Interaction of HA-SR proteins with CLK1 exon 4. RNA immunoprecipitation was carried out on cells transfected with the indicated HA-SR and an HA-EMPTY plasmids. (A) The expression of HA-tagged proteins was validated by immunoblotting an anti-HA antibody. (B) The recovered RNA was quantitated for CLK1 exon 4 using primers shown. The differential between values obtained for each sample is plotted as fold enrichment of recovery. Raw data for RNA immunoprecipitations are provided in Supplemental Table S1. (C) Dual expression of differentially tagged SR proteins as determined by immunoblotting with the relevant antibodies. (D) Impact of the dual expression on the recovery of exon 4 sequences. (E) Impact of dual expression of SR proteins on CLK1 exon 4 splicing. Values plotted correspond to ΔPSI that compare the shift in exon 4 splicing of the dual combination to the shift obtained with each HA protein, and thus the capacity of the Myc-SR or FLAG-SR protein to change the impact of the HA-SR protein alone. Duplicates of RT-PCR reactions are shown below (panel F). The statistical significance of values was evaluated using multiple t-test analysis. (*) P < 0.05, (**) P < 0.01, and (***) P < 0.001.
