Yajima Misako, Miyata Mamiko, Ikuta Kazufumi, Hasegawa Yasuhisa, Oneyama Chitose, Kanda Teru
Division of Microbiology, Faculty of Medicine, Tohoku Medical and Pharmaceutical University, Sendai 983-8536, Japan.
Division of Cancer Cell Regulation, Aichi Cancer Center Research Institute, Nagoya 464-8681, Japan.
Microorganisms. 2019 Apr 30;7(5):119. doi: 10.3390/microorganisms7050119.
Epstein-Barr virus (EBV) genomes, particularly their latent genes, are heterogeneous among strains. The heterogeneity of EBV-encoded latent membrane protein 1 (LMP1) raises the question of whether there are functional differences between LMP1 expressed by cancer-associated EBV and that by non-cancerous strains. Here, we used bacterial artificial chromosome (BAC)-cloned EBV genomes retaining all virally encoded microRNA (miRNA) genes to investigate the functions of cancer-derived LMP1 in the context of the EBV genome. HEK293 cells were stably transfected with EBV-BAC clone DNAs encoding either nasopharyngeal carcinoma (NPC)-derived CAO-LMP1 (LMP1) or LMP1 from a prototype B95-8 strain of EBV (LMP1). When an EBV-BAC clone DNA encoding LMP1 was stably transfected into HEK293 cells, it generated many more stable transformants than the control clone encoding LMP1. Furthermore, stably transfected HEK293 cells exhibited highly efficient production of progeny virus. Importantly, deletion of the clustered viral miRNA genes compromised the ability to produce progeny viruses. These results indicate that cancer-derived LMP1 and viral miRNAs together are necessary for efficient production of progeny virus, and that the resulting increase in efficiency contributes to EBV-mediated epithelial carcinogenesis.
爱泼斯坦-巴尔病毒（EBV）基因组，尤其是其潜伏基因，在不同毒株间具有异质性。EBV编码的潜伏膜蛋白1（LMP1）的异质性引发了一个问题，即癌症相关EBV表达的LMP1与非癌毒株表达的LMP1之间是否存在功能差异。在此，我们使用保留所有病毒编码的微小RNA（miRNA）基因的细菌人工染色体（BAC）克隆的EBV基因组，在EBV基因组背景下研究癌症来源的LMP1的功能。将编码鼻咽癌（NPC）来源的CAO-LMP1（LMP1）或EBV原型B95-8毒株的LMP1（LMP1）的EBV-BAC克隆DNA稳定转染至HEK293细胞。当将编码LMP1的EBV-BAC克隆DNA稳定转染至HEK293细胞时，与编码LMP1的对照克隆相比，它产生了更多的稳定转化子。此外，稳定转染的HEK293细胞表现出高效的子代病毒产生。重要的是，成簇的病毒miRNA基因的缺失损害了产生子代病毒的能力。这些结果表明，癌症来源的LMP1和病毒miRNA共同对于高效产生子代病毒是必需的，并且由此产生的效率提高有助于EBV介导的上皮细胞癌变。
