Mobilization-competent Lentiviral Vector–mediated Sustained Transcriptional Modulation of HIV-1 Expression

Abstract

Current anti-HIV-1 strategies reduce replication through targeting of viral proteins and RNA; meanwhile, targeting at the level of the integrated provirus has been less explored. We show here that mobilization-competent vectors containing small noncoding RNAs targeted to transcriptionally active regions of the human immunodeficiency virus type 1 (HIV-1) long terminal repeat (LTR) can take advantage of integrated virus and modulate HIV-1 replication. Transcriptional silencing of HIV-1 correlates with an increase in silent-state epigenetic marks including histone and DNA methylation, a loss of nuclear factor-kappaB (NF-kappaB) recruitment, and requires Argonaute 1 (Ago-1), histone deacetylase 1 (HDAC-1), and DNA methyltransferase 3a (DNMT3a) localization to the LTR. Long-term suppression of the virus was observed for 1 month with no evidence of viral resistance. These data show that RNA-directed transcriptional silencing of HIV-1 can be delivered by a mobilization-competent vector, suggesting that this system could be used to target long-term selective pressures on conserved promoter elements to evolve less pathogenic variants of HIV-1.

Epigenetics and ncRNAs in Brain Function and Disease: Mechanisms and Prospects for Therapy

“…Subsequently, recruitment of RITS induces heterochromatin formation and promoter DNA methylation at the target promoter. Because RITS is guided to a complementary promoter-associated RNA by its bound small RNA, targeting a promoter with a small RNA TGS effector can lead to chromatin remodelling (promoters become enriched in the silent state chromatin marks H3K9me2 and H3K27me3) and, in some cases, promoter DNA methylation [108,109].…”

“…TGS can be induced using siRNAs, antisense oligonucleotides [109], expressed antisense RNAs [110] or short hairpin RNAs [111,112]. And there are also examples of the successful implementation of this approach in vivo by local delivery in muscle [113], retina [112] and xenograft models [114].…”

The theory of RNA-mediated gene evolution

“…2). The notion presented here is that noncoding RNA-associated recruitment of DNA methyltransferase 3a (DNMT3a) and Enhancer of Zeste 2 (EZH2) to targeted loci can result in those cytosines at the target locus undergoing DNA methylation 17,[19][20][21][22][23] ( Fig. 2A-D).…”

“…This form of silencing involves the action of DNMT3a interacting directly with the small non-coding RNAs 20,21,44,45 and directing both histone and DNA methylation to the homology containing loci. 17,22 This targeting can ultimately result in stable and heritable epigenetically silent loci. 17 (C and D) Long non-coding RNAs that are antisense to particular loci have also been found to interact with DNMT3a 19 and modulate transcription.…”

“…Collectively, data generated over the last decade suggest that: (1) non-coding RNAs direct epigenetic gene silencing and DNA methylation to those CpGs in the RNA targeted locus 17,22,40 ( Fig. 1); (2) methylated cytosines that do not undergo proper deamination and repair can become incorporated into the genome as thymines (Fig.…”