IRON TRANSPORT IN K562 CELLS - A KINETIC-STUDY USING NATIVE GEL-ELECTROPHORESIS AND FE-59 AUTORADIOGRAPHY

The exact mechanisms of iron transport from endosomes to the target ir on containing cellular proteins are currently unknown. To investigate this problem, we used the gradient gel electrophoresis and the sensiti ve detection of Fe-59 by autoradiography to detect separate cellular i ron compounds and their iron kinetics. Cells of human leukemic line K5 62 were labeled with [Fe-59]transferrin for 30-600 s and cellular iron compounds in cell lysates were analyzed by native electrophoretic sep aration followed by Fe-59 autoradiography. Starting with the first 30 s of iron uptake, iron was detectable in a large membrane bound protei n complex (Band I) and in ferritin. Significant amounts of iron were a lso found in labile iron compound(s) with the molecular weight larger than 5000 as judged by ultrafiltration. Iron kinetics in these compart ments was studied. Band I was the only compound with the kinetic prope rties of an intermediate. Transferrin, transferrin receptor and additi onal proteins of the approximate molecular weights of 130 000, 66000 a nd 49 000 were found to be present in Band I. The labile iron compound s and ferritin behaved kinetically as end products. No evidence for lo w molecular weight transport intermediates was found. These results su ggest that intracellular iron transport is highly compartmentalized, t hat iron released from endosomal transferrin passes to its cellular ta rgets in a direct contact with the endosomal membrane complex assigned as Band I. The nature of the labile iron pool and its susceptibility to iron chelation is discussed. (C) 1998 Elsevier Science B.V.