Splicing analysis of 16 PALB2 clinVar variants by minigene assays: identification of six likely pathogenic variants
| Autor: | Alberto Valenzuela-Palomo, Lara Sanoguera-Miralles, Elena Bueno-Martínez, Ada Esteban-Sánchez, Inés Llinares-Burguet, Alicia García-Álvarez, Pedro Pérez-Segura, Susana Gómez-Barrero, Miguel de la Hoya, Eladio A. Velasco-Sampedro |
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| Přispěvatelé: | Ministerio de Ciencia e Innovación (España), Instituto de Salud Carlos III, European Commission, Junta de Castilla y León, CSIC-UVA - Instituto de Biología y Genética Molecular (IBGM), Asociación Española Contra el Cáncer, Universidad de Valladolid, Comunidad de Madrid |
| Rok vydání: | 2022 |
| Předmět: | |
| Zdroj: | Digital.CSIC. Repositorio Institucional del CSIC instname Cancers; Volume 14; Issue 18; Pages: 4541 |
| Popis: | PALB2 loss-of-function variants are associated with significant increased risk of breast cancer as well as other types of tumors. Likewise, splicing disruptions are a common mechanism of disease susceptibility. Indeed, we previously showed, by minigene assays, that 35 out of 42 PALB2 variants impaired splicing. Taking advantage of one of these constructs (mgPALB2_ex1-3), we proceeded to analyze other variants at exons 1 to 3 reported at the ClinVar database. Thirty-one variants were bioinformatically analyzed with MaxEntScan and SpliceAI. Then, 16 variants were selected for subsequent RNA assays. We identified a total of 12 spliceogenic variants, 11 of which did not produce any trace of the expected minigene full-length transcript. Interestingly, variant c.49-1G > A mimicked previous outcomes in patient RNA (transcript ∆(E2p6)), supporting the reproducibility of the minigene approach. A total of eight variant-induced transcripts were characterized, three of which (∆(E1q17), ∆(E3p11), and ∆(E3)) were predicted to introduce a premature termination codon and to undergo nonsense-mediated decay, and five (▼(E1q9), ∆(E2p6), ∆(E2), ▼(E3q48)-a, and ▼(E3q48)-b) maintained the reading frame. According to an ACMG/AMP (American College of Medical Genetics and Genomics/Association for Molecular Pathology)-based classification scheme, which integrates mgPALB2 data, six PALB2 variants were classified as pathogenic/likely pathogenic, five as VUS, and five as likely benign. Furthermore, five ±1,2 variants were catalogued as VUS because they produced significant proportions of in-frame transcripts of unknown impact on protein function. The EAV lab is supported by grants from the Spanish Ministry of Science and Innovation, Plan Nacional de I+D+I 2013–2016, ISCIII (PI20/00225), co‐funded by FEDER (the European Regional Development Fund, European Union) and the Consejería de Educación, Junta de Castilla Cancers 2022, 14, 4541 16 of 18 y León, ref. CSI242P18 (actuación cofinanciada P.O. FEDER 2014–2020 de Castilla y León) and Pro‐ grama Estratégico Instituto de Biología y Genética Molecular (IBGM), Escalera de Excelencia, Junta de Castilla y León (Ref. CLU‐2019‐02). The MdlH lab is supported by a grant from the Spanish Ministry of Science and Innovation, Plan Nacional de I+D+I 2013–2016, ISCIII (PI20/00110), co‐funded by FEDER. L.S.‐M. is supported by a predoctoral fellowship from the AECC Scientific Foundation, Sede Provincial de Valladolid (2019–2023). E.B.‐M. is a postdoctoral researcher funded by the University of Valladolid (POSTDOC‐UVA05, 2022–2025). I.L.‐B. is supported by a predoctoral fellowship from the Consejería de Educación, Junta de Castilla y León (2022–2025). A.E.‐S. is supported through the Operational Program for Youth Employment and the Youth Employment Initiative (YEI), set up by the Community of Madrid in 2020 and co‐financed by the European Social Fund. |
| Databáze: | OpenAIRE |
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