Tn-seq of Caulobacter crescentus under uranium stress reveals genes essential for detoxification and stress tolerance
Abstract
Ubiquitous aquatic bacterium Caulobacter crescentus is highly resistant to uranium (U) and facilitates U biomineralization and thus holds promise as an agent of U bioremediation. In order to gain an understanding of how C. crescentus tolerates U, we employed transposon (Tn) mutagenesis paired with deep sequencing (Tn-seq) in a global screen for genomic elements required for U resistance. Of the 3,879 annotated genes in the C. crescentus genome, 37 were found to be specifically associated with fitness under U stress, 15 of which were subsequently tested through mutational analysis. Systematic deletion analysis revealed that mutants lacking outer membrane transporters (rsaFa and rsaFb), a stress-responsive transcription factor (cztR), or a ppGpp synthetase/hydrolase (spoT) exhibited a significantly lower survival rate under U stress. RsaFa and RsaFb, which are homologues of TolC in Escherichia coli, have previously been shown to mediate S-layer export. Transcriptional analysis revealed upregulation of rsaFa and rsaFb by 4- and 10-fold, respectively, in the presence of U. We additionally show that rsaFa mutants accumulated higher levels of U than the wild type, with no significant increase in oxidative stress levels. These results suggest a function for RsaFa and RsaFb in U efflux and/or maintenance of membrane integrity during Umore »
- Authors:
-
- Lawrence Livermore National Lab. (LLNL), Livermore, CA (United States)
- DOE Joint Genome Inst., Walnut Creek, CA (United States)
- Univ. of British Columbia, Vancouver, BC (Canada)
- California State Univ., Northridge, CA (United States)
- Stanford Univ. School of Medicine, Stanford, CA (United States)
- ETH Zurich (Switzerland)
- Univ. of Wyoming, Laramie, WY (United States)
- Publication Date:
- Research Org.:
- Lawrence Livermore National Lab. (LLNL), Livermore, CA (United States)
- Sponsoring Org.:
- USDOE
- OSTI Identifier:
- 1234593
- Report Number(s):
- LLNL-JRNL-670205
Journal ID: ISSN 0021-9193; TRN: US1600611
- Grant/Contract Number:
- AC52-07NA27344
- Resource Type:
- Accepted Manuscript
- Journal Name:
- Journal of Bacteriology
- Additional Journal Information:
- Journal Volume: 197; Journal Issue: 19; Journal ID: ISSN 0021-9193
- Publisher:
- American Society for Microbiology
- Country of Publication:
- United States
- Language:
- English
- Subject:
- 63 RADIATION, THERMAL, AND OTHER ENVIRONMENTAL POLLUTANT EFFECTS ON LIVING ORGANISMS AND BIOLOGICAL MATERIALS; 59 BASIC BIOLOGICAL SCIENCES
Citation Formats
Yung, Mimi C., Park, Dan M., Overton, K. Wesley, Blow, Matthew J., Hoover, Cindi A., Smit, John R., Murray, Sean R., Ricci, Dante P., Christen, Beat, Bowman, Grant R., and Jiao, Yongqin. Tn-seq of Caulobacter crescentus under uranium stress reveals genes essential for detoxification and stress tolerance. United States: N. p., 2015.
Web. doi:10.1128/JB.00382-15.
Yung, Mimi C., Park, Dan M., Overton, K. Wesley, Blow, Matthew J., Hoover, Cindi A., Smit, John R., Murray, Sean R., Ricci, Dante P., Christen, Beat, Bowman, Grant R., & Jiao, Yongqin. Tn-seq of Caulobacter crescentus under uranium stress reveals genes essential for detoxification and stress tolerance. United States. https://doi.org/10.1128/JB.00382-15
Yung, Mimi C., Park, Dan M., Overton, K. Wesley, Blow, Matthew J., Hoover, Cindi A., Smit, John R., Murray, Sean R., Ricci, Dante P., Christen, Beat, Bowman, Grant R., and Jiao, Yongqin. Mon .
