In vivo analysis of the role of O-glycosylations of von Willebrand factor.

The objective of this project was to study the function of O-glycosylations in von Willebrand factor (VWF) life cycle. In total, 14 different murine Vwf cDNAs mutated on one or several O-glycosylations sites were generated: 9 individual mutants, 2 doublets, 2 clusters and 1 mutant with all 9 murine...

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Autores principales:	Idinath Badirou, Mohamad Kurdi, Paulette Legendre, Julie Rayes, Marijke Bryckaert, Caterina Casari, Peter J Lenting, Olivier D Christophe, Cecile V Denis
Formato:	article
Lenguaje:	EN
Publicado:	Public Library of Science (PLoS) 2012
Materias:	Medicine R Science Q
Acceso en línea:	https://doaj.org/article/80df049f5c7640ac911c0da116a643b6
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spelling	oai:doaj.org-article:80df049f5c7640ac911c0da116a643b62021-11-18T07:18:15ZIn vivo analysis of the role of O-glycosylations of von Willebrand factor.1932-620310.1371/journal.pone.0037508https://doaj.org/article/80df049f5c7640ac911c0da116a643b62012-01-01T00:00:00Zhttps://www.ncbi.nlm.nih.gov/pmc/articles/pmid/22616016/pdf/?tool=EBIhttps://doaj.org/toc/1932-6203The objective of this project was to study the function of O-glycosylations in von Willebrand factor (VWF) life cycle. In total, 14 different murine Vwf cDNAs mutated on one or several O-glycosylations sites were generated: 9 individual mutants, 2 doublets, 2 clusters and 1 mutant with all 9 murine glycosylation sites mutated (Del-O-Gly). We expressed each mutated cDNA in VWF deficient-mice by hydrodynamic injection. An immunosorbent assay with Peanut Agglutinin (PNA) was used to verify the O-glycosylation status. Wild-type (WT) VWF expressed by hepatocytes after hydrodynamic injection was able to bind PNA with slightly higher affinity than endothelial-derived VWF. In contrast, the Del-O-Gly VWF mutant did not bind PNA, demonstrating removal of O-linked glycans. All mutants displayed a normal multimeric pattern. Two mutants, Del-O-Gly and T1255A/T1256A, led to expression levels 50% lower than those induced by WT VWF and their half-life in vivo was significantly reduced. When testing the capacity of each mutant to correct the bleeding time of VWF-deficient mice, we found that S1486A, T1255A, T1256A and the doublet T1255A/T1256A were unable to do so. In conclusion we have shown that O-glycosylations are dispensable for normal VWF multimerization and biosynthesis. It also appears that some O-glycosylation sites, particularly the T1255 and T1256 residues, are involved in the maintenance of VWF plasma levels and are essential for normal haemostasis. As for the S1486 residue, it seems to be important for platelet binding as demonstrated in vitro using perfusion experiments.Idinath BadirouMohamad KurdiPaulette LegendreJulie RayesMarijke BryckaertCaterina CasariPeter J LentingOlivier D ChristopheCecile V DenisPublic Library of Science (PLoS)articleMedicineRScienceQENPLoS ONE, Vol 7, Iss 5, p e37508 (2012)
institution	DOAJ
collection	DOAJ
language	EN
topic	Medicine R Science Q
spellingShingle	Medicine R Science Q Idinath Badirou Mohamad Kurdi Paulette Legendre Julie Rayes Marijke Bryckaert Caterina Casari Peter J Lenting Olivier D Christophe Cecile V Denis In vivo analysis of the role of O-glycosylations of von Willebrand factor.
description	The objective of this project was to study the function of O-glycosylations in von Willebrand factor (VWF) life cycle. In total, 14 different murine Vwf cDNAs mutated on one or several O-glycosylations sites were generated: 9 individual mutants, 2 doublets, 2 clusters and 1 mutant with all 9 murine glycosylation sites mutated (Del-O-Gly). We expressed each mutated cDNA in VWF deficient-mice by hydrodynamic injection. An immunosorbent assay with Peanut Agglutinin (PNA) was used to verify the O-glycosylation status. Wild-type (WT) VWF expressed by hepatocytes after hydrodynamic injection was able to bind PNA with slightly higher affinity than endothelial-derived VWF. In contrast, the Del-O-Gly VWF mutant did not bind PNA, demonstrating removal of O-linked glycans. All mutants displayed a normal multimeric pattern. Two mutants, Del-O-Gly and T1255A/T1256A, led to expression levels 50% lower than those induced by WT VWF and their half-life in vivo was significantly reduced. When testing the capacity of each mutant to correct the bleeding time of VWF-deficient mice, we found that S1486A, T1255A, T1256A and the doublet T1255A/T1256A were unable to do so. In conclusion we have shown that O-glycosylations are dispensable for normal VWF multimerization and biosynthesis. It also appears that some O-glycosylation sites, particularly the T1255 and T1256 residues, are involved in the maintenance of VWF plasma levels and are essential for normal haemostasis. As for the S1486 residue, it seems to be important for platelet binding as demonstrated in vitro using perfusion experiments.
format	article
author	Idinath Badirou Mohamad Kurdi Paulette Legendre Julie Rayes Marijke Bryckaert Caterina Casari Peter J Lenting Olivier D Christophe Cecile V Denis
author_facet	Idinath Badirou Mohamad Kurdi Paulette Legendre Julie Rayes Marijke Bryckaert Caterina Casari Peter J Lenting Olivier D Christophe Cecile V Denis
author_sort	Idinath Badirou
title	In vivo analysis of the role of O-glycosylations of von Willebrand factor.
title_short	In vivo analysis of the role of O-glycosylations of von Willebrand factor.
title_full	In vivo analysis of the role of O-glycosylations of von Willebrand factor.
title_fullStr	In vivo analysis of the role of O-glycosylations of von Willebrand factor.
title_full_unstemmed	In vivo analysis of the role of O-glycosylations of von Willebrand factor.
title_sort	in vivo analysis of the role of o-glycosylations of von willebrand factor.
publisher	Public Library of Science (PLoS)
publishDate	2012
url	https://doaj.org/article/80df049f5c7640ac911c0da116a643b6
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In vivo analysis of the role of O-glycosylations of von Willebrand factor.

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