Figure 5

Effects of Tip110 expression on pre-mRNA splicing-dependent CAT reporter gene expression. A. Scheme of the HIV-1 Rev protein-dependent intron-containing CAT reporter DM138. the CAT gene expression requires nuclear export of intron-containing (unspliced) CAT transcript through the interaction between HIV-1 Rev protein and Rev responsive element (RRE) within the CAT transcript. SD splice donor, SA splice acceptor. b 293T cells were transfected with DM138 (lane 1), DM138 and Rev (lane 2), or DM138, Rev and increasing amounts of Tip110 (lanes 3–7). The cells were harvested 48 h after transfection for CAT assay (top panel) or Western blot for Tip110 (bottom panel). c 293T cells were transfected with DM138 (lane 1), DM138 and Rev (lane 2), or DM138, Rev and increasing amounts of Tip110 siRNA expression plasmids (lanes 3–7). The cells were harvested 48 h after transfection for CAT assay (top panel) or Western blot for Tip110 (bottom panel). The amount of total DNA in all transfections was equalized with pcDNA3 plasmid. A CMVβGal plasmid was included to normalize the transfection variations for CAT assay, while β-actin was included to show equal loading (b, c). Compared to Figure 5b, c only used one fifth of DM138 reporter plasmid in the transfections.