Role of protein methylation in HIV latency

前病毒潜伏期仍然是从感染个体中根除HIV的重要障碍。表观遗传过程在调节HIV基因表达中至关重要。我们假设精氨酸和赖氨酸甲基化在前病毒潜伏期中起重要的沉默作用，并且甲基转移酶的小分子抑制剂可以在治疗上有助于潜伏感染的T细胞中的HIV的诱导/根除（I/E）。这一假设得到了文献中的数据和我们自己的初步未发表的研究的支持，在这些研究中，我们确定了几种甲基转移酶（MT）在潜伏感染的T细胞中对HIV转录具有抑制活性。项目1.5的目标是全面研究MT在持续性HIV感染中的作用，并与Martin Deianey合作实验室的研究人员密切合作，以验证MT作为HIV前病毒潜伏期I/E中的潜在药物靶标。具体目标1：全面分析J-Lat细胞和HIV潜伏期原代细胞培养模型中针对赖氨酸和精氨酸甲基转移酶的小发夹RNA。我们将对11种已知的人精氨酸MT、24种已知的人赖氨酸MT和38种基于与赖氨酸MT的保守SET结构域特征的结构域同源性鉴定的新蛋白进行高度靶向的shRNA筛选，以鉴定一组作为I/E疗法的潜在药物靶标的增强HIV潜伏期的MT。具体目标2：研究甲基转移酶控制HIV潜伏期的分子机制。我们将研究我们前期工作中鉴定的四个抑制性MT（PRMT 1，PRMT 6，Set 1和SETDB 1）的转录活性。我们将研究这些因子在体内向潜伏的HIV LTR的募集，并在潜伏感染的T细胞中鉴定这些酶的底物（组蛋白）。达特、P-TEFb或NF-κ B）。具体目标3：检测甲基转移酶抑制剂对潜伏感染T细胞中HIV转录再激活的影响。我们将评估已知的精氨酸和赖氨酸MT的小分子抑制剂对J-Lat细胞和潜伏期的原代T细胞模型中的HIV前病毒潜伏期的影响。我们将表征这些抑制剂主要靶向的酶，并将此信息用于开发有效和特异性的HIV I/E。总的来说，这些研究将为HIV前病毒潜伏期的基本机制提供新的分子见解，并支持该合作实验室的集体目标，即生产在动物和患者中持续HIV感染的I/E中进行测试的化合物。

Proviral latency remains an important barrier to eradication of HIV from infected individuals. Epigenetic processes are critical in the regulation of HIV gene expression. We hypothesize that arginine and lysine methylation plays an important silencing role in proviral latency and that small molecule inhibitors of methyltransferases can contribute therapeutically to the induction/eradication (l/E) of HIV in latently infected T cells. This hypothesis is supported by data in the literature and our own preliminary unpublished studies, in which we identified several methyltransferases (MTs) with suppressive activity against HIV transcription in latently infected T cells. The goal of Project 1.5 is to comprehensively study the role of MTs in persistent HIV infection and in close collaboration with investigators across the Martin Deianey Collaboratory to validate MTs as potential drug targets in the l/E of HIV proviral latency. Specific Aim 1: Comprehensively analyze small hairpin RNAs directed against lysine and arginine methyltransferases in J-Lat cells and primary cell culture models of HIV latency. We will perform a highly targeted shRNA screen of 11 known human arginine MTs, 24 known human lysine MTs and 38 new proteins identified based on domain homology with a conserved SET domain characteristic for lysine MTs to identify a set of MTs that enforce HIV latency as potential drug targets for l/E therapy. Specific Aim 2: Study the molecular mechanisms how methyltransferases control HIV latency. We will study the transcriptional activities of four suppressive MTs (PRMT1, PRMT6, Set1 and SETDB1) identified by our preliminary work. We will study the in wVo recruitment of these factors to the latent HIV LTR and identify substrates of these enzymes in latently infected T cells (histones. Tat, P-TEFb or NF-KB). Specific Aim 3: Test the effect of methyltransferase inhibitors on the reactivation of HIV transcription in latently infected T cells. We will evaluate the effect of known small molecule inhibitors of arginine and lysine MTs on HIV proviral latency in J-Lat cells and primary T cell models of latency. We will characterize the enzymes primarily targeted by these inhibitors and use this information for the development of effective and specific l/E of HIV, Collectively, these studies will provide novel molecular insight into the basic mechanisms governing HIV proviral latency and support the collective goal of this Collaboratory to produce compounds to be tested in the l/E of persistent HIV infection in animals and patients.