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/ Six3-mediated auto repression and eye development requires its interaction with members of the Groucho-related family of co-repressors. Six3-mediated auto repression and eye development requires its interaction with members of the Groucho-related family of co-repressors. Development (0950-1991) 2002 imex curation Zhu CC., Dyer MA., Uchikawa M., Kondoh H., Lagutin OV., Oliver G. guillermo.oliver@stjude.org 23-Feb-2010: Contacted by JYOTI. E10.5 mouse cDNA orchard Only protein-protein interactions 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 nucleotide sequence nucleotide sequence identification sequence cloning sequence cloning nucleotide sequence Zhu CC., Dyer MA., Uchikawa M., Kondoh H., Lagutin OV., Oliver G. guillermo.oliver@stjude.org Accepted 2009-MAY-6 by ORCHARD. 23-Feb-2010: Contacted by JYOTI. E10.5 mouse cDNA Development (0950-1991) 2002 orchard Only protein-protein interactions imex curation Six3-mediated auto repression and eye development requires its interaction with members of the Groucho-related family of co-repressors. Six3-mediated auto repression and eye development requires its interaction with members of the Groucho-related family of co-repressors. Development (0950-1991) 2002 imex curation Zhu CC., Dyer MA., Uchikawa M., Kondoh H., Lagutin OV., Oliver G. guillermo.oliver@stjude.org 23-Feb-2010: Contacted by JYOTI. E10.5 mouse cDNA orchard Only protein-protein interactions in vitro In vitro pull down pull down weight autoradiogra molecular weight estimation by autoradiography weight autoradiogra Zhu CC., Dyer MA., Uchikawa M., Kondoh H., Lagutin OV., Oliver G. guillermo.oliver@stjude.org Accepted 2009-MAY-6 by ORCHARD. 23-Feb-2010: Contacted by JYOTI. E10.5 mouse cDNA Development (0950-1991) 2002 orchard Only protein-protein interactions imex curation Six3-mediated auto repression and eye development requires its interaction with members of the Groucho-related family of co-repressors. Six3-mediated auto repression and eye development requires its interaction with members of the Groucho-related family of co-repressors. Development (0950-1991) 2002 imex curation Zhu CC., Dyer MA., Uchikawa M., Kondoh H., Lagutin OV., Oliver G. guillermo.oliver@stjude.org 23-Feb-2010: Contacted by JYOTI. E10.5 mouse cDNA orchard Only protein-protein interactions mouse-3t3 Mus musculus embryo cells 3t3 Mouse embryo fibroblast cells anti tag coip anti tag coimmunoprecipitation anti tag coip western blot western blot Immuno blot Immuno blot Zhu CC., Dyer MA., Uchikawa M., Kondoh H., Lagutin OV., Oliver G. guillermo.oliver@stjude.org Accepted 2009-MAY-6 by ORCHARD. 23-Feb-2010: Contacted by JYOTI. E10.5 mouse cDNA Development (0950-1991) 2002 orchard Only protein-protein interactions imex curation Six3-mediated auto repression and eye development requires its interaction with members of the Groucho-related family of co-repressors. Six3-mediated auto repression and eye development requires its interaction with members of the Groucho-related family of co-repressors. Development (0950-1991) 2002 imex curation Zhu CC., Dyer MA., Uchikawa M., Kondoh H., Lagutin OV., Oliver G. guillermo.oliver@stjude.org 23-Feb-2010: Contacted by JYOTI. E10.5 mouse cDNA orchard Only protein-protein interactions mouse-3t3 Mus musculus embryo cells 3t3 Mouse embryo fibroblast cells confocal microscopy confocal microscopy immunostaining immunostaining Zhu CC., Dyer MA., Uchikawa M., Kondoh H., Lagutin OV., Oliver G. guillermo.oliver@stjude.org Accepted 2009-MAY-6 by ORCHARD. 