Multiple sequence variants ofBRCA2exon 7 alter splicing regulation

摘要: Background Exonic variants of unknown biological significance (VUS) identified in patients can affect mRNA splicing, either by changing 5′ or 3′ splice sites modifying splicing regulatory elements. Bioinformatic predictions these elements are still inaccurate and only few such have been functionally mapped BRCA2 . We studied the effect on eight exon 7 VUS, selected from French UMD- mutation database. Methods performed minigene assays analyses patient RNA. also developed a pyrosequencing-based quantitative assay, to measure, RNA, relative contribution each allele production 7-containing transcripts. Moreover, an exonic enhancer (ESE)-dependent assay was used evaluate properties wild-type mutant segments. Results Six out induced defects. In c.517G>T c.631G>A altered natural sites, c.572A>G created new site, c.520C>T, c.587G>A c.617C>G skipping (66%, 25% 46%, respectively). Pyrosequencing RNA confirmed levels for c.520C>T c.617C>G. Results ESE-dependent indicated that disturb elements. Conclusions is regulated multiple sensitive disease-associated sequence variations. Measurements allelic imbalance patient-derived and/or using provide valuable estimates extent partial Assessment pathogenicity with awaits additional evidence especially completion segregation analyses.