Skip to main content
  • ASM
    • Antimicrobial Agents and Chemotherapy
    • Applied and Environmental Microbiology
    • Clinical Microbiology Reviews
    • Clinical and Vaccine Immunology
    • EcoSal Plus
    • Eukaryotic Cell
    • Infection and Immunity
    • Journal of Bacteriology
    • Journal of Clinical Microbiology
    • Journal of Microbiology & Biology Education
    • Journal of Virology
    • mBio
    • Microbiology and Molecular Biology Reviews
    • Microbiology Resource Announcements
    • Microbiology Spectrum
    • Molecular and Cellular Biology
    • mSphere
    • mSystems
  • Log in
  • My alerts
  • My Cart

Main menu

  • Home
  • Articles
    • Current Issue
    • Accepted Manuscripts
    • Archive
    • Minireviews
    • JB Special Collection
    • JB Classic Spotlights
  • For Authors
    • Submit a Manuscript
    • Scope
    • Editorial Policy
    • Submission, Review, & Publication Processes
    • Organization and Format
    • Errata, Author Corrections, Retractions
    • Illustrations and Tables
    • Nomenclature
    • Abbreviations and Conventions
    • Publication Fees
    • Ethics Resources and Policies
  • About the Journal
    • About JB
    • Editor in Chief
    • Editorial Board
    • For Reviewers
    • For the Media
    • For Librarians
    • For Advertisers
    • Alerts
    • RSS
    • FAQ
  • Subscribe
    • Members
    • Institutions
  • ASM
    • Antimicrobial Agents and Chemotherapy
    • Applied and Environmental Microbiology
    • Clinical Microbiology Reviews
    • Clinical and Vaccine Immunology
    • EcoSal Plus
    • Eukaryotic Cell
    • Infection and Immunity
    • Journal of Bacteriology
    • Journal of Clinical Microbiology
    • Journal of Microbiology & Biology Education
    • Journal of Virology
    • mBio
    • Microbiology and Molecular Biology Reviews
    • Microbiology Resource Announcements
    • Microbiology Spectrum
    • Molecular and Cellular Biology
    • mSphere
    • mSystems

User menu

  • Log in
  • My alerts
  • My Cart

Search

  • Advanced search
Journal of Bacteriology
publisher-logosite-logo

Advanced Search

  • Home
  • Articles
    • Current Issue
    • Accepted Manuscripts
    • Archive
    • Minireviews
    • JB Special Collection
    • JB Classic Spotlights
  • For Authors
    • Submit a Manuscript
    • Scope
    • Editorial Policy
    • Submission, Review, & Publication Processes
    • Organization and Format
    • Errata, Author Corrections, Retractions
    • Illustrations and Tables
    • Nomenclature
    • Abbreviations and Conventions
    • Publication Fees
    • Ethics Resources and Policies
  • About the Journal
    • About JB
    • Editor in Chief
    • Editorial Board
    • For Reviewers
    • For the Media
    • For Librarians
    • For Advertisers
    • Alerts
    • RSS
    • FAQ
  • Subscribe
    • Members
    • Institutions
Articles

Hfq Influences Multiple Transport Systems and Virulence in the Plant Pathogen Agrobacterium tumefaciens

Ina Wilms, Philip Möller, Anna-Maria Stock, Rosemarie Gurski, Erh-Min Lai, Franz Narberhaus
Ina Wilms
Lehrstuhl für Biologie der Mikroorganismen, Ruhr-Universität Bochum, Bochum, Germany
  • Find this author on Google Scholar
  • Find this author on PubMed
  • Search for this author on this site
Philip Möller
Lehrstuhl für Biologie der Mikroorganismen, Ruhr-Universität Bochum, Bochum, Germany
  • Find this author on Google Scholar
  • Find this author on PubMed
  • Search for this author on this site
Anna-Maria Stock
Lehrstuhl für Biologie der Mikroorganismen, Ruhr-Universität Bochum, Bochum, Germany
  • Find this author on Google Scholar
  • Find this author on PubMed
  • Search for this author on this site
Rosemarie Gurski
Lehrstuhl für Biologie der Mikroorganismen, Ruhr-Universität Bochum, Bochum, Germany
  • Find this author on Google Scholar
  • Find this author on PubMed
  • Search for this author on this site
Erh-Min Lai
Institute of Plant and Microbial Biology, Academia Sinica, Taipei, Taiwan
  • Find this author on Google Scholar
  • Find this author on PubMed
  • Search for this author on this site
Franz Narberhaus
Lehrstuhl für Biologie der Mikroorganismen, Ruhr-Universität Bochum, Bochum, Germany
  • Find this author on Google Scholar
  • Find this author on PubMed
  • Search for this author on this site
DOI: 10.1128/JB.00510-12
  • Article
  • Figures & Data
  • Info & Metrics
  • PDF
Loading

