Fig. 8. Phosphorylation represses the transactivating function of PPAR. (A) Western blot analysis of phosphorylation levels of PPAR in young and senescent 2BS and WI-38 cells. Total proteins were extracted, and western blotting was performed using specific antibodies against PPAR and phosphorylated PPAR (p-PPAR) as indicated. Arrow indicates phosphorylated PPAR, and the observed doublet is reported as two translation initiation sites. (B) 2BS cells were transfected with expression plasmids pcDNA3.1 (–), pcDNA-PPAR (WT) or mutant pcDNA-PPAR (S84A) as indicated, together with wild-type p16-Luc. Cells were subsequently treated with 20 µM troglitazone or 10 µM pioglitazone. Luciferase activities were measured 24 hours after treatment. Values are the mean ± s.d. of triplicate points from a representative experiment (n=3), which was repeated three times with similar results. ^*P0.05.