IntAct European Bioinformatics Institute IntAct intact http://www.ebi.ac.uk/ http://www.ebi.ac.uk/intact/query/${ac} EBI-[0-9]+|IA:[0-9]+ INTerAction database (IntAct) provides an open source database and toolkit for the storage, presentation and analysis of molecular interactions. http://www.ebi.ac.uk/intact European Bioinformatics Institute; Wellcome Trust Genome Campus; Hinxton, Cambridge; CB10 1SD; United Kingdom http://www.ebi.ac.uk/intact/ http://www.ebi.ac.uk/ zhao-2004-1 Negative regulation of p53 functions by Daxx and the involvement of MDM2. in vitro In vitro anti bait coip anti bait coimmunoprecipitation anti bait coip western blot western blot Immuno blot Immuno blot Zhao LY., Liu J., Sidhu GS., Niu Y., Liu Y., Wang R., Liao D. Apoptosis - Interactions involving proteins with a function related to apoptosis p53-null HCT116 cells were transfected with various combinations of expression plasmids. The authors confirmed by email that the species of origin of the MDM2 construct used was human. dliao@ufl.edu J. Biol. Chem. (0021-9258) 2004 Only protein-protein interactions imex curation Cancer - Interactions investigated in the context of cancer zhao-2004-2 Negative regulation of p53 functions by Daxx and the involvement of MDM2. yeasx Saccharomyces cerevisiae (Baker's yeast) Baker's yeast 2 hybrid two hybrid 2H 2h Gal4 transcription regeneration two-hybrid yeast two hybrid 2-hybrid classical two hybrid classical two hybrid 2H two-hybrid 2-hybrid Gal4 transcription regeneration 2h yeast two hybrid Y2H Y-2H 2 hybrid predetermined predetermined participant predetermined Zhao LY., Liu J., Sidhu GS., Niu Y., Liu Y., Wang R., Liao D. Apoptosis - Interactions involving proteins with a function related to apoptosis p53-null HCT116 cells were transfected with various combinations of expression plasmids. The authors confirmed by email that the species of origin of the MDM2 construct used was human. dliao@ufl.edu J. Biol. Chem. (0021-9258) 2004 Only protein-protein interactions imex curation Cancer - Interactions investigated in the context of cancer zhao-2004-5 Negative regulation of p53 functions by Daxx and the involvement of MDM2. mouse-embryonic_fibroblast Mus musculus embryonic fibroblasts embryonic_fibroblast Embryonic fibroblast Fibroblasts taken from an embryonic as opposed to a foetal or adult source. Foetus is defined as the stage at which tissue diferentiation is initiated. Tissue taken from an embryonic as opposed to a foetal or adult source. Embryo is defined as the development stage prior to organ formation. anti tag coip anti tag coimmunoprecipitation anti tag coip western blot western blot Immuno blot Immuno blot Zhao LY., Liu J., Sidhu GS., Niu Y., Liu Y., Wang R., Liao D. Apoptosis - Interactions involving proteins with a function related to apoptosis p53-null HCT116 cells were transfected with various combinations of expression plasmids. The authors confirmed by email that the species of origin of the MDM2 construct used was human. dliao@ufl.edu J. Biol. Chem. (0021-9258) 2004 Only protein-protein interactions imex curation Cancer - Interactions investigated in the context of cancer zhao-2004-6 Negative regulation of p53 functions by Daxx and the involvement of MDM2. mouse-embryonic_fibroblast Mus musculus embryonic fibroblasts embryonic_fibroblast Embryonic fibroblast Fibroblasts taken from an embryonic as opposed to a foetal or adult source. Foetus is defined as the stage at which tissue diferentiation is initiated. Tissue taken from an embryonic as opposed to a foetal or adult source. Embryo is defined as the development stage prior to organ formation. fluorescence imaging fluorescence microscopy fluorescence imaging immunostaining immunostaining Zhao LY., Liu J., Sidhu GS., Niu Y., Liu Y., Wang R., Liao D. Apoptosis - Interactions involving proteins with a function related to apoptosis