Expression data from CCL185 carcinoma cell line transiently infected with EBV

Epstein-Barr virus(EBV) is associated with malignancies from lymphoid and epithelial origin. In many cases, an incomplete EBV association is noted and confoundswhate role the virus plays in oncogenesis. A number of viral proteins have been shown to interact with epigenetic factors to regulate both viral and host gene expression. Thus, we hypothesize that EBV may inadvertantly induce epigenetic alterations to the host genome that are maintained upon loss of the virus. If proven, these results would broaden EBV's role in tumorigenesis and provide a mechanism for how a tumor virus can act in a "hit-and-run" fashion. We examined EBV-induced epigenetic alterations in a setting of viral redundancy and loss. With mounting evidence that EBV can induce epigenetic alterations, we developed a transient infection model where a clonal derivative (designated cl3) from the CCL185/A549 cell line was infected with a recombinant EBV carrying neomycin resistance cassette in place of the BDLF3 open reading frame. Infected cells were passaged ten times, and then selection pressure was removed for an additional ten passages to allow for viral loss. Three transiently-infected EBV-negative clones were identified by single cell cloning (designated 10-9, 10-10, and 10-14). Uninfected parental clone and cells transfected with pcDNA3 plasmid were passaged alongside the transiently-infected clones. Each sample was analyzed in duplicate.爱泼斯坦-巴尔病毒（Epstein-Barr virus，EBV）与淋巴样及上皮来源的恶性肿瘤密切相关。在多数研究场景中，其与肿瘤的关联并不完全明确，这一现状会混淆该病毒在肿瘤发生过程中所发挥的具体作用。已有多项研究证实，多种病毒蛋白可与表观遗传因子（epigenetic factors）相互作用，进而调控病毒自身及宿主的基因表达。据此，我们提出假说：EBV可能会在无意间诱导宿主基因组发生表观遗传改变，且这种改变会在病毒清除后仍得以稳定维持。若该假说得以验证，将拓展EBV在肿瘤发生中的作用范畴，并为肿瘤病毒如何以“击中即逃逸（hit-and-run）”模式发挥功能提供机制解释。我们在病毒冗余表达及病毒丢失的实验体系中，探究了EBV诱导的表观遗传改变。鉴于越来越多的证据表明EBV可诱导表观遗传改变，我们构建了瞬时感染模型：将携带有替代BDLF3开放阅读框（open reading frame）的新霉素抗性基因盒（neomycin resistance cassette）的重组EBV，感染源自CCL185/A549细胞系的克隆衍生株（命名为cl3）。感染后的细胞传代10次，随后去除筛选压力，再继续传代10次以促使病毒丢失。通过单细胞克隆（single cell cloning）技术，我们筛选得到3株瞬时感染后呈EBV阴性的克隆（分别命名为10-9、10-10及10-14）。未感染的亲代克隆以及转染pcDNA3质粒的细胞，与上述瞬时感染克隆同步传代培养。所有样本均进行双份平行检测。