Regulatory Mechanisms of Cucumber Mosaic Virus Replication

这项提议的主题是黄瓜花叶病毒(CMV)，这是一种寄主范围很广的植物病毒，对它几乎没有天然的抗性。这个项目的目标是：(1)定位病毒基因组中参与复制速度调节和高温抑制复制的序列；(2)确定受各种基因改变影响的植物细胞(原生质体)差异复制的潜在机制；以及(3)确定各种基因改变对病毒复制酶体外调控的影响。这些目标将通过使用CMV株系的生物活性DNA克隆来实现，这些克隆显示出在几个葫芦科寄主中对CMV(和卫星)RNA复制的各个方面的调控差异。控制各种复制相关表型的序列将通过DNA克隆的重组和定点突变来定位。这些序列替换所改变的特定功能将通过分析体外感染的原生质体中的复制产物以及在实验条件下提供适当模板并孵育的纯化的病毒复制酶来确定。这些分析将把病毒蛋白质结构的差异与特定的功能联系起来。

The subject of this proposal is cucumber mosaic virus (CMV), a wide host-range plant virus for which there is very little natural resistance available. The goals of this project are: (1) to localize sequences in the viral genome involved in the regulation of the rate of replication, and the inhibition of replication by high temperatures; (2) to determine the underlying mechanisms of differential replication in plant cells (protoplast) affected by various genetic alterations; and (3) to determine the regulatory effects of various genetic alterations on the viral replicase in vitro. These goals will be achieved by using biologically-active DNA clones of strains of CMV, that show differences in the regulation various aspects of CMV (and satellite) RNA replication in several cucurbit hosts. The sequences controlling the various replicate-related phenotypes will be mapped by recombination and site-directed mutagenesis of the DNA clones. The specific functions altered by these sequence substitutions will be determined by analyses of the replication products both in in vitro infected protoplasts, as well as of the purified viral replicase provided with the appropriate templates and incubated under the experimental conditions. These analyses will relate differences in viral protein structure to specific functions.***//