Epigenomic characterization of latent HIV infection identifies latency regulating transcription factors

Abstract

Transcriptional silencing of HIV in CD4 T cells generates a reservoir of latently infected cells that can reseed infection after interruption of therapy. As such, these cells represent the principal barrier to curing HIV infection, but little is known about their characteristics. To further our understanding of the molecular mechanisms of latency, we characterized a primary cell model of HIV latency in which infected cells adopt heterogeneous transcriptional fates. In this model, we observed that latency is a stable, heritable state that is transmitted through cell division. Using Assay of Transposon-Accessible Chromatin sequencing (ATACseq) we found that latently infected cells exhibit greatly reduced proviral accessibility, indicating the presence of chromatin-based structural barriers to viral gene expression. By quantifying the activity of host cell transcription factors, we observe elevated activity of Forkhead and Kruppel-like factor transcription factors (TFs), and reduced activity of AP-1, RUNX and GATA TFs in latently infected cells. Interestingly, latency reversing agents with different mechanisms of action caused distinct patterns of chromatin reopening across the provirus. We observe that binding sites for the chromatin insulator CTCF are highly enriched in the differentially open chromatin of infected CD4 T cells. Furthermore, depletion of CTCF inhibited HIV latency, identifying this factor as playing a key role in the initiation or enforcement of latency. These data indicate that HIV latency develops preferentially in cells with a distinct pattern of TF activity that promotes a closed proviral structure and inhibits viral gene expression. Furthermore, these findings identify CTCF as a novel regulator of HIV latency.

Genomic profiling of HIV-1 integration in microglia links viral insertions to TAD organization

“…NF-KB subunit displayed increased footprints in productive and latent infection compared to uninfected cells, while STAT1 binding was tracked in active versus latent condition (Figure 4A,B). Interestingly, CTCF, an architectural factor involved in 3D genome organization, had decreased TFBS footprints upon productive microglia infection, and higher occupancy in latently infected microglia when compared to productive infection, in line with a recent study showing evidence for CTCF contribution to HIV-1 latency in T cells (40) (Figure 4A-C). We further focused on differentially occupied CTCF binding sites in the three cell populations, and confirmed an overall reduction of CTCF occupancy in productive infection (Supplementary Figure S5C).…”

“…Likewise, TAD structure prediction was also reinforced by TF with enriched footprints in the latent cell population such as transcriptional repressor REST, also known as Neuron-Restrictive Silencer Factor (NRSF), and CTCF that, as expected, had the most prominent TAD contribution score (Figure 4E) (95, 96). To this end, the role of CTCF in viral replication and latency has been suggested for several DNA viruses, and recently for HIV-1 in primary CD4 + T cells (40, 97). Because HIV-1 proviruses do not harbour CTCF binding sites, it is plausible that CTCF contributes to viral gene expression through its role as an insulator in TAD border formation and looping interactions between nonadjacent segments of DNA (36, 37).…”

“…TADs are characterized by high frequency of internal DNA interactions that facilitate enhancer-promoter contacts, while their margins, referred to as TAD boundaries, are largely conserved among different cell types and are enriched for the insulator binding protein CCCTC-binding factor (CTCF) (36)(37)(38)(39). In the context of HIV-1 infection, a recent study applying Assay for Transposase-Accessible Chromatin using sequencing (ATAC-Seq) to actively and latently infected T cells suggested an increased occupancy and transcriptional repression mediated by architectural protein CTCF in latent cells, thus proposing that dynamic changes in chromatin organization could influence viral expression and persistence (40). Furthermore, genome organization assessment in patient-derived CD4 + T cells suggested that distinct chromatin accessibility adjacent to intact proviruses could contribute to a selective advantage for the virus during long-term ART (41).…”

UHRF1 Suppresses HIV-1 Transcription and Promotes HIV-1 Latency by Competing with p-TEFb for Ubiquitination-Proteasomal Degradation of Tat

“…Thus far, no clinically tested LRAs are able to intensely induce viral expression or diminish the latent reservoir in patients ( 2 , 34 , 35 ). Accumulating evidence demonstrates the critical role of host factors in regulating HIV-1 latency via silencing HIV-1 viral transcription, which are essential for the development of rational therapeutics for the eradication of latent virus ( 4 , 36 – 38 ). In this study, we found that UHRF1 plays a key role in the regulation of HIV-1 transcription and replication.…”

“…Over recent decades, the accumulated evidence shows that a panel of host factors is involved in regulation of HIV-1 latency. Genome-wide CRISPR-Cas9 screens identified FTSJ3, TMEM178A, NICN1, and the integrator complex as host regulators that promote HIV-1 latency ( 3 ), and a total of 471 genes were demonstrated to play key roles in HIV-1 initiation or the enforcement of latency through transposon-accessible chromatin sequencing (ATAC-seq) ( 4 ). Recently, our group identified the host factor heat shock factor 1 (HSF1) as a universal factor in latent HIV-1 reactivation induced by latency-reversing agents (LRAs) that is indispensable in HIV-1 transcription ( 5 ).…”