Table_3_A novel function of the key nitrogen-fixation activator NifA in beta-rhizobia: Repression of bacterial auxin synthesis during symbiosis.docx

Rhizobia fix nitrogen within root nodules of host plants where nitrogenase expression is strictly controlled by its key regulator NifA. We recently discovered that in nodules infected by the beta-rhizobial strain Paraburkholderia phymatum STM815, NifA controls expression of two bacterial auxin synthesis genes. Both the iaaM and iaaH transcripts, as well as the metabolites indole-acetamide (IAM) and indole-3-acetic acid (IAA) showed increased abundance in nodules occupied by a nifA mutant compared to wild-type nodules. Here, we document the structural changes that a P. phymatum nifA mutant induces in common bean (Phaseolus vulgaris) nodules, eventually leading to hypernodulation. To investigate the role of the P. phymatum iaaMH genes during symbiosis, we monitored their expression in presence and absence of NifA over different stages of the symbiosis. The iaaMH genes were found to be under negative control of NifA in all symbiotic stages. While a P. phymatum iaaMH mutant produced the same number of nodules and nitrogenase activity as the wild-type strain, the nifA mutant produced more nodules than the wild-type that clustered into regularly-patterned root zones. Mutation of the iaaMH genes in a nifA mutant background reduced the presence of these nodule clusters on the root. We further show that the P. phymatum iaaMH genes are located in a region of the symbiotic plasmid with a significantly lower GC content and exhibit high similarity to two genes of the IAM pathway often used by bacterial phytopathogens to deploy IAA as a virulence factor. Overall, our data suggest that the increased abundance of rhizobial auxin in the non-fixing nifA mutant strain enables greater root infection rates and a role for bacterial auxin production in the control of early stage symbiotic interactions.

根瘤菌在宿主植物根瘤中固定氮素，其氮化酶的表达严格受关键调节因子NifA的控制。我们最近发现，在感染β-根瘤菌菌株Paraburkholderia phymatum STM815的根瘤中，NifA调控着两种细菌生长素合成基因的表达。无论是iaaM和iaaH的转录本，还是代谢物吲哚乙酰胺（IAM）和吲哚-3-乙酸（IAA），在nifA突变体根瘤中的含量均高于野生型根瘤。在此，我们记录了P. phymatum nifA突变体在普通大豆（Phaseolus vulgaris）根瘤中诱导的结构变化，最终导致瘤结过度生长。为了研究P. phymatum iaaMH基因在共生过程中的作用，我们监测了其在NifA存在与否以及共生不同阶段中的表达。研究发现，在所有共生阶段，iaaMH基因均受到NifA的负调控。尽管P. phymatum iaaMH突变体产生的瘤结数量和固氮酶活性与野生型菌株相同，但nifA突变体产生的瘤结数量多于野生型，并聚集在规则排列的根区。在nifA突变体背景下突变iaaMH基因，降低了这些瘤结在根上的出现。我们进一步证明，P. phymatum iaaMH基因位于共生质粒中GC含量显著较低的区域，且与IAM途径中的两个基因高度相似，该途径常被细菌植物病原体用来部署IAA作为致病因子。总体而言，我们的数据表明，非固氮nifA突变体菌株中根瘤菌生长素的增加丰度，有助于提高根的感染率，并在早期共生相互作用控制中，细菌生长素生产发挥了作用。