The RES domain toxins of RES‐Xre toxin‐antitoxin modules induce cell stasis by degrading NAD⁺

Abstract

Summary Type II toxin‐antitoxin (TA) modules, which are important cellular regulators in prokaryotes, usually encode two proteins, a toxin that inhibits cell growth and a nontoxic and labile inhibitor (antitoxin) that binds to and neutralizes the toxin. Here, we demonstrate that the res‐xre locus from Photorhabdus luminescens and other bacterial species function as bona fide TA modules in Escherichia coli. The 2.2 Å crystal structure of the intact Pseudomonas putida RES‐Xre TA complex reveals an unusual 2:4 stoichiometry in which a central RES toxin dimer binds two Xre antitoxin dimers. The antitoxin dimers each expose two helix‐turn‐helix DNA‐binding domains of the Cro repressor type, suggesting the TA complex is capable of binding the upstream promoter sequence on DNA. The toxin core domain shows structural similarity to ADP‐ribosylating enzymes such as diphtheria toxin but has an atypical NAD+‐binding pocket suggesting an alternative function. We show that activation of the toxin in vivo causes a depletion of intracellular NAD+ levels eventually leading to inhibition of cell growth in E. coli and inhibition of global macromolecular biosynthesis. Both structure and activity are unprecedented among bacterial TA systems, suggesting the functional scope of bacterial TA toxins is much wider than previously appreciated.

The extracellular contractile injection system is enriched in environmental microbes and associates with numerous toxins

“…We iden fied several cases of toxin-immunity gene pairs next to eCIS operons suppor ng the an bacterial role of these eCIS loci as they require immunity genes (an toxins) to protect from self-intoxica on ( Figure 5A). In one case, The toxin, EAT5, resembles RES (Figure 4b), an NADase that is accompanied by an an toxin called Xre 50,51 . We iden fied a gene, EX18DRAFT_03239, encoding an unknown protein located upstream of the toxin.…”

HORMA domain proteins and a Pch2-like ATPase regulate bacterial cGAS-like enzymes to mediate bacteriophage immunity

“…"-LEU -TRP" refers to non-selective growth media, and "-LEU -TRP -HIS -ADE" refers to selective growth media. This operon encodes a RES-Xre type toxin-antitoxin gene pair (Makarova et al 2009;Skjerning et al 2018), labeled A/T, which was not tested for protein-protein interactions. "EV" refers to empty vector controls.…”

Chromosomal toxin-antitoxin systems in Pseudomonas putida are rather selfish than beneficial

“…In order to analyse the benefits and costs of multitude TA systems, the 13 TA loci were deleted from P. putida PaW85 (isogenic to KT2440) chromosome in the following order: graTA (PP_1586-1585), res-xre (PP_2433-2434), higBA (PP_1199-1198), hicAB-1 (PP_1480-1479), relE 2 -higA 2 (PP_5435-PP_0274), mqsRA (PP_4205-4204), mazEF (PP_0770-0771), PP_1716-1717, brnTA (PP_4530-4529), PP_4151-4152, relBE (PP_1268-1267), yefM-yoeB (PP_2940-2939) and relB 2 -parE (PP_2499-2500) ( Table 1). The functionality of only four (graTA, res-xre, mqsRA and mazEF) out of these 13 TA modules has been studied experimentally 28,38,39,[41][42][43] , while the other loci are putative TA pairs predicted in silico only 36 . TADB2 database annotates two more putative TA loci, the hicAB-2 (PP_3900-3899) and PP_3032-3033, which are located in prophage 1 and prophage 2 genomes, respectively 36,44 .…”

“…However, akin to graTA, the absence of the whole mqsRA locus did not affect the persister phenotype 41 . RES toxin of the RES-Xre module is structurally related to NADase toxins and its overexpression results in growth suppression due to NAD + depletion 20,42 . Unfortunately, the consequences of the chromosomal deletion of res-xre genes to P. putida were not examined 42 .…”

“…RES toxin of the RES-Xre module is structurally related to NADase toxins and its overexpression results in growth suppression due to NAD + depletion 20,42 . Unfortunately, the consequences of the chromosomal deletion of res-xre genes to P. putida were not examined 42 . The P. putida MazEF system has been also analysed only in mechanistic sense in a study demonstrating that the purified MazF acts as an mRNase which cleaves UAC sequence 43 .…”