家蚕雏翅(minute wing)发生的分子机制研究

The development and differentiation of wing disc is closely related to the process of insect development, and it has become one of the vital organs in the research of insect development. Insect wings are constantly changing (evolution) to adapt to their survival environment. Analysis of the molecular mechanism of insect wing development can help resolve insect metamorphosis process, and it is useful in the research of classification and evolution of insects. Also it has a deeper understanding of the biological control of pests. The silkworm minute wing (mw) is one of the mutations in silkworm wing, its wings of pupal and adult stages is significantly smaller than wild type, and it is controlled by a recessive gene, mw. mw had been mapped in the silkworm established linkage group 22. On the basis of the fine mapping of mw gene (Ma et al., 2013), and the target gene responsible for mw mutant has been discovered, we want to clone the full length of the gene, then compare the transcriptome of wing disc of the wild type recurrent parent, the mutant parent and the mw-near isogenic line. The mechanism of mw gene influence will be analyzed, and genes which can affect the silkworm wing size and development will be analyzed too. The results will provide reference for the mechanism of silkworm wilng development and lepidopteran pests genetic control.

翅原基的发育与分化和昆虫的个体发育进程密切相关，翅已经成为研究昆虫发育过程的重要器官之一。随着环境的改变，昆虫的翅也在不断发生变化以适应其生存。解析昆虫翅发生发育的分子机理不仅能够有助于昆虫变态发育过程的解析，对昆虫分类和进化能够有更深入的认识，也对害虫的生物防治有重要的指导意义。家蚕雏翅基因（minute wing，mw）突变是家蚕翅突变的一种，由隐性基因mw控制，其突变体在蛹期和成虫期翅明显小于正常翅。mw基因已经被学者证明位于家蚕第22连锁群。本研究在精细定位mw基因（马晓等，2013），并寻找到目标基因的基础上，克隆基因的全长序列，分析突变发生的原因，进而通过转录组技术比较野生型轮回亲本、突变体和近等基因系翅原基表达的差异基因，研究mw基因影响家蚕翅形态的作用机制，进一步解析家蚕翅大小发育的分子机制，并为鳞翅目害虫的遗传防治提供参考。

本项目在精细定位雏翅（mw）基因的基础上（马晓等，2013），分析了定位区域的基因表达及其差异，确定并克隆了决定家蚕雏翅基因（mw）的基因棕榈酰转移酶编码基因（Bm-app）。经对Bm-app基因的基因组测序分析，发现该基因启动子核心区域有10bp的插入，导致其表达量显著下降。对突变体及野生型启动子活性进行了DLR Assay的双荧光素酶测定，证实了该10bp的插入严重影响了启动子活性，进而影响了Hippo通路和蛋白酶体基因的表达，导致家蚕翅型的突变。采用转录组测序技术筛选了mw 近等基因系、轮回亲本和突变体翅原基/翅芽的差异表达基因，表明突变体中Hippo通路和蛋白酶体通路基因表达水平差异显著。对Hippo通路和蛋白酶体中的关键基因BmSd、BmRpt4和Bmprosβ1进行干扰或敲除，也会影响家蚕翅的大小。本研究明确了雏翅发生是由于Bm-app基因的突变影响了蛋白酶体和Hippo通路，进而造成翅膀大小的改变。研究结果对于进一步解析家蚕翅大小发育的分子机制有重要作用，并为鳞翅目害虫的遗传防治提供参考。