The Mycobacterium tuberculosis IdeR is a dual functional regulator that controls transcription of genes involved in iron acquisition, iron storage and survival in macrophages

In this work, we characterize genes in Mycobacterium tuberculosis that are regulated by IdeR ((i) under bar ron-(de) under bar pendent (r) under bar e gulator), an iron-responsive DNA-binding protein of the DtxR family that ha s been shown to regulate iron acquisition in Mycobacterium smegmatis. To id entify some of the genes that constitute the IdeR regulon, we searched the M. tuberculosis genome for promoter regions containing the consensus IdeR/D xR binding sequence. Genes preceded by IdeR boxes included a set encoding p roteins necessary for iron acquisition, such as the biosynthesis of siderop hores (mbtA, mbtB, mbtI), aromatic amino acids (pheA, hisE, hisB-like) and others annotated to be involved in the synthesis of iron-storage proteins ( bfrA, bfrB). Some putative IdeR-regulated genes identified in this search e ncoded proteins predicted to be engaged in the biosynthesis of lipopolysacc haride (LPS)-like molecules (rv3402c), lipids (acpP) and peptidoglycan (mur B). We analysed four promoter regions containing putative IdeR boxes, mbtA- mbtB, mbl, rv3402c and bfrA-bfd, for interaction with IdeR and for iron-dep endent expression. Gel retardation experiments and DNase footprinting analy ses with purified IdeR showed that IdeR binds to these IdeR boxes in vitro. Analysis of the promoters by primer extension indicated that the IdeR boxe s are located near the -10 position of each promoter, suggesting that IdeR acts as a transcriptional repressor by blocking RNA polymerase binding. Usi ng quantitative reverse transcriptase-polymerase chain reaction (RT-PCR) co upled to molecular beacons, we showed that mRNA levels of mbtA, mbtB, mbtl, rv3402c and bfd are induced 14- to 49-fold in cultures of M. tuberculosis starved for iron, whereas mRNA levels of bfrA decreased about threefold. We present evidence that ldeR not only acts as a transcriptional repressor bu t also functions as an activator of bfrA. Three of the IdeR- and iron-repre ssed genes, mbtB, mbtl and rv3402c, were induced during M. tuberculosis inf ection of human THP-1 macrophages.