Structure and function of the Bovine Viral Diarrhea Virus 3' UTR during the virus life cycle.

牛病毒性腹泻病毒（Bovine viral diarrhea virus， BVDV）是影响养牛业的重要病原体，可通过急性感染和持续性感染（PI）。急性感染会减少雄性和雌性牛的繁殖，而PI小牛是感染的来源。急性感染的妊娠女性胎儿在子宫内感染BVDV导致PI小牛。BVDV的控制包括鉴定和扑杀PI小牛以及疫苗接种。然而，如果意外地给怀孕的雌性接种了改良的活疫苗，可能会导致PI小牛的出生。BVDV具有+链RNA基因组，其复制受RNA基因组结构和与其结合的蛋白质的调节。特别是5‘和3’端区域，即非翻译区（UTRs），包含调控病毒转译和复制的序列和结构。此外，少数病毒，如丙型肝炎病毒（HCV）和BVDV的utr也结合促进病毒生命周期的mirna，可能是通过调节它们结合的RNA的功能和改变该区域的RNA结构和/或蛋白质结合。我研究的长期目标是了解BVDV 5‘和3’ utr如何调节病毒复制，以便我们能够确定开发减毒病毒疫苗或控制病毒的细胞趋向性并预防胎儿感染的方法。在本提案中，我们的短期目标是分析BVDV 3 UTR在调节病毒生命周期中的作用。3' UTR是一种结构化的RNA元件，最近的研究表明，宿主细胞microrna miR-17和let-7可连接到3' UTR并促进BVDV的复制。我们有初步的数据表明，除了miR-17或let-7之外，同样粘附在3' UTR上的小rna也可以促进BVDV的复制。这表明，任何结合的小RNA都足以促进病毒复制，只要它与基因组的正确区域结合。我们的目标是确定miRNA退火可以促进BVDV复制的结构域边界。我们还将通过分析该区域发生点突变的病毒的生长来确定该区域在病毒生命周期中的作用。最后，我们将通过分析与该结构域结合的蛋白质，mirna的退火如何改变蛋白质结合，以及如何调节BVDV复制来确定该结构域在BVDV复制中的作用。这些研究将有助于更好地了解BVDV的生命周期以及3' UTR及其结合蛋白和rna的作用。这项工作还可能确定制造新的减毒疫苗的方法，以及限制疫苗复制到特定细胞时间的策略，从而避免疫苗传播到胎儿，并防止接种疫苗的奶牛产下PI小牛。

Bovine viral diarrhea virus (BVDV) is an important pathogen to the cattle industry, through both acute and persistent infections (PI). Acute infections decrease reproduction in both male and females cattle, and PI calves are sources of infection. PI calves result from in utero BVDV infection of the fetus in acutely infected pregnant females. Control of BVDV involves identification and culling of PI calves, and vaccination. However, vaccination with modified live vaccines can lead to the birth of PI calves if vaccines are accidently given to pregnant females. BVDV has a + stranded RNA genome and its replication is regulated by the structure of the RNA genome and protein that bind to it. In particular, the 5' and 3' end regions, the un-translated regions (UTRs), contain sequences and structures that regulate virus translation and replication. In addition, the UTRs of a few viruses, hepatitis C virus (HCV) and BVDV, also bind miRNAs that promote the virus life-cycle, possibly by modulating the functions of the RNA to which they bind and altering RNA structures and/or protein binding at that region. The long-term objective of my research is to understand how the BVDV 5' and 3' UTRs regulate virus replication so that we can identify ways to develop attenuated viral vaccines or control the cellular tropism of the virus and prevent infection of the fetus. In this proposal, our short-term objectives are to analyze the role of the BVDV 3 UTR in regulating the virus' life cycle. The 3' UTR is a structured RNA element and recent research showed that host cellular microRNAs, miR-17 and let-7, anneal to the 3' UTR and promote BVDV replication. We have preliminary data showing that small RNAs other than miR-17 or let-7 that also anneal to the 3' UTR can also promote BVDV replication. This suggests that any small RNA that binds is sufficient to promote virus replication, as long as it binds to the right region of the genome. Our goal is to identify the boundary of the domain to which miRNA annealing can promote BVDV replication. We will also determine the role of this domain in the virus' life cycle by analyzing growth of viruses having point mutations to this region. Finally we will identify the role of this domain in BVDV replication by analyzing the proteins that bind to it, how annealing of the miRNAs changes protein binding, and how this regulates BVDV replication. These studies will lead to a better understanding of the BVDV life cycle and the roles of the 3' UTR and its binding proteins and RNAs. This work may also identify ways to make new attenuated vaccines and strategies to limit vaccine replication to specific cell times and thus avoid vaccine spread to the fetus and prevent birth of PI calves from vaccinated pregnant cows.