23-Feb-2010: Contacted by JYOTI. E10.5 mouse cDNA Development (0950-1991) 2002 orchard Only protein-protein interactions imex curation Six3-mediated auto repression and eye development requires its interaction with members of the Groucho-related family of co-repressors. Six3-mediated auto repression and eye development requires its interaction with members of the Groucho-related family of co-repressors. Development (0950-1991) 2002 imex curation Zhu CC., Dyer MA., Uchikawa M., Kondoh H., Lagutin OV., Oliver G. guillermo.oliver@stjude.org 23-Feb-2010: Contacted by JYOTI. E10.5 mouse cDNA orchard Only protein-protein interactions 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 Zhu CC., Dyer MA., Uchikawa M., Kondoh H., Lagutin OV., Oliver G. guillermo.oliver@stjude.org Accepted 2009-MAY-6 by ORCHARD. 23-Feb-2010: Contacted by JYOTI. E10.5 mouse cDNA Development (0950-1991) 2002 orchard Only protein-protein interactions imex curation Six3-mediated auto repression and eye development requires its interaction with members of the Groucho-related family of co-repressors. Six3-mediated auto repression and eye development requires its interaction with members of the Groucho-related family of co-repressors. Development (0950-1991) 2002 imex curation Zhu CC., Dyer MA., Uchikawa M., Kondoh H., Lagutin OV., Oliver G. guillermo.oliver@stjude.org 23-Feb-2010: Contacted by JYOTI. E10.5 mouse cDNA orchard Only protein-protein interactions mouse-3t3 Mus musculus embryo cells 3t3 Mouse embryo fibroblast cells anti bait coip anti bait coimmunoprecipitation anti bait coip anti tag western anti tag western blot anti tag western Zhu CC., Dyer MA., Uchikawa M., Kondoh H., Lagutin OV., Oliver G. guillermo.oliver@stjude.org Accepted 2009-MAY-6 by ORCHARD. 23-Feb-2010: Contacted by JYOTI. E10.5 mouse cDNA Development (0950-1991) 2002 orchard Only protein-protein interactions imex curation tle4_mouse Transducin-like enhancer protein 4 Tle4 Grg4 Groucho-related protein 4 Grg-4 protein protein mouse Mus musculus Mouse MIRDLSKMYPQTRHPAPHQPAQPFKFTISESCDRIKEEFQFLQAQYHSLKLECEKLASEKTEMQRHYVMYYEMSYGLNIEMHKQAEIVKRLNAICAQVIPFLSQEHQQQVVQAVERAKQVTMAELNAIIGQQLQAQHLSHGHGLPVPLTPHPSGLQPPAIPPIGSSAGLLALSSALGGQSHLPIKDEKKHHDNDHQRDRDSIKSSSVSPSASFRGSEKHRNSTDYSSESKKQKTEEKEIAARYDSDGEKSDDNLVVDVSNEDPSSPRGSPAHSPRENGLDKTRLLKKDAPISPASVASSSSTPSSKSKELSLNEKSTTPVSKSNTPTPRTDAPTPGSNSTPGLRPVPGKPPGVDPLASSLRTPMAVPCPYPTPFGIVPHAGMNGELTSPGAAYAGLHNISPQMSAAAAAAAAAAAYGRSPVVGFDPHHHMRVPAIPPNLTGIPGGKPAYSFHVSADGQMQPVPFPPDALIGPGIPRHARQINTLNHGEVVCAVTISNPTRHVYTGGKGCVKVWDISHPGNKSPVSQLDCLNRDNYIRSCRLLPDGRTLIVGGEASTLSIWDLAAPTPRIKAELTSSAPACYALAISPDSKVCFSCCSDGNIAVWDLHNQTLVRQFQGHTDGASCIDISNDGTKLWTGGLDNTVRSWDLREGRQLQQHDFTSQIFSLGYCPTGEWLAVGMENSNVEVLHVTKPDKYQLHLHESCVLSLKFAHCGKWFVSTGKDNLLNAWRTPYGASIFQSKESSSVLSCDISVDDKYIVTGSGDKKATVYEVIY 5A0090789774CACC so_drome Protein sine oculis so CG11121 protein protein drome Drosophila melanogaster (Fruit fly) Fruit fly