Triggering the Stringent Response: Signals Responsible for Activating (P) Ppgpp Synthesis in Bacteria
Total Page:16
File Type:pdf, Size:1020Kb
Load more
Recommended publications
-
Adaptive Responses by Transcriptional Regulators to Small Molecules in Prokaryotes
Adaptive Responses by Transcriptional Regulators to small molecules in Prokaryotes Structural studies of two bacterial one-component signal transduction systems DntR and HpNikR Cyril Dian Stockholm University Doctoral thesis © Cyril Dian, Stockholm 2007 ISBN 978-91-7155-500-7 Department of Biochemistry and Biophysics The Arrhenius Laboratories for Natural Sciences Stockholm University SE-106 91 Stockholm Sweden All previously published papers are reprinted With permission from the publishers Intellecta Docusys, Stockholm 2007 Abstract Prokaryotes are continually exposed to environmental changes in their physiological conditions. In order to survive such unstable conditions, or to compete with others species for the same environmental niche, prokaryotes must monitor signals about both their extracellular environment and intracellular physiological status and provide rapid and appropriate responses to variations in their surroundings. This adaptive response to environmental signals is triggered mainly by transcriptional regulators via two components, the one- and two-component signal transduction systems. These scan intra- and extracellular small-molecule mixtures and modulate gene expression to provide the appropriate physiological response to the prevailing conditions. Most prokaryotic one component regulators are simple transcription factors comprising of a small-molecule binding domain (SMBD) and a DNA binding domain (DBD). Although the effects of transcription factors on the transcription machinery are well understood, the exact location -
Regulation of Stringent Factor by Branched-Chain Amino Acids
Regulation of stringent factor by branched-chain amino acids Mingxu Fanga and Carl E. Bauera,1 aMolecular and Cellular Biochemistry Department, Indiana University, Bloomington, IN 47405 Edited by Caroline S. Harwood, University of Washington, Seattle, WA, and approved May 9, 2018 (received for review February 21, 2018) When faced with amino acid starvation, prokaryotic cells induce a Under normal growth conditions, the synthetase activity of Rel is stringent response that modulates their physiology. The stringent thought to be self-inhibited; however, during times of amino acid response is manifested by production of signaling molecules starvation, Rel interacts with stalled ribosomes, which activates guanosine 5′-diphosphate,3′-diphosphate (ppGpp) and guanosine synthetase activity to produce (p)ppGpp. The regulation of hy- 5′-triphosphate,3′-diphosphate (pppGpp) that are also called drolase activity is less understood but may involve one or more alarmones. In many species, alarmone levels are regulated by a downstream domains called the TGS and ACT domains. The TGS multidomain bifunctional alarmone synthetase/hydrolase called domain of SpoT has been shown to interact with an acyl carrier Rel. In this enzyme, there is an ACT domain at the carboxyl region protein, so it is presumed to sense the status of fatty acid metab- that has an unknown function; however, similar ACT domains are olism in E. coli (4). The function of the ACT domain is not as clear; present in other enzymes that have roles in controlling amino acid however, recent cryo-EM structures of E. coli RelA show that this metabolism. In many cases, these other ACT domains have been domain is involved in binding deacyl-tRNA as well as the ribosome shown to allosterically regulate enzyme activity through the bind- (5–7). -
Cyclic Di-AMP Signaling in Listeria Monocytogenes by Aaron Thomas
Cyclic di-AMP signaling in Listeria monocytogenes By Aaron Thomas Whiteley A dissertation submitted in partial satisfaction of the requirements for the degree of Doctor of Philosophy in Infectious Diseases and Immunity in the Graduate Division of the University of California, Berkeley Committee in charge: Professor Daniel A. Portnoy, Chair Professor Sarah A. Stanley Professor Russell E. Vance Professor Kathleen R. Ryan Summer 2016 Abstract Cyclic di-AMP signaling in Listeria monocytogenes by Aaron Thomas Whiteley Doctor of Philosophy in Infectious Diseases and Immunity University of California, Berkeley Professor Daniel A. Portnoy, Chair The Gram positive facultative