Methylation in establishment of latency in gHV68

描述(申请人提供)：CpG基序的甲基化已被确认为基因沉默的全球机制，并被假设为在伽马疱疹病毒潜伏期的建立过程中，甲基化是裂解病毒基因沉默的关键组成部分。人类伽玛疱疹病毒，如Epstein Barr病毒和Campos肉瘤疱疹病毒，与几种淋巴增生性疾病有关。体外研究表明，在潜伏期间，裂解病毒复制所必需的基因的启动子区域被高甲基化，而潜伏相关基因的启动子则被低甲基化。由于这些病毒的宿主范围有限，体内研究很困难，因此以伽马疱疹病毒68(GHV68)为模型来研究伽马疱疹病毒在体内的作用。为了探讨细胞甲基转移酶和甲基化在病毒潜伏期的建立和维持中的作用，我们建议研究小鼠感染不同时间点裂解启动子和潜伏启动子的甲基化数量。甲基化水平将与感染后不同天存在的裂解和潜伏特异性病毒转录本的数量进行比较。最后，将使用cre-lox基因打靶系统分析gHV68感染细胞中细胞甲基转移酶Dnmt1的缺失对潜伏期建立的影响。这些研究将促进我们对体内潜伏期建立和维持机制的理解。

DESCRIPTION (provided by applicant): Methylation of CpG motifs has been identified as a global mechanism in the silencing of genes and it has been hypothesized that methylation is a critical component in the silencing of lytic viral genes during the establishment of latency of gammaherpesviruses. Human gammaherpesviruses such as Epstein Barr virus and Campos's Sarcoma Herpes virus are associated with several lymphoproliferative disorders. In vitro research has shown that during latency the promoter regions of genes necessary for lytic viral replication are hypermethylated while the promoters of latency-associated genes are hypomethylated. Since in vivo studies of these viruses are difficult due to their limited host range, gammaherpesvirus 68 (gHV68) is used as a model to study the effects of gammaherpesviruses in vivo. To address the role of cellular methyltransferases and methylation in the establishment and maintenance of latency of the virus, we propose to investigate the amount of methylation of both lytic and latent promoters during different time points of infection in the mouse. The level of methylation will be compared to the amount of lytic- and latency-specific viral transcripts present at the different days postinfection. Finally, the effect of the deletion of the cellular methyltransferase, Dnmt1, in gHV68-infected cells on the establishment of latency will be analyzed using the cre-lox gene-targeting system. These studies will advance our understanding of the mechanisms that govern the establishment and maintenance of latency in vivo.