Ferroportin is a monomer in vivo in mice

Pignatti, Elisa; Mascheroni, Laura; Sabelli, Manuela; Barelli, S.; Biffo, Stefano; Pietrangelo, Antonello

doi:10.1016/j.bcmd.2005.11.001

Cited by 26 publications

(19 citation statements)

References 23 publications

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“…The oligomeric state of Fpn has been debated for several years – the protein has been reported to be a monomer36,38,41 or a dimer/multimer 30,37. Using SEC-LS/UV/RI, a shape- and model-independent method to obtain a molecular mass,34 we determined that purified detergent-solubilized Fpn is monomeric (Figure 1C).…”

Section: Discussionmentioning

confidence: 96%

Investigation of the Biophysical and Cell Biological Properties of Ferroportin, a Multipass Integral Membrane Protein Iron Exporter

Rice

Mendez

Hokanson

et al. 2009

Journal of Molecular Biology

108

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Ferroportin is a multi-pass membrane protein that serves as an iron exporter in many vertebrate cell types. Ferroportin-mediated iron export is controlled by the hormone hepcidin, which binds ferroportin, causing its internalization and degradation. Mutations in ferroportin cause a form of the iron overload disease hereditary hemochromatosis. Relatively little is known about ferroportin's properties or the mechanism by which mutations cause disease. Here we expressed and purified human ferroportin to characterize its biochemical/biophysical properties in solution and conducted cell biological studies in mammalian cells. We show that purified, detergent-solubilized ferroportin was a well-folded monomer that bound hepcidin. In cell membranes, the N-and C-termini were both cytosolic, implying an even number of transmembrane regions, and ferroportin was mainly localized to the plasma membrane. Hepcidin addition resulted in a redistribution of ferroportin to intracellular compartments that labeled with early endosomal and lysosomal, but not Golgi, markers and that trafficked along microtubules. An analysis of 16 disease-related ferroportin mutants revealed that all formed well-folded monomers that localized to the plasma membrane, but some were resistant to hepcidin-induced internalization. The characterizations reported here form a basis upon which models for ferroportin's role in regulating iron homeostasis in health and disease can be interpreted.

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Section: Discussionmentioning

confidence: 96%

Investigation of the Biophysical and Cell Biological Properties of Ferroportin, a Multipass Integral Membrane Protein Iron Exporter

Rice

Mendez

Hokanson

et al. 2009

Journal of Molecular Biology

108

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“…This is in part because the main published evidence supporting the multimeric nature of ferroportin (De Domenico et al, 2007c) is clouded by scientific misconduct (paper #9 in the report by (McCormack et al, 2013)). Several groups reported evidence that ferroportin is a monomer, as determined by a variety of methods (Goncalves et al, 2006; Pignatti et al, 2006; Schimanski et al, 2008) but others detected a dimeric form complexed with hephaestin (Yeh et al, 2009). Detailed physicochemical analysis of recombinant ferroportin extracted from membranes (Rice et al., 2009) led to the conclusion that the monomeric form is stable and functional, and binds hepcidin, but that weak, detergent-sensitive homophilic interactions favouring multimerization in the cell membrane could not definitively be ruled out.…”

Section: Genetic Disorders Of Ferroportin and Their Effects On Systemmentioning

confidence: 99%

Ironing out Ferroportin

2015

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Maintaining physiologic iron concentrations in tissues is critical for metabolism and host defense. Iron absorption in the duodenum, recycling of iron from senescent erythrocytes, and iron mobilization from storage in macrophages and hepatocytes constitute the major iron flows into plasma for distribution to tissues, predominantly for erythropoiesis. All iron transfer to plasma occurs through the iron exporter ferroportin. The concentration of functional membrane-associated ferroportin is controlled by its ligand, the iron-regulatory hormone hepcidin, and fine-tuned by regulatory mechanisms serving iron homeostasis, oxygen utilization, host defense and erythropoiesis. Fundamental questions about the structure and biology of ferroportin remain to be answered.

