Recognition of Chromosomal DNA by Double-Stranded RNA

描述(申请人提供)：短干扰RNA(SiRNAs)识别信使核糖核酸的能力被广泛认可。SiRNA正在进入第一阶段试验，但它们最终影响人类健康的潜力尚不清楚。像任何候选药物一样，需要探索许多选择，以使双链RNA更有效、更多功能。我们观察到，在双链染色体DNA中，与启动子序列互补的21个碱基的抗基因RNA(AgRNAs)可以有效地抑制基因的表达。核连续分析表明agRNAs在转录水平上阻止了基因的表达。最近，我们发现了激活基因表达的agRNAs，为在实验室和临床使用双链RNAs开辟了新的途径。目的：本研究的目的是了解agRNAs的性质，阐明其作用机制，并表征其对基因沉默或激活的潜在作用。在目标1中，我们研究了ArgAerte蛋白在基因激活中的作用。在目标2中，我们研究了RNA介导的染色体DNA识别的机制，并开发了具有改进的效力和特异性的化学修饰的RNA。在目标3中，我们通过建立一个将RNA序列与沉默效力相关联的数据库来开发预测agRNA活性的规则。对于目标4，我们提出了计算和实验策略来识别与人类或病毒基因转录起始点相同的microRNAs。理论基础：我们提议的研究将描述agRNAs的机制。我们的实验将对生物医学研究产生重大影响，因为它们将1)将RNA介导的治疗扩大到染色体DNA内的靶点，2)制定使用agRNAs控制基因表达的规则，3)研究agRNAs在细胞调控基因表达的正常途径中的作用。我们的研究将极大地扩展开发RNA药物的潜力。与公共卫生相关：增加或减少基因表达的药物有可能被开发为治疗大多数疾病的药物。我们的工作是相关的，因为它描述了一种增加或减少与疾病有关的蛋白质水平的新策略。具体地说，我们的工作将扩大可用于设计RNA药物以减少基因表达的选择，并通过提供一种新的策略来设计RNA以增加基因表达来满足未得到满足的需求。

DESCRIPTION (provided by applicant): The ability of short interfering RNAs (siRNAs) to recognize mRNA is widely appreciated. siRNAs are entering Phase I trials, but their ultimate potential to impact human health is unclear. Like any drug candidate, many options will need to be explored to make duplex RNAs more potent and more versatile. We have observed that 21-base antigene RNAs (agRNAs) complementary to promoter sequences within duplex chromosomal DNA act as potent inhibitors of gene expression. Nuclear run-on assays demonstrate agRNAs block gene expression at the level of transcription. Most recently, we have discovered agRNAs that activate gene expression, opening up new avenues for using duplex RNAs in the laboratory and the clinic. Objectives: The objective of this proposal is to understand the properties of agRNAs, elucidate their mechanism of action, and characterize their potential for gene silencing or activation. In Aim 1, we investigate involvement of argonaute proteins in gene activation. In Aim 2, we examine the mechanism of RNA-mediated recognition of chromosomal DNA and develop chemically modified RNAs with improved potencies and specificites. In Aim 3, we develop rules for predicting agRNA activity by building a database correlating RNA sequence with silencing potency. For Aim 4, we propose computational and experimental strategies to identify microRNAs that share identity with transcription start sites for human or viral genes. Rationale: Our proposed research will characterize the mechanism of agRNAs. Our experiments will have a substantial impact on biomedical research because they will 1) expand the reach of RNA-mediated therapy to targets within chromosomal DNA, 2) develop rules for using agRNAs to control gene expression, and 3) examine the role of agRNAs in normal pathways for cellular regulation of gene expression. Our studies will significantly expand the potential for developing RNA drugs. Relevance to Public Health: Agents that increase or decrease gene expression have the potential to be developed as agents for treating most diseases. Our work is relevant becauses it describes a new strategy for increasing or decreasing the levels of proteins involved in disease. Specifically, our work will expand the options available for designing RNA drugs to reduce gene expression and fills an unmet need by providing a new strategy for designing RNAs to increase gene expression.