Role of EBV Lytic Infection in Viral Tumorigenesis

项目摘要/摘要 爱泼斯坦-巴尔病毒(Epstein-Barr Virus，EBV)是世界范围内导致人类癌症的重要原因，包括B细胞和 上皮细胞恶性肿瘤。尽管EBV编码的潜伏蛋白在驱动这些癌症中的作用是 被广泛接受的是，裂解病毒蛋白的重要性仍然相对有争议。尽管如此，Lytic 有效的EBV诱导的淋巴瘤可能需要体内EBV感染细胞亚群的感染 队形。我们之前已经证明了一个裂解缺陷的EBV突变体(缺失BZLF1(Z)-早期 (即，基因)在人源化小鼠模型中形成淋巴瘤的能力受到损害，该模型允许水平 病毒传播。此外，我们和其他人已经确定了生长因子和免疫抑制因子。 从裂解感染的B细胞中释放出来，可能会促进附近潜伏感染的B细胞的生长和存活 B细胞。此外，我们最近发现BZLF1启动子变异体(称为ZP-V3)是 在某些EBV阳性的恶性肿瘤(包括鼻咽癌和艾滋病相关淋巴瘤)中过度表达 其在非恶性组织中的出现频率增强了体外裂解病毒的激活，这是由于结合了 细胞NFAT转录因子到ZP-V3变异体(但不是原型启动子，ZP-P)。尽管 ZP-V3启动子形式在1型EBV毒株中相对罕见，在所有2型EBV毒株中都存在 (这在非洲的疟疾带很常见)。这些结果表明，增强型裂解性EBV感染 增加体内EBV诱导的淋巴瘤的可能性，以及一种特定的癌症相关变体 Z启动子可促进EB病毒感染的B细胞的裂解感染。在这个提案中，我们将使用两个不同的 人源化小鼠模型，比较含有ZP-P形式的EBV表型与癌症的表型- BZLF1启动子的相关(ZP-V3)形式，并探讨溶血性EB病毒感染的机制(S) 促进淋巴增生症。我们还将确定感染疟疾的人的EBV载量是否更高 儿童合并感染含EBV株ZP-V3与含ZP-P株的比较。我们的具体目标是 1)用人源化小鼠模型检测ZP-V3与ZP-P包含型的体内表型 1或2型EBV株；2)比较缺失的BALF5(病毒DNA聚合酶)与 人源化小鼠中BZLF1缺失的EBV突变体，并探讨用BZLF1阻断裂解性EBV DNA复制 抗病毒药物阿昔洛韦抑制EBV诱导的淋巴瘤的发展；以及3)确定 ZP-V3启动子变异是否与感染疟疾的非洲人血浆EBV水平升高有关 孩子们。我们假设EBV ZP-V3变异体将增强人源化EBV诱导的淋巴瘤 通过增加裂解性EBV感染，该变异也与增强的裂解性EBV相关 在感染疟疾的儿童中复制。如果是这样的话，这些结果将表明ZP-V3变体的存在 可能是一种有用的生物标志物，可用于预测EBV诱导人类恶性肿瘤风险的增加。

PROJECT SUMMARY / ABSTRACT Epstein-Barr virus (EBV) is an important cause of human cancers world-wide, including both B cell and epithelial cell malignancies. Although the role of EBV-encoded latency proteins in driving these cancers is well accepted, the importance of lytic viral proteins remains relatively controversial. Nevertheless, lytic infection in a subset of EBV-infected cells in vivo may be required for efficient EBV-induced lymphoma formation. We previously showed that a lytic-defective EBV mutant (missing the BZLF1 (Z) immediate-early (IE) gene) is impaired for the ability to form lymphomas in a humanized mouse model that allows horizontal virus transmission. In addition, we and others have identified growth factors and immunosuppressive factors released from lytically infected B cells that likely enhance the growth and survival of nearby latently infected B cells. Furthermore, we have recently discovered that a BZLF1 promoter variant (known as Zp-V3) that is over-represented in certain EBV-positive malignancies (including NPC and AIDS-related lymphomas) relative to its frequency in non-malignant tissues confers enhanced lytic viral reactivation in vitro due to binding of the cellular NFAT transcription factor to the Zp-V3 variant (but not the prototype promoter, Zp-P). Although the Zp-V3 form of the promoter is relatively uncommon in type 1 EBV strains, it is present in all type 2 strains (which are very common in the malaria belt of Africa). These results suggest that enhanced lytic EBV infection increases the likelihood of EBV-induced lymphomas in vivo, and that a particular cancer-associated variant of the Z promoter promotes lytic infection in EBV-infected B cells. In this proposal, we will use two different humanized mouse models to compare the phenotypes of EBV containing the Zp-P form versus the cancer- associated (Zp-V3) form of the BZLF1 promoter, and to explore mechanism(s) by which lytic EBV infection promotes lymphomagenesis. We will also determine whether EBV loads are higher in malaria-infected children co-infected with Zp-V3 containing EBV strains versus Zp-P containing strains. Our Specific Aims are 1) to use humanized mouse models to examine the in vivo phenotypes of Zp-V3 versus Zp-P containing type 1 or type 2 EBV strains; 2) to compare the phenotypes of BALF5 (the viral DNA polymerase)-deleted versus BZLF1-deleted EBV mutants in humanized mice, and explore whether blocking lytic EBV DNA replication with the antiviral drug, acyclovir, inhibits the development of EBV-induced lymphomas; and 3) to determine whether the Zp-V3 promoter variant is associated with higher plasma levels of EBV in malaria-infected African children. We hypothesize that the EBV Zp-V3 variant will enhance EBV-induced lymphomas in humanized mice by increasing lytic EBV infection, and that this variant is also associated with enhanced lytic EBV replication in malaria-infected children. If so, these results will suggest that the presence of the Zp-V3 variant may be a useful biomarker for predicting increased risk of EBV-induced malignancy in humans.