TMV-COAT PROTEIN IN ENGINEERED RESISTANCE

工程抗性中的 TMV 涂层蛋白

• 批准号: 3141285
• 负责人: Roger Beachy
• 金额: $ 29.72万
• 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/3141285
• 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.

在1986年，该实验室报道（Science 232：738-743），转基因的 表达编码以下衣壳蛋白（CP）的基因的烟草植物 烟草花叶病毒（TMV）基本上被保护免受感染， TMV 这种类型的抗性，“外壳蛋白介导的抗性（CP-MR）"， 已经被用来发展对许多不同类型病毒的抵抗力 在许多植物中。 虽然CP-MR的成功是好的， 转基因植物的测试和评价的已知和高级阶段 株系正在进行中，抗性的细胞和分子机制 目前还不清楚。 最适合进行深入研究的系统是TMV和转基因烟草， 番茄植株。 TMV-CP晶体结构的公开信息 TMV的X射线纤维衍射研究将用于创建突变体 蛋白质研究蛋白质结构，蛋白质：蛋白质， 蛋白质：RNA相互作用对CP-MR的影响。 对病毒脱壳、建立感染以及局部和全身 病毒在转基因植物中传播及同步感染原生质体 将被确定。 为了研究CP-MR中病毒脱壳的早期事件， 将采用一种方法跟踪病毒分解和蛋白质交换 反应. 为了监测CP-MR中感染的局部传播，病毒 复制将通过成像和记录在真实的时间 已经掺入病毒的荧光素酶基因的表达。 探讨烟草花叶病毒CP基因在特定细胞中表达的作用 类型的抗性，将开发转基因植物品系， 在可诱导的、严格调控的 发起人。 启动子受损伤或添加试剂控制， 在表皮和维管木质部细胞中特异性表达的启动子， 维管韧皮部细胞和含叶绿体的细胞将被使用 单独或组合，以检查CP在各个方面的作用， 病毒感染和疾病发展。 接口结构化学，蛋白质建模，分子遗传学， 细胞生物学和病毒学，如这里所描述的代表了一种新的方法， 病毒致病机理的研究。 这也将导致更好的理解 并进一步开发用于食品的转基因抗性 生产以减少限制食物的严重病毒感染 这加剧了世界许多地区的饥饿。