Lntracellular Ca2+ regulates the cellular iron uptake in K562 cells

Fluorescence quenching was used to study the kinetics of the transferrin receptor (TfR)-mediated iron uptake in the calcein-loaded K562 cells. It was found that elevation of intracellular free Ca2+ ([Ca2+](i)) by thapsigargin (TG) speeds up the initial rate of iron uptake and increases the overall capacity of the cells in taking up iron. Depletion of intracellular Ca2+ or complete chelation of extracellular Ca2+ results in complete inhibition of the iron uptake in cells. To gain insight into molecular mechanism, IANBD-labeled transferrin (Tf) and microscopic fluorescence imaging were used to observe the endocytosis and recycling of the Tf-TfR complex in single live cells. The study showed that the preincubation of cells with TG or phorbol myristate acetate (PMA), the direct activator of protein kinase C (PKC), accelerated the endocytosis and recycling of the complex in a dose-dependent manner. W-7, the calmodulin antagonist, and GF109203X, a selected cell-permeant inhibitor of PKC, can reverse the acceleration. Analysis of actin polymerization in controlled, [Ca2+](i)-elevated and W-7-treated cells revealed that the actin polymerization is enhanced as [Ca2+](i) is raised, but reduced by W-7. The results suggest that the regulation of actin polymerization by intracellular Ca2+ may play a central role in Ca2+-dependent iron uptake.Department of Applied Biology and Chemical Technolog