an isotope ratio mass spectrometer operated in the continuous flow mode was utilized. The raw materials were obtained from China. Statistical analysis was performed using descriptive statistics and one-way analysis of variance. The results indicated lower δ(15) N values of range -0.7344‰ to 2.384‰ and 1.032‰ to 2.064‰ for maize and soybean samples, respectively. Whereas, a range of 3.860‰ to 6.011‰ and 5.875‰ to 6.011‰ was, respectively, detected in pig fur and duck feather samples. The δ(15) N difference in plants and animals samples was significant (F = 165.0; P = 1.675 E-10 for maize and pig fur samples; F = 212.8; P = 0.0001284 for soybean and duck feather samples). It was observed that δ(15) N trophic enrichment is helpful in elucidating the respective sources. The authors can emphatically assert that the range of δ(15) N composition of L-leucine derived from plants sources within the study area is -1.000‰ to 3.000‰ whereas the range in animal sources is 4.000‰ to 9.000‰. Practical Application This study provides a reliable approach in verifying the authenticity of not only L-leucine products but also other branched chain amino acid supplements and thereby would help in fraud detection of any economically motivated adulteration and mislabeling of these products. When coupled with H and O stable

  3. HPLC Separation of Sulforaphane Enantiomers in Broccoli and Its Sprouts by Transformation into Diastereoisomers Using Derivatization with (S)-Leucine.

    Science.gov (United States)

    Okada, Makiko; Yamamoto, Atsushi; Aizawa, Sen-Ichi; Taga, Atsushi; Terashima, Hiroyuki; Kodama, Shuji

    2017-01-11

    Racemic sulforaphane, which was derivatized with (S)-leucine (l-leucine), was resolved by reversed phase HPLC with UV detection. The optimum mobile phase conditions were found to be 10 mM citric acid (pH 2.8) containing 22% methanol at 35 °C using detection at 254 nm. Sulforaphane enantiomers in florets and stems of five brands of broccoli and leaves and stems of three brands of broccoli sprouts were analyzed by the proposed HPLC method. Both sulforaphane enantiomers were detected in all of the samples. The S/R ratios of sulforaphane in broccoli samples were 1.5-2.6/97.4-98.5% for florets and 5.0-12.1/87.9-95.0% for stems. The S/R ratios in broccoli sprout samples were higher than those in broccoli samples and were found to be 8.3-19.7/80.3-91.7% for leaves and 37.0-41.8/58.2-63.0% for stems. (S)-Sulforaphane detected in the broccoli and its sprout samples was positively identified by separately using an HPLC with a chiral column (Chiralpak AD-RH) and mass spectrometry.

  4. Effect of chromatolysis upon the incorporation of L-4,5-(3H)-leucine into axotomized neurons

    International Nuclear Information System (INIS)

    Soereide, A.J.

    1981-01-01

    Autoradiography was used to compare the rate of protein synthesis in neurons which showed different types of neuronal change after axotomy. Different types of axon reaction were obtained after crush lesion of the facial nerve in rats and young rabbits and after evulsion of the facial nerve in rats. The incorporation of ( 3 H)-leucine into protein was determined after intraperitoneal injection by counting silver grains over facial neurons in autoradiograms prepared from paraffin sections. Crush lesion of the facial nerve in rats was followed by barely discernible changes in the neurons, no chromatolysis, and full functional recovery. The lesion caused a significant transitory increase in the protein synthesis. After nerve evulsion, the same neurons showed a severe central chromatolysis and ultimate nerve cell disintegration. The protein synthesis was unchanged during the early chromotolytic phase after evulsion and decreased at later stages. Crush lesion of the facial nerve in young rabbits was followed by severe central chromotolysis, full functional recovery, and no loss of neurons. There was a moderate increase in the protein synthesis during chromatolysis and a further increase during the phase of recovery. It is concluded that regeneration after axotomy is accompanied by an increase in protein synthesis, regardless of the type of morphologic change in the neurons. Chromatolysis is not a prerequisite for this increase, and the chromatolytic response per se seems to be of minor importance for the extent of incorporation of ( 3 H)-leucine into neuronal proteins. (orig.)

  5. Effect of insulin on albumin production and incorporation of 14C-leucine into proteins in isolated parenchymal liver cells from normal rats

    DEFF Research Database (Denmark)

    Dich, J; Gluud, C N

    1975-01-01

    the immunologically determined increment in the incubation medium was 1.7 +/- 0.2 mug albumin/min per g liver wet wt. This is about 30% of the rate of production in the perfused liver. Addition of insulin (10(-6)-10(-10) M) enhanced albumin production (50-17%), and incorporation of 14C-leucine both into albumin (50......Parenchymal rat liver cells were isolated by a modification of the collagenase method of Quistorff, Bondesen and Grunnet. The cells secreted albumin into the medium and incorporated 14C-leucine both into cell proteins and proteins secreted into the medium. Albumin production measured from......-8%), secreted proteins (40-9%) and cell proteins (20-8%). Insulin does not increase the production of albumin by depleting the cells. The effect of insulin on albumin production is compatible with an effect on the rate of synthesis as the specific activity of albumin is unaffected by addition of insulin....

  6. Mutations of C19orf12, coding for a transmembrane glycine zipper containing mitochondrial protein, cause mis-localization of the protein, inability to respond to oxidative stress and increased mitochondrial Ca2+.

    Directory of Open Access Journals (Sweden)

    Paola eVenco

    2015-05-01

    Full Text Available Mutations in C19orf12 have been identified in patients affected by Neurodegeneration with Brain Iron Accumulation (NBIA, a clinical entity characterized by iron accumulation in the basal ganglia. By using western blot analysis with specific antibody and confocal studies, we showed that wild-type C19orf12 protein was not exclusively present in mitochondria, but also in the Endoplasmic Reticulum (ER and MAM (Mitochondria Associated Membrane, while mutant C19orf12 variants presented a different localization. Moreover, after induction of oxidative stress, a GFP-tagged C19orf12 wild-type protein was able to relocate to the cytosol. On the contrary, mutant isoforms were not able to respond to oxidative stress. High mitochondrial calcium concentration and increased H2O2 induced apoptosis were found in fibroblasts derived from one patient as compared to controls.C19orf12 protein is a 17kDa mitochondrial membrane-associated protein whose function is still unknown. Our in silico investigation suggests that, the glycine zipper motifs of C19orf12 form helical regions spanning the membrane. The N- and C-terminal regions with respect to the transmembrane portion, on the contrary, are predicted to rearrange in a structural domain, which is homologues to the N-terminal regulatory domain of the magnesium transporter MgtE, suggesting that C19orf12 may act as a regulatory protein for human MgtE transporters. The mutations here described affect respectively one glycine residue of the glycine zipper motifs, which are involved in dimerization of transmembrane helices and predicted to impair the correct localization of the protein into the membranes, and one residue present in the regulatory domain, which is important for protein-protein interaction.

  7. Leucine Supplementation in a Chronically Protein-Restricted Diet Enhances Muscle Weight and Postprandial Protein Synthesis of Skeletal Muscle by Promoting the mTOR Pathway in Adult Rats

    Directory of Open Access Journals (Sweden)

    Bo Zhang

    2017-10-01

    Full Text Available Low protein intake causes a decrease in protein deposition in most animal tissues. The purpose of this study was to investigate whether leucine supplementation would increase the synthesis rate of protein and muscle weight in adult rats, which chronically consume only 58.8% of their protein requirements. Thirty-six male Sprague-Dawley rats were assigned to one of three dietary treatments including a 20% casein diet (CON, a 10% casein + 0.44% alanine diet (R, and a 10% casein + 0.87% leucine diet (RL. After a 10 d dietary treatment, plasma amino acid levels were measured after feeding, the gastrocnemius muscles and soleus muscles were harvested and weighed, and the fractional synthesis rate (FSR and mammalian target of rapamycin (mTOR signaling proteins in skeletal muscle were measured. Regarding the plasma amino acid level, the RL group had the highest concentration of leucine (P < 0.05 and the lowest concentration of isoleucine (P < 0.05 among the three groups, and the CON group had a lower concentration of valine (P < 0.05 than the R and RL groups. Compared with the R and RL groups, the CON group diet significantly increased (P < 0.05 feed intake, protein synthesis rate, and the phosphorylation of eukaryotic initiation factor 4E binding protein 1 (4E-BP1, and decreased the weight of abdominal adipose. Compared with the R group, the RL group significantly increased in gastrocnemius muscle weight, protein synthesis rate, and phosphorylation of both ribosomal protein S6 kinase 1 (S6K1 and 4E-BP1. In conclusion, when protein is chronically restricted in adult rat diets, leucine supplementation moderately improves body weight gain and increases muscle protein synthesis through mTOR activation.

  8. Self-assembling triblock proteins for biofunctional surface modification

    Science.gov (United States)

    Fischer, Stephen E.

    Despite the tremendous promise of cell/tissue engineering, significant challenges remain in engineering functional scaffolds to precisely regulate the complex processes of tissue growth and development. As the point of contact between the cells and the scaffold, the scaffold surface plays a major role in mediating cellular behaviors. In this dissertation, the development and utility of self-assembling, artificial protein hydrogels as biofunctional surface modifiers is described. The design of these recombinant proteins is based on a telechelic triblock motif, in which a disordered polyelectrolyte central domain containing embedded bioactive ligands is flanked by two leucine zipper domains. Under moderate conditions of temperature and pH, the leucine zipper end domains form amphiphilic alpha-helices that reversibly associate into homo-trimeric aggregates, driving hydrogel formation. Moreover, the amphiphilic nature of these helical domains enables surface adsorption to a variety of scaffold materials to form biofunctional protein coatings. The nature and stability of these coatings in various solution conditions, and their interaction with mammalian cells is the primary focus of this dissertation. In particular, triblock protein coatings functionalized with cell recognition sequences are shown to produce well-defined surfaces with precise control over ligand density. The impact of this is demonstrated in multiple cell types through ligand density-dependent cell-substrate interactions. To improve the stability of these physically self-assembled coatings, two covalent crosslinking strategies are described---one in which a zero-length chemical crosslinker (EDC) is utilized and a second in which disulfide bonds are engineered into the recombinant proteins. These targeted crosslinking approaches are shown to increase the stability of surface adsorbed protein layers with minimal effect on the presentation of many bioactive ligands. Finally, to demonstrate the versatility

  9. The Ste20 Family Kinases MAP4K4, MINK1, and TNIK Converge to Regulate Stress-Induced JNK Signaling in Neurons.

    Science.gov (United States)

    Larhammar, Martin; Huntwork-Rodriguez, Sarah; Rudhard, York; Sengupta-Ghosh, Arundhati; Lewcock, Joseph W

    2017-11-15

    The c-Jun- N -terminal kinase (JNK) signaling pathway regulates nervous system development, axon regeneration, and neuronal degeneration after acute injury or in chronic neurodegenerative disease. Dual leucine zipper kinase (DLK) is required for stress-induced JNK signaling in neurons, yet the factors that initiate DLK/JNK pathway activity remain poorly defined. In the present study, we identify the Ste20 kinases MAP4K4, misshapen-like kinase 1 (MINK1 or MAP4K6) and TNIK Traf2- and Nck-interacting kinase (TNIK or MAP4K7), as upstream regulators of DLK/JNK signaling in neurons. Using a trophic factor withdrawal-based model of neurodegeneration in both male and female embryonic mouse dorsal root ganglion neurons, we show that MAP4K4, MINK1, and TNIK act redundantly to regulate DLK activation and downstream JNK-dependent phosphorylation of c-Jun in response to stress. Targeting MAP4K4, MINK1, and TNIK, but not any of these kinases individually, is sufficient to protect neurons potently from degeneration. Pharmacological inhibition of MAP4Ks blocks stabilization and phosphorylation of DLK within axons and subsequent retrograde translocation of the JNK signaling complex to the nucleus. These results position MAP4Ks as important regulators of the DLK/JNK signaling pathway. SIGNIFICANCE STATEMENT Neuronal degeneration occurs in disparate circumstances: during development to refine neuronal connections, after injury to clear damaged neurons, or pathologically during disease. The dual leucine zipper kinase (DLK)/c-Jun- N -terminal kinase (JNK) pathway represents a conserved regulator of neuronal injury signaling that drives both neurodegeneration and axon regeneration, yet little is known about the factors that initiate DLK activity. Here, we uncover a novel role for a subfamily of MAP4 kinases consisting of MAP4K4, Traf2- and Nck-interacting kinase (TNIK or MAP4K7), and misshapen-like kinase 1 (MINK1 or MAP4K6) in regulating DLK/JNK signaling in neurons. Inhibition of

  10. Involvement of transcription factor encoded by the mouse mi locus (MITF) in apoptosis of cultured mast cells induced by removal of interleukin-3.

    Science.gov (United States)

    Tsujimura, T.; Hashimoto, K.; Morii, E.; Tunio, G. M.; Tsujino, K.; Kondo, T.; Kanakura, Y.; Kitamura, Y.

