Les systèmes toxine-antitoxine (STA) sont généralement constitués de deux petites protéines cytoplasmiques : une toxine stable et une antitoxine instable capable de neutraliser la toxine et de réprimer l’expression de l’opéron toxine-antitoxine. Une étude menée au laboratoire avait mis en évidence que la perte du gène hicB3 (ypo3369) de Y. pestis, codant une antitoxine solitaire putative, entraine un retard de la croissance bactérienne in vitro et une atténuation de la virulence dans un modèle murin de peste bubonique (Pradel et al., 2014). Par analyse in silico, nous avons détecté, en amont de hicB3, un petit gène non annoté candidat pour coder la toxine HicA3. La surproduction de HicA3 provoque la bactériostase chez Escherichia coli et Y. pestis et la production subséquente de HicB3 restaure la croissance. HicA3 et HicB3 constituent donc un STA fonctionnel. Cependant, la perte du STA HicA3B3 n’affecte pas la virulence de Y. pestis dans un modèle murin de peste bubonique. Nous avons ensuite purifié et caractérisé les protéines HicA3 et HicB3. La toxine HicA3 est une ribonucléase monomérique de 66 aa qui comporte un résidu histidine catalytique essentiel pour son activité. L’antitoxine HicB3 a une double fonction : elle interagit avec HicA3 pour la neutraliser et elle réprime le promoteur de l’opéron hicA3B3. Des expériences de retard sur gel et de fusions transcriptionnelles avec un gène rapporteur ont révélé que l’antitoxine HicB3 et le complexe HicA3-HicB3 se fixent sur deux opérateurs chevauchant les boîtes -10 et -35 du promoteur PhicA3. Nous avons également résolu la structure cristalline de l’antitoxine HicB3 et celle du complexe HicA3-HicB3. HicB3 est un tétramère qui comporte deux domaines de fixation à l’ADN du type ruban-hélice-hélice et deux domaines de neutralisation de la toxine. / Toxin-antitoxin systems (TAS) are generally constituted by two small cytoplasmic proteins: a stable toxin and an unstable antitoxin which neutralizes the toxin and represses the expression of the toxin-antitoxin operon. In previous research, our lab found that Yersinia pestis lacking the hicB3 (ypo3369) gene, encoding a putative orphan antitoxin, has a growth defect in vitro and is attenuated for virulence in a murine model of bubonic plague (Pradel et al., 2014). In silico analysis revealed a small gene upstream of hicB3, encoding a putative toxin that we called HicA3. HicA3 overproduction generates bacteriostasis of Escherichia coli and Y. pestis, and the subsequent production of HicB3 restores cell growth. HicA3 and HicB3 thus constitute a functional TAS. However, the lack of the HicA3B3 TAS does not affect Y. pestis virulence in a murine model of bubonic plague. We then purified and characterized the HicA3 and HicB3 proteins. The HicA3 toxin is a monomeric 66-aa ribonuclease with a catalytic histidine residue required for its activity. The HicB3 antitoxin has two functions: it binds and neutralizes HicA3 and it represses the hicA3B3 operon promoter. Gel-shift assays and transcriptional reporter fusion experiments showed that both HicB3 and the HicA3-HicB3 complex bind to two operators overlapping the -10 and -35 boxes of the PhicA3 promoter. We also solved the crystal structures of the HicB3 antitoxin and the HicA3-HicB3 complex. HicB3 is a tetramer with two DNA binding domains of the ribbon-helix-helix type and two toxin neutralization domains.

Cell biology is a branch of science with a seemingly infinite abundance of interesting phenomena which are essential to our understanding of life and which may potentially drive the development of technology that improves our lives. Among the open ended questions within the field, an understanding of how gene networks are affected by limited cellular components is both broad and rich with interest. Common to all cellular systems are enzymes which perform many tasks within cells without which organisms could not remain healthy. Here are presented several explorations of enzymatic processing as well as a tool constructed for this purpose. More specifically, these works consider the effect of coupling of gene networks via competition for enzymes found within the cell. It is shown that a limitation on the number of available enzymes permits the formation of bottlenecks which drastically affect molecular dynamics within cells. These effects potentially afford cell behaviors that in part explain the impressive robustness of life to constantly fluctuating environments. / Ph. D.

