线虫病原细菌感知宿主的信号调控机制

病原菌与宿主间的互作关系是生物学研究的前沿和热点。前期发现线虫病原细菌Bacillus nematocida B16可以感知宿主C.elegans的存在，调节相应基因的表达，提前形成芽孢，为诱捕线虫做好准备。虽然细菌感知线虫继而引起两者接触是完成侵染的前提条件，但其中的信号调控机制迄今未见报道。本项目以线虫病原细菌B16和C.elegans为研究对象，采用SPME-GC-MS、纯品验证等鉴定线虫的信号物质；通过转录组、蛋白质组及生物信息学分析筛选B16感知、传递信号物质的蛋白及基因，并通过定量PCR和基因功能实验验证靶基因的功能；采用蛋白纯化、ITC、EMSA、Footprinting、GFP标记和细菌双杂交等技术确定B16关键信号蛋白和信号传导通路；从而揭示病原细菌感知宿主的信号调控机制，拓展人们对病原细菌与宿主的互作机制的认识，为合理利用病原细菌奠定基础，具有重要的科学意义与应用价值。

It is always the focus field in Biology that research on the the interactions between pathogenic microorganisms and hosts. In our previous study, we found that the pathogenic bacteria Bacillus nematocida B16 had the capabilities to sense the presence of nematodes, regulate their metabolism to form the spores, and finally get ready for trapping nematodes. Although it is a prerequisite for the infection of the recognization of the nematodes by bacteria, the process on the bacteria recognizing the signal molecules produced by nematodes and the signal regulation mechanism has not been reported so far. Thus, here using B16 and C. elegans as research targets, we design to investigate the volatile signal compounds emitted by nematodes using SPME-GC-MS, and then the functions of the compounds were tested individually using commercially available standards. The potential proteins (genes) involved in signaling pathway in the recognization process will be screened out by transcriptome, proteome and bioinformatics analysis, and those proteins involved in singaling regulation of recognization will be further identified by real time PCR (QPCR), gene knockout and complementary experiments. The binding of the signal compounds and the target protein will be investigated by protein purification, ITC and structural biology. The transcriptional regulation of the signal proteins will be explored by EMSA、Footprinting et al. Moreover, the localization of signaling proteins, interactions between different proteins in signaling pathway and the crosstalk among different signaling pathways were confirmed by green fluorescence protein fusion gene technique and the bacterial two-hybrid system. Our research has important scientific significance and application value, since it will help reveal the signaling mechanism regulating the recognition of nematodes in pathogenic bacteria, expand the understanding the interaction mechanism between pathogenic bacteria and their hosts, lay the foundation for rational use of pathogenic bacteria.