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A primary CD4+ T cell model of HIV-1 latency established after activation through the T cell receptor and subsequent return to quiescence

Nature Protocols volume 9, pages 2755–2770 (2014)Cite this article

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Abstract

A mechanistic understanding of HIV-1 latency depends on a model system that recapitulates the in vivo condition of latently infected, resting CD4+ T lymphocytes. Latency seems to be established after activated CD4+ T cells, the principal targets of HIV-1 infection, become productively infected and survive long enough to return to a resting memory state in which viral expression is inhibited by changes in the cellular environment. This protocol describes an ex vivo primary cell system that is generated under conditions that reflect the in vivo establishment of latency. Creation of these latency model cells takes 12 weeks and, once established, the cells can be maintained and used for several months. The resulting cell population contains both uninfected and latently infected cells. This primary cell model can be used to perform drug screens, to study cytolytic T lymphocyte (CTL) responses to HIV-1, to compare viral alleles or to expand the ex vivo life span of cells from HIV-1-infected individuals for extended study.

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Figure 1: Representative results of intracellular Bcl-2 expression levels.
Figure 2: Structure of the CM6 HIV-1/GFP reporter virus.
Figure 3: Part I.
Figure 4: Part II.
Figure 5: Part III.
Figure 6: TCGF preparation, as described in Box 1.
Figure 7: Representative FACS measurement of HIV-1/GFP reporter virus infection efficiency.
Figure 8: Representative FACS measurement of frequency of latent infection in Bcl-2 model cells.

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Acknowledgements

We thank all members of the Siliciano laboratory, past and present, who have contributed to the development and testing of the Bcl-2 model cells. This work was supported by the Martin Delaney Collaboratory of AIDS researchers for Eradication (CARE) and Delaney AIDS Research Enterprise (DARE) Collaboratories (US National Institutes of Health (NIH) grants AI096113 and 1U19AI096109), by an amfAR Research Consortium on HIV Eradication (ARCHE) Collaborative Research Grant from the Foundation for AIDS Research (amfAR 108165-50-RGRL), by the Johns Hopkins Center for AIDS Research, by NIH grant 43222 and by the Howard Hughes Medical Institute.

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Authors and Affiliations

  1. Department of Medicine, Johns Hopkins University School of Medicine, Baltimore, Maryland, USA

    Michelle Kim, Nina N Hosmane, C Korin Bullen, Adam Capoferri, Janet D Siliciano & Robert F Siliciano

  2. Howard Hughes Medical Institute, Baltimore, Maryland, USA

    Michelle Kim, Adam Capoferri & Robert F Siliciano

  3. Department of Microbiology, National Taiwan University College of Medicine, Taipei, Taiwan

    Hung-Chih Yang

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  1. Michelle Kim
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Contributions

H.-C.Y. and R.F.S. conceived and developed the original Bcl-2 model cell system; M.K. modified the protocol and wrote the manuscript; M.K., N.N.H. and C.K.B. performed experiments; and all authors reviewed the manuscript.

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Correspondence to Robert F Siliciano.

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Kim, M., Hosmane, N., Bullen, C. et al. A primary CD4+ T cell model of HIV-1 latency established after activation through the T cell receptor and subsequent return to quiescence. Nat Protoc 9, 2755–2770 (2014). https://doi.org/10.1038/nprot.2014.188

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