p53-null HCT116 cells were transfected with various combinations of expression plasmids. The authors confirmed by email that the species of origin of the MDM2 construct used was human. dliao@ufl.edu J. Biol. Chem. (0021-9258) 2004 Only protein-protein interactions imex curation Cancer - Interactions investigated in the context of cancer zhao-2004-3 Negative regulation of p53 functions by Daxx and the involvement of MDM2. human-hct116 Human colon carcinoma HCT116 cells hct116 Human colon carcinoma line HCT116 anti tag coip anti tag coimmunoprecipitation anti tag coip western blot western blot Immuno blot Immuno blot Zhao LY., Liu J., Sidhu GS., Niu Y., Liu Y., Wang R., Liao D. Apoptosis - Interactions involving proteins with a function related to apoptosis p53-null HCT116 cells were transfected with various combinations of expression plasmids. The authors confirmed by email that the species of origin of the MDM2 construct used was human. dliao@ufl.edu J. Biol. Chem. (0021-9258) 2004 Only protein-protein interactions imex curation Cancer - Interactions investigated in the context of cancer zhao-2004-4 Negative regulation of p53 functions by Daxx and the involvement of MDM2. human-hct116 Human colon carcinoma HCT116 cells hct116 Human colon carcinoma line HCT116 anti bait coip anti bait coimmunoprecipitation anti bait coip western blot western blot Immuno blot Immuno blot Zhao LY., Liu J., Sidhu GS., Niu Y., Liu Y., Wang R., Liao D. Apoptosis - Interactions involving proteins with a function related to apoptosis p53-null HCT116 cells were transfected with various combinations of expression plasmids. The authors confirmed by email that the species of origin of the MDM2 construct used was human. dliao@ufl.edu J. Biol. Chem. (0021-9258) 2004 Only protein-protein interactions imex curation Cancer - Interactions investigated in the context of cancer daxx_human Death domain-associated protein 6 DAXX BING2 DAP6 Daxx Fas death domain-associated protein ETS1-associated protein 1 protein protein human Homo sapiens Human MATANSIIVLDDDDEDEAAAQPGPSHPLPNAASPGAEAPSSSEPHGARGSSSSGGKKCYKLENEKLFEEFLELCKMQTADHPEVVPFLYNRQQRAHSLFLASAEFCNILSRVLSRARSRPAKLYVYINELCTVLKAHSAKKKLNLAPAATTSNEPSGNNPPTHLSLDPTNAENTASQSPRTRGSRRQIQRLEQLLALYVAEIRRLQEKELDLSELDDPDSAYLQEARLKRKLIRLFGRLCELKDCSSLTGRVIEQRIPYRGTRYPEVNRRIERLINKPGPDTFPDYGDVLRAVEKAAARHSLGLPRQQLQLMAQDAFRDVGIRLQERRHLDLIYNFGCHLTDDYRPGVDPALSDPVLARRLRENRSLAMSRLDEVISKYAMLQDKSEEGERKKRRARLQGTSSHSADTPEASLDSGEGPSGMASQGCPSASRAETDDEDDEESDEEEEEEEEEEEEEATDSEEEEDLEQMQEGQEDDEEEDEEEEAAAGKDGDKSPMSSLQISNEKNLEPGKQISRSSGEQQNKGRIVSPSLLSEEPLAPSSIDAESNGEQPEELTLEEESPVSQLFELEIEALPLDTPSSVETDISSSRKQSEEPFTTVLENGAGMVSSTSFNGGVSPHNWGDSGPPCKKSRKEKKQTGSGPLGNSYVERQRSVHEKNGKKICTLPSPPSPLASLAPVADSSTRVDSPSHGLVTSSLCIPSPARLSQTPHSQPPRPGTCKTSVATQCDPEEIIVLSDSD p53_human Cellular tumor antigen p53 TP53 P53 Tumor suppressor p53 Phosphoprotein p53 Antigen NY-CO-13 protein protein human Homo sapiens Human MEEPQSDPSVEPPLSQETFSDLWKLLPENNVLSPLPSQAMDDLMLSPDDIEQWFTEDPGPDEAPRMPEAAPPVAPAPAAPTPAAPAPAPSWPLSSSVPSQKTYQGSYGFRLGFLHSGTAKSVTCTYSPALNKMFCQLAKTCPVQLWVDSTPPPGTRVRAMAIYKQSQHMTEVVRRCPHHERCSDSDGLAPPQHLIRVEGNLRVEYLDDRNTFRHSVVVPYEPPEVGSDCTTIHYNYMCNSSCMGGMNRRPILTIITLEDSSGNLLGRNSFEVRVCACPGRDRRTEEENLRKKGEPHHELPPGSTKRALPNNTSSSPQPKKKPLDGEYFTLQIRGRERFEMFRELNEALELKDAQAGKEPGGSRAHSSHLKSKKGQSTSRHKKLMFKTEGPDSD mdm2_human E3 ubiquitin-protein ligase Mdm2 MDM2 p53-binding protein Mdm2 Oncoprotein Mdm2 Double minute 2 protein RING-type E3 ubiquitin transferase Mdm2 protein protein human Homo sapiens Human MCNTNMSVPTDGAVTTSQIPASEQETLVRPKPLLLKLLKSVGAQKDTYTMKEVLFYLGQYIMTKRLYDEKQQHIVYCSNDLLGDLFGVPSFSVKEHRKIYTMIYRNLVVVNQQESSDSGTSVSENRCHLEGGSDQKDLVQELQEEKPSSSHLVSRPSTSSRRRAISETEENSDELSGERQRKRHKSDSISLSFDESLALCVIREICCERSSSSESTGTPSNPDLDAGVSEHSGDWLDQDSVSDQFSVEFEVESLDSEDYSLSEEGQELSDEDDEVYQVTVYQAGESDTDSFEEDPEISLADYWKCTSCNEMNPPLPSHCNRCWALRENWLPEDKGKDKGEISEKAKLENSTQAEEGFDVPDCKKTIVNDSRESCVEENDDKITQASQSQESEDYSQPSTSSSIIYSSQEDVKEFEREETQDKEESVESSLPLNAIEPCVICQGRPKNGCIVHGKTGHLMACFTCAKKLKKRNKPCPVCRQPIQMIVLTYFP