p53 Antiproliferative function is enhanced by aspartate substitution at threonine 18 and serine 20.
Previous studies have demonstrated the irradiation-induced phosphorylation of p53 at Thrl8 and Ser20, residues integral within an a-helical segment of the transactivation domain. Importantly, phosphorylation at either site has been correlated with decreased binding to the inhibitory partner Mdm-2 and enhanced transactivation of p53 target genes. In this study, we investigated the impact of Asp substitution at Thrl8 and Ser20 (p53Tl8D/S20D) on the functional regulation of p53. Asp substitution is commonly accepted as a means of mimicking phosphorylation due to the introduction of negative charge within the functional group. p53T18D/S20D was refractory to in vitro digestion by calpain, a protease recognizing a-helical structure within the transactivation domain. In addition, transfected p53T18D/S20D poorly bound GST-Mdm-2 in vitro, enhanced the endogenous expression of the p53 transactivation targets p21(Waf1/Cip1) and fas/APO-1, and significantly curtailed cell proliferation relative to wild-type p53 transfected cells. Thus, Asp substitution at Thr18 and Ser20 within the a-helical segment of the transactivation domain reduced Mdm-2 interaction, upregulating transactivation of cell-cycle and apoptotic regulatory targets, curtailing cellular proliferation.
Jabbur
James R
JR
Department of Pathology, Cancer Genomics Laboratory, Graduate Program in Cancer Biology, The University of Texas M. D. Anderson Cancer Center, Houston, Texas 77030, USA.
Zhang
Wei
W
D019943:Amino Acid Substitution; D001224:Aspartic Acid / Q000737:chemistry* / Q000235:genetics; D002154:Calpain / Q000494:pharmacology; D002453:Cell Cycle; D050759:Cyclin-Dependent Kinase Inhibitor p21; D016213:Cyclins / Q000378:metabolism; D024202:Electrophoretic Mobility Shift Assay; D015972:Gene Expression Regulation, Neoplastic; D005982:Glutathione Transferase / Q000378:metabolism; D006801:Humans; D006860:Hydrogen Bonding; D007938:Leukemia / Q000378:metabolism / Q000473:pathology; D016297:Mutagenesis, Site-Directed; D009687:Nuclear Proteins; D010766:Phosphorylation; D011233:Precipitin Tests; D011401:Promoter Regions, Genetic; D011518:Proto-Oncogene Proteins / Q000378:metabolism; D051736:Proto-Oncogene Proteins c-mdm2; D012694:Serine / Q000737:chemistry* / Q000235:genetics; D013912:Threonine / Q000737:chemistry* / Q000235:genetics; D014158:Transcription, Genetic; D015533:Transcriptional Activation; D014407:Tumor Cells, Cultured; D016159:Tumor Suppressor Protein p53 / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D019014:fas Receptor / Q000378:metabolism
C493956:CDKN1A protein, human; D050759:Cyclin-Dependent Kinase Inhibitor p21; D016213:Cyclins; D009687:Nuclear Proteins; D011518:Proto-Oncogene Proteins; D016159:Tumor Suppressor Protein p53; D019014:fas Receptor; D013912:Threonine; D001224:Aspartic Acid; D012694:Serine; C497206:MDM2 protein, human; D051736:Proto-Oncogene Proteins c-mdm2; D005982:Glutathione Transferase; D002154:Calpain