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Inducible human immunodeficiency virus type 1 packaging cell lines

H Yu1, A B Rabson, M Kaul

  • 1Department of Molecular Genetics and Microbiology, Robert Wood Johnson Medical School, University of Medicine and Dentistry of New Jersey, Piscataway, New Jersey 08854-5635, USA.

Journal of Virology
|July 1, 1996
PubMed
Summary

Researchers developed an inducible human immunodeficiency virus type 1 (HIV-1)-based packaging cell line to overcome protein cytotoxicity. This novel system allows for controlled expression of HIV-1 proteins, enabling gene transfer into CD4+ cells and aiding HIV-1 biology studies.

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Area of Science:

  • Molecular Biology
  • Virology
  • Gene Therapy

Background:

  • Packaging cell lines are crucial for gene transfer into eukaryotic cells.
  • Human immunodeficiency virus type 1 (HIV-1)-based systems are challenging due to cytotoxic viral proteins.
  • Existing methods struggle with the safe and efficient production of HIV-1 vectors.

Purpose of the Study:

  • To develop a novel HIV-1-based packaging cell line with an inducible expression system.
  • To overcome the cytotoxicity of HIV-1 proteins for improved vector production.
  • To facilitate gene transfer into CD4+ cells and study HIV-1 biology.

Main Methods:

  • Utilized a tetracycline-inducible expression system to control Rev protein expression.
  • Controlled expression of late HIV-1 proteins (Gag, Pol, Env) via the Rev regulatory protein.

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  • Analyzed protein expression (p24gag, gp120env) and reverse transcriptase activity using Western blotting and activity assays.

    • Main Results:

    • Inducible expression of p24gag and gp120env peaked 6-7 days post-induction.
    • Reverse transcriptase activity was detected by day 4 and peaked on days 6-7.
    • High titers of defective vector virus (7 x 10^3 CFU/ml) were produced with no detectable replication-competent virus.
    • An inducible HIV-1 Env-expressing cell line produced env-minus vector virus titers of 2 x 10^4 CFU/ml.

    Conclusions:

    • The developed inducible HIV-1 packaging cell line effectively overcomes protein cytotoxicity.
    • This system enables controlled gene transfer into CD4+ cells and aids HIV-1 research.
    • The inducible Env-expressing cell line is valuable for propagating HIV-1 vectors requiring Env trans-complementation.