The p53 tumor suppressor is a key mediator of the cellular response to stress. By inducing growth arrest, apoptosis, or senescence, it protects cells from malignant transformation. Diverse stress signals trigger phosphorylation of p53 on distinct serine and threonine residues. These modifications have been proposed to reduce p53 interaction with its negative regulator, MDM2, to allow for p53 stabilization, interaction with transcriptional co-activators, and activation of p53 target genes. However, genetic studies suggest that stress-activated phosphorylation may not be essential for p53 activation. We therefore investigated the role of p53 phosphorylation on 6 key serine residues (Ser6, Ser15, Ser20, Ser37, Ser46, Ser392) for p53 activation using Nutlin-3, a recently developed small-molecule MDM2 antagonist (Vassilev et al., 2004, Science 303, 844). Nutlin stabilizes p53 by preventing its interaction with, and subsequent ubiquitin-dependent degradation by MDM2. We show here that nutlin does not induce the phosphorylation of p53. Comparison of the activity of unphosphorylated and phosphorylated p53 induced by the genotoxic drugs doxorubicin and etoposide in HCT116 and RKO cells revealed no difference in their sequence-specific DNA binding, ability to transactivate p53 target genes (p21, MDM2 and MIC-1) and to induce p53-dependent apoptosis. We conclude that p53 phosphorylation on 6 major serine sites is not required for activation of p53 target genes or biological responses in vivo.

[Proc Amer Assoc Cancer Res, Volume 46, 2005]