Outcome of donor splice site mutations accounting for congenital afibrinogenemia reflects order of intron removal in the fibrinogen alpha gene (FGA)

Abstract

Congenital afibrinogenemia (Mendelian Inheritance in Man #202400) is a rare, autosomal recessive disorder characterized by the complete absence of circulating fibrinogen. Our recent studies on the molecular basis of the disease showed that the most common genetic defect is a donor splice mutation in fibrinogen alpha gene (FGA) intron 4, IVS4؉1G>T. Two other FGA donor splice mutations, in intron 1 (IVS1؉3A>G) and intron 3 (IVS3؉1_؉4delGTAA), were identified in afibrinogenemia patients. Because it was impossible to directly study the effect of these mutations on mRNA splicing in patient hepatocytes, we used a transfected cell approach, which previously allowed us to show that the common IVS4 mutation causes afibrinogenemia due to the activation of multiple cryptic donor splice sites. In this study, analysis of the IVS3delGTAA mutation showed exon 3 skipping in 99% of transcripts and exons 2 and 3 skipping in 1% of transcripts. The different outcomes of these donor splice mutations appear to follow the model proposed in a study of fibrillar collagen genes, where donor splice mutations occurring in a rapidly spliced intron with respect to upstream introns lead in most cases to exon skipping, while mutations in later-spliced introns lead to intron inclusion or cryptic splice-site utilization. Indeed, we found that in FGA intron 3 was preferentially spliced first, followed by intron 2, intron 4, and intron 1. (Blood. 2003;

-spectrinBari: a truncated -chain responsible for dominant hereditary spherocytosis

“…There is strong evidence to support the theory that the order of intron removal and speed of splicing are important determinants of mRNA transcripts with multiple exon skipping. [21][22][23] On this basis, our results confirm the proposal that the double exon skipping represents an example of mutually inclusive exons in which the unusually large size of one exon is contingent to the recognition of the following exon by the splicing mechanism. 21 Otherwise, increasing evidence shows that RNA specific secondary structure influences the splicing machinery and plays an important role in exon definition for particular transcripts.…”

Mutations in the fibrinogen gene cluster accounting for congenital afibrinogenemia: an update and report of 10 novel mutations

“…These mutations all affect the conserved donor splice site consensus sequence, with the exception of the FGB c.9581 13C4T mutation that creates a novel donor splice site . For some aberrant splicing with exon skipping [Attanasio et al, 2003;Margaglione et al, 2000], intron inclusion , or usage of cryptic splice sites has been demonstrated. As for other genetic diseases, mutations at acceptor splice sites are less frequent; only two have been identified so far; i.e., one in FGG c.124-3C4G [NeermanArbez et al, 2001] and one in FGB c.1245-1G4C [Horellou et al, 2006].…”

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

“…Reports on autosomal recessive disorder with a molecular base of congenital afibrinogenemia indicated association with the aberrant splicing caused by inactivation of physiologic splice sites [11][12][13][14][15][16][17][18][19][20][21][22][23][24]. Most abnormal RNA splicing was reported to occur near the donor splice site, with five kinds of mutation in FGA, four in FGB and three in the IVS3 +1 to +4 deletions, which were another FGA donor splice site mutation, showed exon 3 skipping in 99% of transcripts, and exon 2 and 3 skipping in 1% of transcripts [23].…”

“…Most abnormal RNA splicing was reported to occur near the donor splice site, with five kinds of mutation in FGA, four in FGB and three in the IVS3 +1 to +4 deletions, which were another FGA donor splice site mutation, showed exon 3 skipping in 99% of transcripts, and exon 2 and 3 skipping in 1% of transcripts [23]. On the other hand, mRNAs from FGB IVS6Δ4b and FGG IVS3-2G, the mutations of both of which were located in acceptor splice sites, showed no splicing out of intron 6 and intron 7 in the Bβ-chain or of intron 3 in the γ-chain, respectively.…”

“…Fibrinogen Matsumoto IX: FGB and FGG gene splicing mutations Some cases of afibrinogenemia caused by aberrant mRNA have been reported to be associated with large deletions [8][9][10] or other genetic mutations that affect transcription [11][12][13][14][15][16][17][18][19][20][21][22][23][24]. We assumed that abnormal transcription of mRNA for FGB IVS6Δ4b and/or FGG IVS3-2G caused hypofibrinogenemia.…”