RirA regulon.
Analysis of upstream regions of candidate iron uptake genes in α-proteobacteria possessing a close RirA ortholog (the Rhizobiales species except the Bradyrhizobiaceae group; Figure 1) yielded a conserved palindromic motif with the consensus 5′-TGA-(N9)-TCA-3′ (Figure 4A). The newly proposed RirA recognition motif, named RirA-box, overlaps and slightly differs from the previously identified IRO motif in the promoter regions of some genes in R. leguminosarum that were shown to be repressed by RirA in Fe-replete conditions [35]. The newly constructed recognition profile was used to scan the genomes of Rhizobiales for additional candidate RirA-boxes, allowing us to tentatively identify many additional candidate targets of RirA (Table S1). For all these species except, notably, the Bradyrhizobiaceae group, we observed many highly significant matches in the upstream regions of genes involved in iron metabolism. The largest class of genes with candidate RirA-boxes in upstream regions is involved in iron uptake and storage.
Figure 4  Sequence Logos for the Predicted Regulatory Sites in α-Proteobacteria
(A) RirA-box (IRO) in eight species from the Rhizobiales order (four Rhizobiaceae, two Mesorhizobium species, Brucella, and Bartonella).
(B) Iron-Rhodo-box in the Rhodobacteraceae.
(C) Mur-box (MRS) in the Rhodobacteraceae/Rhizobiales.
(D) Furα-box in other α-proteobacteria species.
(E) IscRα-I motif in the Rhodobacterales, the Rickettsia, Pelagibacter, Oceanicaulis, Caulobacter, Parvularcula, Rhodospirillum, and Magnetospirillum species.
(F) IscRα-II motif in the Sphingomonadales and Gluconobacter species.

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