TRANSCRIPTION OF HERPES SIMPLEX VIRUS GLYCOPROTEIN GENES

单纯疱疹病毒糖蛋白基因的转录

• 批准号: 3131488
• 负责人: LEWIS I PIZER
• 金额: $ 11.58万
• 依托单位: UNIVERSITY OF COLORADO DENVER
• 依托单位国家: 美国
• 财政年份: 1986
• 资助国家: 美国
• 起止时间: 1986-07-01 至 1989-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/3131488
• 关键词: Alphaherpesvirinae; DNA directed RNA polymerase; HeLa cells; gel filtration chromatography; genetic manipulation; genetic recombination; genetic transcription; messenger RNA; monoclonal antibody; mutant; nucleic acid sequence; plasmids; radiotracer; regulatory gene; thymidine kinase; tissue /cell culture; virus DNA; virus genetics; virus infection mechanism

Herpes simplex virus proteins synthesized in infected cells change in both number and character as the infection proceeds. Underlying these changes in viral proteins are changes in the species and abundance of the viral mRNAs. We proposed that the time of appearance of early and late HSV mRNA is regulated predominantly by the ability to initiate transcription and that several factors control this process. These include primarily DNA sequences upstream from the promotor and viral regulatory proteins. Our goal is to identify the regulatory proteins and the DNA sequences with which they interact. To assay for upstream control elements in viral DNA we will construct recombinant plasmids containing isolated HSV genes, modify them by mutational insertions or deletions and assay their activity in vitro and in vivo. To study transcriptional regulation from these genes, we will use a three-pronged approach that measures RNA synthesis in nuclear extracts, in isolated nuclei, and in cells following transfection with plasmid DNA. The results from the in vivo experiments will be used to assess the validity of the in vitro measurements. Taken together, the data will establish whether transcription of HSV genes is regulated by viral proteins acting either independently or in a coordinate manner and whether specific gene sequences are involved. Long term goals are to study the biochemical interactions between the viral proteins and the enzymes involved in RNA synthesis. The availability of an in vitro transcription system makes the biochemical approach possible.

单纯疱疹病毒感染细胞中合成的蛋白质 随着感染的进行，数字和字符都发生了变化。 在病毒蛋白质的这些变化的基础上， 病毒mRNA的种类和丰度。 我们建议 早期和晚期HSV mRNA的出现时间受到调节 主要是通过启动转录的能力， 有几个因素控制这个过程。 其中主要包括 启动子上游的DNA序列和病毒调节基因 proteins. 我们的目标是确定调控蛋白和DNA 它们相互作用的序列。 测定上游 控制元件的病毒DNA，我们将构建重组 含有分离的HSV基因的质粒，通过 突变插入或缺失，并测定它们在 体外和体内。 为了研究转录调控， 这些基因，我们将使用三管齐下的方法， 核提取物、分离的核和细胞核中的RNA合成 用质粒DNA转染后的细胞。 体内实验的结果将用于评估 体外测量的有效性。 总的来说， 数据将确定HSV基因的转录是否 受病毒蛋白的调节，这些病毒蛋白要么独立发挥作用， 协调方式以及特定基因序列是否 涉案 长期目标是研究 病毒蛋白和酶之间的相互作用 参与RNA合成。 在体外的可用性 转录系统使生物化学方法成为可能。