The Function of EBV LMP1 CTAR3 in Sumoylation and Oncogenesis

项目摘要： 候选人和环境：我的职业目标是成为一名领先的独立科学家 疱疹病毒学。在独立之路奖的指导阶段，我将努力进一步 建立我在EBV领域的出版记录，为我将作为教员进行的研究奠定基础 成员。该项目将提供新技术方面的培训[BAC技术，人性化的老鼠模型 系统，研究蛋白质-蛋白质和蛋白质-DNA相互作用]将适用于这项研究 以独立调查员的身份执行。帕加诺博士和其他教职员工在 北卡罗来纳大学将在学习成为一名成功的独立研究人员所需技能方面发挥不可估量的作用。 研究：EBV是一种普遍存在的疱疹病毒，与几种淋巴系和上皮性恶性肿瘤有关。 潜伏膜蛋白-1(LMP1)是病毒的主要癌蛋白，是一种具有结构性活性的膜蛋白 通过胞浆C末端激活调节多条信号转导途径的信号蛋白 地区(Ctar)。与CTAR1和CTAR2相比，CTAR3的功能没有很好地定义。我最近 发表了LMP1 CTAR3是LMP1与相扑共轭相互作用的必要条件和充分条件 Ubc9酶，并诱导细胞蛋白质的总摩化。LMP1-Ubc9相互作用中断 与肿瘤发生和LMP1相关的细胞行为改变以及LMP1诱导的LMP1缺失 相扑化。进一步了解LMP1 CTAR3的功能和工作机制是我的 下一个优先事项。我假设CTAR3是一个自我调节的结构域，帮助调节致癌 LMP1的潜力。在将在指导阶段完成的具体目标1中，我将确定生物学 LMP1 CTAR3在体内的功能用野生型和LMP1CTAR3缺失病毒感染人源化小鼠， 在LCCC开发的一种新的EBV感染和疾病动物模型，我将检查肿瘤的形成和 病毒的免疫原性。CTAR3的作用是LMP1在细胞内的转运和稳定 也将接受测试。在将在独立阶段完成的具体目标2中，我将审查 CTAR3发挥作用的机制。LMP1诱导相思甲基化的初始靶点包括IRF7、STAT1和 相扑蛋白酶SENP1。我将研究LMP1、CTAR3在这些蛋白的苏莫化中的作用，并 苏莫化如何调节蛋白质功能。最后，因为蛋白质和甲基化可以帮助抑制潜伏裂解 开关，我将研究在LMP1 CTAR3有助于LMP1介导的抑制裂解复制通过诱导 蛋白质总甲基化，特别是PML、ZEB1和ZEB2。拟议的工作将大大增加我们的 了解LMP1 CTAR3的功能，特别是它在调节蛋白质总摩化中的作用，以及如何 CTAR3发挥作用，扩大了对LMP1如何作为癌蛋白的了解。

Project Abstract: Candidate and Environment: My career goal is to become a leading independent scientist in the field of herpesvirology. During the mentored phase of the Pathway to Independence Award, I will work to further establish my publication record in the field of EBV, laying the groundwork for the research I will do as a faculty member. This project will provide training in new techniques [BAC technology, a humanized mouse model system, investigating protein-protein and protein-DNA interactions] that will be applicable to the research performed as an independent investigator. The expertise of Dr. Pagano as well as other faculty members at UNC will be invaluable in learning the necessary skills to become a successful independent researcher. Research: EBV, a ubiquitous ¿-herpesvirus, is associated with several lymphoid and epithelial malignancies. Latent membrane protein-1 (LMP1), the principal viral oncoprotein, is a constitutively active membrane signaling protein that regulates multiple signal transduction pathways via its cytoplasmic C-terminal activating regions (CTAR). In contrast to CTAR1 and CTAR2, the function for CTAR3 is not well defined. I recently published that LMP1 CTAR3 is necessary and sufficient for LMP1 to interact with the SUMO-conjugating enzyme Ubc9 and induce the sumoylation of cellular proteins. Disruption of the LMP1-Ubc9 interaction resulted in changes in cellular behaviors associated with oncogenesis and LMP1 and loss of LMP1-induced sumoylation. Further understanding how LMP1 CTAR3 functions and mechanisms by which it functions is my next priority. I hypothesize that CTAR3 is a self-regulatory domain that helps modulate the oncogenic potential of LMP1. In Specific Aim 1, to be completed during the mentored phase, I will determine biological functions of LMP1 CTAR3 in vivo. Using wildtype and LMP1 CTAR3 deletion viruses to infect humanized mice, a new animal model developed at LCCC for EBV infection and disease, I will examine the tumorigenic and immunogenic potentials of the viruses. The role of CTAR3 is the intracellular trafficking and stability of LMP1 will also be tested. In Specific Aim 2, to be completed during the independent phase, I will examine mechanisms by which CTAR3 functions. Initial targets of LMP1-induced sumoylation include IRF7, STAT1, and the SUMO protease SENP1. I will investigate the role of LMP1 CTAR3 in the sumoylation of these proteins and how sumoylation regulates protein function. Finally, because protein sumoylation can help inhibit the latent-lytic switch, I will examine in LMP1 CTAR3 contributes to LMP1-mediated repression of lytic replication by inducing protein sumoylation, specifically of PML, ZEB1, and ZEB2. The proposed work will greatly increase our understanding of functions for LMP1 CTAR3, specifically its role in regulating protein sumoylation, and how CTAR3 functions, expanding knowledge of how LMP1 acts as an oncoprotein.