| Summary: | <i>Cupriavidus metallidurans</i> CH34 is a well-studied metal-resistant β-proteobacterium and contains a battery of genes participating in metal metabolism and resistance. Here, we generated a mutant (CH34<sup>ZnR</sup>) adapted to high zinc concentrations in order to study how CH34 could adaptively further increase its resistance against this metal. Characterization of CH34<sup>ZnR</sup> revealed that it was also more resistant to cadmium, and that it incurred seven insertion sequence-mediated mutations. Among these, an IS<i>1088</i> disruption of the <i>glpR</i> gene (encoding a DeoR-type transcriptional repressor) resulted in the constitutive expression of the neighboring ATP-binding cassette (ABC)-type transporter. GlpR and the adjacent ABC transporter are highly similar to the glycerol operon regulator and ATP-driven glycerol importer of <i>Rhizobium leguminosarum</i> bv. <i>viciae</i> VF39, respectively. Deletion of <i>glpR</i> or the ABC transporter and complementation of CH34<sup>ZnR</sup> with the parental <i>glpR</i> gene further demonstrated that loss of GlpR function and concomitant derepression of the adjacent ABC transporter is pivotal for the observed resistance phenotype. Importantly, addition of glycerol, presumably by glycerol-mediated attenuation of GlpR activity, also promoted increased zinc and cadmium resistance in the parental CH34 strain. Upregulation of this ABC-type transporter is therefore proposed as a new adaptation route towards metal resistance.
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