Systematic analysis of antiviral microRNA function

在这个提议中，我们研究了特定分子，microrna，如何限制病毒在宿主细胞中复制的能力。病毒（顾名思义）需要宿主蛋白才能进入细胞，在动物体内复制和传播；microrna调节这些蛋白的表达水平。我们之前已经证明，某些宿主microrna抑制多种疱疹病毒（巨细胞病毒、单纯疱疹病毒-1和小鼠γ疱疹病毒）以及一种不相关的单链RNA病毒semlikifforest病毒的复制。我们假设这些microrna的抗病毒特性是基于下调不同病毒所依赖的特定细胞蛋白。这项工作的一个广泛的长期目标是更好地理解microRNA机制（同时对多种宿主蛋白进行微妙的下调）如何用于治疗感染。这种针对宿主的疗法的优势在于，它有可能用于治疗一系列感染，并降低病毒容易发生突变而产生抗药性的可能性。在这里，我们采用尖端的生化技术来鉴定抗病毒microrna的靶标，并确定这些靶标如何抑制病毒复制。使用小鼠和人类巨细胞病毒作为模型系统，我们检查了小鼠和人类细胞（以及可能的其他动物宿主）中存在的mirna的靶标。同时，我们研究巨细胞病毒是否进化出了阻断宿主microRNA功能的机制。最终目标是确定特定抗病毒microrna的作用机制，并阐明将决定在治疗环境中使用这些分子所需的方法的因素（例如，通过多个microrna组合调节基因以及病毒干扰和耐药性的范围）。

In this proposal we examine how specific molecules, microRNAs, limit the capacity of viruses to replicate in host cells. Viruses (by definition) require host proteins to enter a cell, replicate and spread in an animal; microRNAs regulate the expression level of these proteins. We have previously demonstrated that certain host microRNAs suppress replication of multiple herpesviruses (cytomegalovirus, herpes simples virus-1 and mouse gammaherpesvirus) as well as an unrelated single stranded RNA virus, Semlikiforest virus. We hypothesize that the antiviral properties of these microRNAs are based on down-regulating specific cellular proteins that different viruses rely on. A broad, long-term goal in this work is to better understand how the microRNA mechanism (subtle down regulation of multiple host proteins simultaneously) could be used to treat infection. The advantage of this type of host-targeted therapy is that it could potentially be used to treat a range of infections and reduce the likelihood that viruses could readily mutate to become resistant. Here we implement cutting-edge biochemical techniques to identify the targets of antiviral microRNAs and determine how the targets suppress viral replication. Using murine and human cytomegalovirus as model systems, we examine targets of the miRNAs that are present in both mouse and human cells (and likely other animal hosts). In parallel, we examine whether cytomegaloviruses have evolved mechanisms for blocking host microRNA function. The end goal is to determine the mechanism of action of specific antiviral microRNAs and shed light on factors that will dictate the approaches required to use these molecules in a therapeutic context (e.g. combinatorial regulation of genes by multiple microRNAs and scope for viral interference and resistance).