DOCSLIB.ORG

Dynamic Rerouting of the Carbohydrate Flux Is Key to Counteracting

Total Page：16

File Type：pdf, Size：1020Kb

Download full-text PDF Read full-text

Load more

BioMed Central Open Access Research article Dynamic rerouting of the carbohydrate flux is key to counteracting oxidative stress Markus Ralser*, Mirjam M Wamelink†, Axel Kowald*¶, Birgit Gerisch*, Gino Heeren§, Eduard A Struys†, Edda Klipp*, Cornelis Jakobs†, Michael Breitenbach§, Hans Lehrach* and Sylvia Krobitsch* Addresses: *Max Planck Institute for Molecular Genetics, Ihnestrasse 73, 14195 Berlin, Germany. †Department of Clinical Chemistry, Metabolic Unit, VU University Medical Center, Amsterdam, de Boelelaan 1117, 1081 HV Amsterdam, The Netherlands. §Department of Cell Biology, University of Salzburg, Hellbrunnerstrasse 34, 5020 Salzburg, Austria. ¶Current address: Medical Proteome Center, Ruhr University Bochum, Universitätsstrasse 150, 44801 Bochum, Germany. Correspondence: Markus Ralser. Email: [email protected]; Sylvia Krobitsch. Email: [email protected] Published: 21 December 2007 Received: 21 May 2007 Revised: 7 August 2007 Journal of Biology 2007, 6:10 (doi:10.1186/jbiol61) Accepted: 12 October 2007 The electronic version of this article is the complete one and can be found online at http://jbiol.com/content/6/4/10 © 2007 Ralser et al.; licensee BioMed Central Ltd. This is an Open Access article distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/2.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited. Abstract Background: Eukaryotic cells have evolved various response mechanisms to counteract the deleterious consequences of oxidative stress. Among these processes, metabolic alterations seem to play an important role. Results: We recently discovered that yeast cells with reduced activity of the key glycolytic enzyme triosephosphate isomerase exhibit an increased resistance to the thiol-oxidizing reagent diamide. Here we show that this phenotype is conserved in Caenorhabditis elegans and that the underlying mechanism is based on a redirection of the metabolic flux from glycolysis to the pentose phosphate pathway, altering the redox equilibrium of the cytoplasmic NADP(H) pool. Remarkably, another key glycolytic enzyme, glyceraldehyde-3-phosphate dehydrogenase (GAPDH), is known to be inactivated in response to various oxidant treatments, and we show that this provokes a similar redirection of the metabolic flux. Conclusions: The naturally occurring inactivation of GAPDH functions as a metabolic switch for rerouting the carbohydrate flux to counteract oxidative stress. As a consequence, altering the homoeostasis of cytoplasmic metabolites is a fundamental mechanism for balancing the redox state of eukaryotic cells under stress conditions. Journal of Biology 2007, 6:10 10.2 Journal of Biology 2007, Volume 6, Article 10 Krobitsch et al. http://jbiol.com/content/6/4/10 Background observations above suggest that alterations in the carbohy- Reactive oxygen species (ROS) cause damage to cellular drate metabolism could be central for cellular protection processes in all living organisms and contribute to a against ROS and, moreover, that cells reroute the carbohy- number of human disorders such as cancer, cardiovascular drate flux from glycolysis to the PPP to counteract perturba- diseases, stroke, and late-onset neurodegenerative disorders, tions in the cytoplasmic redox state. However, direct and to the aging process itself. To cope with the fatal cellular evidence for this hypothesis is missing so far. By combining consequences triggered by ROS, eukaryotic cells have genetic and quantitative metabolite analyses along with in evolved a number of defense and repair mechanisms, which silico modeling, we present the first direct proof that eukary- are based on enzymatic as well as non-enzymatic processes otic cells indeed actively reroute the metabolic flux from and appear to be highly conserved from unicellular to glycolysis to the PPP as an immediate and protective multicellular eukaryotes. In bacteria and yeast, these anti- response to counteract oxidative stress. oxidant defense mechanisms are partially induced on the basis of changes in global gene expression [1,2]. However, a recent study analyzing a number of genetic and environ- mental perturbations in Escherichia coli demonstrated that Results the changes in the transcriptome and proteome are unex- Reduced intracellular TPI concentration results in pectedly small [3]. Moreover, the transcription of genes enhanced oxidant resistance of Saccharomyces encoding enzymes capable of neutralizing ROS is not gener- cerevisiae and Caenorhabditis elegans ally increased in mammalian cells that are subjected to We reported earlier that a change of the amino acid oxidative stress [4]. isoleucine to valine at position 170 in the human TPI protein (TPIIle170Val) causes a reduction of about 70% in