Tumor Suppressor p53 as a Component of the Tumor Necrosis Factor-induced, Protein Kinase PKR-mediated Apoptotic Pathway in Human Promonocytic U937 Cells*

Despite what is known about the early signaling events in tumor necrosis factor (TNF) α-induced apoptosis, characterization of the downstream events remains largely undefined. It is now known that a cross-talk exists between the interferon and TNF-α pathways. This linkage allows recruitment of the cell proliferation suppressor PKR (dsRNA-dependent protein kinase) from the interferon pathway to play a pivotal role in TNF-α-induced apoptosis. In this study, we took advantage of the differential TNF-α susceptibilities of human promonocytic U937 subclones, deficient in or overexpressing PKR, to further characterize the role of PKR in apoptosis. By reverse transcription-polymerase chain reaction, we demonstrated that TNF-α transiently induces the tumor suppressor p53 in U937 cells. This p53 induction lags behind the TNF-α induction of PKR by 1 h. By cell viability determination, ultrastructural studies, apoptotic DNA laddering, and antisense techniques, it was shown that inhibition of p53 expression in PKR-overexpressing U937 cells abrogates the TNF-α-induced apoptosis in these cells. Conversely, overexpressing wild type p53 in PKR-deficient U937 cells confers the susceptibility of these cells to TNF-α-induced apoptosis. This latter result indicates that p53 induction is an event downstream of TNF-α-induced up-regulation of PKR, thereby further establishing the critical role of p53 in TNF-α-induced apoptosis in U937 cells. PKR-overexpressing U937 cells were found to possess a constitutively higher level of p53, which partly explains why these cells spontaneously undergo apoptosis even without TNF-α treatment. Finally, a model is presented on the interplay between PKR and p53 in effecting TNF-α-induced apoptosis in U937 cells.