tp53-daxx p53 coimmunoprecipitates Daxx in vitro 682111 N/A 682117 unspecified role unspecified role prey prey spofr-sf_9 Spodoptera frugiperda insect cells Fall armyworm sf_9 insect cells Derived from pupal ovarian tissue of the fall armyworm Cloned from the parent cell line, IPLB-SF-21-AE (SF-21) derived from immature ovaries of fall armyworm Spodoptera frugiperda pupae cells are susceptible to Baculovirus infection N/A 682118 unspecified role unspecified role bait bait direct interaction direct interaction false false false 1 A daxx-tp53 p53 coimmunoprecipitates with Daxx in vitro 682111 N/A 682118 unspecified role unspecified role prey prey N/A 682117 unspecified role unspecified role bait bait spofr-sf_9 Spodoptera frugiperda insect cells Fall armyworm sf_9 insect cells Derived from pupal ovarian tissue of the fall armyworm Cloned from the parent cell line, IPLB-SF-21-AE (SF-21) derived from immature ovaries of fall armyworm Spodoptera frugiperda pupae cells are susceptible to Baculovirus infection direct interaction direct interaction false false false 1 A daxx-tp53-1 p53 interacts with Daxx in a yeast two-hybrid assay 682112 N/A 682118 unspecified role unspecified role prey prey N/A 682117 unspecified role unspecified role bait bait region required to bind necessary binding region required to bind deletion analysis deletion analysis certain certain sequence position certain certain certain sequence position certain false physical association physical association false false false 1 B, C This indicates that the Daxx acidic domain (aa 434-496) is required for binding with p53. tp53-daxx-1 p53 interacts with Daxx in a yeast two-hybrid assay 682112 N/A 682118 unspecified role unspecified role bait bait P04637:p.Arg248Trp mutation disrupting mutation disrupting interaction mutation disrupting mutation analysis mutation analysis certain certain sequence position certain certain certain sequence position certain false c-terminal sufficient to bind sufficient binding region sufficient to bind deletion analysis deletion analysis certain certain sequence position certain certain certain sequence position certain false P04637:p.[Lys370Ala;Lys372Ala;Lys373Ala;Lys381Ala;Lys382Ala] mutation disrupting mutation disrupting interaction mutation disrupting mutation analysis mutation analysis certain certain sequence position certain certain certain sequence position certain false certain certain sequence position certain certain certain sequence position certain false certain certain sequence position certain certain certain sequence position certain false certain certain sequence position certain certain certain sequence position certain false certain certain sequence position certain certain certain sequence position certain false P04637:p.Lys386Arg mutation decreasing mutation decreasing interaction hotspot hotspot mutation decreasing mutation analysis mutation analysis certain certain sequence position certain certain certain sequence position certain false P04637:p.Lys386Glu mutation decreasing mutation decreasing interaction hotspot hotspot mutation decreasing mutation analysis mutation analysis certain certain sequence position certain certain certain sequence position certain false P04637:p.Lys319_Lys321delinsAlaAlaAla mutation disrupting mutation disrupting interaction mutation disrupting mutation analysis mutation analysis certain certain sequence position certain certain certain sequence position certain false P04637:p.Lys319_Lys321delinsGluGluGlu mutation disrupting mutation disrupting interaction mutation disrupting mutation analysis mutation analysis certain certain sequence position certain certain certain sequence position certain false P04637:p.