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Proteomic profiling of HIV-1 infection of human CD4+ T cells identifies PSGL-1 as an HIV restriction factor

Abstract

Human immunodeficiency virus (HIV) actively modulates the protein stability of host cells to optimize viral replication. To systematically examine this modulation in HIV infection, we used isobaric tag-based mass spectrometry to quantify changes in the abundance of over 14,000 proteins during HIV-1 infection of human primary CD4+ T cells. We identified P-selectin glycoprotein ligand 1 (PSGL-1) as an HIV-1 restriction factor downregulated by HIV-1 Vpu, which binds to PSGL-1 and induces its ubiquitination and degradation through the ubiquitin ligase SCFβ-TrCP2. PSGL-1 is induced by interferon-γ in activated CD4+ T cells to inhibit HIV-1 reverse transcription and potently block viral infectivity by incorporating in progeny virions. This infectivity block is antagonized by Vpu via PSGL-1 degradation. We further show that PSGL-1 knockdown can significantly abolish the anti-HIV activity of interferon-γ in primary CD4+ T cells. Our study identifies an HIV restriction factor and a key mediator of interferon-γ’s anti-HIV activity.

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Fig. 1: Proteomic profiling of HIV-1 infection in human primary CD4+ T cells.
Fig. 2: Comparison of proteomic profiling results with other large data sets for cross-validation and identification of candidate genes.
Fig. 3: PSGL-1 is degraded by HIV-1 via ubiquitination by the Vpu-hijacked SCFβ-TrCP2 E3 ligase.
Fig. 4: PSGL-1 in target cells inhibits HIV-1 replication, starting from DNA synthesis.
Fig. 5: PSGL-1 in producer cells inhibits the infectivity of HIV-1 progeny virions.
Fig. 6: PSGL-1 mediates the anti-HIV activity of IFN-γ in activated primary CD4+ T cells.

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Code availability

The computational code used in this study is available from the corresponding authors upon request.

Data availability

The data that support the findings of this study are available from the corresponding authors upon request. The raw proteomic data have been uploaded to the ProteomeXchange data repository with the ID number: PXD011233.

References

  1. Malim, M. H. & Bieniasz, P. D. HIV restriction factors and mechanisms of evasion. Cold Spring Harb. Perspect. Med. 2, a006940 (2012).

    Article  Google Scholar 

  2. Yan, N. & Chen, Z. J. Intrinsic antiviral immunity. Nat. Immunol. 13, 214–222 (2012).

    Article  CAS  Google Scholar 

  3. Towers, G. J. & Noursadeghi, M. Interactions between HIV-1 and the cell-autonomous innate immune system. Cell Host Microbe 16, 10–18 (2014).

    Article  CAS  Google Scholar 

  4. Altfeld, M. & Gale, M. Jr. Innate immunity against HIV-1 infection. Nat. Immunol. 16, 554–562 (2015).

    Article  CAS  Google Scholar 

  5. Simon, V., Bloch, N. & Landau, N. R. Intrinsic host restrictions to HIV-1 and mechanisms of viral escape. Nat. Immunol. 16, 546–553 (2015).

    Article  CAS  Google Scholar 

  6. Harris, R. S., Hultquist, J. F. & Evans, D. T. The restriction factors of human immunodeficiency virus. J. Biol. Chem. 287, 40875–40883 (2012).

    Article  CAS  Google Scholar 

  7. Daugherty, M. D. & Malik, H. S. Rules of engagement: molecular insights from host–virus arms races. Annu. Rev. Genet. 46, 677–700 (2012).

    Article  CAS  Google Scholar 

  8. Stremlau, M. et al. The cytoplasmic body component TRIM5α restricts HIV-1 infection in Old World monkeys. Nature 427, 848–853 (2004).

    Article  CAS  Google Scholar 

  9. Sheehy, A. M., Gaddis, N. C., Choi, J. D. & Malim, M. H. Isolation of a human gene that inhibits HIV-1 infection and is suppressed by the viral Vif protein. Nature 418, 646–650 (2002).

