nAChRs Agβ1R81T 突变导致棉蚜对吡虫啉产生抗性的分子机制

项目名称： nAChRs Agβ1R81T 突变导致棉蚜对吡虫啉产生抗性的分子机制

项目编号： No.31301695

项目类型： 青年科学基金项目

立项/批准年度： 2014

项目学科： 农业科学

项目作者： 夏晓明

作者单位： 山东农业大学

项目金额： 23万元

中文摘要： 基于昆虫烟碱型乙酰胆碱受体(nAChRs)的吡虫啉抗性机制是目前国内外研究的热点。已有研究发现，位于nAChRs亚基上Loop D 区域的81位氨基酸高度保守，能与吡虫啉的硝基直接作用，并影响与吡虫啉的结合。我们研究表明，与敏感品系相比，室内筛选的高抗吡虫啉棉蚜品系膜蛋白与[3H]吡虫啉的结合明显降低，并在抗性品系的Agβ1亚基的Loop D区域发现一个点突变，导致81位的苏氨酸取代精氨酸（R81T）。本研究应用免疫共沉淀、昆虫杂合nAChRs表达、放射性配体结合、膜片钳、分子对接技术，重点揭示Agβ1R81T突变对棉蚜nAChRs功能和活性、结合位点结构及结合吡虫啉的影响，从分子水平深入探究Agβ1R81T突变在棉蚜对吡虫啉产生抗性中的作用。研究结果不仅可进一步揭示昆虫对吡虫啉的抗性机制，还为新烟碱类药剂的分子设计、结构改造、产品创新和抗性治理提供新思路和理论依据。

中文关键词： 棉蚜；吡虫啉；靶标突变；抗性；分子机制

英文摘要： Nicotinic acetylcholine receptor (nAChRs) mediated target resistance mechanisms of the insects to imidacloprid are currently a research hotspot at home and abroad. Previous studies indicated that nAChRs β1 subuit are highly conserved at 81 amino acid position, which can directly act with nitro group of the imidacloprid and is a key determinant of imidacloprid binding to nAChRs. Recently we have found that a much higher level of [3H]imidacloprid-specific binding to the susceptible strain than to the resistant strain of Aphis gossypii. Comparison of the nucleotide sequence of nAChRs β1subunit (Agβ1) genes from resistant and susceptible strain revealed a single point mutation in the loop D region of the resistant strain, causing an arginine to threonine substitution (R81T). In order to clarify the influence of Agβ1R81T mutation on the function, activity, and binding site structure of receptor, and the binding ability of nAChRs to imidacloprid, the recombinant hybrid nAChRs will be expressed in heterozygosis, and the co-immunoprecipitation, radioactive ligand binding, bipolar patch clamp, molecular docking study will be adopted. These researches can identify the molecular mechanisms of imidacloprid resistance in A. gossypii. These researches can not only identify the target resistance mechanisms of insects to imidac

英文关键词： Aphis gossypii；imidacloprid；target mutation；resistance；molecular mechanisms