REGULATION OF REPLICATION AND LATENCY BY EBV EBNAS

爱泼斯坦-巴尔病毒(EBV)，一种人类疱疹病毒，建立了潜伏的 B淋巴细胞感染。这些细胞被病毒永生化， 能够在培养中持续增殖。EBV有 传统上与两种恶性疾病有关，伯基特氏症 淋巴瘤和鼻咽癌。最近，人们对EBV的认识 数据增加了导致淋巴增生性疾病的可能性 更清楚地将EBV与霍奇金淋巴瘤的子集联系起来 器官移植受者EB病毒相关性淋巴瘤的发生 和艾滋病患者。 只有有限数量的EBV基因在潜伏感染的B细胞中表达 细胞。本提案中描述的研究集中在两个方面的作用 其中，EBNA-L和EBNA-2，在建立和维护 潜伏期与B细胞永生化生长的建立 表型。EBNA-L在功能上是多功能的，有助于 对其自身表达的调节，并成为唯一需要的病毒蛋白 对于通过延迟源的复制，ORI-P。EBNA-ls函数为 通过直接的DNA结合来调节。然而，通过这些机制 EBNA-L激活转录，OrI-P复制仍未解决。 建议(L)进一步表征DNA识别和 突变的EBNA-L结合所需的二聚化结构域 分析。(2)探讨EBNA-L反式激活的机制 识别转录激活结构域(S)。(3)审查 EBNA-L诱导的结构改变及对OrI-P复制的贡献 确定EBNA在复制和永生化中的作用-L 与细胞蛋白质的相互作用。对EBNA-ls捐款的评估(如果 Any)与R细胞永生化有关，因为EBNA-L/ORI-P 组合被吹捧为人类的非整合载体系统 基因疗法。 EBNA-2是EBV诱导永生化所必需的。EBNA-2上调表达 EBNA和LMP家族潜伏期基因的转录以及 改变特定B细胞基因的表达。建议(4) 描述反式激活所需的EBNA-2结构域和 永生，包括与细胞相互作用的领域 靶向蛋白CBF1；与细胞相互作用的结构域 转录复合体，以及目前未知的功能域 含有进化上保守的氨基酸基序。在最后的细节中 目的(5)靶向EBNA-2反应启动子的细胞CBF1蛋白 将CBF1的特性及其在未感染细胞中的作用 检查过了。

Epstein-Barr virus (EBV), a human herpesvirus, establishes a latent infection in B lymphocytes. These cells are immortalized by the virus and become capable of continuous proliferation in culture. EBV has traditionally been associated with two malignant diseases, Burkitt's lymphoma and nasopharyngeal carcinoma. Recently, awareness of EBVs potential to cause lymphoproliferative disease has been heightened by data more clearly associating EBV with a subset of Hodgkin's lymphomas and by the development of EBV associated lymphomas in organ transplant recipients and AIDS patients. Only a limited number of EBV genes are expressed in latently infected B cells. The research described in this proposal focuses on the role of two of them, EBNA-l and EBNA-2, in the establishment and maintenance of latency and in the establishment of the immortalized B cell growth phenotype. EBNA-l is functionally pleiotropic, contributing to the regulation of its own expression and being the sole viral protein required for replication via the latency origin, ori-P. EBNA-ls function is mediated through direct DNA binding. However the mechanisms by which EBNA-l activates transcription and ori-P replication remain unresolved. It is proposed (l) To further characterize the DNA recognition and dimerization domains that are required for EBNA-l binding by mutational analysis. (2) To determine the mechanism of EBNA-l transactivation by identifying a transcriptional activation domain(s). (3) To examine the contribution to ori-P replication of EBNA-l induced structural changes and determine the role in replication and immortalization of EBNA-l interaction with cell proteins. Assessment of EBNA-ls contribution (if any) to R cell immortalization is pertinent since the EBNA-l/ori-P combination is being touted as a non-integrating vector system for human gene therapy. EBNA-2 is essential for EBV induced immortalization. EBNA-2 up-regulates transcription of both the EBNA and LMP families of latency genes and also alters expression of specific B cell genes. It is proposed to (4) Characterize the domains of EBNA-2 required for transactivation and immortalization including the domain for interaction with the cellular targeting protein, CBF1; the domain for interaction with the cell transcription complex, and the domains of presently unknown function that contain evolutionarily conserved amino acid motifs. In the final specific aim (5) the cell CBF1 protein that targets EBNA-2 to responsive promoters will be characterized and the role of CBF1 in the uninfected cell examined.