Molecular analysis of factor VII deficiency in Italy: a frequent mutation (FVII Lazio) in a repeated intronic region

Abstract

Molecular defects and polymorphic haplotypes of coagulation factor VII gene were studied in eight unrelated Italian subjects with factor VII deficiency, seven having the factor VII- variant, one the factor VIIR variant. An intron 7 mutation, which alters the consensus donor splice site sequence, was found in six subjects. The presence of the founder effect is suggested by their common geographical origin (a mountain area in the Lazio region) and by the identical polymorphic haplotype underlying the mutation. A different mutation, also located in the 5' monomer of the repeated intron 7 sequence, was found in the heterozygous condition in a subject from Northern Italy. New polymorphic alleles were detected in the repeated intron 7 region in subjects from Eastern Africa. Two missense mutations in codon 97 (Gly-->Cys, Gly-->Ser), the first found in the compound heterozygous condition with the frequent intron 7 mutation, suggest the presence of a hot spot mutation site in the second epidermal growth factor domain. Two neutral dimorphisms at codon 333Ser and 115His were detected, the last in linkage disequilibrium with the 353Arg/Gln polymorphism, and showing differences in frequency in the FVII deficient and control subjects.

U1-snRNA–mediated rescue of mRNA processing in severe factor VII deficiency

“…As a model to exploit snRNAs for restoration of correct splicing in coagulation factor deficiencies, we chose the 9726 ϩ 5g/a mutation 17 in FVII occurring in the donor splice site of intron 7 (IVS7) at the ϩ 5 position, whose substitution has been frequently found to be associated with human diseases. [7][8][9] The homozygous patients experienced life-threatening hemorrhagic symptoms and require replacement therapy.…”

Coinheritance of Factor V (FV) Leiden enhances thrombin formation and is associated with a mild bleeding phenotype in patients homozygous for the FVII 9726+5G>A (FVII Lazio) mutation

“…A point mutation (9726ϩ5GϾA, FVII Lazio) affecting nucleotide ϩ5 of intron 7, which is part of the splicing consensus sequence, has been identified as a common cause of CRM Ϫ FVII deficiency in central Italy. 16 FVII Lazio homozygous individuals have undetectable FVII levels and are usually severe bleeders. 16 In vitro expression experiments have shown that the FVII Lazio mutation activates a cryptic splice site in the second minisatellite repeat, leading to a 37-nucleotide insertion in the mature transcript, which in turn causes a frameshift and premature termination of translation.…”

“…16 FVII Lazio homozygous individuals have undetectable FVII levels and are usually severe bleeders. 16 In vitro expression experiments have shown that the FVII Lazio mutation activates a cryptic splice site in the second minisatellite repeat, leading to a 37-nucleotide insertion in the mature transcript, which in turn causes a frameshift and premature termination of translation. Only 0.2% to 1% of all FVII Lazio mRNA is spliced correctly, resulting in a very small amount of normal FVII.…”

“…A point mutation (9726ϩ5GϾA, FVII Lazio) affecting nucleotide ϩ5 of intron 7, which is part of the splicing consensus sequence, has been identified as a common cause of CRM Ϫ FVII deficiency in central Italy. 16 The poor correlation between FVII levels and bleeding tendency 4,5,18 in carriers of FVII defects points at the existence of additional genetic and/or environmental factors that modulate the clinical phenotype. In particular, coinheritance of common thrombophilic mutations might result in a milder clinical phenotype in patients with severe FVII deficiency, as it has been shown for hemophilic disorders.…”

Residual factor VII activity and different hemorrhagic phenotypes in CRM+ factor VII deficiencies (Gly331Ser and Gly283Ser)

“…11,12 Mutagenesis and transfection expression vectors were created by site-directed mutagenesis of plasmid pCDNA3-FVII. 13 Oligonucleotides for mutagenesis spanned the following complementary DNA nucleotides: (1) 1193 through 1220 containing the 1206G Ͼ A transition (Gly331Ser) or the 1207G Ͼ C transversion (Gly331Ala); (2) 1040 through 1073 containing the 1062G Ͼ A transition (Gly283Ser) or the 1063 G Ͼ C transversion (Gly283Ala).…”