Both Corynebacterium diphtheriae DtxR(E175K) and Mycobacterium tuberculosis IdeR(D177K) Are Dominant Positive Repressors of IdeR-Regulated Genes in M. tuberculosis

Abstract

The diphtheria toxin repressor (DtxR) is an important iron-dependent transcriptional regulator of known virulence genes in Corynebacterium diphtheriae. The mycobacterial iron-dependent repressor (IdeR) is phylogenetically closely related to DtxR, with high amino acid similarity in the DNA binding and metal ion binding site domains. We have previously shown that an iron-insensitive, dominant-positive dtxR(E175K) mutant allele from Corynebacterium diphtheriae can be expressed in Mycobacterium tuberculosis and results in an attenuated phenotype in mice (Y. C. Manabe, B. J. Saviola, L. Sun, J. R. Murphy, and W. R. Bishai, Proc. Natl. Acad. Sci. USA 96:12844-12848, 1999). In this paper, we report the M. tuberculosis IdeR(D177K) strain that has the cognate point mutation. We tested four known and predicted IdeR-regulated gene promoters (mbtI, Rv2123, Rv3402c, and Rv1519) using a promoterless green fluorescent protein (GFP) construct. GFP expression from these promoters was abrogated under low-iron conditions in the presence of both IdeR(D177K) and DtxR(E175K), a result confirmed by reverse transcription-PCR. The IdeR regulon can be constitutively repressed in the presence of an integrated copy of ideR containing this point mutation. These data also suggest that mutant IdeR(D177K) has a mechanism similar to that of DtxR(E175K); iron insensitivity occurs as a result of SH3-like domain binding interactions that stabilize the intermediate form of the repressor after ancillary metal ion binding. This construct can be used to elucidate further the IdeR regulon and its virulence genes and to differentiate these from genes regulated by SirR, which does not have this domain.

Genome Sequence of the Streptomycin-Producing MicroorganismStreptomyces griseusIFO 13350

“…Three NRPS gene clusters (SGR443 to SGR455, SGR6709 to SGR6717, and SGR6730 to SGR6742) appear to be involved in the biosynthesis of siderophores because they contain some genes for a siderophore-dependent iron transport system. The presence of IdeR (iron-dependent transcription regulator) box-like sequences (29) in the upstream regions of many genes within these three gene clusters (data not shown) also supports this idea.…”

Differential Proteomic Analysis of theBacillus anthracisSecretome: Distinct Plasmid and Chromosome CO2-Dependent Cross Talk Mechanisms Modulate Extracellular Proteolytic Activities

“…These domains represent bacterial versions of eukaryotic motifs encountered in many signal transduction proteins (38). Furthermore, bacterial proteins possessing SH3 domains were shown to be involved in the virulence of several pathogens, such as the diphtheria toxin repressor DtxR of Corynebacterium diphtheriae (89,149), the invasion protein InlB of Listeria monocytogenes (92), and the iron-dependent repressor IdeR of Mycobacterium tuberculosis (89) ( Table 3). The second abundant protein, BA3660, is HtrA, a chaperone and protease and a well-established virulence factor and potential vaccine candidate in many bacteria, which also has a role in the secretion process (Table 3).…”

Rapid Assessment of Antibacterial Activity against Mycobacterium ulcerans by Using Recombinant Luminescent Strains

“…The promoters used were (i) the widely used hsp60 (48) promoter from Mycobacterium bovis BCG, (ii) the Mycobacterium marinum G13 promoter (6) homolog (MuG13) amplified from M. ulcerans ATCC 19423, and (iii) the artificial mycobacterial optimal promoter (MOP) (22), which was modified and amplified from plasmid pMH29. The E. coli-mycobacterium shuttle vectors included extrachromosomal plasmids based on p60LUX (39), a kind gift from Richard L. Friedman (University of Arizona), and integrative plasmids derived from pMH94 (25) and pCK0601 (27) (Table 1).…”