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/ The PDZ protein tax-interacting protein-1 inhibits beta-catenin transcriptional activity and growth of colorectal cancer cells. The PDZ protein tax-interacting protein-1 inhibits beta-catenin transcriptional activity and growth of colorectal cancer cells. J. Biol. Chem. (0021-9258) 2003 Kanamori M., Sandy P., Marzinotto S., Benetti R., Kai C., Hayashizaki Y., Schneider C., Suzuki H. Apoptosis - Interactions involving proteins with a function related to apoptosis Cancer - Interactions investigated in the context of cancer A mammalian two hybrid assay was performed where interactions were monitored with the pG5luc reporter. rgscerg@gsc.riken.go.jp 27-NOV-2006: Contacted by JYOTI. A library of approximately 15,000 mouse full-length cDNAs fused in frame with the DNA binding domain of Gal4 was used. in vitro In vitro pull down pull down predetermined predetermined participant predetermined Kanamori M., Sandy P., Marzinotto S., Benetti R., Kai C., Hayashizaki Y., Schneider C., Suzuki H. Apoptosis - Interactions involving proteins with a function related to apoptosis A mammalian two hybrid assay was performed where interactions were monitored with the pG5luc reporter. rgscerg@gsc.riken.go.jp Accepted 2006-NOV-16 by JYOTI. 27-NOV-2006: Contacted by JYOTI. A library of approximately 15,000 mouse full-length cDNAs fused in frame with the DNA binding domain of Gal4 was used. J. Biol. Chem. (0021-9258) 2003 The PDZ protein tax-interacting protein-1 inhibits beta-catenin transcriptional activity and growth of colorectal cancer cells. The PDZ protein tax-interacting protein-1 inhibits beta-catenin transcriptional activity and growth of colorectal cancer cells. J. Biol. Chem. (0021-9258) 2003 Kanamori M., Sandy P., Marzinotto S., Benetti R., Kai C., Hayashizaki Y., Schneider C., Suzuki H. Apoptosis - Interactions involving proteins with a function related to apoptosis Cancer - Interactions investigated in the context of cancer A mammalian two hybrid assay was performed where interactions were monitored with the pG5luc reporter. rgscerg@gsc.riken.go.jp 27-NOV-2006: Contacted by JYOTI. A library of approximately 15,000 mouse full-length cDNAs fused in frame with the DNA binding domain of Gal4 was used. human-293 Homo sapiens transformed primary embryonal kidney cells Human 293 Transformed human primary embryonal kidney cells. The Cell type named 293 is synonymous with HEK-293. anti tag coip anti tag coimmunoprecipitation anti tag coip anti tag western anti tag western blot anti tag western Kanamori M., Sandy P., Marzinotto S., Benetti R., Kai C., Hayashizaki Y., Schneider C., Suzuki H. Apoptosis - Interactions involving proteins with a function related to apoptosis A mammalian two hybrid assay was performed where interactions were monitored with the pG5luc reporter. rgscerg@gsc.riken.go.jp Accepted 2006-NOV-16 by JYOTI. 27-NOV-2006: Contacted by JYOTI. A library of approximately 15,000 mouse full-length cDNAs fused in frame with the DNA binding domain of Gal4 was used. J. Biol. Chem. (0021-9258) 2003 The PDZ protein tax-interacting protein-1 inhibits beta-catenin transcriptional activity and growth of colorectal cancer cells. The PDZ protein tax-interacting protein-1 inhibits beta-catenin transcriptional activity and growth of colorectal cancer cells. J. Biol. Chem. (0021-9258) 2003 Kanamori M., Sandy P., Marzinotto S., Benetti R., Kai C., Hayashizaki Y., Schneider C., Suzuki H. Apoptosis - Interactions involving proteins with a function related to apoptosis Cancer - Interactions investigated in the context of cancer A mammalian two hybrid assay was performed where interactions were monitored with the pG5luc reporter. rgscerg@gsc.riken.go.jp 27-NOV-2006: Contacted by JYOTI. A library of approximately 15,000 mouse full-length cDNAs fused in frame with the DNA binding domain of Gal4 was used. human-u2-os Human U2-OS osteosarcoma cell line u2-os U2-OS osteocaroma cell line Not in CAPRI and no primary literature reference fluorescence imaging fluorescence microscopy fluorescence imaging immunostaining immunostaining Kanamori M., Sandy P., Marzinotto S., Benetti R., Kai C., Hayashizaki Y., Schneider C., Suzuki H. Apoptosis - Interactions involving proteins with a function related to apoptosis A mammalian two hybrid assay was performed where interactions were monitored with the pG5luc reporter. rgscerg@gsc.riken.go.jp Accepted 2006-NOV-16 by JYOTI. 27-NOV-2006: Contacted by JYOTI. A library of approximately 15,000 mouse full-length cDNAs fused in frame with the DNA binding domain of Gal4 was used. J. Biol. Chem. (0021-9258) 2003 The PDZ protein tax-interacting protein-1 inhibits beta-catenin transcriptional activity and growth of colorectal cancer cells. The PDZ protein tax-interacting protein-1 inhibits beta-catenin transcriptional activity and growth of colorectal cancer cells. J. Biol. Chem. (0021-9258) 2003 Kanamori M., Sandy P., Marzinotto S., Benetti R., Kai C., Hayashizaki Y., Schneider C., Suzuki H. Apoptosis - Interactions involving proteins with a function related to apoptosis Cancer - Interactions investigated in the context of cancer A mammalian two hybrid assay was performed where interactions were monitored with the pG5luc reporter. rgscerg@gsc.riken.go.jp 27-NOV-2006: Contacted by JYOTI. A library of approximately 15,000 mouse full-length cDNAs fused in frame with the DNA binding domain of Gal4 was used. crigr-cho_k1 Cricetulus griseus ovary K1 cho_k1 Chinese Hamster Ovary K1 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 Kanamori M., Sandy P., Marzinotto S., Benetti R., Kai C., Hayashizaki Y., Schneider C., Suzuki H. Apoptosis - Interactions involving proteins with a function related to apoptosis A mammalian two hybrid assay was performed where interactions were monitored with the pG5luc reporter. rgscerg@gsc.riken.go.jp Accepted 2006-NOV-16 by JYOTI. 27-NOV-2006: Contacted by JYOTI. A library of approximately 15,000 mouse full-length cDNAs fused in frame with the DNA binding domain of Gal4 was used. J. Biol. Chem. (0021-9258) 2003 The PDZ protein tax-interacting protein-1 inhibits beta-catenin transcriptional activity and growth of colorectal cancer cells. The PDZ protein tax-interacting protein-1 inhibits beta-catenin transcriptional activity and growth of colorectal cancer cells. J. Biol. Chem. (0021-9258) 2003 Kanamori M., Sandy P., Marzinotto S., Benetti R., Kai C., Hayashizaki Y., Schneider C., Suzuki H. Apoptosis - Interactions involving proteins with a function related to apoptosis Cancer - Interactions investigated in the context of cancer A mammalian two hybrid assay was performed where interactions were monitored with the pG5luc reporter. rgscerg@gsc.riken.go.jp 27-NOV-2006: Contacted by JYOTI. A library of approximately 15,000 mouse full-length cDNAs fused in frame with the DNA binding domain of Gal4 was used. crigr-cho_k1 Cricetulus griseus ovary K1 cho_k1 Chinese Hamster Ovary K1 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 nucleotide sequence nucleotide sequence identification sequence cloning sequence cloning nucleotide sequence Kanamori M., Sandy P., Marzinotto S., Benetti R., Kai C., Hayashizaki Y., Schneider C., Suzuki H. Apoptosis - Interactions involving proteins with a function related to apoptosis A mammalian two hybrid assay was performed where interactions were monitored with the pG5luc reporter. rgscerg@gsc.riken.go.jp Accepted 2006-NOV-16 by JYOTI. 