羊口疮病毒ORFV118蛋白调控宿主细胞凋亡的分子机制研究

细胞凋亡是宿主细胞对抗病毒感染的重要防御机制之一。近期研究发现羊口疮病毒(ORFV)的新基因ORFV118具有抑制宿主细胞凋亡的作用，并与病毒的毒力相关，但其具体调控机制尚不清楚。本研究拟首先采用流式细胞术、Western blot等方法明确ORFV118对细胞凋亡的影响；再通过微阵列分析、细胞凋亡蛋白膜芯片筛选、capspase活性检测实验分别从基因、蛋白和酶活性水平探索ORFV118调控宿主细胞凋亡的可能途径；并通过蛋白酶激活实验、Western blot检测细胞凋亡相关蛋白、酵母双杂交筛选、免疫共沉淀（Co-IP）和激光共聚焦荧光显微镜等方法阐明ORFV118抑制凋亡的分子机制；最后，用118缺陷病毒NA1/11Δ118感染动物，以观察ORFV118基因在病毒致病机制、病毒毒力以及调节宿主免疫应答和炎症反应中的作用。本研究结果将为ORFV的发病机制、免疫机理以及疾病控制提供理论依据。

Apoptosis is one of the important roles of the host cell defense against viral infection. Our recent studies have found that sheep contagious pustular virus, orf virus (ORFV) encoded a novel protein, and called ORFV118 can inhibit host cell apoptosis and associate with the viral virulence. However, the exact molecular mechanisms are poorly understood. In the proposed study, we will utilize the microarray analysis, apoptotic protein microarray technology, yeast two-hybrid system, co-immunoprecipitation experiments (Co-IP) and flow cytometry to investigate the exact molecular mechanisms of ORFV118, and the role of ORFV118 in host pathogenesis. Specific aim 1: Determine the molecular mechanisms by which ORFV118 regulates apoptosis process. Aim 2: Identify the molecules between ORFV118 and host cells interaction. Aim 3: Determine the role of ORFV118 in virulence and pathogenesis in the natural host. Given that apoptosis is one of the fundamental defensive roles in host cell against viral infection and that ORFV118 is novel, this work is likely to have broad application in further defining cellular apoptosis pathway responses to viral infection and identifying critical junctures in this pathway, therefore, opening to viral manipulation and possible prophylactic-therapeutic intervention. A better understanding of novel viral immunomodulatory function and host responses in the skin may suggest new strategies potentially involving improved vaccines and adjuvant for controlling viral and other diseases.