Neogenin-mediated hemojuvelin shedding occurs after hemojuvelin traffics to the plasma membrane

HFE2 (hemochromatosis type 2 gene) is highly expressed in skeletal muscle and liver hepatocytes. Its encoded protein, hemojuvelin (HJV), is a co-receptor for the bone morphogenetic proteins 2 and 4 (BMP2 and BMP4) and enhances the BMP-induced hepcidin expression. Hepcidin is a central iron regulatory hormone predominantly secreted from hepatocytes. HJV also binds neogenin, a membrane protein widely expressed in many tissues. Neogenin is required for the processing and release of HJV from cells. The role that neogenin plays in HJV trafficking was investigated, using HepG2 cells, a human hepatoma cell line. Knockdown of endogenous neogenin markedly suppresses HJV release but has no evident effect on HJV trafficking to the plasma membrane. The addition of a soluble neogenin ectodomain to cells markedly inhibits HJV release, indicating that the HJV shedding is not processed before trafficking to the cell surface. At the plasma membrane it undergoes endocytosis in a dynamin-independent but cholesterol-dependent manner. The additional findings that HJV release is coupled to lysosomal degradation of neogenin and that cholesterol depletion by filipin blocks both HJV endocytosis and HJV release suggest that neogenin-mediated HJV release occurs after the HJV-neogenin complex is internalized from the cell surface.     

Selective binding of RGMc/hemojuvelin, a key protein in systemic iron metabolism, to BMP-2 and neogenin

Juvenile hemochromatosis is a severe and rapidly progressing hereditary disorder of iron overload, and it is caused primarily by defects in the gene encoding repulsive guidance molecule c/hemojuvelin (RGMc/HJV), a recently identified protein that undergoes a complicated biosynthetic pathway in muscle and liver, leading to cell membrane-linked single-chain and heterodimeric species, and two secreted single-chain isoforms. RGMc modulates expression of the hepatic iron regulatory factor, hepcidin, potentially through effects on signaling by the bone morphogenetic protein (BMP) family of soluble growth factors. To date, little is known about specific pathogenic defects in disease-causing RGMc/HJV proteins. Here we identify functional abnormalities in three juvenile hemochromatosis-linked mutants. Using a combination of approaches, we first show that BMP-2 could interact in biochemical assays with single-chain RGMc species, and also could bind to cell-associated RGMc. Two mouse RGMc amino acid substitution mutants, D165E and G313V (corresponding to human D172E and G320V), also could bind BMP-2, but less effectively than wild-type RGMc, while G92V (human G99V) could not. In contrast, the membrane-spanning protein, neogenin, a receptor for the related molecule, RGMa, preferentially bound membrane-associated heterodimeric RGMc and was able to interact on cells only with wild-type RGMc and G92V. Our results show that different isoforms of RGMc/HJV may play unique physiological roles through defined interactions with distinct signaling proteins and demonstrate that, in some disease-linked RGMc mutants, these interactions are defective.     

Neogenin Interacts with Matriptase-2 to Facilitate Hemojuvelin Cleavage

Hemojuvelin (HJV) and matriptase-2 (MT2) are co-expressed in hepatocytes, and both are essential for systemic iron homeostasis. HJV is a glycosylphosphatidylinositol-linked membrane protein that acts as a co-receptor for bone morphogenetic proteins to induce hepcidin expression. MT2 regulates the levels of membrane-bound HJV in hepatocytes by binding to and cleaving HJV into an inactive soluble form that is released from cells. HJV also interacts with neogenin, a ubiquitously expressed transmembrane protein with multiple functions. In this study, we showed that neogenin interacted with MT2 as well as with HJV and facilitated the cleavage of HJV by MT2. In contrast, neogenin was not cleaved by MT2, indicating some degree of specificity by MT2. Down-regulation of neogenin with siRNA increased the amount of MT2 and HJV on the plasma membrane, suggesting a lack of neogenin involvement in their trafficking to the cell surface. The increase in MT2 and HJV upon neogenin knockdown was likely due to the inhibition of cell surface MT2 and HJV internalization. Analysis of the Asn-linked oligosaccharides showed that MT2 cleavage of cell surface HJV was coupled to a transition from high mannose oligosaccharides to complex oligosaccharides on HJV. These results suggest that neogenin forms a ternary complex with both MT2 and HJV at the plasma membrane. The complex facilitates HJV cleavage by MT2, and release of the cleaved HJV from the cell occurs after a retrograde trafficking through the TGN/Golgi compartments.     

