柞蚕蛹滞育解除期高丰度表达的KK-42结合蛋白基因的功能分析

项目名称： 柞蚕蛹滞育解除期高丰度表达的KK-42结合蛋白基因的功能分析

项目编号： No.31072082

项目类型： 面上项目

立项/批准年度： 2011

项目学科： 轻工业、手工业

项目作者： 刘彦群

作者单位： 沈阳农业大学

项目金额： 8万元

中文摘要： KK-42是一种昆虫生长调节剂，可以加快昆虫（柞蚕、天蚕、家蚕等）幼虫生长发育，尤其能解除天蚕的卵滞育和棉铃虫的蛹滞育。KK-42结合蛋白是从天蚕卵中分离的，并与天蚕卵滞育解除有直接关系。我们从以蛹滞育的柞蚕中也分离到了KK-42结合蛋白基因，本项目分析该蛋白与同源蛋白的系统进化关系，并检测其基因表达谱。卵期蛋白组学研究也证明了该蛋白的存在。BLAST分析表明，目前仅有柞蚕和天蚕的KK-42结合蛋白基因被分离出来，二者在蛋白水平的同源性高达95%。同源比较表明，该蛋白与家蚕等鳞翅目的卵特异蛋白仅有42-48%的相似性，与其它昆虫的脂肪酶仅显示24-36%的相似性。保守功能域和进化分析表明，KK-42结合蛋白是脂肪酶家族的一个新成员。表达谱分析发现，KK-42结合蛋白基因在柞蚕4个发育阶段和所检测的10个组织器官中均有表达，但其表达与柞蚕蛹滞育密切相关（在注定滞育的秋蚕前蛹期正常表达，在滞育期消失或极低量表达，而在滞育解除后又重新开始表达），并且在滞育后发育期间伴随着卵的形成而高丰度表达。研究结果为进一步研究该蛋白在柞蚕生长发育过程中的功能进而理解KK-42的分子作用机制奠定了基础。

中文关键词： 柞蚕；蛹滞育；KK-42结合蛋白；同源比较；表达谱

英文摘要： Imidazole derivative KK-42, an insect growth regulator, not only can promote insect growth and development, such as Antheraea pernyi, A. yamamai and Bombyx mori, but also can induce a precocious termination of diapause in pharate first larvae of A. yamamai and in pupae of Helicoverpa armigera. KK-42 binding protein (KK-42BP) was first isolated from eggs of A. yamamai, and has been shown to be related to termination of diapause of A. yamamai. We have isolated the KK-42BP gene from A. pernyi that enters diapause as a pupa. The aim of this project is to analyze the evolutionary relationships between KK-42BP and other homolgous proteins, and to investigate its expression profile. BLAST analysis showed that KK-42BPs were only isolated from A. pernyi and A. yamamai, with 95% identities at protein sequence level. Homolgous comparison indicated that KK-42BPs only had 42-48% identities with known yolk proteins from lepidopteran insects including B. mori, and had 24-36% identities with lipases from other insects. Conserved motif and phylogenetic analysis suggested that KK-42BPs isolated from A. pernyi and A. yamamai might be a new member of insect lipase superfamily. ApKK-42BP mRNA was expressed in all of the tissues tested, throughout four developmental stages, and in both female and male. ApKK-42BP mRNA was expressed at a low level in pre-diapause period of diapause-destined pupae, disappeared at an undetectable low level in early diapause and diapause stage, and appeared at a low level in post-diapause stage similar to that in pre-diapause stage, suggesting that ApKK-42BP was associated with pupal diapause of A. pernyi. In addition, we also found that expression of ApKK-42BP mRNA increased at a very high level when diapause-activated pupae developed into the late pupal stage, indicating that the KK-42BP gene might also be related to reproductive development of A. pernyi. Our results provide basic information for further studies on function identification of KK-42BP in growth and development of A. pernyi, which will lead to comprehensive understand the molecular mechanism of KK-42 on insects.

英文关键词： Antheraea pernyi; pupal diapause; KK-42 binding protein; homolgous comparison; expression profile