"Tn-seq of Caulobacter crescentus under uranium stress reveals genes essential for detoxification and stress tolerance". United States. https://doi.org/10.1128/JB.00382-15. https://www.osti.gov/servlets/purl/1234593.
@article{osti_1234593,
title = {Tn-seq of Caulobacter crescentus under uranium stress reveals genes essential for detoxification and stress tolerance},
author = {Yung, Mimi C. and Park, Dan M. and Overton, K. Wesley and Blow, Matthew J. and Hoover, Cindi A. and Smit, John R. and Murray, Sean R. and Ricci, Dante P. and Christen, Beat and Bowman, Grant R. and Jiao, Yongqin},
abstractNote = {Ubiquitous aquatic bacterium Caulobacter crescentus is highly resistant to uranium (U) and facilitates U biomineralization and thus holds promise as an agent of U bioremediation. In order to gain an understanding of how C. crescentus tolerates U, we employed transposon (Tn) mutagenesis paired with deep sequencing (Tn-seq) in a global screen for genomic elements required for U resistance. Of the 3,879 annotated genes in the C. crescentus genome, 37 were found to be specifically associated with fitness under U stress, 15 of which were subsequently tested through mutational analysis. Systematic deletion analysis revealed that mutants lacking outer membrane transporters (rsaFa and rsaFb), a stress-responsive transcription factor (cztR), or a ppGpp synthetase/hydrolase (spoT) exhibited a significantly lower survival rate under U stress. RsaFa and RsaFb, which are homologues of TolC in Escherichia coli, have previously been shown to mediate S-layer export. Transcriptional analysis revealed upregulation of rsaFa and rsaFb by 4- and 10-fold, respectively, in the presence of U. We additionally show that rsaFa mutants accumulated higher levels of U than the wild type, with no significant increase in oxidative stress levels. These results suggest a function for RsaFa and RsaFb in U efflux and/or maintenance of membrane integrity during U stress. In addition, we present data implicating CztR and SpoT in resistance to U stress. Together, our findings reveal novel gene targets that are key to understanding the molecular mechanisms of U resistance in C. crescentus.},
doi = {10.1128/JB.00382-15},
journal = {Journal of Bacteriology},
number = 19,
volume = 197,
place = {United States},
year = {Mon Jul 20 00:00:00 EDT 2015},
month = {Mon Jul 20 00:00:00 EDT 2015}
}
Web of Science
Works referenced in this record:
Extracellular reduction of uranium via Geobacter conductive pili as a protective cellular mechanism
journal, September 2011
- Cologgi, D. L.; Lampa-Pastirk, S.; Speers, A. M.
- Proceedings of the National Academy of Sciences, Vol. 108, Issue 37, p. 15248-15252
Modulation of Medium pH by Caulobacter crescentus Facilitates Recovery from Uranium-Induced Growth Arrest
journal, July 2014
- Park, Dan M.; Jiao, Yongqin
- Applied and Environmental Microbiology, Vol. 80, Issue 18
SpoT Regulates DnaA Stability and Initiation of DNA Replication in Carbon-Starved Caulobacter crescentus
journal, August 2008
- Lesley, J. A.; Shapiro, L.
- Journal of Bacteriology, Vol. 190, Issue 20
Detection of Thorium and Uranium
journal, June 1958
- Fritz, J. S.; Bradford, E. C.
- Analytical Chemistry, Vol. 30, Issue 6
Evaluating secretion and surface attachment of SapA, an S-layer-associated metalloprotease of Caulobacter crescentus
journal, May 2012
- Gandham, Lyngrace; Nomellini, John F.; Smit, John
- Archives of Microbiology, Vol. 194, Issue 10
The tolC locus affects the expression of sbmA through σE activity increase: Upregulation of sbmA expression by tolC mutation
journal, August 2010
- Corbalán, Natalia S.; Adler, Conrado; De Cristóbal, Ricardo E.
- FEMS Microbiology Letters, Vol. 311, Issue 2
Identification of host genes that affect acquisition of an integrative and conjugative element in B acillus subtilis: Genes in recipients that affect conjugation
journal, August 2014
- Johnson, Christopher M.; Grossman, Alan D.
