Temporal transcriptomic response during arsenic stress in Herminiimonas arsenicoxydans

Background: Arsenic is present in numerous ecosystems and microorganisms have developed various mechanisms to live in such hostile environments. Herminiimonas arsenicoxydans, a bacterium isolated from arsenic contaminated sludge, has acquired remarkable capabilities to cope with arsenic. In particular our previous studies have suggested the existence of a temporal induction of arsenite oxidase, a key enzyme in arsenic metabolism, in the presence of As(III). Results: Microarrays were designed to compare gene transcription profiles under a temporal As(III) exposure. Transcriptome kinetic analysis demonstrated the existence of two phases in arsenic response. The expression of approximatively 14% of the whole genome was significantly affected by an As(III) early stress and 4% by an As(III) late exposure. The early response was characterized by arsenic resistance, oxidative stress, chaperone synthesis and sulfur metabolism. The late response was characterized by arsenic metabolism and associated mechanisms such as phosphate transport and motility. The major metabolic changes were confirmed by chemical, transcriptional, physiological and biochemical experiments. These early and late responses were defined as general stress response and specific response to As(III), respectively. Conclusion: Gene expression patterns suggest that the exposure to As(III) induces an acute response to rapidly minimize the immediate effects of As(III). Upon a longer arsenic exposure, a broad metabolic response was induced. These data allowed to propose for the first time a kinetic model of the As(III) response in bacteria

Multiple controls affect arsenite oxidase gene expression in Herminiimonas arsenicoxydans

<p>Abstract</p> <p>Background</p> <p>Both the speciation and toxicity of arsenic are affected by bacterial transformations, i.e. oxidation, reduction or methylation. These transformations have a major impact on environmental contamination and more particularly on arsenic contamination of drinking water. <it>Herminiimonas arsenicoxydans </it>has been isolated from an arsenic- contaminated environment and has developed various mechanisms for coping with arsenic, including the oxidation of As(III) to As(V) as a detoxification mechanism.</p> <p>Results</p> <p>In the present study, a differential transcriptome analysis was used to identify genes, including arsenite oxidase encoding genes, involved in the response of <it>H. arsenicoxydans </it>to As(III). To get insight into the molecular mechanisms of this enzyme activity, a Tn<it>5 </it>transposon mutagenesis was performed. Transposon insertions resulting in a lack of arsenite oxidase activity disrupted <it>aoxR </it>and <it>aoxS </it>genes, showing that the <it>aox </it>operon transcription is regulated by the AoxRS two-component system. Remarkably, transposon insertions were also identified in <it>rpoN </it>coding for the alternative N sigma factor (σ⁵⁴) of RNA polymerase and in <it>dnaJ </it>coding for the Hsp70 co-chaperone. Western blotting with anti-AoxB antibodies and quantitative RT-PCR experiments allowed us to demonstrate that the <it>rpoN </it>and <it>dnaJ </it>gene products are involved in the control of arsenite oxidase gene expression. Finally, the transcriptional start site of the <it>aoxAB </it>operon was determined using rapid amplification of cDNA ends (RACE) and a putative -12/-24 σ⁵⁴-dependent promoter motif was identified upstream of <it>aoxAB </it>coding sequences.</p> <p>Conclusion</p> <p>These results reveal the existence of novel molecular regulatory processes governing arsenite oxidase expression in <it>H. arsenicoxydans</it>. These data are summarized in a model that functionally integrates arsenite oxidation in the adaptive response to As(III) in this microorganism.</p

Structure, Function, and Evolution of the Thiomonas spp. Genome

Bacteria of the Thiomonas genus are ubiquitous in extreme environments, such as arsenic-rich acid mine drainage (AMD). The genome of one of these strains, Thiomonas sp. 3As, was sequenced, annotated, and examined, revealing specific adaptations allowing this bacterium to survive and grow in its highly toxic environment. In order to explore genomic diversity as well as genetic evolution in Thiomonas spp., a comparative genomic hybridization (CGH) approach was used on eight different strains of the Thiomonas genus, including five strains of the same species. Our results suggest that the Thiomonas genome has evolved through the gain or loss of genomic islands and that this evolution is influenced by the specific environmental conditions in which the strains live

