A Sinorhizobium meliloti RpoH-Regulated Gene Is Involved in Iron-Sulfur Protein Metabolism and Effective Plant Symbiosis under Intrinsic Iron Limitation

Abstract

In Sinorhizobium meliloti, RpoH-type sigma factors have a global impact on gene expression during heat shock and play an essential role in symbiosis with leguminous plants. Using mutational analysis of a set of genes showing highly RpoH-dependent expression during heat shock, we identified a gene indispensable for effective symbiosis. This gene, designated sufT, was located downstream of the sufBCDS homologs that specify the iron-sulfur (Fe/S) cluster assembly pathway. The identified transcription start site was preceded by an RpoH-dependent promoter consensus sequence. SufT was related to a conserved protein family of unknown molecular function, of which some members are involved in Fe/S cluster metabolism in diverse organisms. A sufT mutation decreased bacterial growth in both rich and minimal media, tolerance to stresses such as iron starvation, and activities of some Fe/S cluster-dependent enzymes. These results support the involvement of SufT in SUF (sulfur mobilization) system-mediated Fe/S protein metabolism. Furthermore, we isolated spontaneous pseudorevertants of the sufT mutant with partially recovered growth; each of them had a mutation in rirA. This gene encodes a global iron regulator whose loss increases the intracellular iron content. Deletion of rirA in the original sufT mutant improved growth and restored Fe/S enzyme activities and effective symbiosis. These results suggest that enhanced iron availability compensates for the lack of SufT in the maintenance of Fe/S proteins. IMPORTANCEAlthough RpoH-type sigma factors of the RNA polymerase are present in diverse proteobacteria, their role as global regulators of protein homeostasis has been studied mainly in the enteric gammaproteobacterium Escherichia coli. In the soil alphaproteobacterium Sinorhizobium meliloti, the rpoH mutations have a strong impact on symbiosis with leguminous plants. We found that sufT is a unique member of the S. meliloti RpoH regulon; sufT contributes to Fe/S protein metabolism and effective symbiosis under intrinsic iron limitation exerted by RirA, a global iron regulator. Our study provides insights into the RpoH regulon function in diverse proteobacteria adapted to particular ecological niches and into the mechanism of conserved Fe/S protein biogenesis. Bacteria of the phylum Proteobacteria have very diverse shape, metabolic capacity, and ecological significance and are adapted to diverse environments. The alphaproteobacterium Sinorhizobium meliloti inhabits the soil and engages in nitrogenfixing symbiosis with its compatible legume hosts (genera Medicago, Melilotus, and Trigonella), in which bacterially fixed nitrogen is exchanged for host photosynthates and enables the host to grow without a nitrogen supply. S. meliloti elicits the formation of nodules from the root cortex and invades the developing nodule cells through a host-derived structure termed an "infection thread." The internalized bacteria terminally differentiate to become nitrogen-fixing bacteroids; their differentiation is driven by a family of ho...

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