27-NOV-2006: Contacted by JYOTI. A library of approximately 15,000 mouse full-length cDNAs fused in frame with the DNA binding domain of Gal4 was used. J. Biol. Chem. (0021-9258) 2003 rtkn_mouse Rhotekin Rtkn protein protein mouse Mus musculus Mouse MFSRNHRSRITVARGSALEMEFKRGRFRLSFFSESPEDTELQRKLDHEIRMRDGACKLLAACSQREQALEATKSLLVCNSRILSYMGELQRRKEAQVLEKTGRRPSDSVQPAQHSPCRGRVCISDLRIPLMWKDTEYFKNKGDLHRWAVFLLLQIGEQIQDTEMVLVDRTLTDISFQNNVLFAEAEPDFELRLELYGACVEEEGALAGAPKRLATKLSSSLGRSSGKRVRASLDSAGASGNSPVLLPTPAVGGPRFHLLAHTTLTLEEVQDGFRTHDLTLTSHEENPAWLPLYGSVCCRLAAQPLCMIQPTASGALRVQQAGELQNGTLVHGVLKGTNLFCYWRSEDADTGQEPLFTIVINKETRVRAGELEQAPEWPFTLSISNKYGDDEVTNTLQLESREALQNWMEALWQLFFDMSQWRHCCDEVMKIETPAPRKPPQALAKQGSLYHEMAIEPLDDIAAVTDILAQREGTRLEPSPPWLAMFTDQPALPSSCSPASVAPVPTWMQPLPWGRPRTFSLDAAPADHSLGPSRSVAPLPPQRSPKSRGFYSKSQLGPWLQSPV DIP protein Q8C6B2 original sequence version: 41 DIP protein Q8C6B2 original sequence version: 57 0934EEBF09B12A3E ctnb1_mouse Catenin beta-1 Beta-catenin Ctnnb1 Catnb protein protein mouse Mus musculus Mouse MATQADLMELDMAMEPDRKAAVSHWQQQSYLDSGIHSGATTTAPSLSGKGNPEEEDVDTSQVLYEWEQGFSQSFTQEQVADIDGQYAMTRAQRVRAAMFPETLDEGMQIPSTQFDAAHPTNVQRLAEPSQMLKHAVVNLINYQDDAELATRAIPELTKLLNDEDQVVVNKAAVMVHQLSKKEASRHAIMRSPQMVSAIVRTMQNTNDVETARCTAGTLHNLSHHREGLLAIFKSGGIPALVKMLGSPVDSVLFYAITTLHNLLLHQEGAKMAVRLAGGLQKMVALLNKTNVKFLAITTDCLQILAYGNQESKLIILASGGPQALVNIMRTYTYEKLLWTTSRVLKVLSVCSSNKPAIVEAGGMQALGLHLTDPSQRLVQNCLWTLRNLSDAATKQEGMEGLLGTLVQLLGSDDINVVTCAAGILSNLTCNNYKNKMMVCQVGGIEALVRTVLRAGDREDITEPAICALRHLTSRHQEAEMAQNAVRLHYGLPVVVKLLHPPSHWPLIKATVGLIRNLALCPANHAPLREQGAIPRLVQLLVRAHQDTQRRTSMGGTQQQFVEGVRMEEIVEGCTGALHILARDVHNRIVIRGLNTIPLFVQLLYSPIENIQRVAAGVLCELAQDKEAAEAIEAEGATAPLTELLHSRNEGVATYAAAVLFRMSEDKPQDYKKRLSVELTSSLFRTEPMAWNETADLGLDIGAQGEALGYRQDDPSYRSFHSGGYGQDALGMDPMMEHEMGGHHPGADYPVDGLPDLGHAQDLMDGLPPGDSNQLAWFDTDL DIP protein Q02248 original sequence version: 128 DIP protein Q02248 original sequence version: 135 DIP protein Q02248 original sequence version: 147 DIP protein Q02248 original sequence version: 187 DIP protein Q02248 original sequence version: 140 D708F170A3FBED6E q82235_9dela tax protein protein humt- Human T-lymphotropic virus 1 MAHFPGFRQSLLFGYPVYVFGDCVQGDWCPISGGLCSARLHRHALLATCPEHQITWDPIDGRVIGSALQFLIPRLPSFPTQRTSKTLKVLTPPITHTTPNIPPSFLQAMRKYSPFRNGYMEPTLGQHLPTLSFPDPGLRPQNLYTLWGGSVVCMYLYQLSPPITWPLLPHVIFCHPGQLGAFLTNVPYKRIEELLYKISLTTGALIILPEDCLPTTLFQPARAPVTLTAWQNGLLPFHSTLTTPGLIWTFTDGTPMISGPCPEDGQPSLVLQSSSFIFHKFQTKAYHPSFLLSHGLIQYSSFHSLHLLFEEYTNIPISLLFNEKEADDNDHEPQISPGGLEPPSEKHFRETEV A59CF71C183E7975 tx1b3_mouse Tax1-binding protein 3 Tax interaction protein 1 Tax1bp3 protein protein mouse Mus musculus Mouse MSYTPGQPVTAVVQRVEIHKLRQGENLILGFSIGGGIDQDPSQNPFSEDKTDKGIYVTRVSEGGPAEIAGLQIGDKIMQVNGWDMTMVTHDQARKRLTKRSEEVVRLLVTRQSLQKAVQQSMLS F3FBD0F475F69FCD tax1bp3-ctnnb1 5 7 unspecified role unspecified role prey prey region tag tag undetermined undetermined sequence position undetermined undetermined undetermined sequence position undetermined 9 unspecified role unspecified role bait bait gal4 dna