Evidence that inhibition of hemojuvelin shedding in response to iron is mediated through neogenin

Hemojuvelin (HJV), encoded by the gene HFE2, is a critical upstream regulator of hepcidin expression. Hepcidin, the central iron regulatory hormone, is secreted from hepatocytes, whereas HFE2 is highly expressed in skeletal muscle and liver. Previous studies demonstrated that HJV is a GPI-anchored protein, binds the proteins neogenin and bone morphogenetic proteins (BMP2 and BMP4), and can be released from the cell membrane (shedding). In this study, we investigated the physiological significance and the underlying mechanism of HJV shedding. In acutely iron-deficient rats with markedly suppressed hepatic hepcidin expression, we detected an early phase increase of serum HJV with no significant change of either HFE2 mRNA or protein levels in gastrocnemius muscle. Studies in both C2C12 (a mouse myoblast cell line) and HepG2 (a human hepatoma cell line) cells showed active HJV shedding, implying that both skeletal muscle and liver could be the source of serum HJV. In agreement with the observations in iron-deficient rats, HJV shedding in these cell lines was down-regulated by holo-transferrin in a concentration-dependent manner. Our present study showing that knock-down of endogenous neogenin, a HJV receptor, in C2C12 cells suppresses HJV shedding and that overexpression of neogenin in HEK293 cells markedly enhances this process, suggests that membrane HJV shedding is mediated by neogenin. The finding that neither BMP4 nor its antagonist, noggin, was able to alter HJV shedding support the lack of involvement of BMP signaling pathway in this process.     

The hereditary hemochromatosis protein, HFE, specifically regulates transferrin-mediated iron uptake in HeLa cells

HFE is the protein product of the gene mutated in the autosomal recessive disease hereditary hemochromatosis (Feder, J. N., Gnirke, A., Thomas, W., Tsuchihashi, Z., Ruddy, D. A., Basava, A., Dormishian, F., Domingo, R. J., Ellis, M. C., Fullan, A., Hinton, L. M., Jones, N. L., Kimmel, B. E., Kronmal, G. S., Lauer, P., Lee, V. K., Loeb, D. B., Mapa, F. A., McClelland, E., Meyer, N. C., Mintier, G. A., Moeller, N., Moore, T., Morikang, E., Prasss, C. E., Quintana, L., Starnes, S. M., Schatzman, R. C., Brunke, K. J., Drayna, D. T., Risch, N. J., Bacon, B. R., and Wolff, R. R. (1996) Nat. Genet. 13, 399-408). At the cell surface, HFE complexes with transferrin receptor (TfR), increasing the dissociation constant of transferrin (Tf) for its receptor 10-fold (Gross, C. N., Irrinki, A., Feder, J. N., and Enns, C. A. (1998) J. Biol. Chem. 273, 22068-22074; Feder, J. N., Penny, D. M., Irrinki, A., Lee, V. K., Lebron, J. A., Watson, N. , Tsuchihashi, Z., Sigal, E., Bjorkman, P. J., and Schatzman, R. C. (1998) Proc. Natl. Acad. Sci. U S A 95, 1472-1477). HFE does not remain at the cell surface, but traffics with TfR to Tf-positive internal compartments (Gross et al., 1998). Using a HeLa cell line in which the expression of HFE is controlled by tetracycline, we show that the expression of HFE reduces 55Fe uptake from Tf by 33% but does not affect the endocytic or exocytic rates of TfR cycling. Therefore, HFE appears to reduce cellular acquisition of iron from Tf within endocytic compartments. HFE specifically reduces iron uptake from Tf, as non-Tf-mediated iron uptake from Fe-nitrilotriacetic acid is not altered. These results explain the decreased ferritin levels seen in our HeLa cell system and demonstrate the specific control of HFE over the Tf-mediated pathway of iron uptake. These results also have implications for the understanding of cellular iron homeostasis in organs such as the liver, pancreas, heart, and spleen that are iron loaded in hereditary hemochromatotic individuals lacking functional HFE.     

