灰飞虱lncRNA调控水稻条纹叶枯病毒复制的作用机制

Laodelphax striatellus Fallén (small brown planthopper, SBPH) is the main vector of rice stripe virus (RSV). Controlling SBPH is an efficient method for the prevention of this notorious disease. RSV can replicate within SBPH, and it is reported that long noncoding RNAs (lncRNA) of vector insects participate in host immune response to the virus and inhibit the excessive replication of virus. This project intend to study the replication mechanism of RSV regulated by lncRNA in SBPH. Here, with available RNA-Seq datasets, the following work will be carried out: 1) Identification of lncRNA gene from RNA-Seq datasets, then to find the lncRNAs differentially expressed between the viruliferous and non-viruliferous SBPH; 2) Performing the target prediction, investigating the co-expression relationship between lncRNAs and their mRNA targets; 3) The confirmed lncRNAs will be selected for full-length analysis, temporal and spatial expression pattern analysis and subcellular localization in the host immune response to the RSV; 4) knockdown of tested lncRNAs and their target gene, then the RSV virus replication and proliferation will be detected, to clarify the roles of lncRNAs during the infection and replication of RSV. The results will reveal the functions of lncRNAs in antiviral immune response, which will open up a new way to control SBPH and RSV.

灰飞虱是水稻条纹叶枯病毒(RSV)的重要传播媒介，控制灰飞虱是当前防治水稻条纹叶枯病的有效方法。RSV在灰飞虱体内能够进行复制增殖，研究表明媒介昆虫的长链非编码RNA(lncRNA)参与对病毒的免疫反应，抑制病毒的过度复制。本项目拟研究灰飞虱lncRNA调控RSV在其体内过度复制增殖的分子机制。从灰飞虱转录组中预测发现lncRNA基因，并比较分析其在带毒和不带毒灰飞虱中的差异表达；预测lncRNA的靶标，分析差异表达的lncRNA与其靶标的表达相关性；对靶向灰飞虱免疫反应相关基因的lncRNA进行全长克隆、组织和龄期及亚细胞定位，明确RSV侵染对其表达的诱导作用；对带毒灰飞虱进行lncRNA及靶标基因的干扰，检测RSV病毒的复制增殖情况，阐明lncRNA在宿主体内RSV增殖复制中的作用。预期成果将揭示灰飞虱lncRNA在RSV免疫反应中的功能，为灰飞虱及水稻条纹叶枯病的防治开辟新途径。

灰飞虱是水稻条纹叶枯病毒(RSV)的重要传播媒介，控制灰飞虱是当前防治水稻条纹叶枯病的有效方法。前期发现灰飞虱卵黄原蛋白受体参与RSV在灰飞虱体内的增殖平衡抑制，而部分长链非编码RNA在RSV侵染后显著高表达。在此基础上，本项目重点研究了灰飞虱lncRNA在调控介体内RSV复制增殖过程中的功能。在灰飞虱转录组中预测发现4786个lncRNA基因，而在带毒灰飞虱唾液腺、中肠和卵巢中发现分别有3、11、25个lncRNA表达量显著升高。其中lncRNA-1473基因与其上游的泛素结合酶基因E2（UBE2）存在顺式调控作用关系，并且具有共表达相关性。定量PCR技术检测和原位杂交实验发现该lncRNA主要在带毒灰飞虱羽化后24小时的卵巢组织巢滤泡细胞中高表达。干扰带毒灰飞虱lncRNA-1473基因及其靶标UBE2后卵巢内RSV病毒基因CP转录本丰度显著下降，后代个体的带毒水平也明显下降，表明lncRNA-1473与靶标顺式作用解除后，RSV对介体灰飞虱抗病毒免疫反应的抑制作用消失，病毒不能在卵巢内完成增殖扩散，暗示lncRNA-1473在调控介体内植物病毒侵染复制具有重要作用。研究结果有助于明确长链非编码RNA在调控植物病毒在介体侵染复制的功能，发现的关键lncRNA基因有望作为靶标基因，在未来借助转基因生物技术或RNA生物农药等绿色害虫控制技术，为灰飞虱及其传播的水稻条纹叶枯病的防治提供新思路。