Skip to main content
  • ASM
    • Antimicrobial Agents and Chemotherapy
    • Applied and Environmental Microbiology
    • Clinical Microbiology Reviews
    • Clinical and Vaccine Immunology
    • EcoSal Plus
    • 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
    • COVID-19 Special Collection
    • Minireviews
    • JVI Classic Spotlights
    • Archive
  • 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 JVI
    • 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
    • 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 Virology
publisher-logosite-logo

Advanced Search

  • Home
  • Articles
    • Current Issue
    • Accepted Manuscripts
    • COVID-19 Special Collection
    • Minireviews
    • JVI Classic Spotlights
    • Archive
  • 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 JVI
    • Editor in Chief
    • Editorial Board
    • For Reviewers
    • For the Media
    • For Librarians
    • For Advertisers
    • Alerts
    • RSS
    • FAQ
  • Subscribe
    • Members
    • Institutions
Cellular Response to Infection

Autophagosome Supports Coxsackievirus B3 Replication in Host Cells

Jerry Wong, Jingchun Zhang, Xiaoning Si, Guang Gao, Ivy Mao, Bruce M. McManus, Honglin Luo
Jerry Wong
The James Hogg iCAPTURE Centre for Cardiovascular and Pulmonary Research, Department of Pathology and Laboratory Medicine, University of British Columbia, Providence Heart + Lung Institute, St. Paul's Hospital, Vancouver, British Columbia, Canada
  • Find this author on Google Scholar
  • Find this author on PubMed
  • Search for this author on this site
Jingchun Zhang
The James Hogg iCAPTURE Centre for Cardiovascular and Pulmonary Research, Department of Pathology and Laboratory Medicine, University of British Columbia, Providence Heart + Lung Institute, St. Paul's Hospital, Vancouver, British Columbia, Canada
  • Find this author on Google Scholar
  • Find this author on PubMed
  • Search for this author on this site
Xiaoning Si
The James Hogg iCAPTURE Centre for Cardiovascular and Pulmonary Research, Department of Pathology and Laboratory Medicine, University of British Columbia, Providence Heart + Lung Institute, St. Paul's Hospital, Vancouver, British Columbia, Canada
  • Find this author on Google Scholar
  • Find this author on PubMed
  • Search for this author on this site
Guang Gao
The James Hogg iCAPTURE Centre for Cardiovascular and Pulmonary Research, Department of Pathology and Laboratory Medicine, University of British Columbia, Providence Heart + Lung Institute, St. Paul's Hospital, Vancouver, British Columbia, Canada
  • Find this author on Google Scholar
  • Find this author on PubMed
  • Search for this author on this site
Ivy Mao
The James Hogg iCAPTURE Centre for Cardiovascular and Pulmonary Research, Department of Pathology and Laboratory Medicine, University of British Columbia, Providence Heart + Lung Institute, St. Paul's Hospital, Vancouver, British Columbia, Canada
  • Find this author on Google Scholar
  • Find this author on PubMed
  • Search for this author on this site
Bruce M. McManus
The James Hogg iCAPTURE Centre for Cardiovascular and Pulmonary Research, Department of Pathology and Laboratory Medicine, University of British Columbia, Providence Heart + Lung Institute, St. Paul's Hospital, Vancouver, British Columbia, Canada
  • Find this author on Google Scholar
  • Find this author on PubMed
  • Search for this author on this site
Honglin Luo
The James Hogg iCAPTURE Centre for Cardiovascular and Pulmonary Research, Department of Pathology and Laboratory Medicine, University of British Columbia, Providence Heart + Lung Institute, St. Paul's Hospital, Vancouver, British Columbia, Canada
  • Find this author on Google Scholar
  • Find this author on PubMed
  • Search for this author on this site
  • For correspondence: hluo@mrl.ubc.ca
DOI: 10.1128/JVI.00641-08
  • Article
  • Figures & Data
  • Info & Metrics
  • PDF
Loading

Article Figures & Data

Figures

    • Open in new tab
    • Download powerpoint
    • Open in new tab
    • Download powerpoint
    FIG. 1.

