P53 AND P21 WAF PROTEINS MODULATE JNK ACTIVITY

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Exposure of mammalian cells to ultraviolet light (UV) and DNA damaging agents triggers the UV response which is characterized by induction of cell cycle regulatory proteins such as p53 and the cyclin-cyclin dependent kinase inhibitor p21WAF. Cells also respond to genotoxic stress by activation of the Jun amino terminal kinase/stress-activated protein kinase pathway (JNK/SAPK). Our preliminary results demonstrate that both wildtype p53 protein and p2lWAF protein can be co- immuneprecipitated with JNK1 enzyme in whole cell extracts. These results represent, to the best of our knowledge, the first report that both p53 and p21WAF proteins are associated with JNK1 enzyme in the cell. In addition, our studies indicate that when cells are expressing wildtype p53 protein and p2lWAF protein, basal JNK1 activity is low and JNK1 can be induced many-fold by UV above control levels. In contrast when cells are expressing mutant p53val135 protein and low levels of p2lWAF, basal JNK1 activity is elevated and fold induction of JNK1 by UV above control is reduced. Further analysis of JNK1 protein has provided preliminary evidence that the phosphorylation status of JNK1 is altered in cells expressing wildtype P53 and p21WAF1 protein (distinct from that normally associated with JNK1 activation). These preliminary results have led us to hypothesize that wildtype p53 protein and p21WAF both bind to JNK1 in the cell and this interaction affects JNK1 signal transduction by inhibiting basal JNK1 activity and modulating the ability of JNK1 to be induced by stress stimuli such as UV irradiation. We further hypothesize that the phosphorylation status of the JNK1 protein is modulated by a p53/p2lWAF-dependent mechanism. The following specific aims will test these hypotheses: Aim number 1 will characterize the interaction of p53 protein and p21WAF protein with JNK protein in cultured cells. Aim number 2 will investigate the effect of p53/p21WAF protein interaction with JNK1 enzyme on JNK1 function in cells. Aim number 3 will investigate the mechanism by which the phosphorylation status of JNK1 protein is altered in cells expressing p53/p21WAF. Aim number 4 will characterize the interaction of p53 protein, p2lWAFprotein, and JNK1 protein in vivo.