Expression of Human Hemojuvelin (HJV) Is Tightly Regulated by Two Upstream Open Reading Frames in HJV mRNA That Respond to Iron Overload in Hepatic Cells

The gene encoding human hemojuvelin (HJV) is one of the genes that, when mutated, can cause juvenile hemochromatosis, an early-onset inherited disorder associated with iron overload. The 5′ untranslated region of the human HJV mRNA has two upstream open reading frames (uORFs), with 28 and 19 codons formed by two upstream AUGs (uAUGs) sharing the same in-frame stop codon. Here we show that these uORFs decrease the translational efficiency of the downstream main ORF in HeLa and HepG2 cells. Indeed, ribosomal access to the main AUG is conditioned by the strong uAUG context, which results in the first uORF being translated most frequently. The reach of the main ORF is then achieved by ribosomes that resume scanning after uORF translation. Furthermore, the amino acid sequences of the uORF-encoded peptides also reinforce the translational repression of the main ORF. Interestingly, when iron levels increase, translational repression is relieved specifically in hepatic cells. The upregulation of protein levels occurs along with phosphorylation of the eukaryotic initiation factor 2α. Nevertheless, our results support a model in which the increasing recognition of the main AUG is mediated by a tissue-specific factor that promotes uORF bypass. These results support a tight HJV translational regulation involved in iron homeostasis.     

The plasmamembrane calmodulin-dependent calcium pump: a major regulator of nitric oxide synthase I

The plasma membrane calcium/calmodulin-dependent calcium ATPase (PMCA) (Shull, G.E., and J. Greeb. 1988. J. Biol. Chem. 263:8646-8657; Verma, A.K., A.G. Filoteo, D.R. Stanford, E.D. Wieben, J.T. Penniston, E.E. Strehler, R. Fischer, R. Heim, G. Vogel, S. Mathews, et al. 1988. J. Biol. Chem. 263:14152-14159; Carafoli, E. 1997. Basic Res. Cardiol. 92:59-61) has been proposed to be a regulator of calcium homeostasis and signal transduction networks of the cell. However, little is known about its precise mechanisms of action. Knock-out of (mainly neuronal) isoform 2 of the enzyme resulted in hearing loss and balance deficits due to severe inner ear defects, affecting formation and maintenance of otoconia (Kozel, P.J., R.A. Friedman, L.C. Erway, E.N. Yamoah, L.H. Liu, T. Riddle, J.J. Duffy, T. Doetschman, M.L. Miller, E.L. Cardell, and G.E. Shull. 1998. J. Biol. Chem. 273:18693-18696). Here we demonstrate that PMCA 4b is a negative regulator of nitric oxide synthase I (NOS-I, nNOS) in HEK293 embryonic kidney and neuro-2a neuroblastoma cell models. Binding of PMCA 4b to NOS-I was mediated by interaction of the COOH-terminal amino acids of PMCA 4b and the PDZ domain of NOS-I (PDZ: PSD 95/Dlg/ZO-1 protein domain). Increasing expression of wild-type PMCA 4b (but not PMCA mutants unable to bind PDZ domains or devoid of Ca2+-transporting activity) dramatically downregulated NO synthesis from wild-type NOS-I. A NOS-I mutant lacking the PDZ domain was not regulated by PMCA, demonstrating the specific nature of the PMCA-NOS-I interaction. Elucidation of PMCA as an interaction partner and major regulator of NOS-I provides evidence for a new dimension of integration between calcium and NO signaling pathways.     

Possible roles of the hereditary hemochromatosis protein, HFE, in regulating cellular iron homeostasis

Hereditary hemochromatosis (HH) is the most common inherited disorder in people of Northern European descent. Over 83% of the cases of HH result from a single mutation of a Cys to Tyr in the HH protein. HFE. This mutation causes a recessive disease resulting in an accumulation of iron in selected tissues. Iron overload damages these organs leading to cirrhosis of the liver, diabetes, cardiomyopathy, and arthritis. The mechanism by which HFE influences iron homeostasis in cells and in the body remains elusive. Lack of functional HFE in humans produces the opposite effects in different cell types in the body. In the early stages of the disease. Kupffer cells in the liver and enterocytes in the intestine cells are iron depleted and have low intracellular ferritin levels, whereas hepatocytes in the liver are iron overloaded and have high intracellular iron levels. This review gives the background and a model as to possible mechanisms of how HFE could exert different effects on iron homeostasis in different cell types.     

