GENE EXPRESSION OF HUMAN RESPIRATORY SYNCYTIAL VIRUS

人类呼吸道合胞病毒的基因表达

• 批准号: 2457821
• 负责人: SAILEN BARIK
• 金额: $ 10.61万
• 依托单位: UNIVERSITY OF SOUTH ALABAMA
• 依托单位国家: 美国
• 财政年份: 1994
• 资助国家: 美国
• 起止时间: 1994-08-01 至 1999-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/2457821
• 关键词: RNA biosynthesis; actins; antibody; gene expression; genetic transcription; laboratory mouse; laboratory rabbit; molecular cloning; molecular pathology; mutant; nucleic acid sequence; phosphoproteins; phosphorylation; protein kinase; protein structure function; recombinant proteins; respiratory syncytial virus; tissue /cell culture; virus RNA; virus genetics; virus protein; virus replication

The long-term goal of this project is to understand the molecular basis of growth and pathogenicity of human respiratory syncytial virus (RSV), the single most important agent of respiratory diseases in the young children throughout the world and a probable agent of sudden infant death syndrome (SIDS). A recurring cause of pneumonia-like disease characterized by rhinitis, pharyngitis, and croup, it accounts for significant morbidity in the first year of human life. It is now clear that a better understanding of the regulation of viral growth at the molecular level is an essential prerequisite to a successful control of this human pathogen, particularly in view of the failure of immune therapy. Although considerable information is available regarding the nucleotide sequence of the viral genome and its constituent genes, our knowledge about the molecular mechanism of expression of the genes remains poor. Recently, we have taken an important step in this direction by establishing an in vitro of transcription system of RSV which efficiently synthesizes all viral mRNAs. The fidelity of the system is underscored by our finding that the mRNAs are capped, methylated and polyadenylated. More importantly, we have demonstrated an essential requirement of cellular protein(s) in viral transcription. Recently, we have dissected the RSV transcription apparatus even further: we have purified viral L protein, phosphoprotein, and the N-RNA template from one another and reconstituted functional transcription using these purified macromolecules. The reconstituted system is now amenable to studies of structure and function of its components. We have recently expressed RSV P protein in E. coli in a phosphate-free form and shown that it can be phosphorylated by cell kinase in vitro. Inhibition of the cell kinase also inhibited RSV transcription suggesting an essential role for P protein phosphorylation in its function. The immediate major goal is to use the reconstituted system to understand the role of P protein phosphorylation in detail, and to characterize the identity of the cellular protein (s) and determine its exact role in RSV transcription. These studies should provide important insights into the functioning of the viral transcription machinery which is an essential component in RSV growth and pathogenesis.

该项目的长期目标是了解 人呼吸道合胞病毒（RSV）的生长和致病性， 幼儿中最重要的呼吸道疾病药物 在世界范围内，可能是猝死综合症的可能的药物 （SIDS）。肺炎样疾病的反复出现的原因是 鼻炎，咽炎和croup，它占明显的发病率 人类生活的第一年。现在很明显，更好的理解 在分子水平上对病毒生长的调节是必不可少的 成功控制这种人类病原体的先决条件，特别是 鉴于免疫疗法的失败。 尽管有关核苷酸的大量信息 病毒基因组及其成分基因的序列，我们的知识 关于基因表达的分子机制仍然很差。 最近，我们通过 建立一个有效的RSV转录系统的体外 合成所有病毒mRNA。系统的保真度强调了 我们的发现，mRNA被封闭，甲基化和聚腺苷酸化。更多的 重要的是，我们已经证明了细胞的重要要求 病毒转录中的蛋白质。最近，我们剖析了RSV 转录设备进一步：我们已经纯化了病毒L蛋白， 磷蛋白和N-RNA模板相互重组 使用这些纯化的大分子的功能转录。 现在，重构系统适合结构研究和 其组件的功能。我们最近在E中表达了RSV P蛋白。 大肠杆菌无磷酸盐形式，表明它可以被磷酸化 体外细胞激酶。 细胞激酶的抑制也抑制了RSV 转录表明P蛋白磷酸化至关重要 在其功能中。直接的主要目标是使用重组 了解P蛋白质磷酸化的作用的系统，并 表征细胞蛋白的身份并确定其身份 在RSV转录中的确切作用。这些研究应该提供重要的 对病毒转录机械功能的见解 是RSV生长和发病机理中的重要组成部分。