REGULATION OF POST-TRANSCRIPTIONAL GENE SILENCING

描述(来自应用程序)：转录后基因沉默(PTGS) 是一种古老的真核生物调节机制，在这种机制中，特定的RNA 序列被锁定并被摧毁。尽管PTGS存在于不同的生物中， 该途径的细胞成分及其调控不是很好。 明白了。植物马铃薯Y病毒的辅助成分蛋白酶(HG-Pro)具有 已被证明能抑制植物中的PTGS。HG-Pro作为诱饵在酵母中的应用 双杂交系统中，我们发现了一种新的钙调蛋白相关蛋白 (称为rgsCaM)与PTGS的这种病毒抑制物相互作用。RGS-Cam， 像HG-Pro本身一样，抑制PTGS的启动和维持。我们的 假设PTGS的HC-Pro抑制是由其相互作用介导的 用RGS-Cam。在这里，我们建议利用HC-Pro和RGS-Cam作为分析工具 植物中沉默的机制。提案的前四个目标重点 关于RGS-CaM在PTGS中的作用，识别需要的蛋白质结构域 抑制PTG以及参与RGS-Cam/HC-Pro相互作用的PTG。 将使用多种方法来干扰RGS-CaM和RGS-CaM的表达 这些操作对PTGS的影响以及HC-Pro对其的抑制作用 化验过了。第三个目标是解决机械性问题，确定如何和 其中HC-Pro和RGS-CaM作用于PTGS中的已知步骤。最后两个 旨在研究其他植物蛋白在HC-Pro介导的蛋白质合成中的作用 抑制PTGS。我们将扩大对另外三个人的调查 使用成功鉴定HC-Pro相互作用蛋白质的方法 RGS-CaM作为PTGS的调节因子。基因芯片技术将用于鉴定 响应PTGS或被HC-Pro逆转而调节的基因。 最后，我们将开发模式遗传生物拟南芥 筛选干扰PTGS抑制的突变体 HC-Pro。植物消声系统可以作为理解类似情况的模型 在动物中的途径，并有潜力被开发用于基因治疗 基因表达的应用和操纵。考虑到它的抗病毒特性 植物中的基因沉默，了解植物中的PTGS很可能导致 人类抗病毒策略的发展。

DESCRIPTION (from the application): Post-transcriptional gene silencing (PTGS) is an ancient eukaryotic regulatory mechanism in which a particular RNA sequence is targeted and destroyed. Although PTGS occurs in diverse organisms, the cellular components of the pathway and its regulation are not well understood. The helper component proteinase (HG-Pro) of plant potyviruses has been shown to suppress PTGS in plants. Using HG-Pro as bait in a yeast two-hybrid system, we have identified a novel calmodulin-related protein (termed rgsCaM) that interacts with this viral suppressor of PTGS. rgs-CaM, like HG-Pro itself, suppresses both initiation and maintenance of PTGS. Our hypothesis is that HC-Pro-suppression of PTGS is mediated by its interaction with rgs-CaM. Here we propose to exploit HC-Pro and rgs-CaM as tools to dissect the mechanism of silencing in plants. The first four aims of the proposal focus on the role of rgs-CaM in PTGS, identifying domains of the protein required to suppress PTGS as well as those involved in the rgs-CaM/HC-Pro interaction. Several approaches will be used to interfere with expression of rgs-CaM and the effect of these manipulations on PTGS and its suppression by HC-Pro will be assayed. The third aim addresses mechanistic questions, determining how and where HC-Pro and rgs-CaM act with regard to known steps in PTGS. The last two aims focus on the role of other plant proteins in the HC-Pro-mediated suppression of PTGS. We will extend investigations of three other HC-Pro-interacting proteins using the approaches that successfully identified rgs-CaM as a regulator of PTGS. Gene array technology will be used to identify genes that are regulated in response to PTGS or its reversal by HC-Pro. Finally, we will exploit the model genetic organism Arabidopsis thalianato screen for mutants that interfere with suppression of PTGS in response to HC-Pro. The plant silencing system serves as a model to understand similar pathways in animals, and has potential to be exploited for gene therapy applications and manipulation of gene expression. Given the antiviral nature of gene silencing in plants, understanding PTGS in plants could well lead to the development of antiviral strategies in humans.