intracellular pathogen Listeria monocytogenes is both ubiquitous in the environment and is a facultative intracellular pathogen. A high degree of adaptability to different growth niches is one reason for the success of this organism. In this dissertation, two facets of L. monocytogenes, growth and gene expression have been investigated. The first portion examines the function and necessity of the nucleotide second messenger c-di-AMP, and the second portion of this dissertation examines the signal transduction network required for virulence gene regulation. Through previous genetic screens and biochemical analysis it was found that the nucleotide second messenger cyclic di-adenosine monophosphate (c-di-AMP) is secreted by the bacterium during intracellular and extracellular growth. Depletion of c-di- AMP levels in L. monocytogenes and related bacteria results in sensitivity to cell wall acting antibiotics such as cefuroxime, decreased growth rate, and decreased virulence. We devised a variety of bacterial genetic screens to identify the function of this molecule in bacterial physiology. The sole di-adenylate cyclase (encoded by dacA) responsible for catalyzing synthesis of c-di-AMP in L. -
Assessing the Role of Spot and Rela in Capsular Polysaccharide
Journal of Experimental Microbiology and Immunology (JEMI) Vol. 15: 52 – 58 Copyright © April 2011, M&I UBC Assessing the Role of SpoT and RelA in Capsular Polysaccharide Synthesis after Treatment with Sub-lethal Concentrations of Kanamycin to Confer Decreased Antibiotic Sensitivity in Escherichia coli Wesley Chenne, Louisa Ng, and Mary Rose Pambid Department of Microbiology & Immunology, University of British Columbia Resistance to various antibiotics has been associated with increased capsular polysaccharide production through activation of RelA and SpoT, regulators of the stringent response, through their respective synthesis and hydrolysis of guanosine tetraphosphate in Escherichia coli. In order to further characterize the role of SpoT and RelA in capsular polysaccharide production and in conferring antibiotic resistance, growth patterns and induced capsular polysaccharide levels of various strains were determined following sub- lethal treatment with kanamycin and further treatment at inhibitory levels of kanamycin. Contrary to previous reports that capsular polysaccharide confers antibiotic resistance, it was found that spoT mutants produced the least capsular polysaccharide but had higher levels of resistance in comparison to relA mutants, which produced the most polysaccharide but exhibited lower resistance. As expected, both these mutants exhibited lower resistance to kanamycin as a result of pre-treatment than the wild-type strain. Thus, we propose that bacterial survival and resistance development upon antibiotic administration -
The Hns Gene of Escherichia Coli Is Transcriptionally Down-Regulated by (P)Ppgpp
microorganisms Article The hns Gene of Escherichia coli Is Transcriptionally Down-Regulated by (p)ppGpp Anna Brandi, Mara Giangrossi, Attilio Fabbretti and Maurizio Falconi * School of Biosciences and Veterinary Medicine, University of Camerino, 62032 Camerino, Italy; [email protected] (A.B.); [email protected] (M.G.); [email protected] (A.F.) * Correspondence: [email protected]; Tel.: +39-0737-403274 Received: 25 August 2020; Accepted: 8 October 2020; Published: 10 October 2020 Abstract: Second messenger nucleotides, such as guanosine penta- or tetra-phosphate, commonly referred to as (p)ppGpp, are powerful signaling molecules, used by all bacteria to fine-tune cellular metabolism in response to nutrient availability. Indeed, under nutritional starvation, accumulation of (p)ppGpp reduces cell growth, inhibits stable RNAs synthesis, and selectively up- or down- regulates the expression of a large number of genes. Here, we show that the E. coli hns promoter responds to intracellular level of (p)ppGpp. hns encodes the DNA binding protein H-NS, one of the major components of bacterial nucleoid. Currently, H-NS is viewed as a global regulator of transcription in an environment-dependent mode. Combining results from relA (ppGpp synthetase) and spoT (ppGpp synthetase/hydrolase) null mutants with those from an inducible plasmid encoded RelA system, we have found that hns expression is inversely correlated with the intracellular concentration of (p)ppGpp, particularly in exponential phase of growth. Furthermore, we have reproduced in an in vitro system the observed in vivo (p)ppGpp-mediated transcriptional repression of hns promoter. Electrophoretic mobility shift assays clearly demonstrated that this unusual nucleotide negatively affects the stability of RNA polymerase-hns promoter complex. -
