EFFECT OF LYTIC EBV REPLICATION PROTEINS ON THE VIRUS

EB病毒（EBV）与B细胞以及上皮细胞恶性肿瘤有关。像所有疱疹病毒一样，EBV以潜伏或裂解形式感染细胞。裂解形式的感染是病毒从宿主到宿主和细胞到细胞传播所必需的。在潜伏感染的细胞中表达两种病毒立即早期（IE）蛋白BZLF1（Z）和BRLF1（R）中的任何一种都足以诱导裂解型感染，其中病毒使用其裂解型复制起点oriLyt进行复制。在这笔拨款中，我们建议继续进行长期研究， EB病毒IE蛋白以及BMRF 1早期蛋白对于溶解性EB病毒复制的重要性。在之前的资助期间，我们发现BMRF 1早期蛋白不仅作为病毒DNA聚合酶辅助蛋白，而且作为激活oriLyt启动子（BHLF 1）以及细胞胃泌素启动子的转录因子。此外，我们最近发现IE Z蛋白具有独特的优先结合能力， 当某些EBV靶启动子处于甲基化形式时，激活这些启动子。我们还表明，Z分散核PML（ND10体），可能因此降低宿主细胞中某些DNA修复蛋白的水平。我们提出以下具体目标。在具体目标#1中，我们将剖析BMRF 1蛋白不仅作为病毒DNA聚合酶辅助因子，而且作为病毒DNA聚合酶辅助因子发挥作用的机制。 转录因子在具体目标#2中，我们将确定oriLyt的DNA甲基化是否影响其复制或转录，反之亦然，如果已知调节oriLyt转录的病毒蛋白（Z，R和BMRF 1）影响oriLyt甲基化。在具体目标#3中，我们将确定一种或多种裂解病毒蛋白（特别是Z、R或BMRF 1）是否抑制细胞DNA修复机制，以及这种抑制细胞DNA修复的能力对于保护病毒末端在裂解形式的感染期间免于多联体化是否重要。这些研究 应该加强我们对Z，R和BMRF1蛋白不仅作为病毒转录因子，而且作为重要的病毒复制蛋白的功能的理解。

Epstein-Barr virus (EBV) is associated with B-cell as well as epithelial cell malignancies. Like all herpesviruses, EBV infects cells in either a latent or lytic form. The lytic form of infection is required for transmission of the virus from host to host, and cell to cell. Expression of either one of the two viral immediate-early (IE) proteins, BZLF1 (Z) and BRLF1(R), in latently infected cells is sufficient to induce the lytic form of infection, in which the virus replicates using its lytic origin of replication, oriLyt. In this grant, we propose to continue our long-term studies regarding the importance of the EBV IE proteins, as well as the BMRF1 early protein, for lytic EBV replication. During the previous period of funding, we showed that the BMRF1 early protein functions not only as the viral DNA polymerase accessory protein, but also as a transcription factor that activates an oriLyt promoter (BHLF1) as well as the cellular gastrin promoter. In addition, we recently discovered that the IE Z protein is uniquely capable of preferentially binding to, and activating, certain EBV target promoters when these promoters are in the methylated form. We have also shown that Z disperses nuclear PML (ND10 bodies), and perhaps as a consequence of this, reduces the level of certain DNA repair proteins in the host cell. We propose the following specific aims. In Specific aim #1, we will dissect the mechanisms by which the BMRF1 protein functions not only as the viral DNA polymerase accessory factor, but also as a transcription factor. In Specific aim #2, we will determine whether DNA methylation of oriLyt affects either its replication, or its transcription, and vice versa if the viral proteins known to regulate oriLyt transcription (Z, R, and BMRF1) affect oriLyt methylation. In Specific Aim #3, we will determine if one or more lytic viral proteins (particularly Z, R or BMRF1) inhibit the cellular DNA repair machinery, and if this ability to inhibit cellular DNA repair is important for protecting the viral ends from concatamerization during the lytic form of infection. These studies should enhance our understanding of how the Z, R and BMRF1 proteins function not only as viral transcription factors, but also as essential viral replication proteins.