Cloning and functional characterization of a mammalian zinc transporter that confers resistance to zinc

A cDNA encoding a zinc transporter (ZnT‐1) was isolated from a rat kidney cDNA expression library by complementation of a mutated, zinc‐sensitive BHK cell line. This cDNA was used to isolate the homologous mouse ZnT‐1 gene. The proteins predicted for these transporters contain six membrane‐spanning domains, a large intracellular loop and a C‐terminal tail. ZnT‐1 is homologous to zinc and cobalt resistance genes of yeast. Immunocytochemistry with an antibody to a myc epitope added to the C‐terminus of ZnT‐1 revealed localization to the plasma membrane. Transformation of normal cells with a mutant ZnT‐1 lacking the first membrane‐spanning domain conferred zinc sensitivity on wild‐type cells, suggesting that ZnT‐1 functions as a multimer. Deletion of the first two membrane‐spanning domains resulted in a non‐functional molecule, whereas deletion of the C‐terminal tail produced a toxic phenotype. Mutant cells have a slightly higher steady‐state level of intracellular zinc and high basal expression of a zinc‐dependent reporter gene compared with normal cells. Mutant cells have a lower turnover of 65Zn compared with normal cells or mutant cells transformed with ZnT‐1. We propose that ZnT‐1 transports zinc out of cells and that its absence accounts for the increased sensitivity of mutant cells to zinc toxicity.