Global Analysis of the Regulon of the Transcriptional Repressor LexA, a Key Component of SOS Response in Mycobacterium tuberculosis

Smollett, Katherine; Smith, Kimberley M.; Kahramanoglou, Christina; Arnvig, Kristine B.; Buxton, Roger S.; Davis, E. A.

doi:10.1074/jbc.m112.357715

Cited by 74 publications

(83 citation statements)

References 48 publications

(113 reference statements)

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“…Tandem configurations resembling the conserved palindromes shown in Fig. 1 have been reported previously for functional LexA-binding sites upstream of lexA and other SOS regulated genes (9,12,14,47,48). Furthermore, the identified palindromes display a consensus sequence (GAACG-N 2 -CGTTC) that is strongly reminiscent of the B. subtilis LexA-binding motif.…”

Section: Resultssupporting

confidence: 75%

“…Furthermore, the identified palindromes display a consensus sequence (GAACG-N 2 -CGTTC) that is strongly reminiscent of the B. subtilis LexA-binding motif. Functional LexA-binding motifs bearing strong similarity to the B. subtilis one have been reported in Firmicutes, Actinobacteria, Cyanobacteria, and chloroflexi and for a LexA paralog in Gammaproteobacteria (26,48,(49)(50)(51)(52)(53)(54). Taken together, these data strongly suggested that the identified tandem palindromes were functional LexA-binding sites.…”

Section: Resultssupporting

confidence: 62%

“…LexA proteins recognizing B. subtilis-type motifs have been reported in the Actinobacteria, the Cyanobacteria, the chloroflexi, and some Gammaproteobacteria (26,48,(49)(50)(51)(52)(53)(54). Even though phylogenetic inference cannot fully resolve the origin of the Gallionellales LexA protein, the fact that it targets a B. subtilis-like binding motif suggested that a detailed analysis of its DNA-binding domain might shed light onto its evolutionary origin.…”

Section: Resultsmentioning

confidence: 99%

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An SOS Regulon under Control of a Noncanonical LexA-Binding Motif in the Betaproteobacteria

et al. 2015

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The SOS response is a transcriptional regulatory network governed by the LexA repressor that activates in response to DNA damage. In the Betaproteobacteria, LexA is known to target a palindromic sequence with the consensus sequence CTGT-N 8 -ACAG. We report the characterization of a LexA regulon in the iron-oxidizing betaproteobacterium Sideroxydans lithotrophicus. In silico and in vitro analyses show that LexA targets six genes by recognizing a binding motif with the consensus sequence GAACGaaCGTTC, which is strongly reminiscent of the Bacillus subtilis LexA-binding motif. We confirm that the closely related Gallionella capsiferriformans shares the same LexA-binding motif, and in silico analyses indicate that this motif is also conserved in the Nitrosomonadales and the Methylophilales. Phylogenetic analysis of LexA and the alpha subunit of DNA polymerase III (DnaE) reveal that the organisms harboring this noncanonical LexA form a compact taxonomic cluster within the Betaproteobacteria. However, their lexA gene is unrelated to the standard Betaproteobacteria lexA, and there is evidence of its spread through lateral gene transfer. In contrast to other reported cases of noncanonical LexA-binding motifs, the regulon of S. lithotrophicus is comparable in size and function to that of many other Betaproteobacteria, suggesting that a convergent SOS regulon has reevolved under the control of a new LexA protein. Analysis of the DNA-binding domain of S. lithotrophicus LexA reveals little sequence similarity with that of other LexA proteins targeting similar binding motifs, suggesting that network structure may limit site evolution or that structural constrains make the B. subtilis-type motif an optimal interface for multiple LexA sequences. IMPORTANCEUnderstanding the evolution of transcriptional systems enables us to address important questions in microbiology, such as the emergence and transfer potential of different regulatory systems to regulate virulence or mediate responses to stress. The results reported here constitute the first characterization of a noncanonical LexA protein regulating a standard SOS regulon. This is significant because it illustrates how a complex transcriptional program can be put under the control of a novel transcriptional regulator. Our results also reveal a substantial degree of plasticity in the LexA recognition domain, raising intriguing questions about the space of protein-DNA interfaces and the specific evolutionary constrains faced by these elements.

