A novel G-to-A mutation in the intron-N of the protein S gene leading to abnormal RNA splicing in a patient with protein S deficiency

Background and Objectives. Hereditary protein S (PS) deficiency is a rare autosomal disorder of the coagulation pathway associated with familial thrombophilia. Design and Methods. We investigated a young propositus with recurrent deep vein thrombosis, a positive family history for thrombotic episodes, and low plasma concentrations of free, but not total PS antigen (12% and 70%, respectively). Results. Sequence analysis of the PS gene showed a heterozygous G-to-A mutation at the first nucleotide of intron N. The patient's father, who had suffered from deep vein thrombosis and had reduced total and free PS antigen (59% and 28%, respectively) was a heterozygote. The G-to-A change predicts the disappearance of a donor splice site. After transfection with a construct, containing either the wild-type or the mutated sequence, cells with the mutant construct showed an aberrant mRNA, consistent with exclusion of exon 14, but not the expected mRNA. Sequencing of the abnormal mRNA showed the complete absence of exon 14. Exclusion of exon 14 predicts the deletion of the amino acid sequence from residue 508 to residue 582, and the shift of the reading frame of the following 8 amino acids with a premature stop codon within exon 15 at position 591. Thus, the truncated PS gene product would not contain the terminal portion of the sex hormone binding globulin-like domain. Interpretation and Conclusions. We have identified a mutation in a highly conserved intronic region of PS gene. The mutation affects in vitro mRNA processing and efficiency of normal splicing.