Protein Interactions with the Glucose Transporter Binding Protein GLUT1CBP That Provide a Link Between GLUT1 and the Cytoskeleton Robert C
Molecular Biology of the Cell Vol. 10, 819–832, April 1999 Protein Interactions with the Glucose Transporter Binding Protein GLUT1CBP That Provide a Link between GLUT1 and the Cytoskeleton Robert C. Bunn, Mari Anne Jensen, and Brent C. Reed* The Department of Biochemistry and Molecular Biology, Louisiana State University School of Medicine, Shreveport, Louisiana 71130-3932 Submitted October 27, 1998; Accepted January 19, 1999 Monitoring Editor: Guido Guidotti Subcellular targeting and the activity of facilitative glucose transporters are likely to be regulated by interactions with cellular proteins. This report describes the identification and characterization of a protein, GLUT1 C-terminal binding protein (GLUT1CBP), that binds via a PDZ domain to the C terminus of GLUT1. The interaction requires the C-terminal four amino acids of GLUT1 and is isoform specific because GLUT1CBP does not interact with the C terminus of GLUT3 or GLUT4. Most rat tissues examined contain both GLUT1CBP and GLUT1 mRNA, whereas only small intestine lacked detectable GLUT1CBP protein. GLUT1CBP is also expressed in primary cultures of neurons and astrocytes, as well as in Chinese hamster ovary, 3T3-L1, Madin–Darby canine kidney, Caco-2, and pheochromocytoma-12 cell lines. GLUT1CBP is able to bind to native GLUT1 extracted from cell membranes, self-associate, or interact with the cytoskeletal proteins myosin VI, a-actinin-1, and the kinesin superfamily protein KIF-1B. The presence of a PDZ domain places GLUT1CBP among a growing family of structural and regulatory proteins, many of which are localized to areas of membrane specialization. This and its ability to interact with GLUT1 and cytoskeletal proteins implicate GLUT1CBP in cellular mechanisms for targeting GLUT1 to specific subcellular sites either by tethering the transporter to cytoskeletal motor proteins or by anchoring the transporter to the actin cytoskeleton. -
Regulation of Stringent Factor by Branched-Chain Amino Acids
Regulation of stringent factor by branched-chain amino acids Mingxu Fanga and Carl E. Bauera,1 aMolecular and Cellular Biochemistry Department, Indiana University, Bloomington, IN 47405 Edited by Caroline S. Harwood, University of Washington, Seattle, WA, and approved May 9, 2018 (received for review February 21, 2018) When faced with amino acid starvation, prokaryotic cells induce a Under normal growth conditions, the synthetase activity of Rel is stringent response that modulates their physiology. The stringent thought to be self-inhibited; however, during times of amino acid response is manifested by production of signaling molecules starvation, Rel interacts with stalled ribosomes, which activates guanosine 5′-diphosphate,3′-diphosphate (ppGpp) and guanosine synthetase activity to produce (p)ppGpp. The regulation of hy- 5′-triphosphate,3′-diphosphate (pppGpp) that are also called drolase activity is less understood but may involve one or more alarmones. In many species, alarmone levels are regulated by a downstream domains called the TGS and ACT domains. The TGS multidomain bifunctional alarmone synthetase/hydrolase called domain of SpoT has been shown to interact with an acyl carrier Rel. In this enzyme, there is an ACT domain at the carboxyl region protein, so it is presumed to sense the status of fatty acid metab- that has an unknown function; however, similar ACT domains are olism in E. coli (4). The function of the ACT domain is not as clear; present in other enzymes that have roles in controlling amino acid however, recent cryo-EM structures of E. coli RelA show that this metabolism. In many cases, these other ACT domains have been domain is involved in binding deacyl-tRNA as well as the ribosome shown to allosterically regulate enzyme activity through the bind- (5–7). -
The Alarmone (P)Ppgpp Is Part of the Heat Shock Response of Bacillus