A major challenge in biology is to understand and predict the effect of mutations on protein structure, stability and function. Chapter 1 provides a general introduction on protein sequence-structure relationships and use of the CcdAB toxin-antitoxin system as a model to study molecular determinants of mutant phenotypes. In Chapter 2, we describe the use of saturation mutagenesis combined with deep sequencing to determine phenotypes for 1664 single-site mutants of the E. coli cytotoxin, CcdB. We examined multiple expression levels, effects of multiple chaperones and proteases and employed extensive in vitro characterization to understand how mutations affect these phenotypes. While general substitution preferences are known, eg polar residues preferred at exposed positions and non-polar ones at buried positions, we show that depth from the surface is important and that there are distinctly different energetic penalties for each specific polar, charged and aromatic amino acid introduced at buried positions. We also show that over-expression of ATP independent chaperones can rescue mutant phenotypes. Other studies have primarily looked at effects of ATP dependent chaperone expression on phenotype, where it is not possible to say whether mutational effects on folding kinetics or thermodynamic stability are the primary determinant of altered phenotypes, since there is energy input with these chaperones. The data suggest that mutational effects on folding rather than stability determine the in vivo phenotype of CcdB mutants. This has important implications for efforts to predict phenotypic effects of mutations and in protein design. While looking at the mutational landscape of a given gene from an evolutionary perspective, it is important to establish the genotype-phenotype relationships under physiologically relevant conditions. At the molecular level, the relationship between gene sequence and fitness has implications for understanding both evolutionary processes and functional constraints on the encoded proteins. Chapter 3 describes a methodology to test the fitness of individual CcdB mutants in E.coli over several generations by monitoring the rate of plasmid loss. We also propose a methodology for high throughput analysis of a pool of CcdB mutants using deep sequencing to quantitate the relative population of each mutant in a population of E.coli cells, grown for several generations and build the fitness landscape. While the F-plasmid based CcdAB system is known to be involved in plasmid maintenance through post-segregational killing, recent identification of ccdAB homologs on the chromosome, including in pathogenic strains of E.coli and other bacteria, has led to speculations on their functional role on the chromosome. In Chapter 4, we show that both the native ccd operon of the E.coli O157 strain as well as the ccd operon from the F- plasmid when inserted on the E.coli chromosome lead to protection from cell death under multiple antibiotic stress conditions through formation of persisters. Both the ccdF and ccdO157 operons may share common mechanisms for activation under stress conditions and also display weak cross activation. The chromosomal toxin shows weaker activity as compared to the plasmidic counterpart and is therefore less efficient in causing cell death. This has important implications in generation of potential therapeutics that target these TA systems. Chapter 5 describes the use of site-saturation mutagenesis coupled with deep sequencing to infer mutational sensitivity for the intrinsically disordered antitoxin, CcdA. The data allows us to make comparisons between overall as well as residue specific mutational sensitivity patterns with that of globular proteins, like CcdB (described in Chapter 2) and study toxin- antitoxin interaction and regulation through saturation suppressor mutagenesis. Interestingly, we found several examples of synonymous point mutations in CcdA that lead to loss of its activity. In Chapter 6 we attempt to explore the molecular bases for some of these synonymous mutations. In most cases the mutated codon has a similar overall codon preference to the WT one. Initial findings suggest a change in mRNA structure leading to change in CcdB: CcdA ratio, thereby causing cell death. These observations have important implications, because TA systems are ubiquitous, highly regulated and are known to be involved in multiple functions including drug tolerance. However a role for RNA structure in their regulation has not been shown previously. Appendix–I lists the mutational sensitivity scores for the CcdB mutants. Phenotypes for CcdA mutants obtained through deep sequencing have been tabulated in Appendix-II. Overall, we provide extensive datasets for mutational sensitivities of a globular (CcdB) and an intrinsically disordered protein (CcdA). Exploration of the molecular determinants of these mutant phenotypes not only provides interesting insights into CcdAB operon function but is also useful in understanding various aspects of protein stability, folding and activity as well as regulation of gene expression in bacteria.

