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/ lu-2000-1 Zhangfei: a second cellular protein interacts with herpes simplex virus accessory factor HCF in a manner similar to Luman and VP16. 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 Lu R., Misra V. Fig 1 vikram.misra@usask.ca A Hela cDNA library was used. Nucleic Acids Res. (0305-1048) 2000 lu-2000-2 Zhangfei: a second cellular protein interacts with herpes simplex virus accessory factor HCF in a manner similar to Luman and VP16. in vitro In vitro pull down pull down predetermined predetermined participant predetermined Lu R., Misra V. Fig 1 vikram.misra@usask.ca A Hela cDNA library was used. Nucleic Acids Res. (0305-1048) 2000 lu-2000-3 Zhangfei: a second cellular protein interacts with herpes simplex virus accessory factor HCF in a manner similar to Luman and VP16. chlae-cos_7 Cercopithecus aethiops simian cells transformed with SV40 Grivet cos_7 cercopithecus aethiops, by transformation with SV40 transcription compl transcriptional complementation assay transcription compl predetermined predetermined participant predetermined Lu R., Misra V. As with the yeast two-hybrid system, a mammalian expression vector that produced the gal4 dbd fusion protein, gal4-hcfc1 was introduced into cos-7 cells with pc-zf or plasmids specifying its mutants. Since zf proteins have their own activation domain, a chimeric zf protein with gal4 activation domain was not needed. Plasmid pG5EC containing gal4-binding motifs was used as the reporter. Plasmid pcDNA3 was used as a control. The CAT activity was measured by ELISA 48hr post-transfection. Fig 1 vikram.misra@usask.ca A Hela cDNA library was used. Nucleic Acids Res. (0305-1048) 2000 lu-2000-4 Zhangfei: a second cellular protein interacts with herpes simplex virus accessory factor HCF in a manner similar to Luman and VP16. ecobb Escherichia coli (strain B / BL21) pull down pull down predetermined predetermined participant predetermined Lu R., Misra V. Fig 1 vikram.misra@usask.ca A Hela cDNA library was used. Nucleic Acids Res. (0305-1048) 2000 lu-2000-5 Zhangfei: a second cellular protein interacts with herpes simplex virus accessory factor HCF in a manner similar to Luman and VP16. chlae-cos_7 Cercopithecus aethiops simian cells transformed with SV40 Grivet cos_7 cercopithecus aethiops, by transformation with SV40 transcription compl transcriptional complementation assay transcription compl predetermined predetermined participant predetermined Lu R., Misra V. As with the yeast two-hybrid system, a mammalian expression vector that produced the gal4 dbd fusion protein, gal4-hcfc1 was introduced into cos-7 cells with pc-zf or plasmids specifying its mutants. Since zf proteins have their own activation domain, a chimeric zf protein with gal4 activation domain was not needed. Plasmid pG5EC containing gal4-binding motifs was used as the reporter. Plasmid pcDNA3 was used as a control. The CAT activity was measured by ELISA 48hr post-transfection. Fig 1 vikram.misra@usask.ca A Hela cDNA library was used. Nucleic Acids Res. (0305-1048) 2000 zhang_human CREB/ATF bZIP transcription factor Host cell factor-binding transcription factor Zhangfei CREBZF ZF protein protein human Homo sapiens Human MRHSLTKLLAASGSNSPTRSESPEPAATCSLPSDLTRAAAGEEETAAAGSPGRKQQFGDEGELEAGRGSRGGVAVRAPSPEEMEEEAIASLPGEETEDMDFLSGLELADLLDPRQPDWHLDPGLSSPGPLSSSGGGSDSGGLWRGDDDDEAAAAEMQRFSDLLQRLLNGIGGCSSSSDSGSAEKRRRKSPGGGGGGGSGNDNNQAATKSPRKAAAAAARLNRLKKKEYVMGLESRVRGLAAENQELRAENRELGKRVQALQEESRYLRAVLANETGLARLLSRLSGVGLRLTTSLFRDSPAGDHDYALPVGKQKQDLLEEDDSAGGVCLHVDKDKVSVEFCSACARKASSSLKM hcfc1_human Host cell factor 1 