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
    • Latest Articles
    • COVID-19 Special Collection
    • Archive
    • Minireviews
  • Topics
    • Applied and Environmental Science
    • Clinical Science and Epidemiology
    • Ecological and Evolutionary Science
    • Host-Microbe Biology
    • Molecular Biology and Physiology
    • Therapeutics and Prevention
  • 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 mBio
    • Editor in Chief
    • Board of Editors
    • AAM Fellows
    • For Reviewers
    • For the Media
    • For Librarians
    • For Advertisers
    • Alerts
    • RSS
    • FAQ
  • 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
mBio
publisher-logosite-logo

Advanced Search

  • Home
  • Articles
    • Latest Articles
    • COVID-19 Special Collection
    • Archive
    • Minireviews
  • Topics
    • Applied and Environmental Science
    • Clinical Science and Epidemiology
    • Ecological and Evolutionary Science
    • Host-Microbe Biology
    • Molecular Biology and Physiology
    • Therapeutics and Prevention
  • 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 mBio
    • Editor in Chief
    • Board of Editors
    • AAM Fellows
    • For Reviewers
    • For the Media
    • For Librarians
    • For Advertisers
    • Alerts
    • RSS
    • FAQ
Research Article

Molecular Dissection of the Essential Features of the Origin of Replication of the Second Vibrio cholerae Chromosome

Matthew A. Gerding, Michael C. Chao, Brigid M. Davis, Matthew K. Waldor
Eric J. Rubin, Editor
Matthew A. Gerding
aProgram in Biological and Biomedical Sciences, Graduate School of Arts and Sciences, Harvard Medical School, Boston, Massachusetts, USA
bDivision of Infectious Diseases, Brigham and Women's Hospital, Boston, Massachusetts, USA
  • Find this author on Google Scholar
  • Find this author on PubMed
  • Search for this author on this site
Michael C. Chao
bDivision of Infectious Diseases, Brigham and Women's Hospital, Boston, Massachusetts, USA
  • Find this author on Google Scholar
  • Find this author on PubMed
  • Search for this author on this site
Brigid M. Davis
bDivision of Infectious Diseases, Brigham and Women's Hospital, Boston, Massachusetts, USA
  • Find this author on Google Scholar
  • Find this author on PubMed
  • Search for this author on this site
Matthew K. Waldor
bDivision of Infectious Diseases, Brigham and Women's Hospital, Boston, Massachusetts, USA
cHoward Hughes Medical Institute, Boston, Massachusetts, USA
  • Find this author on Google Scholar
  • Find this author on PubMed
  • Search for this author on this site
Eric J. Rubin
Harvard School of Public Health
Roles: Editor
  • Find this author on Google Scholar
  • Find this author on PubMed
  • Search for this author on this site
DOI: 10.1128/mBio.00973-15
  • Article
  • Figures & Data
  • Info & Metrics
  • PDF
Loading

ABSTRACT

Vibrionaceae family members are interesting models for studying DNA replication initiation, as they contain two circular chromosomes. Chromosome II (chrII) replication is governed by two evolutionarily unique yet highly conserved elements, the origin DNA sequence oriCII and the initiator protein RctB. The minimum functional region of oriCII, oriCII-min, contains multiple elements that are bound by RctB in vitro, but little is known about the specific requirements for individual elements during oriCII initiation. We utilized undirected and site-specific mutagenesis to investigate the functionality of mutant forms of oriCII-min and assessed binding to various mutant forms by RctB. Our analyses showed that deletions, point mutations, and changes in RctB target site spacing or methylation all impaired oriCII-min-based replication. RctB displayed a reduced affinity for most of the low-efficacy origins tested, although its characteristic cooperative binding was generally maintained. Mutations that removed or altered the relative positions of origin components other than RctB binding sites (e.g., AT-rich sequence, DnaA target site) also abolished replicative capacity. Comprehensive mutagenesis and deep-sequencing-based screening (OriSeq) allowed the identification of a previously uncharacterized methylated domain in oriCII that is required for origin function. Together, our results reveal the remarkable evolutionary honing of oriCII and provide new insight into the complex interplay between RctB and oriCII.

IMPORTANCE The genome of the enteric pathogen Vibrio cholerae consists of two chromosomes. While the chromosome I replication origin and its cognate replication initiator protein resemble those of Escherichia coli, the factors responsible for chromosome II replication initiation display no similarity to any other known initiation systems. Here, to enhance our understanding of how this DNA sequence, oriCII, and its initiator protein, RctB, function, we used both targeted mutagenesis and a new random-mutagenesis approach (OriSeq) to finely map the oriCII structural features and sequences required for RctB-mediated DNA replication. Collectively, our findings reveal the extraordinary evolutionary honing of the architecture and motifs that constitute oriCII and reveal a new role for methylation in oriCII-based replication. Finally, our findings suggest that the OriSeq approach is likely to be widely applicable for defining critical bases in cis-acting sequences.

FOOTNOTES

    • Received 11 June 2015
    • Accepted 25 June 2015
    • Published 28 July 2015
  • Copyright © 2015 Gerding et al.

This is an open-access article distributed under the terms of the Creative Commons Attribution-Noncommercial-ShareAlike 3.0 Unported license, which permits unrestricted noncommercial use, distribution, and reproduction in any medium, provided the original author and source are credited.

View Full Text
PreviousNext
Back to top
Download PDF
Citation Tools
Molecular Dissection of the Essential Features of the Origin of Replication of the Second Vibrio cholerae Chromosome
Matthew A. Gerding, Michael C. Chao, Brigid M. Davis, Matthew K. Waldor
mBio Jul 2015, 6 (4) e00973-15; DOI: 10.1128/mBio.00973-15

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 mBio 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.
Molecular Dissection of the Essential Features of the Origin of Replication of the Second Vibrio cholerae Chromosome
(Your Name) has forwarded a page to you from mBio
(Your Name) thought you would be interested in this article in mBio.
CAPTCHA
This question is for testing whether or not you are a human visitor and to prevent automated spam submissions.
Share
Molecular Dissection of the Essential Features of the Origin of Replication of the Second Vibrio cholerae Chromosome
Matthew A. Gerding, Michael C. Chao, Brigid M. Davis, Matthew K. Waldor
mBio Jul 2015, 6 (4) e00973-15; DOI: 10.1128/mBio.00973-15
del.icio.us logo Digg logo Reddit logo Twitter logo CiteULike logo Facebook logo Google logo Mendeley logo
  • Top
  • Article
    • ABSTRACT
    • INTRODUCTION
    • RESULTS
    • DISCUSSION
    • MATERIALS AND METHODS
    • FOOTNOTES
    • REFERENCES
  • Figures & Data
  • Info & Metrics
  • PDF

Related Articles

Cited By...

About

  • About mBio
  • Editor in Chief
  • Board of Editors
  • AAM Fellows
  • Policies
  • For Reviewers
  • For the Media
  • For Librarians
  • For Advertisers
  • Alerts
  • RSS
  • FAQ
  • Permissions
  • Journal Announcements

Authors

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

Follow #mBio

@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

 

American Society for Microbiology
1752 N St. NW
Washington, DC 20036
Phone: (202) 737-3600

Copyright © 2021 American Society for Microbiology | Privacy Policy | Website feedback

Online ISSN: 2150-7511