    1997-01-01

    Mast cells develop when spleen cells of mice are cultured in the medium containing interleukin (IL)-3. Cultured mast cells (CMCs) show apoptosis when they are incubated in the medium without IL-3. We obtained CMCs from tg/tg mice that did not express the transcription factor encoded by the mi gene (MITF) due to the integration of a transgene at its 5' flanking region. MITF is a member of the basic-helix-loop-helix-leucine zipper (bHLH-Zip) protein family of transcription factors. We investigated the effect of MITF on the apoptosis of CMCs after removal of IL-3. When cDNA encoding normal MITF ((+)-MITF) was introduced into tg/tg CMCs with the retroviral vector, the apoptosis of tg/tg CMCs was significantly accelerated. The mutant mi allele represents a deletion of an arginine at the basic domain of MITF. The apoptosis of tg/tg CMCs was not accelerated by the introduction of cDNA encoding mi-MITF. The overexpression of (+)-MITF was not prerequisite to the acceleration of the apoptosis, as the apoptotic process proceeded faster in +/+ CMCs than in mi/mi CMCs. The Ba/F3 lymphoid cell line is also dependent on IL-3, and Ba/F3 cells show apoptosis after removal of IL-3. The c-myc gene encodes another transcription factor of the bHLH-Zip family, and the overexpression of the c-myc gene accelerated the apoptosis of Ba/F3 cells. However, the overexpression of (+)-MITF did not accelerate the apoptosis of Ba/F3 cells. The (+)-MITF appeared to play some roles for the acceleration of the apoptosis specifically in the mast cell lineage. Images Figure 2 Figure 3 Figure 4 Figure 5 Figure 6 Figure 7 PMID:9327738

  11. Identification of the G13 (cAMP-response-element-binding protein-related protein) gene product related to activating transcription factor 6 as a transcriptional activator of the mammalian unfolded protein response.

    Science.gov (United States)

    Haze, K; Okada, T; Yoshida, H; Yanagi, H; Yura, T; Negishi, M; Mori, K

    2001-04-01

    Eukaryotic cells control the levels of molecular chaperones and folding enzymes in the endoplasmic reticulum (ER) by a transcriptional induction process termed the unfolded protein response (UPR). The mammalian UPR is mediated by the cis-acting ER stress response element consisting of 19 nt (CCAATN(9)CCACG), the CCACG part of which is considered to provide specificity. We recently identified the basic leucine zipper (bZIP) protein ATF6 as a mammalian UPR-specific transcription factor; ATF6 is activated by ER stress-induced proteolysis and binds directly to CCACG. Here we report that eukaryotic cells express another bZIP protein closely related to ATF6 in both structure and function. This protein encoded by the G13 (cAMP response element binding protein-related protein) gene is constitutively synthesized as a type II transmembrane glycoprotein anchored in the ER membrane and processed into a soluble form upon ER stress as occurs with ATF6. The proteolytic processing of ATF6 and the G13 gene product is accompanied by their relocation from the ER to the nucleus; their basic regions seem to function as a nuclear localization signal. Overexpression of the soluble form of the G13 product constitutively activates the UPR, whereas overexpression of a mutant lacking the activation domain exhibits a strong dominant-negative effect. Furthermore, the soluble forms of ATF6 and the G13 gene product are unable to bind to several point mutants of the cis-acting ER stress response element in vitro that hardly respond to ER stress in vivo. We thus concluded that the two related bZIP proteins are crucial transcriptional regulators of the mammalian UPR, and propose calling the ATF6 gene product ATF6alpha and the G13 gene product ATF6beta.

  12. The small leucine-rich proteoglycan, biglycan, is highly expressed in adipose tissue of Psammomys obesus and is associated with obesity and type 2 diabetes

    Directory of Open Access Journals (Sweden)

    Bolton K

    2012-04-01

    Full Text Available Kristy Bolton1, David Segal1, Ken Walder1,21Metabolic Research Unit, School of Medicine, 2Institute for Technology, Research and Innovation, Deakin University, Waurn Ponds, Victoria, AustraliaAbstract: We have previously demonstrated that the small leucine-rich proteoglycan decorin may play a role in adipose tissue homeostasis and the pathophysiology of obesity. Biglycan is highly similar in structure to decorin, therefore we hypothesized it would have a similar expression profile and role to decorin in adipose tissue. Real time polymerase chain reaction was used to measure biglycan mRNA levels in adipose tissue from normal glucose tolerant and impaired glucose tolerant and type 2 diabetic (T2D Psammomys obesus. Biglycan mRNA was found to be highly expressed in adipose tissue, and gene expression was significantly higher in visceral compared to subcutaneous adipose tissue, with elevated levels in obese, T2D compared to lean normal glucose tolerant P. obesus (P < 0.04. Biglycan mRNA was predominantly expressed by stromal/vascular cells of fractionated adipose tissue (P = 0.023. Biglycan expression in adipose tissue, particularly in the obese state, was markedly upregulated. Collectively, our data suggest that the small leucine-rich proteoglycan family proteins biglycan and decorin may play a role in the development of obesity and T2D, possibly by facilitating expansion of adipose tissue mass.Keywords: biglycan, small leucine-rich proteoglycan, Psammomys obesus, adipose tissue, obesity, type 2 diabetes

  13. Differentiating leucine incorporation of Archaea and Bacteria throughout the water column of the eastern Atlantic using metabolic inhibitors

    OpenAIRE

    Yokokawa, Taichi; Sintes, Eva; de Corte, Daniele; Olbrich, Kerstin; Herndl, Gerhard J.

    2012-01-01

    The abundance (based on catalyzed reporter deposition-fluorescence in situ hybrid ization, CARD-FISH) and leucine incorporation rates of Archaea and Bacteria were determined throughout the water column in the eastern Atlantic. Bacteria dominated throughout the water column, although their contribution to total prokaryotic abundance in the bathypelagic layer (1000 to 4000 m depth) was lower than in the surface and mesopelagic layers (0 to 1000 m depth). While marine Crenarchaeota Group I (MCG ...

  14. Impaired Uptake and/or Utilization of Leucine by Saccharomyces cerevisiae Is Suppressed by the SPT15-300 Allele of the TATA-Binding Protein Gene

    DEFF Research Database (Denmark)

    Baerends, RJ; Qiu, Jin-Long; Rasmussen, Simon

    2009-01-01

    Successful fermentations to produce ethanol require microbial strains that have a high tolerance to glucose and ethanol. Enhanced glucose/ethanol tolerance of the laboratory yeast Saccharomyces cerevisiae strain BY4741 under certain growth conditions as a consequence of the expression of a dominant...... us to examine the effect of expression of the SPT15-300 allele in various yeast species of industrial importance. Expression of SPT15-300 in leucine-prototrophic strains of S. cerevisiae, Saccharomyces bayanus, or Saccharomyces pastorianus (lager brewing yeast), however, did not improve tolerance...... to ethanol on complex rich medium (yeast extract-peptone-dextrose). The enhanced growth of the laboratory yeast strain BY4741 expressing the SPT15-300 mutant allele was seen only on defined media with low concentrations of leucine, indicating that the apparent improved growth in the presence of ethanol...

  15. Inhibition of Curcumin on ZAKα Activity Resultant in Apoptosis and Anchorage-Independent Growth in Cancer Cells.

    Science.gov (United States)

    Lee, Jin-Sun; Wang, Tsu-Shing; Lin, Ming Cheng; Lin, Wei-Wen; Yang, Jaw-Ji

    2017-10-31

    Curcumin, a popular yellow pigment of the dietary spice turmeric, has been reported to inhibit cell growth and to induce apoptosis in a wide variety of cancer cells. Although numerous studies have investigated anticancer effects of curcumin, the precise molecular mechanism of action remains unidentified. Whereas curcumin mediates cell survival and apoptosis through mitogen-activated protein kinase (MAPK) and nuclear factor-kappa B (NF-κB) signaling cascades, its impact on the upstream regulation of MAPK is unclear. The leucine-zipper and sterile-α motif kinase alpha (ZAKα), a mitogen-activated protein kinase kinase kinase (MAP3K), activates the c-Jun N-terminal kinase (JNK) and NF-κB pathway. This paper investigated the prospective involvement of ZAKα in curcumin-induced effects on cancer cells. Our results suggest that the antitumor activity of curcumin is mediated via a mechanism involving inhibition of ZAKα activity.

  16. Tunable paramagnetic relaxation enhancements by [Gd(DPA)3]3- for protein structure analysis

    International Nuclear Information System (INIS)

    Yagi, Hiromasa; Loscha, Karin V.; Su, Xun-Cheng; Stanton-Cook, Mitchell; Huber, Thomas; Otting, Gottfried

    2010-01-01

    Paramagnetic relaxation enhancements (PRE) present a powerful source of structural information in nuclear magnetic resonance (NMR) studies of proteins and protein-ligand complexes. In contrast to conventional PRE reagents that are covalently attached to the protein, the complex between gadolinium and three dipicolinic acid (DPA) molecules, [Gd(DPA) 3 ] 3- , can bind to proteins in a non-covalent yet site-specific manner. This offers straightforward access to PREs that can be scaled by using different ratios of [Gd(DPA) 3 ] 3- to protein, allowing quantitative distance measurements for nuclear spins within about 15 A of the Gd 3+ ion. Such data accurately define the metal position relative to the protein, greatly enhancing the interpretation of pseudocontact shifts induced by [Ln(DPA) 3 ] 3- complexes of paramagnetic lanthanide (Ln 3+ ) ions other than gadolinium. As an example we studied the quaternary structure of the homodimeric GCN4 leucine zipper.

  17. Requirement of Sequences outside the Conserved Kinase Domain of Fission Yeast Rad3p for Checkpoint Control

    Science.gov (United States)

    Chapman, Carolyn Riley; Evans, Sarah Tyler; Carr, Antony M.; Enoch, Tamar

    1999-01-01

    The fission yeast Rad3p checkpoint protein is a member of the phosphatidylinositol 3-kinase-related family of protein kinases, which includes human ATMp. Mutation of the ATM gene is responsible for the disease ataxia-telangiectasia. The kinase domain of Rad3p has previously been shown to be essential for function. Here, we show that although this domain is necessary, it is not sufficient, because the isolated kinase domain does not have kinase activity in vitro and cannot complement a rad3 deletion strain. Using dominant negative alleles of rad3, we have identified two sites N-terminal to the conserved kinase domain that are essential for Rad3p function. One of these sites is the putative leucine zipper, which is conserved in other phosphatidylinositol 3-kinase-related family members. The other is a novel motif, which may also mediate Rad3p protein–protein interactions. PMID:10512862

  18. Effect of exercise training on leucine oxidation

    International Nuclear Information System (INIS)

    Hendrix, M.K.; Layman, D.K.

    1986-01-01

    Oxidation of the BCAA leucine is increased during a bout of exhaustive exercise. The purpose of this study was to determine the effects of exercise training on leu oxidation during aerobic exercise. Female Sprague-Dawley rats were fed a commercial diet ad lib and divided into sedentary and two trained groups. Animals were trained to run on a treadmill with a 10 0 incline at 28 m/min for 5 wks for either 50 or 120 min/day. There were no differences in food intake or body weight. After a 12 hr fast, animals were run for 50 or 120 min and changes in leu catabolism determined by measurement of in vivo leu oxidation and activity of branched chain keto acid dehydrogenase (BCKAD). For measurement of leu oxidation, rats were injected IP with 4 μCi 1- 14 C-leu during the last 15 min of exercise, placed in glass metabolic chambers, and 14 CO 2 collected in 1 N NaOH for 30 min periods. Leu oxidation was increased by 40% after 50 min of exercise and by 79% after 120 min of exercise. Five weeks of training reduced the rate of leu oxidation during an exercise bout. The activity of the BCKAD was not increased in the trained animals after either 50 or 120 min of exercise. These data indicate that the rate of leu oxidation during exercises is dependent on the duration of the exercise and that training will reduce the magnitude of this effect

  19. Effect of exercise training on leucine oxidation

    Energy Technology Data Exchange (ETDEWEB)

    Hendrix, M.K.; Layman, D.K.

    1986-03-01

    Oxidation of the BCAA leucine is increased during a bout of exhaustive exercise. The purpose of this study was to determine the effects of exercise training on leu oxidation during aerobic exercise. Female Sprague-Dawley rats were fed a commercial diet ad lib and divided into sedentary and two trained groups. Animals were trained to run on a treadmill with a 10/sup 0/ incline at 28 m/min for 5 wks for either 50 or 120 min/day. There were no differences in food intake or body weight. After a 12 hr fast, animals were run for 50 or 120 min and changes in leu catabolism determined by measurement of in vivo leu oxidation and activity of branched chain keto acid dehydrogenase (BCKAD). For measurement of leu oxidation, rats were injected IP with 4 ..mu..Ci 1-/sup 14/C-leu during the last 15 min of exercise, placed in glass metabolic chambers, and /sup 14/CO/sub 2/ collected in 1 N NaOH for 30 min periods. Leu oxidation was increased by 40% after 50 min of exercise and by 79% after 120 min of exercise. Five weeks of training reduced the rate of leu oxidation during an exercise bout. The activity of the BCKAD was not increased in the trained animals after either 50 or 120 min of exercise. These data indicate that the rate of leu oxidation during exercises is dependent on the duration of the exercise and that training will reduce the magnitude of this effect.

  20. A microsystem to evaluate the synthesis of [3H]leucine labeled proteins by macrophages

    International Nuclear Information System (INIS)

    Varesio, L.; Eva, A.

    1980-01-01

    A method is described for evaluating protein synthesis by adherent MPHI by measuring the incorporation of [ 3 H]leucine into TCA precipitable material. By using guanidine-HCl it was possible to remove completely the radiolabeled proteins from the adherent cells that were cultured in microwells, and retain TCA precipitable material. This procedure enabled the authors to harvest the TCA precipitable proteins with a semiautomatic cell harvester. The guanidine-HCl treatment did not affect the recovery of the radioactive proteins and did not alter the sensitivity of the assay. This method is very simple and rapid and, since it is suitable for processing microcultures, permits detailed studies on the biology of small numbers of MPHI. (Auth.)

