Exploiting Metabolism to Uncloak Epstein-Barr Virus Immunogens in Latently Infected B-cells

项目摘要 EB病毒（EBV）通过唾液传播，在口咽部建立 感染，侵入扁桃体并定植于B细胞室。经口传播的EBV是 传染性单核细胞增多症和多种B细胞和上皮癌的原因，包括 伯基特淋巴瘤（BL）和鼻咽癌。我们还需要了解 病毒基因组的表观遗传调节使其能够定殖口咽和扁桃体， 建立>95%成人的持续感染，每年导致200，000例癌症。扁桃体 在记忆B细胞中，EBV表达单一的免疫原性差的抗原。BL，可以呈现 与颌骨、面部和眼眶的肿瘤一样， 应答相比之下，EB病毒表达近80病毒蛋白在艾滋病患者口腔毛 舌白斑，以及多达8种免疫原性潜伏蛋白， 移植淋巴瘤。在最初的博士后阶段，申请人使用了两个CRISPR 筛选以表征对BL中EBV潜伏期重要的B细胞因子。这些提供了见解 限制B细胞中溶解周期和潜伏基因表达的表观遗传途径， 强调了启动和维持高水平DNA甲基化的表观遗传酶， EB病毒基因组在这种高度限制的潜伏形式。EB病毒DNA甲基化去干扰 抑制EBV抗原表达并使Burkitt细胞对CD 8 + T细胞识别敏感。然而， 关于受感染的B细胞代谢和表观遗传途径之间的相互作用知之甚少， 其必须发生以提供用于DNA和组蛋白修饰的甲基。申请人 假设潜伏性EBV感染的B细胞破坏甲硫氨酸和叶酸代谢 驱动免疫原性EBV沉默所必需的EBV基因组超甲基化的途径 基因产物和T细胞免疫逃避。在辅导K99阶段，申请人将1） 鉴定甲硫氨酸代谢如何促进EBV基因组超甲基化和沉默 在潜伏感染的B细胞和EBV癌蛋白，和2）表征之间的相互作用 叶酸循环和潜伏感染细胞中EBV基因组甲基化。在R 00阶段， 申请人将进行独立研究，以确定潜伏的EBV程序如何改变B细胞 在DNA和组蛋白水平上加强病毒基因组表观遗传状态的代谢途径 甲基化总的来说，这些研究将开辟EBV生物学的新领域，并有望 支持新的治疗方法。职业发展计划将为申请人做好准备 过渡到独立作为一个研究者与多学科的研究方法 病毒/宿主相互作用的代谢组学控制。

Project Summary Epstein-Barr virus (EBV) is spread through the saliva, where it establishes oropharyngeal infection, invades the tonsils and colonizes the B-cell compartment. Orally transmitted EBV is the cause of infectious mononucleosis and of multiple B-cell and epithelial cancers, including Burkitt lymphoma (BL) and nasopharyngeal carcinoma. Much remains to be learned about how epigenetic regulation of the viral genome enables it to colonize the oropharynx and tonsils to establish persistent infection of >95% adults and to cause 200,000 cancers per year. In tonsillar memory B-cells, EBV expresses a single, poorly immunogenic antigen. BL, which can present as tumors of the jaws, face and orbit, use a similar viral program to evade anti-EBV T-cell responses. By contrast, EBV expresses nearly 80 viral proteins in AIDS patients with oral hairy leukoplakia of the tongue, and as many as immunogenic eight latency proteins in post- transplant lymphoma. During the initial post-doctoral period, the applicant used two CRISPR screens to characterize B-cell factors important for EBV latency in BL. These provided insights into epigenetic pathways that restrict expression of lytic cycle and latency genes in B-cells and highlighted epigenetic enzymes that initiate and maintain a high level of DNA methylation of the EBV genome in this highly restricted form of latency. Perturbation of EBV DNA methylation de- represses EBV antigen expression and sensitizes Burkitt cells to CD8+ T-cell recognition. Yet, little is known about cross-talk between infected B-cell metabolism and epigenetic pathways, which must occur to supply methyl groups for DNA and histone modification. The applicant hypothesizes that latently EBV-infected B cells subvert methionine and folate metabolism pathways to drive EBV genome hypermethylation necessary for silencing of immunogenic EBV gene products and T-cell immunoevasion. During the mentored K99 phase, the applicant will 1) identify how methionine metabolism contributes to EBV genome hypermethylation and silencing in latently infected B-cells and EBV oncoproteins, and 2) characterize the interplay between the folate cycle and EBV genome methylation in latently-infected cells. During the R00 phase, the applicant will perform independent research to identify how latent EBV programs alter B-cell metabolic pathways to reinforce viral genome epigenetic states at the level of DNA and histone methylation. Collectively, these studies will open a new area of EBV biology and promise to support novel therapeutic approaches. The career development plan will prepare the applicant for transition to independence as an investigator with a multi-disciplinary approach to study metabolomic control of virus/host interactions.