Project description:Here, we described a novel transcriptional regulator belonging to the MarR family that we named OsbR (oxidative stress response and biofilm formation regulator) in the opportunistic pathogen Chromobacterium violaceum. Transcriptome profiling by DNA microarray using strains with deletion or overexpression of osbR showed that OsbR exert a global regulatory role in C. violaceum, regulating genes involved in oxidative stress response, nitrate reduction, biofilm formation, and several metabolic pathways. EMSA assays showed that OsbR binds to the promoter regions of several OsbR-regulated genes and the in vitro DNA binding activity was inhibited by oxidants. We demonstrated that the overexpression of osbR caused activation of ohrA even in the presence of the repressor OhrR, which resulted in improved growth under organic hydroperoxide treatment. We showed that the proper regulation of the nar genes by OsbR ensures an optimal growth of C. violaceum under anaerobic conditions by tuning the reduction of nitrate to nitrite. Finally, the osbR overexpressing strain showed reduction in biofilm formation and this phenotype correlated with the OsbR-mediated repression of two gene clusters encoding putative adhesins.
Project description:The study investigated the ability of selected (hyper-)thermophilic prokaryotes to grow anaerobically by the reduction of perchlorate and chlorate. Physiological, genomic and proteome analyses suggest that the Crenarchaeon Aeropyrum pernix reduces (per)chlorate with a periplasmic enzyme related to nitrate reductases, while it lacks a functional chlorite-disproportionating enzyme (Cld). A. pernix seems to rely on the chemical reactivity of reduced sulfur compounds with the toxic intermediate chlorite to complete the pathway. The chemical oxidation of thiosulfate (in excessive amounts present in the medium) to sulfate and the concomitant release of chloride anions from the reduction of chlorite are the products of a biotic-abiotic (per)chlorate reduction pathway in A. pernix. The apparent absence of Cld in two other (per)chlorate-reducing microorganisms and their dependence on sulfide for (per)chlorate reduction is consistent with earlier-made observations on (per)chlorate-reducing Archaeoglobus fulgidus. All here discussed microorganisms use strategies for complete (per)chlorate reduction that differ from the physiology of classical (per)chlorate-reducing mesophiles.
Project description:Shewanella spp. possess a broad respiratory versatility, which contributes to the occupation of hypoxic/anoxic environmental or host-associated niches. Here we observed a strain-specific induction of biofilm formation in response to supplementation with the anaerobic electron acceptors dimethyl sulfoxide (DMSO) and nitrate in a panel of Shewanella algae isolates. The respiration-driven biofilm response is not observed in DMSO and nitrate reductase deletion mutants of the type strain S. algae CECT 5071, and can be restored upon complementation with the corresponding reductase operon(s) but not by an operon containing a catalytically inactive nitrate reductase. The distinct transcriptional changes, proportional to the effect of these compounds on biofilm formation, include cyclic di-GMP (c-di-GMP) turnover genes. In support, ectopic expression of the c-di-GMP phosphodiesterase YhjH of Salmonella Typhimurium but not its catalytically inactive variant decreased biofilm formation. The respiration-dependent biofilm response of S. algae may permit differential colonization of environmental or host niches.
Project description:The conversion of nitrate to ammonium, known as nitrate reduction, consumes large amounts of reductants in plants. Previous studies have observed that mitochondrial alternative oxidase (AOX) is upregulated under conditions of limited nitrate reduction, such as low or no nitrate availability, or when ammonium serves as the sole nitrogen (N) source. Electron transfer from ubiquinone to AOX bypasses the proton-pumping complexes III and IV, thereby consuming reductants efficiently. Therefore, the upregulation of AOX under conditions of limited nitrate reduction may help dissipate excessive reductants and mitigate oxidative stress. However, firm evidence supporting this hypothesis is lacking due to the absence of experimental systems capable of directly analyzing the relationship between nitrate reduction and AOX. To address this gap, we developed a novel culturing system that allows for the manipulation of nitrate reduction and AOX activities separately, without inducing N starvation, ammonium toxicity, or disrupting the nitrate signal. Using this system, we investigated genome-wide gene expression with RNA-seq to gain insight into the relationship between AOX and nitrate reduction.
Project description:Geobacter sulfurreducens is a widely explored microorganism recognized by its metabolic versatility able to reduce a number of external electron acceptors. In the present study the capacity of this strain to reduce nitrate was evaluated along with its transcriptomic profile under nitrate-reducing conditions and the catalytic role of Pd nanoparticles on the reductive pathway. Results demonstrated that G. sulfurreducens was able to reduce nitrate and important kinetic differences related to the time response were found among the electron donors used (acetate and hydrogen). When using acetate, a delay response on nitrate reduction of 4 days and reduction of 94% of nitrate was achieved, while nitrite was not detected, and all the nitrogen was recovered as ammonium (79.6 ± 5.7 %). The use of hydrogen as electron donor increased 2-fold the maximum rate of nitrate reduction, leading to 93% reduction of nitrate during the first 20 h with recovery of 45% as ammonium, while nitrite was not detected. In addition, transcriptome profiling analysis of G. sulfurreducens under nitrate-reducing conditions using hydrogen or acetate as an electron donor at 2 and 6 days reveals that a core of 146 genes (69 upregulated and 77 downregulated) are differentially expressed in all conditions. Genes related to nitrogen metabolism, such as nrfA and nrfH, gdhA, and amtB, were upregulated in the incubations and RT-qPCR data confirmed upregulations of these genes. Experiments performed with biologically synthesized Pd (Bio-Pd) + G. sulfurreducens cells demonstrated synergistic input of Bio-Pd and the metabolic capacity of G. sulfurreducens. These results expand the metabolic versatility of G. sulfurreducens, which may have important implications in nitrogen cycling in natural environments and engineered systems.
Project description:The Pae PA1006/nbvF gene was found to be essential for nitrate utilization, biofilm maturation, and virulence. Microarrays were employed to assess global gene expression changes in response to NO3 compared to wild-type.