- Molecular Microbiology
Uranium Interaction with Two Multi-Resistant Environmental Bacteria: Cupriavidus metallidurans CH34 and Rhodopseudomonas palustris
journal, December 2012
- Llorens, Isabelle; Untereiner, Guillaume; Jaillard, Danielle
- PLoS ONE, Vol. 7, Issue 12
Uranium Speciation and Bioavailability in Aquatic Systems: An Overview
journal, January 2002
- Markich, Scott J.
- The Scientific World JOURNAL, Vol. 2
Metals, minerals and microbes: geomicrobiology and bioremediation
journal, December 2009
- Gadd, G. M.
- Microbiology, Vol. 156, Issue 3
Fast and accurate short read alignment with Burrows-Wheeler transform
journal, May 2009
- Li, H.; Durbin, R.
- Bioinformatics, Vol. 25, Issue 14
Uranium and other heavy metal resistance and accumulation in bacteria isolated from uranium mine wastes
journal, March 2012
- Choudhary, Sangeeta; Islam, Ekramul; Kazy, Sufia K.
- Journal of Environmental Science and Health, Part A, Vol. 47, Issue 4
The Caulobacter crescentus Paracrystalline S-Layer Protein Is Secreted by an ABC Transporter (Type I) Secretion Apparatus
journal, June 1998
- Awram, Peter; Smit, John
- Journal of Bacteriology, Vol. 180, Issue 12
Whole-Genome Transcriptional Analysis of Heavy Metal Stresses in Caulobacter crescentus
journal, November 2005
- Hu, P.; Brodie, E. L.; Suzuki, Y.
- Journal of Bacteriology, Vol. 187, Issue 24
The Response of Desulfotomaculum reducens MI-1 to U(VI) Exposure: A Transcriptomic Study
journal, June 2011
- Junier, Pilar; Vecchia, Elena Dalla; Bernier-Latmani, Rizlan
- Geomicrobiology Journal, Vol. 28, Issue 5-6
Caulobacter crescentus as a Whole-Cell Uranium Biosensor
journal, September 2007
- Hillson, N. J.; Hu, P.; Andersen, G. L.
- Applied and Environmental Microbiology, Vol. 73, Issue 23
Two RND proteins involved in heavy metal efflux in Caulobacter crescentus belong to separate clusters within proteobacteria
journal, January 2013
- Valencia, Estela Y.; Braz, Vânia S.; Guzzo, Cristiane
- BMC Microbiology, Vol. 13, Issue 1
Trimmomatic: a flexible trimmer for Illumina sequence data
journal, April 2014
- Bolger, Anthony M.; Lohse, Marc; Usadel, Bjoern
- Bioinformatics, Vol. 30, Issue 15
Genome-Scale Identification of Resistance Functions in Pseudomonas aeruginosa Using Tn-seq
journal, January 2011
- Gallagher, Larry A.; Shendure, Jay; Manoil, Colin
- mBio, Vol. 2, Issue 1
Two Outer Membrane Proteins Are Required for Maximal Type I Secretion of the Caulobacter crescentus S-Layer Protein
journal, November 2004
- Toporowski, M. C.; Nomellini, J. F.; Awram, P.
- Journal of Bacteriology, Vol. 186, Issue 23
UDP- N -Acetylmuramic Acid l-Alanine Ligase (MurC) Inhibition in a tolC Mutant Escherichia coli Strain Leads to Cell Death
journal, August 2014
- Humnabadkar, Vaishali; Prabhakar, K. R.; Narayan, Ashwini
- Antimicrobial Agents and Chemotherapy, Vol. 58, Issue 10
Global Transcriptional Profiling of Shewanella oneidensis MR-1 during Cr(VI) and U(VI) Reduction
journal, November 2005
- Bencheikh-Latmani, R.; Williams, S. M.; Haucke, L.
- Applied and Environmental Microbiology, Vol. 71, Issue 11
Uranium extremophily is an adaptive, rather than intrinsic, feature for extremely thermoacidophilic Metallosphaera species
journal, September 2012
- Mukherjee, A.; Wheaton, G. H.; Blum, P. H.
