Characterization of the CopR regulon of Lactococcus lactis IL1403.
To identify components of the copper homeostatic mechanism of Lactococcus lactis, we employed two-dimensional gel electrophoresis to detect changes in the proteome in response to copper. Three proteins upregulated by copper were identified: glyoxylase I (YaiA), a nitroreductase (YtjD), and lactate oxidase (LctO). The promoter regions of these genes feature cop boxes of consensus TACAnnTGTA, which are the binding site of CopY-type copper-responsive repressors. A genome-wide search for cop boxes revealed 28 such sequence motifs. They were tested by electrophoretic mobility shift assays for the interaction with purified CopR, the CopY-type repressor of L. lactis. Seven of the cop boxes interacted with CopR in a copper-sensitive manner. They were present in the promoter region of five genes, lctO, ytjD, copB, ydiD, and yahC; and two polycistronic operons, yahCD-yaiAB and copRZA. Induction of these genes by copper was confirmed by real-time quantitative PCR. The copRZA operon encodes the CopR repressor of the regulon; a copper chaperone, CopZ; and a putative copper ATPase, CopA. When expressed in Escherichia coli, the copRZA operon conferred copper resistance, suggesting that it functions in copper export from the cytoplasm. Other member genes of the CopR regulon may similarly be involved in copper metabolism.
Magnani
David
D
Department of Clinical Pharmacology, University of Berne, 3010 Berne, Switzerland.
Barré
Olivier
O
Gerber
Simon D
SD
Solioz
Marc
M
D001426:Bacterial Proteins / Q000096:biosynthesis / Q000235:genetics / Q000302:isolation & purification / Q000502:physiology*; D001665:Binding Sites; D016384:Consensus Sequence; D003300:Copper / Q000378:metabolism* / Q000633:toxicity; D004269:DNA, Bacterial / Q000378:metabolism; D004268:DNA-Binding Proteins / Q000235:genetics / Q000302:isolation & purification / Q000502:physiology*; D024881:Drug Resistance, Bacterial; D015180:Electrophoresis, Gel, Two-Dimensional; D024202:Electrophoretic Mobility Shift Assay; D004926:Escherichia coli / Q000187:drug effects / Q000235:genetics; D020869:Gene Expression Profiling; D015964:Gene Expression Regulation, Bacterial / Q000502:physiology*; D013294:Lactococcus lactis / Q000235:genetics / Q000502:physiology*; D007791:Lactoylglutathione Lyase / Q000096:biosynthesis; D006899:Mixed Function Oxygenases / Q000032:analysis; D009601:Nitroreductases / Q000096:biosynthesis; D011401:Promoter Regions, Genetic; D011485:Protein Binding; D020543:Proteome / Q000032:analysis; D018085:Regulon; D015534:Trans-Activators / Q000235:genetics / Q000302:isolation & purification / Q000502:physiology*
D001426:Bacterial Proteins; D004269:DNA, Bacterial; D004268:DNA-Binding Proteins; D020543:Proteome; D015534:Trans-Activators; D003300:Copper; D006899:Mixed Function Oxygenases; C021497:lactate 2-monooxygenase; D009601:Nitroreductases; D007791:Lactoylglutathione Lyase