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.

HIV-1 causes CD4 cell death through DNA-dependent protein kinase during viral integration

Subjects

Abstract

Human immunodeficiency virus-1 (HIV-1) has infected more than 60 million people and caused nearly 30 million deaths worldwide1, ultimately the consequence of cytolytic infection of CD4+ T cells. In humans and in macaque models, most of these cells contain viral DNA and are rapidly eliminated at the peak of viraemia2,3,4, yet the mechanism by which HIV-1 induces helper T-cell death has not been defined. Here we show that virus-induced cell killing is triggered by viral integration. Infection by wild-type HIV-1, but not an integrase-deficient mutant, induced the death of activated primary CD4 lymphocytes. Similarly, raltegravir, a pharmacologic integrase inhibitor, abolished HIV-1-induced cell killing both in cell culture and in CD4+ T cells from acutely infected subjects. The mechanism of killing during viral integration involved the activation of DNA-dependent protein kinase (DNA-PK), a central integrator of the DNA damage response, which caused phosphorylation of p53 and histone H2AX. Pharmacological inhibition of DNA-PK abolished cell death during HIV-1 infection in vitro, suggesting that processes which reduce DNA-PK activation in CD4 cells could facilitate the formation of latently infected cells that give rise to reservoirs in vivo. We propose that activation of DNA-PK during viral integration has a central role in CD4+ T-cell depletion, raising the possibility that integrase inhibitors and interventions directed towards DNA-PK may improve T-cell survival and immune function in infected individuals.

This is a preview of subscription content

Access options

Rent or Buy article

Get time limited or full article access on ReadCube.

from$8.99

All prices are NET prices.

Figure 1: CD4 lymphocytes killed during HIV-1 infection do not express viral gene products.
Figure 2: Dying CD4 lymphocytes lacking viral gene expression had been productively infected prior to cell death.
Figure 3: Proviral DNA integration triggers cell death during HIV-1 infection.
Figure 4: DNA-PK orchestrates a DNA damage response and cell death following proviral DNA integration.

References

  1. 1

    Joint. United Nations Programme on HIV/AIDS. Global Report fact sheet: The global AIDS epidemic http://www.unaids.org/documents/20101123_FS_Global_em_en.pdf (2010)

  2. 2

    Brenchley, J. M. et al. CD4+ T cell depletion during all stages of HIV disease occurs predominantly in the gastrointestinal tract. J. Exp. Med. 200, 749–759 (2004)

    CAS  Article  Google Scholar 

  3. 3

    Mattapallil, J. J. et al. Massive infection and loss of memory CD4+ T cells in multiple tissues during acute SIV infection. Nature 434, 1093–1097 (2005)

    CAS  ADS  Article  Google Scholar 

  4. 4

    Nishimura, Y. et al. Resting naive CD4+ T cells are massively infected and eliminated by X4-tropic simian-human immunodeficiency viruses in macaques. Proc. Natl Acad. Sci. USA 102, 8000–8005 (2005)

    CAS  ADS  Article  Google Scholar 

  5. 5

    Aiken, C. Pseudotyping human immunodeficiency virus type 1 (HIV-1) by the glycoprotein of vesicular stomatitis virus targets HIV-1 entry to an endocytic pathway and suppresses both the requirement for Nef and the sensitivity to cyclosporin A. J. Virol. 71, 5871–5877 (1997)

    CAS  PubMed  PubMed Central  Google Scholar 

  6. 6

    Butler, S. L., Hansen, M. S. & Bushman, F. D. A quantitative assay for HIV DNA integration in vivo. Nature Med. 7, 631–634 (2001)

    CAS  Article  Google Scholar 

  7. 7

    Leavitt, A. D., Robles, G., Alesandro, N. & Varmus, H. E. Human immunodeficiency virus type 1 integrase mutants retain in vitro integrase activity yet fail to integrate viral DNA efficiently during infection. J. Virol. 70, 721–728 (1996)

    CAS  PubMed  PubMed Central  Google Scholar 

  8. 8

    Li, L. et al. Role of the non-homologous DNA end joining pathway in the early steps of retroviral infection. EMBO J. 20, 3272–3281 (2001)

    CAS  Article  Google Scholar 

  9. 9

    Chen, H., Boyle, T. J., Malim, M. H., Cullen, B. R. & Lyerly, H. K. Derivation of a biologically contained replication system for human immunodeficiency virus type 1. Proc. Natl Acad. Sci. USA 89, 7678–7682 (1992)

    CAS  ADS  Article  Google Scholar 

  10. 10

    Doitsh, G. et al. Abortive HIV infection mediates CD4 T cell depletion and inflammation in human lymphoid tissue. Cell 143, 789–801 (2010)

    CAS  Article  Google Scholar 

  11. 11

    Daniel, R., Katz, R. A. & Skalka, A. M. A role for DNA-PK in retroviral DNA integration. Science 284, 644–647 (1999)

