水稻条纹病毒编码蛋白在灰飞虱体内的检测及其与CP体外结合研究

Detection of the Proteins Encoded by Rice Stripe Virus in Laodelphax striatellus Fallén and Interactions in vitro Between CP and the Four Proteins

【目的】检测灰飞虱体内由水稻条纹病毒(RSV)编码的外壳蛋白(CP)、病害特异性蛋白(SP),以及非结构蛋白NS2、NS3和NSvc4等5种蛋白以及病毒粒子CP与SP、NS2、NS3、NSvc4的体外结合,为进一步研究水稻条纹病毒与其介体昆虫互作提供有用信息。【方法】提取高带毒灰飞虱(Q家系)总蛋白,用免疫方法检测昆虫体内CP、SP、NS2、NS3和NSvc4等5种重要蛋白的表达以及病毒粒子CP与SP、NS2、NS3、NSvc4的体外结合情况。【结果】结果表明在带毒灰飞虱体内以上5种蛋白都能检测到,而且CP含量最高,NSvc4、SP、NS3、NS2含量依次降低;用体外原核表达后纯化的蛋白进行了RSV粒子CP与SP、NS2、NS3、NSvc4的体外结合实验,实验结果表明病毒粒子CP与SP、NSvc4之间存在体外结合关系。【结论】检测结果为灰飞虱作为RSV的昆虫寄主并且可以在其体内增殖的结论提供了支持,表达差异表明病毒编码蛋白在介体灰飞虱体内可能存在着一定的协调关系;病毒粒子与SP的结合为前人关于CP与SP可能共同作用于叶绿体进而影响症状的严重度的报道又提供了一个新的证据,病毒粒子与NSvc4之间存在体外结合关系,也为NSvc4可能作为运动蛋白通过与CP的互作以及诱导病毒在胞间运动需要CP的辅助提供了佐证。

【Objective】Detecting the five proteins encoded by RSV（CP,SP,NS2,NS3 and NSvc4） in the small brown plant hopper（Laodelphax striatellus Fallén,SBPH）,investigating the interactions between CP and the other four proteins of RSV in vitro,and providing available data for further studying the relationship between RSV and its insect vector.【Method】The total protein of virus infected SBPH was extracted,and hybridized with specific antibodies of the five RSV proteins.The gel overlay assay was performed.【Result】All the five proteins encoded by RSV presented in the SBPH.Further more,CP has a higher density among the five proteins.Besides,results indicated that there are interactions with CP and SP or CP and NSvc4 in vitro.【Conclusion】The study confirmed that RSV replicates in SBHP,and different concentrations of the five proteins presented in SBHP body indicated the different roles in its replication.The interaction in vitro between CP and NSvc4（the potential movement protein） indicates that the cell-to-cell movement of RSV may depend on this relationship between CP and NSvc4.The results also showed the interaction between CP and SP,which has been reported previously on chloroplast of rice and influence on the severity of symptoms.