Rhizobium purine metabolism and its role in infection of legumes
Abstract
Purine auxotrophs of Rhizobium leguminosarum CFN42 elicit uninfected pseudonodules on bean. Mutations that cause this symbiotic defect mapped to seven partially-sequenced purine biosynthetic genes, all responsible for steps before the production of the intermediate 5-aminoimidazole-4-carboxamide ribonucleotide (AICAR). Supplementing the root medium with AICA riboside, the unphosphorylated form of AICAR, substantially enhanced nodule development. Of the purines tested, only inosine enhanced nodulation, but very high concentrations were required for this effect. Studies with a double mutant indicated that the histidine pathway was required for inosine's effect, presumably because AICAR is produced as a byproduct of histidine synthesis. AICA riboside also promoted infection of soybean by two R. fredii purine auxotrophs, even though these mutants grew poorly with AICA riboside as a purine source. Nodules elicited by purine auxotrophs in the presence of AICA riboside had infection threads and anatomical features characteristic of normal nodules. The nodule-specific proteins, leghemoglobin and uricase, were detected in these nodules, although at lower levels than in nodules elicited by the wild type. Analysis of the temporal requirement for AICA riboside indicated that it was required during the entire infection process, rather than just for infection thread initiation. These studies also suggested that a critical stage of infection is reached six days post-inoculation, at which development becomes committed to forming a true root nodule structure. To better understand how AICA riboside enhances nodule development, AICA riboside utilization and uptake were examined and compared in three Rhizobium genetic backgrounds. A mutant resistant to the toxic AICA riboside analog, pyrazofurin, could not convert AICA riboside to AICAR. Nodulation assays with a purine auxotrophic derivative of this mutant demonstrated that this conversion was required for AICA riboside to promote infection. These results support the hypothesis that rhizobia divert AICAR from purine biosynthesis for the production of a factor that induces infection thread development in the plant. Successful infection by rhizobia triggers the nodule-specific developmental pathway, which leads to the formation of a true nodule structure that is distinct from the root-like structure that develops in the absence of infection.
This paper has been withdrawn.