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Regulation of prohepcidin processing and activity by the subtilisin-like proprotein convertases Furin, PC5, PACE4 and PC7
  1. N Scamuffa1,
  2. A Basak2,
  3. C Lalou1,
  4. A Wargnier3,
  5. J Marcinkiewicz4,
  6. G Siegfried5,
  7. M Chrétien2,
  8. F Calvo1,
  9. N G Seidah4,
  10. A-M Khatib1
  1. 1
    INSERM, u 716, Equipe Avenir, Labelisée La Ligue, IGM, Université Paris 7, Paris, France
  2. 2
    Ottawa Health Research Institute, Ottawa, Canada
  3. 3
    Service de microbiologie, Hopital Saint-Louis, Paris, France
  4. 4
    Clinical Research Institute of Montreal, Montreal, Canada
  5. 5
    INSERM u 770, Faculté de Médecine, Université Paris-Su, Paris, France
  1. Dr A-M Khatib, Laboratoire de Pharmacologie Expérimentale et Clinique, INSERM U 716/Equipe AVENIR, Institut de Génétique Moléculaire, 27 rue Juliette Dodu, 75010 Paris, France; Majid.Khatib{at}inserm.fr

Abstract

Background and aims: Hepcidin is an iron homoeostasis regulator peptide. Loss-of-function mutations cause juvenile haemochromatosis while its over-expression results in anaemia. However, the mechanism and function of preprohepcidin conversion to mature hepcidins (25, 22 and 20 amino acid C-terminal peptides) are not well known. After removal of the signal peptide, the first proteolytic cleavage occurs within the basic motif RRRRR59DT, suggesting the involvement of proprotein convertase (PC) family members in this process.

Methods and results: Using cell transfection experiments, the processing of preprohepcidin in the human hepatocyte line Huh-7 was found to be inhibited by the Furin inhibitors serpin α1-antitrypsin (α1-PDX) and prosegment preproFurin (ppFurin). Site-directed mutagenesis analysis confirmed the RRRRR59DT preprohepcidin cleavage site. In parallel, the lack of preprohepcidin processing found in the PC activity-deficient cell line LoVo was restored by the expression of Furin, paired basic amino acid cleaving enzyme 4 (PACE4), PC5 or PC7. This finding is consistent with the in vitro digestions of a synthetic peptide mimicking the cleavage site of preprohepcidin. In addition, during mouse embryonic development the major expression of hepcidin found in the liver coincided with that of Furin. While hepcidin induces the degradation of the iron transporter ferroportin, its RRRRR59 to SSSSS59 mutant is not active.

Conclusions: These results demonstrate the key role of the convertases Furin, PACE4, PC5 and/or PC7 in the generation and secretion of active hepcidin and suggest that the control of hepcidin processing as a potential therapeutic/diagnostic strategy in hepcidin-related disorders such as haemochromatosis, inflammatory diseases, anaemia and cancer.

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Footnotes

  • Funding: This work was supported by grants from FRSQ-INSERM cooperation, Associaton pour la recherche sur le cancer (ARC) and La Ligue Contre le Cancer, Paris, France to A-MK.

  • Competing interests: None.

  • Ethics approval: All the experiments performed in this study were approved by the Clinical Research Institute of Montreal Animal Ethics Committee in July 2007.

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