Deciphering the complex mechanisms that reprogram gene expression and promote Epstein Barr Virus replication.

EB病毒(Epstein-Barr Virus，EBV)是一种重要的人类病原体，与癌症和其他疾病(鼻咽癌、B细胞淋巴瘤、霍奇金病、淋巴增生性疾病和传染性单核细胞增多症)有关。EBV的繁殖对于产生病毒颗粒至关重要，这些病毒颗粒允许EBV感染细胞并在个体之间传播。EB病毒感染血液中的抗体产生细胞会导致细胞的异常持续生长。大多数情况下，病毒隐藏在这些细胞中，不会复制。最近，病毒的复制与其使这些细胞持续生长的能力有关。一种病毒基因被戏称为“裂解EBV复制的主要调节因子”。该基因产生一种名为Zta的蛋白质，这是复制病毒基因组所必需的。Zta通过直接与基因组中的DNA结合来改变病毒和人类基因的表达。它识别一个非常短的DNA区域，只包含7个DNA碱基对-在2041个潜在的组合中，有32个这种密码的功能变体，每个被称为ZRE。Zta增加了一些基因的表达，但降低了其他基因的表达。Zta增加病毒基因表达的能力是其促进病毒复制能力的关键，而允许Zta增加基因表达的分子也得到了部分了解。相比之下，人们对Zta导致基因表达下降的机制知之甚少。已知的两个被Zta下调的细胞基因通常需要免疫系统来发挥作用，因此Zta可能允许EBV避免免疫反应的破坏。在这里，我们旨在发现决定基因表达是否被Zta上调或下调的分子机制。(I)我们将使用最先进的技术(芯片-序列)来确定人类基因组中Zta的结合位置。我们还将确定Zta结合位点附近的基因是否增加或减少了基因表达。我们还将研究与Zta附近基因结合的其他蛋白质，并将研究Zta结合部位的DNA是否甲基化。然后，我们可以选择有代表性的人类基因进行进一步的研究，以确定哪些邻近的DNA元件导致Zta与ZRE相互作用，从而增加或减少基因表达。(Ii)我们将利用蛋白质组学策略，探索Zta介导的基因表达的增加或减少是否受到Zta上附着的小蛋白分子(SUMO)的影响，还是通过Zta与其他宿主蛋白质的结合来影响。(Iii)该项目将确定在病毒复制过程中受Zta调控的宿主基因，因此很可能突出用于为病毒复制重新编程人类细胞的进一步机制。这将增加我们对EBV复制的基本过程的理解，并有可能将这些作为未来药物发现的目标。

Epstein-Barr virus (EBV) is an important human pathogen associated with cancer and other diseases (nasopharyngeal carcinoma, B-cell-lymphomas, Hodgkin's disease, lymphoproliferative diseases and infectious mononucleosis). Multiplication of EBV is essential to produce the virus particles that allow EBV to infect cells and spread between individuals. Infection of antibody-producing cells in the blood with EBV results in abnormal continual growth of the cells. Most of the time the virus hides in these cells, without replicating. Recently, the replication of the virus has been linked with its ability to cause these cells to grow continuously.One viral gene has been nicknamed "The master regulator of lytic EBV replication". The gene generates a protein, known as Zta, which is required for replication of the viral genome. Zta alters the expression of viral and human genes by binding directly to DNA in the genomes. It recognises a very short region of DNA, containing only 7 DNA base-pairs - of 2041 potential combinations, there are 32 functional variations of this code and each is called a ZRE. Zta increases the expression of some genes but decreases the expression of others. The ability of Zta to increase the expression of viral genes is key to its ability to promote viral replication and the molecules that allow Zta to increase gene expression are partly understood. In contrast little is known of the mechanism of Zta causing decreased gene expression. Two cellular genes known to be down-regulated by Zta are normally required by the immune system to function, so Zta may allow EBV to avoid destruction by the immune response.Here, we aim to discover the molecular mechanisms that determine whether the expression of a gene is increased or decreased by Zta.(i) We will employ state-of-the-art techniques (ChIP-Seq) to identify binding sites for Zta in the human genome. We will also determine whether genes near the Zta binding sites have increased or decreased gene expression. We will also investigate the other proteins that bind to genes near Zta and will investigate whether the DNA at Zta-binding sites is methylated or not. We can then select representative human genes to investigate further to identify which neighbouring DNA elements cause Zta interaction with a ZRE to either increase or decrease gene expression. (ii) We will explore whether Zta-mediated increases or decreases in gene expression are influenced by the attachment of a small protein molecule (SUMO) to Zta or through Zta binding to other host proteins using a proteomics strategy. (iii) This project will identify those host genes that are regulated by Zta during viral replication and so is likely to highlight further mechanisms used to reprogram human cells for viral replication. This will increase our understanding of the basic process of EBV replication and potentially these can be explored as targets for future drug discovery.