Understanding Epstein Barr virus oncogenicity through non-coding RNA structure

 描述（由申请方提供）：EB病毒（EBV）是一种疱疹病毒，可导致约95%的人类终身潜伏感染。在尚未完全了解的方式中，EBV潜伏期与伯基特和霍奇金淋巴瘤等癌症有关。EBV生物学的一个未充分研究的方面，可能有助于我们了解其致癌特性，是RNA结构所起的作用。我在Steitz实验室的研究预测，大约30%的EBV基因组产生具有生物学稳定和进化保守的RNA结构的RNA。这表明EBV转录组的相当一部分可能具有由非编码（nc）RNA结构介导的未知功能作用。同样，结构化的人类宿主转录物也可能在EBV感染和疾病中发挥关键作用。该项目将收集来自活的人类细胞的转录组范围的生化结构探测数据：未感染和感染EBV。这些数据将用于模拟潜伏和裂解病毒/宿主转录物中的RNA结构，并确定重要的RNA结构动力学与EBV感染和癌症的关系。该项目还将探索RNA结构用于治疗干预的可能靶向。在K99期，潜伏的EBV感染的人细胞的转录组将使用RNA结构敏感的细胞渗透性试剂进行化学探测。使用我的共同导师Philip贝维拉夸教授实验室开发的新技术，将使用RNA-Seq读取反应位点。这些数据将为EBV潜伏期的RNA结构提供前所未有的视角。我最近发现的两个高度保守、结构化和丰富的潜伏相关ncRNA的功能也将被研究。这些RNA是在高度致癌的潜伏期（潜伏期III）中产生的，这种分析可能有助于解释潜伏期III的致癌特性。这些努力为未来对EBV感染和癌症重要的病毒和宿主RNA结构的研究奠定了基础，我将成为我未来实验室的重点。在R 00阶段，我将在K99阶段开发的技能和方法将应用于研究EBV阴性和裂解再活化EBV感染细胞的转录组中的RNA结构。除了阐明裂解性EBV转录组中的RNA结构外，这项工作将允许在EBV阴性、潜伏和裂解性宿主和EBV转录组中存在的结构之间进行比较。这将确定病毒和宿主RNA中的潜在结构动力学，这可能对感染很重要。除了提供有关EBV的基本知识外，这项研究还对人类健康具有重要意义。更好地理解EBV感染中的RNA结构可能会识别出可以用治疗剂靶向的致癌RNA。例如，与Matthew迪士尼实验室合作，产生癌症相关病毒miRNA的前体miRNA发夹将被RNA结合小分子靶向以抑制miRNA成熟。通过推进RNA靶向小分子的使用，这项工作有可能产生旨在治疗EBV相关癌症的新疗法。

 DESCRIPTION (provided by applicant): Epstein-Barr virus (EBV) is a herpesvirus that causes lifelong latent infections in ~95% of humans. In ways not yet fully known, EBV latency is implicated in cancers such as Burkitt's and Hodgkin's lymphomas. An understudied aspect of EBV biology, that may help us to understand its oncogenic properties, is the role played by RNA structure. My research in the Steitz lab predicts that approximately 30% of the EBV genome generates RNA with thermodynamically stable and evolutionarily conserved RNA structure. This suggests that a substantial fraction of the EBV transcriptome may have unknown functional roles mediated by non-coding (nc)RNA structure. Likewise, structured human host transcripts may also play key roles in EBV infection and disease. This project will collect transcriptome-wide biochemical structure probing data from living human cells: uninfected and infected with EBV. These data will be used to model RNA structure in latent and lytic virus/host transcripts and to identify important RNA structural dynamics with implications for EBV infection and cancer. This project will also explore the possible targeting of RNA structure for therapeutic intervention. In the K99 phase, the transcriptome of latent EBV-infected human cells will be chemically probed using RNA structure-sensitive cell-permeable reagents. Using novel techniques developed in the lab of my co-mentor Prof. Philip Bevilacqua, reactive sites will be read out using RNA-Seq. These data will provide an unprecedented view of RNA structure in EBV latency. The functions of two highly conserved, structured, and abundant latency- associated ncRNAs, which I recently discovered, will also be investigated. These RNAs are produced during a highly oncogenic form of latency (latency III) and this analysis may help explain the cancer-causing properties of latency III. These endeavors lay the groundwork for future studies of virus and host RNA structures important to EBV infection and cancer, which I will make the focus of my future lab. In the R00 phase, the skills and methods I will develop in the K99 phase will be applied to study RNA structure in the transcriptomes of EBV-negative and lytically re-activated EBV-infected cells. In addition to shedding light on RNA structures in the lytic EBV transcriptome, this work will allow comparisons to be made between structures present in EBV-negative, latent, and lytic host and EBV transcriptomes. This will identify potential structural dynamics in viral and host RNAs, which may be important to infection. In addition to providing fundamentally important basic knowledge about EBV, this research also has significant implications to human health. A better understanding of RNA structure in EBV infection may identify oncogenic RNAs that can be targeted with therapeutics. For example, in collaboration with the Matthew Disney lab, precursor miRNA hairpins generating cancer-associated viral miRNAs will be targeted with RNA-binding small molecules to inhibit miRNA maturation. By advancing the use of RNA-targeting small molecules, this work has the potential to generate new therapeutics aimed at treating EBV-related cancers.