The Immunophenotype and Proviral Landscape of HIV-infected CD4 T Cells During Antiretroviral Therapy

In individuals on effective antiretroviral therapy, integrated HIV proviruses persist within CD4 T cells, forming a viral reservoir that rebounds if treatment is stopped. Identifying and targeting these rare, infected cells is critical for advancing therapies, but methods to study reservoir cells are limited and their unique properties remain largely unknown. We applied DAb-seq, a high-throughput method that combines single-cell DNA and surface protein sequencing, to profile over five hundred and twenty thousand CD4 T cells from the blood of six individuals on ART. Infected cells were unequally distributed in T cell subsets, and differential protein expression between infected and uninfected cells revealed significant heterogeneity across cell subsets. Attempts to identify surface markers that differentiate infected from uninfected cells found antigens that mirrored the enrichment of HIV in central memory subsets. However, while central memory T cells harbored the majority of HIV, cells with intact provirus were enriched relative to their defective counterparts in Naïve and Regulatory T cell subsets, suggesting that they differentially maintain intact proviruses. In summary, we developed DAb-seq as an open-source platform for linking the proviral landscape to diverse cellular phenotypes, revealing heterogeneity in surface protein expression and provirus maintenance across infected subsets.