the In all organisms studied, however, treatment with oxidants enzyme’s catalytic activity [11]. Interestingly, we discovered prompts immediate de novo post-translational modifica- that yeast cells expressing this human TPI variant exhibit tions of a number of proteins, probably affecting their local- increased resistance to the oxidant diamide (N,N,N’,N’- ization and functionality. One of the key targets of those tetramethylazodicarboxamide, Chemical Abstracts Service processes is the glycolytic enzyme glyceraldehyde-3-phos- (CAS) No. 10465-78-8) compared with isogenic yeast cells phate dehydrogenase (GAPDH), which catalyzes the expressing wild-type human TPI, indicating that low TPI reversible oxidative phosphorylation of glyceraldehyde-3- activity confers resistance to specific conditions of oxidative phosphate (gly3p) to 1,3-bisphosphoglycerate. Remarkably, stress. The synthetic reagent diamide is known to oxidize in response to various oxidant treatments this enzyme is cellular thiols, especially protein-integrated cysteines [19], inactivated and transported into the nucleus of the cell, and provoking a rapid decrease in cellular glutathione and has been found S-nitrosylated, S-thiolated, S-glutathiony- hence causing oxidative stress. To dissect the underlying lated, carbonylated and ADP-ribosylated in numerous cell mechanism, we first analyzed whether decreasing the types and organisms under these conditions [5-10]. expression level of wild-type human TPI would result in a Recently, we discovered that yeast cells with reduced cat- similar phenotype. For this, we generated plasmids for the alytic activity of another key glycolytic enzyme, triose- expression of wild-type human TPI under the control of phosphate isomerase (TPI), are highly resistant to the established yeast promoters of different strengths, namely oxidant diamide [11]. This essential enzyme precedes the CYC1, TEF1 and GPD1 promoters [20]. Subsequently, GAPDH in glycolysis, catalyzing the interconversion of di- the ∆tpi1 strain MR100, which is deleted for the yeast TPI1 hydroxyacetone phosphate (dhap) and gly3p, the substrate gene and is inviable on medium containing glucose as sole of GAPDH, and a reduction in its activity results in an ele- carbon source, was transformed with the respective plas- vated cellular dhap concentration [12-14]. In this light, it is mids along with control plasmids encoding yeast TPIIle170Val remarkable that the expression of a subset of glycolytic pro- or yeast TPI. Single colonies were selected and the intracel- teins and proteins implicated in related pathways is repres- lular TPI concentration of plate-grown yeast cells was ana- sed, while the expression of a few enzymes involved in the lyzed (Figure 1a, left panel). As expected, yeast cells pentose phosphate pathway (PPP), which is directly con- expressing the different TPI proteins under the strong GPD1 nected to the glycolytic pathway, is induced under oxidative promoter had a higher TPI concentration compared with stress conditions [1]. Furthermore, enhanced activity of the cells in which the expression was controlled by the interme- PPP has been observed in neonatal rat cardiomyocytes and diate TEF1 or the weak CYC1 promoter. Next, we spotted in human epithelial cells under oxidative stress conditions the respective yeast cells onto medium supplemented with [15,16]. Enzymes of the PPP are crucial for maintaining the differing diamide concentrations. As shown in Figure 1a cytoplasmic NADPH concentration, which provides the (right panel), yeast cells expressing human TPI under the redox power for known antioxidant systems [17,18]. The control of the weakest promoter used, the CYC1 promoter, Journal of Biology 2007, 6:10 http://jbiol.com/content/6/4/10 Journal of Biology 2007, Volume 6, Article 10 Krobitsch et al. 10.3 (a) lle170Val -TPI -yeast TPI-TPI -TPI -TPI GPD -yeast TPI pr pr pr pr pr pr GPD GPD GPD TEF CYC GPDpr -TPIlle170Val TPI GPDpr -TPI TEFpr -TPI G6PDH CYCpr -TPI Without diamide 1.4 mM diamide 1.6 mM diamide 1.8 mM diamide (b) Juglone Diamide RNAi 100 100 Control tpi-1 90 90 80 80 TPI-1 70 70 Wild-type 60 60 50 50 tpi-1 RNAi DAF-21 40 40 30 30 daf-2 (e1370) 20 20 10 10 0 0 Animals alive (percentage) alive Animals 0 1 2 3 4 5 6 7 8 (percentage) alive Animals 0123456789 Time (h) Time (h) Figure 1 Reduced triosephosphate isomerase (TPI) activity increases oxidant resistance of S. cerevisiae and C. elegans. (a) The left panel shows a Western blot analysis of yeast cells expressing wild-type human TPI under the control of promoters of different strengths: GPD1 (GPDpr), TEF1 (TEFpr), and CYC1 (CYCpr). Yeast cells expressing human TPIIle170Val or yeast TPI under the control of the strong
Recommended publications
  • Chuanxiong Rhizoma Compound on HIF-VEGF Pathway and Cerebral Ischemia-Reperfusion Injury’S Biological Network Based on Systematic Pharmacology

