FIGURE 3.

Effects of BPS and splice sites on splicing and leakage of pre-mRNA. (A–C) Boxplots of the strength of BPS (BPS scores), 5′ splice site (5′ss MAXENT), and 3′ splice site (3′ss MAXENT). All introns of human RefGenes, nuclear-retained pre-mRNAs that contain introns in the Nuc category in Figure 1E, and leaked pre-mRNAs that contain introns in the Cyto category in Figure 1E were analyzed. (D) Schematic representation of CDKN1B-derived reporters. Each box shows an exon and a line shows an intron. (E) HEK293T cells were transfected with the reporter plasmids, then treated with SSA (100 ng mL⁻¹, 6 h). RNA samples were purified from the cells. RT-PCR analysis was performed for detecting the spliced and unspliced forms of the reporter constructs. Open triangle and filled triangle indicate spliced mRNA and pre-mRNA, respectively. (F) HEK293T cells were transfected with the reporter plasmids and then treated with SSA (100 ng mL⁻¹, 6 h). Total cell extracts were analyzed by immunoblotting. The antibodies used are indicated to the right of the respective blots.