JNK destabilizes p53 in NPM-ALK–expressing cells. NPM-ALK phosphorylates JNK on Thr183/Tyr185 as demonstrated by down-regulation of NPM-ALK after transfection of SUDHL-1 cells with siRNA targeted to ALK for 48 hours (A). Likewise, NPM-ALK–expressing BaF3 cells express JNK in a phosphorylated form (B). Inhibition of JNK activity with the ATP competitive inhibitor SP600125 for 48 hours at a dose of 20 μM results in an increase in p53 expression in SUDHL-1 cells (C). Confocal microscopy examining p53 cellular location in SUDHL-1 cells treated with 20 μM SP600125 for 24 hours (D). IP of p53 or MDM2 protein after treatment of SUDHL-1 cells with SP600125 for 48 hours at 20 μM. Numbers represent ratios of arbitrary units after densitometric analysis of JNK and p53 protein levels; vertical lines have been inserted to indicate a repositioned gel lane (E). IP for p53 followed by Western blot analysis for ubiquitinated p53 using an anti-ubiquitin antibody. Numbers represent the percentage of p53 protein that is ubiquitinated as determined after densitometric analysis of protein levels (F).