Abstract

The functional analysis of the common prothrombin (F2) 20210*A allele has recently revealed gain-of-function of 3′end processing as a novel genetic mechanism predisposing to human disease. Based on these observations we sought for the functional impact of a novel C>T mutation at position 20209 (F2 20209*T). This mutation affects the penultimate nucleotide of the 3′ untranslated region (UTR) of the F2 mRNA and has been reported to be associated with fetal loss, intrauterine growth retardation and a history of venous thrombosis. We have therefore systematically analyzed the functional effect of this polymorphism and show that the F2 20209*T mutation up-modulates the F2 mRNA expression and protein expression by ~1.4 fold which corresponds to the observation that the reported patients with the F2 20209*T allele tend to have moderate F2 level elevations, slightly above - but also within - the upper normal-range.

Furthermore, because of the critical position of the F2 20209*T allele we have analyzed the putative impact of this mutation on 3′end mRNA formation and show that this mutation represents a gain-of-function mutation, causing increased cleavage site recognition, increased 3′ end processing and increased mRNA accumulation neither affecting the poly(A) tail length nor the site of cleavage. Thus, this novel mutation falls into a general category of mutations in the F2 gene which contribute to thrombophilia by interfering with a tightly balanced architecture of non-canonical 3′end formation signals. This, together with previous results, demonstrates that the F2 cleavage site and adjacent parts of the 3′ flanking sequence represent an error-prone “vulnerable region” in which N>T mutations may give rise to clinically relevant gain-of-function phenotypes.

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