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/ A cancer-associated RING finger protein, RNF43, is a ubiquitin ligase that interacts with a nuclear protein, HAP95. A cancer-associated RING finger protein, RNF43, is a ubiquitin ligase that interacts with a nuclear protein, HAP95. Exp. Cell Res. (0014-4827) 2008 imex curation Sugiura T., Yamaguchi A., Miyamoto K. Apoptosis - Interactions involving proteins with a function related to apoptosis Cancer - Interactions investigated in the context of cancer sugiura.takeyuki.uu@daiichisankyo.co.jp 14-May-2008: Contacted by JYOTI. A human small intestine cDNA library was used. davet 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 Sugiura T., Yamaguchi A., Miyamoto K. Apoptosis - Interactions involving proteins with a function related to apoptosis sugiura.takeyuki.uu@daiichisankyo.co.jp Accepted 2008-APR-18 by DAVET. 14-May-2008: Contacted by JYOTI. A human small intestine cDNA library was used. Exp. Cell Res. (0014-4827) 2008 davet Only protein-protein interactions imex curation A cancer-associated RING finger protein, RNF43, is a ubiquitin ligase that interacts with a nuclear protein, HAP95. A cancer-associated RING finger protein, RNF43, is a ubiquitin ligase that interacts with a nuclear protein, HAP95. Exp. Cell Res. (0014-4827) 2008 imex curation Sugiura T., Yamaguchi A., Miyamoto K. Apoptosis - Interactions involving proteins with a function related to apoptosis Cancer - Interactions investigated in the context of cancer sugiura.takeyuki.uu@daiichisankyo.co.jp 14-May-2008: Contacted by JYOTI. A human small intestine cDNA library was used. davet Only protein-protein interactions 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 Sugiura T., Yamaguchi A., Miyamoto K. Apoptosis - Interactions involving proteins with a function related to apoptosis The authors confirmed that the Ubiquitin used was of human origin. sugiura.takeyuki.uu@daiichisankyo.co.jp Accepted 2008-APR-18 by DAVET. 14-May-2008: Contacted by JYOTI. A human small intestine cDNA library was used. Exp. Cell Res. (0014-4827) 2008 davet Only protein-protein interactions imex curation A cancer-associated RING finger protein, RNF43, is a ubiquitin ligase that interacts with a nuclear protein, HAP95. A cancer-associated RING finger protein, RNF43, is a ubiquitin ligase that interacts with a nuclear protein, HAP95. Exp. Cell Res. (0014-4827) 2008 imex curation Sugiura T., Yamaguchi A., Miyamoto K. Apoptosis - Interactions involving proteins with a function related to apoptosis Cancer - Interactions investigated in the context of cancer sugiura.takeyuki.uu@daiichisankyo.co.jp 14-May-2008: Contacted by JYOTI. A human small intestine cDNA library was used. davet Only protein-protein interactions in vitro In vitro enzymatic study enzymatic study western blot western blot Immuno blot Immuno blot Sugiura T., Yamaguchi A., Miyamoto K. Apoptosis - Interactions involving proteins with a function related to apoptosis Purified E1, E2 and bacterially expressed MBP-RNF43 (aa 1-322) were added to ubiquitin and ATP to assay RNF43 autoubiqutination activity. The E2 enzymes that supported the autoubiquitination were UBE2D2 or UBE2D3. The authors confirmed that the ubiquitin and E2 enzymes used in the ubiquitin ligase assay were of human origin. sugiura.takeyuki.uu@daiichisankyo.co.jp Accepted 2008-APR-18 by DAVET. 