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A two-component regulatory system required for copper-inducible expression of the copper resistance operon of Pseudomonas syringae.

ABSTRACT: Specific induction of the copper resistance operon (cop) promoter from Pseudomonas syringae was measured by beta-galactosidase production from a cop promoter-lacZ fusion. Induction of the cop promoter in P. syringae pv. syringae required trans-acting factors from copper resistance plasmid pPT23D, from which cop was originally cloned. Tn5 mutagenesis of pPT23D was used to localize two complementation groups immediately downstream from copABCD. Cloning and sequencing of the DNA in this region revealed two genes, copR and copS, expressed in the same orientation as the cop operon but from a separate constitutive promoter. The amino acid sequence deduced from these genes showed distinct similarities to known two-component regulatory systems, including PhoB-PhoR and OmpR-EnvZ. In addition, CopR showed strong similarity to copper resistance activator protein PcoR from Escherichia coli. Functional chromosomal homologs to copRS activated the cop promoter, in a copper-inducible manner, in copper-resistant or -sensitive strains of P. syringae pv. tomato and other Pseudomonas species. This implies that copper-inducible gene regulation is associated with a common chromosomally encoded function, as well as plasmid-borne copper resistance, in Pseudomonas spp.

SUBMITTER: Mills SD

PROVIDER: S-EPMC203959 | biostudies-other | 1993 Mar

REPOSITORIES: biostudies-other

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A two-component regulatory system required for copper-inducible expression of the copper resistance operon of Pseudomonas syringae.

Mills S D SD Jasalavich C A CA Cooksey D A DA

Journal of bacteriology 19930301 6

Specific induction of the copper resistance operon (cop) promoter from Pseudomonas syringae was measured by beta-galactosidase production from a cop promoter-lacZ fusion. Induction of the cop promoter in P. syringae pv. syringae required trans-acting factors from copper resistance plasmid pPT23D, from which cop was originally cloned. Tn5 mutagenesis of pPT23D was used to localize two complementation groups immediately downstream from copABCD. Cloning and sequencing of the DNA in this region reve ...[more]

PMID: 8449873

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Project description:Copper resistance mechanisms provide an important adaptive advantage to plant pathogenic bacteria under exposure to copper treatments. Copper resistance determinants have been described in Pseudomonas syringae pv. syringae (Pss) strains isolated from mango intimately associated with 62 kb plasmids belonging to the pPT23A family (PFP). It has been previously described that the indiscriminate use of copper-based compounds promotes the selection of copper resistant bacterial strains and constitutes a selective pressure in the evolution of copper resistance determinants. Hence, we have explored in this study the copper resistance evolution and the distribution of specific genetic determinants in two different Pss mango populations isolated from the same geographical regions, mainly from southern Spain with an average of 20 years of difference. The total content of plasmids, in particular the 62 kb plasmids, and the number of copper resistant Pss strains were maintained at similar levels over the time. Interestingly, the phylogenetic analysis indicated the presence of a phylogenetic subgroup (PSG) in the Pss mango phylotype, mostly composed of the recent Pss population analyzed in this study that was strongly associated with a hyper-resistant phenotype to copper. Genome sequencing of two selected Pss strains from this PSG revealed the presence of a large Tn7-like transposon of chromosomal location, which harbored putative copper and arsenic resistance genes (COARS Tn7-like). Transformation of the copper sensitive Pss UMAF0158 strain with some putative copper resistance genes and RT-qPCR experiments brought into light the role of COARS Tn7-like transposon in the hyper-resistant phenotype to copper in Pss.IMPORTANCECopper compounds have traditionally been used as standard bactericides in agriculture in the past few decades. However, the extensive use of copper has fostered the evolution of bacterial copper resistance mechanisms. Pseudomonas syringae is a plant pathogenic bacterium used worldwide as a model to study plant-pathogen interactions. The adaption of P. syringae to plant surface environment is the most important step prior to an infection. In this scenario, copper resistance mechanisms could play a key role in improving its epiphytic survival. In this work, a novel Tn7-like transposon of chromosomal location was detected in P. syringae pv. syringae strains isolated from mango. This transposon conferred the highest resistance to copper sulfate described to date for this bacterial phytopathogen. Understanding in depth the copper resistance mechanisms and their evolution are important steps to the agricultural industry to get a better improvement of disease management strategies.

Project description:CvsSR is a Ca2+-induced two-component system (TCS) in the plant pathogen Pseudomonas syringae pv. tomato DC3000. Here, we discovered that CvsSR is induced by Fe3+, Zn2+, and Cd2+ However, only supplementation of Ca2+ to medium resulted in rugose, opaque colonies in ΔcvsS and ΔcvsR strains. This phenotype corresponded to formation of calcium phosphate precipitation on the surface of ΔcvsS and ΔcvsR colonies. CvsSR regulated swarming motility in P. syringae pv. tomato in a Ca2+-dependent manner, but swarming behavior was not influenced by Fe3+, Zn2+, or Cd2+ We hypothesized that reduced swarming displayed by ΔcvsS and ΔcvsR strains was due to precipitation of calcium phosphate on the surface of ΔcvsS and ΔcvsR cells grown on agar medium supplemented with Ca2+ By reducing the initial pH or adding glucose to the medium, calcium precipitation was inhibited, and swarming was restored to ΔcvsS and ΔcvsR strains, suggesting that calcium precipitation influences swarming ability. Constitutive expression of a CvsSR-regulated carbonic anhydrase and a CvsSR-regulated putative sulfate major facilitator superfamily transporter in ΔcvsS and ΔcvsR strains inhibited formation of calcium precipitates and restored the ability of ΔcvsS and ΔcvsR bacteria to swarm. Lastly, we found that glucose inhibited Ca2+-based induction of CvsSR. Hence, CvsSR is a key regulator that controls calcium precipitation on the surface of bacterial cells.IMPORTANCE Bacteria are capable of precipitating and dissolving minerals. We previously reported the characterization of the two-component system CvsSR in the plant-pathogenic bacterium Pseudomonas syringae CvsSR responds to the presence of calcium and is important for causing disease. Here, we show that CvsSR controls the ability of the bacterium to prevent calcium phosphate precipitation on the surface of cells. We also identified a carbonic anhydrase and transporter that modulate formation of surface-associated calcium precipitates. Furthermore, our results demonstrate that the ability of the bacterium to swarm is controlled by the formation and dissolution of calcium precipitates on the surface of cells. Our study describes new mechanisms for microbially induced mineralization and provides insights into the role of mineral deposits on bacterial physiology. The discoveries may lead to new technological and environmental applications.

Dataset Information

A two-component regulatory system required for copper-inducible expression of the copper resistance operon of Pseudomonas syringae.

Publications

A two-component regulatory system required for copper-inducible expression of the copper resistance operon of Pseudomonas syringae.

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