Skip to main content

Thank you for visiting nature.com. You are using a browser version with limited support for CSS. To obtain the best experience, we recommend you use a more up to date browser (or turn off compatibility mode in Internet Explorer). In the meantime, to ensure continued support, we are displaying the site without styles and JavaScript.

  • On the Market
  • Published:

A quantitative assay for HIV DNA integration in vivo

Nature Medicine volume 7, pages 631–634 (2001)Cite this article

Abstract

Early steps of infection by HIV-1 involve entry of the viral core into cells, reverse transcription to form the linear viral DNA, and integration of that DNA into a chromosome of the host. The unintegrated DNA can also follow non-productive pathways, in which it is circularized by recombination between DNA long-terminal repeats (LTRs), circularized by ligation of the DNA ends or degraded. Here we report quantitative methods that monitor formation of reverse transcription products, two-LTR circles and integrated proviruses. The integration assay employs a novel quantitative form of Alu-PCR that should be generally applicable to studies of integrating viruses and gene transfer vectors.

This is a preview of subscription content, access via your institution

Relevant articles

Open Access articles citing this article.

Access options

Buy this article

  • Purchase on Springer Link
  • Instant access to full article PDF
Buy now

Prices may be subject to local taxes which are calculated during checkout

Figure 1: Assaying early steps in HIV replication.
Figure 2: DNA metabolism after infection of 293T cells with HIV-based vectors.
Figure 3: Assay of the effects of inhibitors on HIV cDNA metabolism.

References

  1. Heid, C.A., Stevens, J., Livak, K.J. & Williams, P.M. Real time quantitative PCR. Genome Res. 6, 986–994 (1996).

    Article  CAS  Google Scholar 

  2. Lewin, S.R. et al. Use of real-time PCR and molecular beacons to detect virus replication in Human Immunodeficiency Virus type 1-infected individuals on prolonged effective antiretroviral therapy. J. Virol. 73, 6099–6103 (1999).

    CAS  PubMed  PubMed Central  Google Scholar 

  3. Sharkey, M.E. et al. Persistence of episomal HIV-1 infection intermediates in patients on highly active anti-retroviral therapy. Nature Med. 6, 76–81 (2000).

    Article  CAS  Google Scholar 

  4. Benkirane, M., Corbeau, P., Housset, V. & Devaux, C. An antibody that binds the imunoglobulin CDR3-like region of the CD4 molecule inhibits provirus transcription in HIV-infected T cells. EMBO J. 12, 4909–4921 (1993).

    Article  CAS  Google Scholar 

  5. Courcoul, M. et al. Peripheral blood mononuclear cells produce normal amounts of defective Vif- human immunodeficiency virus type 1 particles which are restricted for the preretrotranscription steps. J. Virol. 69, 2068–2074 (1995).

    CAS  PubMed  PubMed Central  Google Scholar 

  6. Sonza, S. et al. Human immunodeficiency virus type 1 replication is blocked prior to revere transcription and integration in freshly isolated peripheral blood monocytes. J. Virol. 70, 3863–3869 (1996).

    CAS  PubMed  PubMed Central  Google Scholar 

  7. Chun, T.-W. et al. Presence of an inducible HIV-1 latent reservoir during highly active antiretroviral therapy. Proc. Natl. Acad. Sci. USA 94, 13193–13197 (1997).

    Article  CAS  Google Scholar 

  8. Carteau, S., Hoffmann, C. & Bushman, F.D. Chromosome structure and HIV-1 cDNA integration: Centromeric alphoid repeats are a disfavored target. J. Virol. 72, 4005–4014 (1998).

    CAS  PubMed  PubMed Central  Google Scholar 

  9. Furtado, M.R. et al. Persistence of HIV-1 transcription in peripheral blood mononuclear cells in patients receiving potent antiretroviral therapy. New Engl. J. Med. 340, 1614–1622 (1999).

    Article  CAS  Google Scholar 

  10. Naldini, L. et al. In vivo gene delivery and stable transduction of nondividing cells by a lentiviral vector. Science 272, 263–267 (1996).

    Article  CAS  Google Scholar 

  11. Kafri, T., van Praag, H., Ouyang, L., Gage, F.H. & Verma, I.M. A packaging cell line for lentiviral vectors. J. Virol. 73, 576–584 (1999).

    CAS  PubMed  PubMed Central  Google Scholar 

  12. Zennou, V. et al. HIV-1 genome nuclear import is mediated by a central DNA flap. Cell 101, 173–185 (2000).

    Article  CAS  Google Scholar 

  13. Follenzi, A., Ailes, L.E., Bakovic, S., Gueuna, M. & Naldini, L. Gene transfer by lentiviral vectors is limited by nuclear translocation and rescued by HIV-1 pol sequences. Nature Genet. 25, 217–222 (2000).

    Article  CAS  Google Scholar 

  14. Hazuda, D.J. et al. Inhibitors of strand transfer that prevent integration and inhibit HIV-1 replication in cells. Science 287, 646–650 (2000).

    Article  CAS  Google Scholar 

Download references

Acknowledgements

This work was supported by NIH grants GM56553 and AI34786 to F.D.B., the James B. Pendleton Charitable Trust, the Berger Foundation, and Cornelia Mackey.

Author information

Authors and Affiliations

  1. Infectious Disease Laboratory, The Salk Institute for Biological Studies, La Jolla, California, USA

    Scott L. Butler, Mark S.T. Hansen & Frederic D. Bushman

Authors
  1. Scott L. Butler
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Mark S.T. Hansen
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Frederic D. Bushman
    View author publications

    You can also search for this author in PubMed Google Scholar

Corresponding author

Correspondence to Frederic D. Bushman.

Rights and permissions

Reprints and permissions

About this article

Cite this article

Butler, S., Hansen, M. & Bushman, F. A quantitative assay for HIV DNA integration in vivo. Nat Med 7, 631–634 (2001). https://doi.org/10.1038/87979

Download citation

  • Issue Date:

  • DOI: https://doi.org/10.1038/87979

This article is cited by

Search

Advanced search

Quick links

Nature Briefing

Sign up for the Nature Briefing newsletter — what matters in science, free to your inbox daily.

Get the most important science stories of the day, free in your inbox. Sign up for Nature Briefing