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Differentiating pathogenic mutations from polymorphic alterations in the splice sites of BRCA1 and BRCA2
Author(s) -
Claes Kathleen,
Poppe Bruce,
Machackova Eva,
Coene Ilse,
Foretova Lenka,
De Paepe Anne,
Messiaen Ludwine
Publication year - 2003
Publication title -
genes, chromosomes and cancer
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.754
H-Index - 119
eISSN - 1098-2264
pISSN - 1045-2257
DOI - 10.1002/gcc.10221
Subject(s) - rna splicing , exon , splice , genetics , biology , nonsense mediated decay , brca2 protein , ovarian cancer , exon skipping , stop codon , cancer research , alternative splicing , splice site mutation , nonsense mutation , mutation , complementary dna , breast cancer , cancer , microbiology and biotechnology , rna , gene , missense mutation , germline mutation
About 4% of all BRCA1 and BRCA2 alterations reported to the Breast Information Core database are splice site variants. Only a limited number of them have been studied at the RNA level. By BRCA1 and BRCA2 mutation analysis of breast/ovarian cancer families, we identified two novel and eight previously reported potential splice site mutations, never characterized at the cDNA level before. RT‐PCR was performed to determine whether these variants disrupted correct splicing. To ensure efficient detection of transcripts containing premature termination codons, a nonsense‐mediated mRNA decay inhibitor was added to the lymphoblastoid cell lines of the patients before RNA extraction. We found that BRCA1 IVS3+3A>C, 4304G>A (in the last codon of exon 12), and IVS19+2delT and BRCA2 IVS6+1G>A, IVS23−2A>G, and IVS24+1G>A lead to aberrant transcripts in lymphocytes. Therefore, they were considered to be true pathogenic mutations, predisposing carriers to cancers of the hereditary breast/ovarian cancer syndrome. BRCA2 IVS24−16T>C is a frequent polymorphism in linkage disequilibrium, with a polymorphic stop codon in exon 27, K3326X. BRCA1 IVS2−14C>T and BRCA2 IVS9−5insT and IVS25+9A>C represent rare variants, not disrupting normal splicing in blood lymphocytes. However, some of the alterations may act differently, qualitatively and/or quantitatively, in breast or ovarian tissues. The data provided in this paper allowed more accurate risk estimation of patients and relatives carrying the mutations described herein and have facilitated genetic counseling. Furthermore, our study is important for a better understanding of splicing mechanisms and revealed new patterns of alternative splicing in BRCA1 and BRCA2 . © 2003 Wiley‐Liss, Inc.