Abstract
Enterococcus hirae possesses two P-type ATPases, CopA and CopB, that are involved in copper homeostasis. These enzymes are induced by extracellular copper concentrations that are either too low or too high for optimal growth. To identify the regulatory proteins involved in induction, the DNA upstream of copA was cloned and sequenced. Following a putative promoter region, it contains two genes, copY and copZ, that encode proteins of 145 and 69 amino acids, respectively. Both proteins contain metal binding motifs and exhibit significant sequence similarity to known regulatory proteins. Gene disruption of copY by reverse genetics caused constitutive overexpression of CopA and CopB, generating a copper-dependent phenotype. In contrast, disruption of copZ suppressed the expression of the two copper-ATPases, rendering the cells copper-sensitive. Both null mutations could be complemented in trans with plasmids bearing copY or copZ. Thus, copY and copZ encode trans-acting metalloregulatory proteins that are required for induction of the cop operon by copper. In this mechanism, CopY apparently acts as a metal-fist type repressor and CopZ as an activator.
Highlights
Escherichia coli, and another gene, synA, from Synechococcus (GenBankTMIEMBL Data Bank accession numbers X15079, L36317, Z36164, L33259, U00039, and U04356, respectively)
The activity of copper-ATPases and associated metal ion binding proteins involved in copper homeostasis must be appropriately controlled in response to the copper concentration; tightly regulated systems are required
The two copperATPases of E. hirae were shown to be regulated in their expression by the extracellular copper concentration: a low level of expression was observed in 10 J-LM copper, and strong induction resulted from either limiting or near toxic levels of copper in the media [11]
Summary
Vol 270, No., Issue of March 3, pp. 4349-4354, 1995 Printed in U.S.A. Two trans-Acting Metalloregulatory Proteins Controlling Expression of the Copper-ATPases of Enterococcus hirae*. CopY and copZ encode trans-acting metalloregulatory proteins that are required for induction of the cop operon by copper In this mechanism, CopY apparently acts as a metal-fist type repressor and CopZ as an activator. At toxic metal ion concentrations, the ACEI protein forms a complex with Cu+ or, with lower stability, Ag+, which binds to a cis-acting copper control sequence upstream of the transcription initiation site of CUPI to induce transcription of the metallothionein gene [15, 16]. We describe two new E. hirae genes, copY and copZ, that are located upstream of the copAB region that encodes the two copper-ATPases They control the expression of the two ATPases, by CopY acting as a repressor and CopZ as an activator. CopY represents a novel, bacterial metal-fist type transcription factor
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