    Chuanxiong Rhizoma Compound on HIF-VEGF Pathway and Cerebral Ischemia-Reperfusion Injury’S Biological Network Based on Systematic Pharmacology

    ORIGINAL RESEARCH published: 25 June 2021 doi: 10.3389/fphar.2021.601846 Exploring the Regulatory Mechanism of Hedysarum Multijugum Maxim.-Chuanxiong Rhizoma Compound on HIF-VEGF Pathway and Cerebral Ischemia-Reperfusion Injury’s Biological Network Based on Systematic Pharmacology Kailin Yang 1†, Liuting Zeng 1†, Anqi Ge 2†, Yi Chen 1†, Shanshan Wang 1†, Xiaofei Zhu 1,3† and Jinwen Ge 1,4* Edited by: 1 Takashi Sato, Key Laboratory of Hunan Province for Integrated Traditional Chinese and Western Medicine on Prevention and Treatment of 2 Tokyo University of Pharmacy and Life Cardio-Cerebral Diseases, Hunan University of Chinese Medicine, Changsha, China, Galactophore Department, The First 3 Sciences, Japan Hospital of Hunan University of Chinese Medicine, Changsha, China, School of Graduate, Central South University, Changsha, China, 4Shaoyang University, Shaoyang, China Reviewed by: Hui Zhao, Capital Medical University, China Background: Clinical research found that Hedysarum Multijugum Maxim.-Chuanxiong Maria Luisa Del Moral, fi University of Jaén, Spain Rhizoma Compound (HCC) has de nite curative effect on cerebral ischemic diseases, *Correspondence: such as ischemic stroke and cerebral ischemia-reperfusion injury (CIR). However, its Jinwen Ge mechanism for treating cerebral ischemia is still not fully explained. [email protected] †These authors share first authorship Methods: The traditional Chinese medicine related database were utilized to obtain the components of HCC. The Pharmmapper were used to predict HCC’s potential targets. Specialty section: The CIR genes were obtained from Genecards and OMIM and the protein-protein This article was submitted to interaction (PPI) data of HCC’s targets and IS genes were obtained from String Ethnopharmacology, a section of the journal database.
  • TPI1 Gene Triosephosphate Isomerase 1

