枯草芽胞杆菌产生羊毛硫细菌素subtilomycin促进其在植物体内定殖的机制

Endophyte can contribute some benefits to plant. They need evade the immune to enters into plants. However, the invasion mechanisms of endophytes and related factors are seldom reported. Previously, an endophyte Bacillus subtilis BSn5 with activity to control soft disease was isolated from Amorphophallus konjac. The lantibiotic subtilomycin gene cluster is locus on a recombination hot spot of B. subtilis genome. By investigating the subtilomycin production of the B. subtilis isolates from distinct environments, we found that its production is related to the origin of plant endosphere. The subtilomycin affects the ability of producer to colonize into Arabidopsis thaliana. The lantibiotic can affect the plant colonization of producer, which has never been reported elsewhere. During the isolation of subtilomycin, the binding of subtilomycin with flagellin, which is encoded by gene hag, was detected. Flagellin is a well studied molecular that can be recognized by plant. These analysis provide us with a new perspective to study the mechanism of endophyte formation. The project intends to carry out the following studies: the interaction mode of subtilomycin and Hag; the affects of subtilomycin on ROS burst, related defense genes expression and metabolites triggered by flg22-FLS2 pathway; the affects of subtilomycin on root surface colonization of producer. This project will provide theoretical basis for agricultural application of endophyte bacteria as biocontrol agent and biofertilizer with stronger environmental tolerance.

内生菌能对植物起到有益功能，它要进入植物体内须突破免疫屏障，但相关机制和影响因素鲜有报道。我们前期从魔芋组织中分离到一株能够抵抗软腐病的内生枯草芽胞杆菌BSn5。其产生的羊毛硫细菌素subtilomycin的基因簇位于枯草芽胞杆菌重组热点处。对不同环境来源的枯草芽胞杆菌分离株的调查发现，该细菌素的产生与植物内生环境相关，并影响BSn5在拟南芥体内定殖，目前尚未见细菌素影响产生菌在植物定殖的报道。分离该细菌素时发现它能与自身鞭毛蛋白结合，已知鞭毛蛋白是能被植物识别的标记物，这为研究内生菌的定殖机制提供了一个新思路。本项目将以BSn5为研究对象，开展以下工作：研究subtilomycin与鞭毛蛋白相互作用方式；该细菌素对拟南芥鞭毛蛋白-FLS2受体识别通路所诱导的活性氧反应、相关免疫基因和代谢物表达的影响；和对产生菌在根表定殖能力的影响。本研究的开展将为研制更稳定的生物农药和肥料提供理论基础。

内生菌能对植物起到有益功能，它要进入植物体内须突破免疫屏障，但相关机制和影响因素鲜有报道。我们前期从魔芋组织中分离到一株能够抵抗软腐病的内生枯草芽胞杆菌BSn5。其产生的羊毛硫细菌素subtilomycin的基因簇位于枯草芽胞杆菌重组热点处。对不同环境来源的枯草芽胞杆菌分离株的调查发现，该细菌素的产生与植物内生环境相关，并影响BSn5在拟南芥体内定殖，目前尚未见细菌素影响产生菌在植物定殖的报道。我们以拟南芥与枯草芽胞杆菌BSn5为对象，开展了subtilomycin对内生菌定殖机制的研究。通过体内的分离和体外的相互作用分析，发现subtilomycin能够结合细菌自身的鞭毛蛋白。这种结合抑制了鞭毛蛋白引起的ROS和气孔闭合等植物防御响应和关键免疫基因的表达。拟南芥定殖分析发现，对植物防御反应的抑制作用提供了产生菌更好的内生定殖能力。此外，我们也发现subtilomycin影响了细菌生物被膜的形成和魔芋根菌群的组成。我们的研究结果提出了一种新的植物内生细菌的内生定殖模式。揭示了细菌素subtilomycin在植物根圈系统中可能发挥多重生态功能赋予产生菌适应优势。这为开发更稳定的生物化肥提供理论指导。