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/ Thymosin beta 4 expression and nuclear transport are regulated by hMLH1. Thymosin beta 4 expression and nuclear transport are regulated by hMLH1. Biochem. Biophys. Res. Commun. (0006-291X) 2007 imex curation Brieger A., Plotz G., Zeuzem S., Trojan J. Cancer - Interactions investigated in the context of cancer a.brieger@em.uni-frankfurt.de 22-Jan-2008: Contacted by JYOTI. A human colon cDNA library was used. orchard Only protein-protein interactions ecoli Escherichia coli (strain K12) 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 Brieger A., Plotz G., Zeuzem S., Trojan J. Cancer - Interactions investigated in the context of cancer a.brieger@em.uni-frankfurt.de Accepted 2007-Not a month!-19 by ORCHARD. A human colon cDNA library was used. Biochem. Biophys. Res. Commun. (0006-291X) 2007 orchard Only protein-protein interactions imex curation Thymosin beta 4 expression and nuclear transport are regulated by hMLH1. Thymosin beta 4 expression and nuclear transport are regulated by hMLH1. Biochem. Biophys. Res. Commun. (0006-291X) 2007 imex curation Brieger A., Plotz G., Zeuzem S., Trojan J. Cancer - Interactions investigated in the context of cancer a.brieger@em.uni-frankfurt.de 22-Jan-2008: Contacted by JYOTI. A human colon cDNA library was used. orchard Only protein-protein interactions human-293t Homo sapiens 293 cells transformed with SV40 large T antigen 293t 293 cells expressing SV40 large T antigen. The original designation of this cell line was 293tsA1609neo. It is not in Cabri,ATCC or HyperCLDB. confocal microscopy confocal microscopy immunostaining immunostaining Brieger A., Plotz G., Zeuzem S., Trojan J. Cancer - Interactions investigated in the context of cancer The authors supplied by email the residues mutated in the nuclear localization signal of hMLH1. a.brieger@em.uni-frankfurt.de Accepted 2007-Not a month!-19 by ORCHARD. 22-Jan-2008: Contacted by JYOTI. A human colon cDNA library was used. Biochem. Biophys. Res. Commun. (0006-291X) 2007 orchard Only protein-protein interactions imex curation Thymosin beta 4 expression and nuclear transport are regulated by hMLH1. Thymosin beta 4 expression and nuclear transport are regulated by hMLH1. Biochem. Biophys. Res. Commun. (0006-291X) 2007 imex curation Brieger A., Plotz G., Zeuzem S., Trojan J. Cancer - Interactions investigated in the context of cancer a.brieger@em.uni-frankfurt.de 22-Jan-2008: Contacted by JYOTI. A human colon cDNA library was used. orchard Only protein-protein interactions human-293t Homo sapiens 293 cells transformed with SV40 large T antigen 293t 293 cells expressing SV40 large T antigen. The original designation of this cell line was 293tsA1609neo. It is not in Cabri,ATCC or HyperCLDB. anti bait coip anti bait coimmunoprecipitation anti bait coip western blot western blot Immuno blot Immuno blot Brieger A., Plotz G., Zeuzem S., Trojan J. Cancer - Interactions investigated in the context of cancer a.brieger@em.uni-frankfurt.de Accepted 2007-Not a month!-19 by ORCHARD. 22-Jan-2008: Contacted by JYOTI. A human colon cDNA library was used. Biochem. Biophys. Res. Commun. (0006-291X) 2007 orchard Only protein-protein interactions imex curation mlh1_human DNA mismatch repair protein Mlh1 MutL protein homolog 1 MLH1 COCA2 protein protein human Homo sapiens Human MSFVAGVIRRLDETVVNRIAAGEVIQRPANAIKEMIENCLDAKSTSIQVIVKEGGLKLIQIQDNGTGIRKEDLDIVCERFTTSKLQSFEDLASISTYGFRGEALASISHVAHVTITTKTADGKCAYRASYSDGKLKAPPKPCAGNQGTQITVEDLFYNIATRRKALKNPSEEYGKILEVVGRYSVHNAGISFSVKKQGETVADVRTLPNASTVDNIRSIFGNAVSRELIEIGCEDKTLAFKMNGYISNANYSVKKCIFLLFINHRLVESTSLRKAIETVYAAYLPKNTHPFLYLSLEISPQNVDVNVHPTKHEVHFLHEESILERVQQHIESKLLGSNSSRMYFTQTLLPGLAGPSGEMVKSTTSLTSSSTSGSSDKVYAHQMVRTDSREQKLDAFLQPLSKPLSSQPQAIVTEDKTDISSGRARQQDEEMLELPAPAEVAAKNQSLEGDTTKGTSEMSEKRGPTSSNPRKRHREDSDVEMVEDDSRKEMTAACTPRRRIINLTSVLSLQEEINEQGHEVLREMLHNHSFVGCVNPQWALAQHQTKLYLLNTTKLSEELFYQILIYDFANFGVLRLSEPAPLFDLAMLALDSPESGWTEEDGPKEGLAEYIVEFLKKKAEMLADYFSLEIDEEGNLIGLPLLIDNYVPPLEGLPIFILRLATEVNWDEEKECFESLSKECAMFYSIRKQYISEESTLSGQQSEVPGSIPNSWKWTVEHIVYKALRSHILPPKHFTEDGNILQLANLPDLYKVFERC DIP protein P40692 original sequence