Toxin ζ Reversible Induces Dormancy and Reduces the UDP-N-Acetylglucosamine Pool as One of the Protective Responses to Cope with Stress

Abstract

Toxins of the ζ/PezT family, found in the genome of major human pathogens, phosphorylate the peptidoglycan precursor uridine diphosphate-N-acetylglucosamine (UNAG) leading to unreactive UNAG-3P. Transient over-expression of a PezT variant impairs cell wall biosynthesis and triggers autolysis in Escherichia coli. Conversely, physiological levels of ζ reversibly induce dormancy produce a sub-fraction of membrane-compromised cells, and a minor subpopulation of Bacillus subtilis cells become tolerant of toxin action. We report here that purified ζ is a strong UNAG-dependent ATPase, being GTP a lower competitor. In vitro, ζ toxin phosphorylates a fraction of UNAG. In vivo, ζ-mediated inactivation of UNAG by phosphorylation does not deplete the active UNAG pool, because expression of the toxin enhances the efficacy of genuine cell wall inhibitors (fosfomycin, vancomycin or ampicillin). Transient ζ expression together with fosfomycin treatment halt cell proliferation, but ε2 antitoxin expression facilitates the exit of ζ-induced dormancy, suggesting that there is sufficient UNAG for growth. We propose that ζ induces diverse cellular responses to cope with stress, being the reduction of the UNAG pool one among them. If the action of ζ is not inhibited, e.g., by de novo ε2 antitoxin synthesis, the toxin markedly enhances the efficacy of antimicrobial treatment without massive autolysis in Firmicutes.

The chromosomal SezAT toxin–antitoxin system promotes the maintenance of the SsPI‐1 pathogenicity island in epidemic Streptococcus suis

“…Therefore, the inhibitory effect of SezT in this study should be interpreted as a consequence of triggering cell death rather than cell stasis. Previously, it was reported that physiological levels of Zeta toxin reversibly induce dormancy in B. subtilis and E. coli cells (Tabone et al ., 2014a,b). Conversely, upon massive overexpression, the PezT toxin triggers autolysis with few intact cells with an ovoid morphology (Mutschler et al ., ), but such massive autolysis was not observed upon Zeta overexpression (Zielenkiewicz and Ceglowski, ).…”

Keeping the Wolves at Bay: Antitoxins of Prokaryotic Type II Toxin-Antitoxin Systems

“…Therefore, ζ functions to inhibit bacterial cell wall synthesis (Mutschler and Meinhart, 2011 ; Mutschler et al, 2011 ). However, a recent paper reported that expression of ζ only reduced the UNAG pool and did not totally deplete it with transient expression of ζ (120 min) reversibly inducing a dormant state that was subsequently rescued by ε expression (Lioy et al, 2012 ; Tabone et al, 2014 ). It was proposed that ζ expression induces diverse responses to cope with stress with reduction in the UNAG levels as one of these responses rather than triggering a latent suicide program by depleting the UNAG pool (Tabone et al, 2014 ).…”

First insights into a type II toxin-antitoxin system from the clinical isolate Mycobacterium sp. MHSD3, similar to epsilon/zeta systems

“…Moreover, two different mechanisms of action for the epsilon/zeta systems have been described so far. In the first one, the cell wall formation is affected through phosphorylation of the UDP-N-acetylglucosamine (UNAG) on its 3’-hydroxyl group [ 20 , 22 ], causing a reversible growth arrest [ 23 , 24 ]. The second mechanism of action implies the phosphorylation of nicotinamide adenine dinucleotide (NAD) or its precursor, nicotinic acid adenine dinucleotide (NAAD), at the adenosine 3’-hydroxil group, affecting NAD/NAAD-dependent pathways [ 25 ].…”