Bacillus subtilis is a gram positive soil bacterium that is surrounded by many other microorganisms its environment. That is why it is necessary for the bacterium to be able to fight with these microorganisms for the nutrients and living space. B. subtilis contains the modules in its genetic make-up that improve its ability to compete. These modules are called the toxin-antitoxin systems. This Diploma Thesis is trying to identify yet undescribed extracellular toxins produced by the wild type BSB1 strain of B. subtilis. The related microorganism Bacillus megaterium was used as a competing bacterium. The contact-dependent or independent manner of killing the competing bacterium was demonstrated using this model. By deletion analysis and comparisons of the genomes of the various strains of B. subtilis, the SPβ prophage was first identified as a region containing an unknown toxin(s). Analysis of the extracellular proteome of B. subtilis subsequently revealed an unknown toxin (or toxin complex, respectively) of the molecular weight exceeding 100 kDa. Even more fascinating was the finding that such a large protein molecule is resistant to the pancreatic protease, trypsin. Subsequent non-enzymatic cyanogen bromide cleavage of the extracellular proteins and their analysis by mass spectrometry revealed...

Streptococcus pyogenes ist ein humanpathogenes Bakterium, welches verschiedene Gewebe besiedeln kann und dadurch unterschiedliche Krankheiten verursacht. Die enorme Anpassungsfähigkeit des Bakteriums beruht auf dessen Fähigkeit, verschiedene, vom Wirt induzierte Stresskonditionen zu ertragen. Genetische Faktoren, die in diesem Zusammenhang eine Rolle spielen, sind Toxin-Antitoxin (TA) Systeme. Typ II TA Systeme kodieren für zwei Proteine, ein Toxin und ein Antitoxin, die einen stabilen TA Komplex bilden. Verschlechtern sich die Wachstumsbedingungen, kann das Antitoxin proteolytisch abgebaut werden, wodurch das freigesetzte Toxin essentielle zelluläre Prozesse des Bakteriums inhibiert. In dieser Studie charakterisierte ich zwei chromosomal kodierte ParDE TA Systeme des pathogenen Bakteriums S. pyogenes. Ähnlich zu anderen Systemen werden das Toxin und das Antitoxin beider hier charakterisierten Systeme co-transkribiert und durch Stresseinwirkung (z.B. Aminosäure-mangel) induziert. Zudem konnten weitere posttranskriptionelle bzw. posttranslationale Mechanismen zur Regulierung der Genexpression beider Systeme nachgewiesen werden. Die extrachromosomale Expression der Toxine ParE1 und ParE2 führten in S. pyogenes und Escherichia coli zum Zelltod, wobei die Co-expression der entsprechenden Antitoxine ParD1 und ParD2 die Toxizität minderte. Allerdings verursachte die Überexpression der Antitoxine allein ebenfalls eine Inhibierung des Zellwachstums. ParD1 hemmte die Zellteilung in E. coli, wobei der N-Terminus des Proteins entscheidend für diesen Effekt zu sein schien. Zusammengefasst erweitern die Ergebnisse dieser Arbeit unser Verständnis von ParE Toxinen und verdeutlichen die diversen Mechanismen, welcher sich TA Systeme bedienen, um die bakterielle Physiologie zu beeinflussen. Zusätzlich gibt diese Arbeit einen Einblick in mögliche Mechanismen, die S. pyogenes implementiert, um Stresskonditionen im Wirt zu überdauern. / Streptococcus pyogenes is a human pathogen with a remarkable ability to colonize different tissues and to endure diverse host-induced stress conditions through mechanisms that have yet to be fully understood. One strategy employed by bacteria to cope with changing environments are toxin-antitoxin (TA) genetic modules. Under non-ideal