14-May-2008: Contacted by JYOTI. A human small intestine cDNA library was used. Exp. Cell Res. (0014-4827) 2008 davet Only protein-protein interactions imex curation rnf43_human E3 ubiquitin-protein ligase RNF43 RNF43 RING finger protein 43 RING-type E3 ubiquitin transferase RNF43 protein protein human Homo sapiens Human MSGGHQLQLAALWPWLLMATLQAGFGRTGLVLAAAVESERSAEQKAIIRVIPLKMDPTGKLNLTLEGVFAGVAEITPAEGKLMQSHPLYLCNASDDDNLEPGFISIVKLESPRRAPRPCLSLASKARMAGERGASAVLFDITEDRAAAEQLQQPLGLTWPVVLIWGNDAEKLMEFVYKNQKAHVRIELKEPPAWPDYDVWILMTVVGTIFVIILASVLRIRCRPRHSRPDPLQQRTAWAISQLATRRYQASCRQARGEWPDSGSSCSSAPVCAICLEEFSEGQELRVISCLHEFHRNCVDPWLHQHRTCPLCMFNITEGDSFSQSLGPSRSYQEPGRRLHLIRQHPGHAHYHLPAAYLLGPSRSAVARPPRPGPFLPSQEPGMGPRHHRFPRAAHPRAPGEQQRLAGAQHPYAQGWGLSHLQSTSQHPAACPVPLRRARPPDSSGSGESYCTERSGYLADGPASDSSSGPCHGSSSDSVVNCTDISLQGVHGSSSTFCSSLSSDFDPLVYCSPKGDPQRVDMQPSVTSRPRSLDSVVPTGETQVSSHVHYHRHRHHHYKKRFQWHGRKPGPETGVPQSRPPIPRTQPQPEPPSPDQQVTRSNSAAPSGRLSNPQCPRALPEPAPGPVDASSICPSTSSLFNLQKSSLSARHPQRKRRGGPSEPTPGSRPQDATVHPACQIFPHYTPSVAYPWSPEAHPLICGPPGLDKRLLPETPGPCYSNSQPVWLCLTPRQPLEPHPPGEGPSEWSSDTAEGRPCPYPHCQVLSAQPGSEEELEELCEQAV 4E87EA0CC359C858 akp8l_human A-kinase anchor protein 8-like AKAP8L HRIHFB2018 NAKAP95 Neighbor of A-kinase-anchoring protein 95 Homologous to AKAP95 protein Helicase A-binding protein 95 NAKAP protein protein human Homo sapiens Human MSYTGFVQGSETTLQSTYSDTSAQPTCDYGYGTWNSGTNRGYEGYGYGYGYGQDNTTNYGYGMATSHSWEMPSSDTNANTSASGSASADSVLSRINQRLDMVPHLETDMMQGGVYGSGGERYDSYESCDSRAVLSERDLYRSGYDYSELDPEMEMAYEGQYDAYRDQFRMRGNDTFGPRAQGWARDARSGRPMASGYGRMWEDPMGARGQCMSGASRLPSLFSQNIIPEYGMFQGMRGGGAFPGGSRFGFGFGNGMKQMRRTWKTWTTADFRTKKKKRKQGGSPDEPDSKATRTDCSDNSDSDNDEGTEGEATEGLEGTEAVEKGSRVDGEDEEGKEDGREEGKEDPEKGALTTQDENGQTKRKLQAGKKSQDKQKKRQRDRMVERIQFVCSLCKYRTFYEDEMASHLDSKFHKEHFKYVGTKLPKQTADFLQEYVTNKTKKTEELRKTVEDLDGLIQQIYRDQDLTQEIAMEHFVKKVEAAHCAACDLFIPMQFGIIQKHLKTMDHNRNRRLMMEQSKKSSLMVARSILNNKLISKKLERYLKGENPFTDSPEEEKEQEEAEGGALDEGAQGEAAGISEGAEGVPAQPPVPPEPAPGAVSPPPPPPPEEEEEGAVPLLGGALQRQIRGIPGLDVEDDEEGGGGAP CBC265BF25996BA2 ubb_human Polyubiquitin-B UBB protein protein human Homo sapiens Human MQIFVKTLTGKTITLEVEPSDTIENVKAKIQDKEGIPPDQQRLIFAGKQLEDGRTLSDYNIQKESTLHLVLRLRGGMQIFVKTLTGKTITLEVEPSDTIENVKAKIQDKEGIPPDQQRLIFAGKQLEDGRTLSDYNIQKESTLHLVLRLRGGMQIFVKTLTGKTITLEVEPSDTIENVKAKIQDKEGIPPDQQRLIFAGKQLEDGRTLSDYNIQKESTLHLVLRLRGGC 33011162F1C48BB1 ub2d3_human Ubiquitin-conjugating enzyme E2 D3 UBC5C Ubiquitin-protein ligase D3 Ubiquitin carrier protein D3 Ubiquitin-conjugating enzyme E2-17 kDa 3 Ubiquitin-conjugating enzyme E2(17)KB 3 UBE2D3 UBCH5C (E3-independent) E2 ubiquitin-conjugating enzyme D3 E2 ubiquitin-conjugating enzyme D3 protein protein human Homo sapiens Human MALKRINKELSDLARDPPAQCSAGPVGDDMFHWQATIMGPNDSPYQGGVFFLTIHFPTDYPFKPPKVAFTTRIYHPNINSNGSICLDILRSQWSPALTISKVLLSICSLLCDPNPDDPLVPEIARIYKTDRDKYNRISREWTQKYAM