决明胰蛋白酶抑制剂调控菜青虫中肠消化酶合成的分子机制研究

项目名称： 决明胰蛋白酶抑制剂调控菜青虫中肠消化酶合成的分子机制研究

项目编号： No.31500276

项目类型： 青年科学基金项目

立项/批准年度： 2016

项目学科： 生物科学

项目作者： 周嘉裕

作者单位： 西南交通大学

项目金额： 20万元

中文摘要： 菜青虫属鳞翅目，是世界性重大致灾害虫之一，给农业生产带来严重威胁。植物胰蛋白酶抑制剂能够通过诱导鳞翅目昆虫肠道内消化酶的过量合成，导致代谢紊乱，发挥抗虫效应，但相关分子机制未见报道。本项目组首次分离得到决明胰蛋白酶抑制剂COTI，其能够抑制菜青虫的取食及生长，并导致30%的致死率。近期，我们已成功克隆得到COTI基因，并表达纯化获得有抗虫生物活性的重组蛋白。转录组数据分析发现，饲喂重组蛋白的菜青虫中肠内有多个消化酶候选基因的表达上调。本项目组拟充分挖掘、获得诱导消化酶的候选基因；结合荧光定量PCR，筛选出重要的诱导消化酶基因，继而考察重要的诱导消化酶对菜青虫生理的影响；克隆重要的诱导消化酶基因上游启动子序列，分析验证启动子中受COTI诱导响应的顺式调控元件及转录因子，以期系统阐释COTI调控菜青虫中肠消化酶合成的分子机制。

中文关键词： 胰蛋白酶抑制剂；菜青虫；消化酶；调控；分子机制

英文摘要： Pieris rapae (Lepidoptera), which spreads widely in the world, brings serious disaster to agricultural production. Although trypsin inhibitor is able to induce excess synthesis of digestive enzymes in the midgut of Lepidoptera insects, the molecular mechanism has not been elucidated to date. Our previous study has shown that the trypsin inhibitor from Cassia obtusifolia, named COTI, could inhibit the growth of Pieris rapae larvae and lead to 30% of mortality rate. Recently, the COTI gene has been cloned, expressed and purified successfully. The obtained recombinat protein showed anti-insect activity. Furthermore, several up-regulated candiate digestive enzymes were found in the transcriptome data of the midgut from Pieris rapae larvae fed on COTI recombinant protien. In this project, the candidate digestive enzymes induce by COTI will be obtained fully from the transcriptome data. Secondly, the important digestive enzymes induced by COTI will be screened using real-time PCR. The influence of those important digestive enzymes on the physiology of Pieris rapae larvae will be studied. Then, the cis-regulatory elements responding to COTI will be cloned and identified. At last, the transcription factor, which binds the cis-acting regulatory element responding to COTI, will be isolated and validated. Therefore, the regulatory mechanism of COTI on the biosyhthesis of digestive enzymes in the midgut of Pieris rapae will be illustrated systematically.

英文关键词： Trypsin inhibitor;Pieris rapae;Digestive enzyme;Regulation;Molecular mechanism