TY - JOUR T1 - Comprehensive RNA and protein functional assessments contribute to the clinical interpretation of <em>MSH2</em> variants causing in-frame splicing alterations JF - Journal of Medical Genetics JO - J Med Genet DO - 10.1136/jmg-2022-108576 SP - jmedgenet-2022-108576 AU - Laëtitia Meulemans AU - Stéphanie Baert Desurmont AU - Marie-Christine Waill AU - Gaia Castelain AU - Audrey Killian AU - Julie Hauchard AU - Thierry Frebourg AU - Florence Coulet AU - Alexandra Martins AU - Martine Muleris AU - Pascaline Gaildrat Y1 - 2022/09/16 UR - http://jmg.bmj.com/content/early/2022/09/15/jmg-2022-108576.abstract N2 - Background Spliceogenic variants in disease-causing genes are often presumed pathogenic since most induce frameshifts resulting in loss of function. However, it was recently shown in cancer predisposition genes that some may trigger in-frame anomalies that preserve function. Here, we addressed this question by using MSH2, a DNA mismatch repair gene implicated in Lynch syndrome, as a model system.Methods Eighteen MSH2 variants, mostly localised within canonical splice sites, were analysed by using minigene splicing assays. The impact of the resulting protein alterations was assessed in a methylation tolerance-based assay. Clinicopathological characteristics of variant carriers were collected.Results Three in-frame RNA biotypes were identified based on variant-induced spliceogenic outcomes: exon skipping (E3, E4, E5 and E12), segmental exonic deletions (E7 and E15) and intronic retentions (I3, I6, I12 and I13). The 10 corresponding protein isoforms exhibit either large deletions (49–93 amino acids (aa)), small deletions (12 or 16 aa) or insertions (3–10 aa) within different functional domains. We showed that all these modifications abrogate MSH2 function, in agreement with the clinicopathological features of variant carriers.Conclusion Altogether, these data demonstrate that MSH2 function is intolerant to in-frame indels caused by the spliceogenic variants analysed in this study, supporting their pathogenic nature. This work stresses the importance of combining complementary RNA and protein approaches to ensure accurate clinical interpretation of in-frame spliceogenic variants.All data relevant to the study are included in the article or uploaded as supplementary information. ER -