Figure 4.

The region downstream of IR exon 11 bound by MBNL1 is required for regulation by MBNL1. (A–J) Diagram of the genomic segments tested in the RHCglo minigene to identify which segment of the 273-nt region contains the MBNL response element. White indicates IR wild-type sequences and black indicates heterologous sequences from the human β-globin gene. X indicates deletion of the 30-nt sequence containing the binding site and the red box indicates the binding site with mutated sequences, which abolish MBNL1 direct interaction (substitutions from probe 8, Figure 3). The minigenes were co-transfected into QT35 quail fibroblasts with plasmids expressing Flag-MBNL1, Flag-MBNL3 or empty vector and splicing was determined by RT–PCR. The numbers indicate the change in percentage inclusion compared to basal inclusion levels (no MBNL). Results are from three independent experiments.