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Toxin-antitoxin systems of Mycobacterium smegmatis are essential for cell survival.

ABSTRACT: The role of chromosomal toxin-antitoxin (TA) modules in bacterial physiology remains enigmatic despite their abundance in the genomes of many bacteria. Mycobacterium smegmatis contains three putative TA systems, VapBC, MazEF, and Phd/Doc, and previous work from our group has shown VapBC to be a bona fide TA system. In this study, we show that MazEF and Phd/Doc are also TA systems that are constitutively expressed, transcribed as leaderless transcripts, and subject to autoregulation, and expression of the toxin component leads to growth inhibition that can be rescued by the cognate antitoxin. No phenotype was identified for deletions of the individual TA systems, but a triple deletion strain (?vapBC, mazEF, phd/doc), designated ?TA(triple), exhibited a survival defect in complex growth medium demonstrating an essential role for these TA modules in mycobacterial survival. Transcriptomic analysis revealed no significant differences in gene expression between wild type and the ?TA(triple) mutant under these conditions suggesting that the growth defect was not at a transcriptional level. Metabolomic analysis demonstrated that in response to starvation in complex medium, both the wild type and ?TA(triple) mutant consumed a wide range of amino acids from the external milieu. Analysis of intracellular metabolites revealed a significant difference in the levels of branched-chain amino acids between the wild type and ?TA(triple) mutant, which are proposed to play essential roles in monitoring the nutritional supply and physiological state of the cell and linking catabolic with anabolic reactions. Disruption of this balance in the ?TA(triple) mutant may explain the survival defect in complex growth medium.

SUBMITTER: Frampton R

PROVIDER: S-EPMC3285314 | biostudies-literature | 2012 Feb

REPOSITORIES: biostudies-literature

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Toxin-antitoxin systems of Mycobacterium smegmatis are essential for cell survival.

Frampton Rebekah R Aggio Raphael B M RB Villas-Bôas Silas G SG Arcus Vickery L VL Cook Gregory M GM

The Journal of biological chemistry 20111223 8

The role of chromosomal toxin-antitoxin (TA) modules in bacterial physiology remains enigmatic despite their abundance in the genomes of many bacteria. Mycobacterium smegmatis contains three putative TA systems, VapBC, MazEF, and Phd/Doc, and previous work from our group has shown VapBC to be a bona fide TA system. In this study, we show that MazEF and Phd/Doc are also TA systems that are constitutively expressed, transcribed as leaderless transcripts, and subject to autoregulation, and expressi ...[more]

PMID: 22199354

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Project description:Toxin-antitoxin (TA) systems are ubiquitous genetic elements in prokaryotes, but their biological importance is poorly understood. Mycobacterium smegmatis contains eight putative TA systems. Previously, seven TAs have been studied, with five of them being verified as functional. Here, we show that Ms0251-0252 is a novel TA system in that expression of the toxin Ms0251 leads to growth inhibition that can be rescued by the antitoxin Ms0252. To investigate the functional roles of TA systems in M. smegmatis, we deleted the eight putative TA loci and assayed the mutants for resistance to various stresses. Deletion of all eight TA loci resulted in decreased survival under starvation conditions and altered fitness when exposed to environmental stresses. Furthermore, we showed that deletion of the eight TA loci decreased resistance to phage infection in Sauton medium compared with the results using 7H10 medium, suggesting that TA systems might have different contributions depending on the nutrient environment. Furthermore, we found that MazEF specifically played a dominant role in resistance to phage infection. Finally, transcriptome analysis revealed that MazEF overexpression led to differential expression of multiple genes, including those related to iron acquisition. Altogether, we demonstrate that TA systems coordinately function to allow M. smegmatis to adapt to changing environmental conditions. IMPORTANCE Toxin-antitoxin (TA) systems are mechanisms for rapid adaptation of bacteria to environmental changes. Mycobacterium smegmatis, a model bacterium for studying Mycobacterium tuberculosis, encodes eight putative TA systems. Here, we constructed an M. smegmatis mutant with deletions of all eight TA-encoding genes and evaluated the resistance of these mutants to environmental stresses. Our results showed that different TA systems have overlapping and, in some cases, opposing functions in adaptation to various stresses. We suggest that complementary TA modules may function together to regulate the bacterial stress response, enabling adaptation to changing environments. Together, this study provides key insights into the roles of TA systems in resistance to various environmental stresses, drug tolerance, and defense against phage infection.

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Toxin-antitoxin systems of Mycobacterium smegmatis are essential for cell survival.

Publications

Toxin-antitoxin systems of Mycobacterium smegmatis are essential for cell survival.

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