Fig. 4. The effect of T-Ag is exon specific. (A) Diagrams showing different alternative exons and their corresponding cis-controlling elements used to evaluate the effects of T-Ag on alternative splicing. ESE, exonic splicing enhancer for SF2/ASF (GAAGAAGAG) and for SRp-55 (GCACGGAC). Positive controls for the activation of exon inclusion by overexpression of SRp-55 and SRp-40 are shown. (B) Effect of T-Ag on alternative splicing of an EDI exon in which the natural ESE was disrupted (lanes 1–4) or replaced by the SRp-55 target site (lanes 5 and 6). Lanes 7–12, effect of T-Ag on EDII alternative splicing. Hep3B cells were co-transfected with pSVEDEDB-Xho (lanes 7–12) and either pBSKS⁺ (Stratagene) (lanes 7 and 8) or pSVEDATot (lanes 9–12). The plasmid expressing T-Ag (Zhu et al., 1991) was co-transfected in the odd lanes. RT–PCRs for EDI (lanes 1–6, 9 and 10) and for EDII (lanes 7, 8, 11 and 12) were as in Materials and methods.