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Section: Resultssupporting

confidence: 75%

Section: Resultssupporting

confidence: 62%

Section: Resultsmentioning

confidence: 99%

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An SOS Regulon under Control of a Noncanonical LexA-Binding Motif in the Betaproteobacteria

et al. 2015

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“…The recA gene encodes a central regulator of the mycobacterial DNA damage (SOS) response and is induced after the exposure of M. tuberculosis to compounds or stimuli that are directly genotoxic (i.e., those which chemically modify or damage the nucleic acid) or which inhibit DNA metabolic functions such as chromosomal replication (15,30,31). In addition to the canonical LexA/RecAdependent SOS response, M. tuberculosis possesses a second, larger DNA damage-inducible regulon which is controlled by a ClpR-like regulator that recognizes a RecA-independent promoter (RecA-NDp) motif (32).…”

mentioning

confidence: 99%

“…In utilizing both radA and recA in the construction of the DNA damage reporters, we sought to ensure our ability to detect a broad range of genotoxic agents. Moreover, since recA itself is included in both LexA/RecA-dependent and RecA-ND regulons (31,32), the use of radA and recA reporters in parallel was intended to maintain the potential to identify compounds eliciting a LexA/RecA response only: such compounds (if they exist) would induce recA promoter expression but not radA.…”

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confidence: 99%

Bioluminescent Reporters for Rapid Mechanism of Action Assessment in Tuberculosis Drug Discovery

Naran

Moosa

Barry

et al. 2016

Antimicrob Agents Chemother

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cThe tuberculosis (TB) drug discovery pipeline is fueled by compounds identified in whole-cell screens against the causative agent, Mycobacterium tuberculosis. Phenotypic screening enables the selection of molecules that inhibit essential cellular functions in live, intact bacilli grown under a chosen in vitro condition. However, deducing the mechanism of action (MOA), which is important to avoid promiscuous targets, often requires significant biological resources in a lengthy process that risks decoupling medicinal chemistry and biology efforts. Therefore, there is a need to develop methods enabling rapid MOA assessment of putative "actives" for triage decisions. Here, we describe a modified version of a bioluminescence reporter assay that allows nondestructive detection of compounds targeting either of two macromolecular processes in M. tuberculosis: cell wall biosynthesis or maintenance of DNA integrity. Coupling the luxCDABE operon from Photorhabdus luminescens to mycobacterial promoters driving expression of the iniBAC operon (PiniB-LUX) or the DNA damage-inducible genes, recA (PrecA-LUX) or radA (PradA-LUX), provided quantitative detection in real time of compounds triggering expression of any of these promoters over an extended 10-to 12-day incubation. Testing against known anti-TB agents confirmed the specificity of each reporter in registering the MOA of the applied antibiotic in M. tuberculosis, independent of bactericidal or bacteriostatic activity. Moreover, profiles obtained for experimental compounds indicated the potential to infer complex MOAs in which multiple cellular processes are disrupted. These results demonstrate the utility of the reporters for early triage of compounds based on the provisional MOA and suggest their application to investigate polypharmacology in known and experimental anti-TB agents.

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The SOS Response Modulates Bacterial Pathogenesis

Bertok

2016

Stress and Environmental Regulation of Gene Expression and Adaptation in Bacteria

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Global Analysis of the Regulon of the Transcriptional Repressor LexA, a Key Component of SOS Response in Mycobacterium tuberculosis

Cited by 74 publications

References 48 publications

An SOS Regulon under Control of a Noncanonical LexA-Binding Motif in the Betaproteobacteria

An SOS Regulon under Control of a Noncanonical LexA-Binding Motif in the Betaproteobacteria

Bioluminescent Reporters for Rapid Mechanism of Action Assessment in Tuberculosis Drug Discovery

The SOS Response Modulates Bacterial Pathogenesis

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