    TPI1 Gene Triosephosphate Isomerase 1

    TPI1 gene triosephosphate isomerase 1 Normal Function The TPI1 gene provides instructions for making an enzyme called triosephosphate isomerase 1. This enzyme is involved in a critical energy-producing process known as glycolysis. During glycolysis, the simple sugar glucose is broken down to produce energy for cells. The triosephosphate isomerase 1 enzyme carries out a specific reaction during glycolysis: the conversion of a molecule called dihydroxyacetone phosphate (DHAP) to glyceraldehyde 3-phosphate. This conversion can go both ways, meaning that the triosephosphate isomerase 1 enzyme can also convert glyceraldehyde 3-phosphate back into DHAP. Additional steps convert glyceraldehyde 3-phosphate into other molecules that ultimately produce energy in the form of a molecule called ATP. For the triosephosphate isomerase 1 enzyme to be turned on (active), it has to attach ( bind) to another triosephosphate isomerase 1 enzyme, forming a two-enzyme complex called a dimer. Health Conditions Related to Genetic Changes Triosephosphate isomerase deficiency At least 12 mutations in the TPI1 gene have been found to cause triosephosphate isomerase deficiency. This condition is characterized by a shortage of red blood cells ( anemia), movement problems, increased susceptibility to infection, and muscle weakness that can affect breathing and heart function. TPI1 gene mutations can lead to the production of an enzyme with decreased activity. As a result, glycolysis is impaired and cells have a decreased supply of energy. One TPI1 gene mutation accounts for approximately 80 percent of triosephosphate isomerase deficiency cases. This change replaces the protein building block (amino acid) glutamic acid with the amino acid aspartic acid at position 104 in the triosephosphate isomerase 1 enzyme (written as Glu104Asp or E104D).
  • Candida Albicans and Candida Glabrata Triosephosphate Isomerase

    Candida Albicans and Candida Glabrata Triosephosphate Isomerase

    Satala et al. BMC Microbiology (2021) 21:199 https://doi.org/10.1186/s12866-021-02235-w RESEARCH Open Access Candida albicans and Candida glabrata triosephosphate isomerase – a moonlighting protein that can be exposed on the candidal cell surface and bind to human extracellular matrix proteins Dorota Satala1, Grzegorz Satala2, Marcin Zawrotniak3 and Andrzej Kozik1,4* Abstract Background: Triosephosphate isomerase (Tpi1) is a glycolytic enzyme that has recently been reported also to be an atypical proteinaceous component of the Candida yeast cell wall. Similar to other known candidal “moonlighting proteins”, surface-exposed Tpi1 is likely to contribute to fungal adhesion during the colonization and infection of a human host. The aim of our present study was to directly prove the presence of Tpi1 on C. albicans and C. glabrata cells under various growth conditions and characterize the interactions of native Tpi1, isolated and purified from the candidal cell wall, with human extracellular matrix proteins. Results: Surface plasmon resonance measurements were used to determine the dissociation constants for the complexes of Tpi1 with host proteins and these values were found to fall within a relatively narrow range of 10− 8- 10− 7 M. Using a chemical cross-linking method, two motifs of the Tpi1 molecule (aa 4–17 and aa 224–247) were identified to be directly involved in the interaction with vitronectin. A proposed structural model for Tpi1 confirmed that these interaction sites were at a considerable distance from the catalytic active site. Synthetic peptides with these sequences significantly inhibited Tpi1 binding to several extracellular matrix proteins suggesting that a common region on the surface of Tpi1 molecule is involved in the interactions with the host proteins.
  • A High Throughput Approach for the Generation of Orthogonally Interacting Protein Pairs Justin Lawrie University of Nebraska-Lincoln, Jlawrie2@Unl.Edu

    A High Throughput Approach for the Generation of Orthogonally Interacting Protein Pairs Justin Lawrie University of Nebraska-Lincoln, [email protected]

