多异瓢虫气味受体HvarOR21的基因克隆及配体结合特性分析  

Gene cloning and ligand binding characterization of the odorant receptor HvarOR21 in the variegated lady beetle,Hippodamia variegata(Coleoptera:Coccinellidae)

作　　者：谢佼昕 唐浩宇 赵丹阳 羿超群 刘晓旭 张永军[2] XIE Jiao-Xin;TANG Hao-Yu;ZHAO Dan-Yang;YI Chao-Qun;LIU Xiao-Xu;ZHANG Yong-Jun(College of Animal Science,Shanxi Agricultural University,Taigu 030801,China;State Key Laboratory for Biology of Plant Diseases and Insect Pests,Institute of Plant Protection,Chinese Academy of Agricultural Sciences,Beijing 100193,China;College of Plant Protection,Agricultural University of Hebei,Baoding 071000,China)

机构地区：[1]山西农业大学动物科学学院,太谷030801 [2]中国农业科学院植物保护研究所,植物病虫害生物学国家重点实验室,北京100193 [3]河北农业大学植物保护学院,保定071000

出　　处：《昆虫学报》2024年第10期1307-1317,共11页Acta Entomologica Sinica

基　　金：国家自然科学基金面上项目(31972338,32372639);国家重点研发计划项目(2021YFD1400700);山西农业大学引进人才科研启动工程项目(2023BQ07)。

摘　　要：【目的】克隆多异瓢虫Hippodamia variegata触角中高表达的气味受体基因HvarOR21,明确HvarOR21的配体结合特性,为揭示多异瓢虫定位猎物栖境的嗅觉识别机制提供理论基础。【方法】基于多异瓢虫成虫触角转录组测序数据与气味受体鉴定结果,通过PCR克隆具有完整开放阅读框(open reading frame,ORF)的HvarOR21 cDNA序列;利用系统发育进化分析和序列分析分别研究HvarOR21的分类及序列结构特征;采用爪蟾Xenopus卵母细胞体外表达结合双电极电压钳记录重组表达蛋白HvarOR21对66种候选气味化合物的电生理反应;通过同源性建模和分子对接模拟预测HvarOR21与癸醛的结合位点。【结果】克隆得到多异瓢虫HvarOR21(GenBank登录号:PP236119)的cDNA全长序列;HvarOR21含有7个跨膜结构域,且N末端位于胞内,C末端位于胞外,符合昆虫气味受体的典型特征,属于鞘翅目(Coleoptera)气味受体G5分支成员。重组HvarOR21对癸醛有反应,且呈剂量依赖性。HvarOR21与癸醛通过疏水作用及范德华力结合多个氨基酸残基,结合力为-22.18 kJ/mol。【结论】癸醛是棉蚜Aphis gossypii为害棉花后释放的虫害诱导植物挥发性化合物。HvarOR21特异性对癸醛有电生理反应,且与癸醛的结合力较强,提示HvarOR21在多异瓢虫定位猎物栖境中发挥作用。【Aim】To clone the odorant receptor(OR)gene HvarOR21 highly expressed in the antennae of the variegated lady beetle,Hippodamia variegata and clarify the ligand binding characteristics of HvarOR21,so as to provide a theoretical basis for revealing the recognition mechanism of the localization of prey habitats for H.variegata.【Methods】Based on the adult antennal transcriptome sequencing data and the identification results of odorant receptors of H.variegata,the cDNA sequence of HvarOR21 with a complete open reading frame(ORF)was cloned using PCR.Phylogenetic analysis and sequence analysis were used to study the classification and sequence structure characteristics of HvarOR21,respectively.Through the heterologous expression in Xenopus oocytes coupled with two-electrode voltage clamp recording,the electrophysiological responses of the recombinant HvarOR21 to 66 candidate odorant compounds were determined.Using homology modeling and molecular docking simulation analysis,the binding sites between HvarOR21 and decanal were predicted.【Results】The full-length cDNA sequence of HvarOR21(GenBank accession no.:PP236119)of H.variegata was cloned and the deduced protein has seven transmembrane domains with an intracellular N-terminus and an extracellular C-terminus,which conforms to the typical structure of insect odorant receptors,belonging to the coleopteran OR group 5 subfamily.The recombinant HvarOR21 specifically tuned to decanal in a dose-dependent manner.HvarOR21 bound multiple amino acid residues with decanal through hydrophobic interactions and van der Waals forces,with the binding energy of-22.18 kJ/mol.【Conclusion】Decanal is a volatile compound emitted from cotton plants infested by Aphis gossypii.HvarOR21 has a specific electrophysiological response to decanal with strong binding affinity,suggesting that HvarOR21 plays an important role in the localization of prey habitats for H.variegata.

关 键 词：多异瓢虫 气味受体 双电极电压钳 癸醛 分子对接 

分 类 号：Q966[生物学—昆虫学]