    Article  CAS  Google Scholar 

  10. Neil, S. J., Zang, T. & Bieniasz, P. D. Tetherin inhibits retrovirus release and is antagonized by HIV-1 Vpu. Nature 451, 425–430 (2008).

    Article  CAS  Google Scholar 

  11. Hrecka, K. et al. Vpx relieves inhibition of HIV-1 infection of macrophages mediated by the SAMHD1 protein. Nature 474, 658–661 (2011).

    Article  CAS  Google Scholar 

  12. Laguette, N. et al. SAMHD1 is the dendritic- and myeloid-cell-specific HIV-1 restriction factor counteracted by Vpx. Nature 474, 654–657 (2011).

    Article  CAS  Google Scholar 

  13. Rosa, A. et al. HIV-1 Nef promotes infection by excluding SERINC5 from virion incorporation. Nature 526, 212–217 (2015).

    Article  CAS  Google Scholar 

  14. Usami, Y., Wu, Y. & Gottlinger, H. G. SERINC3 and SERINC5 restrict HIV-1 infectivity and are counteracted by Nef. Nature 526, 218–223 (2015).

    Article  CAS  Google Scholar 

  15. Yu, X. et al. Induction of APOBEC3G ubiquitination and degradation by an HIV-1 Vif–Cul5–SCF complex. Science 302, 1056–1060 (2003).

    Article  CAS  Google Scholar 

  16. Mitchell, R. S. et al. Vpu antagonizes BST-2-mediated restriction of HIV-1 release via β-TrCP and endo-lysosomal trafficking. PLoS Pathog. 5, e1000450 (2009).

    Article  Google Scholar 

  17. Van Damme, N. et al. The interferon-induced protein BST-2 restricts HIV-1 release and is downregulated from the cell surface by the viral Vpu protein. Cell Host Microbe 3, 245–252 (2008).

    Article  Google Scholar 

  18. Matheson, N. J. et al. Cell surface proteomic map of HIV infection reveals antagonism of amino acid metabolism by Vpu and Nef. Cell Host Microbe 18, 409–423 (2015).

    Article  CAS  Google Scholar 

  19. Greenwood, E. J. et al. Temporal proteomic analysis of HIV infection reveals remodelling of the host phosphoproteome by lentiviral Vif variants. eLife 5, e18296 (2016).

    Article  Google Scholar 

  20. Chan, E. Y. et al. Dynamic host energetics and cytoskeletal proteomes in human immunodeficiency virus type 1-infected human primary CD4 cells: analysis by multiplexed label-free mass spectrometry. J. Virol. 83, 9283–9295 (2009).

    Article  CAS  Google Scholar 

  21. Wojcechowskyj, J. A. et al. Quantitative phosphoproteomics reveals extensive cellular reprogramming during HIV-1 entry. Cell Host Microbe 13, 613–623 (2013).

    Article  CAS  Google Scholar 

  22. Zhou, F. et al. Genome-scale proteome quantification by DEEP SEQ mass spectrometry. Nat. Commun. 4, 2171 (2013).

    Article  Google Scholar 

  23. Sugden, S. M., Bego, M. G., Pham, T. N. & Cohen, E. A. Remodeling of the host cell plasma membrane by HIV-1 Nef and Vpu: a strategy to ensure viral fitness and persistence. Viruses 8, 67 (2016).

    Article  Google Scholar 

  24. van ‘t Wout, A. B. et al. Nef induces multiple genes involved in cholesterol synthesis and uptake in human immunodeficiency virus type 1-infected T cells. J. Virol. 79, 10053–10058 (2005).

    Article  Google Scholar 

  25. Rusinova, I. et al. Interferomev2.0: an updated database of annotated interferon-regulated genes. Nucleic Acids Res. 41, D1040–D1046 (2013).

    Article  CAS  Google Scholar 

  26. McLaren, P. J. et al. Identification of potential HIV restriction factors by combining evolutionary genomic signatures with functional analyses. Retrovirology 12, 41 (2015).

