Harnessing the PIWI piRNA pathway to silence HIV

项目总结 很明显，在接受CART治疗的HIV感染者中，潜伏感染的CD4+T细胞是主要的储存库 以及根除的障碍。这项研究提案特别打算处理题为《沉默》的《FOA》。 与FOA提出的以下具体目标相结合：利用PIWI RNA 消除艾滋病毒的途径“。与其他小的非编码RNA沉默途径(siRNA，shRNA， MiRNA)，piRNA途径具有通过表观遗传永久沉默基因的理论优势 除mRNA降解活性外的修饰(H3K9甲基化)，后者是主要模式 其他小RNA的沉默，从而确保后代细胞中的基因沉默。我们的初步调查结果 表明PIWI蛋白在CD4+T细胞和T细胞系中表达，并似乎受 免疫激活。然而，目前尚不清楚是否可以诱导PIWI-piRNA途径执行 在体细胞中的功能与在生殖细胞中的功能相似。我们假设piRNA-PIWI途径是 在原始的CD4+T细胞中起作用，可以被操纵来沉默HIV。我们将努力确保 通过将基础小非编码RNA和piRNA生物学方面的专家与 艾滋病毒专家，所有人在各自领域都有长期的记录。在建议书的R66部分，我们将测试证明 可以在ACH-2细胞中诱导针对HIV基因的piRNA途径的概念，因此，当他们 被细胞因子激活后，它们已停止产生艾滋病毒病毒粒子。如果成功，则R33部分 该提案将进一步鉴定对多种艾滋病毒毒株具有活性ACH-2 piRNA， 包括取自CART处理个体的体外原代细胞内的交叉分支。作为临床前工作的一部分 将探索开发一种以CD4受体为靶点的嵌合适体-piRNA策略。目前的提案 将揭示关于piRNA途径在人类细胞中的作用的进一步见解，它们是否可以被用作 作为艾滋病毒治愈疗法的一部分的艾滋病毒沉默战略。

PROJECT SUMMARY It is clear that latently infected CD4+ T cells in cART treated HIV-infected individuals are the major reservoir and obstacle to eradication. This research proposal specifically intends to address the FOA entitled “Silencing of HIV-1 Proviruses” with the following specific objective proposed by the FOA: “ to harness the PIWI RNA pathway to silence HIV”. In contrast to other small non-coding RNA silencing pathways (siRNA, shRNA, miRNA), the piRNA pathway has the theoretical advantage of permanently silencing genes through epigenetic modifications (H3K9 methylation) in addition to mRNA degradation activity, the latter being the primary mode of silencing by other small RNAs, thus, ensuring gene silencing in progeny cells. Our preliminary findings indicate that PIWI proteins are expressed in CD4+ T cells and T cell lines, and appear to be regulated by immune activation. It is unknown, however whether the PIWI-piRNA pathway can be induced to perform similar functions in somatic cells as they do in germ cells. We hypothesize that the piRNA-PIWI pathway is functional in primary CD4+ T-cells and can be manipulated to silence HIV. Attempts will be made to ensure success of the project by teaming experts in fundamental small noncoding RNA and piRNA biology with an HIV expert, all having long track records in their fields. In the R66 portion of the proposal we will test the proof of concept that that one can induce a piRNA pathway against HIV genes in ACH-2 cells, so that, when they are activated by cytokine, they have ceased to produce HIV virions. If successful, the R33 portion of the proposal will further characterize ACH-2 piRNAs that have activity against a broad range of HIV strains, including cross Clades within ex vivo primary cells taken from cART treated individuals. As part of pre-clinical development a CD4 receptor targeting chimeric aptamer-piRNA strategy will be explored. The current proposal will reveal further insights on the role of the piRNA pathway in human cells, whether they can be exploited as an HIV silencing strategy as part of the armamentarium toward HIV cure therapy.