Regulation of yodA encoding a novel cadmium-induced protein in Escherichia coli

Institute of Molecular Biology of the Slovak Academy of Sciences, Dúbravská cesta 21, SK-84251 Bratislava, Slovak Republic 1 Department of Cell and Molecular Biology, Göteborg University, Medicinaregatan 9C, SE-41390 Göteborg, Sweden 2 Author for correspondence: P. Ferianc. Tel: +421 2 5930 7427. Fax: +421 2 5930 7416. e-mail: umikferi{at}savba.sk Bacterial accommodation to moderate concentrations of cadmium is accompanied by transient activation of general stress proteins as well as a sustained induction of other proteins of hitherto unknown functions. One of the latter proteins was previously identified as the product of the Escherichia coli yodA ORF. The yodA ORF encodes 216 aa residues (the YodA protein) and the increased synthesis of YodA during cadmium stress was found probably to be a result of transcriptional activation from one single promoter upstream of the structural yodA gene. Analysis of a transcriptional gene fusion, P yodA – lac Z, demonstrated that basal expression of yodA is low during exponential growth and expression is increased greater than 50-fold by addition of cadmium to growing cells. However, challenging cells with additional metals such as zinc, copper, cobalt and nickel did not increase the level of yodA expression. In addition, hydrogen peroxide also increased yodA expression whereas the superoxide-generating agent paraquat failed to do so. Surprisingly, cadmium-induced transcription of yodA is dependent on soxS and fur , but independent of oxyR . Moreover, a double relA spoT mutation abolished induction of yodA during cadmium exposure but ppGpp is not sufficient to induce yodA since expression of the gene is not elevated during stationary phase. After 45 min of cadmium exposure the YodA protein was primarily detected in the cytoplasmic fraction but was later (150 min) found in both the cytoplasmic and periplasmic compartments. Keywords: cadmium stress, yod A characterization, YodA protein, RNA polymerase, S1 nuclease mapping Abbreviations: CDP, cadmium induced protein(s); DIG, digoxigenin; MBP, maltose-binding protein; ppGpp, guanosine 3',5'-bispyrophosphate; RAS, rabbit antiserum; tsp, transcription start point The GenBank accession number for the E. coli yod A gene and the SWISS-PROT accession number for E. coli YodA protein in this paper are AAC75039 and P76344, respectively.