The hemochromatosis proteins HFE, TfR2, and HJV form a membrane-associated protein complex for hepcidin regulation.
OBJECTIVE
The hereditary hemochromatosis-associated membrane proteins HFE, TfR2, and HJV are required for adequate hepatic expression of the iron hormone hepcidin. While the genetic interactions are clear, it remains elusive how bone morphogenetic protein co-receptor HJV functions together with HFE and TfR2 to activate hepcidin transcription via the BMP-SMAD signaling pathway. Here, we investigate whether HFE, TfR2, and HJV physically interact on the surface of hepatocytes.


METHODS
We explore protein-protein interactions by glycerol gradient sedimentation assays and co-immunoprecipitation analyses in transfected HuH7 hepatoma-derived cells.


RESULTS
Our data demonstrate that HFE and TfR2 bind HJV in a non-competitive manner. Co-immunoprecipitation analyses provide direct experimental evidence that HFE, TfR2, and HJV form a multi-protein membrane complex. Our experiments show that like TfR2, HJV competes with TfR1 for binding to HFE, indicating that the expression of TfR2 and HJV may be critical for iron sensing. We identify residues 120-139 of the TfR2 extra-cellular domain as the critical amino acids required for the binding of both HFE and HJV. Interestingly, RGMA, a central nervous system homolog, can substitute for HJV in the complex and promote hepcidin transcription, implicating RGMA in the local control of hepcidin in the CNS.


CONCLUSIONS
Taken together, our findings provide a biochemical basis for hepcidin control by HFE, TfR2, and HJV.
University of Heidelberg, Department of Pediatric Oncology, Hematology and Immunology, Heidelberg, Germany.
Flavia
F
D'Alessio
Matthias W
MW
Hentze
Martina U
MU
Muckenthaler
D000596:Amino Acids / Q000378:metabolism; D023181:Antimicrobial Cationic Peptides / Q000378:metabolism*; D006528:Carcinoma, Hepatocellular / Q000378:metabolism / Q000473:pathology; D045744:Cell Line, Tumor; D002462:Cell Membrane / Q000378:metabolism* / Q000473:pathology; D002490:Central Nervous System / Q000502:physiology; D058851:GPI-Linked Proteins / Q000378:metabolism; D006432:Hemochromatosis / Q000378:metabolism; D000071020:Hemochromatosis Protein; D064451:Hepcidins; D015395:Histocompatibility Antigens Class I / Q000378:metabolism*; D006801:Humans; D008113:Liver Neoplasms / Q000378:metabolism / Q000473:pathology; D008565:Membrane Proteins / Q000378:metabolism*; D046912:Multiprotein Complexes / Q000378:metabolism*; D009419:Nerve Tissue Proteins / Q000378:metabolism; D011485:Protein Binding / Q000502:physiology; D011990:Receptors, Transferrin / Q000378:metabolism*; D015398:Signal Transduction / Q000502:physiology
D000596:Amino Acids; D023181:Antimicrobial Cationic Peptides; D058851:GPI-Linked Proteins; C578702:HAMP protein, human; C103673:HFE protein, human; D000071020:Hemochromatosis Protein; D064451:Hepcidins; D015395:Histocompatibility Antigens Class I; D008565:Membrane Proteins; D046912:Multiprotein Complexes; D009419:Nerve Tissue Proteins; C522183:RGMA protein, human; D011990:Receptors, Transferrin; C503640:TFR2 protein, human