  1. Solvation behaviour of L-leucine in aqueous ionic liquid at different temperatures: Volumetric approach

    Science.gov (United States)

    Sharma, Samriti; Sandarve, Sharma, Amit K.; Sharma, Meena

    2018-05-01

    For the investigation of interactions of L-leucine in aqueous solutions of an ionic liquid (1-butyl-3-methylimidazolium tetra fluoroborate [Bmim][BF4]) at atmospheric pressure over a temperature range of (293.15K to 313.16K), we use the volumetric approach. By using the density data we have calculated the apparent molar volume, VΦ, limiting apparent molar volume, V0Φ, the slope, Sv, partial molar volume of transfer, V0Φ,tr. The values of these acoustical parameters have been used for the interpretation of different interactions like hydrophilic-hydrophilic, hydrophilic-hydrophobic, ion hydrophilic, solute-solvent and solute-solute interactions in the amino acid and ionic liquid solutions.

  2. Arabidopsis leucine-rich repeat extensin (LRX) proteins modify cell wall composition and influence plant growth.

    Science.gov (United States)

    Draeger, Christian; Ndinyanka Fabrice, Tohnyui; Gineau, Emilie; Mouille, Grégory; Kuhn, Benjamin M; Moller, Isabel; Abdou, Marie-Therese; Frey, Beat; Pauly, Markus; Bacic, Antony; Ringli, Christoph

    2015-06-24

    Leucine-rich repeat extensins (LRXs) are extracellular proteins consisting of an N-terminal leucine-rich repeat (LRR) domain and a C-terminal extensin domain containing the typical features of this class of structural hydroxyproline-rich glycoproteins (HRGPs). The LRR domain is likely to bind an interaction partner, whereas the extensin domain has an anchoring function to insolubilize the protein in the cell wall. Based on the analysis of the root hair-expressed LRX1 and LRX2 of Arabidopsis thaliana, LRX proteins are important for cell wall development. The importance of LRX proteins in non-root hair cells and on the structural changes induced by mutations in LRX genes remains elusive. The LRX gene family of Arabidopsis consists of eleven members, of which LRX3, LRX4, and LRX5 are expressed in aerial organs, such as leaves and stem. The importance of these LRX genes for plant development and particularly cell wall formation was investigated. Synergistic effects of mutations with gradually more severe growth retardation phenotypes in double and triple mutants suggest a similar function of the three genes. Analysis of cell wall composition revealed a number of changes to cell wall polysaccharides in the mutants. LRX3, LRX4, and LRX5, and most likely LRX proteins in general, are important for cell wall development. Due to the complexity of changes in cell wall structures in the lrx mutants, the exact function of LRX proteins remains to be determined. The increasingly strong growth-defect phenotypes in double and triple mutants suggests that the LRX proteins have similar functions and that they are important for proper plant development.

  3. Effect of chromatolysis upon the incorporation of L-4,5-(/sup 3/H)-leucine into axotomized neurons

    Energy Technology Data Exchange (ETDEWEB)

    Soereide, A.J.

    1981-01-01

    Autoradiography was used to compare the rate of protein synthesis in neurons which showed different types of neuronal change after axotomy. Different types of axon reaction were obtained after crush lesion of the facial nerve in rats and young rabbits and after evulsion of the facial nerve in rats. The incorporation of (/sup 3/H)-leucine into protein was determined after intraperitoneal injection by counting silver grains over facial neurons in autoradiograms prepared from paraffin sections. Crush lesion of the facial nerve in rats was followed by barely discernible changes in the neurons, no chromatolysis, and full functional recovery. The lesion caused a significant transitory increase in the protein synthesis. After nerve evulsion, the same neurons showed a severe central chromatolysis and ultimate nerve cell disintegration. The protein synthesis was unchanged during the early chromotolytic phase after evulsion and decreased at later stages. Crush lesion of the facial nerve in young rabbits was followed by severe central chromotolysis, full functional recovery, and no loss of neurons. There was a moderate increase in the protein synthesis during chromatolysis and a further increase during the phase of recovery. It is concluded that regeneration after axotomy is accompanied by an increase in protein synthesis, regardless of the type of morphologic change in the neurons. Chromatolysis is not a prerequisite for this increase, and the chromatolytic response per se seems to be of minor importance for the extent of incorporation of (/sup 3/H)-leucine into neuronal proteins.

  4. Activation of the bacterial thermosensor DesK involves a serine zipper dimerization motif that is modulated by bilayer thickness.

    Science.gov (United States)

    Cybulski, Larisa Estefanía; Ballering, Joost; Moussatova, Anastassiia; Inda, Maria Eugenia; Vazquez, Daniela B; Wassenaar, Tsjerk A; de Mendoza, Diego; Tieleman, D Peter; Killian, J Antoinette

    2015-05-19

    DesK is a bacterial thermosensor protein involved in maintaining membrane fluidity in response to changes in environmental temperature. Most likely, the protein is activated by changes in membrane thickness, but the molecular mechanism of sensing and signaling is still poorly understood. Here we aimed to elucidate the mode of action of DesK by studying the so-called "minimal sensor DesK" (MS-DesK), in which sensing and signaling are captured in a single transmembrane segment. This simplified version of the sensor allows investigation of membrane thickness-dependent protein-lipid interactions simply by using synthetic peptides, corresponding to the membrane-spanning parts of functional and nonfunctional mutants of MS-DesK incorporated in lipid bilayers with varying thicknesses. The lipid-dependent behavior of the peptides was investigated by circular dichroism, tryptophan fluorescence, and molecular modeling. These experiments were complemented with in vivo functional studies on MS-DesK mutants. Based on the results, we constructed a model that suggests a new mechanism for sensing in which the protein is present as a dimer and responds to an increase in bilayer thickness by membrane incorporation of a C-terminal hydrophilic motif. This results in exposure of three serines on the same side of the transmembrane helices of MS-DesK, triggering a switching of the dimerization interface to allow the formation of a serine zipper. The final result is activation of the kinase state of MS-DesK.

  5. Small Leucine-Rich Proteoglycans in Renal Inflammation: Two Sides of the Coin.

    Science.gov (United States)

    Nastase, Madalina V; Janicova, Andrea; Roedig, Heiko; Hsieh, Louise Tzung-Harn; Wygrecka, Malgorzata; Schaefer, Liliana

    2018-04-01

    It is now well-established that members of the small leucine-rich proteoglycan (SLRP) family act in their soluble form, released proteolytically from the extracellular matrix (ECM), as danger-associated molecular patterns (DAMPs). By interacting with Toll-like receptors (TLRs) and the inflammasome, the two SLRPs, biglycan and decorin, autonomously trigger sterile inflammation. Recent data indicate that these SLRPs, besides their conventional role as pro-inflammatory DAMPs, additionally trigger anti-inflammatory signaling pathways to tightly control inflammation. This is brought about by selective employment of TLRs, their co-receptors, various adaptor molecules, and through crosstalk between SLRP-, reactive oxygen species (ROS)-, and sphingolipid-signaling. In this review, the complexity of SLRP signaling in immune and kidney resident cells and its relevance for renal inflammation is discussed. We propose that the dichotomy in SLRP signaling (pro- and anti-inflammatory) allows for fine-tuning the inflammatory response, which is decisive for the outcome of inflammatory kidney diseases.

  6. Partial purification of Leucine aminopeptidase (LAP) in Acromegalic Sample of Iraqi Patients

    Science.gov (United States)

    Uloom Mohammad, Taghreed

    2018-05-01

    Acromagaly is a syndrome caused by increased growth hormone secretion from the frontal lobe of the pituitary gland. A Leucine aminopeptidase (EC 34111) activity has been assayed in (30) patients sera samples(15 female and 15 males) with acromegaly age range between (3050) years and (30) sera of healthy as control group (16 femal and 14 male) age range between (3050) years. The goal of the research was partial purified of enzyme from sera patients with acromegaly by dialysis gel filtration by using sephdex G50 and ion exchange chromatography by using DEAE cellulose A50. The results showed a single peak by using gel filtration and the activity was reached to 152 U/L. Two isoenzymes were obtained by using ion exchange chromatography and the purity degree of isoenzymse (I II) were (125) and (128) fold respectively. The current study found that the enzyme showed no significant difference between the healthy and the patients.

  7. Effect of a Herbal-Leucine mix on the IL-1β-induced cartilage degradation and inflammatory gene expression in human chondrocytes

    Directory of Open Access Journals (Sweden)

    Haqqi Tariq M

    2011-08-01

    Full Text Available Abstract Background Conventional treatments for the articular diseases are often effective for symptom relief, but can also cause significant side effects and do not slow the progression of the disease. Several natural substances have been shown to be effective at relieving the symptoms of osteoarthritis (OA, and preliminary evidence suggests that some of these compounds may exert a favorable influence on the course of the disease. The objective of this study was to investigate the anti-inflammatory/chondroprotective potential of a Herbal and amino acid mixture containing extract of the Uncaria tomentosa, Boswellia spp., Lepidium meyenii and L-Leucine on the IL-1β-induced production of nitric oxide (NO, glycosaminoglycan (GAG, matrix metalloproteinases (MMPs, aggrecan (ACAN and type II collagen (COL2A1 in human OA chondrocytes and OA cartilage explants. Methods Primary OA chondrocytes or OA cartilage explants were pretreated with Herbal-Leucine mixture (HLM, 1-10 μg/ml and then stimulated with IL-1β (5 ng/ml. Effect of HLM on IL-1β-induced gene expression of iNOS, MMP-9, MMP-13, ACAN and COL2A1 was verified by real time-PCR. Estimation of NO and GAG release in culture supernatant was done using commercially available kits. Results HLM tested in these in vitro studies was found to be an effective anti-inflammatory agent, as evidenced by strong inhibition of iNOS, MMP-9 and MMP-13 expression and NO production in IL-1β-stimulated OA chondrocytes (p Leucine mixture (HLM up-regulation of ACAN and COL2A1 expression in IL-1β-stimulated OA chondrocytes was also noted (p Conclusion Our data suggests that HLM could be chondroprotective and anti-inflammatory agent in arthritis, switching chondrocyte gene expression from catabolic direction towards anabolic and regenerative, and consequently this approach may be potentially useful as a new adjunct therapeutic/preventive agent for OA or injury recovery.

  8. Anabolic effects of leucine-rich whey protein, carbohydrate, and soy protein with and without β-hydroxy-β-methylbutyrate (HMB) during fasting-induced catabolism: A human randomized crossover trial.

    Science.gov (United States)

    Rittig, Nikolaj; Bach, Ermina; Thomsen, Henrik H; Møller, Andreas B; Hansen, Jakob; Johannsen, Mogens; Jensen, Erik; Serena, Anja; Jørgensen, Jens O; Richelsen, Bjørn; Jessen, Niels; Møller, Niels

    2017-06-01

    Protein-rich beverages are widely used clinically to preserve muscle protein and improve physical performance. Beverages with high contents of leucine or its keto-metabolite β-hydroxy-β-methylbutyrate (HMB) are especially anabolic in muscle, but it is uncertain whether this also applies to catabolic conditions such as fasting and whether common or separate intracellular signaling cascades are involved. To compare a specific leucine-rich whey protein beverage (LWH) with isocaloric carbohydrate- (CHO), soy protein (SOY), and soy protein +3 g HMB (HMB) during fasting-induced catabolic conditions. Eight healthy lean male subjects underwent four interventions (LWH, CHO, SOY, and HMB) using a randomized crossover design. Each trial included a 36 h fast and consisted of a 3 h basal fasting period and a 4 h 'sipping' period. Forearm net balances of phenylalanine (NB phe , measure of net protein loss) improved for all groups (p HMB compared with SOY (p HMB have superior anabolic effects on muscle protein kinetics after 36 h of fasting, and LWH distinctly activates the mTOR pathway. These novel findings suggest that leucine-rich whey protein and/or HMB are specifically beneficial during fasting-induced catabolic conditions. Copyright © 2016 Elsevier Ltd and European Society for Clinical Nutrition and Metabolism. All rights reserved.

  9. Comprehensive Behavioral Analysis of Activating Transcription Factor 5-Deficient Mice

    Directory of Open Access Journals (Sweden)

    Mariko Umemura

    2017-07-01

    Full Text Available Activating transcription factor 5 (ATF5 is a member of the CREB/ATF family of basic leucine zipper transcription factors. We previously reported that ATF5-deficient (ATF5-/- mice demonstrated abnormal olfactory bulb development due to impaired interneuron supply. Furthermore, ATF5-/- mice were less aggressive than ATF5+/+ mice. Although ATF5 is widely expressed in the brain, and involved in the regulation of proliferation and development of neurons, the physiological role of ATF5 in the higher brain remains unknown. Our objective was to investigate the physiological role of ATF5 in the higher brain. We performed a comprehensive behavioral analysis using ATF5-/- mice and wild type littermates. ATF5-/- mice exhibited abnormal locomotor activity in the open field test. They also exhibited abnormal anxiety-like behavior in the light/dark transition test and open field test. Furthermore, ATF5-/- mice displayed reduced social interaction in the Crawley’s social interaction test and increased pain sensitivity in the hot plate test compared with wild type. Finally, behavioral flexibility was reduced in the T-maze test in ATF5-/- mice compared with wild type. In addition, we demonstrated that ATF5-/- mice display disturbances of monoamine neurotransmitter levels in several brain regions. These results indicate that ATF5 deficiency elicits abnormal behaviors and the disturbance of monoamine neurotransmitter levels in the brain. The behavioral abnormalities of ATF5-/- mice may be due to the disturbance of monoamine levels. Taken together, these findings suggest that ATF5-/- mice may be a unique animal model of some psychiatric disorders.