    University of Nebraska - Lincoln DigitalCommons@University of Nebraska - Lincoln Faculty Publications -- Chemistry Department Published Research - Department of Chemistry 1-17-2018 A high throughput approach for the generation of orthogonally interacting protein pairs Justin Lawrie University of Nebraska-Lincoln, [email protected] Xi Song University of Nebraska-Lincoln Wei Niu University of Nebraska-Lincoln, [email protected] Jiantao Guo University of Nebraska-Lincoln, [email protected] Follow this and additional works at: https://digitalcommons.unl.edu/chemfacpub Part of the Analytical Chemistry Commons, Medicinal-Pharmaceutical Chemistry Commons, and the Other Chemistry Commons Lawrie, Justin; Song, Xi; Niu, Wei; and Guo, Jiantao, "A high throughput approach for the generation of orthogonally interacting protein pairs" (2018). Faculty Publications -- Chemistry Department. 122. https://digitalcommons.unl.edu/chemfacpub/122 This Article is brought to you for free and open access by the Published Research - Department of Chemistry at DigitalCommons@University of Nebraska - Lincoln. It has been accepted for inclusion in Faculty Publications -- Chemistry Department by an authorized administrator of DigitalCommons@University of Nebraska - Lincoln. www.nature.com/scientificreports OPEN A high throughput approach for the generation of orthogonally interacting protein pairs Received: 20 September 2017 Justin Lawrie1, Xi Song1, Wei Niu2 & Jiantao Guo1 Accepted: 27 December 2017 In contrast to the nearly error-free self-assembly of protein architectures in nature, artifcial assembly of Published: xx xx xxxx protein complexes with pre-defned structure and function in vitro is still challenging. To mimic nature’s strategy to construct pre-defned three-dimensional protein architectures, highly specifc protein- protein interacting pairs are needed. Here we report an efort to create an orthogonally interacting protein pair from its parental pair using a bacteria-based in vivo directed evolution strategy.
  • Degradation of Functional Triose Phosphate Isomerase Protein Underlies Sugarkill Pathology

    Degradation of Functional Triose Phosphate Isomerase Protein Underlies Sugarkill Pathology

    Copyright Ó 2008 by the Genetics Society of America DOI: 10.1534/genetics.108.087551 Degradation of Functional Triose Phosphate Isomerase Protein Underlies sugarkill Pathology Jacquelyn L. Seigle,1 Alicia M. Celotto1 and Michael J. Palladino2 Department of Pharmacology and Chemical Biology, University of Pittsburgh School of Medicine, Pittsburgh, Pennsylvania 15261 and Pittsburgh Institute for Neurodegenerative Diseases, University of Pittsburgh School of Medicine, Pittsburgh, Pennsylvania 15260 Manuscript received January 26, 2008 Accepted for publication April 12, 2008 ABSTRACT Triose phosphate isomerase (TPI) deficiency glycolytic enzymopathy is a progressive neurodegenerative condition that remains poorly understood. The disease is caused exclusively by specific missense mutations affecting the TPI protein and clinically features hemolytic anemia, adult-onset neurological impairment, degeneration, and reduced longevity. TPI has a well-characterized role in glycolysis, catalyzing the isom- erization of dihydroxyacetone phosphate (DHAP) to glyceraldehyde-3-phosphate (G3P); however, little is known mechanistically about the pathogenesis associated with specific recessive mutations that cause progressive neurodegeneration. Here, we describe key aspects of TPI pathogenesis identified using the TPIsugarkill mutation, a Drosophila model of human TPI deficiency. Specifically, we demonstrate that the mutant protein is expressed, capable of forming a homodimer, and is functional. However, the mutant protein is degraded by the 20S proteasome core leading to loss-of-function pathogenesis. LYCOLYTIC enzymopathies are devastating hu- instability of the protein homodimer, aberrant protein G man diseases that result from heritable mutations complexes, and/or a decrease in isomerase activity con- in genes encoding certain glycolytic enzymes. The un- tribute to pathogenesis of the disease (Olah et al. 2002; derlying pathogenesis of these diseases remains poorly Orosz et al.
  • Cloning and Characterization of the Orotidine-5'-Phosphate Decarboxylase Gene (URA3) from the Osmotolerant Yeast Candida Magnoliae

    Cloning and Characterization of the Orotidine-5'-Phosphate Decarboxylase Gene (URA3) from the Osmotolerant Yeast Candida Magnoliae