conditions, the antitoxin is subject to proteolysis and thus the freed toxin protein can target crucial pathways in the cell modulating bacterial growth. This study, describes the characterization of two chromosomally encoded ParDE-like TA systems from the human pathogen S. pyogenes. The antitoxin-toxin genes of the parDEF1 and parDE2 TA systems are co-transcribed and triggered by stress-induced conditions. The parDE2 TA showed an inspected mRNA processing under amino acid starvation which suggest a putative post-transcriptional regulation. At the post-translational level, both systems are controlled by ClpXP antitoxin-protein degradation in vivo, an important factor for TA triggering. Furthermore, bacterial plasmid-based expression of the toxins ParE1 and ParE2 resulted in effects in cell viability while the antitoxin molecules ParD1 and ParD2 were able to prevent the toxins lethality, respectably. Unlike canonical antitoxins, both ParD1 and ParD2 molecules also displayed deleterious effects, which seemed to be exclusive and related with the N-terminus domain potentially involved in DNA-interaction. Finally, the ParE toxins presented remarkable plasticity, able to harm not only gyrase but also topoisomerase IV, two important bacterial drug targets that modulate DNA-topology. These results expand the view on the ParE molecular targets and highlight the diverse mechanisms TAs employ to modulate bacterial physiology. We also provide more insights into possible mechanisms that S. pyogenes employs to endure stress in the host and efficiently cause disease.

The aim of this study was to identify and characterise type II toxin-antitoxin (TA) systems in Burkholderia pseudomallei, the causative agent of the human disease melioidosis. 8 putative TA systems were identified within the genome of B. pseudomallei K96243. 5 of these were located witihn genome islands. Of the candidate toxins, BPSL0175 (RelE1) or BPSS1060 (RelE2) caused growth to cease when expressed in Escherichia coli, whereas expression of BPSS0390 (HicA) or BPSS1584 (HipA) (in an E. coli ΔhipBA background) caused a reduction in the number of culturable bacteria. HicA also caused growth arrest in B. pseudomallei K96243 ΔhicAB. These toxin induced phenotypes were enhanced by an <3kDa extracellular factor that accumulated in the spent medium during growth. Expression of the cognate antitoxins could restore growth and culturability of cells. Expression of hicA in E. coli gave an increased number of persister cells in response to ciprofloxacin or ceftazidime. Site directed mutagenesis studies identified two key residues within the HicA toxin that were essential for both the reduced culturability and increased persistence phenotypes. Deletion of hicAB from B. pseudomallei K96243 did not affect persister cell or survival frequencies compared to the wild type following treatment with a variety of stress conditions. Deletion of the ΔhipBA locus from B. pseudomallei K96243 also had no affect on bacterial persistence or survival under the conditions tested.

Thesis (DSc)--Stellenbosch University, 2014. / ENGLISH ABSTRACT: Plasmids belonging to the IncQ family have an exceptionally broad host-range and are highly mobilizable in the presence of the self-transmissible IncP plasmids. All IncQ plasmids identified to date have certain features in common. The feature that distinguishes them most from all other plasmids is that they have a unique mechanism of replication. Their replicons consist of repA, repB and repC genes encoding a replicase, primase and DNA-binding proteins respectively. All IncQ plasmids