bd tag tag undetermined undetermined sequence position undetermined undetermined undetermined sequence position undetermined physical association physical association 1 A we screened out the murine homologue of the PDZ domain protein, TIP-1, as a novel binding partner of -catenin tax1bp3-ctnnb1-1 2 7 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 9 unspecified role unspecified role bait bait region myc tag myc tag EUKLISEED epitope tag EUKLISEED epitope tag undetermined undetermined sequence position undetermined undetermined undetermined sequence position undetermined physical association physical association 1 B coimmunoprecipitation experiments were performed on lysates from 293 cells expressing Myc-tagged TIP-1 and HA-tagged -catenin. As shown in Fig. 1B, -catenin was readily observed in the anti-Myc immunocomplex. tax1bp3-ctnnb1-2 3 7 unspecified role unspecified role neutral component neutral component region ha tag ha tag YPYDVPDYA epitope tag YPYDVPDYA epitope tag undetermined undetermined sequence position undetermined undetermined undetermined sequence position undetermined 9 unspecified role unspecified role neutral component neutral component region myc tag myc tag EUKLISEED epitope tag EUKLISEED epitope tag undetermined undetermined sequence position undetermined undetermined undetermined sequence position undetermined colocalization colocalization 1 C These results suggest that -catenin and TIP-1 interact in vivo and that complex formation probably takes place in the nucleus. tax1bp3-rtkn 1 6 unspecified role unspecified role prey prey region 35s radiolabel 35s radiolabel 35S s35 radiolabelled S35 s35 radiolabelled 35S S35 undetermined undetermined sequence position undetermined undetermined undetermined sequence position undetermined region sufficient to bind sufficient binding region sufficient to bind certain certain sequence position certain certain certain sequence position certain in vitro In vitro 9 unspecified role unspecified role bait bait region gst tag glutathione s tranferase tag glutathione S-tranferase tag glutathione S-tranferase tag gst tag undetermined undetermined sequence position undetermined undetermined undetermined sequence position undetermined ecobb Escherichia coli (strain B / BL21) direct interaction direct interaction 1 D we performed pull-down assays with purified GSTTIP- 1 in the presence of in vitro translated -catenin. As positive controls, we used in vitro translated human T-cell lymphotrophic virus-1 Tax and rhotekin, two previously described binding partners of TIP-1 tax1bp3-ctnnb1-3 1 7 unspecified role unspecified role prey prey region 35s radiolabel 35s radiolabel 35S s35 radiolabelled S35 s35 radiolabelled 35S S35 undetermined undetermined sequence position undetermined undetermined undetermined sequence position undetermined c-terminal PDZ-recognition motif DTDL binding region binding-associated region binding region certain certain sequence position certain certain certain sequence position certain in vitro In vitro 9 unspecified role unspecified role bait bait region gst tag glutathione s tranferase tag glutathione S-tranferase tag glutathione S-tranferase tag gst tag undetermined undetermined sequence position undetermined undetermined undetermined sequence position undetermined ecobb Escherichia coli (strain B / BL21) direct interaction direct interaction 1 D, 2 B a strong and specific interaction was detected between TIP-1 and -catenin, suggesting that -catenin physically