title: Naturally occurring BRCA2 alternative mRNA splicing events in clinically relevant samples.
abstract: BRCA1 and BRCA2 are the two principal tumour suppressor genes associated with inherited high risk of breast and ovarian cancer. Genetic testing of BRCA1/2 will often reveal one or more sequence variants of uncertain clinical significance, some of which may affect normal splicing patterns and thereby disrupt gene function. mRNA analyses are therefore among the tests used to interpret the clinical significance of some genetic variants. However, these could be confounded by the appearance of naturally occurring alternative transcripts unrelated to germline sequence variation or defects in gene function. To understand which novel splicing events are associated with splicing mutations and which are part of the normal BRCA2 splicing repertoire, a study was undertaken by members of the Evidence-based Network for the Interpretation of Germline Mutant Alleles (ENIGMA) consortium to characterise the spectrum of naturally occurring BRCA2 mRNA alternate-splicing events.
  
  
  
  mRNA was prepared from several blood and breast tissue-derived cells and cell lines by contributing ENIGMA laboratories. cDNA representing BRCA2 alternate splice sites was amplified and visualised using capillary or agarose gel electrophoresis, followed by sequencing.
  
  
  
  We demonstrate the existence of 24 different BRCA2 mRNA alternate-splicing events in lymphoblastoid cell lines and both breast cancer and non-cancerous breast cell lines.
  
  
  
