Inhibitory Mechanism of Store-operated Ca²⁺ Channels by Zinc

Capacitative calcium influx plays an important role in shaping the Ca ²⁺ response of various tissues and cell types. Inhibition by heavy metals is a hallmark of store-operated calcium channel (SOCC) activity. Paradoxically, although zinc is the only potentially physiological relevant ion, it is the least investigated in terms of inhibitory mechanism. In the present study, we characterize the inhibitory mechanism of the SOCC by Zn²⁺ in the human salivary cell line, HSY, and rat salivary submandibular ducts and acini by monitoring SOCC activity using fluorescence imaging. Analysis of Zn²⁺ inhibition indicated that Zn²⁺ acts as a competitive inhibitor of Ca²⁺ influx but does not permeate through the SOCC, suggesting that Zn²⁺ interacts with an extracellular site of SOCC. Application of the reducing agents, dithiothreitol (DTT) and β-mercaptoethanol, totally eliminated Zn²⁺ and Cd²⁺ inhibition of SOCC, suggesting that cysteines are part of the Zn²⁺ and Cd²⁺ binding site. Interestingly, reducing conditions failed to eliminate the inhibition of SOCC by La³⁺ and Gd³⁺, indicating that the Zn²⁺ and lanthanides binding sites are distinct. Finally, we show that changes in redox potential and Zn²⁺ are regulating, via SOCC activity, the agonist-induced Ca²⁺ response in salivary ducts. The presence of a specific Zn²⁺ site, responsive to physiological Zn²⁺ and redox potential, may not only be instrumental for future structural studies of various SOCC candidates but may also reveal novel physiological aspects of the interaction between zinc, redox potential, and cellular Ca²⁺ homeostasis.