EBV TRANSFORMATION OF B LYMPHOCYTES--EBNA REGULATION

描述：爱泼斯坦-巴尔病毒与伯基特密切相关 淋巴瘤和鼻咽癌，并对 免疫受损个体中的淋巴增生性疾病。长期的 拟议研究的目标是确定 EBV诱导的细胞转化。当这种病毒在体外感染B细胞时， 淋巴细胞变得永生。EB病毒W启动子(WP)的激活 是在感染过程中检测到的第一个病毒转录事件。可湿性粉剂 指导EB病毒核抗原(EBNA)的早期表达。 EBNA蛋白参与细胞和病毒的基因调控， 维持慢性EBV潜伏期。在后来的感染中，有一个开关 到其他EBNA启动子的使用，CP，然后QP。而CP和QP已经被 经过广泛研究，几乎没有关于特定CI的信息 和WP调节的反式成分。我们的初步研究已经 证明在W中至少存在三个顺式元件 启动子，因此这些元素被选为本研究的详细研究对象 求婚。 在第一个特定目标中，特定的细胞转录因子 将描述与可湿性粉剂中的关键顺式元件相互作用的元素。 广泛的启动子突变、凝胶移动分析、启动子足迹和 西南方向的凝胶体将被使用。第二个具体问题 目的探讨可湿性粉剂顺式元件的功能重要性。这将是 通过使用表达载体完成的转染性研究 相关反式激活剂和使用各种WP缺失和突变 构造。在第三个目的中，可湿性粉剂的体内足迹，以及 具有关键顺式元件中断的整个EBV突变体将用于 论证了WP结构域的体内意义。最后，这些研究 将确定作用于TRANS中WP的因素在体内的作用 EBNA启动子转换的时间进程。从这些项目中获得的知识 研究可能最终产生利用EBV基因控制的方法 表达增强EBV感染的恶性细胞的免疫破坏作用 像伯基特淋巴瘤或霍奇金氏病这样的灾难性疾病。

DESCRIPTION: Epstein-Barr virus is strongly associated with Burkitt lymphoma and nasopharyngeal carcinoma and is responsible for lymphoproliferative disease in immunocompromised individuals. The long-term objectives of the studies proposed are to determine the mechanisms of cellular transformation by EBV. When this virus infects B cells in vitro, the lymphocytes become immortalized. Activation of the EBV W promoter (Wp) is the first viral transcriptional event detected during infection. Wp directs the early expression of Epstein-Barr virus nuclear antigens (EBNA). The EBNA proteins are involved in cellular and viral gene regulation and the maintenance of chronic EBV latency. Later in infection, there is a switch to usage of the other EBNA promoters, Cp, then Qp. While Cp and Qp have been studied extensively, almost no information exists regarding the specific cis and trans constituents of Wp regulation. Our preliminary studies have demonstrated the presence of at least three cis elements within the W promoter, so these elements have been selected for detailed study in this proposal. In the first specific aim, the particular cellular transcription factors which interact with critical cis elements in Wp will be characterized. Extensive promoter mutagenesis, gel shift assays, promoter footprinting and southwestern gels of bound factors will be employed. The second specific aim will explore the functional importance of Wp cis elements. This will be accomplished with transfection studies using expression vectors for pertinent transactivators and using various Wp deletional and mutational constructs. In the third aim, in vivo footprinting of Wp, and the use of whole EBV mutants with disruption of key cis elements will be used to demonstrate the in vivo significance of Wp domains. Finally, these studies will ascertain the in vivo role of factors acting upon Wp in trans during the time course of EBNA promoter switching. Knowledge gained from these studies might ultimately produce means of exploiting the control of EBV gene expression to augment immune destruction of EBV-infected malignant cells in such catastrophic diseases as Burkitt lymphoma or Hodgkin's disease.