Expression and characterization of von Willebrand factor dimerization defects in different types of von Willebrand disease

Dimerization defects of von Willebrand factor (vWF) protomers underlie von Willebrand disease (vWD) type 2A, subtype IID (vWD 2A/IID), and correspondi ng mutations have been identified at the 3' end of the vWF gene in exon 52, This study identified and expressed 2 additional mutations in this region, a homozygous defect in a patient with vWD type 3 (C2754W) and a heterozygo us frameshift mutation (8566delC) in a patient with vWD type 2A, subtype II E, Both mutations involve cysteine residues that we propose are possibly es sential for dimerization, To prove this hypothesis, transient recombinant e xpression of each of the 2 mutations introduced in the carboxy-terminal vWF fragment II and in the complete vWF complementary DNA, respectively, were carried out in COS-7 cells and compared with expression of vWD 2A/IID mutat ion C2773R and the wild-type (WT) sequence in COS-7 cells. Recombinant WT v WF fragment II assembled correctly into a dimer, whereas recombinant mutant fragments were monomeric, Homozygous expression of recombinant mutant full -length vWF resulted in additional dimers, probably through disulfide bondi ng at the amino-terminal multimerization site, whereas recombinant WT VWF c orrectly assembled into multimers, Coexpression of recombinant mutant and r ecombinant WT vWF reproduced the multimer patterns observed in heterozygous individuals. Our results suggest that a common defect of vWF biosynthesis- lack of vWF dimerization-may cause diverse types and subtypes of vWD, We al so confirmed previous studies that found that disulfide bonding at the vWF amino-terminal is independent of dimerization at the vWF carboxy-terminal.