    CAS  ADS  Article  Google Scholar 

  12. 12

    Baekelandt, V. et al. DNA-dependent protein kinase is not required for efficient lentivirus integration. J. Virol. 74, 11278–11285 (2000)

    CAS  Article  Google Scholar 

  13. 13

    Daniel, R. et al. Wortmannin potentiates integrase-mediated killing of lymphocytes and reduces the efficiency of stable transduction by retroviruses. Mol. Cell. Biol. 21, 1164–1172 (2001)

    CAS  Article  Google Scholar 

  14. 14

    Ariumi, Y., Turelli, P., Masutani, M. & Trono, D. DNA damage sensors ATM, ATR, DNA-PKcs, and PARP-1 are dispensable for human immunodeficiency virus type 1 integration. J. Virol. 79, 2973–2978 (2005)

    CAS  Article  Google Scholar 

  15. 15

    Callén, E. et al. Essential role for DNA-PKcs in DNA double-strand break repair and apoptosis in ATM-deficient lymphocytes. Mol. Cell 34, 285–297 (2009)

    Article  Google Scholar 

  16. 16

    Engelman, A., Mizuuchi, K. & Craigie, R. HIV-1 DNA integration: mechanism of viral DNA cleavage and DNA strand transfer. Cell 67, 1211–1221 (1991)

    CAS  Article  Google Scholar 

  17. 17

    Morozov, V. E., Falzon, M., Anderson, C. W. & Kuff, E. L. DNA-dependent protein kinase is activated by nicks and larger single-stranded gaps. J. Biol. Chem. 269, 16684–16688 (1994)

    CAS  PubMed  Google Scholar 

  18. 18

    Lau, A., Kanaar, R., Jackson, S. P. & O'Connor, M. J. Suppression of retroviral infection by the RAD52 DNA repair protein. EMBO J. 23, 3421–3429 (2004)

    CAS  Article  Google Scholar 

  19. 19

    Hill, R. & Lee, P. W. The DNA-dependent protein kinase (DNA-PK): More than just a case of making ends meet? Cell Cycle 9, 3460–3469 (2010)

    CAS  Article  Google Scholar 

  20. 20

    Nilsson, A., Sirzen, F., Lewensohn, R., Wang, N. & Skog, S. Cell cycle-dependent regulation of the DNA-dependent protein kinase. Cell Prolif. 32, 239–248 (1999)

    CAS  Article  Google Scholar 

  21. 21

    Chen, B. P. et al. Cell cycle dependence of DNA-dependent protein kinase phosphorylation in response to DNA double strand breaks. J. Biol. Chem. 280, 14709–14715 (2005)

    CAS  Article  Google Scholar 

  22. 22

    Nagasawa, M. et al. Nuclear translocation of the catalytic component of DNA-dependent protein kinase upon growth stimulation in normal human T lymphocytes. Cell Struct. Funct. 22, 585–594 (1997)

    CAS  Article  Google Scholar 

  23. 23

    Perelson, A. S., Neumann, A. U., Markowitz, M., Leonard, J. M. & Ho, D. D. HIV-1 dynamics in vivo: virion clearance rate, infected cell life-span, and viral generation time. Science 271, 1582–1586 (1996)

    CAS  ADS  Article  Google Scholar 

  24. 24

    Igarashi, T. et al. Macrophage are the principal reservoir and sustain high virus loads in rhesus macaques after the depletion of CD4+ T cells by a highly pathogenic simian immunodeficiency virus/HIV type 1 chimera (SHIV): Implications for HIV-1 infections of humans. Proc. Natl Acad. Sci. USA 98, 658–663 (2001)

    CAS  ADS  Article  Google Scholar 

  25. 25

    Shan, L. et al. Stimulation of HIV-1-specific cytolytic T lymphocytes facilitates elimination of latent viral reservoir after virus reactivation. Immunity 36, 491–501 (2012)

    CAS  Article  Google Scholar 

  26. 26

    Ho, D. D. et al. Rapid turnover of plasma virions and CD4 lymphocytes in HIV-1 infection. Nature 373, 123–126 (1995)

    CAS  ADS  Article  Google Scholar 

  27. 27

    Wei, X. et al. Viral dynamics in human immunodeficiency virus type 1 infection. Nature 373, 117–122 (1995)

    CAS  ADS  Article  Google Scholar 

  28. 28

    Martínez, E. et al. Changes in cardiovascular biomarkers in HIV-infected patients switching from ritonavir-boosted protease inhibitors to raltegravir. AIDS 26, 2315–2326 (2012)

    Article  Google Scholar 

  29. 29

    Collman, R. et al. An infectious molecular clone of an unusual macrophage-tropic and highly cytopathic strain of human immunodeficiency virus type 1. J. Virol. 66, 7517–7521 (1992)