contain at least three 22-bp iterons (or 20-bp iterons with 2-bp spacers) that are identical in sequence and to which the RepC DNA-binding protein binds. They replicate by means of a unique strand-displacement mechanism that is considered to place a limit on their size. Replication proceeds by a partially single-stranded intermediate that is believed to result in an increased likelihood of structural instability with an increase in plasmid size. The most compact backbone of IncQ plasmids is approximately 5.9-kb and the largest natural IncQ plasmid reported is 14.2-kb. Although the mobilization regions of IncQ plasmids are not as unique as the replicons, they are all characterized by the primase of the replicon being fused to the relaxase of the mobilization genes. The remainder of the mobilization genes may vary substantially in number and sequence between plasmids and have been subdivided into at least four distinct lineages. This dissertation consists of twenty one manuscripts published during the period 1984 to 2012. The focus is almost entirely on the IncQ plasmid subfamily known as IncQ2. Most of the earlier work was on determining the nature and extent of the replicons, mobilization genes and the toxin-antitoxin plasmid stability system. A strong theme in the latter work focussed on using the natural variation among the IncQ2 plasmids as a means to understand IncQ plasmid evolution. The collection of articles comprises a combination of original research and reviews. / AFRIKAANSE OPSOMMING: Plasmiede wat aan die IncQ familie behoort kom ‘n uitsonderlike wye gasheerselreeks voor en is hoogs mobiliseerbaar deur middel van die selfoordraagbaar IncP plasmiede. Alle IncQ plasmiedes wat tot datum identifiseer is het sekere gemeenskaplike eienskappe. Die eienskap wat hulle van alle ander plasmiedes onderskei is hul unieke dupliseringsmeganisme. Hul dupliseringsmeganisme bestaan uit repA, repB en repC gene wat onderskeidlik ‘n helikase, ‘n ‘primase’ en ‘n DNSbindingsproteïen enkodeer. Die IncQ plasmiede het ten minste drie 22-bp iterone (of 20-bp iterone met 2-bp skeidingsnukleotiede) met ‘n identiese nukleotiedvolgorde en waaraan die RepCbindingsproteïen bind. Hulle dupliseer deur middel van ‘n unieke DNA-string-vervangingsmeganisme wat ‘n beperking op hul grootte plaas. Tydens replikasie word ‘n intermediêre struktuur gevorm wat gedeeltelik enkelstring is en dit is blykbaar die rede vir ‘n verhoging in strukturële onstabilitiet as die plasmied groter word. Die kleinste ruggraat onder die IncQ plasmiede is min of meer 5.9-kb en die grootste natuurlike IncQ plasmied wat gerapporteer is, is 14.2-kb. Alhoewel die mobiliseringsgebied van die IncQ plasmiede nie so duidelik uitkenbaar as die replikons nie, hierdie gebied is gekenmerk deur ‘n ‘primase’ wat aan ‘n ‘relaxase’ in die mobiliseringsgene gekoppel is. Die oorblywende mobiliseringsgene verskil in beide getal en nukleotiedvolgorde tussen plasmiede en is gebruik om die plasmiede in vier duidelike oorsponggroepe in te deel. Hierdie proefskrif bestaan uit een-en-twintig artikels wat tussen 1984 en 2012 gepubliseer is. Die fokus is hoofsaaklik op die IncQ plasmiedsubfamilie wat as IncQ2 bekend is. Baie van die vroeër werk het oor die aard en omvang van die duplisering en mobiliseringsgene asook die toksienteentoksien plasmiedstabiliseringsmeganisme hanteer. ‘n Sterk tema in die latere werk was om die natuurlike variasie onder die IncQ2 plasmiede te bestudeer ten einde IncQ plasmiedevolusie te verstaan. Die publikasie versameling bestaan uit ‘n kombinasie van oorspronklike navorsing en oorsigartikels.

