title: N-linked glycans within the A2 domain of von Willebrand factor modulate macrophage-mediated clearance.
abstract: Enhanced von Willebrand factor (VWF) clearance is important in the etiology of von Willebrand disease. However, the molecular mechanisms underlying VWF clearance remain poorly understood. In this study, we investigated the role of VWF domains and specific glycan moieties in regulating in vivo clearance. Our findings demonstrate that the A1 domain of VWF contains a receptor-recognition site that plays a key role in regulating the interaction of VWF with macrophages. In A1-A2-A3 and full-length VWF, this macrophage-binding site is cryptic but becomes exposed following exposure to shear or ristocetin. Previous studies have demonstrated that the N-linked glycans within the A2 domain play an important role in modulating susceptibility to ADAMTS13 proteolysis. We further demonstrate that these glycans presented at N1515 and N1574 also play a critical role in protecting VWF against macrophage binding and clearance. Indeed, loss of the N-glycan at N1515 resulted in markedly enhanced VWF clearance that was significantly faster than that observed with any previously described VWF mutations. In addition, A1-A2-A3 fragments containing the N1515Q or N1574Q substitutions also demonstrated significantly enhanced clearance. Importantly, clodronate-induced macrophage depletion significantly attenuated the increased clearance observed with N1515Q and N1574Q in both full-length VWF and A1-A2-A3. Finally, we further demonstrate that loss of these N-linked glycans does not enhance clearance in VWF in the presence of a structurally constrained A2 domain. Collectively, these novel findings support the hypothesis that conformation of the VWF A domains plays a critical role in modulating macrophage-mediated clearance of VWF in vivo.
authors.affiliation: Haemostasis Research Group, Institute of Molecular Medicine, Trinity Centre for Health Sciences, St. James's Hospital.
authors.forename: Alain
authors.initials: A
authors.lastname: Chion
authors.affiliation: Haemostasis Research Group, Institute of Molecular Medicine, Trinity Centre for Health Sciences, St. James's Hospital.
authors.forename: Jamie M
authors.initials: JM
authors.lastname: O'Sullivan
authors.affiliation: Haemostasis Research Group, Institute of Molecular Medicine, Trinity Centre for Health Sciences, St. James's Hospital.
authors.forename: Clive
authors.initials: C
authors.lastname: Drakeford
authors.affiliation: Haemostasis Research Group, Institute of Molecular Medicine, Trinity Centre for Health Sciences, St. James's Hospital.
authors.forename: Gudmundur
authors.initials: G
authors.lastname: Bergsson
authors.affiliation: Haemostasis Research Group, Institute of Molecular Medicine, Trinity Centre for Health Sciences, St. James's Hospital.
authors.forename: Niall
authors.initials: N
authors.lastname: Dalton
authors.affiliation: Haemostasis Research Group, Institute of Molecular Medicine, Trinity Centre for Health Sciences, St. James's Hospital.
authors.forename: Sonia
authors.initials: S
authors.lastname: Aguila
authors.affiliation: Haemostasis Research Group, Institute of Molecular Medicine, Trinity Centre for Health Sciences, St. James's Hospital.
authors.forename: Soracha
authors.initials: S
authors.lastname: Ward
authors.affiliation: Inflammation and Immunity Research Group, Institute of Molecular Medicine, and.
authors.forename: Padraic G
authors.initials: PG
authors.lastname: Fallon
authors.affiliation: Haemostasis Research Group, Institute of Molecular Medicine, Trinity Centre for Health Sciences, St. James's Hospital.
authors.forename: Teresa M
authors.initials: TM
authors.lastname: Brophy
authors.affiliation: Department of Clinical Medicine, School of Medicine, Trinity College Dublin, Dublin, Ireland; National Children's Research Centre, Our Lady's Children's Hospital, Crumlin, Dublin, Ireland.
authors.forename: Roger J S
authors.initials: RJ
authors.lastname: Preston
authors.affiliation: Department of Histopathology, Sir Patrick Dun Research Laboratory, St. James's Hospital, Trinity College Dublin, Dublin, Ireland.
authors.forename: Lauren
authors.initials: L
authors.lastname: Brady
authors.affiliation: Department of Histopathology, Sir Patrick Dun Research Laboratory, St. James's Hospital, Trinity College Dublin, Dublin, Ireland.
authors.forename: Orla
authors.initials: O
authors.lastname: Sheils
authors.affiliation: Faculty of Medicine, Imperial College, Hammersmith Hospital, London, United Kingdom.
authors.forename: Michael
authors.initials: M
authors.lastname: Laffan
authors.affiliation: Faculty of Medicine, Imperial College, Hammersmith Hospital, London, United Kingdom.
authors.forename: Thomas A J
authors.initials: TA
authors.lastname: McKinnon
authors.affiliation: Haemostasis Research Group, Institute of Molecular Medicine, Trinity Centre for Health Sciences, St. James's Hospital, National Centre for Hereditary Coagulation Disorders, St. James's Hospital, Dublin, Ireland; and Irish Centre for Vascular Biology, Royal College of Surgeons in Ireland, Dublin, Ireland.
authors.forename: James S
authors.identifier: 0000-0003-0309-3313
authors.initials: JS
authors.lastname: O'Donnell
mesh_terms: D019943:Amino Acid Substitution; D000818:Animals; D045744:Cell Line, Tumor; D006801:Humans; D008264:Macrophages / Q000166:cytology / Q000378:metabolism*; D051379:Mice; D018345:Mice, Knockout; D020125:Mutation, Missense; D011134:Polysaccharides / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D000072417:Protein Domains; D014841:von Willebrand Factor / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*
chemical_list: D011134:Polysaccharides; D014841:von Willebrand Factor