IMPACT OF RNA INTERFERENCE ON QUASISPECIES EVOLUTION IN VECTOR-BORNE FLAVIVI

这个子项目是许多研究子项目中利用 资源由NIH/NCRR资助的中心拨款提供。子项目和 调查员(PI)可能从NIH的另一个来源获得了主要资金， 并因此可以在其他清晰的条目中表示。列出的机构是 该中心不一定是调查人员的机构。 由于它们的高突变率、大的种群规模和快速的复制速度，RNA病毒种群以突变基因组或“准种”的群体形式存在。准种的多样性影响病毒的适应性、疾病的严重性和对抗病毒治疗的反应。确定形成准物种多样性的机制对于更好地预防病毒出现和控制病毒疾病是必要的，特别是节肢动物传播的病毒，所有这些病毒都是RNA病毒，其中许多都是对全球公共卫生的重大新威胁。最近发现的RNA干扰(RNAi)是对与双链RNA触发器高度同源的RNA的靶向破坏，揭示了节肢动物调节病毒复制的第一条一般途径。这项拟议的研究将检验这样一种假设，即载体的RNAi反应对使病毒群体能够逃避RNAi的突变进行选择，从而推动病毒准种的多样化。为了验证RNAi活性下降应该导致准种多样性下降的预测，我们将感染假处理的果蝇细胞和RNAi已被蚊媒登革热病毒特异性下调的果蝇细胞。由此产生的病毒后代的多样性将使用焦磷酸测序来确定。随后将在活的果蝇身上进行类似的实验，方法是用登革热病毒感染缺乏RNAi途径特定成分的野生型果蝇或突变果蝇，并对产生的病毒后代进行测序。

This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. The subproject and investigator (PI) may have received primary funding from another NIH source, and thus could be represented in other CRISP entries. The institution listed is for the Center, which is not necessarily the institution for the investigator. Due to their high rates of mutation, large population sizes, and rapid rates of replication, RNA virus populations exist as a swarm of mutant genomes, or "quasispecies". The diversity of a quasispecies influences viral adaptation, disease severity, and response to antiviral therapies. Identifying the mechanisms that shape quasispecies diversity is necessary to better prevent virus emergence and control virus disease, particularly for arthropod-borne viruses, all of which are RNA viruses and many of which are significant emerging threats to global public health. The recent discovery of RNA interference (RNAi), the targeted destruction of RNA with high homology to a double-stranded RNA trigger, has revealed the first general pathway by which arthropods regulate viral replication. The proposed research will test the hypothesis that the RNAi response of vectors imposes selection for mutations that enable virus populations to escape RNAi, and RNAi thereby drives diversification of viral quasispecies. To test the prediction that decreasing RNAi activity should result in decreased quasispecies diversity, we will infect sham-treated Drosophila cells and Drosophila cells in which RNAi has been specifically downregulated with mosquito-borne dengue virus. Diversity of the resulting virus progeny will be determined using pyrosequencing. A similar experiment will then be conducted in live Drosophila by infecting wild type flies or mutant flies that lack specific components of the RNAi pathway with dengue virus and sequencing the resulting viral progeny.