The Impact of Variants at Branchpoint Splicing Elements in Cancer Genes

Purpose: Branchpoint elements are required for intron removal, and variations at these elements can result in aberrant splicing. We aimed to assess the value of branchpoint annotations generated from recent large-scale studies to select branchpoint-abrogating variants, using hereditary cancer genes as model. Methods: We identified branchpoint elements in 119 genes associated with hereditary cancer from three genome-wide experimentally-inferred and two predicted branchpoint datasets. We then identified variants from public databases that occur within branchpoint elements. We compared conservation, unique variant observations, and population frequencies at different nucleotides within branchpoint motifs. Finally, selected minigene assays were performed to assess the functional impact of variants in branchpoint elements within mismatch repair genes. Results: There was poor overlap between predicted and experimentally-inferred branchpoints. Our analysis of cancer genes suggested that variants at -2nt, -1nt, and branchpoint positions in experimentally-inferred canonical motifs are more likely to be clinically relevant. Minigene assay data showed the -2nt to be more important to branchpoint motif integrity, but also demonstrated fluidity in branchpoint usage. Conclusion: Data from cancer gene analysis suggests that there will be few high-risk alleles that severely impact function via branchpoint abrogation. Results inform a general scheme to prioritize branchpoint motif variants for further study.