The C-type lectin receptor CLEC4M binds, internalizes, and clears von Willebrand factor and contributes to the variation in plasma von Willebrand factor levels

Abstract

Key Points• CLEC4M plays a role in the clearance of VWF.• CLEC4M polymorphisms contribute to the genetic variability of VWF plasma levels.Genetic variation in or near the C-type lectin domain family 4 member M (CLEC4M) has been associated with plasma levels of von Willebrand factor (VWF) in healthy individuals. CLEC4M is a lectin receptor with a polymorphic extracellular neck region possessing a variable number of tandem repeats (VNTR). A total of 491 participants (318 patients with type 1 von Willebrand disease [VWD] and 173 unaffected family members) were genotyped for the CLEC4M VNTR polymorphism. Family-based association analysis on kindreds with type 1 VWD demonstrated an excess transmission of VNTR 6 to unaffected individuals (P 5 .0096) and an association of this allele with increased VWF:RCo (P 5 .029). CLEC4M-Fc bound to VWF. Immunofluorescence and enzyme-linked immunosorbent assay demonstrated that HEK 293 cells transfected with CLEC4M bound and internalized VWF. Cells expressing 4 or 9 copies of the CLEC4M neck region VNTR showed reduced interaction with VWF relative to CLEC4M with 7 VNTR (CLEC4M 4%-60% reduction, P < .001; CLEC4M 9%-45% reduction, P 5 .006). Mice expressing CLEC4M after hydrodynamic liver transfer have a 46% decrease in plasma levels of VWF (P 5 .0094). CLEC4M binds to and internalizes VWF, and polymorphisms in the CLEC4M gene contribute to variable plasma levels of VWF. (Blood. 2013;121(26):5228-5237) Introduction von Willebrand factor (VWF) is a plasma glycoprotein that mediates platelet adhesion and aggregation and acts as the carrier protein for factor VIII (FVIII). VWF synthesis by endothelial cells 1 and megakaryocytes 2 involves complex post-translational modifications including dimerization, glycosylation, sulfation, multimerization, and propeptide cleavage (reviewed by Sadler 3 ). The protein is either constitutively secreted into the plasma and subendothelium or is stored in endothelial Weibel-Palade bodies or platelet a granules from which release can be mediated by a number of chemical and biomechanical stimuli.Plasma VWF levels in healthy participants show a fourfold range (0.50-2.00 IU/mL). 4 These levels are influenced by a variety of genetic and acquired factors. ABO blood group contributes approximately 30% of the genetic influence, 5,6 whereas age, 6,7 acute-phase stimuli, [8][9][10] and several endocrine abnormalities represent acquired determinants of VWF levels. 7,11 In type 1 von Willebrand disease (VWD), which is defined as a partial deficiency of functionally normal VWF, approximately 35% of individuals do not have a putative mutation in the coding region, splice junctions, or proximal promoter of the VWF gene, suggesting that genes other than VWF may contribute to the pathophysiology of this disease. 12,13 VWF circulates in a tight, noncovalently linked complex with FVIII. The mean circulating half-lives of VWF and FVIII are 12 to 18 hours and12 hours, respectively, but details of the fate of both proteins are minimal. Evidence exists that cells in the li...

von Willebrand factor binds to the surface of dendritic cells and modulates peptide presentation of factor VIII

“…Recently, the C-type lectin receptor CLEC4M, the carbohydrate receptor Siglec-5 and scavenger receptor class A member 5 (SCARA5) have been shown to interact with VWF. [30][31][32] Interestingly, transfected HEK293 cells over-expressing CLEC4M or Siglec-5 were shown to efficiently internalize VWF. [30][31][32] The lack of VWF internalization observed in iDCs suggests that CLEC4M, Siglec-5 and SCARA5 are not involved in binding of VWF to dendritic cells.…”

“…VWF specific peptides were identified using Proteome Discoverer and subsequently HLA-DRB1 genotype specific core peptides were predicted using NetMHCpan 2. 30 Our knowledge on the etiology and characteristics of this unwanted immune response remains limited. 36 Genetic studies have shown that allo-antibody development to VWF occurs in patients with deletions, frameshift and nonsense mutations.…”

Life in the shadow of a dominant partner: the FVIII-VWF association and its clinical implications for hemophilia A

“…105 Interestingly, a relationship between the number of tandem repeats in the CLEC4M gene and levels of VWF activity and antigen was observed when analyzing plasma samples from VWD patients. Apparently, sequence variations in VWF receptors may modulate the efficiency by which they remove VWF from circulation.…”

Proteomic identification of Galectin-11 and 14 ligands from Haemonchus contortus

“…The binding of a C-type lectin (CLEC4M), with VWF has previously been shown to enhance the internalisation of VWF by the host cells and alter plasma levels of VWF (Rydz et al 2013). In previous reports, proteins containing the VWF domain are localised in nematode intestine and suggested to play critical roles in cell adhesion and platelet aggregation (Wohner et al 2012).…”