Finding, folding and characterizing the functions of disease-relevant RNA structures

项目摘要/摘要 了解疾病背后的基本分子细节是科学面临的一个关键挑战。这项建议 旨在通过关注疾病中一个未被充分研究的方面来解决这一问题：RNA结构的作用。这个 这个项目的总体目标是优化一条方法学管道，以发现 人类及其病原体的基因组，推断它们的结构，并表征它们的功能。结束 目的是更好地了解RNA结构如何与疾病有关，并利用这一点 以新的方式提供信息，为改善人类健康做出贡献：例如，通过推进基于RNA的 治疗性治疗。在这个项目的过程中，我们将通过关注感染来优化这条管道 EB病毒(EBV)对人B细胞的杀伤作用。EB病毒是一种广泛存在的人类病原体，其感染率超过95%。 在成人人群中，并与多种癌症和自身免疫性疾病有关。其作用机制 EBV致病性的背后仍然难以捉摸，这项提议旨在关注RNA和RNA的作用 感染和疾病的结构。在建立终身潜伏感染的过程中，EBV既产生 其自身的RNA和失调的各种人类RNA。核糖核酸在人类和病原体中起着中心作用 生物学，形成对基因表达必不可少的蛋白质编码和非蛋白质编码分子。 RNA结构发挥着重要的调节作用：例如，通过调节相互作用或改变 功能主题。因此，了解RNA的结构提供了大量的生物学知识，并且可以 用来从基因组中推断出功能序列。我之前的工作结合了计算和 发现EBV基因组中编码的RNA结构的实验方法。其中包括一本小说 病毒RNA的一类，是我发现的稳定的内含子序列(Sis)RNA。在目前的提案中，我们 跟进EBV编码的RNA的功能分析，并将我们的重点扩大到包括人类RNA。 这将通过使用我们开发的创新的新方法重新访问这两个基因组来实现 目前正在开发的新方法。这将与全基因组生物化学相结合 在体外(无细胞系统)和培养的人B细胞内进行结构分析和功能分析 细胞。这项提议的结果将是增强对RNA和RNA结构是如何的基本理解 与EB病毒感染和疾病有关。除了EBV及其相关的影响之外，还有更广泛的影响 病理学：例如，我们正在进行的一部分工作包括分析 病原体(可能找到保守的机制)。此外，许多人发现人类图案受到 EBV可能会在癌症和免疫中发挥作用；因此，这项提议将提供一般 对疾病的洞察。这些功能结构为新兴的RNA靶向提供了有吸引力的靶点 疗法；因此，这一提议在促进人类健康方面具有巨大的潜力：例如，在治疗 癌症和自身免疫性疾病。

PROJECT SUMMARY/ABSTRACT Understanding the basic molecular details behind disease is a critical challenge for science. This proposal aims to approach this by focusing on an understudied aspect of disease: the roles of RNA structure. The overall goal of this project is to optimize a methodological pipeline to discover functional RNA elements in the genomes of humans and their pathogens, deduce their structures, and characterize their functions. The end objective is to gain a better understanding of how RNA structure is implicated in disease and to use this information in novel ways to contribute toward improving human health: for example, by advancing RNA-based therapeutic treatments. Over the course of this project we will optimize this pipeline by focusing on the infection of human B cells by Epstein–Barr virus (EBV). EBV is a widespread human pathogen, which infects over 95% of the adult population, and is implicated in a variety of cancers and autoimmune diseases. The mechanisms behind EBV pathogenicity remain elusive and this proposal aims to focus on the roles of RNA and RNA structure in infection and disease. Over the course of establishing lifelong latent infection, EBV both generates its own RNAs and dysregulates various human RNAs. RNA plays a central role in human and pathogen biology, forming the protein-coding and non-protein-coding molecules that are essential to gene expression. RNA structure plays important regulatory roles: e.g. by mediating interactions or altering accessibility of functional motifs. Thus, knowing the structure of RNA provides a great deal of biological knowledge and can be used to deduce functional sequences from the genome. My previous work combined computational and experimental approaches to discover RNA structures encoded within the EBV genome. This included a novel class of viral RNAs, the stable intronic sequence (sis)RNAs, which I discovered. In this current proposal, we follow up on the functional analyses of EBV-encoded RNAs and expand our focus to include human RNAs. This will be accomplished by revisiting both genomes using innovative new approaches that we developed and novel methods that are currently under development. This will be combined with genome-wide biochemical structure analyses and functional assays performed in vitro (in cell-free systems) and within cultured human B cells. The results of this proposal will be an enhanced basic understanding of how RNA and RNA structure is involved in EBV infection and disease. There will be wider implications beyond EBV and its associated pathologies: for example, part of our pipeline involves the analysis of conservation of RNA structure between pathogens (potentially finding conserved mechanisms). As well, many discovered human motifs affected by EBV will likely have functional roles in cancer and immunity; therefore, this proposal will provide general insights into disease. These functional structures provide attractive targets for emerging RNA-targeting therapies; thus, there is great potential for this proposal to advance human health: e.g. in new treatments for cancer and autoimmune disease.