HOST-VIRUS INTERACTION AND GENE EXPRESSION

宿主-病毒相互作用和基因表达

• 批准号: 2066926
• 负责人: Amiya K. Banerjee
• 金额: $ 23.71万
• 依托单位: CLEVELAND CLINIC FOUNDATION
• 依托单位国家: 美国
• 财政年份: 1992
• 资助国家: 美国
• 起止时间: 1992-02-01 至 1996-01-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/2066926
• 关键词: DNA directed RNA polymerase; Paramyxoviridae disease; Paramyxovirus; RNA biosynthesis; SDS polyacrylamide gel electrophoresis; actin binding protein; actins; autoradiography; capsid; complementary DNA; crosslink; cytoskeletal proteins; enzyme mechanism; gene expression; genetic mapping; genetic transcription; histopathology; host organism interaction; messenger RNA; molecular pathology; monoclonal antibody; phosphorylation; polymerase chain reaction; protein purification; radioimmunoassay; respiratory infections; respiratory syncytial virus; serotyping; site directed mutagenesis; tissue /cell culture; transfection; virus cytopathogenic effect; virus genetics; virus protein; virus replication

The long term goal of the present proposal is to understand the molecular basis of pathogenicity of the respiratory pathogen, human parainfluenza virus 3 (HPIV3), a member of the family paramyxoviridae. The parainfluenza viruses are important respiratory tract pathogens, second to respiratory syncytial virus, as causative agents of common illnesses of children like rhinitis, pharyngitis, and bronchitis. Studies in different parts of the world indicate that these viruses are associated with approximately 40% of severe croup in children. HPIV3 is most pathogenic among the other HPIV types. Effective vaccines to control infection in children have so far been inadequate. Although considerable information regarding the structure and the nucleotide sequence of the genome of HPIV3 is known, our knowledge of the mechanism of gene expression of this important human pathogen is meager. To gain insight into understanding the functions of the viral genes, we recently developed an in vitro transcription system for HPIV3 which efficiently synthesizes virus-specific mRNAs in vitro. More importantly, we demonstrated that cellular actin was needed for effective transcription in vitro to occur, suggesting a possible involvement of cytoskeletal network in the life-cycle of HPIV replication. To delineate the mechanism of gene expression of HPIV3 and study the role of cellular cytoskeletal protein(s) in this process, we propose to study in detail the role of cellular actin and actin-binding proteins in HPIV3 transcription/replication as well as the functional roles of viral RNA polymerase proteins (L and P proteins) in these processes. An in-depth study would be carried out to determine the molecular form of actin required for transcription and whether any other cellular proteins, e.g. actin-binding proteins, are necessary for actin to function. In addition, mode of action of actin in RNA synthesis as well as association of actin with viral nucleocapsid in vitro and in vivo will be studied. The functions of viral RNA polymerase components (L and P proteins) in transcription and replication and their interaction with cellular actin will be studied. Emphasis would be laid on the viral P gene which encodes, in addition to the P protein, C protein and P-D protein, the latter protein arising by a novel editing mechanism. Understanding the functions of these proteins and the unique editing reaction during P gene transcription along with the role of actin in these processes would certainly provide deeper insight into the mechanism of gene expression of HPIV3 and HPIV3-cell protein interactions.

本提案的长期目标是了解分子 呼吸道病原体人副流感病毒的致病性基础 病毒3（HPIV 3），副粘病毒科的成员。 副流感 病毒是重要的呼吸道病原体，仅次于呼吸道 合胞病毒是儿童常见疾病的病原体， 鼻炎、咽炎和支气管炎。 在不同地区的研究 世界表明，这些病毒与大约40%的 儿童重症哮吼。 HPIV 3是所有HPIV中致病性最强的 类型 到目前为止，控制儿童感染的有效疫苗 是不够的。 虽然大量的信息关于结构和 HPIV 3基因组的核苷酸序列是已知的，我们对HPIV 3基因组的了解是已知的。 这种重要的人类病原体的基因表达机制是微薄的。 为了深入了解病毒基因的功能，我们 最近开发了HPIV 3的体外转录系统， 在体外有效合成病毒特异性mRNA。 更重要的是， 我们证明了细胞肌动蛋白是有效转录所必需的， 在体外发生，这表明可能涉及细胞骨架 在HPIV复制的生命周期中的网络。 来描述 HPIV 3基因表达的研究及细胞骨架的作用 蛋白质在这一过程中的作用，我们建议详细研究 HPIV 3细胞肌动蛋白和肌动蛋白结合蛋白 转录/复制以及病毒RNA的功能作用 聚合酶蛋白（L和P蛋白）在这些过程中。 将进行深入研究，以确定 转录所需的肌动蛋白，以及是否有任何其他细胞蛋白， 例如肌动蛋白结合蛋白是肌动蛋白发挥功能所必需的。 在 此外，肌动蛋白在RNA合成中的作用方式以及缔合 将研究肌动蛋白与病毒核衣壳在体外和体内的相互作用。 的 病毒RNA聚合酶组分（L和P蛋白）在 转录和复制及其与细胞肌动蛋白的相互作用 将被研究。 重点将放在病毒P基因上， 除了P蛋白、C蛋白和P-D蛋白外，后一种蛋白 由一种新的编辑机制产生。 了解这些功能 蛋白质和独特的编辑反应，在P基因转录沿着 与肌动蛋白在这些过程中的作用， HPIV 3和HPIV 3-细胞基因表达机制的研究 蛋白质相互作用