花生根瘤菌Bradyrhizobium sp.MM6 Ⅲ型分泌系统的结构和功能

【目的】探究花生根瘤菌Bradyrhizobium sp.MM6的Ⅲ型分泌系统(T3SS)的结构及其在根瘤菌与不同宿主建立共生关系中的作用。【方法】同源比对分析菌株MM6的T3SS基因簇的结构特征,并采用三亲本接合转移的方法构建T3SS调节基因ttsI突变菌株;通过蛭石结瘤和石蜡切片实验,比较突变体与野生型的共生固氮表型差异。【结果】经预测,MM6的T3SS基因簇编码区长约34.1 kb,可分为3个区域,包含10个保守结构基因和8个效应蛋白基因,与B.diazoefficiens USDA110相应基因的序列相似性为83%–93%;成功构建了MM6的ttsI突变株;ttsI突变株与野生型分别与花生(S523和Y45)、野大豆和大豆中黄57结瘤,ttsI突变体在花生中的总瘤数显著增加(P<0.05),根瘤中含菌细胞更多;ttsI突变体在野大豆中平均每株植物增加4个根瘤,根瘤中含菌细胞更多,地上部干重相比野生型MM6显著增加(P<0.05);在大豆中黄57中,野生型MM6能形成红色的有效根瘤,ttsI突变体不结瘤,且植株叶片发黄,地上部干重相比野生型MM6显著降低(P<0.05)。【结论】MM6的T3SS在花生和野大豆共生体系中起着有害的作用,而在大豆中黄57的共生体系中起着有利的作用。

Structure and function of type III secretion system in peanut Bradyrhizobium sp. MM6

[Objective] To reveal the functions of type III secretion system (T3SS) of peanut Bradyrhizobium sp. MM6 interacting with different host plants.[Methods] We analyzed its structural characteristics of T3SS gene clusters by BLASTn in NCBI related database, and constructed the T3SS regulatory gene ttsI mutant by homologous single exchange. We also compared the differences of symbiotic phenotype between the MM6 and its ttsI mutant by nodulating experiment.[Results] The T3SS gene clusters of MM6 included 3 regions containing 10 conserved structural genes and 8 effectors. The similarity was 83% to 93% with B. diazoefficiens USDA110 corresponding genes sequence. The total nodule number and fresh root nodule weight significantly increased in the ttsI mutant nodulating peanut and wild soybean. In soybean Zhonghuang 57, the MM6 formed red effective root nodules, whereas the ttsI mutant only formed 1 empty nodule, and the plant leaves turned yellow, and the dry weight of aboveground part was significantly lower than that of the wild type strain.[Conclusion] The T3SS of MM6 played a negative role in the symbiosis system of MM6 with peanut and wild soybean, whereas it played a positive role when nodulated with the soybean Zhonghuang 57.