Intracellular Ca 2+ regulates the cellular iron uptake in K562 cells

Abstract

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 Ca 2+ ([Ca 2+] 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 Ca 2+ or complete chelation of extracellular Ca 2+ 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, [Ca 2+] i-elevated and W-7-treated cells revealed that the actin polymerization is enhanced as [Ca 2+] i is raised, but reduced by W-7. The results suggest that the regulation of actin polymerization by intracellular Ca 2+ may play a central role in Ca 2+-dependent iron uptake.