MLQHPATDFYDLAAANAAAVLTARHTPPYSPTGLSGSVALHNNNNNNSSTSNNNNSTLDIMAHNGGGAGGGLHLNSSSNGGGGGGVVSGGGSGGRENLPSFGFTQEQVACVCEVLQQAGNIERLGRFLWSLPQCDKLQLNESVLKAKAVVAFHRGQYKELYRLLEHHHFSAQNHAKLQALWLKAHYVEAEKLRGRPLGAVGKYRVRRKFPLPRTIWDGEETSYCFKEKSRSVLRDWYSHNPYPSPREKRDLAEATGLTTTQVSNWFKNRRQRDRAAEHKDGSTDKQHLDSSSDSEMEGSMLPSQSAQHQQQQQQQQHSPGNSSGNNNGLHQQQLQHVAAEQGLQHHPHQPHPASNIANVAATKSSGGGGGGGVSAAAAAQMQMPPLTAAVAYSHLHSVMGAMPMTAMYDMGEYQHL 669E4BD95B857996 six3_mouse Homeobox protein SIX3 Six3 Sine oculis homeobox homolog 3 protein protein mouse Mus musculus Mouse MVFRSPLDLYSSHFLLPNFADSHHCSLLLASSGGGSGASGGGGGAGGGGGGNRAGGGGAGGAGGGSGGGGSRAPPEELSMFQLPTLNFSPEQVASVCETLEETGDIERLGRFLWSLPVAPGACEAINKHESILRARAVVAFHTGNFRDLYHILENHKFTKESHGKLQAMWLEAHYQEAEKLRGRPLGPVDKYRVRKKFPLPRTIWDGEQKTHCFKERTRSLLREWYLQDPYPNPSKKRELAQATGLTPTQVGNWFKNRRQRDRAAAAKNRLQHQAIGPSGMRSLAEPGCPTHGSAESPSTAASPTTSVSSLTERADTGTSILSVTSSDSECDV DIP protein Q62233 original sequence version: 74 1AD7D3C4388043B9 tle5_mouse TLE family member 5 Tle5 Aes Amino-terminal enhancer of split Esp1 Grg Grg-5 Grg5 Groucho-related protein 5 Protein ESP1 Protein GRG protein protein mouse Mus musculus Mouse MMFPQSRHSGSSHLPQQLKFTTSDSCDRIKDEFQLLQAQYHSLKLECDKLASEKSEMQRHYVMYYEMSYGLNIEMHKQAEIVKRLNGICAQVLPYLSQEHQQQVLGAIERAKQVTAPELNSIIRQQLQAHQLSQLQALALPLTPLPVGLQPPSLPAVSAGTGLLSLSALGSQTHLSKEDKNGHDGDTHQEDDGEKSD 2C00233AA6921A9F six6_mouse Homeobox protein SIX6 Six6 Optx2 Six9 Sine oculis homeobox homolog 6 Optic homeobox 2 Six9 protein protein protein mouse Mus musculus Mouse MFQLPILNFSPQQVAGVCETLEESGDVERLGRFLWSLPVAPAACEALNKNESVLRARAIVAFHGGNYRELYHILENHKFTKESHAKLQALWLEAHYQEAEKLRGRPLGPVDKYRVRKKFPLPRTIWDGEQKTHCFKERTRHLLREWYLQDPYPNPSKKRELAQATGLTPTQVGNWFKNRRQRDRAAAAKNRLQQQVLSQGPGRVLRSEGEGTPEVLGVASSPAASLSSKAATSAISITSSDSECDI F1332D5E617B2CF1 optix_drome Protein Optix Optix opt Six3 Homeobox protein SIX3 CG18455 protein protein drome Drosophila melanogaster (Fruit fly) Fruit fly MAVGPTEGKQPPSESFSPTHHQIIAPSPILAVPTLAFSAAQVEIVCKTLEDSGDIERLARFLWSLPVALPNMHEILNCEAVLRARAVVAYHVGNFRELYAIIENHKFTKASYGKLQAMWLEAHYIEAEKLRGRSLGPVDKYRVRKKFPLPPTIWDGEQKTHCFKERTRSLLREWYLQDPYPNPTKKRELAKATGLNPTQVGNWFKNRRQRDRAAAAKNRIQHSQNSSGMGCRSRRADGAASPTPSDSSDSDISLGTHSPVPSSLQLQHSPGSTSNGANDREESLSVDDDKPRDLSGSLPLPLSLPLPLASPTHTPPQLPPGYGGGAGAGPGGPLTGPGCLPPFKLDAATSLFSAGCYLQSFSNLKEMSQQFPIQPIVLRPHPQLPQSLALNGASGGPPLHHPAYAAAYSVECVPGGHGPPHPPPKLRINSPEKLNSTAVAAAASVGGGGGNQHHEPTTTGYHHSGQLMLHRPFSTSPELKHSAPEIT ECA9FD59F171E599 grou_drome Protein groucho gro E(spl)m9/m10 CG8384 Enhancer of split m9/10 protein protein protein drome Drosophila melanogaster (Fruit fly) Fruit fly MYPSPVRHPAAGGPPPQGPIKFTIADTLERIKEEFNFLQAQYHSIKLECEKLSNEKTEMQRHYVMYYEMSYGLNVEMHKQTEIAKRLNTLINQLLPFLQADHQQQVLQAVERAKQVTMQELNLIIGHQQQHGIQQLLQQIHAQQVPGGPPQPMGALNPFGALGATMGLPHGPQGLLNKPPEHHRPDIKPTGLEGPAAAEERLRNSVSPADREKYRTRSPLDIENDSKRRKDEKLQEDEGEKSDQDLVVDVANEMESHSPRPNGEHVSMEVRDRESLNGERLEKPSSSGIKQERPPSRSGSSSSRSTPSLKTKDMEKPGTPGAKARTPTPNAAAPAPGVNPKQMMPQGPPPAGYPGAPYQRPADPYQRPPSDPAYGRPPPMPYDPHAHVRTNGIPHPSALTGGKPAYSFHMNGEGSLQPVPFPPDALVGVGIPRHARQINTLSHGEVVCAVTISNPTKYVYTGGKGCVKVWDISQPGNKNPVSQLDCLQRDNYIRSVKLLPDGRTLIVGGEASNLSIWDLASPTPRIKAELTSAAPACYALAISPDSKVCFSCCSDGNIAVWDLHNEILVRQFQGHTDGASCIDISPDGSRLWTGGLDNTVRSWDLREGRQLQQHDFSSQIFSLGYCPTGDWLAVGMENSHVEVLHASKPDKYQLHLHESCVLSLRFAACGKWFVSTGKDNLLNAWRTPYGASIFQSKETSSVLSCDISTDDKYIVTGSGDKKATVYEVIY DIP protein P16371 original sequence version: 150 DIP protein P16371 original sequence version: 117 9DA08E1ED343B2F5 six3-grg5 1 9 unspecified role unspecified role bait