Fig. 5

Blocking the folding of the SNORD2-intron modulates the level of exon 4 inclusion. A–C Schematic representation of the double strategy to investigate the effect of blocking the SNORD2-intron on the inclusion of exon 4 of the EIF4A2 gene. The strategy includes applying ASOs designed against the SNORD2-intron, highlighted in the blue box (B). ASO1 is entirely located inside the SNORD2 sequence, while ASO2 overlaps with SNORD2 and the intron and ASO3 is mainly in the intron. The second prong of the strategy involves a minigene of the 5′ of the EIF4A2 gene, from the promoter to the 3′ end of its exon 6, highlighted in the red box (C). A mutant of the minigene was also designed with the 30 nucleotides 3′ most in SNORD2 mutated so they are no longer complementary to the branch point regions of intron 3 of EIF4A2. D Box plot showing the modulation of the percent spliced in (PSI) value of exon 4 of EIF4A2 following treatment with different ASOs in 6 different replicates. E Box plot showing the modulation of the PSI value of exon 4 of EIF4A2 in the mutant minigene as compared to the wild-type (WT). **p < 0.01 and ***p < 0.001