E-cadherin, a calcium (Ca2+)-dependent cell-cell adhesion molecule, plays a key role in the maintenance of tissue integrity. We have previously demonstrated that E-cadherin functions in vivo as a cis-dimer through chemical cross-linking reagents. Ca2+ plays an important role in the cis-dimer formation of cadherin. However, the molecular mechanisms by which Ca2+ interacts with the binding sites that regulate cis-dimer structures have not been completely elucidated. As expected for a Ca2+ antagonist, cadmium (Cd2+) disrupts cadherin function by displacing Ca2+ from its binding sites on the cadherin molecules. We used Cd2+ as a probe for investigating the role of Ca2+ in the dynamics of the E-cadherin extracellular region that involve cis-dimer formation and adhesion. While cell-cell adhesion assembly was completely disrupted in the presence of Cd2+, the amount of cis-dimers of E-cadherin that formed at the cell surface was not affected. In our "Cd2+-switch" experiments, we did not find that Cd2+-induced E-cadherin cis-dimer formation in EL cells when they were incubated in low-Ca2+ medium. In the present study, we demonstrated for the first time the effects of Cd2+ on the cis-dimer structure of E-cadherin in living cells using a chemical cross-link analysis. (C) 2014 Elsevier Inc. All rights reserved.