title: Identification of Daxx interacting with p73, one of the p53 family, and its regulation of p53 activity by competitive interaction with PML.
abstract: We performed a yeast two-hybrid screen using p73alpha, which is a member of the p53 family, as bait. We found that the p53 family members were functionally associated with Daxx, which was described originally as a cytoplasmic mediator of Fas signaling, but has been identified recently as a nuclear protein that co-localizes with the promyelocytic leukemia (PML) protein and regulates transcription. Extensive yeast two-hybrid assays indicated a physical interaction between a region including the oligomerization domain (OD) of p73alpha (amino acids 345-380) or p53 (amino acids 319-360) and amino acids 161-311 and 667-740 (C-terminal S/P/T-rich domain) of hDaxx, which is the common binding region of Fas, ASK1 and PML. This interaction was further confirmed by in vitro GST pull-down and in vivo immunoprecipitation assays. Both Daxx and p73/p53 co-localized in nuclear dot-like structures, which are probably nuclear PML oncogenic domains (PODs) or the nuclear domain NB10. Transient co-expression of Daxx resulted in strong inhibition of p73- and p53-mediated transcriptional activation of the synthetic p53-responsive and p21WAF1 promoters. Consequently, Gal4-Daxx repressed basal transcription in a dose-dependent manner. Treatment with trichostatin A, which is an inhibitor of histone deacetylase, or PML over-expression relieved Daxx-mediated transcriptional repression of p53. The mechanism underlying PML-mediated derepression appears to be competitive binding between Daxx, p53 and PML. Taken together, these findings delineate a transcriptional regulatory network that is modulated by differential Daxx-p53-PML interactions in the nuclear PODs. Therefore, Daxx is implicated in the regulation of the cell cycle and apoptosis through transcriptional regulation of p53 and possibly its family members.
authors.affiliation: Department of Bioscience and Biotechnology/Institute of Bioscience, Sejong University, 98 Kunja-dong, Kwangjin-gu, Seoul 143-747, Korea.
authors.forename: Eun-Joo
authors.initials: EJ
authors.lastname: Kim
authors.forename: Jong-Sup
authors.initials: JS
authors.lastname: Park
authors.forename: Soo-Jong
authors.initials: SJ
authors.lastname: Um
mesh_terms: D048868:Adaptor Proteins, Signal Transducing; D001665:Binding Sites / Q000235:genetics; D001667:Binding, Competitive / Q000187:drug effects; D015153:Blotting, Western; D002352:Carrier Proteins / Q000235:genetics / Q000378:metabolism*; D045744:Cell Line, Tumor; D002467:Cell Nucleus / Q000378:metabolism; D015500:Chloramphenicol O-Acetyltransferase / Q000235:genetics / Q000378:metabolism; D056970:Co-Repressor Proteins; D004268:DNA-Binding Proteins / Q000235:genetics / Q000378:metabolism*; D004791:Enzyme Inhibitors / Q000494:pharmacology; D016147:Genes, Tumor Suppressor; D005982:Glutathione Transferase / Q000235:genetics / Q000378:metabolism; D049452:Green Fluorescent Proteins; D056572:Histone Deacetylase Inhibitors; D006801:Humans; D006877:Hydroxamic Acids / Q000494:pharmacology; D047908:Intracellular Signaling Peptides and Proteins; D008164:Luminescent Proteins / Q000235:genetics / Q000378:metabolism; D018613:Microscopy, Confocal; D018832:Molecular Chaperones; D009363:Neoplasm Proteins / Q000235:genetics / Q000378:metabolism*; D009687:Nuclear Proteins / Q000235:genetics / Q000378:metabolism*; D000071156:Promyelocytic Leukemia Protein; D011485:Protein Binding / Q000187:drug effects; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism; D012441:Saccharomyces cerevisiae / Q000235:genetics; D014157:Transcription Factors / Q000235:genetics / Q000378:metabolism*; D014162:Transfection; D000072160:Tumor Protein p73; D016159:Tumor Suppressor Protein p53 / Q000235:genetics / Q000378:metabolism*; D025521:Tumor Suppressor Proteins; D020798:Two-Hybrid System Techniques
chemical_list: D048868:Adaptor Proteins, Signal Transducing; D002352:Carrier Proteins; D056970:Co-Repressor Proteins; C106985:DAXX protein, human; D004268:DNA-Binding Proteins; D004791:Enzyme Inhibitors; D056572:Histone Deacetylase Inhibitors; D006877:Hydroxamic Acids; D047908:Intracellular Signaling Peptides and Proteins; D008164:Luminescent Proteins; D018832:Molecular Chaperones; D009363:Neoplasm Proteins; D009687:Nuclear Proteins; D000071156:Promyelocytic Leukemia Protein; D011993:Recombinant Fusion Proteins; C000606460:TP73 protein, human; D014157:Transcription Factors; D000072160:Tumor Protein p73; D016159:Tumor Suppressor Protein p53; D025521:Tumor Suppressor Proteins; C070576:PML protein, human; D049452:Green Fluorescent Proteins; C012589:trichostatin A; D015500:Chloramphenicol O-Acetyltransferase; D005982:Glutathione Transferase