褐飞虱自噬相关基因NlATG4的表达与功能

doi:10.16409/j.cnki.2095-039x.2022.01.011

摘要/Abstract

摘要： ATG4是一种半胱氨酸蛋白酶，负责ATG8蛋白羧基末端的切割，在自噬体形成过程中发挥重要作用。本研究克隆得到褐飞虱ATG4基因（NlATG4）的cDNA全长序列（GenBank登录号:MF062502.1）。RT-qPCR分析结果表明，NlATG4基因在褐飞虱的若虫和成虫时期均有表达，1～2龄若虫的表达量最高，在其他发育阶段的表达量稍低。RNAi结果显示，在dsRNA注射后的第4 d，靶标基因NlATG4的表达量显著下降（仅为dsGFP对照组的22%），同时，NlATG4的RNA干扰导致虫体ATP含量显著降低，仅为1.0 μmol/(L·mg蛋白^-1)，为对照组的25%；在注射后的第6 d，dsNlATG4处理组的褐飞虱存活率出现显著降低，其脂肪体组织变得松散，大型脂滴（直径≥5.0 μm）数量增多，类酵母共生菌周围泡状结构的界限基本消失；在注射后的第8 d，dsNlATG4处理的褐飞虱的存活率下降为36.7%，比dsGFP对照组的存活率下降53.3%。本研究表明靶向NlATG4基因的RNA干扰会破坏褐飞虱脂肪体的结构完整性和类酵母共生菌的微环境，影响脂类物质代谢和ATP含量，并导致褐飞虱存活率降低。因此，NlATG4基因在褐飞虱正常生长发育中发挥重要作用，在褐飞虱防治中具有一定的潜力。

关键词: 褐飞虱, NlATG4基因, RNAi, 脂肪体

Abstract: ATG4, a cysteine protease, is responsible for the cleavage of the carboxyl terminal of ATG8 protein and plays an important role in the formation of autophagosomes. In this study, the full-length cDNA of ATG4 gene in the Nilaparvata lugens (NlATG4) was cloned (GenBank Accession No. MF062502.1). RT-qPCR analysis showed that the NlATG4 gene was expressed in both nymphs and adults and the expression level in the 1^st to 2^nd instar nymphs was the highest and slightly lower at other developmental stages. RNAi results showed that, at 4 d post treatment of dsRNA injection, the expression level of the target gene NlATG4 decreased significantly (about 22% of that of the dsGFP control) and the ATP content also significantly decreased to only 1.0 μmol/(L·mg protein^-1), about 25% of that of the dsGFP control. At 6 d post dsRNA treatment, the survival rate of the dsNlATG4-treated N. lugens decreased significantly in contrast to that of the dsGFP control, the fat body became looser, and the number of large lipid droplets (diameter ≥5.0 μm) increased, and the boundary of the bubble structure enclosing the yeast-like symbiont almost disappeared in dsNlATG4 treatment group. At 8 d post dsRNA treatment, the survival rate of dsNlATG4-treated N. lugens was 36.7%, 53.3% lower than that for the dsGFP control. This study shows that RNA interference targeting NlATG4 gene breaks the structural integrity of fat body and the microenvironment of yeast-like symbiont, affecting lipid metabolism and ATP content, and reduces the survival of N. lugens. Therefore, NlATG4 gene plays an important role in the normal growth and development of N. lugens and has some potential in the control of N. lugens.

Key words: Nilaparvata lugens, NlATG4, RNAi, fat body

中图分类号:

矫启启, 吴建艮, 冯娅琳, 俞飞飞, 陈桐桐, 郑园园, 郝培应, 俞晓平. 褐飞虱自噬相关基因NlATG4的表达与功能[J]. 中国生物防治学报, 2023, 39(1): 111-121.

JIAO Qiqi, WU Jiangen, FENG Yalin, YU Feifei, CHEN Tongtong, ZHENG Yuanyuan, HAO Peiying, YU Xiaoping. Expression and Function Analysis of Autophagy-related Gene NlATG4 in Brown Planthopper, Nilaparvata lugens (Hemiptera: Delphacidae)[J]. Chinese Journal of Biological Control, 2023, 39(1): 111-121.

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