海帕西啶铁调节和血色素沉着

血色素沉着是一种血浆铁沉积过多而导致的器官损伤性疾病,多种铁调节基因如HFE、HJV、HAMP和TfR2等的突变均可导致该病的发生,其中HAMP是最为重要的一种。HAMP基因编码一种名为海帕西啶的小肽,是小肠铁重吸收和巨噬细胞铁释放的负调节因子。海帕西啶含量的减少将导致血清铁过负荷和血色素沉着的发生,HFE、HJV和TfR2等基因可影响海帕西啶的表达,从而使海帕西啶成为血色素沉着的中央调节者。这些研究对血色素沉着发生机制的理解及其诊断和治疗具有重要意义。     

Molecular Evolution of Hemojuvelin and the Repulsive Guidance Molecule Family

Repulsive guidance molecules (RGMs) are found in vertebrates and chordates and are involved in embryonic development and iron homeostasis. Members of this family are GPI-linked membrane proteins that contain an N-terminal signal peptide, a C-terminal propeptide, and a conserved RGD motif. Vertebrates are known to possess three paralogues; RGMA and RGMB (sometimes called Dragon) are expressed in the nervous system and are thought to play various roles in neural development. Hemojuvelin (HJV; also called repulsive guidance molecule c, RGMC) is the third member of this family, and mutations in this gene result in a form of juvenile hemochromatosis (type 2A). Phylogenetic analyses of 55 different RGM family sequences from 21 different species support the existence of a novel gene, found only in fish, which we have labeled RGMD. The pattern of conserved residues in each family identifies new candidates for important functional roles, including ligand binding. [Reviewing Editor: Dr. Gail Simmons]     

Expression studies of neogenin and its ligand hemojuvelin in mouse tissues.

Juvenile hemochromatosis is a severe hereditary iron overload disease caused by mutations in the HJV (hemojuvelin) and HAMP (hepcidin) genes. Hepcidin is an important iron regulatory hormone, and hemojuvelin may regulate hepcidin synthesis via the multifunctional membrane receptor neogenin. We explored the expression of murine hemojuvelin and neogenin mRNAs and protein. Real-time RT-PCR analysis of 18 tissues from male and female mice was performed to examine the mRNA expression profiles. To further study protein expression and localization we used immunohistochemistry on several tissues from three mouse strains. Mouse Neo1 mRNA was detectable in the 18 tissues tested, the highest signals being evident in the ovary, uterus, and testis. Neogenin protein was observed in the brain, skeletal muscle, heart, liver, stomach, duodenum, ileum, colon, renal cortex, lung, testis, ovary, oviduct, and uterus. The spleen, thymus, and pancreas were negative for neogenin. The highest signals for Hjv mRNA were detectable in the skeletal muscle, heart, esophagus, and liver. The results indicate that Neo1 mRNA is widely expressed in both male and female mouse tissues with the highest signals detected in the reproductive system. Moreover, Hjv and Neo1 mRNAs are simultaneously expressed in skeletal muscle, heart, esophagus, and liver.     

Ruk is ubiquitinated but not degraded by the proteasome.

The regulator of ubiquitous kinase (Ruk) protein, also known as CIN85 or SETA, is an adaptor-type protein belonging to the CD2AP/CMS family. It was found in complexes with many signaling proteins, including phosphoinositol (PtdIns) 3-kinase (EC 2.7.1.137), Cbl, GRB2, p130Cas and Crk. Functional analysis of these interactions, implicated Ruk in the regulation of apoptosis, receptor endocytosis and cytoskeletal rearrangements. We have recently demonstrated that overexpression of Ruk induces apoptotic death in neurons, which could be reversed by activated forms of PtdIns 3-kinase and PKB/Akt. Furthermore, Ruk was shown to be a negative regulator of PtdIns 3-kinase activity through binding to its P85 regulatory subunit [Gout, I., Middleton, G., Adu, J., Ninkina, N. N., Drobot, L. B., Filonenko, V., Matsuka, G., Davies, A.M., Waterfield, M. & Buchman, V. L. (2000) Embo J.19, 4015-4025]. Here, we report for the first time, that all three isoforms of Ruk (L, M and S) are ubiquitinated. Specific interaction between the E3 ubiquitin ligase Cbl and all three Ruk isoforms was demonstrated by coexpression studies in Hek293 cells. The interaction of Ruk M and S isoforms with Cbl was found to be mediated via heterodimerization with Ruk L. The use of proteosomal and lysosomal inhibitors clearly indicated that ubiquitination of Ruk L does not lead to its degradation. Based on this study, we propose a possible mechanism for the regulation of Ruk function by ubiquitination.     

