The p53 Isoform Deltap53 lacks intrinsic transcriptional activity and reveals the critical role of nuclear import in dominant-negative activity.
The transcription factor p53 is one of the most frequently mutated tumor suppressors. Recent progress has unraveled several novel isoforms of p53. Intriguingly, one of the p53 isoform, Deltap53, which lacks part of the DNA binding domain, was reported to be transcriptionally active toward some p53 target genes and is critical for the intra-S phase checkpoint. Here, we show that, in contrast to full-length p53, ectopically expressed Deltap53 neither transactivated the promoters of p21(CIP1/WAF1) or murine double minute-2 (MDM2) nor repressed the cyclin B1 promoter in unstressed H1299 cells. Due to the deletion of a nuclear localization signal, Deltap53 was not imported into the nucleus. Engineering of nuclear localization signals to Deltap53 restored nuclear accumulation. However, the nuclear-targeting Deltap53 remained inactive, indicating that the lack of intrinsic activity of Deltap53 was not simply due to subcellular localization but to its incomplete DNA binding domain. Similar to p53, Deltap53 was subjected to MDM2-mediated ubiquitination/proteolysis. The cytoplasmic localization of Deltap53 correlated with the instability of the protein because forcing Deltap53 into the nucleus increased its stability. Although Deltap53 could form a complex with p53 and stimulated the cytoplasmic retention of p53, it was not a robust inhibitor of p53. Targeting Deltap53 into the nucleus enhanced the dominant-negative activity of Deltap53. These observations underscore the critical role of subcellular localization in the dominant-negative action of p53.
Chan
Wan Mui
WM
Department of Biochemistry, Hong Kong University of Science and Technology, Clear Water Bay, Hong Kong.
Poon
Randy Y C
RY
D021581:Active Transport, Cell Nucleus / Q000502:physiology; D002467:Cell Nucleus / Q000378:metabolism*; D050759:Cyclin-Dependent Kinase Inhibitor p21 / Q000378:metabolism; D003593:Cytoplasm / Q000378:metabolism; D006801:Humans; D019913:Nuclear Localization Signals; D011485:Protein Binding; D020033:Protein Isoforms; D011499:Protein Processing, Post-Translational; D051736:Proto-Oncogene Proteins c-mdm2 / Q000378:metabolism; D015533:Transcriptional Activation; D014407:Tumor Cells, Cultured; D016159:Tumor Suppressor Protein p53 / Q000378:metabolism*; D025801:Ubiquitin / Q000378:metabolism
C493956:CDKN1A protein, human; D050759:Cyclin-Dependent Kinase Inhibitor p21; D019913:Nuclear Localization Signals; D020033:Protein Isoforms; D016159:Tumor Suppressor Protein p53; D025801:Ubiquitin; C497206:MDM2 protein, human; D051736:Proto-Oncogene Proteins c-mdm2