Figure 8.

SAK overexpression attenuated p53-induced apoptosis. (A) Establishment of SAK stably expressing clones: U2-OS (p53-wt) cells were transfected with plasmid-expressing FLAG-tagged SAK, along with the pcDNA3 vector control. Stable clones were isolated after G418 selection and ring cloning. Cell lysates were prepared and subjected to Western blot analysis using anti-FLAG antibody. Two independent clones (Cl. 5 and Cl. 8) with SAK expression as a 98-kDa band were shown, along with a vector control. (B) SAK overexpression attenuated p53-induced apoptosis: three stable lines, shown in (A), were subjected to etoposide (25 µM) treatment for 36 hours to induce p53 or left untreated. Both attached and detached cells were harvested and subjected to DNA fragmentation gel assay. Induction of DNA fragmentation by etoposide-activated p53 was obviously seen in the control cells, but to a much lesser extent, in two SAK-overexpressing clones.