Comparison of the interactions of transferrin receptor and transferrin receptor 2 with transferrin and the hereditary hemochromatosis protein HFE

The transferrin receptor (TfR) interacts with two proteins important for iron metabolism, transferrin (Tf) and HFE, the protein mutated in hereditary hemochromatosis. A second receptor for Tf, TfR2, was recently identified and found to be functional for iron uptake in transfected cells (Kawabata, H., Germain, R. S., Vuong, P. T., Nakamaki, T., Said, J. W., and Koeffler, H. P. (2000) J. Biol. Chem. 275, 16618-16625). TfR2 has a pattern of expression and regulation that is distinct from TfR, and mutations in TfR2 have been recognized as the cause of a non-HFE linked form of hemochromatosis (Camaschella, C., Roetto, A., Cali, A., De Gobbi, M., Garozzo, G., Carella, M., Majorano, N., Totaro, A., and Gasparini, P. (2000) Nat. Genet. 25, 14-15). To investigate the relationship between TfR, TfR2, Tf, and HFE, we performed a series of binding experiments using soluble forms of these proteins. We find no detectable binding between TfR2 and HFE by co-immunoprecipitation or using a surface plasmon resonance-based assay. The affinity of TfR2 for iron-loaded Tf was determined to be 27 nm, 25-fold lower than the affinity of TfR for Tf. These results imply that HFE regulates Tf-mediated iron uptake only from the classical TfR and that TfR2 does not compete for HFE binding in cells expressing both forms of TfR.     

The ubiquitin-homology protein, DAP-1, associates with tumor necrosis factor receptor (p60) death domain and induces apoptosis

The tumor necrosis factor receptor, p60 (TNF-R1), transduces death signals via the association of its cytoplasmic domain with several intracellular proteins. By screening a mammalian cDNA library using the yeast two-hybrid cloning technique, we isolated a ubiquitin-homology protein, DAP-1, which specifically interacts with the cytoplasmic death domain of TNF-R1. Sequence analysis reveals that DAP-1 shares striking sequence homology with the yeast SMT3 protein that is essential for the maintenance of chromosome integrity during mitosis (Meluh, P. B., and Koshland, D. (1995) Mol. Biol. Cell 6, 793-807). DAP-1 is nearly identical to PIC1, a protein that interacts with the PML tumor suppressor implicated in acute promyelocytic leukemia (Boddy, M. N., Howe, K., Etkin, L. D., Solomon, E., and Freemont, P. S. (1996) Oncogene 13, 971-982), and the sentrin protein, which associates with the Fas death receptor (Okura, T., Gong, L., Kamitani, T., Wada, T., Okura, I., Wei, C. F., Chang, H. M., and Yeh, E. T. (1996) J. Immunol. 157, 4277-4281). The in vivo interaction between DAP-1 and TNF-R1 was further confirmed in mammalian cells. In transient transfection assays, overexpression of DAP-1 suppresses NF-kappaB/Rel activity in 293T cells, a human kidney embryonic carcinoma cell line. Overexpression of either DAP-1 or sentrin causes apoptosis of TNF-sensitive L929 fibroblast cell line, as well as TNF-resistant osteosarcoma cell line, U2OS. Furthermore, the dominant negative Fas-associated death domain protein (FADD) protein blocks the cell death induced by either DAP-1 or FADD. Collectively, these observations highly suggest a role for DAP-1 in mediating TNF-induced cell death signaling pathways, presumably through the recruitment of FADD death effector.     