Article Figures & Data

Figures

  • Additional Files
  • Fig 1
    • Open in new tab
    • Download powerpoint
    Fig 1

    Complex formation between Hfq, the sRNA AbcR1, and its target atu2422. (A) Gel retardation experiments with purified Hfq protein and AbcR1. 32P-labeled AbcR1 sRNA (0.07 pmol) was incubated with increasing concentrations of Hfq protein at 30°C for 20 min. Final concentrations (fold excess) of added Hfq are shown above the gel. Lane 1 contains a water control. (B) Gel retardation experiments with AbcR1, Hfq, and the target atu2422 mRNA fragment (100 nucleotides). 32P-labeled AbcR1 sRNA (0.07 pmol) was incubated with purified Hfq (1,000-fold excess) and increasing concentrations of atu2422. Final concentrations (fold excess) of added atu2422 RNA are shown above the gel. Lane 1 contains a water control. Samples were run on a 6% native gel at 250 V. The asterisks indicate radioactive (32P) labeling of the sRNA AbcR1.

  • Fig 2
    • Open in new tab
    • Download powerpoint
    Fig 2

    Effect of Hfq on AbcR1-mediated gene regulation in vivo. (A) Northern blot analysis of AbcR1 and atu2422 in A. tumefaciens wild type (WT) and the Δhfq strain. Hybridizations were performed with 8 μg of total RNA from cells grown in YEB complex medium to the optical densities indicated above the gels. Primers used for RNA probe generation are listed in Table S2 in the supplemental material. Ethidium bromide-stained 23S RNAs or tRNAs were used as loading controls. nt, nucleotides. (B) Stability of AbcR1. WT and Δhfq mutant cells were grown to an OD600 of 1.0 in YEB medium. Samples were taken before (0) and 1, 5, and 10 min after the addition of rifampin (250 μg/ml). Amounts of 8 μg of total RNA were separated on a 10% polyacrylamide gel containing 7 M urea and were detected by Northern analysis using a DNA probe (generated by primers DNAprobe_C2A_fw and DNAprobe_C2A_rv). The size of AbcR1 is given on the right. Ethidium bromide-stained tRNAs were used as loading controls. (C) Relative amounts of AbcR1 in A. tumefaciens WT and the hfq mutant after the addition of rifampin. Northern blot signals (B) were quantified using AlphaEaseFC software.

  • Fig 3
    • Open in new tab
    • Download powerpoint
    Fig 3

    Altered protein levels in the hfq deletion mutant. (A) Total protein samples from different growth phases (indicated above the gels) of A. tumefaciens wild type (WT), the Δhfq strain, the complemented mutant (Δhfq+hfq), and control strains containing the empty vector (v) (WT+v and Δhfq+v) were loaded on a 12% SDS–PAGE gel. The positions of marker proteins are given on the left in kDa. Eight prominent protein bands accumulating in the mutant were analyzed via MALDI-TOF. The proteins identified with the highest confidence are given on the right. ABC transporter components are underlined. (B) Identification of overexpressed proteins (A) in the Δhfq mutant by MALDI-TOF. Proteins that were ranked at the first position after MALDI-TOF analysis are in boldface and marked by asterisks. All other listed proteins were additionally identified during mass spectrometry.

  • Fig 4
    • Open in new tab
    • Download powerpoint
    Fig 4

    Altered RNA levels in the hfq deletion mutant. Northern blots determining RNA levels of some of the proteins identified by SDS-PAGE in Fig. 3A. Hybridizations were performed with amounts of 8 μg of total RNA from A. tumefaciens wild type (WT) and the Δhfq mutant grown to the optical densities indicated above. Primers used for RNA probe generation are listed in Table S2 in the supplemental material. Ethidium bromide-stained 23S RNAs were used as loading controls.

  • Fig 5
    • Open in new tab
    • Download powerpoint
    Fig 5

    Growth defect and altered cell morphology of the Δhfq strain. (A) A. tumefaciens wild type (WT), the Δhfq mutant, the complemented mutant (Δhfq+hfq), and corresponding control strains harboring the empty vector (v) (WT+v and Δhfq+v) were grown in YEB complex medium, and OD600 values were plotted over time. Three independent experiments gave comparable results. (B) Microscopic images were taken of A. tumefaciens strains from cultures grown in YEB medium to optical densities indicated above the images. Y-shaped cells are indicated by arrows.

