Fattore di von Willebrand
E' una proteina multimerica presente nel plasma fondamentale nell'adesione ed aggregazione delle piastrine nelle prime fasi della formazione del tappo emostatico. La carenza di questa proteina è alla base della malattia di von Willebrand, la patologia dell'emostasi più frequente (incidenza dell'1% nella popolazione mondiale).
Blood. 2010 Dec 9;116(24):5371-6. Epub 2010 Aug 27.
The dominant-negative von Willebrand factor gene deletion p.P1127_C1948delinsR: molecular mechanism and modulation.
Casari C, Pinotti M, Lancellotti S, Adinolfi E, Casonato A, De Cristofaro R, Bernardi F.
Department of Biochemistry and Molecular Biology, University of Ferrara, Ferrara;
Abstract
Understanding molecular mechanisms in the dominant inheritance of von Willebrand disease would improve our knowledge of pathophysiologic processes underlying its prevalence. Cellular models of severe type 2 von Willebrand disease, caused by a heterozygous deletion in the von Willebrand factor (VWF) gene, were produced to investigate the altered biosynthesis. Coexpression of the wild-type and in-frame deleted (p.P1127_C1948delinsR) VWF forms impaired protein secretion, high molecular weight multimer formation and function (VWF collagen-binding 1.9% ± 0.5% of wild-type), which mimicked the patient's phenotype. mRNA, protein, and cellular studies delineated the highly efficient dominant-negative mechanism, based on the key role of heterodimers as multimer terminators. The altered VWF, synthesized in large amounts with the correctly encoded "cysteine knot" domain, formed heterodimers and heterotetramers with wild-type VWF, in addition to deleted homodimers. Impaired multimerization was associated with reduced amounts of VWF in late endosomes. Correction of the dominant-negative effect was explored by siRNAs targeting the mRNA breakpoint, which selectively inhibited the in-frame deleted VWF expression. Although the small amount of the deleted protein synthesized after inhibition still exerted dominant, even though weakened, negative effects, the siRNA treatment restored secretion of large multimers with improved function (VWF collagen-binding 28.0% ± 3.3% of wild-type).
PMID: 20570857 [PubMed - in process]
Br J Haematol. 1998 Dec;103(3):885-7.
Two novel mutations (Pro864His, Val867Glu) causing type 2A von Willebrand disease and affecting a single restriction site in exon 28.
Bernardi F, Casonato A, Marchetti G, Gemmati D, Bizzaro N, Pontara E, Girolami A.
Dipartimento di Biochimica e Biologia Molecolare, Centro Studi Biochimici della Patologie del Genoma Umano, Universita' di Ferrara, Italy.
We detected two transversions in two unrelated Italian patients with type 2A von Willebrand disease (VWD): a C to A at nucleotide 8821 and a T to A at nucleotide 8830, resulting in the missense mutations Pro864His and Val867Glu respectively. Both mutations were in the heterozygous form and abolished the BstXI restriction site in exon 28 of the VWF gene. In both mutations plasma VWF multimer pattern improved by antiproteases. Moreover, DDAVP normalized plasma VWF multimers in the Pro864His patient, especially when protease inhibitors were present. These new mutations appear to be of the 2A VWD subtype due to the increased susceptibility to proteases.
PMID: 9858250 [PubMed - indexed for MEDLINE]
Br J Haematol. 1996 Jan;92(1):241-3.
A novel mutation (Leu817Pro) causing type 2A von Willebrand disease.
Gemmati D, Serino ML, Moratelli S, Ballerini G, Furbetta M, Lunghi B, Marchetti G, Bernardi F.
Centro per lo Studio dell'Emostasi e Trombosi, Università degli Studi di Ferrara, Italy.
We studied a patient affected by von Willebrand disease type 2A who experienced several mild bleeding episodes and was characterized by markedly reduced haemostatic parameters. In the exon 28 of von Willebrand factor (vWF) gene a T to C transition at nucleotide 8680, resulting in the missense mutation Leu817Pro, was found in the heterozygous form in the patient and in two affected relatives. As suggested by the presence in platelets of a complete spectrum of vWF multimers as well as by the increased vWF antigen levels and improved haemostasis after DDAVP treatment, the mutation is compatible with normal multimerization, and could be responsible for a reduced stability or an impaired physiological secretion of vWF.
