Calibration of Pathogenicity Due to Variant-Induced Leaky Splicing Defects by Using BRCA2 Exon 3 as a Model System
Year: 2020
Type: article
Abstract: BRCA2 is a clinically actionable gene implicated in breast and ovarian cancer predisposition that has become a high priority target for improving the classification of variants of unknown significance (VUS). Among all BRCA2 VUS, those causing partial/leaky splicing defects are the most challenging to classify because the minimal level of full-length (FL) transcripts required for normal function remains to be established. Here, we explored BRCA2 exon 3 (BRCA2e3) as a model for calibrating variant-induced spliceogenicity and estimating thresholds for BRCA2 haploinsufficiency. In silico predictions, minigene splicing assays, patients' RNA analyses, a mouse embryonic stem cell (mESC) complementation assay and retrieval of patient-related information were combined to determine the minimal requirement of FL BRCA2 transcripts. Of 100 BRCA2e3 variants tested in the minigene assay, 64 were found to be spliceogenic, causing mild to severe RNA defects. Splicing defects were also confirmed in patients' RNA when available. Analysis of a neutral leaky variant (c.231T>G) showed that a reduction of approximately 60% of FL BRCA2 transcripts from a mutant allele does not cause any increase in cancer risk. Moreover, data obtained from mESCs suggest that variants causing a decline in FL BRCA2 with approximately 30% of wild-type are not pathogenic, given that mESCs are fully viable and resistant to DNA-damaging agents in those conditions. In contrast, mESCs producing lower relative amounts of FL BRCA2 exhibited either null or hypomorphic phenotypes. Overall, our findings are likely to have broader implications on the interpretation of BRCA2 variants affecting the splicing pattern of other essential exons. Significance: These findings demonstrate that BRCA2 tumor suppressor function tolerates substantial reduction in full-length transcripts, helping to determine the pathogenicity of BRCA2 leaky splicing variants, some of which may not increase cancer risk.
Source: Cancer Research
Authors: Hélène Tubeuf, Sandrine M. Caputo, Teresa Sullivan, Julie Rondeaux, Sophie Krieger, Virginie Caux-Moncoutier, Julie Hauchard, Gaia Castelain, Alice Fiévet, Laëtitia Meulemans, Françoise Révillion, Mélanie Léoné, Nadia Boutry-Kryza, Capucine Delnatte, Marine Guillaud-Bataille, Linda Cleveland, Susan Reid, Eileen Southon, Omar Soukarieh, Aurélie Drouet, Daniela Di Giacomo, Myriam Vezain, Françoise Bonnet-Dorion, Violaine Bourdon, Hélène Larbre, Danièle Muller, Pascal Pujol, Fátima Vaz, Séverine Audebert-Bellanger, Chrystelle Colas, Laurence Venat-Bouvet, Angela R. Solano, Dominique Stoppa-Lyonnet, Claude Houdayer, Thierry Frebourg, Pascaline Gaildrat, Shyam K. Sharan, Alexandra Martins
Institutions: Inserm, Université de Rouen Normandie, Université Paris Sciences et Lettres, Institut Curie, National Cancer Institute, Center for Cancer Research, Normandie Université, Centre François Baclesse, Université de Caen Normandie, Institut Gustave Roussy, Université Paris Descartes, Centre Oscar Lambret, Hospices Civils de Lyon, Centre Hospitalier Universitaire de Nantes, Institut Bergonié, Institut Paoli-Calmettes, Institut Jean Godinot, Institut Strauss, Hôpital Arnaud de Villeneuve, Instituto Português de Oncologia Francisco Gentil, Hôpital Universitaire Dupuytren, Centro de Educación Médica e Investigaciones Clínicas Norberto Quirno
Language: English
Subfield: Genetics
Domain: Life Sciences
Open Access status: hybrid
Funders: European Commission, Institut National de la Santé et de la Recherche Médicale, Association Nationale de la Recherche et de la Technologie, Ligue Contre le Cancer, Institut National Du Cancer, Direction Générale de l’offre de Soins, Groupement des Entreprises Françaises dans la lutte contre le Cancer, Région Normandie, National Institutes of Health, European Regional Development Fund, National Cancer Institute
OpenAlex ID: W3041221269