  10. A credit-card library approach for disrupting protein-protein interactions.

    Science.gov (United States)

    Xu, Yang; Shi, Jin; Yamamoto, Noboru; Moss, Jason A; Vogt, Peter K; Janda, Kim D

    2006-04-15

    Protein-protein interfaces are prominent in many therapeutically important targets. Using small organic molecules to disrupt protein-protein interactions is a current challenge in chemical biology. An important example of protein-protein interactions is provided by the Myc protein, which is frequently deregulated in human cancers. Myc belongs to the family of basic helix-loop-helix leucine zipper (bHLH-ZIP) transcription factors. It is biologically active only as heterodimer with the bHLH-ZIP protein Max. Herein, we report a new strategy for the disruption of protein-protein interactions that has been corroborated through the design and synthesis of a small parallel library composed of 'credit-card' compounds. These compounds are derived from a planar, aromatic scaffold and functionalized with four points of diversity. From a 285 membered library, several hits were obtained that disrupted the c-Myc-Max interaction and cellular functions of c-Myc. The IC50 values determined for this small focused library for the disruption of Myc-Max dimerization are quite potent, especially since small molecule antagonists of protein-protein interactions are notoriously difficult to find. Furthermore, several of the compounds were active at the cellular level as shown by their biological effects on Myc action in chicken embryo fibroblast assays. In light of our findings, this approach is considered a valuable addition to the armamentarium of new molecules being developed to interact with protein-protein interfaces. Finally, this strategy for disrupting protein-protein interactions should prove applicable to other families of proteins.

  11. OsLOL1, a C2C2-type zinc finger protein, interacts with OsbZIP58 to promote seed germination through the modulation of gibberellin biosynthesis in Oryza sativa.

    Science.gov (United States)

    Wu, Jiahe; Zhu, Chuanfeng; Pang, Jinhuan; Zhang, Xiangrong; Yang, Chunlin; Xia, Guixian; Tian, Yingchuan; He, Chaozu

    2014-12-01

    Seed germination is a key developmental process in the plant life cycle that is influenced by various environmental cues and phytohormones through gene expression and a series of metabolism pathways. In the present study, we investigated a C2C2-type finger protein, OsLOL1, which promotes gibberellin (GA) biosynthesis and affects seed germination in Oryza sativa (rice). We used OsLOL1 antisense and sense transgenic lines to explore OsLOL1 functions. Seed germination timing in antisense plants was restored to wild type when exogenous GA3 was applied. The reduced expression of the GA biosynthesis gene OsKO2 and the accumulation of ent-kaurene were observed during germination in antisense plants. Based on yeast two-hybrid and firefly luciferase complementation analyses, OsLOL1 interacted with the basic leucine zipper protein OsbZIP58. The results from electrophoretic mobility shift and dual-luciferase reporter assays showed that OsbZIP58 binds the G-box cis-element of the OsKO2 promoter and activates LUC reporter gene expression, and that interaction between OsLOL1 and OsbZIP58 activates OsKO2 gene expression. In addition, OsLOL1 decreased SOD1 gene expression and accelerated programmed cell death (PCD) in the aleurone layer of rice grains. These findings demonstrate that the interaction between OsLOL1 and OsbZIP58 influences GA biosynthesis through the activation of OsKO2 via OsbZIP58, thereby stimulating aleurone PCD and seed germination. © 2014 The Authors The Plant Journal © 2014 John Wiley & Sons Ltd.

  12. Cell type-specific deficiency of c-kit gene expression in mutant mice of mi/mi genotype.

    Science.gov (United States)

    Isozaki, K.; Tsujimura, T.; Nomura, S.; Morii, E.; Koshimizu, U.; Nishimune, Y.; Kitamura, Y.

    1994-01-01

    The mi locus of mice encodes a novel member of the basic-helix-loop-helix-leucine zipper protein family of transcription factors (hereafter called mi factor). In addition to microphthalmus, osteopetrosis, and lack of melanocytes, mice of mi/mi genotype are deficient in mast cells. Since the c-kit receptor tyrosine kinase plays an important role in the development of mast cells, and since the c-kit expression by cultured mast cells from mi/mi mice is deficient in both mRNA and protein levels, the mast cell deficiency of mi/mi mice has been attributed at least in part to the deficient expression of c-kit. However, it remained to be examined whether the c-kit expression was also deficient in tissues of mi/mi mice. In the present study, we examined the c-kit expression by mi/mi skin mast cells using in situ hybridization and immunohistochemistry. Moreover, we examined the c-kit expression by various cells other than mast cells in tissues of mi/mi mice. We found that the c-kit expression was deficient in mast cells but not in erythroid precursors, testicular germ cells, and neurons of mi/mi mice. This suggested that the regulation of the c-kit transcription by the mi factor was dependent on cell types. Mice of mi/mi genotype appeared to be a useful model to analyze the function of transcription factors in the whole-animal level. Images Figure 1 Figure 2 Figure 3 Figure 4 Figure 5 Figure 6 PMID:7524330

  13. Overexpression of a wheat (Triticum aestivum L.) bZIP transcription factor gene, TabZIP6, decreased the freezing tolerance of transgenic Arabidopsis seedlings by down-regulating the expression of CBFs.

    Science.gov (United States)

    Cai, Wangting; Yang, Yaling; Wang, Weiwei; Guo, Guangyan; Liu, Wei; Bi, Caili

    2018-03-01

    The basic leucine zipper (bZIP) proteins play important roles against abiotic stress in plants, including cold stress. However, most bZIPs involved in plant freezing tolerance are positive regulators. Only a few bZIPs function negatively in cold stress response. In this study, TabZIP6, a Group C bZIP transcription factor gene from common wheat (Triticum aestivum L.), was cloned and characterized. The transcript of TabZIP6 was strongly induced by cold treatment (4 °C). TabZIP6 is a nuclear-localized protein with transcriptional activation activity. Arabidopsis plants overexpressing TabZIP6 showed decreased tolerance to freezing stress. Microarray as well as quantitative real-time PCR (qRT-PCR) analysis showed that CBFs and some key COR genes, including COR47 and COR15B, were down-regulated by cold treatment in TabZIP6-overexpressing Arabidopsis lines. TabZIP6 was capable of binding to the G-box motif and the CBF1 and CBF3 promoters in yeast cells. A yeast two-hybrid assay revealed that TabZIP6, as well as the other two Group S bZIP proteins involved in cold stress tolerance in wheat, Wlip19 and TaOBF1, can form homodimers by themselves and heterodimers with each other. These results suggest that TabZIP6 may function negatively in the cold stress response by binding to the promoters of CBFs, and thereby decreasing the expression of downstream COR genes in TabZIP6-overexpressing Arabidopsis seedlings. Copyright © 2018. Published by Elsevier Masson SAS.

  14. Evaluation of the role of TAX, HBZ, and HTLV-1 proviral load on the survival of ATLL patients.

    Science.gov (United States)

    Akbarin, Mohammad Mehdi; Shirdel, Abbas; Bari, Alireza; Mohaddes, Seyedeh Tahereh; Rafatpanah, Houshang; Karimani, Ehsan Ghayour; Etminani, Kobra; Golabpour, Amin; Torshizi, Reza

    2017-06-01

    Adult T-cell leukemia/lymphoma (ATLL) is an aggressive malignancy with very poor prognosis and short survival, caused by the human T-lymphotropic virus type-1 (HTLV-1). The HTLV-1 biomarkers trans-activator x (TAX) and HTLV-1 basic leucine zipper factor (HBZ) are main oncogenes and life-threatening elements. This study aimed to assess the role of the TAX and HBZ genes and HTLV-1 proviral load (PVL) in the survival of patients with ATLL. Forty-three HTLV-1-infected individuals, including 18 asymptomatic carriers (AC) and 25 ATLL patients (ATLL), were evaluated between 2011 and 2015. The mRNA expression of TAX and HBZ and the HTLV-1 PVL were measured by quantitative PCR. Significant differences in the mean expression levels of TAX and HBZ were observed between the two study groups (ATLL and AC, P =0.014 and P =0.000, respectively). In addition, the ATLL group showed a significantly higher PVL than AC ( P =0.000). There was a significant negative relationship between PVL and survival among all study groups ( P =0.047). The HTLV-1 PVL and expression of TAX and HBZ were higher in the ATLL group than in the AC group. Moreover, a higher PVL was associated with shorter survival time among all ATLL subjects. Therefore, measurement of PVL, TAX , and HBZ may be beneficial for monitoring and predicting HTLV-1-infection outcomes, and PVL may be useful for prognosis assessment of ATLL patients. This research demonstrates the possible correlation between these virological markers and survival in ATLL patients.

  15. A minireview of E4BP4/NFIL3 in heart failure.

    Science.gov (United States)

    Velmurugan, Bharath Kumar; Chang, Ruey-Lin; Marthandam Asokan, Shibu; Chang, Chih-Fen; Day, Cecilia-Hsuan; Lin, Yueh-Min; Lin, Yuan-Chuan; Kuo, Wei-Wen; Huang, Chih-Yang

    2018-06-01

    Heart failure (HF) remains a major cause of morbidity and mortality worldwide. The primary cause identified for HF is impaired left ventricular myocardial function, and clinical manifestations may lead to severe conditions like pulmonary congestion, splanchnic congestion, and peripheral edema. Development of new therapeutic strategies remains the need of the hour for controlling the problem of HF worldwide. Deeper insights into the molecular mechanisms involved in etiopathology of HF indicate the significant role of calcium signaling, autocrine signaling pathways, and insulin-like growth factor-1 signaling that regulates the physiologic functions of heart growth and development such as contraction, metabolism, hypertrophy, cytokine signaling, and apoptosis. In view of these facts, a transcription factor (TF) regulating the myriad of these signaling pathways may prove as a lead candidate for development of therapeutics. Adenovirus E4 promoter-binding protein (E4BP4), also known as nuclear-factor, interleukin 3 regulated (NFIL3), a type of basic leucine zipper TF, is known to regulate the signaling processes involved in the functioning of heart. The current review discusses about the expression, structure, and functional role of E4BP4 in signaling processes with emphasis on calcium signaling mechanisms, autocrine signaling, and insulin-like growth factor II receptor-mediated processes regulated by E4BP4 that may regulate the pathogenesis of HF. We propose that E4BP4, being the critical component for the regulation of the above signaling processes, may serve as a novel therapeutic target for HF, and scientific investigations are merited in this direction. © 2018 Wiley Periodicals, Inc.

  16. The Effects of Leucine, Zinc, and Chromium Supplements on Inflammatory Events of the Respiratory System in Type 2 Diabetic Rats

    OpenAIRE

    Kolahian, Saeed; Sadri, Hassan; Shahbazfar, Amir Ali; Amani, Morvarid; Mazadeh, Anis; Mirani, Mehdi

    2015-01-01

    Diabetes mellitus is a major cause of serious micro- and macrovascular diseases that affect nearly every system in the body, including the respiratory system. Non-enzymatic protein glycation due to hyperglycaemic stress has fundamental implications due to the large capillary network and amount of connective tissue in the lung. The current study was designed to determine whether leucine, zinc, and chromium supplementations influence the function and histological structure of the respiratory tr...

  17. MicroRNA-193b-3p acts as a tumor suppressor by targeting the MYB oncogene in T-cell acute lymphoblastic leukemia.

    Science.gov (United States)

    Mets, E; Van der Meulen, J; Van Peer, G; Boice, M; Mestdagh, P; Van de Walle, I; Lammens, T; Goossens, S; De Moerloose, B; Benoit, Y; Van Roy, N; Clappier, E; Poppe, B; Vandesompele, J; Wendel, H-G; Taghon, T; Rondou, P; Soulier, J; Van Vlierberghe, P; Speleman, F

    2015-04-01

    The MYB oncogene is a leucine zipper transcription factor essential for normal and malignant hematopoiesis. In T-cell acute lymphoblastic leukemia (T-ALL), elevated MYB levels can arise directly through T-cell receptor-mediated MYB translocations, genomic MYB duplications or enhanced TAL1 complex binding at the MYB locus or indirectly through the TAL1/miR-223/FBXW7 regulatory axis. In this study, we used an unbiased MYB 3'untranslated region-microRNA (miRNA) library screen and identified 33 putative MYB-targeting miRNAs. Subsequently, transcriptome data from two independent T-ALL cohorts and different subsets of normal T-cells were used to select miRNAs with relevance in the context of normal and malignant T-cell transformation. Hereby, miR-193b-3p was identified as a novel bona fide tumor-suppressor miRNA that targets MYB during malignant T-cell transformation thereby offering an entry point for efficient MYB targeting-oriented therapies for human T-ALL.

  18. Abscisic acid signaling is controlled by a BRANCHED1/HD-ZIP I cascade in Arabidopsis axillary buds.

    Science.gov (United States)

    González-Grandío, Eduardo; Pajoro, Alice; Franco-Zorrilla, José M; Tarancón, Carlos; Immink, Richard G H; Cubas, Pilar

    2017-01-10

    Shoot-branching patterns determine key aspects of plant life and are important targets for crop breeding. However, we are still largely ignorant of the genetic networks controlling locally the most important decision during branch development: whether the axillary bud, or branch primordium, grows out to give a lateral shoot or remains dormant. Here we show that, inside the buds, the TEOSINTE BRANCHED1, CYCLOIDEA, PCF (TCP) transcription factor BRANCHED1 (BRC1) binds to and positively regulates the transcription of three related Homeodomain leucine zipper protein (HD-ZIP)-encoding genes: HOMEOBOX PROTEIN 21 (HB21), HOMEOBOX PROTEIN 40 (HB40), and HOMEOBOX PROTEIN 53 (HB53). These three genes, together with BRC1, enhance 9-CIS-EPOXICAROTENOID DIOXIGENASE 3 (NCED3) expression, lead to abscisic acid accumulation, and trigger hormone response, thus causing suppression of bud development. This TCP/HD-ZIP genetic module seems to be conserved in dicot and monocotyledonous species to prevent branching under light-limiting conditions.

