ENHANCER-INDEPENDENT AND DEPENDENT ACNPV GENE ACTIVATION

增强子独立和依赖性 ACNPV 基因激活

• 批准号: 3454999
• 负责人: Linda A. Guarino
• 金额: $ 8.57万
• 依托单位: TEXAS A&M UNIVERSITY HEALTH SCIENCE CTR
• 依托单位国家: 美国
• 财政年份: 1989
• 资助国家: 美国
• 起止时间: 1989-03-01 至 1994-02-28
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/3454999
• 关键词: Cypovirus; DNA binding protein; DNA footprinting; Insecta; chromatography; gene deletion mutation; genetic enhancer element; genetic mapping; genetic promoter element; insect virus; nucleic acid sequence; point mutation; tissue /cell culture; virus DNA; virus protein

Gene expression from the delayed early 39K promoter of Autographa californica nuclear polyhedrosis virus (AcNPV) is regulated by both a trans-activating viral factor and a cis-acting enhancer. The trans-activating factor is IE1, an immediate early gene product absolutely required for 39K expression in a transient assay analysis, but insufficient for optimal expression under certain conditions. When the concentration of IE1 is limiting, a viral enhancer element cis-linked to the 39K promoter is also required. Enhancer function is also dependent upon IE1. Thus, the IE1 gene product plays two roles in the regulation of this delayed early gene; in trans-activates both the 39K promoter and the viral enhancer that cis-activates the 39k promoter. The mechanism of action of the enhancer-dependent and enhancer-independent regulation of 39K will be defined. Specifically, this work includes five parts: (1) identification of the enhancer sequences which influence the interactions between viral DNA and trans- activating host and viral factors, (2) the identification of factors which bind the viral enhancer in normal Spodoptera frugiperda cells (host specific factors) and in cells which express the viral IE1 gene (IE1-specific factors), (3) analysis of the 39K promoter sequences that are important for transcription in the presence and absence of the cis-linked enhancer, (4) identification of factors which bind the 39K promoter in normal and in IE1- expressing cells, and (5) confirmation of in vitro binding analysis by transient assay analysis and in vivo susceptibility of viral DNA to chemical and enzymatic probes.

来自延迟的39K启动子的基因表达 加州核多面病毒（ACNPV）都受到两者的调节 反式激活病毒因子和顺式作用增强子。 这 反式激活因子是IE1，即直接的早期基因产物 瞬态测定中39K表达的绝对需要 分析，但不足以在特定下进行最佳表达 状况。 当IE1的浓度限制时，病毒 还需要与39K启动子连接的增强子元件顺式连接。 增强子功能也取决于IE1。 因此，IE1基因 产品在该延迟的早期调节中扮演两个角色 基因;在反式激活中，既有39K启动子和病毒 顺式激活39K启动子的增强子。 机制 增强子依赖和增强子无关的动作 将定义39K的法规。 具体来说，这项工作 包括五个部分：（1）识别增强子序列 影响病毒DNA与反式的相互作用 激活宿主和病​​毒因素，（2）鉴定 在正常脚翅目中结合病毒增强子的因素 frugiperda细胞（宿主特定因素）和在表达的细胞中 病毒IE1基因（IE1特异性因素），（3）对39K的分析 对转录很重要的启动子序列 顺式连接增强子的存在和不存在，（4）识别 在正常和IE1中结合39K启动子的因素 表达细胞，（5）确认体外结合分析 通过瞬时测定分析和病毒DNA的体内敏感性 进行化学和酶探针。