Decidualization of the endometrial stromal compartment is critical for embryo implantation. Initiation of this differentiation process requires elevated intracellular cAMP levels. We now report a massive and sustained up-regulation of p53 tumor suppressor protein during cAMP-induced decidualization of cultured endometrial stromal cells. Nuclear accumulation of p53 was not accompanied by increased mRNA expression, suggesting stabilization of the protein as the underlying mechanism. Proteasomal degradation of p53 is known to be mediated by nuclear Mdm2. Nuclear translocation of Mdm2, in turn, is dependent on phosphorylation by protein kinase B/Akt (PKB/Akt). In cAMP-treated decidualized cells, p53 accumulation was associated with decreased nuclear Mdm2 and cytoplasmic PKB/Akt levels. Conversely, withdrawal of the decidualization stimulus resulted in morphological and biochemical dedifferentiation, disappearance of p53, but increased abundance of PKB/Akt. Furthermore, Western blot and immunohistochemical analyses of endometrial biopsies confirmed that p53 is expressed in vivo in the stromal compartment during the late secretory phase of the cycle. The observation that p53 protein expression is closely associated with decidual transformation indicates a novel role for this tumor suppressor in regulating human endometrial function.