- Proceedings of the National Academy of Sciences, Vol. 109, Issue 41
Importance of c-Type cytochromes for U(VI) reduction by Geobacter sulfurreducens
journal, January 2007
- Shelobolina, Evgenya S.; Coppi, Maddalena V.; Korenevsky, Anton A.
- BMC Microbiology, Vol. 7, Issue 1
The porin and the permeating antibiotic: a selective diffusion barrier in Gram-negative bacteria
journal, November 2008
- Pagès, Jean-Marie; James, Chloë E.; Winterhalter, Mathias
- Nature Reviews Microbiology, Vol. 6, Issue 12
The essential genome of a bacterium
journal, January 2011
- Christen, Beat; Abeliuk, Eduardo; Collier, John M.
- Molecular Systems Biology, Vol. 7, Issue 1
The Challenge of Efflux-Mediated Antibiotic Resistance in Gram-Negative Bacteria
journal, March 2015
- Li, Xian-Zhi; Plésiat, Patrick; Nikaido, Hiroshi
- Clinical Microbiology Reviews, Vol. 28, Issue 2
Multiplexed Digital mRNA Profiling of the Inflammatory Response in the West Nile Swiss Webster Mouse Model
journal, October 2014
- Peña, José; Plante, Jessica A.; Carillo, Alda Celena
- PLoS Neglected Tropical Diseases, Vol. 8, Issue 10
Biomineralization of Uranium by PhoY Phosphatase Activity Aids Cell Survival in Caulobacter crescentus
journal, May 2014
- Yung, Mimi C.; Jiao, Yongqin
- Applied and Environmental Microbiology, Vol. 80, Issue 16
A fine scale phenotype-genotype virulence map of a bacterial pathogen
journal, July 2012
- van Opijnen, T.; Camilli, A.
- Genome Research, Vol. 22, Issue 12
Genes required for alleviation of uranium toxicity in sulfate reducing bacterium Desulfovibio alaskensis G20
journal, February 2014
- Li, Xiangkai; Zhang, He; Ma, Yantian
- Ecotoxicology, Vol. 23, Issue 4
Yeast Genes Involved in Uranium Tolerance and Uranium Accumulation: A Functional Screening Using the Nonessential Gene Deletion Collection
journal, June 2012
- Sakamoto, Fuminori; Nankawa, Takuya; Ohnuki, Toshihiko
- Geomicrobiology Journal, Vol. 29, Issue 5
The complex logic of stringent response regulation in Caulobacter crescentus: starvation signalling in an oligotrophic environment: Characterization of the stringent response in a freshwater oligotroph
journal, March 2011
- Boutte, Cara C.; Crosson, Sean
- Molecular Microbiology, Vol. 80, Issue 3
NIH Image to ImageJ: 25 years of image analysis
journal, June 2012
- Schneider, Caroline A.; Rasband, Wayne S.; Eliceiri, Kevin W.
- Nature Methods, Vol. 9, Issue 7
BLAST: a more efficient report with usability improvements
journal, April 2013
- Boratyn, Grzegorz M.; Camacho, Christiam; Cooper, Peter S.
- Nucleic Acids Research, Vol. 41, Issue W1
CztR, a LysR-Type Transcriptional Regulator Involved in Zinc Homeostasis and Oxidative Stress Defense in Caulobacter crescentus
journal, August 2010
- Braz, Vânia S.; da Silva Neto, José F.; Italiani, Valéria C. S.
- Journal of Bacteriology, Vol. 192, Issue 20
Genes involved in cadmium resistance in Caulobacter crescentus
journal, October 2005
- Braz, Vânia S.; Marques, Marilis V.
- FEMS Microbiology Letters, Vol. 251, Issue 2
Evidence-Based Annotation of Gene Function in Shewanella oneidensis MR-1 Using Genome-Wide Fitness Profiling across 121 Conditions
journal, November 2011
- Deutschbauer, Adam; Price, Morgan N.; Wetmore, Kelly M.