[Lys372Ala;Lys373Ala] mutation disrupting mutation disrupting interaction mutation disrupting mutation analysis mutation analysis certain certain sequence position certain certain certain sequence position certain false certain certain sequence position certain certain certain sequence position certain false P04637:p.[Lys372Glu;Lys373Glu] mutation disrupting mutation disrupting interaction mutation disrupting mutation analysis mutation analysis certain certain sequence position certain certain certain sequence position certain false certain certain sequence position certain certain certain sequence position certain false P04637:p.[Lys381Ala;Lys382Ala] mutation disrupting mutation disrupting interaction mutation disrupting mutation analysis mutation analysis certain certain sequence position certain certain certain sequence position certain false certain certain sequence position certain certain certain sequence position certain false P04637:p.[Lys381Glu;Lys382Glu] mutation disrupting mutation disrupting interaction mutation disrupting mutation analysis mutation analysis certain certain sequence position certain certain certain sequence position certain false certain certain sequence position certain certain certain sequence position certain false P04637:p.[Lys370Ala;Lys372Ala;Lys373Ala] mutation disrupting mutation disrupting interaction mutation disrupting mutation analysis mutation analysis certain certain sequence position certain certain certain sequence position certain false certain certain sequence position certain certain certain sequence position certain false certain certain sequence position certain certain certain sequence position certain false P04637:p.[Lys370Glu;Lys372Glu;Lys373Glu] mutation disrupting mutation disrupting interaction mutation disrupting mutation analysis mutation analysis certain certain sequence position certain certain certain sequence position certain false certain certain sequence position certain certain certain sequence position certain false certain certain sequence position certain certain certain sequence position certain false P04637:p.[Lys372Glu;Lys373Glu;Lys381Glu;Lys382Glu] mutation disrupting mutation disrupting interaction mutation disrupting mutation analysis mutation analysis certain certain sequence position certain certain certain sequence position certain false certain certain sequence position certain certain certain sequence position certain false certain certain sequence position certain certain certain sequence position certain false certain certain sequence position certain certain certain sequence position certain false P04637:p.Ser376Glu mutation disrupting mutation disrupting interaction mutation disrupting mutation analysis mutation analysis certain certain sequence position certain certain certain sequence position certain false P04637:p.[Ser376Glu;Ser392Glu] mutation disrupting mutation disrupting interaction mutation disrupting mutation analysis mutation analysis certain certain sequence position certain certain certain sequence position certain false certain certain sequence position certain certain certain sequence position certain false P04637:p.[Lys319Arg;Lys320Arg;Lys321Arg;Lys372Arg;Lys373Arg;Lys381Arg;Lys382Arg] mutation decreasing mutation decreasing interaction hotspot hotspot mutation decreasing mutation analysis mutation analysis certain certain sequence position certain certain certain sequence position certain false certain certain sequence position certain certain certain sequence position certain false certain certain sequence position certain certain certain sequence position certain false certain certain sequence position certain certain certain sequence position certain false certain certain sequence position certain certain certain sequence position certain false certain certain sequence position certain certain certain sequence position certain false certain