家蚕细胞质型多角体病毒受体鉴定及入侵细胞的分子机理研究

项目名称： 家蚕细胞质型多角体病毒受体鉴定及入侵细胞的分子机理研究

项目编号： No.31272500

项目类型： 面上项目

立项/批准年度： 2013

项目学科： 农业科学

项目作者： 贡成良

作者单位： 苏州大学

项目金额： 80万元

中文摘要： 为了筛选和鉴定家蚕质型多角体病毒(BmCPV)的受体，探讨BmCPV有效入侵的分子机制，我们在已完成BmCPV基因组测序、通过生物信息学分析家蚕免疫球蛋白超家族(IgSF)预测BmCPV受体、SRC激酶基因克隆和家蚕转基因平台建立的基础上，拟通过BmCPV与中肠细胞表面蛋白互作和酵母双杂交捕获、验证BmCPV受体候选基因；通过抑制或缺失候选基因表达以及异位表达候选基因,研究家蚕对BmCPV的感染性和组织特异性，鉴定受体；通过特异性抑制剂研究细胞内吞BmCPV方式；通过RNAi抑制β1整联蛋白、SRC激酶的表达，抑制剂抑制SRC激酶活性，根据家蚕对BmCPV的感染性及BmCPV入侵细胞过程观察，探讨β1整联蛋白和SRC激酶在BmCPV入侵细胞中的功能。本项目不仅能明确BmCPV受体和入侵的基本分子机制，并可为敲除或抑制病毒入侵相关的家蚕基因，提高家蚕对BmCPV抗性以及研发防治药物提供靶标。

中文关键词： 家蚕；质型多角体病毒；受体；内吞；抑制剂

英文摘要： In our previous studies, sequencing of Bombyx mori cypovirus (BmCPV) genome, predicting of BmCPV receptor genes through the bioinformatics analysis of B. mori immunoglobulin superfamily (IgSF) genes, cloning of B. mori SRC kinase gene and establishing platform for generating transgenic silkworm have be accomplished. In this study, we are going to screen and identify BmCPV receptor and characterize functional BmCPV entry so as to understand the molecular mechanism of BmCPV cell entry. Based on the former studies, we will capture BmCPV receptor candidate genes with the interaction between BmCPV structural protein VP3 and proteins located on the midgut cell surface, and validate with yeast two-hybrid system. The receptor candidate genes will be further determined by the changes of BmCPV infectivity to silkworm after inhibiting or deleting the expression and ectopic expression of the candidate genes. Endocytosis pathway of BmCPV will be studied by using specific inhibitors for internalization. The roles of β1 integrin and SRC kinase in the course of BmCPV cell entry are characterized according to the changes of BmCPV infectivity to silkworm and observation of BmCPV cell entry with electron microscope after both of suppressing the expression of β1 integrin and SRC kinase genes through RNAi and inhibiting the activit

英文关键词： Bombyx mori；Cypovirus；Receptor；Endocytosis；Inhibitor