Department of Entomology, Cornell University, Geneva, NY, 14456, USA; School of Life and Environmental Sciences, Hangzhou Normal University, Hangzhou, China.
Department of Entomology, Cornell University, Geneva, NY, 14456, USA.
Insect Biochem Mol Biol. 2018 Nov;102:59-66. doi: 10.1016/j.ibmb.2018.09.013. Epub 2018 Sep 29.
Adoption of biotech crops engineered to express insecticidal toxins from Bacillus thuringiensis (Bt) has revolutionized insect pest control in agriculture. For continuing effective application and development of the environmentally friendly Bt biotechnology, it is fundamental to understand pathways of toxicity of Bt toxins in insects. In this study, mutations were introduced in the midgut cadherin gene in the cabbage looper, Trichoplusia ni, by CRISPR/Cas9 mutagenesis. T. ni strains with mutations in the genes of two major receptors for Bt toxins, the midgut cadherin and ABCC2, and three Cry1A toxins with shared and differential midgut binding sites were used as an experimental system to dissect the roles of the cadherin and ABCC2 in the pathways of toxicity of Bt toxins. Results from assays of responses of the T. ni strains to different Bt toxins revealed that the cadherin and ABCC2 play independent roles in the mode of action of Cry1A toxins and that Bt toxins exert insecticidal activity through multiple redundant pathways of toxicity in insects. Besides the cadherin and ABCC2 pathways, there exists an additional major pathway of toxicity to be identified for Cry1Aa. The results also confirmed that the toxicity of Cry2Ab involves neither the cadherin nor the ABCC2 protein. The multiple pathway model for Bt toxins clarified from this study provided new insights into the molecular modes of action of Bt toxins and mechanisms of insect resistance to Bt toxins.
采用苏云金芽孢杆菌（Bt）表达的杀虫毒素工程化的生物技术作物的采用彻底改变了农业害虫防治。为了继续有效应用和开发环保的 Bt 生物技术，了解 Bt 毒素在昆虫中的毒性途径是至关重要的。在这项研究中，通过 CRISPR/Cas9 诱变技术引入了小菜蛾（Trichoplusia ni）中肠钙粘蛋白基因的突变。使用具有 Bt 毒素两个主要受体（中肠钙粘蛋白和 ABCC2）基因突变和三个具有共享和差异的中肠结合位点的 Cry1A 毒素的 T. ni 菌株作为实验系统，剖析钙粘蛋白和 ABCC2 在 Bt 毒素毒性途径中的作用。对不同 Bt 毒素的 T. ni 菌株反应的测定结果表明，钙粘蛋白和 ABCC2 在 Cry1A 毒素的作用模式中发挥独立的作用，并且 Bt 毒素通过昆虫中多个冗余的毒性途径发挥杀虫活性。除了钙粘蛋白和 ABCC2 途径外，还存在有待鉴定的 Cry1Aa 的另一个主要毒性途径。该结果还证实了 Cry2Ab 的毒性既不涉及钙粘蛋白也不涉及 ABCC2 蛋白。本研究阐明的 Bt 毒素多途径模型为 Bt 毒素的分子作用模式和昆虫对 Bt 毒素的抗性机制提供了新的见解。
