重要农业害虫RNA干扰效率差异机制研究

RNAi是一种新型的害虫防控策略。但RNAi效率在不同昆虫类群中差异显著，例如鞘翅目和直翅目等昆虫RNAi效率大大高于双翅目和鳞翅目昆虫，这种差异极大地制约了该技术在害虫防治中的适用性，但其差异机制尚缺乏系统研究。本项目拟选择飞蝗和亚洲玉米螟这两种干扰效率差异较大的重要农业害虫为模式，以制约RNAi效率的关键节点为线条，综合运用细胞生物学、生物化学、分子生物学和植物生物技术等多学科的方法开展比较研究：1)揭示双链RNA酶对两种昆虫中肠和血淋巴dsRNA稳定性及RNAi效率的影响；2）比较两种昆虫对外源dsRNA的吸收途径及内涵体逃逸机制；3）鉴定RNAi通路关键因子并研究其对RNAi效率的影响；4）以表达dsRNA的转基因玉米为材料，探讨植物介导表达dsRNA的抗虫机理。本项目将系统解析昆虫RNAi差异的分子机制，探索改进干扰效率的策略，以提高RNAi效率并促进其在农业害虫控制中的应用。

RNA interference (RNAi) has been widely recognized as a novel strategy for insect pest management. However, there are significant differences in RNAi efficiency among different insect taxonomic groups. For example, coleopterans and orthopterans show significantly higher RNAi efficiency than dipterans and lepidopterans. Such a difference has been regarded as one of the major limiting factors which restrict the application of RNAi for controlling different insect pests. However, we know very little about the molecular mechanisms conferring such differences. This proposed research chooses two important agricultural insect pests, Locusta migratoria (orthopteran) and Ostrinia furnacalis (lepidopteran), as models for comparative studies as they show significantly different RNAi efficiencies. We will focus on the key steps limiting the RNAi efficiency by using integrated approaches, including cell biology, biochemistry, molecular biology and plant biotechnology. We propose to: 1) investigate the biological function of different dsRNases to clarify their effect on the stability of dsRNA in the midgut and hemolymph of the two insect species; 2) compare the cellular uptake and endosomal escape of dsRNA between the two insect species; 3)identify the key genes involved in the systemic RNAi pathway, quantify their expression levels, and study their effect on RNAi efficiency; and 4) utilize the transgenic maize expressing dsRNA to elucidatethe silencing mechanism of plant-mediated RNAi in L.migratoria. This project is expected to elucidate the molecular mechanisms causing differential RNAi efficiencies in different insects, formulate new strategies to overcome the obstacle of RNAi, improve the efficiency of RNAi, and facilitate the application of RNAi-based technologies for insect pest management.