Figure 3.
The RBM20 RRM and C-terminus are sufficient for titin splicing repression. (A) Schematic representation of RBM20 and the deletion mutants. The two zinc finger (ZnF) domains are depicted in blue, the RRM green, the RS-like domain in orange. Unstructured regions are indicated as small gray boxes. The WT protein and all deletion mutants carry C-terminal myc- and his-epitope tags. RBM20 amino acid residues are indicated underneath the schemes and the corresponding molecular weight of proteins is depicted in brackets. (B) RT-PCR validation of RBM20-mediated changes in processing the human titin TTN241-3 splicing reporter in HEK293 cells. Gray boxes represent mRNA species with and without the alternative exon 242 (top and bottom band, respectively). Quantification of three independent transfections are shown as percentages of mRNA excluding the alternative exon. (C) qRT-PCR (SYBR Green) analysis of HEK293 cells transfected with the TTN241-3 splicing reporter and the RBM20 expression constructs with activity in panel B: n = 3; *P < 0.05; **P < 0.01; ***P < 0.001.