Feline Immunodeficiency Virus Orf-A A Model for HIV Vpr

描述（由申请人提供）：猫免疫缺陷病毒（FIV）动物模型为评估人类艾滋病毒/艾滋病的新型抗病毒策略提供了独特且重要的机会。然而，与 HIV 编码蛋白共享保守功能的 FIV 辅助基因的表征对于开发这种动物模型至关重要，以表征可用于治疗人类免疫缺陷病毒 1 (HIV-1) 感染的有针对性的干预策略。 FIV 基因 OH-A，也称为 orf-2，编码 77 个氨基酸的辅助蛋白。我们最近的研究表明 Orf-A 在涉及病毒粒子形成的 FIV 生命周期的后期步骤和涉及病毒感染性的早期步骤中非常重要，并绘制了复制中这些步骤所需的关键 Orf-A 结构域。在另一项研究中，我们报告了以前未被识别的 Orf-A 的新功能。 GFP-Orf-A 融合蛋白在哺乳动物细胞中的表达证明了这种 FIV 辅助蛋白的核定位，并促进了对 FIV Orf-A 核输入至关重要的核定位信号 (NLS) 的绘制。最后，对瞬时表达 GFP-Orf-A 的细胞的细胞周期概况的评估表明，Orf-A 会导致细胞周期的第二个间隙 (G2) 停滞。基于这些新发现，我们假设 FIV oH-A 编码一种病毒蛋白，其表达的特定特性与 HIV-1 Vpr 表达的特性相似。此外，我们假设特定的 oH.A 蛋白-蛋白相互作用可能是非灵长类慢病毒（FIV 和山羊关节炎脑炎病毒）或灵长类慢病毒（HIV 和猿猴免疫缺陷病毒）编码的辅助基因的共同特性。我们建议通过研究 Orf-A 在病毒复制中的功能、对复制至关重要的 Orf-A 结构域以及 Off-A 影响病毒复制的机制来进一步表征 FIV Orf-A。重要的是，我们将比较 RV Orf-A 和 HIV Vpr 表达以及亚细胞定位对宿主细胞功能（包括细胞周期和细胞活力）的影响。最后，我们将通过实验猫接种研究测试编码 Orf-A 突变的 FIV 分子克隆来检查 Orf-A 功能对病毒发病机制的影响。 FIV动物模型将用于分析FIV Off-A和HIV-1 Vpr共有的特定病毒基因活性（G2阻滞、细胞凋亡等）对体内病毒发病机制的影响，并相应地检验特定Vpr结构域作为基于HIV-1 Vpr的抗病毒治疗靶点的价值。

DESCRIPTION (provided by applicant): The feline immunodeficiency virus (FIV) animal model presents a unique and significant opportunity for assessing novel antiviral strategies for HIV/AIDS in humans. However, characterization of FIV accessory genes that share conserved functions with HIV-encoded proteins will be crucial to development of this animal model for characterizing targeted intervention strategies useful for treatment of human immunodeficiency virus-1 (HIV-1) infection of humans. FIV gene OH-A, also designated orf-2, encodes a 77 amino acid accessory protein. Our recent studies have shown Orf-A to be important in late steps of the FIV life cycle involved in virion formation and in early steps involved in virus infectivity and have mapped critical Orf-A domains needed for these steps in replication. In a separate study we reported novel previously unrecognized Orf-A functions. Expression of GFP-Orf-A fusion proteins in mammalian cells demonstrated nuclear localization for this FIV accessory protein and facilitated mapping of a nuclear localization signal (NLS) critical for nuclear import of FIV Orf-A. Lastly, assessment of cell cycle profiles of cells transiently expressing GFP-Orf-A demonstrated that Orf-A causes an arrest at the second gap (G2) of the cell cycle. Based on these novel findings, we hypothesize that FIV oH-A encodes a viral protein that expresses specific properties similar to those expressed by HIV-1 Vpr. Furthermore, we hypothesize that specific oH.A protein-protein interactions may be common properties of accessory genes encoded by either non-primate lentiviruses (FIV and caprine arthritis encephalitis virus) or primate lentiviruses (HIV and simian immunodeficiency virus). We propose to further characterize FIV Orf-A by investigating Orf-A functions in virus replication, Orf-A domains critical for replication, and mechanisms by which Off-A affects virus replication. Importantly, we will compare effects of RV Orf-A and HIV Vpr expression and subcellular localization on host cell functions including cell cycle and cell viability. Lastly, we will examine effects of Orf-A function on viral pathogenesis through experimental cat inoculation studies testing FIV molecular clones encoding Orf-A mutations. The FIV animal model will be used to analyze the impact of specific viral gene activities (G2 arrest, apoptosis, etc.) shared by FIV Off-A and HIV-1 Vpr on viral pathogenesis in vivo and to accordingly examine the value of specific Vpr domains as targets for antiviral therapeutics based on HIV-1 Vpr.