棉铃虫V-ATPase亚基B是Cry1Ac的功能受体

doi:10.16409/j.cnki.2095-039x.2022.09.005

摘要/Abstract

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Ligand blot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。

关键词: V-ATPase B, 棉铃虫, Cry1Ac, 细胞毒力

Abstract: 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.

Key words: V-ATPase B, Helicoverpa armigera, Cry1Ac, cytotoxicity

中图分类号:

S476.1

段云鹏, 姚雪, 李品, 王萌涵, 胡守印, 胡恩静, 刘永刚, 杨淑芳, 魏纪珍. 棉铃虫V-ATPase亚基B是Cry1Ac的功能受体[J]. 中国生物防治学报, 2022, 38(5): 1094-1102.

DUAN Yunpeng, YAO Xue, LI Pin, WANG Menghan, HU Shouyin, HU Enjing, LIU Yonggang, YANG Shufang, WEI Jizhen. V-ATPase Subunit B is a Functional Receptor of Cry1Ac in Helicoverpa armigera (Lepidoptera:Noctuidae)[J]. Chinese Journal of Biological Control, 2022, 38(5): 1094-1102.

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