Figure 6

Functional role of p53 acetylation in nutlin-sensitivity. (A) SAOS-2 and H1299 cells were transiently transfected with empty vector, p53 full length (FL) or the actylation defective mutant p53 6KR and treated with 20 μM nutlin-3 for 24 hours. Cell viability was determined using the WST-1 viability/proliferation assay (***p < 0.001, **p < 0.01). Results were analyzed in triplicates in tree independent experiments and error bars represent standard error of mean. Transfections were verified in Western blots with antibodies against p53 and actin. (B) H1299 cells were transiently transfected with empty vector (EV), p53 full length (FL), or p53 6KR and treated with DMSO or 20 μM nutlin-3 for 6 hours. Western blotting was performed using antibodies against p53, MDM2, acetylated p53 (Lys382), Hsp27, Hsp90 and acetylated Hsp90 (Lys294) and actin. Bands were quantified using region of interest imaging analysis, and values are given as fold induction of control (DMSO treated sample for each transfection for EV/p53 FL/p53 6KR) relative to actin.