Determining the functions of novel genes for influenza A and Ebola viruses

甲型流感病毒(LAV)和埃博拉病毒(EBOV)是导致人类 分别与呼吸道疾病或出血热有关的传染病。两种病毒都有很小的 基因组，仅编码已知的病毒蛋白11-12(LAV)或8(EBOV)；然而，新的 近年来的蛋白质表明，这些病毒可能编码额外的未鉴定基因，这些基因 通过非常规战略转录或转录/翻译。我们假设这些基因和 他们的产品可能对病毒的生命周期做出了贡献。因此，研究项目2(RP 2)的目标是 鉴定并确定未鉴定的LAV和EBOV基因的功能。在爱尼一号，我们将 实验证明，假设的基因可以从LAV和EBOV的基因组中表达出来 蛋白质水平。对于所有未表征的基因类别：1)已证实的假想候选基因，2)未知 候选开放阅读框(即已知已表达但尚未具有功能的病毒蛋白 特征)和3)非编码候选病毒RNA，我们将评估它们的生化功能 未确定的候选基因，包括使用我们实验室的细胞凋亡分析。附加生化 将在RP 1-Baric中进行分析，以检查未知基因在干扰素和Tolllike中的作用 受体信号转导和在RP3-Damania中的作用 炎症体和P53信号转导。总而言之，所有三个RPS都将贡献各种分析来确定 这一建议中未描述的基因的生化功能。通过使用反向基因组学技术 我们实验室为LAV和EBOV开发的病毒，我们将产生不再编码 未经鉴定的基因产物。然后，我们将比较细菌的生长动力学和宿主对感染的反应 野生型病毒和突变病毒在细胞培养中(目标2)和在小鼠模型中(目标3)。此外，我们还将 进行共聚焦显微镜成像(AIM 2)以确定潜在的 新的LAV和EBOV基因产物，以及潜在的新型SARS冠状病毒(RP1)和人类 疱疹病毒8型(Rp3)基因产物。这样的显微镜研究将有助于确定这些细胞的功能

Influenza A viruses (lAV) and Ebola viruses (EBOV) are negafive-sense RNA viruses that cause human infecfions associated with respiratory disease or hemorrhagic fever, respectively. Both viruses have small genomes, encoding only 11-12 (lAV) or 8 (EBOV) known viral proteins; however, the idenfification of novel proteins in recent years suggests that these viruses may encode additional uncharacterized genes that are transcribed or transcribed/translated, via non-conventional strategies. We hypothesize that these genes and their products likely contribute to the viral life cycle. Therefore, the goal of research project 2 (RP 2) is to identify and determine the functions of uncharacterized lAV and EBOV genes. In Aini 1, we will experimentally prove that hypothetical genes can be expressed from the genomes of lAV and EBOV at the protein level. For all uncharacterized gene categories; 1) proven hypothefical gene candidates, 2) unknown open reading frame candidates (i.e., viral proteins already known to be expressed but not yet functionally characterized), and 3) noncoding viral RNA candidates, we will assess the biochemical function of these uncharacterized gene candidates, including using apoptosis assays in our laboratory. Additional biochemical assays will be perfonned in RP 1 - Baric to examine the role of uncharacterized genes in interferon and tolllike receptor signaling and in RP 3 - Damania to examine the role of uncharacterized genes in inflammasome and p53 signaling. Collecfively, all three RPs will contribute various assays to determine the biochemical funcfion of the uncharacterized genes in this proposal. By using reverse genefics techniques developed in our laboratory for lAV and EBOV, we will generate mutant viruses that no longer encode the uncharacterized gene product. We will then compare the growth kinetics and host responses to infection of wild-type virus and mutant viruses in cell culture (Aim 2) and in a mouse model (Aim 3). In addition, we will perform confocal microscopy imagining (Aim 2) to determine the subcellular localization of the potentially novel lAV and EBOV gene products, and also of potentially novel SARS coronavirus (RP 1) and human herpesviruses-8 (RP 3) gene products. Such microscopy studies will aid in determining the function of these