Cellular Determinants of Apoptosis in Virus-Infected Cells

病毒感染细胞凋亡的细胞决定因素

• 批准号: 8242019
• 负责人: GANES C. SEN
• 金额: $ 34.62万
• 依托单位: CLEVELAND CLINIC LERNER COM-CWRU
• 依托单位国家: 美国
• 财政年份: 2008
• 资助国家: 美国
• 起止时间: 2008-04-01 至 2013-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8242019
• 关键词: 1-Phosphatidylinositol 3-Kinase; Ablation; Apoptosis; Apoptotic; Biochemical; Biological; Candidate Disease Gene; Caspase; Cell Death; Cell Line; Cell Survival; Cells; Cessation of life; Cytolysis; End Point Assay; Family member; Gene Activation; Genes; Genetic; Genetic Screening; Health; Hour; Human Genome; Immune response; Individual; Infection; Insertional Mutagenesis; Isoenzymes; Libraries; Life; Life Cycle Stages; Literature; Lytic; Lytic Phase; Mammalian Cell; Measles virus; Mediating; Mediator of activation protein; Mitochondria; Molecular; Mumps virus; Nature; Organism; Outcome; Paramyxoviridae; Paramyxovirus; Pathogenesis; Pathway interactions; Phase; Process; Production; Property; Protein Biosynthesis; Protein Kinase; Proteins; RNA chemical synthesis; Regulation; Respiratory syncytial virus; Risk; Role; Sendai virus; Severities; Signal Transduction; Small Interfering RNA; Societies; Stress; System; Testing; Time; Viral; Virion; Virus; Virus Diseases; Virus Replication; Work; cell type; human IRF3 protein; interferon regulatory factor-3; killings; mutant; novel; receptor; research study; response; small hairpin RNA; transcription factor; viral RNA

DESCRIPTION (provided by applicant): Because viral diseases are major health risks to the society, it is important to understand how they are caused and how our natural defense systems can protect us. A virus-infected cell can either die or it can live; if it lives it may be permanently infected with the virus. This proposal is to investigate the molecular mechanism that dictates the above choice, namely, whether a virus-infected cell dies or lives and produces more viruses. Many major viral diseases are caused by Paramyxoviridae, such as respiratory syncytial virus, parainfluzenza virus, measles virus and mumps virus. The nature and the severity of pathogenesis is determined by the host response, a major component of which is the innate response of the infected cell which may undergo apoptosis or live and be capable to establish persistent infection, the two outcomes having opposite effects on the host organism. We have made the novel observation that in cells not expressing the cellular transcription factor IRF-3, paramyxoviruses are not lytic because IRF-3 initiates an apoptotic response in infected cells. Moreover, this response is greatly accelerated by inhibiting the activity of PI3 kinases. These observations led us to formulate the hypothesis that IRF-3 determines the choice between lytic and non-lytic infection and PI3 kinases determine the timing of the lysis. Three specific aims will be pursued to test the hypothesis. In the first aim, we will determine which known properties of IRF-3 are needed and which known IRF-3 regulated genes may be mediating apoptosis. These experiments will use over- expression of individual proteins or their ablation by siRNA. Finally, unbiased genetic screens of the whole human genome will be conducted by using insertional mutagenesis or an appropriate shRNA library and cell survival as the end point assay. In the second aim, we will determine whether the intrinsic or the extrinsic pathway is used in viral apoptosis. We will also identify the specific isozyme of PI3 kinase and its downstream targets that are required for blocking the rapid apoptosis. In the third aim, we will compare the properties of the persistently infected cell with those of the acutely infected cells. For this purpose, the different steps of the viral life cycle will be examined. Moreover, the status of the innate immune response in the persistently infected cells will be evaluated.

描述（由申请人提供）：由于病毒疾病是社会的主要健康风险，因此了解它们的造成方式以及我们的自然防御系统如何保护我们很重要。感染病毒的细胞可以死亡或可以生存；如果生存，它可能会永久感染该病毒。该建议是研究决定上述选择的分子机制，即病毒感染的细胞死亡还是生命并产生更多的病毒。许多主要的病毒疾病是由paramyxoviridae引起的，例如呼吸道合胞病毒，parainfluzenza病毒，麻疹病毒和腮腺炎病毒。发病机理的性质和严重程度取决于宿主反应，其中一个主要组成部分是感染细胞的先天反应，该反应可能患有凋亡或生命并能够建立持续的感染，这两个结果对宿主生物体有相反的影响。我们已经做出了新的观察结果，即在不表达细胞转录因子IRF-3的细胞中，帕托氏病毒不是裂解的，因为IRF-3在感染细胞中启动了凋亡反应。此外，通过抑制PI3激酶的活性大大加速了这种反应。这些观察结果使我们提出了以下假设：IRF-3决定了裂解和非裂解感染之间的选择，而PI3激酶决定了裂解的时机。将追求三个具体目标以检验该假设。在第一个目标中，我们将确定需要哪些已知的IRF-3特性，哪些已知的IRF-3调节基因可能正在介导凋亡。这些实验将使用单个蛋白质或siRNA消融的过度表达。最后，将通过使用插入诱变或适当的shRNA文库和细胞存活作为终点测定法进行整个人类基因组的公正遗传筛选。在第二个目标中，我们将确定固有或外在途径是否用于病毒凋亡。我们还将确定阻断快速凋亡所需的PI3激酶的特定同工酶及其下游靶标。在第三个目标中，我们将比较持续感染的细胞的性质与急性感染细胞的特性。为此，将检查病毒生命周期的不同步骤。此外，将评估持续感染细胞中先天免疫反应的状态。