Viral Noncoding RNAs and Cell Transformation

项目总结/摘要 非编码（nc）RNA在携带三种致癌疱疹病毒的淋巴细胞中的作用正在研究中。 研究了EB病毒（EBV）感染并转化人B细胞;它是EB病毒感染的病原体。 传染性单核细胞增多症，并与几种人类癌症有关。松鼠猴疱疹病毒（HVS）诱导 在新世界猴中的致命淋巴瘤和白血病，并在培养中转化人类T淋巴细胞。 卡波济肉瘤相关疱疹病毒（KSHV）折磨免疫功能低下的个体，并持续存在于一个 潜伏形式直到裂解活化。近年来，我们把工作重点放在了 两种EBV编码的EBER，七种HVS编码的HSUR和六种HVS microRNA，以及KSHV PAN RNA。这些病毒ncRNA都是丰富的，在相关病毒之间是保守的，并结合宿主蛋白， 形成ncRNP。我们的功能研究揭示了microRNA生物发生和衰变的新机制， 揭示了病毒ncRNP可能是病毒DNA复制（EBER 2）等多种核过程所必需的 或mRNA输出到细胞质（PAN），确定了三螺旋在RNA稳定化中的作用， 为病毒进化提供了重要的见解最引人注目的是，我们对病毒ncRNA的研究 发现并开始阐明新的细胞机制的存在，如细胞的调节， microRNA群体以及polyA尾和3′UTR如何合作稳定细胞mRNA。 提出的目标将利用这些进展进一步研究潜在的分子机制。我们 将扩展我们最初发现的靶向microRNA衰变（TDMD），以识别蛋白质， 另外的RNA信号有助于细胞microRNA降解，以及调查的作用， 来自SARS-CoV-2基因组的推定小RNA在调节宿主免疫应答中具有潜在的 诊断/治疗意义。我们将确定它的polyA尾以及内部序列 有助于PAN RNA的能力，使核出口的晚期裂解mRNA，导致病毒体蛋白 生产和病毒体释放。我们将搜索细胞转录组数据库， 序列/结构基序有助于polyA-3′UTR相互作用（以及可能的RNA稳定化），如 最近通过我们的高分辨率X射线分析揭示了这一点。新发现的三链体形成元素（ENEs） 将分析冠状病毒RNA的稳定活性和对病毒蛋白的可能贡献 合成，具有潜在的治疗应用。EBV中病毒转录本之间的广泛相互作用- 将验证并进一步分析通过peptien交联发现的感染细胞。

Project Summary/Abstract The roles of noncoding (nc)RNAs in lymphoid cells harboring each of three oncogenic herpesviruses are being investigated. Epstein-Barr virus (EBV) infects and transforms human B cells; it is the causative agent of infectious mononucleosis and is associated with several human cancers. Herpesvirus saimiri (HVS) induces fatal lymphomas and leukemias in New World monkeys and transforms human T lymphocytes in culture. Kaposi's sarcoma-associated herpesvirus (KSHV) afflicts immunocompromised individuals and persists in a latent form until lytic activation. In recent years, we have focused our efforts on the structure and functions of the two EBV-encoded EBERs, the seven HVS-encoded HSURs and six HVS microRNAs, as well as the KSHV PAN RNA. These viral ncRNAs are all abundant, conserved between related viruses and bind host proteins to form ncRNPs. Our functional studies have uncovered novel mechanisms of microRNA biogenesis and decay, revealed that viral ncRNPs can be essential for nuclear processes as diverse as viral DNA replication (EBER2) or mRNA export to the cytoplasm (PAN), identified the role of triple helices in RNA stabilization, and contributed important insights into viral evolution. Most compelling is that our studies of viral ncRNAs have uncovered the existence of and begun to elucidate novel cellular mechanisms such as the regulation of cellular microRNA populations and how the polyA tail and 3′UTR may collaborate to stabilize cellular mRNAs. Proposed aims will exploit these advances to further investigate the underlying molecular mechanisms. We shall extend our original discovery of target-directed microRNA decay (TDMD) to identify proteins and additional RNA signals contributing to cellular microRNA degradation, as well as investigate the role of a putative small RNA derived from the SARS-CoV-2 genome in regulating host immune responses, with potential diagnostic/therapeutic implications. We shall establish how its polyA tail as well as internal sequences contribute to PAN RNA's ability to enable the nuclear export of late lytic mRNAs, leading to virion protein production and virion release. We shall search cellular transcriptome databases for the presence of RNA sequence/structure motifs contributing to polyA-3′UTR interactions (and presumably RNA stabilization), as recently revealed by our high-resolution X-ray analyses. Newly discovered triplex-forming elements (ENEs) in coronavirus RNAs will be analyzed for their stabilization activity and possible contributions to viral protein synthesis, with potential therapeutic applications. Extensive interactions between viral transcripts in EBV- infected cells discovered by psoralen crosslinking will be validated and further analyzed.