Intronic variants can abrogate a BP site or activate a pseudoexon
Only a limited number of variants in hereditary cancer genes have been reported to cause aberrant splicing through the alteration of BP sites (Table 5). Of these, experimental validation of BP abrogation has been conducted for only two XPC variants detected in patients with xeroderma pigmentosum, a condition that increases the risk of skin cancers: LRG_472t1:c.413-24A>G resulted in partial skipping of exon 4 skipping in patient-derived mRNA (Khan et al., 2004); and LRG_472t1:c.413-9T>A, located within the acceptor motif but also annotated as a BP site variant, was found to lead to complete exon 4 skipping (Khan et al., 2004). The -9 and -24 nucleotides in intron 3 of XPC were subsequently shown to be functional BP sites necessary for the efficient and accurate splicing ofXPC pre-mRNA using U2 small nuclear ribonucleoprotein-BP interaction assays (Khan et al., 2010). Two substitutions predicted to alter a BP motif within the RB1 gene, identified in patients with retinoblastoma, result in the skipping of a downstream exon (Houdayer et al., 2008; K. Zhang et al., 2008). Similarly, a substitution at the -18 nucleotide upstream of exon 5 in BRCA1(LRG_292t1:c.135-18T>G) resulted in a three-fold increase of the Δ5 transcript isoform in an analysis of mRNA from a hereditary breast and ovarian cancer patient (Wappenschmidt et al., 2012). This variant was not captured by in silico prediction methods but it is within the -18 to -44 nucleotide window of high-confidence annotated BPs (Signal, Gloss, Dinger, & Mercer, 2018). Nine other variants within the BP window in BRCA1 , MLH1 and RAD51C have been reported to lead to exon skipping (Leman et al., 2020).
Pseudoexon-activating variants have been documented mostly in rare monogenic disorders, but several examples have also been reported in genes causing hereditary cancer syndromes (Vaz-Drago et al., 2017). To date, there are at least 13 documented pseudoexon-activating variants in hereditary cancer genes (Table 5): 12 are single nucleotide variants that create a new 5’ splice site or strengthen an existing cryptic 5’ splice site, and the other is a 4-bp deletion that introduces an intron-splicing processing element.