Figure 1

Dichromatic fluorescence alternative splicing reporter for Ttn to monitor the RBM20 activity in splicing regulation. (A) Schematic representation of the Ttn reporter minigene TtnE50-E51E218-E219-EGFP/mCherry (top) and mRNAs derived from it (bottom). Two genomic fragments Ttn E50-E51 and E218-E219 were inserted between human histone H2B cDNA and the EGFP/mCherry cassette. Expression of E51E218-EGFP and ΔE51E218-mCherry indicates inclusion and skipping of a chimeric exon E51E218, respectively. (B) Microphotographs of HeLa cells co-transfected with the fluorescence Ttn splicing reporter minigene and an empty vector or an expression vector for the wild-type (WT) or mutant RBM20 protein. Fluorescence of EGFP and mCherry is pseudo-colored in green and magenta, respectively. Scale bar, 100 µm. (C) RT-PCR analysis of the Ttn splicing reporter minigene co-expressed with an empty vector or an expression vector for the wild-type (WT) or mutant RBM20 protein in HeLa cells. Representative gel-like presentation (left) and calculated inclusion levels (right) are indicated. Error bars indicate standard errors of the means. ^#p < 0.001 and **p < 0.01 to WT (n = 3 biological replicates, one-way ANOVA followed by Dunnett’s test).