Molecular Mechanism of Epstein-Barr virus DNA replication in viral infected tumors and cellular immunity for viral associated cancers

EBV编码7个病毒基因，这些基因对orilyt依赖性DNA复制至关重要。BZLF1蛋白是一种orilyt结合蛋白，也是一种裂解反激活因子。BALF5基因编码DNA Pol催化亚基，BMRF1基因编码DNA Pol辅助亚基。BALF2基因编码单链dna结合蛋白。BBLF4、BSLF1和BBLF2/3基因编码的其余三种蛋白的功能尚未证实，但预测分别为解旋酶、引物酶和解旋酶-引物酶相关蛋白。在本研究中，我们通过免疫沉淀分析发现EBV DNA Pol催化亚基与BBLF4/BSLF1/BBLF2/3复合物相互作用。EBV DNA聚合酶与EBV推测的解旋酶-引物酶复合物的相互作用值得特别关注，因为它们被认为协调复制叉上的前导链和滞后链DNA合成。我们也一直在研究潜伏期EBV DNA复制的细胞周期调控，重点关注hORC， hCDC6和hMCM蛋白，它们都被认为是DNA复制起始的关键调控因子。我们根据自己和其他人的数据假设的当前模型如下。ORC和CDC6被认为在整个细胞周期中与基质相关。MCM异六聚体复合物主要由ORC/CDC6装载到与基质无关的染色质区域。染色质结合MCM的丰度是结合CDC6的数倍，MCM结合染色质的程度相对较大，可以解释哺乳动物细胞中起始的模式，即所谓的“起始区”。结合的mcm可能被CDC7和CDK2激酶磷酸化激活。激活的MCM在DNA复制中起着未知但重要的作用，同时从染色质中转移，与DNA复制耦合。携带eb病毒的胃腺癌通常伴有大量淋巴细胞浸润。为了表征肿瘤浸润淋巴细胞（til），我们从手术切除的ebv相关胃癌中分离并培养了这种细胞。分离的TILs由hla - i类限制性CD8+细胞毒性T淋巴细胞（ctl）组成，ctl能杀死自体ebv转化的细胞，但不能杀死PHA母细胞，并识别HLA-A24为限制性分子。我们的数据表明，一些细胞蛋白可能参与了对ebv相关胃癌的强T细胞反应。少

EBV encodes seven viral genes that are essential for the oriLyt-dependent DNA replication. The BZLF1 protein is an oriLyt-binding protein and acts also the lytic transactivator. The BALF5 gene encodes the DNA Pol catalytic subunit and the BMRF1 gene encodes the DNA Pol accessory subunit. A single-stranded DNA-binding protein is encoded by the BALF2 gene . The functions of the remaining three proteins encoded by the genes BBLF4 , BSLF1, and BBLF2/3 are not demonstrated but predicted to be helicase, primase, and helicase-primase associated proteins, respectively. In the present study, we found that the EBV DNA Pol catalytic subunit interacts with the BBLF4/BSLF1/BBLF2/3 complex by immunoprecipitation analyses. The interactions of the EBV DNA polymerase with the EBV putative helicase-primase complex warrant particular attention because they are thought to coordinate leading and lagging strand DNA synthesis at the replication fork.We have been also studying cell cycle regulation of latent … More phase EBV DNA replication, focusing on hORC, hCDC6 and hMCM proteins, all of which are thought to be key regulators of initiation of DNA replication. Current model which we hypothesize from our and others data is as follows. ORC and CDC6 are assumed to be associated with the matrix throughout the cell cycle. MCM heterohexameric complexes are loaded, by ORC/CDC6, mainly onto chromatin regions not associated with the matrix. The abundance of chromatin-bound MCM is several times that of bound CDC6, and the relatively large extent of MCM-bound chromatin could explain the pattern of initiation in mammalian cells, namely the so-called "initiation zone". The bound MCMs might be activated through phosphorylation possibly by CDC7 and CDK2 kinase. The activated MCM plays an unknown but essential role in DNA replication, and is simultaneously displaced from chromatin, coupled to DNA replication.Gastric adenocarcinomas carrying EBV are known to be accompanied by massive lymphocyte infiltration. To characterize the tumor infiltrating lymphocytes(TILs), we isolated and cultured such cells from surgically resected EBV-associated gastric carcinoma. The isolated TILs consisted of HLA-class I-restricted CD8+ cytotoxic T lymphocytes(CTLs) which killed autologous EBV-transformed cells but not PHA blast cells and recognized HLA-A24 as restriction molecules. Our data indicated that some cellular proteins may be involved in the strong T cell response to EBV-associated gastric carcinoma. Less