HCFC1 HCF1 C1 factor VP16 accessory protein VCAF CFF HFC1 protein protein human Homo sapiens Human MASAVSPANLPAVLLQPRWKRVVGWSGPVPRPRHGHRAVAIKELIVVFGGGNEGIVDELHVYNTATNQWFIPAVRGDIPPGCAAYGFVCDGTRLLVFGGMVEYGKYSNDLYELQASRWEWKRLKAKTPKNGPPPCPRLGHSFSLVGNKCYLFGGLANDSEDPKNNIPRYLNDLYILELRPGSGVVAWDIPITYGVLPPPRESHTAVVYTEKDNKKSKLVIYGGMSGCRLGDLWTLDIDTLTWNKPSLSGVAPLPRSLHSATTIGNKMYVFGGWVPLVMDDVKVATHEKEWKCTNTLACLNLDTMAWETILMDTLEDNIPRARAGHCAVAINTRLYIWSGRDGYRKAWNNQVCCKDLWYLETEKPPPPARVQLVRANTNSLEVSWGAVATADSYLLQLQKYDIPATAATATSPTPNPVPSVPANPPKSPAPAAAAPAVQPLTQVGITLLPQAAPAPPTTTTIQVLPTVPGSSISVPTAARTQGVPAVLKVTGPQATTGTPLVTMRPASQAGKAPVTVTSLPAGVRMVVPTQSAQGTVIGSSPQMSGMAALAAAAAATQKIPPSSAPTVLSVPAGTTIVKTMAVTPGTTTLPATVKVASSPVMVSNPATRMLKTAAAQVGTSVSSATNTSTRPIITVHKSGTVTVAQQAQVVTTVVGGVTKTITLVKSPISVPGGSALISNLGKVMSVVQTKPVQTSAVTGQASTGPVTQIIQTKGPLPAGTILKLVTSADGKPTTIITTTQASGAGTKPTILGISSVSPSTTKPGTTTIIKTIPMSAIITQAGATGVTSSPGIKSPITIITTKVMTSGTGAPAKIITAVPKIATGHGQQGVTQVVLKGAPGQPGTILRTVPMGGVRLVTPVTVSAVKPAVTTLVVKGTTGVTTLGTVTGTVSTSLAGAGGHSTSASLATPITTLGTIATLSSQVINPTAITVSAAQTTLTAAGGLTTPTITMQPVSQPTQVTLITAPSGVEAQPVHDLPVSILASPTTEQPTATVTIADSGQGDVQPGTVTLVCSNPPCETHETGTTNTATTTVVANLGGHPQPTQVQFVCDRQEAAASLVTSTVGQQNGSVVRVCSNPPCETHETGTTNTATTATSNMAGQHGCSNPPCETHETGTTNTATTAMSSVGANHQRDARRACAAGTPAVIRISVATGALEAAQGSKSQCQTRQTSATSTTMTVMATGAPCSAGPLLGPSMAREPGGRSPAFVQLAPLSSKVRLSSPSIKDLPAGRHSHAVSTAAMTRSSVGAGEPRMAPVCESLQGGSPSTTVTVTALEALLCPSATVTQVCSNPPCETHETGTTNTATTSNAGSAQRVCSNPPCETHETGTTHTATTATSNGGTGQPEGGQQPPAGRPCETHQTTSTGTTMSVSVGALLPDATSSHRTVESGLEVAAAPSVTPQAGTALLAPFPTQRVCSNPPCETHETGTTHTATTVTSNMSSNQDPPPAASDQGEVESTQGDSVNITSSSAITTTVSSTLTRAVTTVTQSTPVPGPSVPPPEELQVSPGPRQQLPPRQLLQSASTALMGESAEVLSASQTPELPAAVDLSSTGEPSSGQESAGSAVVATVVVQPPPPTQSEVDQLSLPQELMAEAQAGTTTLMVTGLTPEELAVTAAAEAAAQAAATEEAQALAIQAVLQAAQQAVMGTGEPMDTSEAAATVTQAELGHLSAEGQEGQATTIPIVLTQQELAALVQQQQLQEAQAQQQHHHLPTEALAPADSLNDPAIESNCLNELAGTVPSTVALLPSTATESLAPSNTFVAPQPVVVASPAKLQAAATLTEVANGIESLGVKPDLPPPPSKAPMKKENQWFDVGVIKGTNVMVTHYFLPPDDAVPSDDDLGTVPDYNQLKKQELQPGTAYKFRVAGINACGRGPFSEISAFKTCLPGFPGAPCAIKISKSPDGAHLTWEPPSVTSGKIIEYSVYLAIQSSQAGGELKSSTPAQLAFMRVYCGPSPSCLVQSSSLSNAHIDYTTKPAIIFRIAARNEKGYGPATQVRWLQETSKDSSGTKPANKRPMSSPEMKSAPKKSKADGQ creb3_human Cyclic AMP-responsive element-binding protein 3 Luman Transcription factor LZIP-alpha CREB3 LZIP Leucine zipper protein protein protein human Homo sapiens Human MELELDAGDQDLLAFLLEESGDLGTAPDEAVRAPLDWALPLSEVPSDWEVDDLLCSLLSPPASLNILSSSNPCLVHHDHTYSLPRETVSMDLESESCRKEGTQMTPQHMEELAEQEIARLVLTDEEKSLLEKEGLILPETLPLTKTEEQILKRVRRKIRNKRSAQESRRKKKVYVGGLESRVLKYTAQNMELQNKVQLLEEQNLSLLDQLRKLQAMVIEISNKTSSSSTCILVLLVSFCLLLVPAMYSSDTRGSLPAEHGVLSRQLRALPSEDPYQLELPALQSEVPKDSTHQWLDGSDCVLQAPGNTSCLLHYMPQAPSAEPPLEWPFPDLFSEPLCRGPILPLQANLTRKGGWLPTGSPSVILQDRYSG hcfc1-zf-1 Y2H of interaction between hcfc1 and zf 207458 N/A 207463 unspecified role unspecified role prey prey N/A 207464 unspecified role unspecified role bait bait physical association physical association false false false hcfc1-zf-2 GST pull-down of interaction between hcfc1 and zf 207459 N/A 207463 unspecified role unspecified role bait bait asp221ala required to bind necessary binding region required to bind mutation analysis mutation analysis certain certain sequence position certain certain certain sequence position certain false his222ala required to bind necessary binding region required to bind mutation analysis mutation analysis certain certain sequence position certain certain certain sequence position certain false tyr224ala required to bind necessary binding region required to bind mutation analysis mutation analysis certain certain sequence position certain certain certain sequence position certain false 