绿盲蝽G蛋白偶联受体激酶2基因（AlGRK2）的表达分析及在绿盲蝽生长发育中的功能

doi:10.3864/j.issn.0578-1752.210335

摘要/Abstract

摘要： 【目的】克隆绿盲蝽（Apolygus lucorum）G蛋白偶联受体激酶2基因（AlGRK2）cDNA序列，明确其时空表达谱，阐明外源蜕皮激素（20E）对AlGRK2表达的影响，分析AlGRK2在绿盲蝽若虫发育中的作用，为进一步研究其在蜕皮激素信号通路中的功能打下基础。【方法】RACE法克隆获得AlGRK2全长，qRT-PCR分析不同日龄绿盲蝽及雌成虫不同组织中AlGRK2基因的表达谱，分析外源20E诱导及RNAi处理后，AlGRK2的应答反应及对绿盲蝽生长发育的影响。【结果】AlGRK2全长2 715 bp，开放阅读框2 106 bp，编码701个氨基酸；蛋白结构分析显示AlGRK2包含4个结构域，具有GRK2蛋白的典型结构域（PH结构域）；AlGRK2在绿盲蝽1—16日龄虫体内均有表达，并以在1日龄表达量最高，在14日龄表达量最低；AlGRK2在绿盲蝽雌成虫卵巢、中肠及脂肪体中高表达，在胸与足中的表达较低；外源20E处理后，AlGRK2在绿盲蝽1日龄和3日龄表达量显著下调，AlGRK2在雌成虫各组织中均表达上调，在卵巢及脂肪体中上调幅度最大；而绿盲蝽若虫发育历期、末龄若虫体重和成虫羽化率均显著下降；此外，与dsGFP处理组相比，dsAlGRK2处理后绿盲蝽的AlGRK2表达水平显著下降，若虫死亡率及发育历期显著增加，而成虫羽化率和5龄若虫体重显著下降。【结论】AlGRK2在绿盲蝽体内的表达谱显示出发育阶段特异性和组织特异性；外源20E处理可抑制AlGRK2的表达，并对绿盲蝽生长发育有不利影响，表现为加快绿盲蝽的发育进度、降低5龄若虫体重及成虫羽化率；此外，RNAi也显示出相似的效应。

关键词: 绿盲蝽, G蛋白偶联受体激酶2, 基因克隆, 表达量, RNA干扰

Abstract: 【Objective】In this study, we cloned the full-length cDNA of the G protein-coupled receptor kinase 2 (AlGRK2) in Apolygus lucorum, and analyzed its expression profiles. We also clarified the effect of AlGRK2 expression under exogenous ecdysterone hormone 20E in A. lucorum and the role of AlGRK2 in the development of A. lucorum nymphs, laying the foundation for further research on its function in the ecdysone signaling pathway.【Method】We cloned and obtained the AlGRK2 in A. lucorum by RACE method. Using the qRT-PCR method, we determined the relative expression levels of AlGRK2 in different days old (1-16 day) and different tissues (head, thorax, wings, legs, midgut, ovary and fat body) in female adults in A. lucorum. Finally, We analyzed the response of the AlGRK2 in A. lucorum after exogenous 20E induction and RNAi treatment and its effects on the developmental duration of the nymphs, the weight of the 5th instar nymphs and the emergence rate of adults.【Result】The full length of AlGRK2 cDNA is 2 715 bp, and ORF is 2 106 bp. ExPASy predicts that the protein molecular weight is 80.2 kD. Protein structure analysis shows that AlGRK2 contains 4 domains and has the typical domain in the GRK2 protein (PH domain). qRT-PCR results showed that AlGRK2 was expressed in 1-day-old to 16 day-old nymphs of A. lucorum, in which the expression level was the highest at 1th day-old and the lowest at 14-day-old. The AlGRK2 gene was highly expressed in the ovary, midgut, and fat body of female adults and less expressed in the thorax and legs. After exogenous 20E treatment, the expression of AlGRK2 was significantly down-regulated in nymphs at the 1-day-old and the 3-day-old. The relative expression of AlGRK2 in all tissues of females was up-regulated after treating with 20E which compared with control, especially in ovary and fat body. Compared with CK, the developmental duration of nymph, the body weight of the last instar nymph and the emergence rate of adult in A. lucorum were significantly decreased after 20E treatment. In addition, AlGRK2 expression level of A. lucorum significantly decreased in the dsAlGRK2 group compared with the dsGFP group. Compared with the dsGFP group, the mortality rate and the 5th instar developmental period of the dsAlGRK2 group of A. lucorum nymph increased significantly, while the adult emergence rate and the 5th instar nymph body weight decreased significantly.【Conclusion】The expression profile of AlGRK2 in A. lucorum showed the specificity of the developmental stage and the tissue. Exogenous 20E treatment can inhibit the expression of AlGRK2 gene in A. lucorum and delay the growth and development of A. lucorum, and RNAi also shows similar effects. The results provide a theoretical basis for revealing the function of GRK2 and perfecting the ecdysone signal pathway, and laying a foundation for providing new targets for pest control.

Key words: Apolygus lucorum, G protein-coupled receptor kinase 2, gene cloning, relative expression, RNAi

谭永安, 姜义平, 赵静, 肖留斌. 绿盲蝽G蛋白偶联受体激酶2基因（AlGRK2）的表达分析及在绿盲蝽生长发育中的功能[J]. 中国农业科学, 0, (): 1-.

TAN YongAn, Jiang YiPing, ZHAO Jing, XIAO LiuBin. Expression profile of G protein-coupled receptor kinase 2 gene (AlGRK2) and its function in the development of Apolygus lucorum[J]. Scientia Agricultura Sinica, 0, (): 1-.

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绿盲蝽G蛋白偶联受体激酶2基因（AlGRK2）的表达分析及在绿盲蝽生长发育中的功能

Expression profile of G protein-coupled receptor kinase 2 gene (AlGRK2) and its function in the development of Apolygus lucorum

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