Linkage analysis in autosomal inherited von Willebrand disease (VWD) is important to diagnose the carriers and reduce the burden of severe type VWD. The study was designed to identify the carriers and estimate the frequency of variable number of tandem repeats (VNTR) instability in VWD families. Carrier detection was performed in eight recessive type 3 VWD (VWD3) families using VNTRs VWF1 and VWF2, RsaI (789Thr/Ala) linkage markers, multimer analysis and DNA sequencing. Moreover, five dominant VWD families were studied through DNA sequencing and multimer analysis. Frequency of VWF VNTR instability was investigated in 20 VWD families. In VWD3 families, a total of 22 (81.5%) carriers were identified using VWF1 and VWF2 markers. However, only 13(48.1%) carriers were identified through RsaI markers. Mutation screening revealed 22(81.5%) carriers in VWD3 and 4 (33.3%) carriers in VWD2 families. In comparison to DNA sequencing, the accuracy of VWF1 and VWF2 markers in VWD3 was 85.7% while RsaI could identify 68.2% carriers accurately. Mutations p.R1205H and p.C1272R were identified as de novo in families. Multimer analysis confirmed the identified carriers in VWD2 families. Three VWD families were found to be carrying VNTR instability for VWF1 and VWF2 locus. VNTRs could be an effective linkage markers for carrier detection in VWD3 families. However, in the event of germline de novo mutations and VNTR instability, it may confound risk of misdiagnosis of carriers. Multimer analysis could be an alternative way of carrier detection in dominant type 2A and type 2B VWD families.
Type 2 von Willebrand disease (VWD) is characterised by qualitative defects in von Willebrand factor (VWF). Exon 28 of the VWF gene is known to be a hot spot for type 2 VWD mutations. The goal of this study was to characterise the mutations in VWF exon 28 and understand the molecular basis of phenotypes through in vitro and in silico studies. Mutation screening was performed in 56 type 2 VWD patients through direct sequencing. Expression vectors for five mutations were transiently expressed in 293-EBNA cells to understand the mutations pathology. Furthermore, in silico structure analysis was performed for 13 missense mutations. A total of 16 including eight novel mutations were detected in 23 (41%) patients. Of these, 15 were missense (including seven V1439M, A1464P, M1495L, I1509V, R1527Q, N1635I and A1647D novel ones) and one was a novel gene conversion. Expression studies and characterisation of recombinant VWF suggested the loss of VWF function for mutants P1266Q, V1439M and N1635I and gain of function for mutant R1308C. No apparent defect was seen in mutant N1231S. In silico structure analysis suggested the probable gain or loss of hydrogen/van der Waals interactions in 10 mutant proteins. In conclusion, type 2A mutations and gene conversion were found to be a common cause of type 2 VWD. Expression studies suggest the mutations N1635I for type 2A(II), P1266Q and V1439M for type 2M, R1308C for type 2B VWD and N1231S as a non-causative variant. Moreover, in silico studies of the mutants show the probable cause of respective phenotypes.
scite is a Brooklyn-based organization that helps researchers better discover and understand research articles through Smart Citations–citations that display the context of the citation and describe whether the article provides supporting or contrasting evidence. scite is used by students and researchers from around the world and is funded in part by the National Science Foundation and the National Institute on Drug Abuse of the National Institutes of Health.
hi@scite.ai
10624 S. Eastern Ave., Ste. A-614
Henderson, NV 89052, USA
Copyright © 2024 scite LLC. All rights reserved.
Made with 💙 for researchers
Part of the Research Solutions Family.