bait 10 unspecified role unspecified role prey prey physical association physical association In the initial screening, two of 1.2 million clones encoded a protein that specifically interacted with Six3 but not with the unrelated protein Raf1 or the pc97 vector containing only the Gal4-DB (data not shown). Sequence analysis identified one of the Gal4-AD in-frame fusion proteins as the 197 amino acid full-length Grg5 grg5-six3 2 9 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 10 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 ecoli Escherichia coli (strain K12) direct interaction direct interaction 2 GST pull-down experiments were performed to examine the interaction between Grg5 and the murine Six/So family members Six6 and Six2, and Drosophila sine oculis (So). Six6 and Drosophila So, like Six3, interacted with Grg5; grg5-six6 2 10 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 ecoli Escherichia coli (strain K12) 11 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 deacetylation deacetylation reaction deacetylation 2 GST pull-down experiments were performed to examine the interaction between Grg5 and the murine Six/So family members Six6 and Six2, and Drosophila sine oculis (So). Six6 and Drosophila So, like Six3, interacted with Grg5; grg5-so 2 10 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 ecoli Escherichia coli (strain K12) 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 direct interaction direct interaction 2 GST pull-down experiments were performed to examine the interaction between Grg5 and the murine Six/So family members Six6 and Six2, and Drosophila sine oculis (So). Six6 and Drosophila So, like Six3, interacted with Grg5; grg4-six3 2 9 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 Q62233:p.Phe88Glu mutation decreasing mutation decreasing interaction hotspot hotspot mutation decreasing certain certain sequence position certain certain certain sequence position certain F E Wed May 02 17:03:50 BST 2018 This feature has been corrected as a result of our quality control procedures. The original label was 'phe88glu' Tue Jun 30 18:03:19 BST 2020 This feature has been corrected as a result of our quality control procedures. The original label was 'p.Phe88Glu' in vitro In vitro 7 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 ecoli Escherichia coli (strain K12) direct interaction direct interaction 2 We extended these initial experiments and demonstrated that mouse Grg4 and fly Groucho proteins can also interact strongly with mouse Six3 and Six6 (Fig. 2B). Although much weaker, interactions between mouse Grg4 and fly optix were also detected (Fig. 2B). These results suggest that mammalian Six3 can directly interact with members of the Groucho family in vitro. grg4-six6 2 7 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 ecoli Escherichia coli (strain K12) 11 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 direct interaction direct interaction 2 We extended these initial experiments and demonstrated that mouse Grg4 and fly Groucho proteins can also interact strongly with mouse Six3 and Six6 (Fig. 2B). Although much weaker, interactions between mouse Grg4 and fly optix were also detected (Fig. 2B). These results suggest that mammalian Six3 can directly interact with members of the Groucho family