| 斜纹夜蛾蜕皮激素合成通路相关CYP450基因的鉴定及表达分析 |
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| 引用本文： | 谷峻,叶艳,李时宇,袁雅菲,黄立华. 斜纹夜蛾蜕皮激素合成通路相关CYP450基因的鉴定及表达分析[J]. 昆虫学报, 2022, 65(12): 1606-1614. DOI: 10.16380/j.kcxb.2022.12.006 |
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| 作者姓名： | 谷峻 叶艳 李时宇 袁雅菲 黄立华 |
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| 作者单位： | (华南师范大学生命科学学院, 广东省昆虫发育生物学与应用技术重点实验室, 广州 510631) |
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| 摘 要： | 【目的】本研究旨在探讨蜕皮激素合成通路相关CYP450基因的表达规律，为害虫防治提供潜在的作用靶标。【方法】以家蚕Bombyx mori CYP450基因为查询序列，通过同源比对的方法从斜纹夜蛾Spodoptera litura基因组中鉴定蜕皮激素合成代谢通路中的CYP450基因，并构建其系统进化树；采用qPCR法检测鉴定的CYP450基因在斜纹夜蛾不同发育阶段(6龄幼虫、预蛹和蛹)、这3个发育阶段的不同组织(中肠、表皮和脂肪体)以及4龄幼虫取食不同寄主植物(辣椒Capsicum annuum、黄瓜Cucumis sativus、番薯Ipomoea batatas和花生Arachis hypogaea)叶片后5龄幼虫中肠中的表达量，计算4龄幼虫取食不同寄主植物叶片后发育至蛹的历期；应用PITA, miRanda, microTar和RNAhybrid 4种软件，预测调控鉴定的CYP450基因的微小RNA (miRNA)。【结果】鉴定到斜纹夜蛾蜕皮激素合成通路相关的6个直系同源CYP450基因CYP307A1, CYP306A1, CYP302A1, CYP315A1, CYP314A1和CYP18A1；系统进化树显示，斜纹夜蛾这6个CYP450基因分别归属于CYP2和线粒体CYP两个亚家族。CYP306A1, CYP314A1和CYP18A1分别在6龄幼虫、预蛹和蛹期具有最高的表达量，并且分别在6龄幼虫中肠、预蛹脂肪体和蛹表皮中表达量最高。相比取食人工饲料的对照组，4龄幼虫取食番薯和花生叶片后斜纹夜蛾4龄幼虫发育至蛹的历期显著延长， CYP306A1在5龄幼虫中肠中的表达量显著上调。在CYP307A1，CYP315A1, CYP314A1和CYP18A1上鉴定出了多种miRNA结合位点。【结论】蜕皮激素合成代谢通路中，CYP306A1，CYP314A1和CYP18A1可能分别在斜纹夜蛾幼虫、预蛹和蛹期起着关键的调控作用，同时也参与宿主植物次生代谢产物的解毒代谢，并且受到miRNA的严密调控。研究结果不仅有助于深入理解昆虫变态发育调控的复杂机制，还为将来的害虫防治提供了潜在的作用靶标，有利于斜纹夜蛾等害虫的可持续治理。
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| 关 键 词： | 斜纹夜蛾 变态发育 蜕皮激素 寄主植物 基因表达 miRNA |
Identification and expression profiling of CYP450 genes involved in the ecdysone synthesis pathway inSpodoptera litura(Lepidoptera: Noctuidae) |
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| GU Jun,YE Yan,LI Shi-Yu,YUAN Ya-Fei,HUANG Li-Hua. Identification and expression profiling of CYP450 genes involved in the ecdysone synthesis pathway inSpodoptera litura(Lepidoptera: Noctuidae)[J]. Acta Entomologica Sinica, 2022, 65(12): 1606-1614. DOI: 10.16380/j.kcxb.2022.12.006 |
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| Authors: | GU Jun YE Yan LI Shi-Yu YUAN Ya-Fei HUANG Li-Hua |
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| Affiliation: | (Guangdong Provincial Key Laboratory of Insect Developmental Biology and Applied Technology, School of Life Sciences, South China Normal University, Guangzhou 510631, China) |
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| Abstract: | 【Aim】 This study aims to explore the expression profiles of CYP450 genes involved in the ecdysone synthesis pathway, so as to provide potential targets for the control of pest insects. 【Methods】The CYP450 genes were identified from the genome of Spodoptera litura by homologous blast using the Bombyx mori CYP450 genes as queries, and their phylogenetic tree was constructed. qPCR was carried out to examine the expression levels of the identified CYP450 genes in different developmental stages (6th instar larva, prepupa and pupa), various tissues (midgut, cuticle and fat body) of these three developmental stages, and the midgut of the 5th instar larvae after the 4th instar larvae fed with the leaves of different host plants ［pepper (Capsicum annuum), cucumber (Cucumis sativus), sweet potato (Ipomoea batatas) and peanut (Arachis hypogaea)］. The developmental duration from the 4th instar larva to pupa was recorded after the 4th instar larvae were fed with the leaves of different host plants. In addition, microRNAs (miRNAs) targeting the identified CYP450 genes were predicted using four programs, PITA, miRanda, microTar and RNAhybrid. 【Results】 Six CYP450 orthologs, CYP307A1, CYP306A1, CYP302A1, CYP315A1, CYP314A1 and CYP18A1, involved in the ecdysone synthesis pathway were identified in S. litura, and the phylogenetic tree showed that the six genes belong to two clans, CYP2 and mitochondrial CYP, respectively. CYP306A1, CYP314A1 and CYP18A1 showed the highest expression levels in the 6th instar larvae, prepupae and pupae, respectively, and the highest expression levels in the midgut of the 6th instar larvae, the fat body of prepupae and the cuticle of pupae, respectively. After the 4th instar larvae of S. litura were fed with the leaves of sweet potato and peanut, the developmental duration from the 4th instar larva to pupa was significantly prolonged, and the expression level of CYP306A1 in the midgut of the 5th instar larvae was significantly upregulated, as compared to those fed with the artificial diet. Multiple miRNA binding sites were identified on CYP307A1, CYP315A1, CYP314A1 and CYP18A1. 【Conclusion】 CYP306A1, CYP314A1 and CYP18A1 in the ecdysone synthesis pathway play key regulatory roles in the larva, prepupa and pupa of S. litura, respectively, and are also involved in detoxification of the secondary metabolites in host plants and subject to the control of various miRNAs. These findings not only help to understand the complex mechanism of insect metamorphosis, but also provide potential targets for pest insect control, which will benefit for the sustainable management of pest insects including S. litura. |
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