    J. Microbiol. Biotechnol. (2012), 22(5), 642–648 http://dx.doi.org/10.4014/jmb.1111.11071 First published online February 17, 2012 pISSN 1017-7825 eISSN 1738-8872 Cloning and Characterization of the Orotidine-5'-Phosphate Decarboxylase Gene (URA3) from the Osmotolerant Yeast Candida magnoliae Park, Eun-Hee1, Jin-Ho Seo2, and Myoung-Dong Kim1* 1School of Biotechnology and Bioengineering, Kangwon National University, Chuncheon 200-701, Korea 2Department of Agricultural Biotechnology, Seoul National University, Seoul 151-921, Korea Received: November 28, 2011 / Revised: January 10, 2012 / Accepted: January 11, 2012 We determined the nucleotide sequence of the URA3 gene amounts of erythritol [10, 12, 21]. A gene transformation/ encoding orotidine-5'-phosphate decarboxylase (OMPDCase) disruption system might be essential to understand the of the erythritol-producing osmotolerant yeast Candida underlying molecular mechanisms of osmotolerance and magnoliae by degenerate polymerase chain reaction and erythritol production in C. magnoliae. However, the genetic genome walking. Sequence analysis revealed the presence manipulation of C. magnoliae is limited by a lack of markers of an uninterrupted open-reading frame of 795 bp, encoding and efficient transformation methods, which implies that a 264 amino acid residue protein with the highest identity manipulation of this strain should involve the use of to the OMPDCase of the yeast Kluyveromyces marxianus. dominant drug-resistance markers. Although several dominant Phylogenetic analysis of the deduced amino acid sequence drug-resistance markers are available [33], little success revealed that it shared a high degree of identity with other has been achieved in Candida species, which frequently yeast OMPDCase homologs. The cloned URA3 gene exhibit drug resistance and show different codon usage successfully complemented the ura3 null mutation in preferences [1, 15].
  • Datasheet: VPA00146 Product Details

    Datasheet: VPA00146 Product Details

    Datasheet: VPA00146 Description: GOAT ANTI TPI1 Specificity: TPI1 Format: Purified Product Type: PrecisionAb™ Polyclonal Isotype: Polyclonal IgG Quantity: 100 µl Product Details Applications This product has been reported to work in the following applications. This information is derived from testing within our laboratories, peer-reviewed publications or personal communications from the originators. Please refer to references indicated for further information. For general protocol recommendations, please visit www.bio-rad-antibodies.com/protocols. Yes No Not Determined Suggested Dilution Western Blotting 1/1000 PrecisionAb antibodies have been extensively validated for the western blot application. The antibody has been validated at the suggested dilution. Where this product has not been tested for use in a particular technique this does not necessarily exclude its use in such procedures. Further optimization may be required dependant on sample type. Target Species Human Species Cross Reacts with: Mouse, Rat Reactivity N.B. Antibody reactivity and working conditions may vary between species. Product Form Purified IgG - liquid Preparation Goat polyclonal antibody purified by affinity chromatography. Buffer Solution TRIS buffered saline Preservative 0.02% Sodium Azide (NaN ) 0.5 % BSA Stabilisers 3 Immunogen Peptide with the sequence C-LKPEFVDIINAKQ, from the C Terminus of the protein sequence. External Database Links UniProt: P60174 Related reagents Entrez Gene: 7167 TPI1 Related reagents Synonyms TPI Page 1 of 2 Specificity Goat anti Human TPI1 antibody recognizes triosephosphate isomerase, also known as TIM or triose-phosphate isomerase. Encoded by the TPI1 gene, this enzyme consists of two identical proteins and catalyzes the isomerization of glyceraldehydes 3-phosphate (G3P) and dihydroxy-acetone phosphate (DHAP) in glycolysis and gluconeogenesis.
  • A Bioinformatics Model of Human Diseases on the Basis Of

    A Bioinformatics Model of Human Diseases on the Basis Of

    SUPPLEMENTARY MATERIALS A Bioinformatics Model of Human Diseases on the basis of Differentially Expressed Genes (of Domestic versus Wild Animals) That Are Orthologs of Human Genes Associated with Reproductive-Potential Changes Vasiliev1,2 G, Chadaeva2 I, Rasskazov2 D, Ponomarenko2 P, Sharypova2 E, Drachkova2 I, Bogomolov2 A, Savinkova2 L, Ponomarenko2,* M, Kolchanov2 N, Osadchuk2 A, Oshchepkov2 D, Osadchuk2 L 1 Novosibirsk State University, Novosibirsk 630090, Russia; 2 Institute of Cytology and Genetics, Siberian Branch of Russian Academy of Sciences, Novosibirsk 630090, Russia; * Correspondence: [email protected]. Tel.: +7 (383) 363-4963 ext. 1311 (M.P.) Supplementary data on effects of the human gene underexpression or overexpression under this study on the reproductive potential Table S1. Effects of underexpression or overexpression of the human genes under this study on the reproductive potential according to our estimates [1-5]. ↓ ↑ Human Deficit ( ) Excess ( ) # Gene NSNP Effect on reproductive potential [Reference] ♂♀ NSNP Effect on reproductive potential [Reference] ♂♀ 1 increased risks of preeclampsia as one of the most challenging 1 ACKR1 ← increased risk of atherosclerosis and other coronary artery disease [9] ← [3] problems of modern obstetrics [8] 1 within a model of human diseases using Adcyap1-knockout mice, 3 in a model of human health using transgenic mice overexpressing 2 ADCYAP1 ← → [4] decreased fertility [10] [4] Adcyap1 within only pancreatic β-cells, ameliorated diabetes [11] 2 within a model of human diseases
  • A Methodological Approach for the Construction of a Radiation Hybrid Map of Bovine Chromosome 5