    Article  Google Scholar 

  27. Kosiol, C. et al. Patterns of positive selection in six mammalian genomes. PLoS Genet. 4, e1000144 (2008).

    Article  Google Scholar 

  28. Cohen, G. B. et al. The selective downregulation of class I major histocompatibility complex proteins by HIV-1 protects HIV-infected cells from NK cells. Immunity 10, 661–671 (1999).

    Article  CAS  Google Scholar 

  29. Zahoor, M. A. et al. HIV-1 Vpr induces interferon-stimulated genes in human monocyte-derived macrophages. PLoS ONE 9, e106418 (2014).

    Article  Google Scholar 

  30. Maitra, R. K. & Silverman, R. H. Regulation of human immunodeficiency virus replication by 2′,5′-oligoadenylate-dependent RNase L. J. Virol. 72, 1146–1152 (1998).

    CAS  PubMed  PubMed Central  Google Scholar 

  31. Shah, A. H. et al. Degranulation of natural killer cells following interaction with HIV-1-infected cells is hindered by downmodulation of NTB-A by Vpu. Cell Host Microbe 8, 397–409 (2010).

    Article  CAS  Google Scholar 

  32. Grover, J. R., Veatch, S. L. & Ono, A. Basic motifs target PSGL-1, CD43, and CD44 to plasma membrane sites where HIV-1 assembles. J. Virol. 89, 454–467 (2015).

    Article  Google Scholar 

  33. Mangeat, B. et al. HIV-1 Vpu neutralizes the antiviral factor tetherin/BST-2 by binding it and directing its β-TrCP2-dependent degradation. PLoS Pathog. 5, e1000574 (2009).

    Article  Google Scholar 

  34. Margottin, F. et al. A novel human WD protein, h-βTrCp, that interacts with HIV-1 Vpu connects CD4 to the ER degradation pathway through an F-box motif. Mol. Cell 1, 565–574 (1998).

    Article  CAS  Google Scholar 

  35. Shirogane, T., Jin, J., Ang, X. L. & Harper, J. W. SCFβ-TrCP controls clock-dependent transcription via casein kinase 1-dependent degradation of the mammalian period-1 (Per1) protein. J. Biol. Chem. 280, 26863–26872 (2005).

    Article  CAS  Google Scholar 

  36. Vigan, R. & Neil, S. J. Determinants of tetherin antagonism in the transmembrane domain of the human immunodeficiency virus type 1 Vpu protein. J. Virol. 84, 12958–12970 (2010).

    Article  CAS  Google Scholar 

  37. Cavrois, M., De Noronha, C. & Greene, W. C. A sensitive and specific enzyme-based assay detecting HIV-1 virion fusion in primary T lymphocytes. Nat. Biotechnol. 20, 1151–1154 (2002).

    Article  CAS  Google Scholar 

  38. Wu, Y., Beddall, M. H. & Marsh, J. W. Rev-dependent indicator T cell line. Curr. HIV Res. 5, 394–402 (2007).

    Article  CAS  Google Scholar 

  39. Lam, S. S. et al. Directed evolution of APEX2 for electron microscopy and proximity labeling. Nat. Methods 12, 51–54 (2015).

    Article  CAS  Google Scholar 

  40. Lamsoul, I., Uttenweiler-Joseph, S., Moog-Lutz, C. & Lutz, P. G. Cullin 5-RING E3 ubiquitin ligases, new therapeutic targets? Biochimie 122, 339–347 (2016).

    Article  CAS  Google Scholar 

  41. Sauter, D. & Kirchhoff, F. Multilayered and versatile inhibition of cellular antiviral factors by HIV and SIV accessory proteins. Cytokine Growth Factor Rev. 40, 3–12 (2018).

    Article  CAS  Google Scholar 

  42. Hotter, D. & Kirchhoff, F. Interferons and beyond: induction of antiretroviral restriction factors. J. Leukoc. Biol. 103, 465–477 (2018).

