棉铃虫V-ATPase亚基B是Cry1Ac的功能受体  

V-ATPase Subunit B is a Functional Receptor of Cry1Ac in Helicoverpa armigera(Lepidoptera:Noctuidae)

作　　者：段云鹏 姚雪 李品 王萌涵 胡守印 胡恩静 刘永刚 杨淑芳[1] 魏纪珍 DUAN Yunpeng;YAO Xue;LI Pin;WANG Menghan;HU Shouyin;HU Enjing;LIU Yonggang;YANG Shufang;WEI Jizhen(College of Plant Protection,Henan Agricultural University/Henan International Laboratory for Green Pest Control,Zhengzhou 450002,China)

机构地区：[1]河南农业大学植物保护学院/河南省害虫绿色防控国际联合实验室

出　　处：《中国生物防治学报》2022年第5期1094-1102,共9页Chinese Journal of Biological Control

基　　金：国家自然科学基金(32172401);河南省农业领域科技攻关项目(212102110137);河南省大学生创业训练项目(S202110466023)。

摘　　要：目前对转苏云金芽胞杆菌Bacillus thuringiensis(Bt)的研究发现,液泡型ATP酶(Vacuolar-type proton ATPase,V-ATPase)可能是Bt的一类新型受体。我们前期通过构建棉铃虫的中肠酵母文库筛选Cry1Ac的结合蛋白发现棉铃虫V-ATPase亚基B(V-ATPase B)可以与Cry1Ac结合。为明确V-ATPase B在Cry1Ac毒力和昆虫对Cry1Ac抗性机制中的作用,本研究首先采用实时荧光定量PCR技术分析了该基因在抗感Cry1Ac棉铃虫幼虫中及其受到Cry1Ac诱导时的基因表达情况;通过Ligandblot进一步地证实了其与Cry1Ac的结合特性;并通过在Sf9细胞中表达V-ATPase B的试验验证了其功能。结果表明V-ATPase B在抗性品系及受到Cry1Ac诱导时均下调表达,Ligand blot证实了V-ATPase B与Cry1Ac特异性结合;并且在昆虫细胞内过表达该基因,会增强Cry1Ac的细胞毒力。研究结果表明棉铃虫V-ATPase B是Cry1Ac的功能受体,并有可能通过降低基因表达来参与棉铃虫对Cry1Ac的抗性形成。Current studies on the mode of action of Bacillus thuringiensis(Bt) have found that vacuolar-type proton ATPase(V-ATPase) may be a new receptor of Bt proteins.In our previous study of screening for Cry1Ac binding protein from the midgut yeast library of cotton bollworm,we found that V-ATPase subunit B(V-ATPase B) was a Cry1Ac binding receptor.To clarify the role of V-ATPase B in Cry1Ac toxicity and insect resistance to Cry1Ac,we measured the expression levels of V-ATPase B in Cry1Ac-susceptible and-resistance strains and in Cry1Ac induced larvae using real-time quantitative PCR(RT-PCR),characterized the binding of V-ATPase B with Cry1Ac protein using Ligand blot,and tested the functions of V-ATPase B in the toxicity of Cry1Ac by expressing V-ATPase B in Sf9 cells.The results showed significantly reduced expression levels of V-ATPase B in Cry1Ac-resistant and Cry1Ac-feeding H.armigera larvae.Ligand blot experiments verified the interaction between V-ATPase B and Cry1Ac protein.Importantly,overexpressing V-ATPase B in cells caused higher cytotoxicity of Cry1Ac.These results suggest that V-ATPase subunit B is a functional receptor of Cry1Ac and may be involved in resistance evolution of cotton bollworm to Cry1Ac through reducing its expression.

关 键 词：V-ATPase B 棉铃虫 CRY1AC 细胞毒力 

分 类 号：S476.1[农业科学—农业昆虫与害虫防治]