associates with TIP-1. the readily observed interaction between -catenin and TIP-1 was almost completely abolished by the removal of the DTDL sequence from the C terminus of -catenin tax1bp3-tax 1 8 unspecified role unspecified role prey prey region 35s radiolabel 35s radiolabel 35S s35 radiolabelled S35 s35 radiolabelled 35S S35 undetermined undetermined sequence position undetermined undetermined undetermined sequence position undetermined in vitro In vitro 9 unspecified role unspecified role bait bait region gst tag glutathione s tranferase tag glutathione S-tranferase tag glutathione S-tranferase tag gst tag undetermined undetermined sequence position undetermined undetermined undetermined sequence position undetermined ecobb Escherichia coli (strain B / BL21) direct interaction direct interaction 1 D As positive controls, we used in vitro translated human T-cell lymphotrophic virus-1 Tax and rhotekin, two previously described binding partners of TIP-1 tax1bp3-ctnnb1-4 4 7 unspecified role unspecified role prey prey region tag tag undetermined undetermined sequence position undetermined undetermined undetermined sequence position undetermined 9 unspecified role unspecified role bait bait gal4 dna bd tag tag undetermined undetermined sequence position undetermined undetermined undetermined sequence position undetermined region sufficient to bind sufficient binding region sufficient to bind certain certain sequence position certain certain certain sequence position certain physical association physical association 2 A Various deletion mutants of TIP-1 were tested in an in vivo interaction assay with coexpressed -catenin, and the obtained results clearly demonstrated the requirement of the intact PDZ domain for complex formation (Fig. 2A). Interestingly, although the removal of the first 12 amino acids of TIP-1 had no significant effect on the interaction with -catenin, deletion of the last eight residues resulted in a 3-fold reduction of binding efficiency, suggesting a possible role for this protein region in the formation or stability of the complex. These results indicate the PDZ domain of TIP-1 is required for complex formation with beta-catenin and that the last 8 residues of TIP-1 may be involved in complex formation. tax1bp3-ctnnb1-5 4 9 unspecified role unspecified role bait bait gal4 dna bd tag tag undetermined undetermined sequence position undetermined undetermined undetermined sequence position undetermined 7 unspecified role unspecified role prey prey region tag tag undetermined undetermined sequence position undetermined undetermined undetermined sequence position undetermined c-terminal contains PDZ-recognition motif DTDL sufficient to bind sufficient binding region sufficient to bind certain certain sequence position certain certain certain sequence position certain region Armadillo repeats 2-8 sufficient to bind sufficient binding region sufficient to bind certain certain sequence position certain certain certain sequence position certain physical association physical association 2 D these results clearly demonstrate a direct physical association between the extreme C terminus of -catenin and the PDZ domain of TIP-1 and reveal an alternative mechanism for complex formation in vivo. These results indicate that as well as the interaction between beta-catenin and TIP-1 involving the 4 C-terminal resicues of beta-catenin, in vivo the residues 173-483 containing Armadillo repeats 2-8 can also allow complex formation.