Sinorhizobium fredii HH103 RirA Is Required for Oxidative Stress Resistance and Efficient Symbiosis with Soybean

Members of Rhizobiaceae contain a homologue of the iron-responsive regulatory protein RirA. In different bacteria, RirA acts as a repressor of iron uptake systems under iron-replete conditions and contributes to ameliorate cell damage during oxidative stress. In Rhizobium leguminosarum and Sinorhizobium meliloti, mutations in rirA do not impair symbiotic nitrogen fixation. In this study, a rirA mutant of broad host range S. fredii HH103 has been constructed (SVQ780) and its free-living and symbiotic phenotypes evaluated. No production of siderophores could be detected in either the wild-type or SVQ780. The rirA mutant exhibited a growth advantage under iron-deficient conditions and hypersensitivity to hydrogen peroxide in iron-rich medium. Transcription of rirA in HH103 is subject to autoregulation and inactivation of the gene upregulates fbpA, a gene putatively involved in iron transport. The S. fredii rirA mutant was able to nodulate soybean plants, but symbiotic nitrogen fixation was impaired. Nodules induced by the mutant were poorly infected compared to those induced by the wild-type. Genetic complementation reversed the mutant's hypersensitivity to H₂O₂, expression of fbpA, and symbiotic deficiency in soybean plants. This is the first report that demonstrates a role for RirA in the Rhizobium-legume symbiosis.