A functional map of HIV-host interactions in primary human T cells

Abstract

Human Immunodeficiency Virus (HIV) relies on host molecular machinery for replication. Systematic attempts to genetically or biochemically define these host factors have yielded hundreds of candidates, but few have been functionally validated in primary cells. Here, we target 426 genes previously implicated in the HIV lifecycle through protein interaction studies for CRISPR-Cas9-mediated knock-out in primary human CD4+ T cells in order to systematically assess their functional roles in HIV replication. We achieve efficient knockout (>50% of alleles) in 364 of the targeted genes and identify 86 candidate host factors that alter HIV infection. 47 of these factors validate by multiplex gene editing in independent donors, including 23 factors with restrictive activity. Both gene editing efficiencies and HIV-1 phenotypes are highly concordant among independent donors. Importantly, over half of these factors have not been previously described to play a functional role in HIV replication, providing numerous novel avenues for understanding HIV biology. These data further suggest that host-pathogen protein-protein interaction datasets offer an enriched source of candidates for functional host factor discovery and provide an improved understanding of the mechanics of HIV replication in primary T cells.

Directing HIV-1 for degradation by non-target cells, using bi-specific single-chain llama antibodies

“…In general however, since earlier controverses about the fate of HIV-1 in endosomal vesicles, much evidence indicated that HIV-1 is degraded in lysosomes [53][54][55] and generally requires CD4 and a coreceptor for productive infection [56][57][58] . In addition, not all cell types will provide all the gene products required to support viral propagation [59][60][61] . Note that also the natural Fc-mediated degradation of HIV-1 bound by conventional antibodies was recently indicated to occur by endocytosis (into liver sinusoidal endothelial cells) and subsequent degradation in lysosomes 62 .…”

Closing the Door with CRISPR: Genome Editing of CCR5 and CXCR4 as a Potential Curative Solution for HIV

“…For instance, Rebensburg et al showed that HIV-1 depends on Sec24C for replication through interactions with the viral core, and is important for reverse transcription, nuclear import and infectivity [ 39 ]. Other studies have similarly investigated other intracellular targets that are important for HIV infection and replication after cellular entry [ 40 , 41 ].…”

“…Nonetheless, CXCR4 knockout was shown to be feasible in cell lines [ 86 , 87 ] and primary CD4 + T cells [ 58 , 94 , 95 ]. Furthermore, CXCR4 knockout in primary human CD4 + T cells conferred in vitro resistance to HIV infection [ 25 , 40 , 74 ].…”

“…CRISPR has also been used as a research tool in cell lines to further understand HIV biology and host proteins involved in the anti-viral response, or contributing to viral success [ 86 , 87 ]. Recently, a CRISPR-screen was performed to identify host proteins that interact with HIV in primary human T cells [ 40 ]. Here, authors identified 62 proteins that are so-called dependency factors, proteins that HIV requires in order to infect and/or propagate in cells.…”

“…Nonetheless, CXCR4 knockout was shown to be feasible in cell lines [86,87] and primary CD4 + T cells [58,94,95]. Furthermore, CXCR4 knockout in primary human CD4 + T cells conferred in vitro resistance to HIV infection [25,40,74]. In contrast to CCR5, no studies are currently available describing the use of CXCR4 knockout cells in murine models or humans.…”