Hepcidin Expression Is Regulated by a Complex of Hemochromatosis-Associated Proteins.

Hemochromatosis is a common genetic disease resulting from increased dietary iron absorption and tissue iron deposition. Mutations in five unrelated genes are known to cause hemochromatosis in humans and mice. These encode the classic hemochromatosis protein (HFE), transferrin receptor 2 (TFR2), the iron exporter ferroportin (FPN), hemojuvelin (HJV), and the circulating anti-microbial peptide hepcidin (HAMP). Hepcidin binds to FPN, causing its internalization and degradation, thus decreasing cellular iron release. A basic understanding of the pathophysiology of FPN and hepcidin mutations has recently been elucidated; however, it was still unclear how mutations in HFE, TFR2, and HJV cause hemochromatosis. All are associated with decreased hepcidin production and inappropriately high levels of ferroportin activity. HFE, TFR2 and HJV are normally expressed in the hepatic cells that produce hepcidin. With collaborators, we showed that HJV acts as a bone morphogenetic protein (BMP) co-receptor. HJV binds to the BMP ligands and forms a complex with Type I BMP receptors, resulting in signaling through a SMAD pathway and induction of hepcidin expression. Disease causing mutations in HJV abrogate BMP co-receptor activity, and hepatocytes from Hjv−/ − mice have a blunted response to BMP2. HFE was known to form a complex with the classical transferrin receptor, TFR1. Several models have been proposed implicating this complex in the regulation of normal iron homeostasis, but they have not taken the role of hepcidin into account. To examine the HFE/TFR1 interaction in vivo, we developed mice expressing a mutant form of TFR1 that should constitutively interact with HFE. We found that these transgenic animals have a phenotype similar to Hfe−/ − mice, suggesting that TFR1 serves to sequester HFE to silence its activity. We next asked whether HFE might also participate in BMP signaling. We found that forced expression of HFE in a hepatoma cell line induces transcription of a reporter gene linked to the hepcidin promoter. It also induces transcription from a heterologous promoter containing BMP-responsive elements, suggesting that HFE works through the BMP pathway. In contrast, forced expression of TFR2 did not amplify expression of either reporter, but it prevented cellular release of a soluble cleavage product of HJV. Furthermore, we showed that both HFE and TFR2 are associated with HJV in a stable protein complex that can be isolated by co-immunoprecipitation or Ni-affinity chromatography. TFR2 appears to aid in the recruitment of HFE to this complex. We conclude that HFE and TFR2 thus serve to amplify BMP signaling through an HJV/BMP receptor pathway. Our findings provide a compelling explanation for the similar clinical hemochromatosis phenotypes resulting from mutations in these genes.