PMID: 8562403 [PubMed - indexed for MEDLINE]
Hum Genet. 1992 Nov;90(3):297-8.
Characterization and mapping of the 5' portion of von Willebrand factor pseudogene.
Patracchini P, Marchetti G, Aiello V, Croci G, Calzolari E, Bernardi F.
Centro di Studi Biochimici delle Patologie del Genoma Umano, Università di Ferrara, Italy.
A genomic fragment containing the 5' boundary of the von Willebrand factor pseudogene was cloned, partially sequenced and used for in situ hybridization experiments on metaphase spreads from a Philadelphia chromosome (Ph1)-positive chronic myelogenous leukemia patient. Data obtained indicate that the von Willebrand factor pseudogenic region is centromeric to the breakpoint cluster region on 22q11.2. This probe could be used for the study of deletions in the DiGeorge syndrome.
PMID: 1487245 [PubMed - indexed for MEDLINE]
Br J Haematol. 1991 May;78(1):71-9.
Characterization of the pseudogenic and genic homologous regions of von Willebrand factor.
Marchetti G, Patracchini P, Volinia S, Aiello V, Schiavoni M, Ciavarella N, Calzolari E, Schwienbacher C, Bernardi F.
Centro Studi Biochimici delle Patologie del Genoma Umano-Istituto Chimica Biologica, Università di Ferrara, Italy.
The homologous pseudogenic and genic regions of von Willebrand factor (vWF) were studied in DNA from a patient with homozygous deletion of vWF genes and compared with a normal control. This analysis indicates informative restriction patterns for the investigation of restriction fragment length polymorphisms (RFLPs) and gene lesions, and for molecular cloning. A useful new genic XbaI RFLP was found and characterized. A large BgIII fragment of the pseudogenic region was cloned and mapped, and single sequences (9 kb) were used as probes. Corresponding genic and pseudogenic fragments, which contain exons 23-28, and specific restriction patterns were identified, including a new polymorphic TaqI site that was mapped in the gene. A cloned fragment contains the 5' boundary of the pseudogene and recognizes an additional and unknown homologous sequence in the genome. The chromosomal localization of the vWF pseudogene and of the breakpoint cluster region (BCR) gene were compared by 'in situ' hybridization: overlapping patterns were detected. The cloning, characterization and mapping of the pseudogenic region improves the analysis of this portion of chromosome 22 affected by several somatic and constitutional alterations, and also of the corresponding genic region on chromosome 12.
PMID: 2043485 [PubMed - indexed for MEDLINE]
Br J Haematol. 1990 Mar;74(3):282-9.
Characterization of polymorphic markers in the von Willebrand factor gene and pseudogene.
Bernardi F, Marchetti G, Casonato A, Gemmati D, Patracchini P, Legnani C, DeRosa V, Girolami A, Conconi F.
Centro Studi Biochimici delle Patologie del Genoma Umano, Istituto di Chimica Biologica, Università di Ferrara, Italy.
Three TaqI restriction fragment length polymorphisms (RFLP) detected by the central portion of von Willebrand factor cDNA, which recognizes the true gene and in addition pseudogenic sequences, were characterized and mapped. Small cDNA fragments which hybridized with DNA from families with von Willebrand disease were used. Two of the RFLP, recognized by 1.7 and 0.45 kb cDNA fragments, are not in linkage either with von Willebrand disease or with RFLP located in the von Willebrand factor (vWF) gene, which indicates their pseudogenic location. These markers located in 22q11, near to the bcr gene, provide new tools for the study of several somatic and constitutional alterations affecting this chromosomal region. The third RFLP is recognized by a cDNA fragment corresponding to the N-terminal portion of mature vWF and is localized in the true gene. Since significant linkage disequilibrium with other informative RFLP is not present, this marker contributes to the definition of family haplotypes associated with von Willebrand disease.
