P40: Deep intronic mutations may explain unbalanced expression of APC alleles in familial adenomatous polyposis

Taina Nieminen1, Walter Pavicic2, Noora Porkka3, Heikki Järvinen4, Anna  Lepistö5, Päivi Peltomäki6.

1 – University of Helsinki, Department of Medical and Clinical Genetics, Helsinki, Finland. 2 – Laboratorio de Citogenética y Mutagénesis, Instituto Multidisciplinario de Biología Celular (IMBICE-CONICET-CICPBA), La Plata, Argentina. 3 – University of Helsinki, Department of Medical and Clinical Genetics, Helsinki, Finland. 4 – Second Department of Surgery, Helsinki University Central Hospital, Helsinki, Finland. 5 – Department of Colorectal Surgery, Abdominal Center, Helsinki University Hospital, Helsinki, Finland. 6 – University of Helsinki, Department of Medical and Clinical Genetics, Helsinki, Finland.

Aim

Familial Adenomatous Polyposis (FAP) is caused by germline mutations in the APC gene. We investigated 56 FAP families from Finland that had remained APC mutation-negative after traditional tests. Of particular interest were four families with allele-specific expression (ASE) of APC alleles.

Method

These families were interrogated by next generation sequencing of the whole genomes (WGS) and RNA (RNA-seq).

Results

RNA-seq raised the suspicion of a pseudoexon (inclusion of intronic sequence in the mRNA) in three families. A pseudoexon between exons 5 and 6 of the APC gene was present in one family and a pseudoexon between exons 10 and 11 in two families. By WGS, the exon 5-6 pseudoexon generated a cryptic splice site leading to a 127-bp intronic insertion in the APC mRNA. Two alternative genetic changes underlay the exon 10-11 pseudoexon and both created a new splice donor site leading to an identical 83-bp insertion in APC mRNA. All three pseudoexons were predicted to cause frameshifts and premature stop codons leading to APC protein truncation.

Conclusion

Pseudoexon mutations accounted for 3/4 families with APC-ASE in our study. We conclude that RNA-seq is an effective method to reveal pseudoexons and is worth considering in mutation-negative families.

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