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/ Transcriptional repression of target genes by LEUNIG and SEUSS, two interacting regulatory proteins for Arabidopsis flower development. Transcriptional repression of target genes by LEUNIG and SEUSS, two interacting regulatory proteins for Arabidopsis flower development. Proc. Natl. Acad. Sci. U.S.A. (0027-8424) 2004 imex curation Sridhar VV., Surendrarao A., Gonzalez D., Conlan RS., Liu Z. zl17@umail.umd.edu 25-Jun-2008: Contacted by JYOTI. 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 Sridhar VV., Surendrarao A., Gonzalez D., Conlan RS., Liu Z. zl17@umail.umd.edu Accepted 2008-JUN-15 by ORCHARD. Proc. Natl. Acad. Sci. U.S.A. (0027-8424) 2004 orchard Only protein-protein interactions imex curation Transcriptional repression of target genes by LEUNIG and SEUSS, two interacting regulatory proteins for Arabidopsis flower development. Transcriptional repression of target genes by LEUNIG and SEUSS, two interacting regulatory proteins for Arabidopsis flower development. Proc. Natl. Acad. Sci. U.S.A. (0027-8424) 2004 imex curation Sridhar VV., Surendrarao A., Gonzalez D., Conlan RS., Liu Z. zl17@umail.umd.edu 25-Jun-2008: Contacted by JYOTI. orchard Only protein-protein interactions in vitro In vitro pull down pull down anti tag western anti tag western blot anti tag western Sridhar VV., Surendrarao A., Gonzalez D., Conlan RS., Liu Z. zl17@umail.umd.edu Accepted 2008-JUN-15 by ORCHARD. 25-Jun-2008: Contacted by JYOTI. Proc. Natl. Acad. Sci. U.S.A. (0027-8424) 2004 orchard Only protein-protein interactions imex curation seuss_arath Transcriptional corepressor SEUSS SEU F28H19.10 At1g43850 protein protein arath Arabidopsis thaliana (Mouse-ear cress) Mouse-ear cress MVPSEPPNPVGGGENVPPSILGGQGGAPLPSQPAFPSLVSPRTQFGNNMSMSMLGNAPNISSLLNNQSFVNGIPGSMISMDTSGAESDPMSNVGFSGLSSFNASSMVSPRSSGQVQGQQFSNVSANQLLAEQQRNKKMETQSFQHGQQQSMQQQFSTVRGGGLAGVGPVKMEPGQVSNDQQHGQVQQQQQKMLRNLGSVKLEPQQIQAMRNLAQVKMEPQHSEQSLFLQQQQRQQQQQQQQQFLQMPGQSPQAQMNIFQQQRLMQLQQQQLLKSMPQQRPQLPQQFQQQNLPLRPPLKPVYEPGMGAQRLTQYMYRQQHRPEDNNIEFWRKFVAEYFAPNAKKRWCVSMYGSGRQTTGVFPQDVWHCEICNRKPGRGFEATAEVLPRLFKIKYESGTLEELLYVDMPRESQNSSGQIVLEYAKATQESVFEHLRVVRDGQLRIVFSPDLKIFSWEFCARRHEELIPRRLLIPQVSQLGSAAQKYQQAAQNATTDSALPELQNNCNMFVASARQLAKALEVPLVNDLGYTKRYVRCLQISEVVNSMKDLIDYSRETRTGPIESLAKFPRRTGPSSALPGPSPQQASDQLRQQQQQQQQQQQQQQQQQQQQQQQQTVSQNTNSDQSSRQVALMQGNPSNGVNYAFNAASASTSTSSIAGLIHQNSMKGRHQNAAYNPPNSPYGGNSVQMQSPSSSGTMVPSSSQQQHNLPTFQSPTSSSNNNNPSQNGIPSVNHMGSTNSPAMQQAGEVDGNESSSVQKILNEILMNNQAHNNSSGGSMVGHGSFGNDGKGQANVNSSGVLLMNGQVNNNNNTNIGGAGGFGGGIGQSMAANGINNINGNNSLMNGRVGMMVRDPNGQQDLGNQLLGAVNGFNNFDWNA 636D83BAD3946467 leung_arath Transcriptional corepressor LEUNIG LUG Protein ROTUNDA2 RON2 L23H3.30 At4g32551 protein protein arath Arabidopsis thaliana (Mouse-ear cress) Mouse-ear cress