Defective p53 post-translational modification required for wild type p53 inactivation in malignant epithelial cells with mdm2 gene amplification.
Mdm2 gene amplification occurs in benign and chemotherapy-responsive malignant tumors with wtp53 genes as well as in breast and epithelial cancers. Mdm2 amplification in benign tumors suggests that it is not sufficient for p53 inactivation in cancer, implying that other defects in the p53 pathway are required for malignancy. We investigated mechanisms of wtp53 protein inactivation in malignant conversion of epithelial cells by comparing clonally related initiated cells with their derivative cancerous cells that have mdm2 amplification. Deficiencies in p53 accumulation and activities in response to DNA damage were not due simply to Mdm2 destabilization of p53 protein, but to continued association of DNA-bound p53 with Mdm2 protein and lack of binding and acetylation by p300 protein. The aberrant interactions were not because of mdm2 amplification alone, because DNA-bound p53 protein from initiated cells failed to bind ectopically expressed Mdm2 or endogenous overexpressed Mdm2 from cancerous cells. Phosphorylations of endogenous p53 at Ser18, -23, or -37 were insufficient to dissociate Mdm2, because each was induced by UV in cancerous cells. Interestingly, phospho-mimic p53-T21E did dissociate the Mdm2 protein from DNA-bound p53 and recovered p300 binding and p21 induction in the cancerous cells. Thus wtp53 in malignant cells with mdm2 amplification can be inactivated by continued association of DNA-bound p53 protein with Mdm2 and failure of p300 binding and acetylation, coupled with a defect in p53 phosphorylation at Thr21. These findings suggest therapeutic strategies that address both p53/Mdm2 interaction and associated p53 protein defects in human tumors that have amplified mdm2 genes.
Knights
Chad D
CD
Department of Dermatology, Oregon Health & Science University, Portland, Oregon 97239, USA.
Liu
Yuangang
Y
Appella
Ettore
E
Kulesz-Martin
Molly
M
D000123:Acetyltransferases / Q000378:metabolism; D000818:Animals; D018797:Cell Cycle Proteins / Q000378:metabolism; D002454:Cell Differentiation; D002460:Cell Line; D002461:Cell Line, Transformed; D045744:Cell Line, Tumor; D002471:Cell Transformation, Neoplastic; D004247:DNA / Q000378:metabolism; D004249:DNA Damage; D018076:DNA, Complementary / Q000378:metabolism; D004307:Dose-Response Relationship, Radiation; D004847:Epithelial Cells / Q000378:metabolism*; D005434:Flow Cytometry; D005784:Gene Amplification; D017930:Genes, Reporter; D016158:Genes, p53; D049452:Green Fluorescent Proteins; D018840:HSP70 Heat-Shock Proteins / Q000378:metabolism; D051548:Histone Acetyltransferases; D006801:Humans; D015151:Immunoblotting; D008164:Luminescent Proteins / Q000378:metabolism; D051379:Mice; D009687:Nuclear Proteins; D010766:Phosphorylation; D010957:Plasmids / Q000378:metabolism; D011233:Precipitin Tests; D011485:Protein Binding; D011499:Protein Processing, Post-Translational; D011518:Proto-Oncogene Proteins / Q000235:genetics*; D051736:Proto-Oncogene Proteins c-mdm2; D016283:Proto-Oncogene Proteins p21(ras) / Q000378:metabolism; D013912:Threonine / Q000737:chemistry; D013997:Time Factors; D014157:Transcription Factors; D015533:Transcriptional Activation; D014162:Transfection; D016159:Tumor Suppressor Protein p53 / Q000737:chemistry* / Q000378:metabolism; D014466:Ultraviolet Rays; D050880:p300-CBP Transcription Factors; D000099224:p300-CBP-Associated Factor
D000123:Acetyltransferases; D018797:Cell Cycle Proteins; D004247:DNA; D018076:DNA, Complementary; D049452:Green Fluorescent Proteins; D018840:HSP70 Heat-Shock Proteins; D051548:Histone Acetyltransferases; D008164:Luminescent Proteins; D009687:Nuclear Proteins; D011518:Proto-Oncogene Proteins; D051736:Proto-Oncogene Proteins c-mdm2; D016283:Proto-Oncogene Proteins p21(ras); D013912:Threonine; D014157:Transcription Factors; D016159:Tumor Suppressor Protein p53; D050880:p300-CBP Transcription Factors; D000099224:p300-CBP-Associated Factor; C497206:MDM2 protein, human; C497207:Mdm2 protein, mouse; C501009:HRAS protein, human