version: 99 C9231FC406C2CA20 tyb4_human Thymosin beta-4 Fx TMSB4X TB4X THYB4 TMSB4 protein protein human Homo sapiens Human MSDKPDMAEIEKFDKSKLKKTETQEKNPLPSKETIEQEKQAGES 440C6158482DAAD0 mlh1-tmsb4x0 1 4 unspecified role unspecified role bait bait 5 unspecified role unspecified role prey prey physical association physical association During the last years several hMLH1-interacting partners have been identified by yeast two-hybrid. To circum-vent the limitations inherent to the yeast system, like delicate identification of nuclear proteins, we applied a bacterial two-hybrid screening. We now report the identification of Tb4, an actin-binding, and cell motility regulating protein tmsb4x-mlh1 3 4 unspecified role unspecified role prey prey 5 unspecified role unspecified role bait bait physical association physical association 1 The physical interaction of hMLH1 with Tb4 was verified by coimmunoprecipitation in a mammalian system P62328; P40692 transfection mlh1-tmsb4x 3 4 unspecified role unspecified role bait bait 5 unspecified role unspecified role prey prey physical association physical association 1 The physical interaction of hMLH1 with Tb4 was verified by coimmunoprecipitation in a mammalian system P62328; P40692 transfection mlh1-tmsb4x-1 3 5 unspecified role unspecified role prey prey 4 unspecified role unspecified role bait bait physical association physical association 1 The physical interaction of hMLH1 with Tb4 was verified by coimmunoprecipitation in a mammalian system P40692 transfection tmsb4x-mlh1-3 3 5 unspecified role unspecified role bait bait 4 unspecified role unspecified role prey prey physical association physical association 1 The physical interaction of hMLH1 with Tb4 was verified by coimmunoprecipitation in a mammalian system 5-aza-2'-deoxycytidine (5-Aza-CdR) - This interaction is enhanced by treatment of cells with 5-Aza-CdR, which restored hMLH1 expression in 293T cells by demethylation of the hMLH1 promoter. tmsb4x-mlh1-1 3 4 unspecified role unspecified role prey prey 5 unspecified role unspecified role bait bait physical association physical association 1 The physical interaction of hMLH1 with Tb4 was verified by coimmunoprecipitation in a mammalian system P40692 transfection mlh1-tmsb4x-2 3 4 unspecified role unspecified role bait bait 5 unspecified role unspecified role prey prey physical association physical association 1 The physical interaction of hMLH1 with Tb4 was verified by coimmunoprecipitation in a mammalian system 5-aza-2'-deoxycytidine (5-Aza-CdR) - This interaction is enhanced by treatment of cells with 5-Aza-CdR, which restored hMLH1 expression in 293T cells by demethylation of the hMLH1 promoter. mlh1-tmsb4x-3 2 4 unspecified role unspecified role neutral component neutral component 5 unspecified role unspecified role neutral component neutral component colocalization colocalization 2 Transfection of 293T cells with hMLH1 wt (Fig. 2, third panel) or restoration of endogenous hMLH1 expression by promoter demethylation using 5-aza-2-deoxycytidine (Fig. 2, bottom panel) prompted strong expression of endogenous Tb4 and colocalization with hMLH1 mainly in the nucleus. P40692 transfection When the NLS (nuclear localization signal) of hMLH1 was mutated (K471N or R472G), hMLH1 and thymosin beta 4 remained in the cytoplasm, which indicates that translocation of thymosin beta 4 relies on interaction with hMLH1 harboring a functional NLS. mlh1-tmsb4x-4 2 5 unspecified role unspecified role neutral component neutral component 4 unspecified role unspecified role neutral component neutral component colocalization colocalization 2 Transfection of 293T cells with hMLH1 wt (Fig. 2, third panel) or restoration of endogenous hMLH1 expression by promoter demethylation using 5-aza-2-deoxycytidine (Fig. 2, bottom panel) prompted strong expression of endogenous Tb4 and colocalization with hMLH1 mainly in the nucleus. This finding was confirmed in HCT-116 mlh 1-2 cells (data 5-aza-2'-deoxycytidine (5-Aza-CdR) - This interaction is enhanced by treatment of cells with 5-Aza-CdR, which restored hMLH1 expression in 293T cells by demethylation of the hMLH1 promoter.