Enhanced Cd²⁺-selective root-tonoplast-transport in tobaccos expressing Arabidopsis cation exchangers

Several Arabidopsis CAtion eXchangers (CAXs) encode tonoplast-localized transporters that appear to be major contributors to vacuolar accumulation/sequestration of cadmium (Cd²⁺), an undesirable pollutant ion that occurs in man largely as a result of dietary consumption of aerial tissues of food plants. But, ion-selectivity of individual CAX transporter types remains largely unknown. Here, we transformed Nicotiana tabacum with several CAX genes driven by the Cauliflower Mosaic Virus (CaMV) 35S promoter and monitored divalent cation transport in root-tonoplast vesicles from these plants in order to select particular CAX genes directing high Cd²⁺ antiporter activity in root tonoplast. Comparison of seven different CAX genes indicated that all transported Cd²⁺, Ca²⁺, Zn²⁺, and Mn²⁺ to varying degrees, but that CAX4 and CAX2 had high Cd²⁺ transport and selectivity in tonoplast vesicles. CAX4 driven by the CaMV 35S and FS3 [figwort mosaic virus (FMV)] promoters increased the magnitude and initial rate of Cd²⁺/H⁺ exchange in root-tonoplast vesicles. Ion selectivity of transport in root-tonoplast vesicles isolated from FS3::CAX4-expressing plant lines having a range of gene expression was Cd²⁺>Zn²⁺»Ca²⁺»Mn²⁺ and the ratios of maximal Cd²⁺ (and Zn²⁺) versus maximal Ca²⁺ and Mn²⁺ transport were correlated with the levels of CAX4 expression. Root Cd accumulation in high CAX4 and CAX2 expressing lines was increased in seedlings grown with 0.02 μM Cd. These observations are consistent with a model in which expression of an Arabidopsis-gene-encoded, Cd²⁺-efficient antiporter in host plant roots results in greater root vacuole Cd²⁺ transport activity, increased root Cd accumulation, and a shift in overall root tonoplast ion transport selectivity towards higher Cd²⁺ selectivity. Results support a model in which certain CAX antiporters are somewhat more selective for particular divalent cations.