PMID: 1970740 [PubMed - indexed for MEDLINE]
Blood. 1990 Feb 1;75(3):677-83.
A de novo and heterozygous gene deletion causing a variant of von Willebrand disease.
Bernardi F, Marchetti G, Guerra S, Casonato A, Gemmati D, Patracchini P, Ballerini G, Conconi F.
Centro Studi Biochimici delle Patologie del Genoma Umano-Istituto di Chimica Biologica, Università di Ferrara, Italy.
An abnormal von Willebrand factor (vWF) gene restriction pattern has been found in a patient with von Willebrand disease. Because this gene alteration is not present in his parents or in 50 normal and 25 affected subjects, and the restriction fragment length polymorphism haplotypes are inherited normally in the patient's family, we suggest that a de novo mutation is present. Bands with reduced intensity and additional fragments, observed in several restriction digests, hybridize with noncontiguous copy DNA (cDNA) portions, thus indicating the presence of a heterozygous gene deletion. The deletion removes a genomic region containing at least codons 1147 through 1854 and corresponding to the D3-A3 homologous protein domains. The extent of the vWF pseudogene on chromosome 22 is roughly similar to that of the deleted area. However, the pseudogenic nature of the deletion is excluded by the mapping of bands with reduced intensity in the patient to the true vWF gene. The vWF antigen levels are one fourth of normal and ristocetin cofactor activity is severely impaired. The reduction of high molecular weight multimers in plasma and platelets and the altered triplet morphology are compatible with the presence of a dominant variant of type II von Willebrand disease.
PMID: 1967540 [PubMed - indexed for MEDLINE]
Hum Genet. 1989 Oct;83(3):264-6.
Sublocalization of von Willebrand factor pseudogene to 22q11.22-q11.23 by in situ hybridization in a 46,X,t(X;22)(pter;q11.21) translocation.
Patracchini P, Calzolari E, Aiello V, Palazzi P, Banin P, Marchetti G, Bernardi F.
Centro di Studi Biochimici delle Patologie del Genoma Umano, Università di Ferrara, Italy.
The von Willebrand factor pseudogene, previously mapped to chromosome 22, was sublocalized by in situ hybridization using as probe a von Willebrand factor cDNA fragment completely contained in the pseudogenic region. Chromosome spreads were from a patient carrying a unique balanced de novo translocation 46,X,t(X;22)(pter;q11.21). Silver grain analysis indicated that the human von Willebrand factor pseudogene is located on 22q,11,22-q11,23, a region relevant for several somatic and constitutional chromosomal alterations.
PMID: 2793170 [PubMed - indexed for MEDLINE]
Br J Haematol. 1988 Feb;68(2):243-8.
von Willebrand disease investigated by two novel RFLPs.
Bernardi F, Guerra S, Patracchini P, Volinia S, Buzzoni D, Ballerini G, Casonato A, Marchetti G.
Studi Biochimici Morbo di Cooley, Università di Ferrara, Italy.
Two partial cDNAs for von Willebrand factor (vWF) were used to investigate gene lesions and restriction fragment length polymorphisms (RFLPs) in vW disease (vWd) and normal controls. No gene alteration was detected but two TaqI RFLPs, likely to be intronic and originating from point mutations, were found in the 3' part of vWF gene. The two TaqI RFLPs, identified by the same probe, are informative in approximately 50% of the subjects. Used in combination with two other known RFLPs, they define several haplotypes similarly distributed in vWd and normals. Linkage disequilibrium between loci identified by the RFLPs is present. In a family study the RFLP patterns demonstrate homozygosity for the affected vWF gene in a severe (type III) patient and identify several heterozygous subjects. The RFLPs analysis has been related to the haemostatic values and multimer distribution. In two of the four unrelated patients with severe vWd examined the RFLPs study indicates double heterozygosity for the affected vWF genes.
PMID: 2894837 [PubMed - indexed for MEDLINE]
