Xiao Yutao, Dai Qing, Hu Ruqin, Pacheco Sabino, Yang Yongbo, Liang Gemei, Soberón Mario, Bravo Alejandra, Liu Kaiyu, Wu Kongming
From the State Key Laboratory for Biology of Plant Disease and Insect Pests, Chinese Academy of Agricultural Sciences, West Yuanmingyuan Road, Beijing 100193, China.
the Agricultural Genomics Institute at Shenzhen, Chinese Academy of Agricultural Sciences, Shenzhen 518120, China.
J Biol Chem. 2017 Feb 17;292(7):2933-2943. doi: 10.1074/jbc.M116.768671. Epub 2017 Jan 12.
Transgenic plants that produce (Bt) crystalline (Cry) toxins are cultivated worldwide to control insect pests. Resistance to toxins threatens this technology, and although different resistance mechanisms have been identified, some have not been completely elucidated. To gain new insights into these mechanisms, we performed multiple back-crossing from a 3000-fold Cry1Ac-resistant BtR strain from cotton bollworm (), isolating a 516-fold Cry1Ac-resistant strain (96CAD). Cry1Ac resistance in 96CAD was tightly linked to a mutant cadherin allele (mHaCad) that contained 35 amino acid substitutions compared with HaCad from a susceptible strain (96S). We observed significantly reduced levels of the mHaCad protein on the surface of the midgut epithelium in 96CAD as compared with 96S. Expression of both cadherin alleles from 96CAD and 96S in insect cells and immunofluorescence localization in insect midgut tissue sections showed that the HaCAD protein from 96S localizes on the cell membrane, whereas the mutant 96CAD-mHaCad was retained in the endoplasmic reticulum (ER). Mapping of the mutations identified a D172G substitution mainly responsible for cadherin mislocalization. Our finding of a mutation affecting membrane receptor trafficking represents an unusual and previously unrecognized resistance mechanism.
产生苏云金芽孢杆菌（Bt）晶体（Cry）毒素的转基因植物在全球范围内种植以防治害虫。对Bt毒素的抗性威胁到这项技术，尽管已经确定了不同的抗性机制，但有些尚未完全阐明。为了深入了解这些机制，我们对棉铃虫的一个对Cry1Ac具有3000倍抗性的BtR品系进行了多次回交，分离出一个对Cry1Ac具有516倍抗性的品系（96CAD）。96CAD中对Cry1Ac的抗性与一个突变的钙粘蛋白等位基因（mHaCad）紧密相关，与敏感品系（96S）的HaCad相比，该等位基因含有35个氨基酸替换。与96S相比，我们观察到96CAD中肠上皮细胞表面的mHaCad蛋白水平显著降低。在昆虫细胞中表达96CAD和96S的钙粘蛋白等位基因，并在昆虫中肠组织切片中进行免疫荧光定位，结果表明96S的HaCAD蛋白定位于细胞膜，而突变体96CAD - mHaCad则保留在内质网（ER）中。对突变的定位确定了一个主要导致钙粘蛋白错误定位的D172G替换。我们发现一个影响膜受体运输的突变代表了一种不同寻常且以前未被认识到的Bt抗性机制。