  19. GILZ Promotes Production of Peripherally Induced Treg Cells and Mediates the Crosstalk between Glucocorticoids and TGF-β Signaling

    Directory of Open Access Journals (Sweden)

    Oxana Bereshchenko

    2014-04-01

    Full Text Available Regulatory T (Treg cells expressing the transcription factor forkhead box P3 (FoxP3 control immune responses and prevent autoimmunity. Treatment with glucocorticoids (GCs has been shown to increase Treg cell frequency, but the mechanisms of their action on Treg cell induction are largely unknown. Here, we report that glucocorticoid-induced leucine zipper (GILZ, a protein induced by GCs, promotes Treg cell production. In mice, GILZ overexpression causes an increase in Treg cell number, whereas GILZ deficiency results in impaired generation of peripheral Treg cells (pTreg, associated with increased spontaneous and experimental intestinal inflammation. Mechanistically, we found that GILZ is required for GCs to cooperate with TGF-β in FoxP3 induction, while it enhances TGF-β signaling by binding to and promoting Smad2 phosphorylation and activation of FoxP3 expression. Thus, our results establish an essential GILZ-mediated link between the anti-inflammatory action of GCs and the regulation of TGF-β-dependent pTreg production.

  20. Leucine Biosynthesis Is Involved in Regulating High Lipid Accumulation in Yarrowia lipolytica

    Energy Technology Data Exchange (ETDEWEB)

    Kerkhoven, Eduard J.; Kim, Young-Mo; Wei, Siwei; Nicora, Carrie D.; Fillmore, Thomas L.; Purvine, Samuel O.; Webb-Robertson, Bobbie-Jo; Smith, Richard D.; Baker, Scott E.; Metz, Thomas O.; Nielsen, Jens; Lee, Sang Yup

    2017-06-20

    ABSTRACT

    The yeastYarrowia lipolyticais a potent accumulator of lipids, and lipogenesis in this organism can be influenced by a variety of factors, such as genetics and environmental conditions. Using a multifactorial study, we elucidated the effects of both genetic and environmental factors on regulation of lipogenesis inY. lipolyticaand identified how two opposite regulatory states both result in lipid accumulation. This study involved comparison of a strain overexpressing diacylglycerol acyltransferase (DGA1) with a control strain grown under either nitrogen or carbon limitation conditions. A strong correlation was observed between the responses on the transcript and protein levels. Combination ofDGA1overexpression with nitrogen limitation resulted in a high level of lipid accumulation accompanied by downregulation of several amino acid biosynthetic pathways, including that of leucine in particular, and these changes were further correlated with a decrease in metabolic fluxes. This downregulation was supported by the measured decrease in the level of 2-isopropylmalate, an intermediate of leucine biosynthesis. Combining the multi-omics data with putative transcription factor binding motifs uncovered a contradictory role for TORC1 in controlling lipid accumulation, likely mediated through 2-isopropylmalate and a Leu3-like transcription factor.

    IMPORTANCEThe ubiquitous metabolism of lipids involves refined regulation, and an enriched understanding of this regulation would have wide implications. Various factors can influence lipid metabolism, including the environment and genetics. We demonstrated, using a multi-omics and multifactorial experimental setup, that multiple factors affect lipid accumulation in the yeastYarrowia lipolytica. Using integrative analysis, we identified novel interactions between nutrient restriction and genetic factors

  1. Influence of l-Leucine and l-Alanine on Lrp Regulation of foo, Coding for F1651, a Pap Homologue

    OpenAIRE

    Berthiaume, Frédéric; Crost, Cécile; Labrie, Vincent; Martin, Christine; Newman, Elaine B.; Harel, Josée

    2004-01-01

    The foo operon encodes F1651 fimbriae that belong to the P-regulatory family and are synthesized by septicemic Escherichia coli. Using an Lrp-deficient host and the lrp gene cloned under the arabinose pBAD promoter, we demonstrated that foo was transcribed proportionally to the amount of Lrp synthesized. l-Leucine and l-alanine decreased drastically the steady-state transcription of foo and modified phase variation, independently of the presence of FooI. Specific mutations in the C-terminal r...

  2. Leucine-rich repeat-containing synaptic adhesion molecules as organizers of synaptic specificity and diversity.

    Science.gov (United States)

    Schroeder, Anna; de Wit, Joris

    2018-04-09

    The brain harbors billions of neurons that form distinct neural circuits with exquisite specificity. Specific patterns of connectivity between distinct neuronal cell types permit the transfer and computation of information. The molecular correlates that give rise to synaptic specificity are incompletely understood. Recent studies indicate that cell-surface molecules are important determinants of cell type identity and suggest that these are essential players in the specification of synaptic connectivity. Leucine-rich repeat (LRR)-containing adhesion molecules in particular have emerged as key organizers of excitatory and inhibitory synapses. Here, we discuss emerging evidence that LRR proteins regulate the assembly of specific connectivity patterns across neural circuits, and contribute to the diverse structural and functional properties of synapses, two key features that are critical for the proper formation and function of neural circuits.

  3. Leucine-Enriched Essential Amino Acids Augment Mixed Protein Synthesis, But Not Collagen Protein Synthesis, in Rat Skeletal Muscle after Downhill Running

    OpenAIRE

    Kato, Hiroyuki; Suzuki, Hiromi; Inoue, Yoshiko; Suzuki, Katsuya; Kobayashi, Hisamine

    2016-01-01

    Mixed and collagen protein synthesis is elevated for as many as 3 days following exercise. Immediately after exercise, enhanced amino acid availability increases synthesis of mixed muscle protein, but not muscle collagen protein. However, the potential for synergic effects of amino acid ingestion with exercise on both mixed and collagen protein synthesis remains unclear. We investigated muscle collagen protein synthesis in rats following post-exercise ingestion of leucine-enriched essential a...

  4. Isolation and characterization of cDNA encoding the 80-kDa subunit protein of the human autoantigen Ku (p70/p80) recognized by autoantibodies from patients with scleroderma-polymyositis overlap syndrome

    International Nuclear Information System (INIS)

    Mimori, Tsuneyo; Ohosone, Yasuo; Hama, Nobuaki; Suwa, Akira; Akizuki, Masashi; Homma, Mitsuo; Griffith, A.J.; Hardin, J.A.

    1990-01-01

    Anti-Ku (p70/p80) autoantibodies in patients with scleroderma-polymyositis overlap syndrome recognize a 70-kDa/80-kDa protein heterodimer which binds to terminal regions of double-stranded DNA. In the present study, the authors isolated full-length cDNAs that encode the 80-kDa Ku subunit. Initial screening of a human spleen cDNA library with anti-Ku antibodies yielded a cDNA of 1.0 kilobase (kb) (termed K71) encoding a portion of the 80-kDa Ku polypeptide (identification based on immunological criteria). In RNA blots, this cDNA hybridized with two mRNAs of 3.4 and 2.6 kb. In vitro transcription and translation experiments produced an immunoprecipitable polypeptide which comigrated with the 80-kDa Ku subunit. The Ku80-6 cDNA proved to be 3304 nucleotides in length, with an additional poly(A) tail, closely approximating the size of the larger mRNA. It contains a single long open reading frame encoding 732 amino acids. The putative polypeptide has a high content of acidic amino acids and a region with periodic repeat of leucine in every seventh position which may form the leucine zipper structure. In genomic DNA blots, probes derived from the opposite ends of cDNA Ku80-6 hybridized with several nonoverlapping restriction fragments from human leukocyte DNA, indicating that the gene encoding the 80-kDa Ku polypeptide is divided into several exons by intervening sequences

  5. Leucine-Enriched Essential Amino Acids Augment Mixed Protein Synthesis, But Not Collagen Protein Synthesis, in Rat Skeletal Muscle after Downhill Running

    Science.gov (United States)

    Kato, Hiroyuki; Suzuki, Hiromi; Inoue, Yoshiko; Suzuki, Katsuya; Kobayashi, Hisamine

    2016-01-01

    Mixed and collagen protein synthesis is elevated for as many as 3 days following exercise. Immediately after exercise, enhanced amino acid availability increases synthesis of mixed muscle protein, but not muscle collagen protein. However, the potential for synergic effects of amino acid ingestion with exercise on both mixed and collagen protein synthesis remains unclear. We investigated muscle collagen protein synthesis in rats following post-exercise ingestion of leucine-enriched essential amino acids. We determined fractional protein synthesis rates (FSR) at different time points following exercise. Mixed protein and collagen protein FSRs in skeletal muscle were determined by measuring protein-bound enrichments of hydroxyproline and proline, and by measuring the intracellular enrichment of proline, using injections of flooding d3-proline doses. A leucine-enriched mixture of essential amino acids (or distilled water as a control) was administrated 30 min or 1 day post-exercise. The collagen protein synthesis in the vastus lateralis was elevated for 2 days after exercise. Although amino acid administration did not increase muscle collagen protein synthesis, it did lead to augmented mixed muscle protein synthesis 1 day following exercise. Thus, contrary to the regulation of mixed muscle protein synthesis, muscle collagen protein synthesis is not affected by amino acid availability after damage-inducing exercise. PMID:27367725

  6. Leucine-Enriched Essential Amino Acids Augment Mixed Protein Synthesis, But Not Collagen Protein Synthesis, in Rat Skeletal Muscle after Downhill Running

    Directory of Open Access Journals (Sweden)

    Hiroyuki Kato

    2016-06-01

    Full Text Available Mixed and collagen protein synthesis is elevated for as many as 3 days following exercise. Immediately after exercise, enhanced amino acid availability increases synthesis of mixed muscle protein, but not muscle collagen protein. However, the potential for synergic effects of amino acid ingestion with exercise on both mixed and collagen protein synthesis remains unclear. We investigated muscle collagen protein synthesis in rats following post-exercise ingestion of leucine-enriched essential amino acids. We determined fractional protein synthesis rates (FSR at different time points following exercise. Mixed protein and collagen protein FSRs in skeletal muscle were determined by measuring protein-bound enrichments of hydroxyproline and proline, and by measuring the intracellular enrichment of proline, using injections of flooding d3-proline doses. A leucine-enriched mixture of essential amino acids (or distilled water as a control was administrated 30 min or 1 day post-exercise. The collagen protein synthesis in the vastus lateralis was elevated for 2 days after exercise. Although amino acid administration did not increase muscle collagen protein synthesis, it did lead to augmented mixed muscle protein synthesis 1 day following exercise. Thus, contrary to the regulation of mixed muscle protein synthesis, muscle collagen protein synthesis is not affected by amino acid availability after damage-inducing exercise.

  7. New poly(ester urea) derived from L-leucine: Electrospun scaffolds loaded with antibacterial drugs and enzymes

    Energy Technology Data Exchange (ETDEWEB)

    Díaz, Angélica; Valle, Luis J. del [Departament d' Enginyeria Química, Universitat Politècnica de Catalunya, Av. Diagonal 647, Barcelona E-08028 (Spain); Tugushi, David; Katsarava, Ramaz [Institute of Chemistry and Molecular Engineering, Agricultural University of Georgia, 13 km. David Aghmashenebeli Alley, Tblisi 0131, Georgia (United States); Puiggalí, Jordi, E-mail: Jordi.Puiggali@upc.edu [Departament d' Enginyeria Química, Universitat Politècnica de Catalunya, Av. Diagonal 647, Barcelona E-08028 (Spain)

    2015-01-01

    Electrospun scaffolds from an amino acid containing poly(ester urea) (PEU) were developed as promising materials in the biomedical field and specifically in tissue engineering applications. The selected poly(ester urea) was obtained with a high yield and molecular weight by reaction of phosgene with a bis(α-aminoacyl)-α,ω-diol-diester monomer. The polymer having L-leucine, 1,6-hexanediol and carbonic acid units had a semicrystalline character and relatively high glass transition and melting temperatures. Furthermore it was highly soluble in most organic solvents, an interesting feature that facilitated the electrospinning process and the effective incorporation of drugs with bactericidal activity (e.g. biguanide derivatives such as clorhexidine and polyhexamethylenebiguanide) and enzymes (e.g. α-chymotrypsin) that accelerated the degradation process. Continuous micro/nanofibers were obtained under a wide range of processing conditions, being diameters of electrospun fibers dependent on the drug and solvent used. Poly(ester urea) samples were degradable in media containing lipases and proteinases but the degradation rate was highly dependent on the surface area, being specifically greater for scaffolds with respect to films. The high hydrophobicity of new scaffolds had repercussions on enzymatic degradability since different weight loss rates were found depending on how samples were exposed to the medium (e.g. forced or non-forced immersion). New scaffolds were biocompatible, as demonstrated by adhesion and proliferation assays performed with fibroblast and epithelial cells. - Highlights: • Electrospun scaffolds from a biodegradable poly(ester urea) have been prepared. • Scaffolds were effectively loaded with bactericide agents. • Enzymatic degradability of the L-leucine derived poly(ester urea) was demonstrated. • Enzymes that accelerate degradation were incorporated in the electrospun fibers. • Cell adhesion/proliferation assays demonstrated

  8. The CD3 gamma leucine-based receptor-sorting motif is required for efficient ligand-mediated TCR down-regulation

    DEFF Research Database (Denmark)

    von Essen, Marina; Menné, Charlotte; Nielsen, Bodil L

    2002-01-01

    . The other pathway is dependent on protein kinase C (PKC)-mediated activation of the CD3 gamma di-leucine-based receptor-sorting motif. Previous studies have failed to demonstrate a connection between ligand- and PKC-induced TCR down-regulation. Thus, although an apparent paradox, the dogma has been...... that ligand- and PKC-induced TCR down-regulations are not interrelated. By analyses of a newly developed CD3 gamma-negative T cell variant, freshly isolated and PHA-activated PBMC, and a mouse T cell line, we challenged this dogma and demonstrate in this work that PKC activation and the CD3 gamma di...

