In vitro transcription of compound heterozygous hypofibrinogenemia Matsumoto IX; first identification of FGB IVS6 deletion of 4 nucleotides and FGG IVS3-2A > G causing abnormal RNA splicing

BackgroundWe reported a case of hypofibrinogenemia Matsumoto IX (M IX) caused by a novel compound heterozygous mutation involving an FGB IVS6 deletion of 4 nucleotides (Δ4b) (three T, one G; between FGB IVS6-10 and -16) and FGG IVS3-2A/G, which are both identified for the first time. To examine the transcription of mRNA from the M IX gene, we cloned the wild-type and mutant genes into expression vectors.MethodsThe vectors were transfected into CHO cells and transiently produced wild-type, Bβ- or γ-mRNA in the cells. The mRNAs amplified with RT-PCR were analyzed by agarose gel electrophoresis and nucleotide sequencing.ResultsThe RT-PCR product from FGB IVS6Δ4b showed aberrant mRNA that included both introns 6 and 7, and that from FGG IVS3-2G showed two aberrant mRNAs, a major one including intron 3 and a minor in which intron 3 was spliced by a cryptic splice site in exon 4. We speculated that the aberrant mRNAs are degraded before translation into proteins, and/or translated variant chains are subjected to quality control and degraded in the cytoplasm.ConclusionThe reduced plasma fibrinogen level of the M IX patient was caused by abnormal RNA splicing of one or both of the FGB and FGG genes.