The Irr and RirA Proteins Participate in a Complex Regulatory Circuit and Act in Concert To Modulate Bacterioferritin Expression in Ensifer meliloti 1021

Abstract

In this work we found that the <i>bfr</i> gene of the rhizobial species <i>Ensifer meliloti</i>, encoding a bacterioferritin iron storage protein, is involved in iron homeostasis and the oxidative stress response. This gene is located downstream of and overlapping the <i>smc03787</i> open reading frame (ORF). No well-predicted RirA or Irr boxes were found in the region immediately upstream of the <i>bfr</i> gene although two presumptive RirA boxes and one presumptive Irr box were present in the putative promoter of <i>smc03787</i> We demonstrate that <i>bfr</i> gene expression is enhanced under iron-sufficient conditions and that Irr and RirA modulate this expression. The pattern of <i>bfr</i> gene expression as well as the response to Irr and RirA is inversely correlated to that of <i>smc03787</i> Moreover, our results suggest that the small RNA SmelC759 participates in RirA- and Irr-mediated regulation of <i>bfr</i> expression and that additional unknown factors are involved in iron-dependent regulation.<b>IMPORTANCE</b><i>E. meliloti</i> belongs to the <i>Alphaproteobacteria</i>, a group of bacteria that includes several species able to associate with eukaryotic hosts, from mammals to plants, in a symbiotic or pathogenic manner. Regulation of iron homeostasis in this group of bacteria differs from that found in the well-studied <i>Gammaproteobacteria</i> In this work we analyzed the effect of <i>rirA</i> and <i>irr</i> mutations on <i>bfr</i> gene expression. We demonstrate the effect of an <i>irr</i> mutation on iron homeostasis in this bacterial genus. Moreover, results obtained indicate a complex regulatory circuit where multiple regulators, including RirA, Irr, the small RNA SmelC759, and still unknown factors, act in concert to balance <i>bfr</i> gene expression.