  9. Effect of leucine-to-methionine substitutions on the diffraction quality of histone chaperone SET/TAF-Ibeta/INHAT crystals.

    Science.gov (United States)

    Senda, Miki; Muto, Shinsuke; Horikoshi, Masami; Senda, Toshiya

    2008-10-01

    One of the most frequent problems in crystallization is poor quality of the crystals. In order to overcome this obstacle several methods have been utilized, including amino-acid substitutions of the target protein. Here, an example is presented of crystal-quality improvement by leucine-to-methionine substitutions. A variant protein with three amino-acid substitutions enabled improvement of the crystal quality of the histone chaperone SET/TAF-Ibeta/INHAT when combined with optimization of the cryoconditions. This procedure improved the resolution of the SET/TAF-Ibeta/INHAT crystals from around 5.5 to 2.3 A without changing the crystallization conditions.

  10. A virtual high-throughput screening approach to the discovery of novel inhibitors of the bacterial leucine transporter, LeuT

    DEFF Research Database (Denmark)

    Simmons, Katie J; Gotfryd, Kamil; Billesbølle, Christian B

    2013-01-01

    Abstract Membrane proteins are intrinsically involved in both human and pathogen physiology, and are the target of 60% of all marketed drugs. During the past decade, advances in the studies of membrane proteins using X-ray crystallography, electron microscopy and NMR-based techniques led to the e...... this is a virtual high-throughput screening (vHTS) technique initially developed for soluble proteins. This paper describes application of this technique to the discovery of inhibitors of the leucine transporter (LeuT), a member of the neurotransmitter:sodium symporter (NSS) family....

  11. Similar effects of leucine rich and regular dairy products on muscle mass and functions of older polymyalgia rheumatica patients: a randomized crossover trial.

    Science.gov (United States)

    Björkman, M P; Pilvi, T K; Kekkonen, R A; Korpela, R; Tilvis, R S

    2011-06-01

    Leucine-rich milk and whey proteins have been suggested for prevention of age related loss of muscle mass and strength i.e. sarcopenia. The effects of milk protein supplementation and low intensity home based physical exercise on body composition and muscle functions were investigated. A randomized double blind crossover trial. Community dwelling members of Helsinki rheumatoid association. Older people (N=47, mean age 69.5 years) suffering from polymyalgia rheumatica. Patients performed as many stand ups as possible twice a day after which they ingested a regular (Control) or a whey protein enriched dairy product with high leucine content (Test). The 8-week intervention periods were separated by a 4-week wash-out. Body composition was measured by dual x-ray absorptiometry and muscle functions by hand grip strength, force platform countermovement jump performance, chair stand test, and walking speed. The 16-week home-based post-exercise supplementation resulted in a 1.8% increase (p = 0.052) in lower limb muscle mass. Walking speed (+5.3%, p = 0.007) and chair stand test performance (-12.2 %, p products prevent or treat age-associated sarcopenia and whether they are superior to the present commercial milk products.

  12. Effect of inhibitory avoidance trainning, ACTH, beta-endorphin and adrenaline on the incorporation of 14C-leucine into synaptosomal proteins of rat hypothalamus, amygdala and hippocampus

    International Nuclear Information System (INIS)

    Dalmaz, C.; Maia, H.M.M.; Izquierdo, I.

    1986-01-01

    'In vitro' incorporation of leucine to protein was studied in synaptosomes isolated from the hypothalamus, amygdala and hippocampus of rats submitted to inhibitory avoidance training or to the i.p. injection of ACTH, beta-endorphin or adrenaline; or in synaptosomes incubated with these substances. (M.A.C.) [pt

  13. Hygiene Basics

    Science.gov (United States)

    ... Staying Safe Videos for Educators Search English Español Hygiene Basics KidsHealth / For Teens / Hygiene Basics What's in this article? Oily Hair Sweat ... smell, anyway? Read below for information on some hygiene basics — and learn how to deal with greasy ...

  14. Experimental and modeling studies on the enantio-separation of 3,5-dinitrobenzoyl-(R),(S)-leucine by continuous liquid-liquid extraction in a cascade of centrifugal contactor separators

    NARCIS (Netherlands)

    Schuur, Boelo; Winkelman, Jozef G.M.; Vries, Johannes G. de; Heeres, Hero J.

    2010-01-01

    The continuous enantioselective liquid–liquid extraction of aqueous 3,5-dinitrobenzoyl-(R),(S)-leucine (AR,S) using O-(1-t-butylcarbamoyl)-11-octadecylsulfinyl-10,11-dihydro-quinine (C,a cinchona alkaloid) as extractant in 1,2-dichloroethane (DCE) was studied experimentally in a countercurrently

  15. Native whey protein with high levels of leucine results in similar post-exercise muscular anabolic responses as regular whey protein: a randomized controlled trial.

    Science.gov (United States)

    Hamarsland, Håvard; Nordengen, Anne Lene; Nyvik Aas, Sigve; Holte, Kristin; Garthe, Ina; Paulsen, Gøran; Cotter, Matthew; Børsheim, Elisabet; Benestad, Haakon B; Raastad, Truls

    2017-01-01

    Protein intake is essential to maximally stimulate muscle protein synthesis, and the amino acid leucine seems to possess a superior effect on muscle protein synthesis compared to other amino acids. Native whey has higher leucine content and thus a potentially greater anabolic effect on muscle than regular whey (WPC-80). This study compared the acute anabolic effects of ingesting 2 × 20 g of native whey protein, WPC-80 or milk protein after a resistance exercise session. A total of 24 young resistance trained men and women took part in this double blind, randomized, partial crossover, controlled study. Participants received either WPC-80 and native whey ( n  = 10), in a crossover design, or milk ( n  = 12). Supplements were ingested immediately (20 g) and two hours after (20 g) a bout of heavy-load lower body resistance exercise. Blood samples and muscle biopsies were collected to measure plasma concentrations of amino acids by gas-chromatography mass spectrometry, muscle phosphorylation of p70S6K, 4E-BP1 and eEF-2 by immunoblotting, and mixed muscle protein synthesis by use of [ 2 H 5 ]phenylalanine-infusion, gas-chromatography mass spectrometry and isotope-ratio mass spectrometry. Being the main comparison, differences between native whey and WPC-80 were analysed by a one-way ANOVA and comparisons between the whey supplements and milk were analysed by a two-way ANOVA. Native whey increased blood leucine concentrations more than WPC-80 and milk ( P  whey ingestion induced a greater phosphorylation of p70S6K than milk 180 min after exercise ( P  = 0.03). Muscle protein synthesis rates increased 1-3 h hours after exercise with WPC-80 (0.119%), and 1-5 h after exercise with native whey (0.112%). Muscle protein synthesis rates were higher 1-5 h after exercise with native whey than with milk (0.112% vs. 0.064, P  = 0.023). Despite higher-magnitude increases in blood leucine concentrations with native whey, it was not superior to WPC-80

  16. A high whey protein-, leucine-, and vitamin D-enriched supplement preserves muscle mass during intentional weight loss in obese older adults: a double-blind randomized controlled trial

    NARCIS (Netherlands)

    Verreijen, A.M.; Verlaan, S.; Engberink, M.F.; Swinkels, S.; Bosch, J.; Weijs, P.J.M.

    2015-01-01

    Background: Intentional weight loss in obese older adults is a risk factor for muscle loss and sarcopenia. Objective: The objective was to examine the effect of a high whey protein-, leucine-, and vitamin D-enriched supplement on muscle mass preservation during intentional weight loss in obese older

  17. Basic electrotechnology

    CERN Document Server

    Ashen, R A

    2013-01-01

    BASIC Electrotechnology discusses the applications of Beginner's All-purpose Symbolic Instruction Code (BASIC) in engineering, particularly in solving electrotechnology-related problems. The book is comprised of six chapters that cover several topics relevant to BASIC and electrotechnology. Chapter 1 provides an introduction to BASIC, and Chapter 2 talks about the use of complex numbers in a.c. circuit analysis. Chapter 3 covers linear circuit analysis with d.c. and sinusoidal a.c. supplies. The book also discusses the elementary magnetic circuit theory. The theory and performance of two windi

  18. Differentiating leucine incorporation of Archaea and Bacteria throughout the water column of the eastern Atlantic using metabolic inhibitors

    OpenAIRE

    Yokokawa, T.; Sintes, E.; de Corte, D.; Olbrich, K.; Herndl, G.J.

    2012-01-01

    The abundance (based on catalyzed reporter deposition-fluorescence in situ hybrid ization, CARD-FISH) and leucine incorporation rates of Archaea and Bacteria were determined throughout the water column in the eastern Atlantic. Bacteria dominated throughout the water column, although their contribution to total prokaryotic abundance in the bathypelagic layer (1000 to 4000 m depth) was lower than in the surface and mesopelagic layers (0 to 1000 m depth). While marine Crenarchaeota Group I (MCG ...

  19. Effects of steroids and sex reversal on intestinal absorption of L-[14C]leucine in vivo, in rainbow trout, Salmo gairdneri

    International Nuclear Information System (INIS)

    Habibi, H.R.; Ince, B.W.

    1983-01-01

    The effects of steroids (17 alpha-methyltestosterone (MT), 17 beta-oestradiol (E2)), and of sex reversal (XX male) on intestinal absorption and accumulation of L-[ 14 C]leucine (5 mM), were investigated in unanaesthetized rainbow trout (Salmo gairdneri), using an in vivo gut perfusion technique. Each steroid was luminally perfused through the gut at a concentration of 50 micrograms/ml perfusate, during five separate perfusions carried out on the same fish at 30-min intervals (perfusion periods 1 to 5), for a total of 120 min at 14 degrees. Experiments were also conducted on masculinized, genetically female trout (XX male) with steroid-free perfusate. MT treatment significantly increased the intestinal absorption of radioleucine during periods 1 and 2, whilst E2 was without effect. Neither MT nor E2 influenced intestinal accumulation (mid- and hindgut) of radioleucine, and accumulation of 14 C-solutes in skeletal muscle. Sex reversal, however, whilst having no effect on leucine absorption, nevertheless significantly increased intestinal accumulation of radioleucine, and accumulation of 14 C-solutes in skeletal muscle. The effects observed in the present study are in agreement with previous work in trout using everted gut sac preparations. It is suggested that the growth-promoting effects of anabolic-androgenic steroids in fish may be partly explained by their action on gastrointestinal function

  20. Targeting Inflammation and Downstream Protein Metabolism in Sarcopenia: A Brief Up-Dated Description of Concurrent Exercise and Leucine-Based Multimodal Intervention

    Directory of Open Access Journals (Sweden)

    Zhi Xia

    2017-06-01

    Full Text Available Sarcopenia is defined as the progressive loss of muscle mass with age, and poses a serious threat to the physiological and psychological health of the elderly population with consequential economic and social burdens. Chronic low-grade inflammation plays a central role in the development of sarcopenia such that it alters cellular protein metabolism to favor proteolysis over synthesis, and thereby accelerates muscular atrophy. The purpose of this review is to highlight how exercise and nutrition intervention strategies can attenuate or treat sarcopenia. Resistance exercise increases not only muscle mass but also muscle strength, while aerobic exercise is able to ameliorate the age-related metabolic disorders. Concurrent exercise training integrates the advantages of both aerobic and resistance exercise, and may exert a significant synergistic effect in the aging organism. Higher protein intakes rich in the amino acid leucine appear to restore skeletal muscle protein metabolism balance by rescuing protein synthesis in older adults. There is good reason to believe that a multimodal treatment, a combination of exercise and increased leucine consumption in the diet, can combat some of the muscle loss associated with aging. Future research is needed to consolidate these findings to humans, and to further clarify to what extent and by which mechanisms protein metabolism might be directly involved in sarcopenia pathogenesis and the multimodal treatment responses.

  1. A Quantitative Tool to Distinguish Isobaric Leucine and Isoleucine Residues for Mass Spectrometry-Based De Novo Monoclonal Antibody Sequencing

    Science.gov (United States)

    Poston, Chloe N.; Higgs, Richard E.; You, Jinsam; Gelfanova, Valentina; Hale, John E.; Knierman, Michael D.; Siegel, Robert; Gutierrez, Jesus A.

    2014-07-01

    De novo sequencing by mass spectrometry (MS) allows for the determination of the complete amino acid (AA) sequence of a given protein based on the mass difference of detected ions from MS/MS fragmentation spectra. The technique relies on obtaining specific masses that can be attributed to characteristic theoretical masses of AAs. A major limitation of de novo sequencing by MS is the inability to distinguish between the isobaric residues leucine (Leu) and isoleucine (Ile). Incorrect identification of Ile as Leu or vice versa often results in loss of activity in recombinant antibodies. This functional ambiguity is commonly resolved with costly and time-consuming AA mutation and peptide sequencing experiments. Here, we describe a set of orthogonal biochemical protocols, which experimentally determine the identity of Ile or Leu residues in monoclonal antibodies (mAb) based on the selectivity that leucine aminopeptidase shows for n-terminal Leu residues and the cleavage preference for Leu by chymotrypsin. The resulting observations are combined with germline frequencies and incorporated into a logistic regression model, called Predictor for Xle Sites (PXleS) to provide a statistical likelihood for the identity of Leu at an ambiguous site. We demonstrate that PXleS can generate a probability for an Xle site in mAbs with 96% accuracy. The implementation of PXleS precludes the expression of several possible sequences and, therefore, reduces the overall time and resources required to go from spectra generation to a biologically active sequence for a mAb when an Ile or Leu residue is in question.

