Epigenetic Silencing of HIV-1 by the Histone H3 Lysine 27 Methyltransferase Enhancer of Zeste 2

Abstract

Latent HIV proviruses are silenced as the result of deacetylation and methylation of histones located at the viral long terminal repeat (LTR). Inhibition of histone deacetylases (HDACs) leads to the reemergence of HIV-1 from latency, but the contribution of histone lysine methyltransferases (HKMTs) to maintaining HIV latency remains uncertain. Chromatin immunoprecipitation experiments using latently infected Jurkat T-cell lines demonstrated that the HKMT enhancer of Zeste 2 (EZH2) was present at high levels at the LTR of silenced HIV proviruses and was rapidly displaced following proviral reactivation. Knockdown of EZH2, a key component of the Polycomb repressive complex 2 (PRC2) silencing machinery, and the enzyme which is required for trimethyl histone lysine 27 (H3K27me3) synthesis induced up to 40% of the latent HIV proviruses. In contrast, there was less than 5% induction of latent proviruses following knockdown of SUV39H1, which is required for H3K9me3 synthesis. Knockdown of EZH2 also sensitized latent proviruses to external stimuli, such as T-cell receptor stimulation, and slowed the reversion of reactivated proviruses to latency. Similarly, cell populations that responded poorly to external stimuli carried HIV proviruses that were enriched in H3K27me3 and relatively depleted in H3K9me3. Treating latently infected cells with the HKMT inhibitor 3-deazaneplanocin A, which targets EZH2, led to the reactivation of silenced proviruses, whereas chaetocin and BIX01294 showed only minimal reactivation activities. These findings suggest that PRC2-mediated silencing is an important feature of HIV latency and that inhibitors of histone methylation may play a useful role in induction strategies designed to eradicate latent HIV pools.

Post-transcriptional gene silencing, transcriptional gene silencing and human immunodeficiency virus

“…Additionally, EZH2, one of the PCR2 subunits that establish H3K27me3, has been found to be present at the LTR of latent provirus. Knockdown of EZH2 resulted in higher transcriptional activation of the HIV promoter than when knocking down Suv39H1 [177] , indicating that the former is associated with a more responsive epigenetic silencing.…”

“…In HIV-1 infection, H3K27me3 has been found enriched in the 5'LTR promoter in cell line models of latent infection in which the virus reactivates upon stimulation [177] . This is consistent with H3K27me3 being generally a more flexible epigenetic repressive mark and with the likelihood that most of the inducible latent provirus is silenced through pathways involving H3K27me3, rather than H3K9me3.…”

Progress and challenges in the use of latent HIV-1 reactivating agents

“…Exposure of cells to 10 µmol/L DZNep led to global reductions in H3K27me3 (44% reduction) and H3K9me2 (70% reduction), accompanied by a reactivation of latent virus. A synergistic effect was observed when DZNep and SAHA were combined [54] . The selective combination of DZNep and an HDACi provided the most effective means to reverse the epigenetic gene silencing; therefore, DZNep, which targets H3K27me3 formation, has great potential as a selective inducer of latent HIV-1 proviruses.…”

“…BIX-01294 reactivated HIV-1 expression in 80% of resting CD4 + T cell cultures isolated from similar patients [32] . However, in another experimental system, BIX-01294 was a comparatively poor inducer of latent proviruses and was only able to induce 21.1% of the latent proviruses in the E4 cell line after overnight exposure to the drug [54] . The broad-spectrum agent DZNep was reported to downregulate several HKMTs and to target EZH2 [58] .…”

“…H3K9 trimethylation (H3K9me3), mediated by Suv39H1 [53] ; H3K27 trimethylation (H3K27me3) [54] and H3K9 dimethylation (H3K9me2), mediated by G9α [19] lead to HIV transcriptional silencing in different cell models, including peripheral blood mononuclear cells (PBMCs) from infected individuals. Therefore, HMTIs could also be used to purge HIV-1 latent reservoirs.…”