    Article  CAS  Google Scholar 

  43. Rihn, S. J. et al. The envelope gene of transmitted HIV-1 resists a late interferon γ-induced block. J. Virol. 91, e02254-16 (2017).

    Article  Google Scholar 

  44. Zhang, Y. et al. A robust error model for iTRAQ quantification reveals divergent signaling between oncogenic FLT3 mutants in acute myeloid leukemia. Mol. Cell. Proteomics 9, 780–790 (2010).

    Article  CAS  Google Scholar 

  45. Jiang, C. et al. A non-viral CRISPR/Cas9 delivery system for therapeutically targeting HBV DNA and pcsk9 in vivo. Cell Res. 27, 440–443 (2017).

    Article  CAS  Google Scholar 

  46. Roesch, F. et al. Hyperthermia stimulates HIV-1 replication. PLoS Pathog. 8, e1002792 (2012).

    Article  CAS  Google Scholar 

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

    Article  CAS  Google Scholar 

  48. Martell, J. D., Deerinck, T. J., Lam, S. S., Ellisman, M. H. & Ting, A. Y. Electron microscopy using the genetically encoded APEX2 tag in cultured mammalian cells. Nat. Protoc. 12, 1792–1816 (2017).

    Article  CAS  Google Scholar 

Download references

Acknowledgements

This work was supported by grants from the State Key Research Development Program of China to X.T. (no. 2016YFC1200303) and F.Z. (no. 2017YFA0505100), the China National Funds for Excellent Young Scientists (no. 31722030), grants from the National Natural Science Foundation of China to X.-F.Y. (no. 81772169) and F.Z. (no. 31670836), a grant from the Shanghai Institute of Higher Learning to F.Z. (no. TP2015003) and a grant from the NIMH, US Public Health Service to Y.W. (no. 5R01MH102144). We thank Q. Wang and X. Ye (Institute of Microbiology of the Chinese Academy of Sciences) for the use of the Biosafety level 3 Tissue Culture Facility. We thank H. Qi, N. Yan, H. Deng, A. Elia and N. Zheng for critical reading of the manuscript.

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Authors and Affiliations

Authors

Contributions

X.T. conceived and supervised the project. F.Z. conducted the mass spectrometry experiment and data analysis. Y.L. conducted most of the experiments with help from C.F., H.Z., S.L., T.Z. and X.K. Y.F., Z.Z. and D.D. conducted experiments on PSGL-1-blocking viral infectivity and western blot detection of PSGL-1 in virions. Y.W. supervised experiments on PSGL-1-blocking viral infectivity. J.Su and X.-F.Y. contributed to supervision and data analysis of the project. J.Sun and Y.Z. contributed key reagents and performed clinical analysis of the study. Q.W., J.G., M.L., W.Z. and Z.S. contributed to the data analysis. X.T. and F.Z. wrote and Y.W. edited the manuscript.

Corresponding authors

Correspondence to Feng Zhou, Yuntao Wu or Xu Tan.

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The authors declare no competing interests.

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Supplementary information

Supplementary Information

Supplementary Figures 1–6, Supplementary Table 1 and legends for Supplementary Datasets.

Reporting Summary

Supplementary Dataset 1

Raw mass spectrometry intensities of all identified proteins.

Supplementary Dataset 2

List of proteins with differential abundances among cell populations.

Supplementary Dataset 3

Gene ontology analysis of proteins with differential abundances among cell populations.

Supplementary Dataset 4

List of interferon-stimulated genes.

Supplementary Dataset 5

List of read counts of RNA-seq analysis.

Supplementary Dataset 6

List of genes with differential mRNA levels among cell populations.

Supplementary Dataset 7

List of positively selected genes.

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Liu, Y., Fu, Y., Wang, Q. et al. Proteomic profiling of HIV-1 infection of human CD4+ T cells identifies PSGL-1 as an HIV restriction factor. Nat Microbiol 4, 813–825 (2019). https://doi.org/10.1038/s41564-019-0372-2

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