MSQTNWEADKMLDVYIHDYLVKRDLKATAQAFQAEGKVSSDPVAIDAPGGFLFEWWSVFWDIFIARTNEKHSEVAASYIETQMIKAREQQLQQSQHPQVSQQQQQQQQQQIQMQQLLLQRAQQQQQQQQQQHHHHQQQQQQQQQQQQQQQQQQQQHQNQPPSQQQQQQSTPQHQQQPTPQQQPQRRDGSHLANGSANGLVGNNSEPVMRQNPGSGSSLASKAYEERVKMPTQRESLDEAAMKRFGDNVGQLLDPSHASILKSAAASGQPAGQVLHSTSGGMSPQVQTRNQQLPGSAVDIKSEINPVLTPRTAVPEGSLIGIPGSNQGSNNLTLKGWPLTGFDQLRSGLLQQQKPFMQSQSFHQLNMLTPQHQQQLMLAQQNLNSQSVSEENRRLKMLLNNRSMTLGKDGLGSSVGDVLPNVGSSLQPGGSLLPRGDTDMLLKLKMALLQQQQQNQQQGGGNPPQPQPQPQPLNQLALTNPQPQSSNHSIHQQEKLGGGGSITMDGSISNSFRGNEQVLKNQSGRKRKQPVSSSGPANSSGTANTAGPSPSSAPSTPSTHTPGDVISMPNLPHSGGSSKSMMMFGTEGTGTLTSPSNQLADMDRFVEDGSLDDNVESFLSQEDGDQRDAVTRCMDVSKGFTFTEVNSVRASTTKVTCCHFSSDGKMLASAGHDKKAVLWYTDTMKPKTTLEEHTAMITDIRFSPSQLRLATSSFDKTVRVWDADNKGYSLRTFMGHSSMVTSLDFHPIKDDLICSCDNDNEIRYWSINNGSCTRVYKGGSTQIRFQPRVGKYLAASSANLVNVLDVETQAIRHSLQGHANPINSVCWDPSGDFLASVSEDMVKVWTLGTGSEGECVHELSCNGNKFQSCVFHPAYPSLLVIGCYQSLELWNMSENKTMTLPAHEGLITSLAVSTATGLVASASHDKLVKLWK 7CB879744496A8AA seu-lug 1 3 unspecified role unspecified role prey prey 4 unspecified role unspecified role bait bait n-terminal required to bind necessary binding region required to bind certain certain sequence position certain certain certain sequence position certain physical association physical association Figure 1 The LUG protein can be divided into three domains: the N-terminal LUFS domain, the central Q-rich domain, and the C-terminal 7-WD repeat domain. Full-length LUG, LUFS, LUFS+Q, and Q+WD were each fused to the DNA-binding domain of GAL4 (GAL4BD) and were tested for interaction with full-length SEU which was fused to the activation domain of GAL4 (GAL4AD). Whereas full-length LUG, LUFS, and LUFS+Q interacted with full-length SEU, Q+WD failed to interact with SEU (Fig. 1A). This finding suggested that the LUFS domain is both necessary and sufficient for interacting with SEU. Interestingly, the strongest interaction was observed between the LUFS and SEU seu-lug-1 2 3 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) 4 unspecified role unspecified role prey prey n-terminal required to bind necessary binding region required to bind certain certain sequence position certain certain certain sequence position certain region mbp tag maltose binding protein tag maltose binding protein tag maltose binding protein tag mbp tag undetermined undetermined sequence position undetermined undetermined undetermined sequence position undetermined ecobb Escherichia coli (strain B / BL21) direct interaction direct interaction Figure 2 To establish whether the two-hybrid interactions between LUG and SEU represented a direct physical contact in vitro, we carried out GST affinity chromatography. Immobilized GSTSEU was incubated with MBP, MBP-LUFS, and MBPLUFS+Q. In agreement with the two-hybrid data, GST-SEU (but not GST alone) interacted specifically with MBP-LUFS and MBP-LUFS+Q (Fig. 2), demonstrating a direct protein-protein interaction between LUG and SEU.