家蚕翅原基变态发育的分子调控机理

Based on a research model of ecdysone and juvenile hormone regulation of wing disc cuticle protein (BmWCPs), this project will study the molecular mechanisms of hormone signal transduction pathways in regulation of metamorphosis in the domesticated silkworm (Bombyx mori) and wild silkworm (B. mandarina) by using approaches of molecular biology, developmental biology, transcriptomics, proteomics and biotechnology. It will determine the differences in gene and protein expression in the wing dics between B. mori and B. mandarina. It will also try to develop silkworm strain with elongated pupal stage by transgenetic method. The research contents include: 1) molecualr regulation of B. mori wing disc development by ecdysone hormone signal transduction pathway; 2) molecualr regulation of B. mori wing disc development by juvenile hormone signal transduction pathway; 3) effects of other nuclear transcription factors on the development of the B. mori wing disc; 4) transcriptomics and proteomics analysis of the wing dics of B. mori and B. mandarina during metamorphosis and 5) construction of transgenetic silkworm with elgonated pupal stage. The study is important in explanation of the molecular mechanisms of insect metamorphosis, differences in wing disc development of B. mori and B. mandarina and its effects on the flight ability. The study will also play a role in generation of silkworm strains with elongated pupal stage by taking the advantage of the character of pupa-specific expression of BmWCPs genes to solve the issue of silk damage due to the early emergence of an adult from a cocoon.

以家蚕和野桑蚕为研究对象，以受蜕皮激素和保幼激素调控的蛹期翅原基表皮几丁质蛋白BmWCPs基因为主要研究模型，采用分子生物学、发育生物学、转录组学和蛋白组学等方法，研究激素信号传导途径对家蚕变态发育调控的分子机理，分析家蚕与野桑蚕翅原基发育过程中的差异基因，并尝试通过转基因技术获得蛹期延长的家蚕品系。研究内容包括：1）蜕皮激素信号传导途径对家蚕翅原基发育的分子调控；2）保幼激素信号传导途径对家蚕翅原基发育的分子调控；3）其它核转录因子对家蚕和野桑蚕翅原基发育的影响；4）家蚕和野桑蚕翅原基变态发育的转录组学和蛋白组学分析；5）蛹期延长的转基因家蚕素材构建。研究对阐释昆虫变态发育分子调控机制，解释家蚕与野桑蚕翅原基发育的差异及其对飞翔能力影响等科学问题具有重要意义，并为利用BmWCPs基因蛹期专一表达的特性以延长蚕蛹发育，解决成虫过早破茧而出、降低蚕丝质量的重大生产问题提供理论指导。

变态发育是昆虫最显著的发育特征和奇特的生物学现象，使得昆虫能使昆虫更好地适应环境、生存和繁殖。翅原基变态发育是昆虫变态发育最典型的事件之一。家蚕重要的经济昆虫，同时是鳞翅目的模式昆虫，对翅发育的研究可为寻求昆虫生长调节剂打下基础。本项目以阐明昆虫激素调控家蚕翅原基变态发育的分子机制为出发，分析了家蚕翅原基发育的基因表达调控，家蚕与野桑蚕翅原基发育过程中的转录组差异，通过GAL4/UAS转基因技术构建蛹期延长的家蚕品系，研究了DNA二级结构调控基因表达的分子机制。在国际重要SCI刊物发表10篇论文，申报国家发明专利2项，圆满完成了计划任务。主要取得如下重要结果：(1) 阐明了蜕皮激素和保幼激素调控翅原基变态发育的分子机制，主要是通过转录因子( BmBR-C Z4、BmPOUM2、BmAbd-A等)的级联反应调控蛹期专一表达的翅原基表皮蛋白基因BmWCPs的表达，促使幼虫翅原基完成向蛹翅的转变；(2) 发现BmFoxA是调控翅原基变态发育的分子开关之一，与BmSAGE一起通过调控BmWCPs的表达与不表达来调控翅原基变态发育；(3) 揭示了家蚕飞行能力退化的分子机制，主要由于大量与翅发育相关基因表达历时短且表达量低和背板肌动蛋白带细胞骨架调节与紧密连接相关通路下调导致背板肌肉细胞细小且排列疏松等造成的；(4) 获得了以BmWCP4启动子和致死/凋亡相关基因构建的蛹期延长的转基因家蚕素材；(5) 发现了一种新的转录调控机制，BmPOUM2启动子上的二级结构G4和i-motif及其它们的结合蛋白参与了BmPOUM2的表达调控。研究结果不仅可提供昆虫变态发育的新理论，也可为理解昆虫对人工驯化环境的适应性和新的基因表达调控机制提供数据，为今后研制新型昆虫生长调节剂奠定基础，更为解决成虫过早破茧而出，降低蚕丝质量的重大生产问题提供了素材。