褐飞虱神经肽及其受体基因的功能筛查

doi:10.3785/j.issn.1008-9209.2022.06.292

浙江大学学报（农业与生命科学版）

2022, Vol. 48

Issue (6): 766-775 DOI: 10.3785/j.issn.1008-9209.2022.06.292

研究论文

褐飞虱神经肽及其受体基因的功能筛查

王斯亮^1,²(

),罗序梅¹,张传溪^1,³(

)

^1.浙江大学农业与生物技术学院昆虫科学研究所，杭州 310058
^2.温州科技职业学院农业与生物技术学院，浙江温州 325006
^3.宁波大学植物病毒学研究所，农产品质量安全危害因子与风险防控国家重点实验室/农业农村部和浙江省植物保护生物技术重点实验室，浙江宁波 315211

Screening the function of genes encoding neuropeptides and their receptors in Nilaparvata lugens

Siliang WANG^1,²(

),Xumei LUO¹,Chuanxi ZHANG^1,³(

)

^1.Institute of Insect Sciences, College of Agriculture and Biotechnology, Zhejiang University, Hangzhou 310058, China
^2.College of Agriculture and Biotechnology, Wenzhou Vocational College of Science and Technology, Wenzhou 325006, Zhejiang, China
^3.State Key Laboratory for Managing Biotic and Chemical Threats to the Quality and Safety of Agro-Products/Key Laboratory of Biotechnology in Plant Protection of Ministry of Agriculture and Rural Affairs and Zhejiang Province, Institute of Plant Virology, Ningbo University, Ningbo 315211, Zhejiang, China

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摘要：

神经肽对昆虫的生命活动和环境适应性具有重要影响，是害虫防治的潜在靶标。褐飞虱（Nilaparvata lugens）是亚洲地区一种重要的水稻害虫。本研究对褐飞虱神经肽及其受体基因的功能进行了筛查，通过聚合酶链反应扩增并验证得到了41个神经肽基因（含1个可变剪接本）和44个受体基因。通过RNA干扰方法，得到沉默后造成高死亡率的4个神经肽基因（NlCCAP、NlETH、NlOKA和NlPK）和2个受体基因（NlA36和NlA46），这些基因在褐飞虱3龄若虫阶段注射双链RNA（double-stranded RNA, dsRNA）进行沉默后，成虫相对存活率＜0.2，具有成为害虫防治靶标的潜力。RNA干扰结果表明：几种蜕皮前后行为调控神经肽，如甲壳类动心肽、蜕皮诱导激素、鞣化激素的基因与其他昆虫研究中报道的功能相似，而另一些与蜕皮调控相关的神经肽（如促前胸腺激素、羽化激素）在褐飞虱中的生理功能则不明显。本研究为褐飞虱神经肽生理功能的深入探索奠定了基础。

关键词： 褐飞虱; 神经肽; RNA干扰; 蜕皮

Abstract:

Neuropeptides are crucial for life activities and environmental fitness of insects, which are potential targets for pest control. The brown planthopper (Nilaparvata lugens) is an important pest on rice in Asia. Thus, functional screening analysis of genes encoding neuropeptides and their receptors in N. lugens were conducted. The cDNA sequences of 41 neuropeptides (containing one alternative splicing transcript) and 44 receptors were verified by polymerase chain reaction (PCR). Four neuropeptide genes (NlCCAP, NlETH, NlOKA and NlPK) and two receptor genes (NlA36 and NlA46) were identified to cause high mortality through RNA interference (RNAi), showing potential in pest control. These genes knockdown resulted in the relative survival rate of adult less than 0.2 when injected with double-stranded RNA (dsRNA) at the third instar, and had the potential to be target genes for pest control. The results of RNAi also demonstrated that neuropeptides, such as crustacean cardioactive peptide (CCAP), ecdysis triggering hormone (ETH) and bursicon were conserved in physiological function, while other neuropeptides (eg. prothoracicotropic hormone, eclosion hormone) which related to ecdysis in other insects were undefined in physiological function in N. lugens. Our study provides a basis for further exploration of neuropeptides’ physiological function in N. lugens.

Key words: Nilaparvata lugens neuropeptide RNA interference ecdysis

收稿日期: 2022-06-29 出版日期: 2022-12-27

CLC:

S 433.3

基金资助: 国家自然科学基金项目(31871954);温州科技职业学院博士科研启动项目

通讯作者: 张传溪 E-mail: wsl512935@163.com;chxzhang@zju.edu.cn

作者简介: 王斯亮（https://orcid.org/0000-0001-6831-0652），E-mail：wsl512935@163.com

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引用本文:

王斯亮,罗序梅,张传溪. 褐飞虱神经肽及其受体基因的功能筛查[J]. 浙江大学学报（农业与生命科学版）, 2022, 48(6): 766-775.

Siliang WANG,Xumei LUO,Chuanxi ZHANG. Screening the function of genes encoding neuropeptides and their receptors in Nilaparvata lugens. Journal of Zhejiang University (Agriculture and Life Sciences), 2022, 48(6): 766-775.

链接本文:

https://www.zjujournals.com/agr/CN/10.3785/j.issn.1008-9209.2022.06.292 或 https://www.zjujournals.com/agr/CN/Y2022/V48/I6/766

表1 褐飞虱神经肽基因的RNA干扰效果

表2 褐飞虱神经肽受体基因的RNA干扰效果

图1 褐飞虱在 NlETH 基因经RNA干扰后的存活率和表型

图2 褐飞虱在 NlCCAP 基因经RNA干扰后的存活率和表型A. 褐飞虱在不同龄期注射dsNlCCAP后的存活率；B. 褐飞虱在NlCCAP基因经RNA干扰后的表型。

图3 注射不同的dsRNA后编码鞣化激素的2个亚基的基因表达量变化

图4 褐飞虱在 Nlburs - a 和 Nlburs - b 基因经RNA干扰后的存活率和表型A.褐飞虱在不同龄期注射dsNlburs-a和dsNlburs-b后的存活率；B.褐飞虱在Nlburs-a和Nlburs-b基因经RNA干扰后的表型。

图5 褐飞虱不同龄期干扰鞣化激素基因对成虫表型（A）及成虫卷翅率（B）的影响

图6 褐飞虱在 NlA46 基因经RNA干扰后的存活率和表型

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