爪哇根结线虫效应蛋白Mj-1-1在寄生过程中的功能研究

Meloidogyne javanica is one of the most damaging root-knot nematodes9（RKN）and effector protein plays an important role during the interaction between RKN and host, but only a few of effector proteins of M.javanica were identified and studied. A new effector (Mj-1-1) was isolated in our team and some basic results of Mj-1-1 were obtained in my doctoral dissertation. In this study, We hope through the in situ hybridization and immunofluorescence localization, confirm the localization of Mj-1-1 in M.javanica, and combined the histologic section demonstrate the secretion of Mj-1-1 in plant during the parasitism. On the results of yeast two hybrid, verify the interaction between MJ-1-1 and host proteins. Further more, analyse the function of host proteins during the infection and parasitism by over expression and RNAi. We hope based on the above results, make clear the function of Mj-1-1 during the parasitism and provide the theoretical basis to the control of nematode.

爪哇根结线虫是危害最严重的根结线虫之一，效应子蛋白在根结线虫与寄主植物的互作中起着非常重要的作用，但是只有很少的爪哇根结线虫效应子蛋白得到研究，Mj-1-1是本实验室从爪哇根结线虫中分离到的一个新的效应子蛋白，也是本申请人博士论文的一部分，本申请项目是在已有数据的基础上，希望对Mj-1-1开展进一步的研究工作。一方面通过原位杂交和免疫定位确定Mj-1-1在爪哇根结线虫中的组织定位，同时，采用植物组织切片技术，确定Mj-1-1在线虫寄生过程中植物体内的分泌情况。另一方面BIFC和CoIP技术，验证MJ-1-1与寄主蛋白之间的相互作用，以及通过过表达和RNAi等技术研究寄主蛋白对爪哇根结线虫侵染和寄生的影响。希望通过以上研究结果，深入揭示Mj-1-1在爪哇根结线虫侵染和寄生中的功能，为根结线虫的防治提供理论依据。

本项目以爪蛙根结线虫为研究对象，对Mj-1-1基因在爪蛙根结线虫体内的组织定位、MJ-1-1蛋白与拟南芥蛋白的相互作用等做了进一步的分析。研究结果表明，Mj-1-1基因在爪蛙根结线虫的背食道腺中表达；在拟南芥体内，通过与KA1和MELP蛋白之间相互作用来促进自身侵染和寄生；亚细胞定位表明MJ-1-1蛋白定位于拟南芥原生质体的细胞核中，MELP蛋白定位于拟南芥整个细胞中，在拟南芥原生质体中没有观察到KAI蛋白的荧光信号；通过共转化的方法，在酵母细胞中验证了拟南芥两个蛋白KA1和MELP与MJ-1-1之间的相互作用；用Melp基因对应的拟南芥突变体进行接种试验表明，突变体拟南芥与野生型拟南芥相比，感病性显著降低。这些结果为研究MJ-1-1蛋白与MELP蛋白之间的相互作用提供了分子基础，也为研究爪蛙根结线虫在植物体内治致病的分子机制提供了理论依据。