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The two-component signal transduction system CopRS of Corynebacterium glutamicum is required for adaptation to copper-excess stress.

Schelder S, Zaade D, Litsanov B, Bott M, Brocker M - PLoS ONE (2011)

Bottom Line: Using comparative transcriptome analysis of the ΔcopRS mutant and the wild type in combination with electrophoretic mobility shift assays and reporter gene studies the CopR regulon and the DNA-binding motif of CopR were identified.Evidence was obtained that CopR binds only to the intergenic region between cg3285 (copR) and cg3286 in the genome of C. glutamicum and activates expression of the divergently oriented gene clusters cg3285-cg3281 and cg3286-cg3289.Altogether, our data suggest that CopRS is the key regulatory system in C. glutamicum for the extracytoplasmic sensing of elevated copper ion concentrations and for induction of a set of genes capable of diminishing copper stress.

View Article: PubMed Central - PubMed

Affiliation: Institut für Bio-und Geowissenschaften, IBG-1: Biotechnologie, Forschungszentrum Jülich, Jülich, Germany.

ABSTRACT
Copper is an essential cofactor for many enzymes but at high concentrations it is toxic for the cell. Copper ion concentrations ≥50 µM inhibited growth of Corynebacterium glutamicum. The transcriptional response to 20 µM Cu(2+) was studied using DNA microarrays and revealed 20 genes that showed a ≥ 3-fold increased mRNA level, including cg3281-cg3289. Several genes in this genomic region code for proteins presumably involved in the adaption to copper-induced stress, e. g. a multicopper oxidase (CopO) and a copper-transport ATPase (CopB). In addition, this region includes the copRS genes (previously named cgtRS9) which encode a two-component signal transduction system composed of the histidine kinase CopS and the response regulator CopR. Deletion of the copRS genes increased the sensitivity of C. glutamicum towards copper ions, but not to other heavy metal ions. Using comparative transcriptome analysis of the ΔcopRS mutant and the wild type in combination with electrophoretic mobility shift assays and reporter gene studies the CopR regulon and the DNA-binding motif of CopR were identified. Evidence was obtained that CopR binds only to the intergenic region between cg3285 (copR) and cg3286 in the genome of C. glutamicum and activates expression of the divergently oriented gene clusters cg3285-cg3281 and cg3286-cg3289. Altogether, our data suggest that CopRS is the key regulatory system in C. glutamicum for the extracytoplasmic sensing of elevated copper ion concentrations and for induction of a set of genes capable of diminishing copper stress.

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Influence of increasing copper ion concentrations on growth of C. glutamicum wild type (▪) and C. glutamicum ΔcopRS (•).For experimental details see legend to Fig. 1. The time points of CuSO4 addition are indicated by arrows.
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pone-0022143-g003: Influence of increasing copper ion concentrations on growth of C. glutamicum wild type (▪) and C. glutamicum ΔcopRS (•).For experimental details see legend to Fig. 1. The time points of CuSO4 addition are indicated by arrows.

Mentions: In order to verify the involvement of the CopRS two-component system in copper homoeostasis, the resistance of a copRS deletion mutant [21] to copper ions and other heavy metal salts was compared to the wild type using agar diffusion assays. An increased susceptibility was only observed for copper ions (Fig. 2A) but not for all other heavy metal ions tested (nickel, manganese, zinc, silver, cobalt, lead or cadmium; data not shown). Cultivation of ΔcopRS and C. glutamicum wild type in CGXII minimal medium to which different CuSO4 concentrations (5–500 µM) were added when the cultures had reached an OD600 of 5–6 corroborated the reduced resistance to copper ions of the mutant strain (Fig. 3). The phenotype could be complemented by plasmid-borne copRS expression (Fig. 2B). These results confirmed the assumption that CopRS is involved in copper homeostasis of C. glutamicum.


The two-component signal transduction system CopRS of Corynebacterium glutamicum is required for adaptation to copper-excess stress.

Schelder S, Zaade D, Litsanov B, Bott M, Brocker M - PLoS ONE (2011)

Influence of increasing copper ion concentrations on growth of C. glutamicum wild type (▪) and C. glutamicum ΔcopRS (•).For experimental details see legend to Fig. 1. The time points of CuSO4 addition are indicated by arrows.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC3140484&req=5

pone-0022143-g003: Influence of increasing copper ion concentrations on growth of C. glutamicum wild type (▪) and C. glutamicum ΔcopRS (•).For experimental details see legend to Fig. 1. The time points of CuSO4 addition are indicated by arrows.
Mentions: In order to verify the involvement of the CopRS two-component system in copper homoeostasis, the resistance of a copRS deletion mutant [21] to copper ions and other heavy metal salts was compared to the wild type using agar diffusion assays. An increased susceptibility was only observed for copper ions (Fig. 2A) but not for all other heavy metal ions tested (nickel, manganese, zinc, silver, cobalt, lead or cadmium; data not shown). Cultivation of ΔcopRS and C. glutamicum wild type in CGXII minimal medium to which different CuSO4 concentrations (5–500 µM) were added when the cultures had reached an OD600 of 5–6 corroborated the reduced resistance to copper ions of the mutant strain (Fig. 3). The phenotype could be complemented by plasmid-borne copRS expression (Fig. 2B). These results confirmed the assumption that CopRS is involved in copper homeostasis of C. glutamicum.

Bottom Line: Using comparative transcriptome analysis of the ΔcopRS mutant and the wild type in combination with electrophoretic mobility shift assays and reporter gene studies the CopR regulon and the DNA-binding motif of CopR were identified.Evidence was obtained that CopR binds only to the intergenic region between cg3285 (copR) and cg3286 in the genome of C. glutamicum and activates expression of the divergently oriented gene clusters cg3285-cg3281 and cg3286-cg3289.Altogether, our data suggest that CopRS is the key regulatory system in C. glutamicum for the extracytoplasmic sensing of elevated copper ion concentrations and for induction of a set of genes capable of diminishing copper stress.

View Article: PubMed Central - PubMed

Affiliation: Institut für Bio-und Geowissenschaften, IBG-1: Biotechnologie, Forschungszentrum Jülich, Jülich, Germany.

ABSTRACT
Copper is an essential cofactor for many enzymes but at high concentrations it is toxic for the cell. Copper ion concentrations ≥50 µM inhibited growth of Corynebacterium glutamicum. The transcriptional response to 20 µM Cu(2+) was studied using DNA microarrays and revealed 20 genes that showed a ≥ 3-fold increased mRNA level, including cg3281-cg3289. Several genes in this genomic region code for proteins presumably involved in the adaption to copper-induced stress, e. g. a multicopper oxidase (CopO) and a copper-transport ATPase (CopB). In addition, this region includes the copRS genes (previously named cgtRS9) which encode a two-component signal transduction system composed of the histidine kinase CopS and the response regulator CopR. Deletion of the copRS genes increased the sensitivity of C. glutamicum towards copper ions, but not to other heavy metal ions. Using comparative transcriptome analysis of the ΔcopRS mutant and the wild type in combination with electrophoretic mobility shift assays and reporter gene studies the CopR regulon and the DNA-binding motif of CopR were identified. Evidence was obtained that CopR binds only to the intergenic region between cg3285 (copR) and cg3286 in the genome of C. glutamicum and activates expression of the divergently oriented gene clusters cg3285-cg3281 and cg3286-cg3289. Altogether, our data suggest that CopRS is the key regulatory system in C. glutamicum for the extracytoplasmic sensing of elevated copper ion concentrations and for induction of a set of genes capable of diminishing copper stress.

Show MeSH
Related in: MedlinePlus