certain sequence position certain certain certain sequence position certain false N/A 682117 unspecified role unspecified role prey prey physical association physical association false false false 1 D, E, F, G Point mutations in the NH2 terminus of p53 did not affect binding to Daxx and nor did mutations in the core domain. However, C-terminal truncations of p53 abolished this interaction (aa 1-315 and 1-355), indicating the C-terminal regulatory domain is required for this interaction. daxx-tp53-3 Daxx coimmunoprecipitated p53 in DKO MEF lysates 682113 N/A 682118 unspecified role unspecified role prey prey P04637:p.[Leu22Gln;Trp23Ser] mutation decreasing mutation decreasing interaction hotspot hotspot mutation decreasing mutation analysis mutation analysis certain certain sequence position certain certain certain sequence position certain false certain certain sequence position certain certain certain sequence position certain false P04637:p.[Lys381Ala;Lys382Ala] mutation decreasing mutation decreasing interaction hotspot hotspot mutation decreasing mutation analysis mutation analysis certain certain sequence position certain certain certain sequence position certain false certain certain sequence position certain certain certain sequence position certain false N/A 682117 unspecified role unspecified role bait bait n-terminal flag tag flag tag FLAG-tagged FLAG DYKDDDDKV epitope tag DYKDDDDKV epitope tag FLAG FLAG-tagged n-term range n-terminal range n-term range n-term range n-terminal range n-term range false physical association physical association false false false 5 C MDM2 - Cotransfection with MDM2 significantly enhanced this interaction. Q9UER7; P04637 transfection daxx-mdm2-3 Daxx coimmunoprecipitated MDM2 in DKO MEF lysates 682113 N/A 682119 unspecified role unspecified role prey prey N/A 682117 unspecified role unspecified role bait bait n-terminal flag tag flag tag FLAG-tagged FLAG DYKDDDDKV epitope tag DYKDDDDKV epitope tag FLAG FLAG-tagged n-term range n-terminal range n-term range n-term range n-terminal range n-term range false physical association physical association false false false 5 D p53 - In the absence of p53 expression the amount of coprecipitated MDM2 was slightly reduced. Q9UER7; Q00987 transfection daxx-tp53-4 GFP-Daxx and p53 colocalize in DKO MEF cells 682114 N/A 682118 unspecified role unspecified role prey prey N/A 682117 unspecified role unspecified role bait bait region gfp tag green fluorescent protein tag GFP green fluorescent protein GFP green fluorescent protein gfp tag undetermined undetermined sequence position undetermined undetermined undetermined sequence position undetermined false colocalization colocalization false false false 5 E Daxx appeared in nuclear body-like structures as well as in the nucleolus, whereas both p53 and MDM2 were largely excluded from these locations. The immunostaining patterns of p53 or MDM2 were essentially the same, whether both together or each individually was coexpressed with GFP-Daxx. daxx-mdm2-4 GFP-Daxx and MDM2 colocalize in DKO MEF cells 682114 N/A 682117 unspecified role unspecified role bait bait region gfp tag green fluorescent protein tag GFP green fluorescent protein GFP green fluorescent protein gfp tag undetermined undetermined sequence position undetermined undetermined undetermined sequence position undetermined false N/A 682119 unspecified role unspecified role prey prey colocalization colocalization false false false 5 E Daxx appeared in nuclear body-like structures as well as in the nucleolus, whereas both p53 and MDM2 were largely excluded from these locations. The immunostaining patterns of p53 or MDM2 were essentially the same, whether both together or each individually was coexpressed with GFP-Daxx. daxx-tp53-2 FLAG-Daxx coimmunoprecipitates p53 and MDM2 in HCT116 lysates 682115 N/A 682119 unspecified role unspecified