    CVB3 infection increases autophagosome formation. (A) Representative electron micrographs of sham- and CVB3-infected HeLa (top) and HEK293A (bottom) cells at 7 h postinfection. Arrows indicate representative autophagosomes that would be scored positive in panel B. (B) Quantitation of the numbers of autophagosomes in sham- and CVB3-infected HEK293A cells. Data shown represent the number of autophagosomes per cell under each condition (mean ± standard error [SE]). #, P < 0.001 compared to sham infection, by Student's t test. (C) The LC3-II/LC3-I ratio, which is a hallmark of autophagy, increases with the progression of CVB3 infection. (Left) Western blot analysis of LC3 and viral capsid protein VP1 expression in CVB3-infected cells at various times postinfection. β-Actin expression was examined as a protein loading control. (Right) Expression of LC3-II and LC3-I and ratio of LC3-II to LC3-I at the indicated times after CVB3 infection (means ± SE; n = 5 or 7 [as indicated]). LC3 expression was quantitated by densitometric analysis using NIH ImageJ v1.37 and normalized to sham infection, which was arbitrarily set to a value of 1.0. &, P < 0.01 compared to sham infection. (D) Representative confocal images and quantitation of autophagosomes in GFP-LC3-transfected HEK293A cells after CVB3 infection. HEK293A cells were transfected with GFP-LC3 plasmid for 24 h, followed by CVB3 infection. The GFP fluorescence was analyzed by confocal fluorescence microscopy. The percentage of cells with punctate GFP-LC3 localization relative to all GFP-positive cells was calculated and is presented as the mean ± SE (n = 5). #, P < 0.001 compared to sham infection.

  • FIG. 2.
    • Open in new tab
    • Download powerpoint
    FIG. 2.

    CVB3 infection induces eIF2α phosphorylation. Western blot analysis of phosphorylated eIF2α and β-actin (loading control) expression was performed at the indicated times postinfection. Similar results were obtained in two independent experiments.

  • FIG. 3.
    • Open in new tab
    • Download powerpoint
    FIG. 3.

    Autophagy inhibitor 3-MA reduces CVB3 replication. (A) (Top) Western blot analysis of viral protein VP1 and β-actin (loading control) expression 7 h following CVB3 infection, with or without 3-MA treatment, as indicated. (Bottom) VP1 expression was quantitated by densitometric analysis using NIH ImageJ v1.37 and normalized to β-actin expression. The VP1/β-actin ratio was further normalized to the CVB3-infected group without treatment, which was arbitrarily set to a value of 1.0. The data shown are means ± SE (n = 3), and significance was determined by Student's t test. *, P < 0.05; #, P < 0.001 (compared to nontreated cells). (B) Plaque assay measuring the CVB3 viral progeny titers in supernatants collected from infected cells treated with or without 3-MA. The data shown are means ± SE for three independent experiments. *, P < 0.05 compared to nontreated cells. (C) Western blot analysis of CVB3-infected cells treated with 10 mM 3-MA at different times, as indicated.

  • FIG. 4.
    • Open in new tab
    • Download powerpoint
    FIG. 4.

    Induction of autophagy enhances viral replication. Western blot analysis of VP1 and β-actin expression was performed at 7 h postinfection. (A) Prior to CVB3 infection, cells were incubated in Hank's balanced salt solution for 2 h to induce cell starvation. (B) Cells were pretreated with rapamycin (50 nM) for 3 h. VP1 expression was quantitated, normalized, and analyzed as described in the legend to Fig. 3 (data are means ± SE; n = 3). *, P < 0.05; #, P < 0.001 (compared to nontreated cells).

  • FIG. 5.
    • Open in new tab
    • Download powerpoint
    FIG. 5.

