TMV-COAT PROTEIN IN ENGINEERED RESISTANCE

工程抗性中的 TMV 涂层蛋白

• 批准号: 2063723
• 负责人: Roger Beachy
• 金额: $ 29.94万
• 依托单位: SCRIPPS RESEARCH INSTITUTE, THE
• 依托单位国家: 美国
• 财政年份: 1989
• 资助国家: 美国
• 起止时间: 1989-02-01 至 1997-04-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/2063723
• 关键词: RNA biosynthesis; RNase protection assay; capsid; computer simulation; electron microscopy; gene expression; genetically modified plants; host organism interaction; mutant; physical model; plant genetics; protein engineering; protein structure function; protoplast /spheroplast; site directed mutagenesis; tobacco; tobacco mosaic virus; transposon /insertion element; virion; virus RNA; virus cytopathogenic effect; virus genetics; virus infection mechanism; virus replication

In 1986 this laboratory reported (Science 232:738-743) that transgenic tobacco plants that express a gene encoding the capsid protein (CP) of tobacco mosaic virus (TMV) are substantially protected against infection by TMV. This type of resistance, "coat-protein mediated resistance (CP-MR)", has been used to develop resistance against many different types of viruses in a number of types of plants. Although the successes of CP-MR are well known, and advanced stages of testing and evaluation of transgenic plant lines are in progress, the cellular and molecular mechanisms of resistance are not yet known. The system best-suited for in-depth study is TMV and transgenic tobacco and tomato plants. Published information of the crystal structure of TMV-CP and x-ray fiber diffraction studies of TMV will be used to create mutant proteins to investigate the role of protein structure, protein:protein, and protein:RNA interactions on CP-MR. The effects of wild-type and mutant CPs on virus uncoating, establishment of infection, and local and systemic spread of virus in transgenic plants and synchronously infected protoplasts will be determined. To study early events in virus uncoating in CP-MR an epitope-tagging approach will be taken to follow virus disassembly and protein exchange reactions. To monitor localized spread of the infection in CP-MR, virus replication will be followed in real time by imaging and recording the expression of a luciferase gene that has been incorporated into the virus. To explore the role that expression of the TMV-CP gene in specific cell types has on resistance, transgenic plant lines will be developed that express the CP gene under control of inducible, tightly regulated promoters. Promoters controlled by wounding or added reagents and promoters expressed specifically in epidermal and vascular xylem cells, vascular phloem cells, and in chloroplast containing cells will be used singly or in combination to examine the role of CP in various aspects of virus infection and disease development. Interfacing structural chemistry, protein modeling, molecular genetics, cell biology, and virology as described here represents a novel approach to studies of viral pathogenesis. It will also lead to better understanding of CP-MR and further develop transgenic resistance for use in food production to decrease the often severe virus infections that limit food availability and exacerbate hunger in many parts of the world.

1986年，该实验室报告(Science 232：738-743)转基因 表达编码烟草衣壳蛋白(CP)基因的烟草 烟草花叶病毒(TMV)实质上是通过 烟草花叶病毒。这种抗性称为“外壳蛋白介导的抗性(CP-MR)”， 已经被用来对许多不同类型的病毒产生抵抗力 在许多类型的植物中。尽管CP-MR的成功之处在于 转基因植物检测和评估的已知和高级阶段 品系研究进展，抗性的细胞和分子机制 还不为人所知。 最适合深入研究的系统是烟草花叶病毒和转基因烟草以及 番茄植株。已发表的TMV-CP晶体结构信息 而烟草花叶病毒的x射线纤维衍射研究将被用于创造突变体 蛋白质研究蛋白质结构的作用，蛋白质：蛋白质，和 蛋白质：CP-MR上的RNA相互作用野生型和突变型CPS的作用 关于病毒的脱壳，感染的建立，以及局部和系统的 病毒在转基因植物和同步感染原生质体中的传播 将会被确定。 CP-MR An表位标记法研究病毒脱壳的早期事件 将采取方法来跟踪病毒的分解和蛋白质交换 反应。监测CP-MR病毒感染的局部传播 复制之后将实时成像和记录 已经整合到病毒中的荧光素酶基因的表达。 探讨TMV-CP基因在特定细胞中表达的作用 没有抗性的类型，将开发出转基因植物系 在可诱导的、严格调控的控制下表达CP基因 推动者。受伤害或添加试剂控制的启动子和 启动子在表皮和维管木质部细胞中特异表达， 维管束韧皮部细胞，以及在叶绿体中含有的细胞都会用到 单独或组合研究CP在不同方面的作用 病毒感染和疾病发展。 界面结构化学、蛋白质建模、分子遗传学、 这里描述的细胞生物学和病毒学代表了一种新的方法 病毒致病机制的研究。这也将带来更好的理解 CP-MR和进一步发展转基因抗性在食品中的应用 生产以减少通常严重的限制食物的病毒感染 在世界许多地区，粮食供应不足并加剧了饥饿。