Toksino-antitoksino (TA) sistemos – tai poros viename operone esančių bakterijų ir archėjų genų, kurių vienas koduoja toksišką baltymą, o antras – jį neutralizuojantį baltymą-antitoksiną. Tol, kol ląstelėje gaminamas pakankamas abiejų baltymų kiekis, antitoksinas jungiasi su toksinu ir jį išaktyvina. Tačiau, esant nepalankioms aplinkos sąlygoms, labilesnis antitoksinas suardomas aktyvintų proteazių, o likęs laisvas stabilesnis toksinas slopina gyvybiškai svarbius ląstelinius procesus – baltymų arba DNR biosintezę, dėl ko stabdomas ląstelių augimas arba jos žūva. Escherichia coli chromosomoje aprašyta daugiau nei dešimt TA sistemų, kurių viena yra dinJ-yafQ, apie kurią žinoma labai nedaug. Anksčiau laboratorijoje atliktuose darbuose nustatyta, kad dinJ-yafQ koduoja transliaciją slopinantį toksiną YafQ, o DinJ ir YafQ baltymai sudaro stiprų baltymų kompleksą, slopinantį YafQ toksišką poveikį. Kol kas nieko nėra žinoma apie YafQ molekulės sritis, svarbias sąveikai su antitoksinu DinJ. Šiame darbe sekai atrankios mutagenezės metodu buvo tirtos YafQ baltymo sritys, svarbios sąveikai su „savuoju“ toksinu DinJ. TA sistemoms būdinga savo operono transkripcijos autoreguliacija. DNR sulėtinimo gelyje eksperimentais parodėme atrankią DNR ir antitoksino DinJ bei DinJ-YafQ baltymų komplekso sąveiką. Laisvas antitoksinas DinJ silpniau sąveikauja su DNR nei būdamas komplekse su YafQ, o sąveikai su DNR svarbi DinJ baltymo N galinė dalis. Iš dviejų dinJ-yafQ operono promotoriaus srityje... [toliau žr. visą tekstą] / Prokaryotic toxin antitoxin systems consist of two adjacent genes, where one encodes a stable toxin harmful to essential cellular processes (translation or DNA synthesis), and the other a labile antitoxin, capable of blocking the toxin's activity by binding into stable protein complex. TA systems are proposed to be involved in bacterial adaptation to stress conditions by modulating the level of essential biological processes. There are at least ten characterized chromosome-encoded TA loci in Escherichia coli. The dinJ-yafQ operon codes for YafQ toxin which is neutralized by its cognate antitoxin, DinJ. YafQ is known to inhibit translation in vivo and belongs to the RelE toxin family of toxin ribonucleases. By using site-specific mutagenesis of YafQ, we have investigated the protein regions important for its interaction with DinJ antitoxin. Transcriptional autoregulation has been reported for members of all known TA gene families and appears to be general characteristic of regulation of TA loci. In this work electrophoretic mobility shift assay was used to investigate the interaction between the antitoxin DinJ and DinJ-YafQ complex and dinJ-yafQ operon promoter DNA. Antitoxin DinJ in the complex with YafQ had an enhanced DNA-binding affinity compared to free DinJ. N-terminal domain of antitoxin is crucial for interaction with DNA. Bioinformatic analysis of dinJ-yafQ operon promoter region revealed several palindromic DNA islands and their importance for interaction with DinJ... [to full text]

L'interaction symbiotique entre la bactérie du sol Sinorhizobium meliloti et la plante de la famille des légumineuses Medicago sp. conduit au développement d’un nouvel organe racinaire: la nodosité. Au sein de cet organe, les bactéries différenciées en bactéroïdes, réduisant l’azote atmosphérique en ammoniac directement assimilable par la plante, favorisant ainsi sa nutrition azotée. En échange, la plante, grâce à son activité photosynthétique, fournit aux bactéroïdes des composés carbonés. Cette association à bénéfice mutuel n’est toutefois pas permanente. En effet, quelques semaines seulement après l'établissement de la symbiose, une sénescence définie par une dégradation des bactéroïdes puis des cellules végétales, est observée. Cette étape du développement nodositaire est aujourd’hui encore peu étudiée et mal comprise.L’objectif premier de ce travail était donc d’analyser le rôle du bactéroïde dans cette rupture symbiotique. Pour cela, nous nous sommes plus particulièrement intéressés au rôle des systèmes Toxine Antitoxine (TA) de type VapBC de S. meliloti. En effet, ces opérons sont, dans la littérature, connus pour être impliqués dans la réponse aux stress, la persistance et/ou la mort bactérienne ainsi que la survie de la bactérie au sein de la cellule hôte. Dans un premier temps, nous avons développé une analyse globale du rôle des 11 systèmes VapBC chromosomiques de S. meliloti dans l’interaction