VIRAL TRANSCRIPTION IN EBV TRANSFORMED B CELLS

这个子项目是利用由NIH/NCRR资助的中心拨款提供的资源的许多研究子项目之一。子项目和调查员(PI)可能从另一个NIH来源获得了主要资金，因此可能会出现在其他CRISE条目中。列出的机构是针对中心的，而不一定是针对调查员的机构。长期目标是了解EB病毒(EBV)在永生潜伏期内基因表达是如何调节的。EB病毒在被感染的宿主体内建立了一种终生感染，与地方性Burkitt？S淋巴瘤、鼻咽癌、30-50%的霍奇金？S病以及近一半发生在免疫抑制患者的淋巴瘤的发生密切相关。值得注意的是，EB病毒感染外周静息B细胞会导致生长转化，导致体外产生永生化的淋巴母细胞系。根据最近对血清阳性个体中EBV感染的分析，EBV驱动B细胞增殖和随后分化的能力似乎在病毒感染的B细胞的扩散和记忆B细胞(其中病毒基因表达非常有限)中建立长期潜伏库方面发挥了重要作用。了解EBV如何在永生潜伏期调节病毒基因的表达，最终可能为干扰病毒生命周期的这一阶段提供策略，这可能会干扰记忆B细胞室潜伏期的建立。我们继续重点分析在EBV永生潜伏期程序中参与调控EBNA基因表达的顺式元件的识别和特征。目的1.用包装缺陷和复制零的EBV产生永生化的棉顶绒猴LCL，作为非永生化EBV突变体的诱导库。目的2.EBV永生化淋巴母细胞系中具有顺式元件突变的EBV的产生和鉴定，这些顺式元件被认为参与调节EBNA基因转录。目的3.分析原代B细胞感染初期对WP活性的要求。目的4.B细胞永生化潜伏期建立和维持过程中WP甲基化的分析？甲基化在调节淋巴母细胞系WP活性中的作用。

This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. The subproject and investigator (PI) may have received primary funding from another NIH source, and thus could be represented in other CRISP entries. The institution listed is for the Center, which is not necessarily the institution for the investigator. The long term goal is to understand how Epstein-Barr virus (EBV) gene expression is regulated during immortalizing latency. EBV establishes a life-long infection within the infected host, and is closely associated with the development of endemic Burkitt?s lymphoma, nasopharyngeal carcinoma, 30-50% of Hodgkin?s disease, and nearly half of the lymphomas that arise in immunosuppressed patients. Notably, EBV infection of peripheral resting B cells results in growth transformation resulting, ex vivo, in the generation of immortalized lymphoblastoid cell lines. Based on recent analyses of EBV infection in seropositive individuals, it seems likely that the ability of EBV to drive B cell proliferation, and subsequent differentiation, plays an important role in the dissemination of virus infected B cells and the establishment of a long-lived latency reservoir in memory B cells (in which there is very limited viral gene expression). Understanding how EBV regulates viral gene expression during immortalizing latency may ultimately provide strategies for interfering with this phase of the virus life cycle, which could interfere with the establishment of latency in the memory B cell compartment. We continue to focus analyses on identifying and characterizing cis-elements involved in regulated EBNA gene expression during the immortalizing latency program of EBV. Aim 1. Generation of a cottontop marmoset LCL immortalized with a packaging defective and replication null EBV to serve as an inducible reservoir for non-immortalizing EBV mutants. Aim 2. Generation and characterization of EBV harboring mutations in cis-elements thought to be involved in regulating EBNA gene transcription in EBV immortalized lymphoblastoid cell lines. Aim 3. Analysis of the requirements for Wp activity during initial stages of infection of primary B cells. Aim 4. Analysis of Wp methylation during the establishment and maintenance of immortalizing latency in B cells ? role of methylation in regulating Wp activity in lymphoblastoid cell lines.