对Cry毒素作用机理尤其是受体蛋白的研究是其抗性治理的理论基础。已报道的Cry毒素结合蛋白有多种，但进行系统证实为受体蛋白的仍不多。前期工作中，申请人发现小菜蛾V-ATP酶A亚基与Cry1Ac具有结合作用，但其是否为Cry毒素的受体蛋白并不清楚。本项目拟以小菜蛾V-ATP酶A亚基为研究对象，在分析A亚基基因在Cry毒素作用下表达量变化和体外测定毒素对V-ATPase酶活影响的基础上，利用原核表达及分子对接分析A亚基与Cry毒素作用的结合位点；并利用真核细胞表达结合细胞毒性进行功能分析，明确A亚基的作用；采用RNA干扰下调A亚基基因来探讨小菜蛾对Cry毒素的敏感性变化，分别从基因、体外、细胞和活体水平分析A亚基作为Cry毒素受体蛋白的功能，旨在为阐明Cry毒素作用机理及其抗药性治理提供理论支持。

Research on action mechanism of Cry toxins, especially on the receptor proteins, is the theoretical foundation of resistance management. Many binding proteins of Cry toxins has been identified, but a few of them have been validated as functional receptors. Our previous work found the binding of Cry1Ac with subunit A of V-ATPase from Plutella xylostella. But whether subunit A is the receptor of Cry toxins or not needs to be further validated. Hence, in this project, based on the analysis of gene expression changing of subunit A induced by Cry toxins and the test of enzyme activity of V-ATPase impacted by Cry toxins, prokaryotic expression and molecular docking will be used to analyze the binding sites of subunit A and Cry toxins. In addition, cell expression system and cytotoxicity will be carried out to verify the function of subunit A during the process of Cry toxins action,. Furthermore, RNAi will be used to study the susceptible changing of P. xylostella to toxins through down-regulation of subunit A expression. In general, the receptor function of subunit A will be identified by validation at genetic and celluar level as well as in vitro and vivo, which will provide us the theoretical basis for explanation the mode action of Cry toxins and insect resistance management.