bioRxiv preprint doi: https://doi.org/10.1101/688689; this version posted July 1, 2019. The copyright holder for this preprint (which was not certified by peer review) is the author/funder, who has granted bioRxiv a license to display the preprint in perpetuity. It is made available under aCC-BY 4.0 International license. 1 The alarmone (p)ppGpp is part of the heat shock response of Bacillus 2 subtilis 3 4 Heinrich Schäfer1,2, Bertrand Beckert3, Wieland Steinchen4, Aaron Nuss5, Michael 5 Beckstette5, Ingo Hantke1, Petra Sudzinová6, Libor Krásný6, Volkhard Kaever7, Petra 6 Dersch5,8, Gert Bange4*, Daniel Wilson3* & Kürşad Turgay1,2* 7 8 Author affiliations: 9 1 Institute of Microbiology, Leibniz Universität Hannover, Herrenhäuser Str. 2, D-30419, 10 Hannover, Germany. 11 2 Max Planck Unit for the Science of Pathogens, Charitéplatz 1, 10117 Berlin, Germany 12 3 Institute for Biochemistry and Molecular Biology, Martin-Luther-King Platz 6, University of 13 Hamburg, 20146 Hamburg, Germany. 14 4 Philipps-University Marburg, Center for Synthetic Microbiology (SYNMIKRO) and 15 Department of Chemistry, Hans-Meerwein-Strasse, 35043 Marburg, Germany. 16 5 Department of Molecular Infection Biology, Helmholtz Centre for Infection Research, 17 Braunschweig, Germany 18 6 Institute of Microbiology, Czech Academy of Sciences, Vídeňská 1083, 142 20, Prague, 19 Czech Republic. 20 7 Hannover Medical School, Research Core Unit Metabolomics, Carl-Neuberg-Strasse 1, 21 30625, Hannover, Germany. 22 8 Institute of Infectiology, Von-Esmarch-Straße 56, University of Münster, Münster, Germany 23 24 Short title: The interplay of heat shock and stringent response 25 1 bioRxiv preprint doi: https://doi.org/10.1101/688689; this version posted July 1, 2019. -
(P)Ppgpp Synthesis by the Ca2+-Dependent Rela-Spot Homolog
bioRxiv preprint doi: https://doi.org/10.1101/767004; this version posted September 12, 2019. The copyright holder for this preprint (which was not certified by peer review) is the author/funder. All rights reserved. No reuse allowed without permission. Plastidial (p)ppGpp synthesis by the Ca2+-dependent RelA-SpoT homolog regulates the adaptation of chloroplast gene expression to darkness in Arabidopsis Sumire Ono1,4, Sae Suzuki1,4, Doshun Ito1 Shota Tagawa2, Takashi Shiina2, and Shinji Masuda3,5 1School of Life Science & Technology, Tokyo Institute of Technology, Yokohama 226-8501, Japan 2Graduate School of Life and Environmental Sciences, Kyoto Prefectural University, Sakyo-ku, Kyoto 606-8522, Japan 3Center for Biological Resources & Informatics, Tokyo Institute of Technology, Yokohama 226-8501, Japan 4These authors contributed equally to the study 5Corresponding author: E-mail, [email protected] Abbreviations: CRSH, Ca2+-activated RSH; flg22, flagellin 22; (p)ppGpp, 5'-di(tri)phosphate 3'-diphosphate; PAMPs; pathogen-associated molecular patterns; RSH, RelA-SpoT homolog; qRT-PCR, quantitative real-time PCR 1 bioRxiv preprint doi: https://doi.org/10.1101/767004; this version posted September 12, 2019. The copyright holder for this preprint (which was not certified by peer review) is the author/funder. All rights reserved. No reuse allowed without permission. Abstract In bacteria, the hyper-phosphorylated nucleotides, guanosine 5'-diphosphate 3'-diphosphate (ppGpp) and guanosine 5'-triphosphate 3'-diphosphate (pppGpp), function as secondary messengers in the regulation of various metabolic processes of the cell, including transcription, translation, and enzymatic activities, especially under nutrient deficiency. The activity carried out by these nucleotide messengers is known as the stringent response. -
Tetracycline Modulates the Valine Induced Stringent Response and Decreases Expressed Rpos in Escherichia Coli
Journal of Experimental Microbiology and Immunology (JEMI) Vol. 15: 117 – 124 Copyright © April 2011, M&I UBC Tetracycline Modulates the Valine Induced Stringent Response and Decreases Expressed RpoS in Escherichia coli Natasha Castillon, Krysta Coyle, June Lai, and Cindy Yang Department of Microbiology & Immunology, University of British Columbia The stringent response in Escherichia coli is a physiological response to stress characterized by accumulation of the alarmone (p)ppGpp and the down-regulation of various processes involved in cellular growth and protein synthesis. It has been previously shown that activation of the stringent response by amino acid starvation is able to confer protection against various classes of antibiotics, including tetracycline and ampicillin. However, no observations have