    A Methodological Approach for the Construction of a Radiation Hybrid Map of Bovine Chromosome 5

    Genetics and Molecular Biology, 27, 1, 22-32 (2004) Copyright by the Brazilian Society of Genetics. Printed in Brazil www.sbg.org.br Research Article A methodological approach for the construction of a radiation hybrid map of bovine chromosome 5 M. De Donato1,2*, R.A. Brenneman1,3*, D.M. Stelly5, J.E. Womack5 and J.F. Taylor1,6 1Texas A&M University, Department of Animal Science, College Station, USA. 2Present address: Departamento de Biomedicina, IIBCA, Universidad de Oriente, Venezuela. 3Present address: Omaha’s Henry Doorly Zoo, Omaha, USA. 4Laboratory of Plant Molecular Cytogenetics, Department of Soil and Crop Sciences, Texas A&M University, College Station, USA. 5Laboratory of Genetics, Department of Veterinary Pathobiology, Texas A&M University, College Station, USA. 6Present address: University of Missouri, Campus Drive, Columbia, USA. Abstract A bovine 5,000 rad WG-RH panel was used to construct an RH map of bovine chromosome 5 (BTA5). Twenty-one microsatellites and thirteen genes were scored in the panel using PAGE and radioactive labeling. Marker retention ranged from 8.9%-25.8% and averaged 17.8%. Pairwise locus analysis placed all markers in a single syntenic group with a LOD support of 4.0. At a LOD support of 8.0, a centromeric group of 23 syntenic markers was formed. Telomeric groups of 11 and 9 markers were assembled with a LOD support of 6.0 and 8.0, respectively. All markers were ordered by maximum likelihood methods using the program RHMAP. Only 13 markers were ordered with a LOD support of at least 3.0, while 25 and 29 markers were ordered with a support of at least 2.0 and 1.0, respectively.
  • Ncomms8838.Pdf

    Ncomms8838.Pdf

    ARTICLE Received 2 Feb 2015 | Accepted 17 Jun 2015 | Published 21 Jul 2015 DOI: 10.1038/ncomms8838 OPEN Functional genomics identifies negative regulatory nodes controlling phagocyte oxidative burst Daniel B. Graham1,2, Christine E. Becker3, Aivi Doan1, Gautam Goel3, Eduardo J. Villablanca1,2,3, Dan Knights4, Amanda Mok1, Aylwin C.Y. Ng1,5, John G. Doench1, David E. Root1, Clary B. Clish1 & Ramnik J. Xavier1,2,3,5,6 The phagocyte oxidative burst, mediated by Nox2 NADPH oxidase-derived reactive oxygen species, confers host defense against a broad spectrum of bacterial and fungal pathogens. Loss-of-function mutations that impair function of the Nox2 complex result in a life-threatening immunodeficiency, and genetic variants of Nox2 subunits have been implicated in pathogenesis of inflammatory bowel disease (IBD). Thus, alterations in the oxidative burst can profoundly impact host defense, yet little is known about regulatory mechanisms that fine-tune this response. Here we report the discovery of regulatory nodes controlling oxidative burst by functional screening of genes within loci linked to human inflammatory disease. Implementing a multi-omics approach, we define transcriptional, metabolic and ubiquitin-cycling nodes controlled by Rbpj, Pfkl and Rnf145, respectively. Furthermore, we implicate Rnf145 in proteostasis of the Nox2 complex by endoplasmic reticulum-associated degradation. Consequently, ablation of Rnf145 in murine macrophages enhances bacterial clearance, and rescues the oxidative burst defects associated with Ncf4 haploinsufficiency. 1 Broad Institute of MIT and Harvard, Cambridge, Massachusetts 02142, USA. 2 Department of Medicine, Massachusetts General Hospital, Harvard Medical School, Boston, Massachusetts 02114, USA. 3 Center for Computational and Integrative Biology, Massachusetts General Hospital, Harvard Medical School, Boston, Massachusetts 02114, USA.
  • A Curated Biochemical Pathways Database for the Laboratory Mouse Alexei V Evsikov, Mary E Dolan, Michael P Genrich, Emily Patek and Carol J Bult

