mRBMIR interacts with a functionally active population of SR proteins in vitro. (A) Depletion of NEs with beads coated with mRBMIR but not thioredoxin alone inhibited the splicing of a panel of SR protein-dependent pre-mRNA substrates. (B) Quantitation of data from independent pre-mRNA splicing assays showing the percentage of splicing inhibition for each substrate. In one of the experiments, there was slight RNA degradation in the ASF/SF2 sample, resulting in the increased size of the error bar. (C) Add-back of individual SR proteins to mRBMIR-depleted extract can restore splicing of specific substrates. The individual SR species 9G8, ASF/SF2, or SC35 (150–200 μg) or SRp20 (400 μg) were added to the depleted extract. Pre-mRNA alone is shown in lane 1. The percentage of splicing efficiency of each of the pre-mRNA substrates is shown underneath. (D) 9G8, SRp20, and ASF/SF2 interact specifically with mRBMIR in an S100 extract. The same amount of each specific SR species that was added initially was readded after depletion in the add-back experiments. Pre-mRNA alone is shown in lane 1. The percentage of splicing efficiency in each lane is shown underneath.