    CAS  PubMed  PubMed Central  Google Scholar 

  30. 30

    Adachi, A. et al. Production of acquired immunodeficiency syndrome-associated retrovirus in human and nonhuman cells transfected with an infectious molecular clone. J. Virol. 59, 284–291 (1986)

    CAS  PubMed  PubMed Central  Google Scholar 

  31. 31

    Bolton, D. L. & Lenardo, M. J. Vpr cytopathicity independent of G2/M cell cycle arrest in human immunodeficiency virus type 1-infected CD4+ T cells. J. Virol. 81, 8878–8890 (2007)

    CAS  Article  Google Scholar 

  32. 32

    Yamamoto, T. et al. Selective transmission of R5 HIV-1 over X4 HIV-1 at the dendritic cell-T cell infectious synapse is determined by the T cell activation state. PLoS Pathog. 5, e1000279 (2009)

    Article  Google Scholar 

  33. 33

    O'Doherty, U., Swiggard, W. J. & Malim, M. H. Human immunodeficiency virus type 1 spinoculation enhances infection through virus binding. J. Virol. 74, 10074–10080 (2000)

    CAS  Article  Google Scholar 

  34. 34

    Willmore, E. et al. A novel DNA-dependent protein kinase inhibitor, NU7026, potentiates the cytotoxicity of topoisomerase II poisons used in the treatment of leukemia. Blood 103, 4659–4665 (2004)

    CAS  Article  Google Scholar 

  35. 35

    Leahy, J. J. et al. Identification of a highly potent and selective DNA-dependent protein kinase (DNA-PK) inhibitor (NU7441) by screening of chromenone libraries. Bioorg. Med. Chem. Lett. 14, 6083–6087 (2004)

    CAS  Article  Google Scholar 

  36. 36

    Komarov, P. G. et al. A chemical inhibitor of p53 that protects mice from the side effects of cancer therapy. Science 285, 1733–1737 (1999)

    CAS  Article  Google Scholar 

  37. 37

    Suzuki, Y. et al. Quantitative analysis of human immunodeficiency virus type 1 DNA dynamics by real-time PCR: integration efficiency in stimulated and unstimulated peripheral blood mononuclear cells. Virus Genes 27, 177–188 (2003)

    CAS  Article  Google Scholar 

  38. 38

    Li, Y. et al. Complete nucleotide sequence, genome organization, and biological properties of human immunodeficiency virus type 1 in vivo: evidence for limited defectiveness and complementation. J. Virol. 66, 6587–6600 (1992)

    CAS  PubMed  PubMed Central  Google Scholar 

  39. 39

    O'Doherty, U., Swiggard, W. J., Jeyakumar, D., McGain, D. & Malim, M. H. A sensitive, quantitative assay for human immunodeficiency virus type 1 integration. J. Virol. 76, 10942–10950 (2002)

    CAS  Article  Google Scholar 

Download references

Acknowledgements

We thank D. Ambrozak, R. Nguyen, and S. Perfetto for help with cell sorting, U. Olshevsky, J. Casazza, D. Bolton, A. Pegu and M. Louder for discussions and technical help, and A. Tislerics and B. Hartman for manuscript preparation. This research was supported by the Intramural Research Program of the Vaccine Research Center, NIAID, National Institutes of Health. The findings and conclusions in this report are those of the authors and do not necessarily reflect the views of the funding agency.

Author information

Affiliations

Authors

Contributions

A.C. and G.J.N. designed the research studies; A.C., M.G, C.P. and T.Y. performed the research; A.C., M.G. and T.Y. contributed to development and generation of vectors; A.C., C.P., R.A.K. and G.J.N. analysed data; and A.C., R.A.K. and G.J.N. wrote the paper.

Corresponding author

Correspondence to Gary J. Nabel.

Ethics declarations

Competing interests

The authors declare no competing financial interests.

Supplementary information

Supplementary Information

This file contains Supplementary Figures 1-4 and Supplementary References. These figures show that HIV-1 induced cell death depends on provirus integration and is observed in p24- CD4+ lymphocytes from healthy donors infected in vitro and patient samples ex vivo. Blocking death by DNA-PK inhibitors does not result from diminished viral replication or general resistance to cell death. (PDF 621 kb)

PowerPoint slides

Rights and permissions

Reprints and Permissions

About this article

Cite this article

Cooper, A., García, M., Petrovas, C. et al. HIV-1 causes CD4 cell death through DNA-dependent protein kinase during viral integration. Nature 498, 376–379 (2013). https://doi.org/10.1038/nature12274

Download citation

Further reading

Comments

By submitting a comment you agree to abide by our Terms and Community Guidelines. If you find something abusive or that does not comply with our terms or guidelines please flag it as inappropriate.

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