Characterization of a Novel F5 Intronic Variant Associated with Aberrant Splicing and Severe Factor V Deficiency

Congenital factor V (FV) deficiency is a rare autosomal recessive bleeding disorder caused by defects in the F5 gene associated with bleeding manifestations of variable severity. In this study we report molecular and functional characterization of a novel F5 variant that causes aberrant splicing and significantly reduces protein expression in a patient with severe FV deficiency. We performed F5 mutation screening and functional study in a proband (FV:C, 0.4%) with a history of gastrointestinal bleeding, post-traumatic bleeding, hematomas, ecchymoses, and discomfort in his ankle joints since infancy. Sequencing revealed a novel homozygous F5 gene variant NC_000001.10:169519985G>C (or NM_000130.5:c.1297 –8C>G). Bioinformatics sequence analysis predicted that this variant would lead to the acceptor site loss of the intron 8–exon 9 junction. However, mRNA analysis identified that it also activated an aberrant splice site located 7 nucleotides upstream of the normal one and was associated with the production of an anomalous F5 transcript with retention of seven nucleotides of intron 8 resulting in a premature stop codon. We revealed no traces of normal transcript in the patient. Our findings confirm that it is not only changes in canonical splicing dinucleotides that can significantly disrupt splicing sites and impair pre-mRNA processing.