- PLoS Genetics, Vol. 7, Issue 11
c-Type Cytochrome-Dependent Formation of U(IV) Nanoparticles by Shewanella oneidensis
journal, August 2006
- Marshall, Matthew J.; Beliaev, Alexander S.; Dohnalkova, Alice C.
- PLoS Biology, Vol. 4, Issue 8
Shotgun Proteomic Analysis Unveils Survival and Detoxification Strategies by Caulobacter crescentus during Exposure to Uranium, Chromium, and Cadmium
journal, March 2014
- Yung, Mimi C.; Ma, Jincai; Salemi, Michelle R.
- Journal of Proteome Research, Vol. 13, Issue 4
New horizons for (p)ppGpp in bacterial and plant physiology
journal, January 2006
- Braeken, Kristien; Moris, Martine; Daniels, Ruth
- Trends in Microbiology, Vol. 14, Issue 1
Bio-precipitation of uranium by two bacterial isolates recovered from extreme environments as estimated by potentiometric titration, TEM and X-ray absorption spectroscopic analyses
journal, December 2011
- Merroun, Mohamed L.; Nedelkova, Marta; Ojeda, Jesus J.
- Journal of Hazardous Materials, Vol. 197
Effects of uranium and phosphate concentrations on oxidative stress related responses induced in Arabidopsis thaliana
journal, November 2008
- Vanhoudt, Nathalie; Vandenhove, Hildegarde; Smeets, Karen
- Plant Physiology and Biochemistry, Vol. 46, Issue 11
Uranium phosphate biomineralization by fungi: Uranium biomineralization by fungi
journal, March 2015
- Liang, Xinjin; Hillier, Stephen; Pendlowski, Helen
- Environmental Microbiology, Vol. 17, Issue 6
Additive effect of tolC and rfa mutations on the hydrophobic barrier of the outer membrane of Escherichia coli K-12.
journal, January 1994
- Fralick, J. A.; Burns-Keliher, L. L.
- Journal of Bacteriology, Vol. 176, Issue 20
ESSENTIALS: Software for Rapid Analysis of High Throughput Transposon Insertion Sequencing Data
journal, August 2012
- Zomer, Aldert; Burghout, Peter; Bootsma, Hester J.
- PLoS ONE, Vol. 7, Issue 8
Microbial heavy-metal resistance
journal, June 1999
- Nies, D. H.
- Applied Microbiology and Biotechnology, Vol. 51, Issue 6
Efflux-mediated heavy metal resistance in prokaryotes
journal, June 2003
- Nies, Dietrich H.
- FEMS Microbiology Reviews, Vol. 27, Issue 2-3
Structure and Function of TolC: The Bacterial Exit Duct for Proteins and Drugs
journal, June 2004
- Koronakis, Vassilis; Eswaran, Jeyanthy; Hughes, Colin
- Annual Review of Biochemistry, Vol. 73, Issue 1
Escherichia coli Response to Uranyl Exposure at Low pH and Associated Protein Regulations
journal, February 2014
- Khemiri, Arbia; Carrière, Marie; Bremond, Nicolas
- PLoS ONE, Vol. 9, Issue 2
Genome-Wide Mutant Fitness Profiling Identifies Nutritional Requirements for Optimal Growth of Yersinia pestis in Deep Tissue
journal, August 2014
- Palace, Samantha G.; Proulx, Megan K.; Lu, Shan
- mBio, Vol. 5, Issue 4
U(VI) reduction by spores of Clostridium acetobutylicum
journal, November 2010
- Vecchia, Elena Dalla; Veeramani, Harish; Suvorova, Elena I.
- Research in Microbiology, Vol. 161, Issue 9
Two-Component Signal Transduction Pathways Regulating Growth and Cell Cycle Progression in a Bacterium: A System-Level Analysis
journal, September 2005
- Skerker, Jeffrey M.; Prasol, Melanie S.; Perchuk, Barrett S.
- PLoS Biology, Vol. 3, Issue 10
MochiView: versatile software for genome browsing and DNA motif analysis
journal, April 2010
- Homann, Oliver R.; Johnson, Alexander D.
