TRANSPORT OF IRON AND OTHER TRANSITION-METALS INTO CELLS AS REVEALED BY A FLUORESCENT-PROBE

Transport of nontransferrin-bound iron into cells is thought to be med iated by a facilitated mechanism involving either the trivalent form F e(III) or the divalent form Fe(II) following reduction of Fe(III) at t he cell surface. We have made use of the probe calcein, whose fluoresc ence is rapidly and stoichiometrically quenched by divalent metals suc h as Fe(II), Cu(II), Co(II), and Ni(II) and is minimally affected by v ariations in ionic strength, Ca(II) and Mg(II). Addition of Fe(II) sal ts to calcein-loaded human erythroleukemia K-562 cells elicited a slow quenching response that was markedly accelerated by the ionophore A-2 3187 and was reversed by membrane-permeant but not by impermeant chela tors. These observations were confirmed by fluorescence imaging of cel ls. Other divalent metals such as Co(II), Ni(II), and Mn(II) permeated into cells at roughly similar rates, and their uptake, like that of F e(II), was blocked by trifluoperazine, bepridil, and impermeant sulfhy dryl-reactive organomercurials, indicating the operation of a common t ransport mechanism. This method could provide a versatile tool for stu dying the transport of iron and other transition metals into cells.