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Identification of TMPRSS6 cleavage sites of hemojuvelin
Hemojuvelin (HJV), the coreceptor of the BMP-SMAD pathway that up-regulates hepcidin transcription, is a repulsive guidance molecule (RGMc) which undergoes a complex intracellular processing. Following autoproteolysis, it is exported to the cell surface both as a full-length and a heterodimeric prot...
Autores principales: | , , , , , , , |
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Formato: | Online Artículo Texto |
Lenguaje: | English |
Publicado: |
BlackWell Publishing Ltd
2015
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Materias: | |
Acceso en línea: | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4395201/ https://www.ncbi.nlm.nih.gov/pubmed/25704252 http://dx.doi.org/10.1111/jcmm.12462 |
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author | Rausa, Marco Ghitti, Michela Pagani, Alessia Nai, Antonella Campanella, Alessandro Musco, Giovanna Camaschella, Clara Silvestri, Laura |
author_facet | Rausa, Marco Ghitti, Michela Pagani, Alessia Nai, Antonella Campanella, Alessandro Musco, Giovanna Camaschella, Clara Silvestri, Laura |
author_sort | Rausa, Marco |
collection | PubMed |
description | Hemojuvelin (HJV), the coreceptor of the BMP-SMAD pathway that up-regulates hepcidin transcription, is a repulsive guidance molecule (RGMc) which undergoes a complex intracellular processing. Following autoproteolysis, it is exported to the cell surface both as a full-length and a heterodimeric protein. In vitro membrane HJV (m-HJV) is cleaved by the transmembrane serine protease TMPRSS6 to attenuate signalling and to inhibit hepcidin expression. In this study, we investigated the number and position of HJV cleavage sites by mutagenizing arginine residues (R), potential TMPRSS6 targets, to alanine (A). We analysed translation and membrane expression of HJV R mutants and the pattern of fragments they release in the culture media in the presence of TMPRSS6. Abnormal fragments were observed for mutants at arginine 121, 176, 218, 288 and 326. Considering that all variants, except HJV(R121A), lack autoproteolytic activity and some (HJV(R176A) and HJV(R288A)) are expressed at reduced levels on cell surface, we identified the fragments originating from either full-length or heterodimeric proteins and defined the residues 121 and 326 as the TMPRSS6 cleavage sites in both isoforms. Using the N-terminal FLAG-tagged HJV, we showed that residue 121 is critical also in the rearrangement of the N-terminal heterodimeric HJV. Exploiting the recently reported RGMb crystallographic structure, we generated a model of HJV that was used as input structure for all-atoms molecular dynamics simulation in explicit solvent. As assessed by in silico studies, we concluded that some arginines in the von Willebrand domain appear TMPRSS6 insensitive, likely because of partial protein structure destabilization. |
format | Online Article Text |
id | pubmed-4395201 |
institution | National Center for Biotechnology Information |
language | English |
publishDate | 2015 |
publisher | BlackWell Publishing Ltd |
record_format | MEDLINE/PubMed |
spelling | pubmed-43952012015-04-20 Identification of TMPRSS6 cleavage sites of hemojuvelin Rausa, Marco Ghitti, Michela Pagani, Alessia Nai, Antonella Campanella, Alessandro Musco, Giovanna Camaschella, Clara Silvestri, Laura J Cell Mol Med Original Articles Hemojuvelin (HJV), the coreceptor of the BMP-SMAD pathway that up-regulates hepcidin transcription, is a repulsive guidance molecule (RGMc) which undergoes a complex intracellular processing. Following autoproteolysis, it is exported to the cell surface both as a full-length and a heterodimeric protein. In vitro membrane HJV (m-HJV) is cleaved by the transmembrane serine protease TMPRSS6 to attenuate signalling and to inhibit hepcidin expression. In this study, we investigated the number and position of HJV cleavage sites by mutagenizing arginine residues (R), potential TMPRSS6 targets, to alanine (A). We analysed translation and membrane expression of HJV R mutants and the pattern of fragments they release in the culture media in the presence of TMPRSS6. Abnormal fragments were observed for mutants at arginine 121, 176, 218, 288 and 326. Considering that all variants, except HJV(R121A), lack autoproteolytic activity and some (HJV(R176A) and HJV(R288A)) are expressed at reduced levels on cell surface, we identified the fragments originating from either full-length or heterodimeric proteins and defined the residues 121 and 326 as the TMPRSS6 cleavage sites in both isoforms. Using the N-terminal FLAG-tagged HJV, we showed that residue 121 is critical also in the rearrangement of the N-terminal heterodimeric HJV. Exploiting the recently reported RGMb crystallographic structure, we generated a model of HJV that was used as input structure for all-atoms molecular dynamics simulation in explicit solvent. As assessed by in silico studies, we concluded that some arginines in the von Willebrand domain appear TMPRSS6 insensitive, likely because of partial protein structure destabilization. BlackWell Publishing Ltd 2015-04 2015-02-22 /pmc/articles/PMC4395201/ /pubmed/25704252 http://dx.doi.org/10.1111/jcmm.12462 Text en © 2015 The Authors. Journal of Cellular and Molecular Medicine published by John Wiley & Sons Ltd and Foundation for Cellular and Molecular Medicine. http://creativecommons.org/licenses/by/4.0/ This is an open access article under the terms of the Creative Commons Attribution License, which permits use, distribution and reproduction in any medium, provided the original work is properly cited. |
spellingShingle | Original Articles Rausa, Marco Ghitti, Michela Pagani, Alessia Nai, Antonella Campanella, Alessandro Musco, Giovanna Camaschella, Clara Silvestri, Laura Identification of TMPRSS6 cleavage sites of hemojuvelin |
title | Identification of TMPRSS6 cleavage sites of hemojuvelin |
title_full | Identification of TMPRSS6 cleavage sites of hemojuvelin |
title_fullStr | Identification of TMPRSS6 cleavage sites of hemojuvelin |
title_full_unstemmed | Identification of TMPRSS6 cleavage sites of hemojuvelin |
title_short | Identification of TMPRSS6 cleavage sites of hemojuvelin |
title_sort | identification of tmprss6 cleavage sites of hemojuvelin |
topic | Original Articles |
url | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4395201/ https://www.ncbi.nlm.nih.gov/pubmed/25704252 http://dx.doi.org/10.1111/jcmm.12462 |
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