Project 1, Miller: Novel Functions of the Epstein-Barr Viral Lytic Cycle Activator Protein ZEBRA

项目1，摘要/摘要 EB病毒（EBV）是一种与全球重要的癌症相关的人类致癌病毒， 如胃癌和鼻咽癌，几种淋巴瘤，包括伯基特，霍奇金和弥漫性大细胞 淋巴瘤发生在免疫活性个体中，多克隆淋巴瘤发生在免疫缺陷宿主中。 我们的实验室专注于EBV裂解周期，这是细胞间病毒传播所必需的， 并直接导致病毒致癌性。EB病毒裂解周期由以下启动 两种病毒蛋白质ZEBRA和Rta的表达，这两种转录因子也在以下方面发挥不同的直接作用： 裂解病毒DNA复制。我们提出的工作是基于新认识的ZEBRA，主要功能 裂解性周期激活蛋白最初是在我们的实验室发现的。在AIM 1中，与Steitz合作， 实验室，我们将探索ZEBRA在介导病毒宿主关闭细胞中的新作用的机制。 细胞基因表达我们将研究ZEBRA如何控制胞质多聚腺苷酸的易位 结合蛋白（PABPC）的细胞核，ZEBRA如何选择性地分配PABPC在细胞核内， ZEBRA在细胞核中保留多聚腺苷酸化的mRNA，以及ZEBRA如何选择性地抑制多聚腺苷酸化的mRNA的合成。 细胞蛋白而不是病毒蛋白。在AIM 2中，我们将与勇雄实验室一起研究结构基础， ZEBRA对甲基化DNA的优先识别。我们将研究甲基化单倍型， ZEBRA靶向的病毒和细胞基因的启动子。我们将揭示甲基化的 病毒和细胞DNA中的ZEBRA反应元件以及ZEBRA蛋白本身介导的特性 甲基化DNA的优先识别。这些研究最终将产生ZEBRA的晶体结构 与甲基化DNA结合。我们提出的实验将阐明控制两种 以前未知的主要EBV裂解性周期激活蛋白的功能，即病毒宿主关闭和 通过识别甲基化DNA的转录激活。这些建议的研究具有特别的 与EBV阳性胃癌的相关性，其中EBV基因组的存在诱导了一种 细胞基因超甲基化

Project 1, Summary/Abstract Epstein-Barr virus (EBV) is a human oncogenic virus associated with carcinomas of global importance, such as gastric and nasopharyngeal cancer, several lymphomas, including Burkitt, Hodgkin and diffuse large cell lymphoma that occur in immunocompetent individuals and polyclonal lymphomas in immunodeficient hosts. Our laboratory focuses on the EBV lytic cycle that is essential for transmission of virus among cells and between individuals and directly contributes to viral oncogenicity. The EBV lytic cycle is initiated by expression of two viral proteins, ZEBRA and Rta, transcription factors that also play distinct direct roles in lytic viral DNA replication. Our proposed work is based on newly recognized functions of ZEBRA, the major lytic cycle activator protein originally discovered in our laboratory. In AIM1, in collaboration with the Steitz lab, we will explore mechanisms underlying the novel role of ZEBRA in mediating viral host shut off of cellular gene expression. We will investigate how ZEBRA controls the translocation of cytoplasmic poly A binding protein (PABPC) to the nucleus, how ZEBRA selectively distributes PABPC within the nucleus, how ZEBRA retains poly-adenylated mRNA in the nucleus and how ZEBRA selectively inhibits synthesis of cellular but not viral proteins. In AIM2, together with the lab of Yong Xiong, we will study the structural basis of preferential recognition of methylated DNA by ZEBRA. We will investigate the methylation haplotype of promoters of viral and cellular genes targeted by ZEBRA. We will uncover features of the methylated ZEBRA response elements in viral and cellular DNA and properties of the ZEBRA protein itself that mediate preferential recognition of methylated DNA. These studies will eventually yield a crystal structure of ZEBRA bound to methylated DNA. Our proposed experiments will elucidate molecular mechansims that govern two previously unknown functions of the major EBV lytic cycle activator protein, namely viral host shutoff and transcriptional activation via recognition of methylated DNA. These proposed studies have particular relevance for EBV-positive gastric cancer where the presence of the EBV genome induces a state of cellular gene hypermethylation.