Impact of the Mg2+-citrate transporter CitM on heavy metal toxicity in Bacillus subtilis

Bacillus subtilis possesses a secondary transporter, CitM, that is specific for the complex of citrate and Mg2+ but is also capable of transporting citrate in complex with the heavy metal ions Zn2+, Ni2+ and Co2+. We report on the impact of CitM activity on the toxicity of Zn2+, Ni2+ and Co2+ in B. subtilis. In a citM deletion mutant or under conditions in which CitM is not expressed, the toxic effects of the metals were reduced by the presence of citrate in the medium. In contrast, the presence of citrate dramatically enhanced toxicity when the Mg2+-citrate transporter was present in the membrane. It is demonstrated that the complex of Ni2+ and citrate is transported into the cell and that the uptake is responsible for the enhanced toxicity. At toxic concentrations of the metal ions, the cultures adapted by developing tolerance against these ions. Tolerant cells isolated by exposure to one of the metal ions remained tolerant after growth in the absence of toxic metal ions and were cross-tolerant against the other two toxic ions. Tolerant strains were shown to contain point mutations in the citM gene, which resulted in premature termination of translation.