Inhibitory Role of Plk1 in the Regulation of p73-dependent Apoptosis through Physical Interaction and Phosphorylation

• Published in: The Journal of biological chemistry 2008-03, Vol.283 (13), p.8555-8563
• Main Authors: Koida, Nami, Ozaki, Toshinori, Yamamoto, Hideki, Ono, Sayaka, Koda, Tadayuki, Ando, Kiyohiro, Okoshi, Rintaro, Kamijo, Takehiko, Omura, Ken, Nakagawara, Akira
• Format: Article
• Language: English
• Subjects: Animals; Apoptosis; Cell Cycle Proteins - genetics; Cell Cycle Proteins - metabolism; Cell Line; Chlorocebus aethiops; DNA-Binding Proteins - genetics; DNA-Binding Proteins - metabolism; Enzyme Activation; Humans; Nuclear Proteins - genetics; Nuclear Proteins - metabolism; Phosphorylation; Polo-Like Kinase 1; Protein Binding; Protein Serine-Threonine Kinases - genetics; Protein Serine-Threonine Kinases - metabolism; Proto-Oncogene Proteins - genetics; Proto-Oncogene Proteins - metabolism; RNA, Small Interfering - genetics; Simian virus 40; Transcriptional Activation - genetics; Tumor Protein p73; Tumor Suppressor Protein p53 - genetics; Tumor Suppressor Protein p53 - metabolism; Tumor Suppressor Proteins - genetics; Tumor Suppressor Proteins - metabolism
• ISSN: 0021-9258; 1083-351X
• DOI: 10.1074/jbc.M710608200

In response to DNA damage, p73 plays a critical role in cell fate determination. In this study, we have found that Plk1 (polo-like kinase 1) associates with p73, phosphorylates p73 at Thr-27, and thereby inhibits its pro-apoptotic activity. During cisplatin-mediated apoptosis in COS7 cells in which the endogenous p53 is inactivated by SV40 large T antigen, p73 was induced to accumulate in association with a significant down-regulation of Plk1. Consistent with these observations, Plk1 reduced the stability of the endogenous p73. Immunoprecipitation and in vitro pulldown assay demonstrated that p73 binds to the kinase domain of Plk1 through its NH2-terminal region. Luciferase reporter assay and reverse transcription-PCR analysis revealed that Plk1 is able to block the p73-mediated transcriptional activation. Of note, kinase-deficient Plk1 mutant (Plk1(K82M)) retained an ability to interact with p73; however, it failed to inactivate the p73-mediated transcriptional activation, suggesting that kinase activity of Plk1 is required for the inhibition of p73. Indeed, in vitro kinase assay indicated that p73 is phosphorylated at Thr-27 by Plk1. Furthermore, small interference RNA-mediated knockdown of the endogenous Plk1 in p53-deficient H1299 cells resulted in a significant increase in the number of cells with sub-G1 DNA content accompanied by the up-regulation of p73 and pro-apoptotic p53AIP1 as well as the proteolytic cleavage of poly(ADP-ribose) polymerase. Thus, our present results suggest that Plk1-mediated dysfunction of p73 is one of the novel molecular mechanisms to inhibit the p53-independent apoptosis, and the inhibition of Plk1 might provide an attractive therapeutic strategy for cancer treatment.