绿盲蝽足近距离感受性信息素的分子机制

The nonvolatile or low volatile sex pheromones on insect body surface play important roles in close-range courtship. The chemosensory proteins expressed in legs may be involved in perception of close-range sex pheromones in insect mating. Present study will be conducted to comprehend the molecular basis of Apolygus lucorum legs in perception of close-range sex pheromones. Specific objectives are: using HPLC, GC-MS, combined with electrophysiological and behavioral techniques to identify the compounds released from insect body surface, which can trigger electrophysiological and behavioral responses of A.lucorum in mating behavior; identifying the sensilla types on insect legs by using scanning and transmission electron microscope; screening the specific chemosensory genes expressed in legs by using RNA-seq and RT-qPCR methods; investigating the interactions between recombinant chemosensory proteins and the candidate ligands by using ligand-binding assays and two-electrode-voltage-clamp recordings; verifying the roles of A.lucorum legs in perception of close-range sex pheromones by RNAi as well as electrophysiological and behavioral tests in vivo. Our data will provide theoretical basis for developing new compounds to regulate the mating behavior of mirid bugs.

昆虫体表非挥发性或低挥发性性信息素在两性近距离求偶过程中发挥重要作用。许多昆虫求偶时，表达于足上的化学感受相关蛋白参与了对近距离性信息素的感受。本申请以绿盲蝽为研究对象，通过HPLC、GC-MS、电生理及行为学检测等技术鉴定雌雄绿盲蝽成虫体表具有电生理和行为活性的化合物组分；利用扫描及透射电镜鉴定绿盲蝽足部化学感器类型，结合转录组测序及组织表达谱分析筛选足特异表达的化学感受相关基因；体外重组表达化学感受相关蛋白并利用结合实验及双电极电压钳筛选其配体化合物；RNAi结合电生理及求偶行为学解析化学感受相关蛋白识别低挥发及非挥发性性信息素的功能特征，阐明绿盲蝽足近距离感受性信息素的分子机制，揭示非挥发性或低挥发性性信息素对绿盲蝽短距离求偶行为的作用，为设计新型性信息素调控化合物及干扰昆虫交配策略提供理论依据。

绿盲蝽前中后足共有3种锥形感器、3种刺型感器、2种毛形感器和蒲姆氏鬃毛。绿盲蝽足部转录组中共鉴定出32个化学感受相关基因，包括20个OBPs、8个CSPs、1个AlucOR109、2个IRs、1个SNMP。其中AluOBP3、AlucOBP35以及AlucOR109在绿盲蝽足部特异表达。配体结合显示，AlucOBP3和AlucOBP35有着比较宽的结合谱，AlucOBP3能够特异性结合性信息素类似物丙烯酸异丁酯，而AlucOBP35特异结合性信息素类似物四氧代反二己烯醛，表明AlucOBP35可参与绿盲蝽的近距离求偶。AlucOBP3和AlucOBP35均能够结合花香气味β-紫罗兰酮以及蒿类活性物质乙酸2-甲基丁酯等。另外AlucOBP3和AlucOBP35对驱避剂二甲基二硫醚也有结合能力，显示AlucOBP3和AlucOBP35在绿盲蝽足部化学感受过程中的重要作用。电压钳测定显示AlucOR109对植物及昆虫体表挥发物组分反-2-己烯醛反应最大，对盲蝽类主要性信息组分及素类似物等有较大反应，另外对驱避剂二甲基二硫硫醚也有较大反应。足部电位技术（ELG）测定表明，绿盲蝽足部对乙酸叶醇酯、(E)-乙酸-2-己烯-1-醇酯、丁酸反-2-己烯酯、反-2-己烯酸4种化合物均有电生理反应。绿盲蝽足部注射dsOR109后，OR109基因表达量极显著降低。RNAi的绿盲蝽成虫足对 (E)-乙酸-2-己烯-1-醇酯的ELG反应值也显著性下降。综上，化学感受相关蛋白调控绿盲蝽足部的近距离化学感受，是绿盲蝽近距离感受行为的重要调控靶标。. 在Journal of Pest Science， Pest Management Science，Journal of Agricultural and Food Chemistry等学术期刊发表SCI论文14篇，中文核心期刊论文1篇，授权专利2项。