Regulation of ubiquitination and degradation of p53 in unstressed cells through C-terminal phosphorylation.
Previously, we found that the protein kinase C (PKC) inhibitor H7 stimulates p53 to accumulate in a form incapable of inducing transcription from p53-dependent promoters. We concluded that H7 inhibits constitutive C-terminal phosphorylation of p53, which regulates its turnover in unstressed cells. We now show that p53 and its inhibitor MDM2 (HDM2 in human cells) are together in the nuclei of H7-treated cells and can be co-immunoprecipitated. Despite this association of p53 with the ubiquitin ligase MDM2, ubiquitinated p53 was not detected in H7-treated cells. Furthermore, co-treatment with H7 and the proteosome inhibitor LLnL prevented the accumulation of ubiquitinated p53 that was observed in cells treated solely with LLnL. In addition, treatment of cells with the PKC activator phorbol ester stimulated the ubiquitination of p53 and reduced its ability to accumulate after stress. H7 did not induce the phosphorylation of human p53 on Ser-15 (Ser-18 in mouse protein), a modification that occurs in response to DNA damage and leads to the release of MDM2 and to transactivation by p53. We conclude that phosphorylation of the C-terminal domain of p53 by PKC increases its ubiquitination and degradation in unstressed cells.
Chernov
M V
MV
Department of Molecular Biology, Lerner Research Institute, The Cleveland Clinic Foundation, Cleveland, Ohio 44195, USA.
Bean
L J
LJ
Lerner
N
N
Stark
G R
GR
D019307:1-(5-Isoquinolinesulfonyl)-2-Methylpiperazine / Q000494:pharmacology; D000818:Animals; D004789:Enzyme Activation; D004791:Enzyme Inhibitors / Q000494:pharmacology; D006801:Humans; D006868:Hydrolysis; D051379:Mice; D009687:Nuclear Proteins; D010766:Phosphorylation; D011493:Protein Kinase C / Q000037:antagonists & inhibitors / Q000378:metabolism; D011518:Proto-Oncogene Proteins / Q000378:metabolism; D051736:Proto-Oncogene Proteins c-mdm2; D012694:Serine / Q000378:metabolism; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology; D016159:Tumor Suppressor Protein p53 / Q000737:chemistry / Q000378:metabolism*; D014452:Ubiquitins / Q000378:metabolism*
D004791:Enzyme Inhibitors; D009687:Nuclear Proteins; D011518:Proto-Oncogene Proteins; D016159:Tumor Suppressor Protein p53; D014452:Ubiquitins; D012694:Serine; D019307:1-(5-Isoquinolinesulfonyl)-2-Methylpiperazine; C497206:MDM2 protein, human; C497207:Mdm2 protein, mouse; D051736:Proto-Oncogene Proteins c-mdm2; D011493:Protein Kinase C; D013755:Tetradecanoylphorbol Acetate