Zinc‐metallothionein ratios reflect cellular zinc status

Abstract

Animal and human studies show that intracellular zinc levels are sensitive to changes in dietary zinc. The protein metallothionein (MT) is maintained in proportion to intracellular zinc content, and therefore may be a useful marker of zinc status. Measuring MT levels is difficult due to the unique properties of the protein, so most studies focus on MT gene expression. Our objective was to develop new methods to quantify MT protein and MT‐bound zinc content. Zinc deficiency or excess was modeled in the Jurkat leukocytic cell line using the zinc‐selective chelator TPEN or exogenous zinc sulfate, respectively. Cellular protein was separated using size‐exclusion filtration to isolate an MT‐rich fraction. Then, endogenous zinc content was released, apo‐MT was loaded with cadmium, and MT‐bound cadmium levels were quantified by ICP; this method permitted the determination of cellular MT‐bound zinc content (MBZC) and MT zinc saturation (%MTsat). Gene expression of the major MT and zinc transporter isoforms were measured in parallel by RT‐PCR, which reflected the change in zinc balance in these cells. MBZC and %MTsat levels correlated with intracellular zinc content and changes in MT gene expression, showing bimodal responses to zinc deficiency and zinc excess. These measures of the cellular zinc‐metallothionein ratio may have utility as markers of zinc homeostasis. This work was supported by the HarvestPlus.