"Project 3" Defining the in vivo function of ncRNAs during MHV68 latency and lymphomagenesis

项目概要 转化的人类伽马疱疹病毒 EBV 和 KSHV 在 B 细胞中建立稳定的潜伏感染， 提供可能导致恶性疾病发展的终生病毒库。因此，定义 控制长期潜伏期和肿瘤发生的机制对于设计合理的策略至关重要 预防疾病。人类伽玛疱疹病毒的体内研究因困难而受到严重限制 在自然宿主中工作。鼠伽马疱疹病毒 68 (MHV68) 与 EBV 和 KSHV 相关并引起 小鼠淋巴瘤和淋巴增殖性疾病，为研究提供了易于操作的小动物模型 体内病毒/宿主关系的机制研究。与 EBV 和 KSHV 一样，MHV68 表达多种形式 ncRNA 在感染和发病机制中的功能很大程度上未知。与大多数人相比 伽马疱疹病毒蛋白编码基因、伽马疱疹病毒 ncRNA 在体内大量表达 慢性感染和淋巴增殖性疾病期间的增生性病变，表明这些 ncRNA 可能在潜伏期和肿瘤发生中发挥关键的保守作用。为了支持这一点，我们已经证明了 MHV68 TMER4 和 EBV EBER1 在血行传播中具有保守的功能，并且在体内 抑制保守的 miRNA 靶点会促进 B 细胞潜伏期。此外，我们的新初步发现表明 小 ncRNA 和 miRNA 在病毒诱导的 B 细胞淋巴瘤的发生和进展中的作用。因此，我们 假设伽马疱疹病毒利用广泛的 ncRNA 来塑造关键的 B 细胞信号通路 从而促进病毒传播、潜伏和肿瘤发生。目标 1（与项目 2 合作 在核心 C 和 D 的支持下，我们将定义伽玛疱疹病毒小 RNA 的机制 促进受感染 B 细胞的流出和传播，促进 B 细胞在周围部位的潜伏期，并驱动 淋巴瘤发生的关键阶段。目标 2（与项目 2 和 3 合作，并得到 Cores 的支持） B、C 和 D) 我们将定义伽玛疱疹病毒 miRNA 对宿主 mRNA 和 lncRNA 抑制的贡献 B 细胞潜伏期和淋巴瘤发生。该计划的高度协作性以及系统性 对携带 EBV 或 KSHV 基因的 ncRNA 突变体和重组病毒进行体内分析，提供了 确定 ncRNA 贡献的特定分子机制的极其强大的方法 伽玛疱疹病毒潜伏期和肿瘤发生。

Project Summary The transforming human gammaherpesviruses EBV and KSHV establish stable latent infections in B cells, providing a lifelong reservoir of virus that can contribute to the development of malignant disease. Thus, defining the mechanisms that govern long-term latency and tumorigenesis is critical for designing rational strategies to prevent disease. In vivo studies of gammaherpesviruses in humans have been severely limited by the difficulties of working in the natural host. Murine gammaherpesvirus 68 (MHV68) is related to EBV and KSHV and causes lymphomas and lymphoproliferative disease in mice, providing a readily manipulatable small animal model for mechanistic studies of the virus/host relationship in vivo. Like EBV and KSHV, MHV68 expresses multiple forms of ncRNAs whose functions during infection and pathogenesis are largely unknown. In contrast to most gammaherpesvirus protein-coding genes, gammaherpesvirus ncRNAs are abundantly expressed in vivo during chronic infection and in hyperplastic lesions during lymphoproliferative disease, suggesting that these ncRNAs may play key conserved roles in latency and tumorigenesis. In support of this, we have demonstrated that MHV68 TMER4 and EBV EBER1 share a conserved function in hematogenous dissemination, and that in vivo suppression of a conserved miRNA target promotes B cell latency. Further, our new preliminary findings implicate both small ncRNAs and miRNAs in the genesis and progression of virus-induced B cell lymphoma. Thus, we hypothesize that gammaherpesviruses utilize a broad array of ncRNAs to shape critical B cell signaling pathways and thereby promote virus dissemination, latency and tumorigenesis. In Aim 1 (in collaboration with Project 2 and with support from Cores C and D), we will define the mechanisms by which gammaherpesvirus small RNAs promote the egress and dissemination of infected B cells, facilitate B cell latency at peripheral sites, and drive key stages of lymphomagenesis. In Aim 2 (in collaboration with Projects 2 and 3, and with support from Cores B, C and D) we will define the contribution of gammaherpesvirus miRNA repression of host mRNAs and lncRNAs to B cell latency and lymphomagenesis. The highly collaborative nature of this program, along with the systematic in vivo analyses of ncRNA mutants and recombinant viruses carrying EBV or KSHV genes, provides an extremely powerful means to determine the specific molecular mechanisms by which ncRNAs contribute to gammaherpesvirus latency and tumorigenesis.