Clonal CD4+ T cells in the HIV-1 latent reservoir display a distinct gene profile upon reactivation

Abstract

Despite suppressive combination antiretroviral therapy (ART), latent HIV-1 proviruses persist in patients. This latent reservoir is established within 48–72 h after infection, has a long half-life1,2, enables viral rebound when ART is interrupted, and is the major barrier to a cure for HIV-13. Latent cells are exceedingly rare in blood (1 per 1 × 106 CD4+ T cells) and are typically enumerated by indirect means, such as viral outgrowth assays4,5. We report a new strategy to purify and characterize single reactivated latent cells from HIV-1-infected individuals on suppressive ART. Surface expression of viral envelope protein was used to enrich reactivated latent T cells producing HIV RNA, and single-cell analysis was performed to identify intact virus. Reactivated latent cells produce full-length viruses that are identical to those found in viral outgrowth cultures and represent clones of in vivo expanded T cells, as determined by their T cell receptor sequence. Gene-expression analysis revealed that these cells share a transcriptional profile that includes expression of genes implicated in silencing the virus. We conclude that reactivated latent T cells isolated from blood can share a gene-expression program that allows for cell division without activation of the cell death pathways that are normally triggered by HIV-1 replication.

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Fig. 1: Latency capture enriches for cells producing HIV RNA.
Fig. 2: Full-length virus sequences recovered by scRNA-seq.
Fig. 3: Captured cells express Env that is identical to latent virus emerging in Q2VOAs and represent expanded clones.
Fig. 4: A distinct gene signature defines reactivated latent cells.

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Acknowledgements

We thank all participants who contributed to this study; members of the Nussenzweig laboratory for helpful discussions, particularly E. Kara and T. Oliveira; our lab manager Z. Jankovic; L. Mesin and M. Biton for advice on scRNA-seq; A. Gazumyan for bNAb production; G. Breton for help with FACS; K. Gordon and N. Thomas for performing all FACSorting experiments; A. Han and M. Davis for TCR sequencing advice; D. Mucida, C. Rice and P. Bieniasz for helpful discussion; K. Millard for recruitment of study subjects; and M. Deal for assistance with figures. This work was supported by the Bill and Melinda Gates Foundation Collaboration for AIDS Vaccine Discovery (OPP1033115 and OPP1124068), the National Institutes of Health (NIH) Center for HIV/AIDS Vaccine Immunology and Immunogen Discovery (CHAVI-ID) (1UM1 AI100663), BEAT-HIV Delaney Collaboratory (UM1 AI126620), the National Institute of Allergy and Infectious Diseases of the NIH (AI100148, AI037526), the Robertson Foundation, and the Rockefeller University. M.C. is supported by NIH grant U01AI118536. M.C.N. is a Howard Hughes Medical Institute (HHMI) investigator.

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Contributions

L.B.C., M.J., and M.C.N. wrote the manuscript; L.B.C., M.J., and M.C.N. designed and analyzed experiments; L.B.C. and M.J. performed LURE experiments, RNA sequencing, Q2VOAs, TCR sequencing, and virus SGA; R.V. and I.T.d.S performed bioinformatics analysis of RNA-seq data; A.S.H. performed TCR sequencing and virus SGA; J.C.C.L. and Y.Z.C. performed Q2VOAs; J.A.P. performed phylogenetic analysis of env sequencing and gene enrichment analysis; A.L.B. and M.C. performed study subject recruitment and oversaw sample collection.

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Correspondence to Michel C. Nussenzweig.

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Supplementary Text and Figures

Supplementary Figures 1–11 and Supplementary Table 1

Reporting Summary

Supplementary Table 2

Genes that segregate cells into clusters by PCA

Supplementary Table 3

Differentially expressed gene list. Genes differently expressed by Env+ compared to control cells with P < 0.01

Supplementary Table 4

Enriched biological processes and molecular functions using Gene Ontology database.

Supplementary Table 5

Genes included in Gene Ontology categories.

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Cohn, L.B., da Silva, I.T., Valieris, R. et al. Clonal CD4+ T cells in the HIV-1 latent reservoir display a distinct gene profile upon reactivation. Nat Med 24, 604–609 (2018). https://doi.org/10.1038/s41591-018-0017-7

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