been made on partial activation of stringency. This investigation attempted to repress the level of bacterial stringent response in E. coli CP78 through the use of tetracycline. Tetracycline is an antibiotic which has been shown to restrict the cellular accumulation of (p)ppGpp in CP78 Escherichia coli. The effectiveness of the starvation by valine was monitored by measuring turbidity. The modulation of stringent response was monitored by the level of RpoS, since RpoS production is known to be enhanced by activated stringent response. As measured by turbidity, we successfully established an amino acid starvation condition with valine treatment as seen in the inhibited growth of CP78 cells as compared to untreated cells. The valine inhibition of the growth of the bacteria appeared to be reduced by the addition of tetracycline. However, cultures treated with high concentrations of tetracycline seemed to become more turbid than the untreated control even though they had similar levels of protein. -
Guanosine Pentaphosphate Phosphohydrolase of Escherichia Coli Is a Long-Chain Exopolyphosphatase J
Proc. Natl. Acad. Sci. USA Vol. 90, pp. 7029-7033, August 1993 Biochemistry Guanosine pentaphosphate phosphohydrolase of Escherichia coli is a long-chain exopolyphosphatase J. D. KEASLING*, LEROY BERTSCHt, AND ARTHUR KORNBERGtI *Department of Chemical Engineering, University of California, Berkeley, CA 94720-9989; and tDepartment of Biochemistry, Stanford University School of Medicine, Stanford, CA 94305-5307 Contributed by Arthur Kornberg, April 14, 1993 ABSTRACT An exopolyphosphatase [exopoly(P)ase; EC MATERIALS AND METHODS 3.6.1.11] activity has recently been purified to homogeneity from a mutant strain of Escherichia coi which lacks the Reagents and Proteins. Sources were as follows: ATP, principal exopoly(P)ase. The second exopoly(P)ase has now ADP, nonradiolabeled nucleotides, poly(P)s, bovine serum been identified as guanosine pentaphosphate phosphohydro- albumin, and ovalbumin from Sigma; [y-32P]ATP at 6000 lase (GPP; EC 3.6.1.40) by three lines of evidence: (i) the Ci/mmol (1 Ci = 37 GBq) and [y-32P]GTP at 6000 Ci/mmol sequences of five btptic digestion fragments of the purified from ICN; Q-Sepharose fast flow, catalase, aldolase, Super- protein are found in the translated gppA gene, (u) the size ofthe ose-12 fast protein liquid chromatography (FPLC) column, protein (100 kDa) agrees with published values for GPP, and and Chromatofocusing column and reagents from Pharmacia (iu) the ratio of exopoly(P)ase activity to GPP activity remains LKB; DEAE-Fractogel, Pll phosphocellulose, and DE52 constant throughout a 300-fold purification in the last steps of DEAE-cellulose from Whatman; protein standards for SDS/ the procedure. -
Diana Manuela Pinto Barros Classification and Structure-Based Inference of Transcriptional Regulatory Proteins
Universidade do Minho Departamento de Informatica´ Diana Manuela Pinto Barros Classification and Structure-Based Inference of Transcriptional Regulatory Proteins June, 2016 Universidade do Minho Departamento de Informatica´ Diana Manuela Pinto Barros Classification and Structure-Based Inference of Transcriptional Regulatory Proteins Tese de Mestrado Mestrado em Bioinformatica´ Trabalho efetuado sobre a orientac¸ao˜ de Sonia´ Carneiro Analia´ Lourenc¸o June, 2016 Acknowledgements I’d like to thank my advisors, Sonia´ Carneiro, for all of the essential input, guidance and time she has conceded this work, otherwise impossible to develop and Analia´ Lourenc¸o, for her valuable time and advices throughout this journey. I want to give the biggest thanks to my family. To my parents, Fernando and Inesˆ Barros for always believing in me and for giving me opportunities that weren’t given to them. Thank you for not giving up on me and for making me the person I am today. To my brothers, Sergio´ e Pedro. You are examples of perseverance and success that I can only hope to achieve one day. To my grandparents, Maria da Conceic¸ao˜ Silva e Antonio´ Ferreira Pinto for being the type of people I want to become some day and for being an example that I could always look up to. To my aunts: Em´ılia Pinto and Luz Pinto, my extra mothers, for being light in times of need and providing me the hope and strength when I had none left. I wouldn’t have made it this far without you. I want to thank all my friends that have accompanied me throughout this adventure.