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
Virus-Cell Interactions

Antiviral Activity of the Proteasome on Incoming Human Immunodeficiency Virus Type 1

Olivier Schwartz, Valérie Maréchal, Bertrand Friguet, Fernando Arenzana-Seisdedos, Jean-Michel Heard
Olivier Schwartz
Laboratoire Rétrovirus et Transfert Génétique, URA CNRS 1157,
  • Find this author on Google Scholar
  • Find this author on PubMed
  • Search for this author on this site
Valérie Maréchal
Laboratoire Rétrovirus et Transfert Génétique, URA CNRS 1157,
  • Find this author on Google Scholar
  • Find this author on PubMed
  • Search for this author on this site
Bertrand Friguet
Unité de Biochimie Cellulaire, and
  • Find this author on Google Scholar
  • Find this author on PubMed
  • Search for this author on this site
Fernando Arenzana-Seisdedos
Unitéd’Immunologie Virale, Institut Pasteur, 75724 Paris Cedex 15, France
  • Find this author on Google Scholar
  • Find this author on PubMed
  • Search for this author on this site
Jean-Michel Heard
Laboratoire Rétrovirus et Transfert Génétique, URA CNRS 1157,
  • Find this author on Google Scholar
  • Find this author on PubMed
  • Search for this author on this site
DOI: 10.1128/JVI.72.5.3845-3850.1998
  • Article
  • Figures & Data
  • Info & Metrics
  • PDF
Loading

Article Figures & Data

Figures

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

    The proteasome inhibitor MG132 increases HIV infection. (A) P4 indicator cells (HeLa CD4+LTR-lacZ +) were incubated with HIV-1 (left panels) or with HIVΔenv(VSV), a defective HIV-1 strain with a deletion in the env gene and pseudotyped with the VSV-G envelope, in the absence and in the presence of MG132 (25 μM). Five hours later, virus and MG132 were removed. After 24 h, HIV-infected cells were revealed by in situ staining for β-galactosidase (β-gal) activity. (B) The effect of MG132 is independent of the viral input. HIV infection was quantified in P4 cells in a β-galactosidase based colorimetric assay. P4 cells were infected with the indicated amounts of HIV-1 (left panel) or HIVΔenv(VSV) (right panel) with or without MG132 (50 μM) for 1 h. After 24 h, cells were lysed, and β-galactosidase activity (optical density [O.D.] units) in cell extracts was measured. Values are means of triplicates. Variation between each point of triplicate determinations was below 10%. Data are representative of three independent experiments.

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

    Analysis of HIV-1 infection in the presence of MG132 on various cell lines. HeLa-CD4 cells and the Jurkat, CEM, and HUT78 T-lymphoid cell lines were infected with the HIV-HSA reporter virus pseudotyped with the VSV-G envelope, which contains, in the place ofnef, the gene coding for HSA (CD24). Following integration and proviral expression, cells synthesize CD24, which can be detected at the cell surface. Cell lines were infected with HIV-HSA(VSV) (50 ng of p24Gag) for 1 h, with or without MG132 (50 μM). Infection was revealed 24 h later by staining with anti-CD24 monoclonal antibodies and flow cytometry analysis of gated living cells. Data are representative of three independent experiments.

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

    Analysis of the effect of various protease inhibitors on HIV infection and on LTR activity. (A) Effect of MG132 on HIV-1 LTR activity. P4 cells were transfected with 1 μg of pLTRX-Luc and the indicated amounts of pCMV-Tat DNA. After 24 h, cells were pulse-incubated for 1 h with or without MG132 (50 μM), and 5 h later, luciferase activities in cell extracts were measured. Results are expressed as RLU per microgram of cellular protein. Values are means of triplicate determinations, and variation between each point was below 10%. (B) P4 cells were infected with HIV-1 (left panel) or HIVΔenv(VSV) (right panel) in the presence of the proteasome inhibitors MG132 (50 μM) or lactacystin (40 μM) or with calpain inhibitor I (50 μM) or calpain inhibitor II (50 μM) for 1 h and washed. Infections were revealed 24 h later by measuring β-galactosidase activity (optical density [O.D.] units). Data are representative of three independent experiments.

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

    MG132 induces an accumulation of proviral DNA and of cytosolic p24Gag proteins in target cells. (A) Synthesis of proviral DNA in newly infected cells. P4 cells were infected by a 5-h incubation period with HIV at the indicated amounts of p24Gag in the absence or in the presence of MG132 (25 μM). Seventeen hours later, low-molecular-weight DNA was extracted and analyzed by Southern blotting with an HIV probe (upper panel). Digestion with EcoRI produced diagnostic fragments with sizes of 5.7 and 9.1 kb from linear DNA (L) and DNA containing one LTR circle (C), respectively. Since variations in the yield of cellular low-molecular-weight DNA were observed, samples were normalized by hybridization with the mitochondrial gene coding for cytochromeb (cyt. b) (lower panel). (B) Accumulation of cytosolic p24Gag proteins. P4 cells were incubated with HIV-1 (1 μg of p24) in the absence (untreated columns) and in the presence of MG132 (50 μM) or lactacystin (40 μM) for 1 h at 37°C. Cells were then washed to remove unbound virus and further incubated at 37°C in medium containing the indicated inhibitors. At each time point, cells were treated with Pronase to eliminate virus adsorbed at the cell surface and lysed. Postnuclear supernatants were separated in cytosol and pellet fractions. The pellet fraction corresponds to cellular membranes and vesicles. p24Gag contents (in picograms) were measured in the cytosolic fraction. Time zero corresponds to the beginning of infection. Data are representative of three independent experiments.

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

    In vitro degradation of HIV-1 Gag proteins by purified 20S proteasome. HIV-1 virions were pelleted by ultracentrifugation and resuspended in a buffer containing 0.4% Nonidet P-40 to remove the viral membrane. HIV-1 virions (10 ng of p24Gag) were then incubated with purified 20S proteasome (0.4 μg) for 30 min at 37°C, with or without MG132 (20 μM) or the serine protease inhibitor phenylmethylsulfonyl fluoride (PMSF) (50 μM). Viral proteins were then revealed in a Western blot assay with an anti-HIV-1 serum. Molecular mass markers are indicated (in kilodaltons) on the right. Data are representative of three independent experiments.

PreviousNext
Back to top
Download PDF
Citation Tools
Antiviral Activity of the Proteasome on Incoming Human Immunodeficiency Virus Type 1
Olivier Schwartz, Valérie Maréchal, Bertrand Friguet, Fernando Arenzana-Seisdedos, Jean-Michel Heard
Journal of Virology May 1998, 72 (5) 3845-3850; DOI: 10.1128/JVI.72.5.3845-3850.1998

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.
Antiviral Activity of the Proteasome on Incoming Human Immunodeficiency Virus Type 1
(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
Antiviral Activity of the Proteasome on Incoming Human Immunodeficiency Virus Type 1
Olivier Schwartz, Valérie Maréchal, Bertrand Friguet, Fernando Arenzana-Seisdedos, Jean-Michel Heard
Journal of Virology May 1998, 72 (5) 3845-3850; DOI: 10.1128/JVI.72.5.3845-3850.1998
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 AND DISCUSSION
    • ACKNOWLEDGMENTS
    • FOOTNOTES
    • REFERENCES
  • Figures & Data
  • Info & Metrics
  • PDF

KEYWORDS

Cysteine Endopeptidases
HIV-1
Multienzyme Complexes

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