symbiotique par des analyses in silico et de phénotypes de mutants d'invalidation du gène de la toxine en interactions avec Medicago sp. Deux études ont été réalisés de façon plus détaillées sur deux modules vapBC (VapBC5 et VapBC7). / The symbiotic interaction between the soil bacterium Sinorhizobium meliloti and the legumes plant Medicago sp. led to the development of a new root organ: the nodule. In this nodule differenciated bacteria into bacteroids, reducing atmospheric nitrogen into ammonia directly assimilated by the plant, thus promoting its nitrogen nutrition. In exchange, the plant, thanks to its photosynthetic activity, provides carbon compounds to the bacteroids. This mutual benefit association is however not permanent. Indeed, just weeks after the establishment of the symbiosis, senescence defined by a degradation of Bacteroides and plant cells, is observed. This stage of development is poorly understood in particularly about bacterial signal.The primary objective of this study was therefore to analyze the role of bacteroids in this symbiotic rupture. For this, we are particularly interested in the role of VapBC toxin antitoxin systems (TA) of S. meliloti. Indeed, in the literature, they are known to be involved in the stress response, persistence and / or bacterial death and the survival of the bacteria within the host cell. At first, we developed a global analysis of the role of 11 VapBC chromosomal systems in S. meliloti symbiotic interaction. After an in silico study, we studied the symbiotic phenotype with Medicago sp., Of each of the bacterial toxin mutants invalidation. Given the results, we, as a second step, developed a detail analysis of phenotypes obtained with two of these mutants: vapC5- and vapC7-.

L'îlot génomique Salmonella Genomic Island 1 (SGI1) est un élément intégratif et mobilisable, support de nombreux gènes de résistance aux antibiotiques, et identifié chez de nombreux genres bactériens. Le transfert de SGI1 requiert spécifiquement la présence d'un plasmide conjugatif du groupe d'incompatibilité IncA/C. Les régulateurs globaux AcaCD des plasmides IncA/C activent l’excision de SGI1 qui, une fois sous forme d’un intermédiaire extrachromosomique circulaire, va pouvoir être transféré en utilisant la machinerie de conjugaison encodée par les plasmides IncA/C (mobilisation conjugative en trans). Depuis la description de SGI1, plusieurs études ont relaté une apparente stabilité de SGI1 au cours des générations bactériennes. Cependant, des observations préliminaires indiquaient des difficultés de cohabitation entre SGI1 et les plasmides IncA/C. L’objectif de ce travail était d’étudier la stabilité de SGI1 et sa compatibilité avec les plasmides conjugatifs IncA/C dont dépend sa mobilité. L’opéron putatif S026- S025 de SGI1 a été identifié comme constituant un système Toxine-Antitoxine (TA) qui a été appelé sgiAT. Le rôle de ce système TA dans la stabilité de SGI1 a été mis en évidence en présence d'un plasmide IncA/C. De plus, l’incompatibilité entre SGI1 et les plasmides IncA/C a été démontrée expérimentalement pour la première fois. La stabilité de SGI1 est liée à son intégration chromosomique. Cependant, lorsque SGI1 est excisé du chromosome et donc vulnérable (il peut être perdu), c’est-à-dire en présence d’un plasmide IncA/C, le système TA sgiAT joue un rôle important dans le maintien de SGI1 dans les populations bactériennes. / The multidrug resistance Salmonella Genomic Island 1 (SGI1) is an integrative mobilizable element identified in several enterobacterial pathogens. This chromosomal island requires specifically the presence of a conjugative IncA/C plasmid to be excised and transfered by conjugation (mobilization in trans). Preliminary observations suggest stable maintenance of SGI1 in the bacterial host but paradoxically also incompatibility between SGI1 and IncA/C plasmids. Here, using a Salmonella enterica serovar Agona clonal bacterial population as model, we demonstrate that a Toxin-Antitoxin (TA) system encoded by SGI1 plays a critical role in its stable host maintenance when an IncA/C plasmid is concomitantly present. This system, designated sgiAT for Salmonella genomic island 1 Antitoxin and Toxin respectively, thus seems to play a stabilizing role in a situation where SGI1 is susceptible to be lost through plasmid IncA/C-mediated excision. Moreover and for the first time, the incompatibility between SGI1 and IncA/C plasmids was experimentally confirmed.