固氮施氏假单胞菌RpoN蛋白对鞭毛合成的影响

The Regulatory Effect of RpoN on Flagellar Biosynthesis in Nitrogen-Fixing Pseudomonas stutzeri

选择性σ因子σ^(54)(又称为RpoN)参与到许多细菌特定基因的转录起始过程,如氮同化基因、四碳二羧酸转运基因以及鞭毛基因。为了研究施氏假单胞菌A1501(Pseudomonas stutzeri,A1501)中σ因子RpoN对鞭毛合成的影响,构建了A1501的rpoN缺失突变株与功能回补株,并对野生型、突变株与回补株的鞭毛结构、swimming运动、生物膜形成、根际定殖能力以及鞭毛基因的表达水平进行了比较分析。结果表明:rpo N突变株丧失了鞭毛结构与swimming运动能力,根际定殖与生物膜形成能力相对野生型分别下降了25倍与10倍;此外,rpoN缺失后A1501中大量鞭毛基因发生显著下调;启动子分析发现RpoN除了可能参与调控Ⅱ级、Ⅲ级鞭毛基因外,还可能调控一级基因fli A以及四级基因fliC。以上结果说明RpoN在不同调控等级影响A1501鞭毛的合成,进而影响该菌运动、生物膜形成、根际定殖这些与水稻建立起联合固氮体系息息相关的过程。

Many bacteria use the alternative sigma factor σ^（54）（ also known as Rpo N） to transcribe specific genes,such as nitrogen assimilation genes,C4-dicarboxylate transport genes and flagellar genes. In this study,the global regulator was investigated to explore its regulatory effect on flagellar biosynthesis in Pseudomonas stutzeri A1501. Deletion mutant and complementary strain were constructed in rpo N to analyze phenotypes in flagellar structure,swimming motility,biofilm formation,rhizosphere colonization and relative expression levels of flagellar genes. The results showed that loss of rpo N could cause impaired flagellar structure and swimming motility,25-and 10-fold decrease in competitive root colonization and biofilm-forming ability,respectively. Moreover,relative expression levels of flagellar genes in rpo N mutant could significantly decrease when compared to the wild-type. In Pseudomonas,Class Ⅱ and Class Ⅲ flagellar genes were Rpo N-dependent. And in this work,putative Rpo N binding site was observed in fli C,fli A promoter regions,indicating that Rpo N in A1501 could also participate in Class I and Class IV genes regulatory networks. These results suggested that Rpo N affected flagellar biosynthesis process by participating different flagellar gene transcriptional hierarchy, thus affecting swimming motility, biofilm formation and root colonization in Pseudomonas stutzeri A1501.