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. 2022 Jul 4;23(13):7446.
doi: 10.3390/ijms23137446.

Identification of Spliceogenic Variants beyond Canonical GT-AG Splice Sites in Hereditary Cancer Genes

Vita Šetrajčič Dragoš  1   2 , Ksenija Strojnik  3 , Gašper Klančar  1 , Petra Škerl  1 , Vida Stegel  1 , Ana Blatnik  2   3 , Marta Banjac  3 , Mateja Krajc  3 , Srdjan Novaković  1   4
Affiliations

Identification of Spliceogenic Variants beyond Canonical GT-AG Splice Sites in Hereditary Cancer Genes

Vita Šetrajčič Dragoš et al. Int J Mol Sci. .

Abstract

Pathogenic/likely pathogenic variants in susceptibility genes that interrupt RNA splicing are a well-documented mechanism of hereditary cancer syndromes development. However, if RNA studies are not performed, most of the variants beyond the canonical GT-AG splice site are characterized as variants of uncertain significance (VUS). To decrease the VUS burden, we have bioinformatically evaluated all novel VUS detected in 732 consecutive patients tested in the routine genetic counseling process. Twelve VUS that were predicted to cause splicing defects were selected for mRNA analysis. Here, we report a functional characterization of 12 variants located beyond the first two intronic nucleotides using RNAseq in APC, ATM, FH, LZTR1, MSH6, PALB2, RAD51C, and TP53 genes. Based on the analysis of mRNA, we have successfully reclassified 50% of investigated variants. 25% of variants were downgraded to likely benign, whereas 25% were upgraded to likely pathogenic leading to improved clinical management of the patient and the family members.

Keywords: RNAseq; VUS; VUS reclassification; aberrant splicing; deep intronic variants; hereditary cancer syndrome; in silico prediction tools; pseudoexon; reclassification; splice variant; spliceogenic; spliceogenic variant.

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Conflict of interest statement

The authors declare no conflict of interest.

Figures

Figure 1
Figure 1
Sashimi plots representing variant’s impact on splicing. Variant (A) uLZTR1:c.1942G>T caused complete splicing impairment (Class 3S), variants (B) FH:c.1237-11C>G; (C) ATM:c.5763-1056G>A; (D) APC:c.1408+743_1408+745delinsACG; (E) RAD51C:c.1027-3C>G; (F) ATM:c.172G>T; (G) MSH6:c.3646+5G>A; (H) ATM:c.7816A>G; (I) TP53:c.74+23C>T; (J) PALB2:c.2379C>T caused incomplete or uncertain splicing impairment (Class 2S) and variants (K) RAD51C:c.779G>A; (L) PALB2:c.3495G>A have not impaired splicing. Red arrows represent variant’s location. Red graphs depict carriers, orange graphs depict control samples. RPKM—reads per kilobase of exon model, represents the number of mapped reads in the sample.
See this image and copyright information in PMC

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