硫激肽及其受体调控褐飞虱取食行为  

Sulfakinin and its receptor regulate the feeding behavior of Nilaparvata lugens (Hemiptera: Delphacidae)

作　　者：郭迪 张素 李剑 高聪芬[1,2] 吴顺凡 GUO Di;ZHANG Su;LI Jian;GAO Cong-Fen;WU Shun-Fan(College of Plant Protection,Nanjing Agricultural University,Nanjing 210095,China;State&Local Joint Engineering Research Center of Green Pesticide Invention and Application,Nanjing 210095,China;Sanya Institute of Nanjing Agricultural University,Sanya 572025,China)

机构地区：[1]南京农业大学植物保护学院,南京210095 [2]绿色农药创制与应用技术国家地方联合工程研究中心,南京210095 [3]南京农业大学三亚研究院,三亚572025

出　　处：《昆虫学报》2023年第3期277-291,共15页Acta Entomologica Sinica

基　　金：国家自然科学基金优秀青年科学基金项目(32022011);国家自然科学基金面上项目(31772205)。

摘　　要：【目的】明确硫激肽(sulfakinin, SK)及硫激肽受体(sulfakinin receptor, SKR)在褐飞虱Nilaparvata lugens取食行为中的作用。【方法】PCR克隆褐飞虱硫激肽基因Nlsk及其受体基因Nlskr cDNA全长序列并进行生物信息学分析;利用qRT-PCR检测Nlsk及Nlskr在褐飞虱不同发育阶段(卵、1-5龄若虫、雄成虫和雌成虫)和雌成虫不同组织(头、触角、翅、口针、足、肠道和马氏管)中的表达量;褐飞虱3龄若虫注射dsNlskr进行基因沉默,qRT-PCR检测4龄若虫中Nlskr的表达量,测定4龄若虫的取食量;基于已构建的Nlskr RNAi后的4龄若虫转录组数据库进行差异表达基因(differentially expressed genes, DEGs)的GO和KEGG分析以及取食相关基因的qRT-PCR验证。【结果】PCR克隆得到了褐飞虱Nlsk(GenBank登录号:AB817281)及Nlskr(GenBank登录号:BAO01059.1)的cDNA全长序列。序列比对结果显示,褐飞虱NlSK成熟肽具有与其他物种保守的C端FMRFamide型多肽结构。NlSKR与其他昆虫同源受体存在高度保守的跨膜结构域。qRT-PCR检测结果表明,Nlsk与Nlskr在卵和1龄若虫中高表达,两基因均主要在头部表达,Nlskr还在褐飞虱口针中高表达。RNAi沉默Nlskr可显著提高褐飞虱4龄若虫的取食量;基于转录组数据DEGs的GO和KEGG分析结果表明,RNAi沉默Nlskr可显著影响褐飞虱嗅觉、味觉、能量代谢以及取食相关神经肽及受体基因的表达;取食相关基因的qRT-PCR验证结果表明,沉默Nlskr降低Nl7tmOR,NlOAR-3R,NlUH-FAF和NlTRP-161A的表达量,而提高NlGr64f,NlUE-E2和NlTHR的表达量。【结论】本研究明确了硫激肽及其受体参与调控褐飞虱取食行为,为害虫取食行为抑制剂研发提供了潜在靶点。【Aim】To clarify the function of sulfakinin(SK)and sulfakinin receptor(SKR)in the feeding behavior of Nilaparvata lugens.【Methods】The full-length cDNA sequences of sulfakinin gene Nlsk and sulfakinin receptor gene Nlskr of N.lugens were cloned by PCR and subjected to bioinformatical analysis.The expression levels of Nlsk and Nlskr in different developmental stages(egg,1st-5th instar nymphs,and male and female adults)and different tissues(head,antenna,wing,proboscis,leg,gut and Malpighian tubules)of the female adult of N.lugens were analyzed by qRT-PCR.The 3rd instar nymphs of N.lugens were injected with ds Nlskr for gene silencing and the expression level of Nlskr in the 4th instar nymph was detected by qRT-PCR.The food intake of the 4th instar nymph after the Nlskr silencing was measured.GO and KEGG analyses of differentially expressed genes(DEGs)and qRT-PCR verification of feeding-related genes based on the previously constructed transcriptome database of the 4th instar nymphs after RNAi of Nlskr were performed.【Results】The full-length cDNA sequences of Nlsk(GenBank accession number:AB817281)and Nlskr(GenBank accession number:BAO01059.1)of N.lugens were cloned by PCR.Sequence alignment results showed that the NlSK mature peptide of N.lugens has a C-terminal FMRFamide polypeptide structure that is conserved with other species.NlSKR has a highly conserved transmembrane domain with homologous receptors of other insects.The results of qRT-PCR showed that Nlsk and Nlskr were highly expressed in the egg and 1st instar nymph,and mainly in the head.Nlskr was also highly expressed in the proboscis of N.lugens.Silencing Nlskr significantly increased the food intake of the 4th instar nymphs of N.lugens.Based on transcriptome data,GO and KEGG analysis result of DEGs showed that silencing of Nlskr by RNAi significantly affected the expression of the olfactory,gustation,energy metabolism,and feeding-related neuropeptides and receptor genes.The qRT-PCR verification results of feeding-related genes showed that silen

关 键 词：褐飞虱 硫激肽 硫激肽受体 转录组 取食行为 

分 类 号：Q966[生物学—昆虫学]