  2. Leucine-enkephalin-like immunoreactivity is localized in luteinizing hormone-producing cells in the axolotl (Ambystoma mexicanum) pituitary.

    Science.gov (United States)

    Suzuki, Hirohumi; Yamamoto, Toshiharu

    2014-02-01

    In this study, we used immunohistochemical techniques to determine the cell type of leucine-enkephalin (Leu-ENK)-immunoreactive cells in the axolotl (Ambystoma mexicanum) pituitary. Immunoreactive cells were scattered throughout the pars distalis except for the dorso-caudal portion. These cells were immuno-positive for luteinizing hormone (LH), but they were immuno-negative for adrenocorticotrophic, growth, and thyroid-stimulating hormones, as well as prolactin. Immunoelectron microscopy demonstrated that Leu-ENK-like substance and LH co-localized within the same secretory granules. Leu-ENK secreted from gonadotrophs may participate in LH secretion in an autocrine fashion, and/or may participate in the release of sex steroids together with LH. Copyright © 2013 Elsevier Ltd. All rights reserved.

  3. Altered metabolic incorporation of fucose and leucine into PNS myelin of 25-week-old diabetic (C57BL/Ks [db/db]) mice: effects of untreated diabetes on nerve metabolism

    International Nuclear Information System (INIS)

    Chez, M.G.; Peterson, R.G.

    1983-01-01

    Sciatic nerves of 25-week-old genetically diabetic (C57BL/Ks [db/db]) mice and their litter-mate controls were removed, and their metabolic incorporation of [ 3 H]fucose and [ 14 C]leucine into myelin was studied in vitro. Untreated diabetic animals showed significant increases (p less than 0.05) in the fucose/leucine incorporation into myelin when compared to values found for their litter-mates. These results correlated well with previous experiments performed on alloxan or streptozotocin-diabetic rats and thus show the in vitro incubation procedure to be a good indicator of altered metabolic conditions in peripheral nerves due to diabetes mellitus. The resulting ratio increases seen in diabetic animals is at variance with the decrease in ratios found in animals undergoing typical Wallerian degeneration. These results suggest that different metabolic processes operate in untreated diabetics than in normals or in those undergoing other degenerative nerve processes

  4. Signal peptide cleavage is essential for surface expression of a regulatory T cell surface protein, leucine rich repeat containing 32 (LRRC32)

    OpenAIRE

    Chan, Derek V; Somani, Ally-Khan; Young, Andrew B; Massari, Jessica V; Ohtola, Jennifer; Sugiyama, Hideaki; Garaczi, Edina; Babineau, Denise; Cooper, Kevin D; McCormick, Thomas S

    2011-01-01

    Abstract Background Elevated numbers of regulatory T cells (Tregs) have been implicated in certain cancers. Depletion of Tregs has been shown to increase anti-tumor immunity. Tregs also play a critical role in the suppression of autoimmune responses. The study of Tregs has been hampered by a lack of adequate surface markers. Leucine Rich Repeat Containing 32 (LRRC32), also known as Glycoprotein A Repetitions Predominant (GARP), has been postulated as a novel surface marker of activated Tregs....

  5. Nuclear export of cutaneous HPV8 E7 oncoprotein is mediated by a leucine-rich nuclear export signal via a CRM1 pathway

    Energy Technology Data Exchange (ETDEWEB)

    Onder, Zeynep; Chang, Vivian; Moroianu, Junona, E-mail: moroianu@bc.edu

    2015-01-01

    We recently determined that the nuclear import of cutaneous beta genus HPV8 E7 oncoprotein it is mediated by its zinc-binding domain via direct hydrophobic interactions with the FG nucleoporins Nup62 and Nup153 (Onder and Moroianu, 2014). Here we investigated the nuclear export of HPV8 E7 oncoprotein using confocal microscopy after transfections of HeLa cells with EGFP–8cE7 and mutant plasmids and treatment with Ratjadone A nuclear export inhibitor. We determined that HPV8 E7 contains a leucine-rich nuclear export signal (NES), {sub 76}IRTFQELLF{sub 84}, within its zinc-binding domain that mediates its nuclear export via a CRM1 pathway. We found that HPV8 E7 interacts with CRM1 and that the hydrophobic amino acid residues I76, F79 and L82 of the NES are essential for this interaction and for nuclear export of HPV8 E7 oncoprotein. - Highlights: • HPV8 E7 has a leucine-rich NES within its zinc-binding domain that mediates its nuclear export. • CRM1 nuclear export receptor interacts with HPV8 E7 and mediates its export. • Identification of the critical hydrophobic amino acids of the NES of HPV8 E7.

  6. Solid-state conformation of copolymers of ß-benzyl-L-aspartate with L-alanine, L-leucine, L-valine, γ-benzyl-L-glutamate, or ε-carbobenzoxy-L-lysine

    NARCIS (Netherlands)

    Sederel, Willem L.; Bantjes, Adriaan; Feijen, Jan; Anderson, James M.

    1980-01-01

    The solid-state conformation of copolymers of ß-benzyl-L-aspartate [L-Asp(OBzl)] with L-leucine (L-Leu), L-alanine (L-Ala), L-valine (L-Val), γ-benzyl-L-glutamate [L-Glu(OBzl)], or ε-carbobenzoxy-L-lysine (Cbz-L-Lys) has been studied by ir spectroscopy and circular dichroism (CD). The ir spectra in

  7. Volumetric study of aqueous proline-leucine solutions from (283.15 to 318.15) K

    International Nuclear Information System (INIS)

    Mendonca, Angela F.S.S.; Barbas, Maria Joao A.; Freitas, Jose M.; Lampreia, Isabel M.S.

    2004-01-01

    Density data for very diluted aqueous solutions of proline-leucine are reported for eight different temperatures from (283.15 to 318.15) K. Measurements were made using a vibrating tube densimeter. Apparent molar volumes of the dipeptide were calculated, at each temperature, and least squares fitted to linear equations. A weighted method with weighting factors inversely proportional to the square of the individual uncertainties has been used. Limiting partial molar volumes, V m,2 ∞ and S v values were obtained from the intercepts and slopes of those equations. Negative slopes were obtained for the lower temperatures against positive values for the higher. Trends of the values obtained are an evidence of the overwhelming contribution of the hydrophobic interactions at lower temperatures. Limiting partial molar isobaric expansions, E P,m,2 ∞ , have been also calculated using the temperature derivatives of the corresponding volumes, showing negative values. The decrease of their absolute values, with increasing temperature, points out to predominant structure maker behaviour of the solute

  8. Pharmacokinetic and imaging studies of the hepatobiliary agent sup(99m)Tc-pyridoxal leucine

    International Nuclear Information System (INIS)

    Sawas-Dimopoulou, C.; Chiotellis, E.; Dassiou, A.; Papanicolaou, N.; Simitzis, G.; Koutoulidis, K.; Hadzilouka-Mantaka, A.

    1978-01-01

    An investigation into the usefulness of sup(99m)Tc-pyridoxal leucine has demonstrated its advantages over 131 I-rose bengal in the diagnosis of patients with several liver and gall bladder complaints. Toxicity studies in mice, rabbits and dogs showed no histological signs of tissular lesions at doses of up to 5000 to 25,000 times the clinical dose. Visualization of the liver, gall bladder and biliary excretion into intestines was rapid. The appearance of activity into the intestines was delayed in patients with partial obstruction of the common bile duct. Insufficient diagnostic information was provided in jaundiced patients with higher levels of bilirubin (7 to 12 mg%). These patients showed reduced liver uptake with high background, and intestinal activity was not always clearly visualized in an 18 h study. At bilirubin levels higher than 12 mg% no liver uptake was usually observed, but only renal activity. 131 I-rose bengal was preferable for the differential diagnosis of obstructive jaundice in these patients. (U.K.)

  9. Transcriptional regulation of rod photoreceptor homeostasis revealed by in vivo NRL targetome analysis.

    Directory of Open Access Journals (Sweden)

    Hong Hao

    Full Text Available A stringent control of homeostasis is critical for functional maintenance and survival of neurons. In the mammalian retina, the basic motif leucine zipper transcription factor NRL determines rod versus cone photoreceptor cell fate and activates the expression of many rod-specific genes. Here, we report an integrated analysis of NRL-centered gene regulatory network by coupling chromatin immunoprecipitation followed by high-throughput sequencing (ChIP-Seq data from Illumina and ABI platforms with global expression profiling and in vivo knockdown studies. We identified approximately 300 direct NRL target genes. Of these, 22 NRL targets are associated with human retinal dystrophies, whereas 95 mapped to regions of as yet uncloned retinal disease loci. In silico analysis of NRL ChIP-Seq peak sequences revealed an enrichment of distinct sets of transcription factor binding sites. Specifically, we discovered that genes involved in photoreceptor function include binding sites for both NRL and homeodomain protein CRX. Evaluation of 26 ChIP-Seq regions validated their enhancer functions in reporter assays. In vivo knockdown of 16 NRL target genes resulted in death or abnormal morphology of rod photoreceptors, suggesting their importance in maintaining retinal function. We also identified histone demethylase Kdm5b as a novel secondary node in NRL transcriptional hierarchy. Exon array analysis of flow-sorted photoreceptors in which Kdm5b was knocked down by shRNA indicated its role in regulating rod-expressed genes. Our studies identify candidate genes for retinal dystrophies, define cis-regulatory module(s for photoreceptor-expressed genes and provide a framework for decoding transcriptional regulatory networks that dictate rod homeostasis.

  10. The Arabidopsis thaliana mutant air1 implicates SOS3 in the regulation of anthocyanins under salt stress

    KAUST Repository

    Van Oosten, Michael James

    2013-08-08

    The accumulation of anthocyanins in plants exposed to salt stress has been largely documented. However, the functional link and regulatory components underlying the biosynthesis of these molecules during exposure to stress are largely unknown. In a screen of second site suppressors of the salt overly sensitive3-1 (sos3-1) mutant, we isolated the anthocyanin-impaired-response-1 (air1) mutant. air1 is unable to accumulate anthocyanins under salt stress, a key phenotype of sos3-1 under high NaCl levels (120 mM). The air1 mutant showed a defect in anthocyanin production in response to salt stress but not to other stresses such as high light, low phosphorous, high temperature or drought stress. This specificity indicated that air1 mutation did not affect anthocyanin biosynthesis but rather its regulation in response to salt stress. Analysis of this mutant revealed a T-DNA insertion at the first exon of an Arabidopsis thaliana gene encoding for a basic region-leucine zipper transcription factor. air1 mutants displayed higher survival rates compared to wild-type in oxidative stress conditions, and presented an altered expression of anthocyanin biosynthetic genes such as F3H, F3′H and LDOX in salt stress conditions. The results presented here indicate that AIR1 is involved in the regulation of various steps of the flavonoid and anthocyanin accumulation pathways and is itself regulated by the salt-stress response signalling machinery. The discovery and characterization of AIR1 opens avenues to dissect the connections between abiotic stress and accumulation of antioxidants in the form of flavonoids and anthocyanins. © 2013 Springer Science+Business Media Dordrecht.

  11. Resveratrol stimulates c-Fos gene transcription via activation of ERK1/2 involving multiple genetic elements.

    Science.gov (United States)

    Thiel, Gerald; Rössler, Oliver G

    2018-06-05

    The polyphenol resveratrol is found in many plant and fruits and is a constituent of our diet. Resveratrol has been proposed to have chemopreventive and anti-inflammatory activities. On the cellular level, resveratrol activates stimulus-regulated transcription factors. To identify resveratrol-responsive elements within a natural gene promoter, the molecular pathway leading to c-Fos gene expression by resveratrol was dissected. The c-Fos gene encodes a basic region leucine zipper transcription factor and is a prototype of an immediate-early gene that is regulated by a wide range of signaling molecules. We analyzed chromatin-integrated c-Fos promoter-luciferase reporter genes where transcription factor binding sites were destroyed by point mutations or deletion mutagenesis. The results show that mutation of the binding sites for serum response factor (SRF), activator protein-1 (AP-1) and cAMP response element binding protein (CREB) significantly reduced reporter gene transcription following stimulation of the cells with resveratrol. Inactivation of the binding sites for signal transducer and activator of transcription (STAT) or ternary complex factors did not influence resveratrol-regulated c-Fos promoter activity. Thus, the c-Fos promoter contains three resveratrol-responsive elements, the cAMP response element (CRE), and the binding sites for SRF and AP-1. Moreover, we show that the transcriptional activation potential of the c-Fos protein is increased in resveratrol-stimulated cells, indicating that the biological activity of c-Fos is elevated by resveratrol stimulation. Pharmacological and genetic experiments revealed that the protein kinase ERK1/2 is the signal transducer that connects resveratrol treatment with the c-Fos gene. Copyright © 2018 Elsevier B.V. All rights reserved.

  12. A regulatory network-based approach dissects late maturation processes related to the acquisition of desiccation tolerance and longevity of Medicago truncatula seeds.

