Evidence for the multimeric structure of ferroportin.
Ferroportin (Fpn) (IREG1, SLC40A1, MTP1) is an iron transporter, and mutations in Fpn result in a genetically dominant form of iron overload disease. Previously, we demonstrated that Fpn is a multimer and that mutations in Fpn are dominant negative. Other studies have suggested that Fpn is not a multimer and that overexpression or epitope tags might affect the localization, topology, or multimerization of Fpn. We generated wild-type Fpn with 3 different epitopes, GFP, FLAG, and c-myc, and expressed these constructs in cultured cells. Co-expression of any 2 different epitope-tagged proteins in the same cell resulted in their quantitative coimmunoprecipitation. Treatment of Fpn-GFP/Fpn-FLAG-expressing cells with crosslinking reagents resulted in the crosslinking of Fpn-GFP and Fpn-FLAG. Western analysis of rat glioma C6 cells or mouse bone marrow macrophages exposed to crosslinking reagents showed that endogenous Fpn is a dimer. These results support the hypothesis that the dominant inheritance of Fpn-iron overload disease is due to the dominant-negative effects of mutant Fpn proteins.
De Domenico
Ivana
I
Department of Pathology, School of Medicine, University of Utah, Salt Lake City, UT 84132, USA.
Ward
Diane McVey
DM
Musci
Giovanni
G
Kaplan
Jerry
J
D000818:Animals; D027682:Cation Transport Proteins / Q000235:genetics / Q000276:immunology / Q000378:metabolism*; D002460:Cell Line; D000939:Epitopes / Q000276:immunology; D006801:Humans; D051379:Mice; D011485:Protein Binding; D051381:Rats; D000098742:Ferroportin
D027682:Cation Transport Proteins; D000939:Epitopes; D000098742:Ferroportin