Interactions of Monocytes, HIV, and ART Identified by an Innovative scRNAseq Pipeline: Pathways to Reservoirs and HIV-Associated Comorbidities

HIV reservoirs persist despite successful antiretroviral therapy (ART) and are a major obstacle to the eradication and cure of HIV. The mature monocyte subset, CD14(+)CD16(+), contributes to viral reservoirs and HIV-associated comorbidities. Only a subset of monocytes harbors HIV (HIV(+)), while the rest remain uninfected, exposed cells (HIV(exp)). We developed an innovative single cell RNA sequencing (scRNAseq) pipeline that detects HIV and host transcripts simultaneously, enabling us to examine differences between HIV(+) and HIV(exp) mature monocytes. Using this, we characterized uninfected, HIV(+), and HIV(exp) primary human mature monocytes with and without ART. We showed that HIV(+) mature monocytes do not form their own cluster separately from HIV(exp) but can be distinguished by significant differential gene expression. We found that ART decreased levels of unspliced HIV transcripts potentially by modulating host transcriptional regulators shown to decrease viral infection and replication. We also identified and characterized mature monocyte subpopulations differentially impacted by HIV and ART. We identified genes dysregulated by ART in HIV(exp) monocytes compared to their uninfected counterpart and, of interest, the junctional protein ALCAM, suggesting that ART impacts monocyte functions. Our data provide a novel method for simultaneous detection of HIV and host transcripts. We identify potential targets, such as those genes whose expression is increased in HIV(+) mature monocytes compared to HIV(exp), to block their entry into tissues, preventing establishment/replenishment of HIV reservoirs even with ART, thereby reducing and/or eliminating viral burden and HIV-associated comorbidities. Our data also highlight the heterogeneity of mature monocyte subsets and their potential contributions to HIV pathogenesis in the ART era.