Gene regulatory role of vIRF1 in KSHV infection

摘要 人类感染致癌性卡波西肉瘤相关疱疹病毒(KSHV)始于口腔 通过在口腔上皮细胞中复制。口腔上皮细胞裂解性KSHV感染的调节仍然很差 这阻碍了有效的抗病毒疗法的发展，以阻断KSHV的传播。其中之一 KSHV的独特特征是它编码四种不同的病毒干扰素调节因子(VIRF)，这四种因子是 与细胞干扰素调节因子(IRF)同源。其中之一是vIRF1，它表示为 在KSHV感染期间最早和最高水平，并被KSHV用来控制许多不同方面的 发病机制，如阻断抗病毒反应或刺激肿瘤发生和感染细胞的生长。 而vIRF1在抑制不同免疫应答信号通路中的作用已有研究 详细地说，vIRF1在调节KSHV和宿主细胞基因方面的转录功能是 在病毒发病机制中的重要作用仍然知之甚少。VIRF1是由449个氨基酸组成的长蛋白质。它有一个推定的 DNA结合域(DBD)，也包含核定位信号(NLS)，它具有C-末端的IRF 交互作用域(IAD)。我的初步实验表明，vIRF1的DNA结合域(DBD)是 VIRF1介导的宿主和病毒基因诱导所需的，表明vIRF1同时诱导病毒和宿主基因 通过与它们的发起人绑定。我们的研究小组还发现了与病毒和宿主蛋白相互作用的 VIRF1通过蛋白质复合体纯化。这揭示了vIRF1与组蛋白乙酰转移酶的相互作用。 (HATS)，在基因诱导中经常作为各种DNA结合转录因子的辅助因子。我 假设vIRF1可以与特定的宿主和病毒启动子结合，并使用HATS诱导其 促进口腔上皮细胞中KSHV裂解感染的相应基因。我提出了两个具体目标： 测试我的假设。目标1专注于在缺少vIRF1的情况下识别vIRF1直接靶向的宿主基因 在KSHV感染的口腔上皮细胞中的其他vIRFs。目标2是确定中的vIRF1反应元件 VIRF1靶向KSHV启动子，而KSHV启动子是vIRF1介导的启动子激活所必需的。这项工程的完成 该项目将提供病毒学、表观遗传学和基因组学方法方面的深入培训，因为这些方法将 用来阐明这种关键的寄主病原体相互作用。口腔生物学综合训练计划 在佛罗里达大学牙科学院将为我提供一个出色的环境，让我为 通过提供个性化的指导和出色的临床和研究培训，成为一名牙医科学家。

Abstract Infection of humans by the oncogenic Kaposi's sarcoma-associated herpesvirus (KSHV) starts in the oral cavity by replicating in oral epithelial cells. Regulation of lytic KSHV infection of the oral epithelium is still poorly understood, which hampers the development of effective antiviral therapies to block KSHV transmission. One of the unique features of KSHV is that it encodes four different viral interferon regulatory factors (vIRFs), which are homologous with cellular interferon regulatory factors (IRFs). One of them is vIRF1, which is expressed the earliest and at the highest level during KSHV infection and is used by KSHV to control many different aspects of the pathogenesis such as blocking antiviral response or stimulating oncogenesis and the growth of infected cells. While the function of vIRF1 in the inhibition of different immune response signaling pathways have been studied in detail, the transcriptional function of vIRF1 in the regulation of both KSHV and host cellular genes that are important in viral pathogenesis is still poorly understood. vIRF1 is 449 amino acid long protein. It has a putative DNA-binding domain (DBD), which also contains a nuclear localization signal (NLS), and it has a C-terminal IRF interaction domain (IAD). My preliminary experiments revealed that the DNA-binding domain (DBD) of vIRF1 is required for vIRF1-mediated host and viral gene induction, indicating vIRF1 induces both viral and host genes by binding to their promoters. Our research group has also identified the viral and host proteins interacting with vIRF1 by protein complex purification. This revealed the interaction of vIRF1 with histone acetyltransferases (HATs) that often serve as co-factors for various DNA-binding transcription factors in gene induction. I hypothesize that vIRF1 can bind to specific host and viral promoters and uses HATs to induce their corresponding genes to enhance the lytic KSHV infection in oral epithelial cells. I propose two specific aims to test my hypothesis. Aim 1 focuses on identifying the host genes that are directly targeted by vIRF1 in the absence of the other vIRFs in KSHV-infected oral epithelial cells. Aim 2 is to determine the vIRF1 response element in vIRF1 target KSHV promoters that is required for vIRF1-mediated promoter activation. The completion of this project will provide in-depth training in virology, epigenetics and genomics approaches as these methods will be used to elucidate this critical host pathogen interactions. The Comprehensive Training Program in Oral Biology at the University of Florida College of Dentistry will provide me an outstanding environment to prepare me for a career as a dentist scientist by providing personalized mentorship and excellent clinical and research training.