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 false ecobb Escherichia coli (strain B / BL21) N/A 207464 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 false in vitro In vitro physical association physical association false false false Mutagenesis study of the hcfc1-binding motif in zf showed that each of the residues at AA positions 221, 222 and 224 were essential for interaction with hcfc1. hcfc1-zf-3 Mammalian two-hybrid assay of interaction between hcfc1 and zf 207460 N/A 207464 unspecified role unspecified role bait bait gal4 dbd tag tag undetermined undetermined sequence position undetermined undetermined undetermined sequence position undetermined false chlae-cos_7 Cercopithecus aethiops simian cells transformed with SV40 Grivet cos_7 cercopithecus aethiops, by transformation with SV40 N/A 207463 unspecified role unspecified role prey prey his222ala required to bind necessary binding region required to bind mutation analysis mutation analysis certain certain sequence position certain certain certain sequence position certain false tyr224ala required to bind necessary binding region required to bind mutation analysis mutation analysis certain certain sequence position certain certain certain sequence position certain false asp221ala required to bind necessary binding region required to bind mutation analysis mutation analysis certain certain sequence position certain certain certain sequence position certain false chlae-cos_7 Cercopithecus aethiops simian cells transformed with SV40 Grivet cos_7 cercopithecus aethiops, by transformation with SV40 physical association physical association false false false This was an in-vivo mutagenesis study of the hcfc1-binding motif in zf showing that each of the residues at AA positions 221, 222 and 224 were essential for interaction with hcfc1. hcfc1-zf-4 GST pull-down of interaction between hcfc1 and zf and the effect of p134s mutation in hcfc1 207461 N/A 207464 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 false pro134ser required to bind necessary binding region required to bind mutation analysis mutation analysis certain certain sequence position certain certain certain sequence position certain false in vitro In vitro N/A 207463 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 false ecobb Escherichia coli (strain B / BL21) physical association physical association false false false Demonstrates that zf associates with hcfc1 but not its mutant hcfc1(p134s). hcfc1-zf-5 Mammalian two-hybrid assay of interaction between hcfc1 and zf 207462 N/A 207465 unspecified role unspecified role prey prey chlae-cos_7 Cercopithecus aethiops simian cells transformed with SV40 Grivet cos_7 cercopithecus aethiops, by transformation with SV40 N/A 207464 unspecified role unspecified role bait bait gal4 tag tag undetermined undetermined sequence position undetermined undetermined undetermined sequence position undetermined false pro134ser required to bind necessary binding region required to bind mutation analysis mutation analysis certain certain sequence position certain certain certain sequence position certain false chlae-cos_7 Cercopithecus aethiops simian cells transformed with SV40 Grivet cos_7 cercopithecus aethiops, by transformation with SV40 N/A 207463 unspecified role unspecified role prey prey chlae-cos_7 Cercopithecus aethiops simian cells transformed with SV40 Grivet cos_7 cercopithecus aethiops, by transformation with SV40 physical association physical association false false false This was an in-vivo study demonstrating that zf and creb can interact with wild-type hcfc1 but not with the hcfc1(p134s)mutant.