Bacterial response to arsenic stress

L'arsenic est un métalloïde largement distribué dans l'environnement. Sous l'action des micro-organismes, cet élément peut passer d'une forme oxydée à une forme réduite et inversement chez certaines bactéries. Herminiimonas arsenicoxydans est une bactérie hétérotrophe qui est capable de réduire l'arséniate (As(V) et d'oxyder l'arsénite (As(lll)). La réduction de l'As(V) a été largement étudiée ces dernières années, alors que l'oxydation de l'As(III) catalysée par l'arsénite oxydase (Aox) est un processus qui a fait l'objet de peu d'études. Ce travail de thèse a eu pour objectif principal l'étude de stress arsénié chez les bactéries, en particulier la régulation de l'arsénite oxydase. L'analyse de mutants obtenus par transposition aléatoire d'un mini-Tn5 a conduit à l'identification des gènes dont les produits jouent un rôle dans le contrôle de l'arsénite oxydase : aoxA, aoxB, aoxRS, rpoN et dnaJ. En parallèle la réponse globale au stress arsénié a été étudiée. Nous avons ainsi montrés que les gènes et protéines régulés par l'arsenic interviennent dans une grande variété de fonctions telles que le transport du phosphate, la motilité, la résistance à l'arsenic via ars, le stress oxydatif... Néanmoins, l'arsénite oxydase n'a pas été identifiée lors de ces travaux. Aussi, la cinétique de l'oxydation et les mécanismes de régulation sous-jacents ont été étudiées. Ces travaux ont ensuite été étendus à des bactéries chimiolithoautotrophes telles que Thiomonas sp. et Rhizobium sp. NT-26. L'As(III) peut servir de donneur d'électrons chez ces micro-organismes et son oxydation est aussi catalysée par l'arsénite oxydase. L'analyse de la réponse globale au stress arsénié semble indiquer que certains mécanismes de réponses sont conservés tels que la motilité, le transport de phosphate, la résistance à l'arsenic via ars. Néanmoins, des spécificités ont pu être observées telles que la biosynthèse d'acide gras, des transporteurs ABC... De la même manière, l'analyse des mutants a permis de valider le rôle de AroR (AoxR) dans la régulation de l'arsénite oxydase chez une bactérie chimiolithoautotrophe et d'autres régulateurs ont également été identifiés : PtxB et MoeB.Arsenic is a metalloid which is largely distributed in the environment. The implication of microorganisms in oxidoreduction reactions between the bioavailable As(III) and the less mobile As(V) oxidation states has been demonstrated. The heterotrophic bacteria, Herminiimonas arsenicoxydans, overcome the toxic effects of arsenic stress by either reducing arsenate (As(V)), or by oxidizing As(III). The reduction of As(V) has been largely studied, but the mechanism of oxidation of arsenite catalysed by arsenite oxidase is still unknown. In a first part, to identify genes possibly involved in the control of arsenite oxidation in H. arsenicoxydans, a library of 10,000 mutants was constructed by transposon mutagenesis. Transposon insertions resulting in a lack of arsenite oxidase activity disrupted aoxA, aoxB, aoxRS, rpoN and dnaJ genes. In parallel, a differential transcriptome combined to a proteome analysis was performed. This functional analysis indicated that H arsenicoxydans expressed genes and proteins required not only for arsenic detoxification or stress response but also involved in motility, exopolysaccharide synthesis, phosphate import or energetic metabolism However, no variation was found in the genes coding for arsenite oxidase. To address this process, physiological analyses coupled with Western immunoblotting experiments and DNA microarrays were used to examine the temporal changes in transcriptome profiles during the transition from As(III) to As(V) species due to As(III) oxidation. In a second part, the study was transposed to chemolithotrophic bacteria such as Thiomonas sp. and Rhizobium sp. NT-26. In these bacteria, the oxidation of As(III) is also catalysed by arsenite oxidase and As(III) can be utilized as electron donor. The analysis of the global response to arsenic stress, indicated that some mechanisms are conserved such as motility, phosphate import, arsenic resistance and other are specific such as fatty acids biosynthesis, ABC-type transporters Moreover, the involvement of AroR (AoxR) in the regulation of arsenite oxidase activity in a chemolitotrophic bacteria was validated in this work and others regulators were identified: PtxB and MoeB

Bacterial response to arsenic stress

L'arsenic est un métalloïde largement distribué dans l'environnement. Sous l'action des micro-organismes, cet élément peut passer d'une forme oxydée à une forme réduite et inversement chez certaines bactéries. Herminiimonas arsenicoxydans est une bactérieArsenic is a metalloid which is largely distributed in the environment. The implication of microorganisms in oxidoreduction reactions between the bioavailable As(III) and the less mobile As(V) oxidation states has been demonstrated. The heterotrophic bac

Enhanced structural and functional genome elucidation of the arsenite-oxidizing strain Herminiimonas arsenicoxydans by proteomics data.

International audienceThe arsenite-oxidizing strain Herminiimonas arsenicoxydans proteome was investigated with gel electrophoresis and tandem mass spectrometry analyses. The comparison of experimental and theoretical M(r) and pI, as well as that of peptide sequences identified by MS and predicted protein sequences, allowed the correction of five protein annotations. More importantly, the functional analysis of SDS- and 2D-PAGE proteome maps obtained in the presence of arsenic, combined with partial transcriptomic results indicate that H. arsenicoxydans expressed genes and proteins required not only for arsenic detoxification or stress response but also involved in motility, exopolysaccharide synthesis, phosphate import or energetic metabolism. This study provides therefore new insights into the adaptation processes of H. arsenicoxydans in response to arsenic