The KSHV Ubiquitome: Developing a CURE (Course-based undergraduate research experience)

摘要 泛素和泛素样蛋白，如相扑-1和相扑-2/3，具有广泛的作用。 蛋白质的稳定性、定位、相互作用和整体功能。许多病毒已经进化出一种机制来 通过操纵泛素蛋白酶体系统影响细胞过程。卡波西肉瘤 疱疹病毒(KSHV)编码具有E3泛素连接酶活性和相扑连接酶活性的蛋白，并与 细胞泛素蛋白酶体系统，已报道对信号、凋亡、免疫反应和 细胞周期调节。KSHV是一种人类肿瘤病毒，与三种不同的癌症和两种炎症性疾病有关 综合症。潜伏期和裂解复制都在疾病的建立和维持中发挥作用，因此 了解调节这一转变的机制很重要。 我们的长期目标是识别和表征KSHV通过病毒靶向的细胞通路 泛素和类泛素修饰系统的操纵。这将有助于我们理解 KSHV感染的细胞生物学，并使进一步研究和最终干预的目标得以确定。 我们对Trex BCBL1细胞的泛素组以及表达该基因的细胞进行了比较分析 只有RTA一个人。我们已经在转RTA的细胞中的146个蛋白质中确定了193个差异泛素化位点， 在两个数据中，多西环素处理的Trex BCBL1细胞的206个蛋白质中有274个位点，98个蛋白质中有209个位点 集合，代表具有RTA诱导的泛素化改变的蛋白质。在这笔赠款的AIM1中，我们建议进行 在开发治疗药物的同时，对我们的发现进行验证和初步表征(课程- 基于本科生的研究经验)。在AIM 2中，我实验室的本科生和硕士生 将从裂解重新激活、基因组复制和感染病毒的角度评估感染细胞的结果 制作。这些研究将产生重大影响，因为我们将识别和表征小说 在本科生开设课程时病毒逃避和宿主细胞过程的操纵策略 为不同群体的学生提供研究体验。

ABSTRACT Ubiquitin and ubiquitin like (Ubl) proteins such as SUMO-1 and SUMO-2/3 have wide ranging effects on protein stability, localization, interactions, and overall function. Many viruses have evolved mechanisms to influence cellular processes through manipulation of the ubiquitin proteasome system. Kaposi’s sarcoma herpesvirus (KSHV) encodes proteins with E3 ubiquitin ligase activity, SUMO ligase activity and interacts with the cellular ubiquitin proteasome system, with reported effects on signaling, apoptosis, immune response and cell cycle regulation. KSHV is a human tumor virus associated with three different cancers and two inflammatory syndromes. Both latency and lytic replication play roles in establishing and maintaining disease, therefore it is important to understand the mechanisms that regulate this transition. Our long-term goal is to identify and characterize cellular pathways targeted by KSHV through viral manipulation of the ubiquitin and ubiquitin-like modification systems. This will contribute to our understanding of the cell biology of KSHV infection and enable identification of targets for further study and ultimately intervention. We have carried out a comparative analysis of the ubiquitome in TREx BCBL1 cells as well as cells expressing RTA alone. We have identified 193 differentially ubiquitinated sites in 146 proteins in cells transfected with RTA, 274 sites in 206 proteins in doxycycline treated TREx BCBL1 cells, and 209 sites in 98 proteins in both data sets, representing proteins with RTA induced ubiquitination alterations. In AIM1 of this grant, we propose to carry out validation and initial characterization of our findings in concert with the development of a CURE (course- based undergraduate research experience). In AIM 2, undergraduate and masters students in my laboratory will evaluate outcomes in infected cells in terms of lytic reactivation, genome replication and infectious virus production. These studies will have a significant impact as we will identify and characterize novel strategies of viral evasion and manipulation of host cellular processes while providing undergraduate research experiences to a diverse population of students.