- BMC Biology, Vol. 8, Issue 1
Tn-seq: high-throughput parallel sequencing for fitness and genetic interaction studies in microorganisms
journal, September 2009
- van Opijnen, Tim; Bodi, Kip L.; Camilli, Andrew
- Nature Methods, Vol. 6, Issue 10
Accumulation of periplasmic enterobactin impairs the growth and morphology of Escherichia coli tolC mutants: Δ tolC phenotypes caused by periplasmic enterobactin
journal, December 2013
- Vega, Daniel E.; Young, Kevin D.
- Molecular Microbiology, Vol. 91, Issue 3
Mechanism and Inhibition of LpxC: An Essential Zinc-Dependent Deacetylase of Bacterial Lipid A Synthesis
journal, February 2008
- Zhou, Pei; Barb, Adam
- Current Pharmaceutical Biotechnology, Vol. 9, Issue 1
Reduction of Cellular Stress by TolC-Dependent Efflux Pumps in Escherichia coli Indicated by BaeSR and CpxARP Activation of spy in Efflux Mutants
journal, December 2012
- Rosner, J. L.; Martin, R. G.
- Journal of Bacteriology, Vol. 195, Issue 5
Metabolic shutdown in Escherichia coli cells lacking the outer membrane channel TolC: Metabolic shutdown in TolC mutants
journal, June 2010
- Dhamdhere, Girija; Zgurskaya, Helen I.
- Molecular Microbiology, Vol. 77, Issue 3
Decrease of U(VI) Immobilization Capability of the Facultative Anaerobic Strain Paenibacillus sp. JG-TB8 under Anoxic Conditions Due to Strongly Reduced Phosphatase Activity
journal, August 2014
- Reitz, Thomas; Rossberg, Andre; Barkleit, Astrid
- PLoS ONE, Vol. 9, Issue 8
An nptI-sacB-sacR cartridge for constructing directed, unmarked mutations in Gram-negative bacteria by marker exchange-eviction mutagenesis
journal, January 1987
- Ried, Jeffrey L.; Collmer, Alan
- Gene, Vol. 57, Issue 2-3
Complete genome sequence of Caulobacter crescentus
journal, March 2001
- Nierman, W. C.; Feldblyum, T. V.; Laub, M. T.
- Proceedings of the National Academy of Sciences, Vol. 98, Issue 7
Complexation of Uranium by Cells and S-Layer Sheets of Bacillus sphaericus JG-A12
journal, September 2005
- Merroun, M. L.; Raff, J.; Rossberg, A.
- Applied and Environmental Microbiology, Vol. 71, Issue 9
A cell cycle-regulated bacterial DNA methyltransferase is essential for viability.
journal, February 1996
- Stephens, C.; Reisenauer, A.; Wright, R.
- Proceedings of the National Academy of Sciences, Vol. 93, Issue 3
Cutadapt removes adapter sequences from high-throughput sequencing reads
journal, May 2011
- Martin, Marcel
- EMBnet.journal, Vol. 17, Issue 1
Works referencing / citing this record:
The Use of Transposon Insertion Sequencing to Interrogate the Core Functional Genome of the Legume Symbiont Rhizobium leguminosarum
journal, November 2016
- Perry, Benjamin J.; Akter, Mir S.; Yost, Christopher K.
- Frontiers in Microbiology, Vol. 7
Molecular insight into the expression of metal transporter genes in Chryseobacterium sp. PMSZPI isolated from uranium deposit
journal, May 2019
- Nongkhlaw, Macmillan; Joshi, Santa Ram
- PLOS ONE, Vol. 14, Issue 5
Complete genome sequence of Caulobacter flavus RHGG3T, a type species of the genus Caulobacter with plant growth-promoting traits and heavy metal resistance
journal, January 2019
- Yang, Endong; Sun, Leni; Ding, Xiaoyuan
- 3 Biotech, Vol. 9, Issue 2
Transposon insertional mutagenesis in Saccharomyces uvarum reveals trans -acting effects influencing species-dependent essential genes
journal, January 2019
- Sanchez, Monica R.; Payen, Celia; Cheong, Frances
- Genome Research, Vol. 29, Issue 3