Decreased hephaestin expression and activity leads to decreased iron efflux from differentiated Caco2 cells

Chen, Huijun; Attieh, Zouhair; Dang, Thi Mai; Huang, Gang; Hee, R. M. van der; Vulpe, Chris D.

doi:10.1002/jcb.22178

Cited by 26 publications

(17 citation statements)

References 30 publications

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“…The Caco-2 intestinal epithelial cell line was used as a positive control in our immunoblots. Caco-2 cells express both Hp and Fpn and have been established as a model for iron transport across the intestinal barrier (21,24,25). A distinct band at ϳ130 kDa is observed on immunoblots of Caco-2 and hBMVEC extracts probed for Hp; two distinct bands are observed at ϳ63 and 70 kDa when probed for Fpn (Fig.…”

Section: Resultsmentioning

confidence: 91%

Ferroportin and Exocytoplasmic Ferroxidase Activity Are Required for Brain Microvascular Endothelial Cell Iron Efflux

McCarthy

Kosman

2013

Journal of Biological Chemistry

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Background: Cellular iron efflux occurs via cooperation of ferroportin and a ferroxidase. Results: Depletion of ferroxidase and ferroportin from human brain microvascular endothelial cells decreases their ability to efflux iron. Conclusion: Iron efflux from brain microvascular endothelial cell ferroportin requires exocytoplasmic ferroxidase activity. Significance: The mechanism of iron efflux from endothelial cells of the blood-brain barrier is fundamental to how iron enters the brain.

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Section: Resultsmentioning

confidence: 91%

Ferroportin and Exocytoplasmic Ferroxidase Activity Are Required for Brain Microvascular Endothelial Cell Iron Efflux

McCarthy

Kosman

2013

Journal of Biological Chemistry

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“…The cytoplasmic Fe(II) can efflux from the enterocyte at the basolateral surface via the Fe(II) permease Fpn. As noted above, this efflux is dependent on the ferroxidation due to Hp (27,29,54,61,73,74); in plasma, the product of this coupled permeation-oxidation is likely ferric Tf. The redox cycling in Ft (Fig.…”

Section: Ferrireductases and Ferroxidases Are The Iron Metabolic Pathwaymentioning

confidence: 91%

Redox Cycling in Iron Uptake, Efflux, and Trafficking

Kosman

2010

Journal of Biological Chemistry

106

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Aerobic organisms are faced with a dilemma. Environmental iron is found primarily in the relatively inert Fe(III) form, whereas the more metabolically active ferrous form is a strong pro-oxidant. This conundrum is solved by the redox cycling of iron between Fe(III) and Fe(II) at every step in the iron metabolic pathway. As a transition metal ion, iron can be "metabolized" only by this redox cycling, which is catalyzed in aerobes by the coupled activities of ferric iron reductases (ferrireductases) and ferrous iron oxidases (ferroxidases).Following the first crystallization of a protein, jack bean urease, reported by Sumner in 1926 (1), the evolution of protein chemistry remained a slow process, hampered by limitations in techniques for protein resolution and analysis. Not surprisingly, many proteins obtained in relatively pure form were those with prosthetic metal ion groups that imparted a visible absorbance to guide the purification. Some were copper proteins whose Cu(II) spectral properties gave their protein host a greenish blue to bright blue hue. Two of these were from mammalian whole blood: hemocuprein (cupric species from erythrocytes, superoxide dismutase) and ceruloplasmin (cerulean blue species from plasma). Superoxide dismutase was first isolated in 1938 by Mann and Keilin (5); Cp 2 was first isolated in 1948 by Holmberg and Laurell (6). Both proteins play essential roles in aerobic homeostasis. In the case of ceruloplasmin and the other copper proteins that share its substrate specificity, this role is to manage the redox state of iron in the metabolism of this essential yet potentially cytotoxic transition metal ion. These proteins, all of which are members of the MCO family (7-9), and the mechanism by which they provide this function are a focus of this minireview.

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“…Membranes were blocked for one hour at room temperature with shaking in blocking buffer (10% non-fat milk in Tris-buffered saline with 0.1% Tween- 20 (TBST)). Membranes were then incubated with the relevant primary antibody diluted in blocking buffer for one hour at room temperature (FTN [rabbit anti-human FTN, 1∶5000, cat #650771, ICN Biomedicals, Seven Hills, Australia]; HP [rabbit anti-HP C-terminus, 1∶1000, cat #HEPH11-A, Alpha Diagnostics, Owings Mills, MD]; HP [rabbit anti-HP D4 center, 1∶3000, produced in-house [27]]; Actin [mouse monoclonal anti-beta actin, 1∶10,000, cat #ab6276, Abcam, Cambridge, UK]; FPN1 [rabbit anti-FPN1, 1∶2500, cat #MTP11-A, Alpha Diagnostics]). The membranes were then washed with TBST and incubated for one hour at room temperature with secondary antibody (anti-rabbit HRP, 1∶8000, Merck, or anti-mouse HRP, 1∶10,000).…”

Section: Methodsmentioning

confidence: 99%

The Multicopper Ferroxidase Hephaestin Enhances Intestinal Iron Absorption in Mice

Fuqua

Darshan

et al. 2014

PLoS ONE

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Hephaestin is a vertebrate multicopper ferroxidase important for the transfer of dietary iron from intestinal cells to the blood. Hephaestin is mutated in the sex-linked anemia mouse, resulting in iron deficiency. However, sex-linked anemia mice still retain some hephaestin ferroxidase activity. They survive, breed, and their anemia improves with age. To gain a better understanding of the role of hephaestin in iron homeostasis, we used the Cre-lox system to generate knockout mouse models with whole body or intestine-specific (Villin promoter) ablation of hephaestin. Both types of mice were viable, indicating that hephaestin is not essential and that other mechanisms, multicopper ferroxidase-dependent or not, must compensate for hephaestin deficiency. The knockout strains, however, both developed a microcytic, hypochromic anemia, suggesting severe iron deficiency and confirming that hephaestin plays an important role in body iron acquisition. Consistent with this, the knockout mice accumulated iron in duodenal enterocytes and had reduced intestinal iron absorption. In addition, the similarities of the phenotypes of the whole body and intestine-specific hephaestin knockout mice clarify the important role of hephaestin specifically in intestinal enterocytes in maintaining whole body iron homeostasis. These mouse models will serve as valuable tools to study the role of hephaestin and associated proteins in iron transport in the small intestine and other tissues.

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Decreased hephaestin expression and activity leads to decreased iron efflux from differentiated Caco2 cells

Cited by 26 publications

References 30 publications

Ferroportin and Exocytoplasmic Ferroxidase Activity Are Required for Brain Microvascular Endothelial Cell Iron Efflux

Ferroportin and Exocytoplasmic Ferroxidase Activity Are Required for Brain Microvascular Endothelial Cell Iron Efflux

Redox Cycling in Iron Uptake, Efflux, and Trafficking

The Multicopper Ferroxidase Hephaestin Enhances Intestinal Iron Absorption in Mice

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