  These naturally occurring alternate-splicing events contribute to the array of cDNA fragments that may be seen in assays for mutation-associated splicing defects. Caution must be observed in assigning alternate-splicing events to potential splicing mutations.
authors.affiliation: Department of Medicine, University of Chicago, Chicago, Illinois, USA.
authors.forename: James D
authors.initials: JD
authors.lastname: Fackenthal
authors.affiliation: Department of Medicine, University of Chicago, Chicago, Illinois, USA.
authors.forename: Toshio
authors.initials: T
authors.lastname: Yoshimatsu
authors.affiliation: Department of Medicine, University of Chicago, Chicago, Illinois, USA.
authors.forename: Bifeng
authors.initials: B
authors.lastname: Zhang
authors.affiliation: Department of Medicine, University of Chicago, Chicago, Illinois, USA.
authors.forename: Gorka R
authors.initials: GR
authors.lastname: de Garibay
authors.affiliation: Unit of Molecular Bases of Genetic Risk and Genetic Testing, Department of Preventive and Predictive Medicine, Fondazione IRCCS Istituto Nazionale dei Tumori (INT), Milano, Italy.
authors.forename: Mara
authors.initials: M
authors.lastname: Colombo
authors.affiliation: Unit of Molecular Bases of Genetic Risk and Genetic Testing, Department of Preventive and Predictive Medicine, Fondazione IRCCS Istituto Nazionale dei Tumori (INT), Milano, Italy.
authors.forename: Giovanna
authors.initials: G
authors.lastname: De Vecchi
authors.affiliation: Department of Medicine, University of Chicago, Chicago, Illinois, USA.
authors.forename: Samantha C
authors.initials: SC
authors.lastname: Ayoub
authors.affiliation: Department of Medicine, University of Chicago, Chicago, Illinois, USA.
authors.forename: Kumar
authors.initials: K
authors.lastname: Lal
authors.affiliation: Department of Medicine, University of Chicago, Chicago, Illinois, USA.
authors.forename: Olufunmilayo I
authors.initials: OI
authors.lastname: Olopade
authors.affiliation: Fundación Pública Galega de Medicina Xenómica-SERGAS, Grupo de Medicina Xenómica-USC, CIBERER, IDIS, Santiago de Compostela, Spain.
authors.forename: Ana
authors.initials: A
authors.lastname: Vega
authors.affiliation: Fundación Pública Galega de Medicina Xenómica-SERGAS, Grupo de Medicina Xenómica-USC, CIBERER, IDIS, Santiago de Compostela, Spain.
authors.forename: Marta
authors.initials: M
authors.lastname: Santamariña
authors.affiliation: Fundación Pública Galega de Medicina Xenómica-SERGAS, Grupo de Medicina Xenómica-USC, CIBERER, IDIS, Santiago de Compostela, Spain.
authors.forename: Ana
authors.initials: A
authors.lastname: Blanco
authors.affiliation: Medical Faculty, Center for Hereditary Breast and Ovarian Cancer, Center for Integrated Oncology (CIO) and Center for Molecular Medicine Cologne (CMMC), University of Cologne and University Hospital Cologne, Germany.
authors.forename: Barbara
authors.initials: B
authors.lastname: Wappenschmidt
authors.affiliation: Medical Faculty, Center for Hereditary Breast and Ovarian Cancer, Center for Integrated Oncology (CIO) and Center for Molecular Medicine Cologne (CMMC), University of Cologne and University Hospital Cologne, Germany.
authors.forename: Alexandra
authors.initials: A
authors.lastname: Becker
authors.affiliation: Service de Génétique and INSERM U830, Institut Curie and Université Paris Descartes, Sorbonne Paris Cité, Paris, France.
authors.forename: Claude
authors.initials: C
authors.lastname: Houdayer
authors.affiliation: Department of Pathology, University of Otago, Christchurch, New Zealand.
authors.forename: Logan C
authors.initials: LC
authors.lastname: Walker
authors.affiliation: Laboratorio de Oncología Molecular, Instituto de Investigación Sanitaria San Carlos (IdISSC), Hospital Clínico San Carlos, Madrid, Spain.
authors.forename: Irene
authors.initials: I
authors.lastname: López-Perolio
authors.affiliation: Department of Clinical Genetics, Odense University Hospital, Odense C, Denmark.
authors.forename: Mads
authors.initials: M
authors.lastname: Thomassen
authors.affiliation: Genetics and Computational Biology Division, QIMR Berghofer Medical Research Institute, Brisbane, Queensland, Australia.
authors.forename: Michael
authors.initials: M
authors.lastname: Parsons
authors.affiliation: Genetics and Computational Biology Division, QIMR Berghofer Medical Research Institute, Brisbane, Queensland, Australia.
authors.forename: Phillip
authors.initials: P
authors.lastname: Whiley
authors.affiliation: Department of Clinical Genetics, Maastricht University Medical Center, Maastricht, The Netherlands.
authors.forename: Marinus J
authors.initials: MJ
authors.lastname: Blok
authors.affiliation: Maastricht Science Programme, Faculty of Humanities and Sciences, Maastricht University, Maastricht, The Netherlands.
authors.forename: Rita D
authors.initials: RD
authors.lastname: Brandão
authors.affiliation: Department of Clinical Genetics, Maastricht University Medical Center, Maastricht, The Netherlands.
authors.forename: Demis
authors.initials: D
authors.lastname: Tserpelis
authors.affiliation: Human Development and Health Academic Unit, Faculty of Medicine, University of Southampton, Southampton General Hospital, Southampton, UK.
authors.forename: Diana
authors.initials: D
authors.lastname: Baralle
authors.affiliation: Oncogenetics Group, Vall d'Hebron Institute of Oncology (VHIO) and Universitat Autonoma de Barcelona, Barcelona, Spain.
authors.forename: Gemma
authors.initials: G
authors.lastname: Montalban
authors.affiliation: Oncogenetics Group, Vall d'Hebron Institute of Oncology (VHIO) and Universitat Autonoma de Barcelona, Barcelona, Spain.
authors.forename: Sara
authors.initials: S
authors.lastname: Gutiérrez-Enríquez
authors.affiliation: Oncogenetics Group, Vall d'Hebron Institute of Oncology (VHIO) and Universitat Autonoma de Barcelona, Barcelona, Spain Clinical and Molecular Genetics Area, University Hospital Vall d'Hebron, Barcelona, Spain.
authors.forename: Orland
authors.initials: O
authors.lastname: Díez
authors.affiliation: Molecular Diagnostic Unit, Hereditary Cancer Program, IDIBELL-Catalan Institute of Oncology, Barcelona, Spain.
authors.forename: Conxi
authors.initials: C
authors.lastname: Lazaro
authors.affiliation: Genetics and Computational Biology Division, QIMR Berghofer Medical Research Institute, Brisbane, Queensland, Australia.
authors.forename: Amanda B
authors.initials: AB
authors.lastname: Spurdle
authors.affiliation: Unit of Molecular Bases of Genetic Risk and Genetic Testing, Department of Preventive and Predictive Medicine, Fondazione IRCCS Istituto Nazionale dei Tumori (INT), Milano, Italy.
authors.forename: Paolo
authors.initials: P
authors.lastname: Radice
authors.affiliation: Laboratorio de Oncología Molecular, Instituto de Investigación Sanitaria San Carlos (IdISSC), Hospital Clínico San Carlos, Madrid, Spain.
authors.forename: Miguel
authors.initials: M
authors.lastname: de la Hoya
mesh_terms: D017398:Alternative Splicing / Q000235:genetics*; D019313:BRCA1 Protein / Q000235:genetics; D024682:BRCA2 Protein / Q000235:genetics*; D001943:Breast Neoplasms / Q000235:genetics; D002460:Cell Line; D045744:Cell Line, Tumor; D005260:Female; D020022:Genetic Predisposition to Disease / Q000235:genetics; D005820:Genetic Testing / Q000379:methods; D006801:Humans; D061986:MCF-7 Cells; D009154:Mutation / Q000235:genetics; D010051:Ovarian Neoplasms / Q000235:genetics; D022821:RNA Splice Sites / Q000235:genetics; D012333:RNA, Messenger / Q000235:genetics*
chemical_list: D019313:BRCA1 Protein; C492913:BRCA1 protein, human; D024682:BRCA2 Protein; C551750:BRCA2 protein, human; D022821:RNA Splice Sites; D012333:RNA, Messenger
keywords: BRCA2; alternate splicing; breast cancer; tumor suppressor