Viral-host interactions that influence the life cycle of DNA tumor viruses

项目2 项目摘要摘要 致癌病毒导致了15%-20%的人类癌症。高危HPV与5%的癌症有关， 包括宫颈癌(CC)以及越来越多的口咽鳞状细胞癌 (OSCC)。RNA分子，如长非编码RNA(LncRNAs)和共转录R-环(RNA：DNA 杂交种)与基因组不稳定和癌症的其他特征有关，但在 HPV感染的背景。我们的初步数据显示，高危HPV31阳性细胞增加了 R-环与未感染细胞的比较。此外，我们已经确定了几个可以被 影响上皮细胞分化、增殖和存活的HPV31。这些RNA分子是否具有 促病毒作用和/或在HPV发病机制中的作用尚不清楚。DNA肿瘤病毒EBV和KSHV也是 在口腔中发现，并导致口腔癌。与HPV类似，EBV和KSHV也使用 口腔上皮作为一种激活裂解复制和病毒产生的机制。口腔上皮细胞可能是 唾液中传染性病毒的来源，也是病毒致病的关键组成部分。然而，这些机制 调节HPV的生产性复制，以及口腔中EBV和KSHV的潜伏期/裂解期 上皮细胞的特征不是很好。这些病毒可能使用类似的策略来解开增殖。 从分化到促进病毒持续和病毒传播。KSHV和EBV改变宿主lncRNA 在其他细胞类型中的分布，以及R环形成序列已经在KSHV和EBV病毒基因组中被鉴定。 此外，我们还发现KSHV可诱导口腔上皮细胞R环形成和DNA损伤。我们 假设HPV、EBV和KSHV通过R环重新编程上皮细胞环境 形成/分布和lncRNA表达，以支持病毒的生命周期。具体目标是测试这一点 假设为：(1)通过绘制R-环的分布图，确定R-环在HPV和KSHV发病中的作用 并确定R-环是否对感染细胞的DNA损伤有贡献；(2)确定 细胞内的lncRNAs如何通过一种途径在HPV以及KSHV和EBV在上皮细胞中的生命周期中发挥作用 对特定lncRNA的过度表达/缺失研究的偏颇方法，以及通过 高通量测序以检测lncRNA表达的全局变化。了解R-循环是如何 而有助于复制这些DNA肿瘤病毒的lncRNAs可能会揭示RNA分子在 促进病毒的持久性以及确定可用于治疗病毒的细胞途径- 以及可能与病毒无关的癌症。

Project 2 Project Summary Abstract Oncogenic viruses cause 15-20% of human cancers. High-risk HPVs are associated with 5% of cancers, including cervical cancer (CC) as well as an increasing number of oropharyngeal squamous cell carcinomas (OSCC). RNA molecules, such as long non-coding RNAs (lncRNAs) and co-transcriptional R-loops (RNA:DNA hybrids) are associated with genomic instability and other hallmarks of cancer, but are understudied in the context of HPV infection. Our preliminary data indicate high-risk HPV31 positive cells have increased levels of R-loops compared to uninfected cells. Furthermore, we have identified several lncRNAs that are altered by HPV31 that influence epithelial differentiation, proliferation and survival. Whether these RNA molecules serve a pro-viral role and/or contribute to HPV pathogenesis is unclear. The DNA tumor viruses EBV and KSHV are also found in the oral cavity and cause oral cancers. Similarly to HPV, EBV and KSHV also use differentiation of the oral epithelium as a mechanism to activate lytic replication and virus production. Oral epithelial cells are the likely source of infectious virus in the saliva and a critical component of viral pathogenesis. However, the mechanisms that regulate the productive replication of HPV, as well as the latent/lytic phases of EBV and KSHV in the oral epithelium are not well characterized. These viruses likely employ similar strategies to uncouple proliferation from differentiation to promote both viral persistence and viral spread. KSHV and EBV alter the host lncRNA profile in other cell types, and R-loop forming sequences have been identified in KSHV and EBV viral genomes. Additionally, we have found that KSHV induces R-loop formation and DNA damage in oral epithelial cells. We hypothesize that HPV, EBV and KSHV reprogram the epithelial cell environment through R-loop formation/distribution and lncRNA expression in order to support the viral life cycle. Specific Aims to test this hypothesis are: (1) Determine the role of R-loops in HPV and KSHV pathogenesis by mapping R-loop distribution on cellular and viral DNA and determining if R-loops contribute to DNA damage in infected cells; (2) Determine how cellular lncRNAs contribute to the life cycle of HPV as well as KSHV and EBV in epithelial cells through a biased approach of overexpression/depletion studies of specific lncRNAs, and an unbiased approach through high throughput sequencing to examine global alterations in lncRNA expression. Understanding how R-loops and lncRNAs contribute to replication of these DNA tumor viruses may uncover a role for RNA molecules in facilitating viral persistence as well as identify cellular pathways that may be exploited for the treatment of viral- and potentially non-viral-associated cancers.