    A Curated Biochemical Pathways Database for the Laboratory Mouse Alexei V Evsikov, Mary E Dolan, Michael P Genrich, Emily Patek and Carol J Bult

    Open Access Software2009EvsikovetVolume al. 10, Issue 8, Article R84 MouseCyc: a curated biochemical pathways database for the laboratory mouse Alexei V Evsikov, Mary E Dolan, Michael P Genrich, Emily Patek and Carol J Bult Address: The Jackson Laboratory, Main Street, Bar Harbor, ME 04609, USA. Correspondence: Carol J Bult. Email: [email protected] Published: 14 August 2009 Received: 22 May 2009 Revised: 17 July 2009 Genome Biology 2009, 10:R84 (doi:10.1186/gb-2009-10-8-r84) Accepted: 14 August 2009 The electronic version of this article is the complete one and can be found online at http://genomebiology.com/2009/10/8/R84 © 2009 Evsikov et al,; licensee BioMed Central Ltd. This is an open access article distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/2.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited. MouseCyc<p>MouseCyc database is a database of curated metabolic pathways for the laboratory mouse.</p> Abstract Linking biochemical genetic data to the reference genome for the laboratory mouse is important for comparative physiology and for developing mouse models of human biology and disease. We describe here a new database of curated metabolic pathways for the laboratory mouse called MouseCyc http://mousecyc.jax.org. MouseCyc has been integrated with genetic and genomic data for the laboratory mouse available from the Mouse Genome Informatics database and with pathway data from other organisms, including human. Rationale Biochemical interactions and transformations among organic The availability of the nearly complete genome sequence for molecules are arguably the foundation and core distinguish- the laboratory mouse provides a powerful platform for pre- ing feature of all organic life.
  • 9Kosinski Proof2

    9Kosinski Proof2

    Tested Studies for Laboratory Teaching Proceedings of the Association for Biology Laboratory Education Vol. 37, Article 9, 2016 An Introduction to Bioinformatics Robert J. Kosinski Department of Biological Sciences, 132 Long Hall, Clemson University, Clemson SC 29634- 0314 USA ([email protected]) This exercise is used in introductory biology for majors at Clemson University. Students download DNA and protein sequences from a Web site and apply common bioinformatics tools to identifying and researching them. The students identify a piece of a gene using BLAST, explore their gene’s genomic “neighborhood,” determine the percent of DNA that is transcribed in this neighborhood, identify a protein using BLAST, research this protein using UniProt, search for literature on the role of the protein in disease, and finally, determine if DNA isolates from a mass disease outbreak show evidence of bioterrorism. Keywords: bioinformatics, gene, protein, BLAST, GenBank, UniProt, PubMed, bioterrorism Introduction Each student is given a code letter (A-Q). Another code (Z) is not given to students because it is used as an Overview of the Laboratory example in the exercise. Each of these letters represents a This is a “dry lab” introduction to some common group of files. Using the Web site techniques in bioinformatics using DNA and protein files http://www.ableweb.org/volumes/vol−37/kosinski/bioinfo downloaded from a Clemson Web site. An older version rmatics.htm each student will download: of this laboratory has been used at Clemson for about 10 a) a fragment of human genomic DNA years, and was presented at ABLE in 2006. Aside from corrupted by sequencing ambiguities (Ns).