Processing and Function of Polyoma RNA

描述(申请人提供)：这个项目的总体目标是了解多瘤病毒RNA是如何在受感染的细胞中制造和调节的，并将该病毒作为一个模型系统来学习哺乳动物RNA处理和功能的更多一般规则。我们发现，病毒基因表达的调控在很大程度上依赖于双链RNA的形成和细胞内dsRNA特异性腺苷脱氨酶(ADAR1)的编辑。在前一个资助时期完成的实验表明，病毒早-晚转换的长期寻找的触发因素可能是重叠，也可能是对早和晚多聚腺苷信号的编辑。这代表了一种新的基因调控机制。我们还了解了许多关于dsRNAs的核反应的新知识，包括一个长的核保留的非编码RNA(NEAT1)参与了被称为paraspeckles的核体的组装，它在保留编辑的RNA方面发挥着作用。我们将把这些研究扩展到三个相关的目标。首先，我们将使用新的方法来更好地表征病毒生命周期中的RNA表达，并将确定促进受调控基因表达的DNA序列元件的性质和相互作用。在第二个目标中，我们将更仔细地研究多瘤dsRNAs在细胞核中的命运，以及包括Paraspeckles在内的细胞dsRNA反应通路对感染的影响。在最终目标中，我们将进行研究，以了解更多关于反义RNA如何在哺乳动物细胞核中发挥作用的研究。这项工作将包括研究单个DNA分子是否可以在两个方向上同时转录，以及一种新的方法，使用小分子指导的异源二聚化来改变DNA和RNA分子的核内邻近性。

DESCRIPTION (provided by applicant): The overall goals of this project are to understand how polyoma virus RNAs are made and regulated in infected cells and to use this virus as a model system to learn more general rules for mammalian RNA processing and function. We have discovered that much regulation of viral gene expression depends on the formation of double-stranded RNAs and their editing by the cellular dsRNA specific adenosine deaminase (ADAR1). Experiments completed during the previous funding period suggested that the long-sought trigger for the viral early-late switch might be the overlap and perhaps also the editing of the early and late polyadenylation signals. This represents a new mechanism of gene regulation. We also learned much new about nuclear responses to dsRNAs, including the involvement of a long nuclear-retained noncoding RNA (NEAT1) in the assembly of nuclear bodies called paraspeckles, which function in the retention of edited RNAs. We will extend these studies in three related aims. In the first, we will use new methods to better characterize RNA expression in the viral life cycle and will determine the nature and interplay of DNA sequence elements that promote regulated gene expression. In the second aim we will examine more closely the fate of polyoma dsRNAs in the nucleus, and the impact of cellular dsRNA response pathways including paraspeckles on infection. In the final aim we will carry out studies to learn more about how antisense RNA works in mammalian nuclei. This work will involve studying whether single DNA molecules can allow concurrent transcription in both directions as well as a new approach using small molecule-directed heterodimerization to alter the intranuclear proximity of DNA and RNA molecules.