  • Fig 6
    • Open in new tab
    • Download powerpoint
    Fig 6

    Virulence defect of the A. tumefaciens Δhfq strain on potato discs. Quantitative tumor formation assay on potato tuber discs. (A) Amounts of 104 and 106 cells/disc of A. tumefaciens wild type (WT), the Δhfq mutant, the complemented mutant (Δhfq+hfq), and control strains harboring the empty vector (v) (WT+v and Δhfq+v) were examined for their tumor formation efficiency on potato discs. Tumorigenesis efficiency is scored by the number of tumors per disc (mean value calculated from results of 60 potato tuber discs for each strain in each independent experiment; error bars show standard errors of the means). (B) Representative pictures of tumors on potato discs. Two independent experiments gave similar results.

  • Fig 7
    • Open in new tab
    • Download powerpoint
    Fig 7

    Presence of T4SS proteins in the WT and hfq mutant. Cells were grown under non-virulence-inducing (− AS) or virulence-inducing conditions (+ AS [1 mM]) in AB minimal medium (AS, acetosyringone). Cell lysates were subjected to SDS-PAGE, followed by Western blotting and detection with specific antisera.

  • Fig 8
    • Open in new tab
    • Download powerpoint
    Fig 8

    Current model of Hfq-dependent processes in A. tumefaciens. A schematic A. tumefaciens cell is depicted with inner and outer membrane (IM and OM, respectively). mRNAs influenced by Hfq mostly code for periplasmic proteins. Hfq-mediated regulation is exerted via the sRNA AbcR1 (69) (continuous lines) or via yet-to-be-determined sRNAs (dashed lines).

Additional Files

  • Figures
  • Supplemental material

    Files in this Data Supplement:

    • Supplemental file 1 -

      Table S1, strains and plasmids

      Table S2, oligonucleotides

      Fig. S1, purification of the Hfq protein

      Fig. S2, verification of the chromosomal hfq deletion

      Fig. S3, altered tumor formation by the Δhfq mutant on plants

      PDF, 2.8M

PreviousNext
Back to top
Download PDF
Citation Tools
Hfq Influences Multiple Transport Systems and Virulence in the Plant Pathogen Agrobacterium tumefaciens
Ina Wilms, Philip Möller, Anna-Maria Stock, Rosemarie Gurski, Erh-Min Lai, Franz Narberhaus
Journal of Bacteriology Sep 2012, 194 (19) 5209-5217; DOI: 10.1128/JB.00510-12

Citation Manager Formats

  • BibTeX
  • Bookends
  • EasyBib
  • EndNote (tagged)
  • EndNote 8 (xml)
  • Medlars
  • Mendeley
  • Papers
  • RefWorks Tagged
  • Ref Manager
  • RIS
  • Zotero
Print

Alerts
Sign In to Email Alerts with your Email Address
Email

Thank you for sharing this Journal of Bacteriology article.

NOTE: We request your email address only to inform the recipient that it was you who recommended this article, and that it is not junk mail. We do not retain these email addresses.

Enter multiple addresses on separate lines or separate them with commas.
Hfq Influences Multiple Transport Systems and Virulence in the Plant Pathogen Agrobacterium tumefaciens
(Your Name) has forwarded a page to you from Journal of Bacteriology
(Your Name) thought you would be interested in this article in Journal of Bacteriology.
Share
Hfq Influences Multiple Transport Systems and Virulence in the Plant Pathogen Agrobacterium tumefaciens
Ina Wilms, Philip Möller, Anna-Maria Stock, Rosemarie Gurski, Erh-Min Lai, Franz Narberhaus
Journal of Bacteriology Sep 2012, 194 (19) 5209-5217; DOI: 10.1128/JB.00510-12
del.icio.us logo Digg logo Reddit logo Twitter logo CiteULike logo Facebook logo Google logo Mendeley logo
  • Top
  • Article
    • ABSTRACT
    • INTRODUCTION
    • MATERIALS AND METHODS
    • RESULTS
    • DISCUSSION
    • ACKNOWLEDGMENTS
    • FOOTNOTES
    • REFERENCES
  • Figures & Data
  • Info & Metrics
  • PDF

Related Articles

Cited By...

About

  • About JB
  • Editor in Chief
  • Editorial Board
  • Policies
  • For Reviewers
  • For the Media
  • For Librarians
  • For Advertisers
  • Alerts
  • RSS
  • FAQ
  • Permissions
  • Journal Announcements

Authors

  • ASM Author Center
  • Submit a Manuscript
  • Article Types
  • Ethics
  • Contact Us

Follow #Jbacteriology

@ASMicrobiology

       

ASM Journals

ASM journals are the most prominent publications in the field, delivering up-to-date and authoritative coverage of both basic and clinical microbiology.

About ASM | Contact Us | Press Room

 

ASM is a member of

Scientific Society Publisher Alliance

Copyright © 2019 American Society for Microbiology | Privacy Policy | Website feedback

Print ISSN: 0021-9193; Online ISSN: 1098-5530