The Escherichia coli mqsR and ygiT Genes Encode a New Toxin-Antitoxin Pair

Abstract

Toxin-antitoxin (TA) systems are plasmid-or chromosome-encoded protein complexes composed of a stable toxin and a short-lived inhibitor of the toxin. In cultures of Escherichia coli, transcription of toxin-antitoxin genes was induced in a nondividing subpopulation of bacteria that was tolerant to bactericidal antibiotics. Along with transcription of known toxin-antitoxin operons, transcription of mqsR and ygiT, two adjacent genes with multiple TA-like features, was induced in this cell population. Here we show that mqsR and ygiT encode a toxin-antitoxin system belonging to a completely new family which is represented in several groups of bacteria. The mqsR gene encodes a toxin, and ectopic expression of this gene inhibits growth and induces rapid shutdown of protein synthesis in vivo. ygiT encodes an antitoxin, which protects cells from the effects of MqsR. These two genes constitute a single operon which is transcriptionally repressed by the product of ygiT. We confirmed that transcription of this operon is induced in the ampicillin-tolerant fraction of a growing population of E. coli and in response to activation of the HipA toxin. Expression of the MqsR toxin does not kill bacteria but causes reversible growth inhibition and elongation of cells.Bacterial toxin-antitoxin (TA) systems (for reviews, see references 16 and 54) are complexes consisting of a stable toxin component and a short-lived antitoxin. Toxins of TA systems are autotoxic; they target vital functions of the producing bacterium itself. TA complexes were discovered because they are plasmid-stabilizing entities. When a bacterium carries a plasmid which encodes a toxin and an antitoxin, both molecules are produced continuously and have no effect on the activities of the cell. When the plasmid is lost during cell division, the toxin is released and kills or inhibits the cell, because the unstable antitoxin is degraded faster. Chromosomal TA systems were found later (36), and comparative studies determined that they are widespread in free-living bacteria (35, 41).There are two different types of bacterial TA systems, which depend on the nature of the antitoxin. In type I TA systems, the antitoxin is a small regulatory RNA (15, 18). In type II TA systems, which are relevant to this study, both the toxin and the antitoxin are proteins. Protein antitoxins neutralize toxins by direct interaction, forming catalytically inactive complexes. All known TA genes are organized as operons. The antitoxin is usually encoded by the first gene and always acts as a transcriptional autorepressor of the operon either alone or in a complex with the toxin molecule. Thus, antitoxins control toxin activity in two ways: through direct binding and through transcriptional regulation (17).The toxins of type II systems attack essential functions of a bacterial cell, either protein synthesis through cleavage of free or ribosome-bound mRNA (e.g., RelE, MazF, HigB, and HicA) (10,11,25,43) or the replication and integrity of DNA through interference with DNA gyrase (e.g., CcdB and ...

Toxin-Antitoxin Systems Influence Biofilm and Persister Cell Formation and the General Stress Response

“…The three-dimensional structures show that toxin MqsR is an RNase similar to RelE and YoeB (10) that cleaves mRNA primarily at GCU sites (78) and that antitoxin MqsA binds DNA via its helix-turn-helix motif in its C-terminal domain while binding MqsR at its N-terminal domain (10). These initial results linking MqsR/MqsA to biofilm formation were confirmed recently using 48-h biofilms in which deletion of mqsRA reduced biofilm formation (34) as found in the original report (26). Hence, these studies served to identify a 10th role for TA modules in cell physiology: influencing biofilm formation.…”

Global Expression Profile of Biofilm Resistance to Antimicrobial Compounds in the Plant-Pathogenic Bacterium Xylella fastidiosa Reveals Evidence of Persister Cells

“…Therefore, bacteria have evolved two complementary and highly redundant strategies to protect themselves from antimicrobials: multidrug efflux and, when this fails, tolerance of persister cells (20,21). Few genes related to MDR were induced in our experimental conditions and, moreover, subinhibitory and inhibitory concentrations of both antimicrobial compounds led to induction of TA genes, which has been associated with the persistence phenotype (8,16,32). The majority of induced TA systems were found in phage regions, but their roles in bacteriophages have not been confirmed.…”

“…Since the expression of TA systems directly correlates to the persistence phenotype in other bacteria (16,17,39), and since we observed the induction of expression of TA genes even in subinhibitory concentrations of copper and tetracycline, we hypothesize that a pretreatment with antimicrobial compounds could lead to an increase in the number of persister cells. To test this hypothesis, we performed an experiment using three different conditions.…”

“…The expression of the toxin reduces metabolic functions, leading the cells to enter in a dormant, multidrug-tolerant state (21), which could be reversible (16). In addition, TA systems influence biofilm formation and the general stress response in different bacterial species (39).…”

Structure of the Escherichia coli Antitoxin MqsA (YgiT/b3021) Bound to Its Gene Promoter Reveals Extensive Domain Rearrangements and the Specificity of Transcriptional Regulation

“…The antitoxin MqsA is even more unusual because it is the first antitoxin that requires a metal, zinc, for structural stability (19); it is the only E. coli antitoxin that is structured throughout its entire sequence; and it is the first antitoxin demonstrated to bind DNA via its C-terminal and not N-terminal domain (19, 27, 36 -39). Critically, in addition to binding its own promoter (19,34,35,40), MqsA and the MqsR⅐MqsA complex are also the only known antitoxin/TA pair that bind not only their own promoter but also the promoters of other genes that play important roles in E. coli physiology, including mcbR and spy (19) as well as cspD (31). Moreover, although 14 antitoxin structures have been solved to date, their primary sequences are highly divergent when compared with MqsA (supplemental Table S1) because the sequence identity between MqsA and the most closely related antitoxin, the E. coli protein HigA, is only 13%.…”