Proviral location affects cognate peptide–induced virus production and immune recognition of HIV-1–infected T cell clones
Filippo Dragoni, Abena K. Kwaa, Caroline C. Traut, Rebecca T. Veenhuis, Bezawit A. Woldemeskel, Angelica Camilo-Contreras, Hayley E. Raymond, Arbor G. Dykema, Eileen P. Scully, Amanda M. Rosecrans, Kellie N. Smith, Frederic D. Bushman, Francesco R. Simonetti, Joel N. Blankson
Filippo Dragoni, Abena K. Kwaa, Caroline C. Traut, Rebecca T. Veenhuis, Bezawit A. Woldemeskel, Angelica Camilo-Contreras, Hayley E. Raymond, Arbor G. Dykema, Eileen P. Scully, Amanda M. Rosecrans, Kellie N. Smith, Frederic D. Bushman, Francesco R. Simonetti, Joel N. Blankson
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Clinical Research and Public Health AIDS/HIV

Proviral location affects cognate peptide–induced virus production and immune recognition of HIV-1–infected T cell clones

Abstract

BACKGROUND HIV-1–infected CD4+ T cells contribute to latent reservoir persistence by proliferating while avoiding immune recognition. Integration features of intact proviruses in elite controllers (ECs) and people on long-term therapy suggest that proviruses in specific chromosomal locations can evade immune surveillance. However, direct evidence of this mechanism is missing.METHODS In this case report, we characterized integration sites and full genome sequences of expanded T cell clones in an EC before and after chemoradiation. We identified the cognate peptide of infected clones to investigate cell proliferation and virus production induced by T cell activation, and susceptibility to autologous CD8+ T cells.RESULTS The proviral landscape was dominated by 2 large clones with replication-competent proviruses integrated into zinc finger (ZNF) genes (ZNF470 and ZNF721) in locations previously associated with deeper latency. A third nearly intact provirus, with a stop codon in Pol, was integrated into an intergenic site. Upon stimulation with cognate Gag peptides, infected clones proliferated extensively and produced virus, but the provirus in ZNF721 was 200-fold less inducible. While autologous CD8+ T cells decreased the proliferation of cells carrying the intergenic provirus, they had no effect on cells with the provirus in the ZNF721 gene.CONCLUSIONS We provide direct evidence that upon activation of infected clones by cognate antigen, the lower inducibility of intact proviruses in ZNF genes can result in immune evasion and persistence.FUNDING Office of the NIH Director and National Institute of Dental & Craniofacial Research; NIAID, NIH; Johns Hopkins University Center for AIDS Research.

Authors

Filippo Dragoni, Abena K. Kwaa, Caroline C. Traut, Rebecca T. Veenhuis, Bezawit A. Woldemeskel, Angelica Camilo-Contreras, Hayley E. Raymond, Arbor G. Dykema, Eileen P. Scully, Amanda M. Rosecrans, Kellie N. Smith, Frederic D. Bushman, Francesco R. Simonetti, Joel N. Blankson

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Figure 3

PBMC stimulation with HIV-1 Gag peptides induces proliferation of infected cells and virus production.

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PBMC stimulation with HIV-1 Gag peptides induces proliferation of infect...
(A) CD8-depleted PBMCs collected on day 926 after CRT were cultured for 9 days in the presence of CMV lysate, a Gag peptide pool, or left untreated (No Tx); total LTR (R-U5) copies were quantified from genomic DNA on day 9. The same Gag peptide pool was used in 2 independent experiments from the same time point. (B) Stimulation with Gag peptides led to a marked increase in LTR copies, which was paralleled by virus production detected in the culture supernatant (C). (D) Maximum likelihood phylogeny analysis of HIV-1 RNA U5-gag single genome sequences; star symbols indicate node with bootstrap >75; black arrow indicates 11-nt deletion; highlighter plot on the right shows nucleotide mutations relative to ZNF470i. (E) Characterization of paired full genome sequence and integration site analysis of a new near-intact variant with a premature stop codon in Pol (indicated by asterisk). (F) Design of 2 competition probes used to distinguish intact U5-PBS from the Chr7.d11sc variant; the panel at the bottom shows a dPCR 2D plot of a sample containing both targets. (G) HIV-1 DNA copies per million cells from day 9 of culture by measuring ZNF470i, ZNF721i, or the 11-nt deletion from Chr7.d11sc; numbers above bar charts indicate fold-change from No Tx. (H) HIV-1 RNA copies per mL of supernatant at day 9 of culture; symbols represent 2 independent stimulations; error bars indicate SD.