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“…While studies have supported the view that Fpn is a multimer 9,13,24 and that the mutant allele can affect the behavior of the wild-type allele, other studies have suggested that Fpn is monomeric. 10,12,25 All human Fpn mutations are missense mutations. If haploinsufficiency was the explanation for Fpn disease, then nonsense mutations should also result in the disorder; however, none have been found.…”

Section: Discussionmentioning

confidence: 99%

The flatiron mutation in mouse ferroportin acts as a dominant negative to cause ferroportin disease

Zohn

Domenico

Pollock

et al. 2007

Blood

105

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IntroductionHereditary hemochromatosis is a common disorder in humans, characterized by iron overload resulting in tissue injury and ultimately organ failure. Typically, hemochromatosis exhibits an autosomal-recessive pattern of inheritance and is associated with mutations in HFE, hemojuvelin, hepcidin, or transferrin receptor 2. 1,2 Targeted deletion of these genes in the mouse results in hemochromatosis, providing mouse models for most forms of the disease. Hemochromatosis type IV, also referred to as ferroportin (Fpn) disease, results from mutations in the iron transporter ferroportin. Fpn is the only known iron exporter in mammalian cells and is present on the surface of macrophages, intestinal enterocytes, hepatocytes, and placental cells. [3][4][5] The level of cell surface Fpn is regulated by its interaction with hepcidin, a peptide secreted by the liver in response to iron stores and inflammation. Hepcidin binds to Fpn, inducing its internalization and degradation, thus regulating the export of iron from cells to plasma. 6 Mutations in Fpn lead to iron-overload disease but, in contrast to other forms of hemochromatasis, ferroportin disease exhibits an autosomal-dominant pattern of inheritance. 7 The disorder has different presentations depending on the Fpn mutation. Mutations leading to Fpn that is not internalized by hepcidin result in iron accumulation in hepatocytes and high transferrin saturation. 8,9 Mutations leading to Fpn that is not appropriately targeted to the cell surface result in iron accumulation in Kupffer cells and low transferrin saturation. [9][10][11] The mechanism by which the disease mutations exert a dominant effect is unclear. Some groups that study the disease suggest that it results from haploinsufficiency, 10,12 whereas others suggest that the disorder results from a dominantnegative effect of the mutant allele. 9,13 Importantly, all human mutations are missense mutations and mice that are heterozygous for a targeted deletion of Fpn do not show the disease. 14 Treatment for hemochromatosis aims to decrease iron load by repeated phlebotomy and this treatment works well for most patients. Many patients with ferroportin disease, however, become anemic with phlebotomy, highlighting the need for a mouse model to develop better treatments.We report here on a missense mutation in mouse Fpn that results in a disorder that is identical to classic human ferroportin disease. We show that macrophages isolated from mutant mice have no Fpn on their cell surface and that expression of Fpn constructs containing the missense mutation (H32R) affects the behavior of wild-type Fpn. These results show that Fpn disease is due to a dominant-negative effect of the mutant allele and provide the first mouse model for this disorder. Materials and methods Generation of mutant mice and identification of Fpn mutationThe ffe mouse line was identified in a screen for recessive ethylnitrosourea (ENU)-induced mutations that cause morphologic abnormalities at embryonic day (E) 12.5. [15][16][17] The ffe mutation...

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Ferroportin is a monomer in vivo in mice

Cited by 26 publications

References 23 publications

Investigation of the Biophysical and Cell Biological Properties of Ferroportin, a Multipass Integral Membrane Protein Iron Exporter

Investigation of the Biophysical and Cell Biological Properties of Ferroportin, a Multipass Integral Membrane Protein Iron Exporter

Ironing out Ferroportin

The flatiron mutation in mouse ferroportin acts as a dominant negative to cause ferroportin disease

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