Ceruloplasmin Ferroxidase Activity Stimulates Cellular Iron Uptake by a Trivalent Cation-specific Transport Mechanism

Abstract

The balance required to maintain appropriate cellular and tissue iron levels has led to the evolution of multiple mechanisms to precisely regulate iron uptake from transferrin and low molecular weight iron chelates. A role for ceruloplasmin (Cp) in vertebrate iron metabolism is suggested by its potent ferroxidase activity catalyzing conversion of Fe 2؉ to Fe 3؉ , by identification of yeast copper oxidases homologous to Cp that facilitate high affinity iron uptake, and by studies of "aceruloplasminemic" patients who have extensive iron deposits in multiple tissues. We have recently shown that Cp increases iron uptake by cultured HepG2 cells. In this report, we investigated the mechanism by which Cp stimulates cellular iron uptake. Cp stimulated the rate of non-transferrin 55 Fe uptake by iron-deficient K562 cells by 2-3-fold, using a transferrin receptor-independent pathway. Induction of Cp-stimulated iron uptake by iron deficiency was blocked by actinomycin D and cycloheximide, consistent with a transcriptionally induced or regulated transporter. Cp-stimulated iron uptake was completely blocked by unlabeled Fe 3؉ and by other trivalent cations including Al 3؉ , Ga 3؉ , and Cr 3؉ , but not by divalent cations. These results indicate that Cp utilizes a trivalent cation-specific transporter. Cp ferroxidase activity was required for iron uptake as shown by the ineffectiveness of two ferroxidase-deficient Cp preparations, copper-deficient Cp and thiomolybdate-treated Cp. We propose a model in which iron reduction and subsequent re-oxidation by Cp are essential for an iron uptake pathway with high ion specificity.

Targeted gene disruption reveals an essential role for ceruloplasmin in cellular iron efflux

“…While recent studies in vitro have suggested that ceruloplasmin may function to facilitate non-transferrin-dependent cellular iron influx (15,16), no differences were observed in iron uptake in Cp ϩ/ϩ and Cp Ϫ/Ϫ mice (Table 2), even under experimental conditions designed to favor this uptake pathway when examined over a 72-hr time frame (Fig. 4A).…”

“…Early studies in copper-deficient animals revealed impaired reticuloendothelial cell iron release that was corrected by the administration of ceruloplasmin, implying a role for this protein as a plasma ferroxidase regulating tissue iron efflux (12)(13)(14). However, such studies were clouded by the effects of copper deficiency in these animals, and more recent work with cultured cell lines in vitro has suggested a role for ceruloplasmin in cellular iron influx (15,16). Understanding how ceruloplasmin affects iron homeostasis is essential for the development of effective therapies in aceruloplasminemia and may provide new insight into the pathogenesis of iron overload.…”

Role of Ceruloplasmin in Macrophage Iron Efflux during Hypoxia

“…Interestingly, astrocyte Cp is glycosylphosphatidylinositol-linked suggesting that both free and membrane-bound forms of Cp are involved in iron transport processes. Three other laboratories reported that Cp did not increase iron release (even in the presence of apotransferrin) from HepG2 or erythroleukemic K562 cells (19,20), placental BeWo cells (25), or glioblastoma BT325 cells (21).…”

Multivalent Metal-Induced Iron Acquisition from Transferrin and Lactoferrin by Myeloid Cells

“…In addition, Cu regulates Fe uptake from low-m.w. complexes in both yeast and some human cells through its role in enzymes with ferrioxidase activity that contribute to Fe acquisition (45,(63)(64)(65). However, we have previously shown that Fe reductase activity in HL-60 cells is not altered by exposure to Ga and other metals and, thus, the ability of the tri/ tetravalent metals to alter cellular Fe acquisition cannot be due to effects on Fe reductase activity (50).…”

“…Many cell types have been shown to acquire Fe bound to a variety of low-m.w. chelating agents in vitro (9,12,(43)(44)(45)(46)(47). Several mechanisms have been proposed for this acquisition.…”