Selective regulation of endogenous G protein-coupled receptors by arrestins in HEK293 cells

Arrestins play an important role in regulating desensitization and trafficking of G protein-coupled receptors (GPCRs). However, limited insight into the specificity of arrestin-mediated regulation of GPCRs is currently available. Recently, we used an antisense strategy to reduce arrestin levels in HEK293 cells and characterize the role of arrestins on endogenous G(s)-coupled receptors (Mundell, S. J., Loudon, R. B., and Benovic, J. L. (1999) Biochemistry 38, 8723-8732). Here, we characterized GPCRs coupled to either G(q) (M(1) muscarinic acetylcholine receptor (M(1)AchR) and P2y(1) and P2y(2) purinergic receptors) or G(i) (somatostatin and AT1 angiotensin receptors) in wild type and arrestin antisense HEK293 cells. The agonist-specific desensitization of the M(1)Ach and somatostatin receptors was significantly attenuated in antisense-expressing cells, whereas desensitization of P2y(1) and P2y(2) purinergic and AT1 angiotensin receptors was unaffected by reduced arrestin levels. To further examine arrestin/GPCR specificity, we studied the effects of endogenous GPCR activation on the redistribution of arrestin-2 epitope tagged with the green fluorescent protein (arrestin-2-GFP). These studies revealed a receptor-specific movement of arrestin-2-GFP that mirrored the arrestin-receptor specificity observed in the antisense cells. Thus, agonist-induced activation of endogenous beta(2)-adrenergic, prostaglandin E(2), M(1)Ach, and somatostatin receptors induced arrestin-2-GFP redistribution to early endosomes, whereas P2y(1) and P2y(2) purinergic and AT1 angiotensin receptor activation did not. Thus, endogenous arrestins mediate the regulation of selective G(q)- and G(i)-coupled receptors in HEK293 cells.     

RNA沉默抑制子p19调控细胞周期相关蛋白表达

番茄丛矮病毒的P19蛋白不仅是一个重要的病毒致病因子,而且还可作为RNA干扰(RNAi)的抑制子．这种作用是通过限制细胞内的小RNA,比如小干扰RNA(siRNAs)和微RNA(miRNAs)来实现．但是目前对P19蛋白在哺乳动物细胞上的作用还未见报道．构建了一株p19稳定表达的293细胞系,即293-p19．流式细胞仪分析发现在293细胞中过量表达P19蛋白可显著引发细胞周期的G2/M阻滞．细胞增殖实验显示,293-p19细胞的DNA复制及细胞生长均受到显著的抑制． 此外,研究还发现p19可使人胚肾293细胞内的细胞周期调控子的表达谱发生改变． 其中包括上调cyclin A1,CDK2,CDK4,CDK6,p18,cyclin D2,p19INK4d和E2F1,及下调p15,cyclin A,cyclin B1和cyclin E1的表达．上述研究结果提示,p19有可能靶向多个G2/M调控蛋白从而引发细胞的G2/M阻滞．     

Hfe acts in hepatocytes to prevent hemochromatosis

Hereditary hemochromatosis (HH) is a prevalent, potentially fatal disorder of iron metabolism hallmarked by intestinal hyperabsorption of iron, hyperferremia, and hepatic iron overload. In both humans and mice, type I HH is associated with mutations in the broadly expressed HFE/Hfe gene. To identify where Hfe acts to prevent HH, we generated mice with tissue-specific Hfe ablations. This work demonstrates that local Hfe expression in hepatocytes serves to maintain physiological iron homeostasis, answering a long-standing question in medicine and explaining earlier clinical observations.     

Regulation of type II transmembrane serine proteinase TMPRSS6 by hypoxia-inducible factors: new link between hypoxia signaling and iron homeostasis

Hepcidin is a liver-derived hormone with a key role in iron homeostasis. In addition to iron, it is regulated by inflammation and hypoxia, although mechanisms of hypoxic regulation remain unclear. In hepatocytes, hepcidin is induced by bone morphogenetic proteins (BMPs) through a receptor complex requiring hemojuvelin (HJV) as a co-receptor. Type II transmembrane serine proteinase (TMPRSS6) antagonizes hepcidin induction by BMPs by cleaving HJV from the cell membrane. Inactivating mutations in TMPRSS6 lead to elevated hepcidin levels and consequent iron deficiency anemia. Here we demonstrate that TMPRSS6 is up-regulated in hepatic cell lines by hypoxia and by other activators of hypoxia-inducible factor (HIF). We show that TMPRSS6 expression is regulated by both HIF-1α and HIF-2α. This HIF-dependent up-regulation of TMPRSS6 increases membrane HJV shedding and decreases hepcidin promoter responsiveness to BMP signaling in hepatocytes. Our results reveal a potential role for TMPRSS6 in hepcidin regulation by hypoxia and provide a new molecular link between oxygen sensing and iron homeostasis.