    Science.gov (United States)

    Verdier, Jerome; Lalanne, David; Pelletier, Sandra; Torres-Jerez, Ivone; Righetti, Karima; Bandyopadhyay, Kaustav; Leprince, Olivier; Chatelain, Emilie; Vu, Benoit Ly; Gouzy, Jerome; Gamas, Pascal; Udvardi, Michael K; Buitink, Julia

    2013-10-01

    In seeds, desiccation tolerance (DT) and the ability to survive the dry state for prolonged periods of time (longevity) are two essential traits for seed quality that are consecutively acquired during maturation. Using transcriptomic and metabolomic profiling together with a conditional-dependent network of global transcription interactions, we dissected the maturation events from the end of seed filling to final maturation drying during the last 3 weeks of seed development in Medicago truncatula. The network revealed distinct coexpression modules related to the acquisition of DT, longevity, and pod abscission. The acquisition of DT and dormancy module was associated with abiotic stress response genes, including late embryogenesis abundant (LEA) genes. The longevity module was enriched in genes involved in RNA processing and translation. Concomitantly, LEA polypeptides accumulated, displaying an 18-d delayed accumulation compared with transcripts. During maturation, gulose and stachyose levels increased and correlated with longevity. A seed-specific network identified known and putative transcriptional regulators of DT, including ABSCISIC ACID-INSENSITIVE3 (MtABI3), MtABI4, MtABI5, and APETALA2/ ETHYLENE RESPONSE ELEMENT BINDING PROTEIN (AtAP2/EREBP) transcription factor as major hubs. These transcriptional activators were highly connected to LEA genes. Longevity genes were highly connected to two MtAP2/EREBP and two basic leucine zipper transcription factors. A heat shock factor was found at the transition of DT and longevity modules, connecting to both gene sets. Gain- and loss-of-function approaches of MtABI3 confirmed 80% of its predicted targets, thereby experimentally validating the network. This study captures the coordinated regulation of seed maturation and identifies distinct regulatory networks underlying the preparation for the dry and quiescent states.

  13. Cross-talk between abscisic acid-dependent and abscisic acid-independent pathways during abiotic stress.

    Science.gov (United States)

    Roychoudhury, Aryadeep; Paul, Saikat; Basu, Supratim

    2013-07-01

    Salinity, drought and low temperature are the common forms of abiotic stress encountered by land plants. To cope with these adverse environmental factors, plants execute several physiological and metabolic responses. Both osmotic stress (elicited by water deficit or high salt) and cold stress increase the endogenous level of the phytohormone abscisic acid (ABA). ABA-dependent stomatal closure to reduce water loss is associated with small signaling molecules like nitric oxide, reactive oxygen species and cytosolic free calcium, and mediated by rapidly altering ion fluxes in guard cells. ABA also triggers the expression of osmotic stress-responsive (OR) genes, which usually contain single/multiple copies of cis-acting sequence called abscisic acid-responsive element (ABRE) in their upstream regions, mostly recognized by the basic leucine zipper-transcription factors (TFs), namely, ABA-responsive element-binding protein/ABA-binding factor. Another conserved sequence called the dehydration-responsive element (DRE)/C-repeat, responding to cold or osmotic stress, but not to ABA, occurs in some OR promoters, to which the DRE-binding protein/C-repeat-binding factor binds. In contrast, there are genes or TFs containing both DRE/CRT and ABRE, which can integrate input stimuli from salinity, drought, cold and ABA signaling pathways, thereby enabling cross-tolerance to multiple stresses. A strong candidate that mediates such cross-talk is calcium, which serves as a common second messenger for abiotic stress conditions and ABA. The present review highlights the involvement of both ABA-dependent and ABA-independent signaling components and their interaction or convergence in activating the stress genes. We restrict our discussion to salinity, drought and cold stress.

  14. Autoimmune encephalitis associated with voltage-gated potassium channels-complex and leucine-rich glioma-inactivated 1 antibodies

    DEFF Research Database (Denmark)

    Celicanin, Marko; Blaabjerg, M; Maersk-Moller, C

    2017-01-01

    BACKGROUND AND PURPOSE: The aim of this study was to describe clinical and paraclinical characteristics of all Danish patients who tested positive for anti-voltage-gated potassium channels (VGKC)-complex, anti-leucine-rich glioma-inactivated 1 (LGI1) and anti-contactin-associated protein-2......, electroencephalography and (18) F-fluorodeoxyglucose positron emission tomography scans were re-evaluated by experts in the field. RESULTS: A total of 28/192 patients tested positive for VGKC-complex antibodies by radioimmunoassay and indirect immunofluorescence; 17 had antibodies to LGI1 and 6/7 of the available....... CONCLUSIONS: Patients diagnosed with anti-LGI1 autoimmune encephalitis increased significantly from 2009 to 2014, probably due to increased awareness. In contrast to seropositive anti-VGKC-complex patients, all anti-LGI1-positive patients presented with a classical limbic encephalitis. The majority...

  15. Effects of steroids and sex reversal on intestinal absorption of L-(/sup 14/C)leucine in vivo, in rainbow trout, Salmo gairdneri

    Energy Technology Data Exchange (ETDEWEB)

    Habibi, H.R.; Ince, B.W.

    1983-12-01

    The effects of steroids (17 alpha-methyltestosterone (MT), 17 beta-oestradiol (E2)), and of sex reversal (XX male) on intestinal absorption and accumulation of L-(/sup 14/C)leucine (5 mM), were investigated in unanaesthetized rainbow trout (Salmo gairdneri), using an in vivo gut perfusion technique. Each steroid was luminally perfused through the gut at a concentration of 50 micrograms/ml perfusate, during five separate perfusions carried out on the same fish at 30-min intervals (perfusion periods 1 to 5), for a total of 120 min at 14 degrees. Experiments were also conducted on masculinized, genetically female trout (XX male) with steroid-free perfusate. MT treatment significantly increased the intestinal absorption of radioleucine during periods 1 and 2, whilst E2 was without effect. Neither MT nor E2 influenced intestinal accumulation (mid- and hindgut) of radioleucine, and accumulation of /sup 14/C-solutes in skeletal muscle. Sex reversal, however, whilst having no effect on leucine absorption, nevertheless significantly increased intestinal accumulation of radioleucine, and accumulation of /sup 14/C-solutes in skeletal muscle. The effects observed in the present study are in agreement with previous work in trout using everted gut sac preparations. It is suggested that the growth-promoting effects of anabolic-androgenic steroids in fish may be partly explained by their action on gastrointestinal function.

  16. Molecular cloning and characterization of leucine aminopeptidase from Fasciola gigantica.

    Science.gov (United States)

    Changklungmoa, Narin; Chaithirayanon, Kulathida; Kueakhai, Pornanan; Meemon, Krai; Riengrojpitak, Suda; Sobhon, Prasert

    2012-07-01

    M17 leucine aminopeptidase (LAP) is one of a family of metalloexopeptidases, of which short peptide fragments are cleaved from the N-terminals. In this study, the full length of cDNA encoding Fasciola gigantica LAP (FgLAP) was cloned from adult parasites. The amino acid sequences of FgLAP showed a high degree of identity (98%) with that from Fasciola hepatica and a low degree of identities (11% and 9%) with those from cattle and human. Phylogenetic analysis revealed that the FgLAP was closely related and grouped with F. hepatica LAP (FhLAP). Northern analysis showed that FgLAP transcriptional products have 1800 base pairs. Analysis by RNA in situ hybridization indicated that LAP gene was expressed in the cecal epithelial cells of adult parasites. A polyclonal antibody to a recombinant FgLAP (rFgLAP) detected the native LAP protein in various developmental stages of the parasite. In a functional test, this rFgLAP displayed aminolytic activity using a fluorogenic Leu-MCA substrate, and was significantly inhibited by bestatin. Its maximum activity was at pH 8.0 and enhanced by Mn(2+) ions. Localization of LAP proteins by immunohistochemistry and immunofluorescence techniques indicated that the enzyme was distributed in the apical cytoplasm of cecal epithelial cells. Because of its important metabolic role and fairly exposed position, FgLAP is a potential drug target and a possible vaccine candidate against fasciolosis. Copyright © 2012 Elsevier Inc. All rights reserved.

  17. Tyrosine kinase, aurora kinase and leucine aminopeptidase as attractive drug targets in anticancer therapy - characterisation of their inhibitors.

    Science.gov (United States)

    Ziemska, Joanna; Solecka, Jolanta

    Cancers are the leading cause of deaths all over the world. Available anticancer agents used in clinics exhibit low therapeutic index and usually high toxicity. Wide spreading drug resistance of cancer cells induce a demanding need to search for new drug targets. Currently, many on-going studies on novel compounds with potent anticancer activity, high selectivity as well as new modes of action are conducted. In this work, we describe in details three enzyme groups, which are at present of extensive interest to medical researchers and pharmaceutical companies. These include receptor tyrosine kinases (e.g. EGFR enzymes) and non-receptor tyrosine kinases (Src enzymes), type A, B and C Aurora kinases and aminopeptidases, especially leucine aminopeptidase. We discuss classification of these enzymes, biochemistry as well as their role in the cell cycle under normal conditions and during cancerogenesis. Further on, the work describes enzyme inhibitors that are under in vitro, preclinical, clinical studies as well as drugs available on the market. Both, chemical structures of discovered inhibitors and the role of chemical moieties in novel drug design are discussed. Described enzymes play essential role in cell cycle, especially in mitosis (Aurora kinases), cell differentiation, growth and apoptosis (tyrosine kinases) as well as G1/S transition (leucine aminopeptidase). In cancer cells, they are overexpressed and only their inhibition may stop tumor progression. This review presents the clinical outcomes of selected inhibitors and argues the safety of drug usage in human volunteers. Clinical studies of EGFR and Src kinase inhibitors in different tumors clearly show the need for molecular selection of patients (to those with mutations in genes coding EGFR and Src) to achieve positive clinical response. Current data indicates the great necessity for new anticancer treatment and actions to limit off-target activity.

  18. Valine but not leucine or isoleucine supports neurotransmitter glutamate synthesis during synaptic activity in cultured cerebellar neurons

    DEFF Research Database (Denmark)

    Bak, Lasse Kristoffer; Johansen, Maja L.; Schousboe, Arne

    2012-01-01

    Synthesis of neuronal glutamate from a-ketoglutarate for neurotransmission necessitates an amino group nitrogen donor; however, it is not clear which amino acid(s) serves this role. Thus, the ability of the three branched-chain amino acids (BCAAs), leucine, isoleucine, and valine, to act as amino...... group nitrogen donors for synthesis of vesicular neurotransmitter glutamate was investigated in cultured mouse cerebellar (primarily glutamatergic) neurons. The cultures were superfused in the presence of (15) N-labeled BCAAs, and synaptic activity was induced by pulses of N-methyl-D-aspartate (300 µ......]valine was able to maintain the amount of vesicular glutamate during synaptic activity. This indicates that, among the BCAAs, only valine supports the increased need for synthesis of vesicular glutamate. © 2012 Wiley Periodicals, Inc....

  19. Application of 15N-leucine in the study of the contribution rate of diet and each tissue to nitrogen excretion

    International Nuclear Information System (INIS)

    Yamamoto, Shigeru; Rikimaru, Tohru; Kamiesu, Noriko; Inoue, Goro

    1980-01-01

    Experimental diets (protein-free diet and 2% or 10% lactoalbumin diet) were given to male SD rats, and the urinary N content from diet was obtained from the amount of urine 15 N 24 hours after the oral administration of 15 N-leucine. N excretion from each tissue was obtained from the contents of tissue-synthesized protein N and tissue-increased N. The synthetic protein N content was obtained from the recovery rate 24 hours after 15 N-leucine administration, and the tissue-increased N content from the change in the quantity of N in each tissue on the 14th and 21st days of experimental diet. Body weight increased in the 10% diet group, decreased in the 0% diet group, and showed no change in the 2% group. The mean daily urinary N excretion was inhibited more in the 2% group than in the 0% group. The tissue 15 N level was high in the urine, followed by the order of digestive tract, liver and feces, and low in the skeletal muscle and skin. N excretion from tissues was greatest for the skeletal muscle; the ratio to total N excretion was high for the skeletal muscle and low for visceral organs in the 0% diet group, compared with the 10% diet group. With the 2% diet, most tissues showed intermediate values, and excretion from the skeletal muscle was lowest among the 3 groups, accounting for the greater inhibition of urinary N excretion than that in the 0% group. (Chiba, N.)

  20. Interaction of Prevotella intermedia strain 17 leucine-rich repeat domain protein AdpF with eukaryotic cells promotes bacterial internalization.

    Science.gov (United States)

    Sengupta, Dipanwita; Kang, Dae-Joong; Anaya-Bergman, Cecilia; Wyant, Tiana; Ghosh, Arnab K; Miyazaki, Hiroshi; Lewis, Janina P

    2014-06-01

    Prevotella intermedia is an oral bacterium implicated in a variety of oral diseases. Although internalization of this bacterium by nonphagocytic host cells is well established, the molecular players mediating the process are not well known. Here, the properties of a leucine-rich repeat (LRR) domain protein, designated AdpF, are described. This protein contains a leucine-rich region composed of 663 amino acid residues, and molecular modeling shows that it folds into a classical curved solenoid structure. The cell surface localization of recombinant AdpF (rAdpF) was confirmed by electron and confocal microscopy analyses. The recombinant form of this protein bound fibronectin in a dose-dependent manner. Furthermore, the protein was internalized by host cells, with the majority of the process accomplished within 30 min. The internalization of rAdpF was inhibited by nystatin, cytochalasin, latrunculin, nocodazole, and wortmannin, indicating that microtubules, microfilaments, and signal transduction are required for the invasion. It is noteworthy that preincubation of eukaryotic cells with AdpF increased P. intermedia 17 internalization by 5- and 10-fold for HeLa and NIH 3T3 fibroblast cell lines, respectively. The addition of the rAdpF protein was also very effective in inducing bacterial internalization into the oral epithelial cell line HN4, as well as into primary cells, including human oral keratinocytes (HOKs) and human umbilical vein endothelial cells (HUVECs). Finally, cells exposed to P. intermedia 17 internalized the bacteria more readily upon reinfection. Taken together, our data demonstrate that rAdpF plays a role in the internalization of P. intermedia 17 by a variety of host cells.