Toxin-antitoxin systems and their role in disseminating and maintaining antimicrobial resistance

Abstract

Toxin-antitoxin systems (TAs) are ubiquitous among bacteria and play a crucial role in the dissemination and evolution of antibiotic resistance, such as maintaining multi-resistant plasmids and inducing persistence formation. Generally, activities of the toxins are neutralised by their conjugate antitoxins. In contrast, antitoxins are more liable to degrade under specific conditions such as stress, and free active toxins interfere with essential cellular processes including replication, translation and cell-wall synthesis. TAs have also been shown to be responsible for plasmid maintenance, stress management, bacterial persistence and biofilm formation. We discuss here the recent findings of these multifaceted TAs (type I-VI) and in particular examine the role of TAs in augmenting the dissemination and maintenance of multi-drug resistance in bacteria.

Keywords: addictive systems; antimicrobial resistance; persistence; toxin–antitoxins.

Figure 1.

The intracellular targets of TAloci. TA loci usually encode two genes: one is a stable toxin and the other one is an unstable antitoxin. The antitoxins sequester the toxins but are subjected to proteolytic degradation by cellular proteases (Lon or ClpXP) under stress condition. Consequently, free active toxins alter cellular processes including DNA replication, translation or cell-wall synthesis, which ultimately results in slow growth or the formation of highly drug-tolerant persisters. TAs examples for the cellular targets are given below. (1) Zeta toxin inhibits cell-wall synthesis by specific phosphorylation of peptidoglycan precursor UNAG. (2) TisB, HokB and GhoT: the products of TisB and HokB can decrease the level of membrane potential motive force (pmf) and ATP by inserting into cytoplasmic membrane, while protein GhoT can lyse cell membrane and change cell morphologies. (3) CcdB and ParE inhibit DNA replication by poison DNA gyrase. (4) Doc inhibits translation by phosphoralation of elongation factor Tu (EF-Tu). (5–7) MazF, RelB and VapC inhibit translation by cleavage of mRNAs like single-stranded mRNA, A-site on ribosome and initiator tRNA^fMet, respectively. (8) HipA inhibits translation by phosphoration of GltX. tRNA:fMet indicates initiator tRNA at P site carried formyl methionine; ‘p’ indicates phosphorylation.

Figure 2.

Model of the TisB toxin induced SOS response and persistence formation. (1) Antibiotics (like ciprofloxacin) kill bacteria by damaging their DNA; (2) the SOS response gene recA is activated by the accumulation of single-stranded DNA (ssDNA). (3) The induced RecA interacts with the LexA repressor, leading to facilitate the LexA autocleavage. (4) Once the degradation of LexA repressor, the SOS genes are induced to repair DNA damage. (5) Concurrently, the SOS induction results in cleavage of the istR-1 pool. (6) The expression of tisB is activated by degrading the level of antitoxin IstR-1, this causes membrane damage and the loss of membrane proton motive force (pmf) and ATP level (7); as a result, drugs were drive to out of the cells, leading to persister formation (8). The green and purple arrowheads representing the promoters under LexA control and istR-1 constitutive promoter, respectively.

Figure 3.

(p)ppGpp-hipA mediated persister pathway. In response to particular stresses, SpoT and RelA are activated to synthesise the nucleotide alarmone (p)ppGpp, The increased (p)ppGpp levels lead to the accumulation of inoganic polyphosphate (PolyP) through inhibition of exopolyphosphatase (PPX), that the cellular enzyme to degrades PolyP. The accumulated PolyP combines with Lon protease preferentially to cleave the antitoxin HipB, resulting in an excess of toxin HipA. In return, free active toxin HipA inactivates GltX by phosphorylation of its ATP-binding site Ser²³⁹, with the consequence of uncharged tRNA with glutamate (tRNA^Glu) accumulation in the cell. Uncharged tRNA^Glu loads at empty ribosomal sites and triggers the activation of RelA to more (p)ppGpp synthesis, promoting cells entry into dormant state. Note that SpoT and RelA are bifunctional synthetase-hydrolase enzyme, if the stresses have been removed, they can hydrolase (p)ppGpp and bring cells to normal growth (Dalebroux and Swanson 2012). The red box labelled with ‘?’ indicates that the link between stringent response-associated genes (including ppGpp, Lon, PolyP) and TAs has been exploring in some TAs, such as relBE, mazEF and yefM-yeoB. It has been proved that the activation of toxin MazF and YoeB is dependent on the Lon-mediated degradation of their cognates, antitoxins, but not on the accumulation of PolyP and ppGpp (Christensen et al. , ; Ramisetty et al. 2016).