Iron Uptake Mediated by Binding of H-Ferritin to the TIM-2 Receptor in Mouse Cells

Jian Han,William E Seaman,Mark Willingham,Xiumin Di,Suzy V Torti,Frank M Torti,Wei Wang,Martin W Brechbiel

doi:10.1371/journal.pone.0023800

Abstract

Ferritin binds specifically and saturably to a variety of cell types, and recently several ferritin receptors have been cloned. TIM-2 is a specific receptor for H ferritin (HFt) in the mouse. TIM-2 is a member of the T cell immunoglobulin and mucin domain containing (TIM) protein family and plays an important role in immunity. The expression of TIM-2 outside of the immune system indicates that this receptor may have broader roles. We tested whether ferritin binding to TIM-2 can serve as an iron delivery mechanism. TIM-2 was transfected into normal (TCMK-1) mouse kidney cells, where it was appropriately expressed on the cell surface. HFt was labeled with 55Fe and 55Fe-HFt was incubated with TIM-2 positive cells or controls. 55Fe-HFt uptake was observed only in TIM-2 positive cells. HFt uptake was also seen in A20 B cells, which express endogenous TIM-2. TIM-2 levels were not increased by iron chelation. Uptake of 55Fe-HFt was specific and temperature-dependent. HFt taken up by TIM-2 positive cells transited through the endosome and eventually entered a lysosomal compartment, distinguishing the HFt pathway from that of transferrin, the classical vehicle for cellular iron delivery. Iron delivered following binding of HFt to TIM-2 entered the cytosol and became metabolically available, resulting in increased levels of endogenous intracellular ferritin. We conclude that TIM-2 can function as an iron uptake pathway.

Highlights

Ferritin is a ubiquitously distributed protein principally known for its role in iron storage and detoxification [1,2]
We compared levels of cell surface TIM-2 in transfectants to A20 cells, a mouse lymphocyte cell line that expresses endogenous TIM-2, as well as to A20 cells that had been transfected with a TIM-2 expression vector to increase expression of TIM-2 [22]
Immunoprecipitation confirmed that levels of TIM-2 in untreated TCMK-TIM-2 transfectants were approximately equivalent to levels of endogenous TIM-2 in untreated A20 cells (Fig. 2C)

Summary

Introduction

Ferritin is a ubiquitously distributed protein principally known for its role in iron storage and detoxification [1,2]. It is composed of two subunit types, termed H and L; twenty-four of these assemble to form the ferritin protein. Ferritin is regulated post-transcriptionally by iron through the action of iron regulatory proteins (IRPs), which act as ferritin translational repressors (see [4,5] for review). IRPs control the levels of transferrin receptor 1 (TfR1), a protein that mediates uptake of iron bound to transferrin, the principal source of iron in mammalian cells. TfR1 mRNA and protein increase under conditions of iron depletion, such as that induced by an iron chelator

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Results

Conclusion