Figure 3
Effect of SRSF1 knockdown or the reduction of H3K36me3 levels on the alternative splicing of apoptotic gene transcripts. (a) Huh-7 cells were transfected with 40 nM si-SRSF1 in the SRSF1 knockdown, or 40 nM scramble RNA was used as the negative control for 72 h. The efficiency of the SRSF1 knockdown was assessed through Western blotting. (b) mRNAs were extracted and detected using RT-PCR for the alternative splicing of the HIPK3, SMAC/DIABLO, and SURVIVIN transcripts. (c) Huh-7 cells were transfected with 20 nM si-SETD2 for the reduction of H3K36me3 levels or 20 nM scramble RNA was used as the negative control for 72 h, followed by exposure to 10 μM 4bHWE for 24 h. The efficiency of the SETD2 knockdown and the reduction of H3K36me3 levels was assessed through Western blotting. (d) mRNAs were extracted and detected using RT-PCR for the alternative splicing of the HIPK3, SMAC/DIABLO, and SURVIVIN transcripts. Western blotting: equal amounts of whole cell lysates (20 μg) were separated using SDS-PAGE and immunoblotted with various antibodies as indicated. Tubulin and Histone H3 are shown as internal standards. The fold-change values are presented below each band. RT-PCR: the ratios of the densities of the two bands (alternative exon-containing isoforms to alternative exon-lacking isoforms) are presented below each group. Data are the mean of three independent experiments. *p < 0.05 compared with the control. a p < 0.05 for the comparison between scramble RNA + 4bHWE and si-SETD2 + 4bHWE. M = marker. Full-length blots are presented in Supplementary Fig. S2.
