Splicing Analysis of 16 PALB2 ClinVar Variants by Minigene Assays: Identification of Six Likely Pathogenic Variants

Abstract

PALB2 pathogenic variants confer high risk of breast cancer. Here, we have analyzed the impact of PALB2 variants on splicing, a gene expression step that removes introns to form the mature messenger RNA. This process is performed by the splicing machinery through the recognition of specific sequences, namely the 30 and 50 splice sites, which determine the exon ends. Variants at these sequences may trigger anomalous splicing and aberrant transcripts that may be associated with a disease. To test the impact of variants on splicing, we used a biotechnological tool called minigene, which replicates, at small-scale, the human gene of interest. Thus, we checked 16 PALB2 variants at the intron/exon boundaries using the minigene mgPALB2_ex1-3. We found that twelve variants disrupted splicing, six of which could be classified as likely pathogenic. These results facilitate the clinical management of carrier patients and families since they may benefit from tailored prevention protocols and therapies.