Five novel mutations in the gene for human blood coagulation factor V associated with type I factor V deficiency

Abstract

IntroductionHuman coagulation factor V (FV) is a single-chain glycoprotein that plays an important role in maintaining the hemostatic balance. It circulates in blood as an inactive procoagulant with a M r of 330 kd and a structure consisting of 3 homologous A-type domains and 2 homologous C-type domains connected by a heavily glycosylated B domain in the order A1-A2-B-A3-C1-C2. Proteolytic cleavage by thrombin at R709, R1018, and R1545 (single-letter amino acid codes) results in removal of the B domain and converts the procofactor into the fully active cofactor FVa, which consists of a M r 105-kd heavy chain (A1-A2) and a M r 74-or 71-kd light chain (A3-C1-C2), associated via a single Ca ϩϩ ion. [1][2][3] The difference in molecular weight of the light chain reflects the presence of 2 isoforms of FVa (FVa 1 and FVa 2 ) due to alternative glycosylation of the C2 domain, which leads to different affinities for biologic membranes and subsequent overall procoagulant activity. 4,5 In its active form, FVa forms an essential part of the prothrombinase complex that catalyzes the conversion of prothrombin to thrombin by factor Xa in the presence of calcium and a phospholipid membrane. 1-3 Activated protein C (APC) inactivates FVa through cleavage of the active cofactor at R306, R506, and R679 and requires FV as a cofactor in the APC-mediated inactivation of factor VIIIa (FVIIIa). 6,7 Thus, FV plays an important role in the procoagulant pathway as well as in the protein C anticoagulant pathway. The structure of FV is similar to FVIII (both cofactors share approximately 40% homology in their heavy and light chains) and ceruloplasmin, the copper-binding protein in plasma. 8,9 Recently, the crystal structure of the C2 domain of FV has been established 10 and molecular models for the A and C domains of FV have been proposed. 11,12 The gene for coagulation FV has been mapped to chromosome 1q23 13 and spans more than 80 kilobases (kb). It consists of 25 exons and the messenger RNA (mRNA) encodes a leader peptide of 28 amino acids and a mature protein of 2196 amino acids. Roughly, the heavy chain is encoded by exons 1 to 12 and the light chain by exons 14 to 25. The entire B domain is encoded by exon 13, which contains 2 tandem repeats of 17 amino acids and 31 tandem repeats of 9 amino acids that are absent in the B domain of FVIII. 14,15 Deficiency of FV, or parahemophilia, was first described in 1947 by Owren. 16 It is a rare autosomal recessive bleeding disorder with an estimated frequency of one in one million. The phenotypic expression of FV deficiency is variable; heterozygotes are usually asymptomatic, whereas homozygous patients show mild, moderate, or severe bleeding symptoms. Identifying the molecular basis underlying this disease will help to obtain more insight into the mechanisms involved in this variable clinical expression. The recently published complete nucleotide sequence of the FV gene (GenBank accession number Z99572) has facilitated the molecular characterization underlying FV deficiency and reports have ide...

Factor V Deficiency Associated with Congenital Cardiac Disorder and Intracranial Hemorrage

“…Most factor V-deficient patients have low factor V activity and antigen level. More than 200 factor V-deficient cases have been reported in the literature, but the molecular basis for factor V deficiency has been established in only a few cases [5,6]. Hereditary factor V deficiency is a relatively uncommon disorder characterized by a variable course but with a uniformly favorable response to transfusion with fresh plasma or freshly frozen plasma.…”

Factor V I359T: a novel mutation associated with thrombosis and resistance to activated protein C

“…Of these substitutions, the previously unreported c.529G>T transversion is predicted to result in the substitution of Glu 119 by a termination codon (FV E119X). This allele was inherited in cis with the c.6533C>T transition that is predicted to result in the substitution of Met 2120 by Thr and was previously reported (van Wijk et al, 2001). The primary consequence of a nonsense mutation such as c.529G>T (FV E119X) is not the synthesis of a truncated protein but recognition and degradation of the mRNA by the cell via a process termed nonsense mediated decay.…”

“…This analysis showed heterozygosity for seven nucleotide substitutions: c.495G>A (exon 4), c.529G>T (exon 4), c.642G>T (exon 4), c.1250T>C (exon 8), c.1332A>G (exon 8), c.4185C>T (exon 13) and c.6533C>T (exon 24). Four of these (c.495G>A, c.642G>T, c.1332A>G and c.4185C>T) were previously reported polymorphisms that were not predicted to cause amino-acid substitutions (Zivelin et al, 1997;Cargill et al, 1999;Irizarry et al, 2000;Le et al, 2000;Ohnishi et al, 2000;van Wijk et al, 2001). The remaining substitutions (c.529G>T, c.1250T>C and c.6533C>T) were predicted to result in the substitutions FV E119X, FV I359T and FV M2120T respectively.…”

“…The FV gene exons and intronexon junctions were amplified by PCR from genomic DNA obtained from II-2 using oligonucleotide primers and experimental conditions described previously (van Wijk et al, 2001). All nucleotide substitutions were confirmed by direct DNA sequence analysis performed in forward and reverse directions.…”

Arg2074Cys missense mutation in the C2 domain of factor V causing moderately severe factor V deficiency: molecular characterization by expression of the recombinant protein

“…5 The genetic basis of severe or moderately severe type I FV deficiency is still poorly explored (up to now, a total of 12 mutations scattered through F5 have been identified). [8][9][10][11][12][13] Most mutations are nonsense, frameshift, or splicing mutations, giving rise to null alleles, whereas only 3 missense mutations have been identified. An additional 8 mutations causing FV deficiency have been reported in the heterozygous state, 5 of which were found in patients who carried the FV Leiden mutation on the second allele, leading to the "pseudohomozygous APC-resistance" condition.…”