    Gene silencing of ATG7 by siRNA inhibits CVB3 replication. (A) Representative light microscopic images of CVB3-infected cells, with or without ATG7 siRNA transfection. (B) Western blot probing for VP1 and ATG7 expression, with or without ATG7 knockdown. VP1 expression was quantitated, normalized, and analyzed as described in the legend to Fig. 3 (data are means ± SE; n = 3). *, P < 0.05 compared to scramble siRNA-transfected cells.

  • FIG. 6.
    • Open in new tab
    • Download powerpoint
    FIG. 6.

    Knockdown of class III PI3K components reduces CVB3 replication. (A) Western blot analysis of VP1 and Beclin-1 or VPS34 expression in CVB3-infected cells transfected with Beclin-1 (left) or VPS34 (right) siRNA prior to CVB3 infection. VP1 expression was quantitated, normalized, and analyzed as described in the legend to Fig. 3 (data are means ± SE; n = 3). *, P < 0.05 compared to scramble siRNA control. (B) Plaque assay results for scramble or Beclin-1 and VPS34 siRNA-transfected cells (means ± SE; n = 3). *, P < 0.05; #, P < 0.001 (compared to scramble siRNA-transfected cells).

  • FIG. 7.
    • Open in new tab
    • Download powerpoint
    FIG. 7.

    Expression of p62 is not affected by CVB3 infection. (A) Western blot analysis of p62, a marker of autophagic protein degradation activity, in cells treated with the lysosome inhibitors bafilomycin A (BFLA; 0.1 μM) and NH4Cl (2.5 mM) for 16 h. (B) Expression of p62 in CVB3-infected cells at the indicated times after virus infection. Similar results were obtained in two independent experiments.

  • FIG. 8.
    • Open in new tab
    • Download powerpoint
    FIG. 8.

    Lysosomal inhibition enhances CVB3 replication. (A) Western blot analysis of VP1 and LAMP2 expression in CVB3-infected cells transiently transfected with scramble or LAMP2 siRNA. VP1 expression was quantitated, normalized, and analyzed as described in the legend to Fig. 3 (data are means ± SE; n = 3 or 4 [as indicated]). (B) Virus titer in scramble or LAMP2 siRNA-transfected cells, determined by plaque assay (mean ± SE; n = 3). *, P < 0.05 compared to scramble siRNA control.

PreviousNext
Back to top
Download PDF
Citation Tools
Autophagosome Supports Coxsackievirus B3 Replication in Host Cells
Jerry Wong, Jingchun Zhang, Xiaoning Si, Guang Gao, Ivy Mao, Bruce M. McManus, Honglin Luo
Journal of Virology Aug 2008, 82 (18) 9143-9153; DOI: 10.1128/JVI.00641-08

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 Virology 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.
Autophagosome Supports Coxsackievirus B3 Replication in Host Cells
(Your Name) has forwarded a page to you from Journal of Virology
(Your Name) thought you would be interested in this article in Journal of Virology.
CAPTCHA
This question is for testing whether or not you are a human visitor and to prevent automated spam submissions.
Share
Autophagosome Supports Coxsackievirus B3 Replication in Host Cells
Jerry Wong, Jingchun Zhang, Xiaoning Si, Guang Gao, Ivy Mao, Bruce M. McManus, Honglin Luo
Journal of Virology Aug 2008, 82 (18) 9143-9153; DOI: 10.1128/JVI.00641-08
del.icio.us logo Digg logo Reddit logo Twitter logo CiteULike logo Facebook logo Google logo Mendeley logo
  • Top
  • Article
    • ABSTRACT
    • MATERIALS AND METHODS
    • RESULTS
    • DISCUSSION
    • ACKNOWLEDGMENTS
    • FOOTNOTES
    • REFERENCES
  • Figures & Data
  • Info & Metrics
  • PDF

KEYWORDS

autophagy
Enterovirus B, Human
Phagosomes
virus replication

Related Articles

Cited By...

About

  • About JVI
  • 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 #Jvirology

@ASMicrobiology

       

 

JVI in collaboration with

American Society for Virology

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

Print ISSN: 0022-538X; Online ISSN: 1098-5514