A deep intronic mutation in FGB creates a consensus exonic splicing enhancer motif that results in afibrinogenemia caused by aberrant mRNA splicing, which can be corrected in vitro with antisense oligonucleotide treatment

Hum Mutat. 2009 Feb;30(2):221-7. doi: 10.1002/humu.20839.


We previously described a novel homozygous point mutation (FGB c.115-600A>G) located deep within intron 1 of the fibrinogen beta gene (FGB), as a likely cause of afibrinogenemia. While this was the only mutation detected, its pathological mechanism was unclear. Here we show the mutation causes the inclusion of a 50-bp cryptic exon by creating a consensus heptad motif recognized by the spliceosome recruiting protein pre-mRNA splicing factor (SF2)/arginine/serine-rich alternative splicing factor (ASF) splicing factor 2/alternative splicing factor (SF2/ASF). Translation of the aberrant mRNA would result in truncation of the Bbeta chain, preventing fibrinogen synthesis. Selective introduction of a second mutation into the enhancer motif abolished the SF2/ASF binding motif and re-established normal pre-mRNA splicing. Subsequent introduction of antisense phosphorodiamidate morpholino oligonucleotides (PMOs) into transfected cells containing the mutant construct blocked the protein-RNA interaction and successfully restored normal splicing ( approximately 50% at 2 microM and approximately 90% at 10 microM). The molecular characterization of this case has revealed a unique disease mechanism, shown the importance of screening for deep intronic mutations, and provided evidence that antisense gene therapy is potentially practical for the treatment of diseases caused by this class of mutation.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Afibrinogenemia / genetics*
  • Animals
  • Base Sequence
  • COS Cells
  • Chlorocebus aethiops
  • Consensus Sequence
  • Enhancer Elements, Genetic*
  • Exons / genetics*
  • Fibrinogen / genetics*
  • Humans
  • Introns / genetics*
  • Molecular Sequence Data
  • Mutation / genetics*
  • Oligonucleotides, Antisense / pharmacology
  • RNA Splicing / drug effects
  • RNA Splicing / genetics*
  • RNA, Messenger / genetics
  • RNA, Messenger / metabolism
  • Reverse Transcriptase Polymerase Chain Reaction
  • Sequence Analysis, DNA


  • Oligonucleotides, Antisense
  • RNA, Messenger
  • Fibrinogen