role prey prey N/A 682118 unspecified role unspecified role prey prey N/A 682117 unspecified role unspecified role bait bait n-terminal flag tag flag tag FLAG-tagged FLAG DYKDDDDKV epitope tag DYKDDDDKV epitope tag FLAG FLAG-tagged n-term range n-terminal range n-term range n-term range n-terminal range n-term range false n-terminal sufficient to bind sufficient binding region sufficient to bind certain certain sequence position certain certain certain sequence position certain false association association false false false 2 B, 3 B, C MDM2 - This interaction only occurred when cells were tansfected with MDM2. Q9UER7; P04637; Q00987 transfection Daxx constructs aa 1-574, 1-501, and a construct with deletion of aa 191-242 (a putative amphipathic helix) bound to p53 less effectively than wild-type Daxx. In addition, the Daxx construct with a deletion between aa 502 and 574 exhibited weak binding to p53. The Daxx NH2-terminal domain (aa 1-130) and the COOH-terminal constructs (aa 516-740 and 573-740) did not bind to p53. These results indicate that the acidic domain of Daxx is required for binding to p53. More MDM2 was precipitated with WT Daxx than with the Daxx fragments (aa 1-574 and 1-502), in parallel with the amounts of precipitated p53. Interestingly, although p53 was precipitated with Daxx Delta191-242 and Delta502-574 constructs, little MDM2 was coprecipitated with them. These results suggest that MDM2 may mediate the Daxx-p53 interaction, as a physical bridge and stabilizer of the complex, and by inhbiting covalent modifications of p53. Or, MDM2 may alter the conformation of Daxx, so exposing its p53-binding domain. tp53-daxx-2 p53 coimmunoprecipitates Daxx in HCT116 lysates 682116 N/A 682117 unspecified role unspecified role prey prey region ha tag ha tag YPYDVPDYA epitope tag YPYDVPDYA epitope tag undetermined undetermined sequence position undetermined undetermined undetermined sequence position undetermined false N/A 682118 unspecified role unspecified role bait bait P04637:p.[Lys381Ala;Lys382Ala] mutation decreasing mutation decreasing interaction hotspot hotspot mutation decreasing mutation analysis mutation analysis certain certain sequence position certain certain certain sequence position certain false certain certain sequence position certain certain certain sequence position certain false P04637:p.[Lys381Arg;Lys382Arg] mutation decreasing mutation decreasing interaction hotspot hotspot mutation decreasing mutation analysis mutation analysis certain certain sequence position certain certain certain sequence position certain false certain certain sequence position certain certain certain sequence position certain false P04637:p.Ser376Ala mutation decreasing mutation decreasing interaction hotspot hotspot mutation decreasing mutation analysis mutation analysis certain certain sequence position certain certain certain sequence position certain false P04637:p.Ser376Glu mutation decreasing mutation decreasing interaction hotspot hotspot mutation decreasing mutation analysis mutation analysis certain certain sequence position certain certain certain sequence position certain false P04637:p.[Lys381Glu;Lys382Glu] mutation disrupting mutation disrupting interaction mutation disrupting mutation analysis mutation analysis certain certain sequence position certain certain certain sequence position certain false certain certain sequence position certain certain certain sequence position certain false P04637:p.[Leu22Glu;Trp23Ser] mutation disrupting mutation disrupting interaction mutation disrupting mutation analysis mutation analysis certain certain sequence position certain certain certain sequence position certain false certain certain sequence position certain certain certain sequence position certain false physical association physical association false false false 4 B Q9UER7; P04637 transfection