in vitro. grg4-optix 2 12 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 7 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 ecoli Escherichia coli (strain K12) direct interaction direct interaction 2 We extended these initial experiments and demonstrated that mouse Grg4 and fly Groucho proteins can also interact strongly with mouse Six3 and Six6 (Fig. 2B). Although much weaker, interactions between mouse Grg4 and fly optix were also detected (Fig. 2B). These results suggest that mammalian Six3 can directly interact with members of the Groucho family in vitro. six3-groucho 2 13 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 ecoli Escherichia coli (strain K12) direct interaction direct interaction 2 We extended these initial experiments and demonstrated that mouse Grg4 and fly Groucho proteins can also interact strongly with mouse Six3 and Six6 (Fig. 2B). Although much weaker, interactions between mouse Grg4 and fly optix were also detected (Fig. 2B). These results suggest that mammalian Six3 can directly interact with members of the Groucho family in vitro. six3-grg5-1 5 10 unspecified role unspecified role prey prey 9 unspecified role unspecified role bait bait region sufficient to bind sufficient binding region sufficient to bind deletion analysis deletion analysis certain certain sequence position certain certain certain sequence position certain physical association physical association 3A Mapping of the interaction domains of Six3 and Grg5 using a yeast two-hybrid assay. grg5-six3-1 5 9 unspecified role unspecified role bait bait 10 unspecified role unspecified role prey prey region sufficient to bind sufficient binding region sufficient to bind deletion analysis deletion analysis certain certain sequence position certain certain certain sequence position certain physical association physical association 3B The fragment containing the Q domain and four amino acids of the SP domain of Grg5 (Grg51-134) interacted with Six3 grg5-six3-2 4 9 unspecified role unspecified role neutral component neutral component 10 unspecified role unspecified role neutral component neutral component region flag tag flag tag FLAG-tagged FLAG DYKDDDDKV epitope tag DYKDDDDKV epitope tag FLAG FLAG-tagged undetermined undetermined sequence position undetermined undetermined undetermined sequence position undetermined colocalization colocalization 4B Confocal images showing the colocalization of Grg5 and Six3 in the nuclei of transfected NIH3T3 cells. six3-grg4 3 9 unspecified role unspecified role prey prey 7 unspecified role unspecified role bait bait region flag tag flag tag FLAG-tagged FLAG DYKDDDDKV epitope tag DYKDDDDKV epitope tag FLAG FLAG-tagged undetermined undetermined sequence position undetermined undetermined undetermined sequence position undetermined physical association physical association 2C Mouse Six3 bound to Grg4 in mammalian cells. six3-groucho-1 6 9 unspecified role unspecified role bait bait 13 unspecified role unspecified role prey prey region flag tag flag tag FLAG-tagged FLAG DYKDDDDKV epitope tag DYKDDDDKV epitope tag FLAG FLAG-tagged undetermined undetermined sequence position undetermined undetermined undetermined sequence position undetermined physical association physical association 2D Fly Groucho immunoprecipitated with mouse Six3.