ADD74A8A708EFEE3 ub2d2_human Ubiquitin-conjugating enzyme E2 D2 UBC5B UBCH4 UBE2D2 UBC4 UBCH5B Ubiquitin-protein ligase D2 Ubiquitin carrier protein D2 Ubiquitin-conjugating enzyme E2-17 kDa 2 Ubiquitin-conjugating enzyme E2(17)KB 2 PUBC1 p53-regulated ubiquitin-conjugating enzyme 1 (E3-independent) E2 ubiquitin-conjugating enzyme D2 E2 ubiquitin-conjugating enzyme D2 protein protein human Homo sapiens Human MALKRIHKELNDLARDPPAQCSAGPVGDDMFHWQATIMGPNDSPYQGGVFFLTIHFPTDYPFKPPKVAFTTRIYHPNINSNGSICLDILRSQWSPALTISKVLLSICSLLCDPNPDDPLVPEIARIYKTDREKYNRIAREWTQKYAM DIP protein P62837 original sequence version: 81 DIP protein P62837-1 original sequence version: 134 DIP protein P62837 original sequence version: 95 DIP protein P62837 original sequence version: 139 DIP protein P62837 original sequence version: 37 DIP protein P62837 original sequence version: 115 DIP protein P62837 original sequence version: 58 DIP protein P62837 original sequence version: 74 DIP protein P62837 original sequence version: 103 DIP protein P62837 original sequence version: 101 C942BE7853CBC355 uba1_rabit Ubiquitin-like modifier-activating enzyme 1 Ubiquitin-activating enzyme E1 UBA1 UBE1 protein protein rabit Oryctolagus cuniculus (Rabbit) Rabbit MSSSPLSKKRRVSGPDPKPGSNCSPAQSVLPQVPSAPTNGMAKNGSEADIDEGLYSRQLYVLGHEAMKRLQTSSVLVSGLRGLGVEIAKNIILGGVKAVTLHDQGTAQWADLSSQFYLREEDIGKNRAEVSQPRLAELNSYVPVTAYTGPLVEDFLSGFQVVVLTNSPLEDQLRVGEFCHSRGIKLVVADTRGLFGQLFCDFGEEMILTDSNGEQPLSTMVSMVTKDNPGVVTCLDEARHGFESGDFVSFSEVQGMTELNGNQPIEIKVLGPYTFSICDTSNFSDYIRGGIVSQVKVPKKISFKSLSASLAEPDFVMTDFAKFSRPAQLHIGFQALHKFCAQHSRPPRPRNEEDAAELVTLARAVNSKASSAVQQDSLDEDLIRNLAFVAAGDLAPINAFIGGLAAQEVMKACSGKFMPIMQWLYFDALECLPEDKESLTEDKCLPRQNRYDGQVAVFGSDLQEKLGRQKYFLVGAGAIGCELLKNFAMIGLGCGENGEIIVTDMDTIEKSNLNRQFLFRPWDVTKLKSDTAAAAVHQMNPHIRVTSHQNRVGPDTERIYDDDFFQTLDGVANALDNVDARMYMDRRCVYYRKPLLESGTLGTKGNVQVVIPFLTESYSSSQDPPEKSIPICTLKNFPNAIEHTLQWARDEFEGLFKQPAENVNQYLTDPKFVERTLRLAGTQPLEVLEAVQRSLVLQLPQSWADCVTWACHHWHTQYSNNIRQLLHNFPPDQLTSSGAPFWSGPKRCPHPLTFDVSNPLHLDYVMAAANLFAQTYGLAGSQDRAAVATLLQSVQVPEFTPKSGVKIHVSDQELQSANASVDDSRLEELKATLPSPDKLPGFKMYPIDFEKDDDSNFHMDFIVAASNLRAENYDIPPADRHKSKLIAGKIIPAIATTTAAVVGLVCLELYKVVQGHRHLDSYKNGFLNLALPFFGFSEPLAAPRHQYYNQEWTLWDRFEVQGLQPNGEEMTLKQFLDYFKTEHKLEITMLSQGVSMLYSFFMPAAKLKERLDQPMTEIVSRVSKRKLGRHVRALVLELCCNDESGEDVEVPYVRYTIR 1196EAC5EEEC06A5 rnf43-akap8l 1 5 unspecified role unspecified role prey prey 4 unspecified role unspecified role bait bait region sufficient to bind sufficient binding region sufficient to bind certain certain sequence position certain certain certain sequence position certain physical association physical association Among them we paid our attention to one clone (number 13584) that showedstrong interactionwith RNF43 asmeasured by a -galactosidase assay (Fig. 6A). This clone harbored a contiguous open reading frame of HAP95 The authors state that their RNF43 cDNA clone contains two differences from the predicted protein sequence for NM_017763 (which is the same as Q68DV7): Ile47Val and Arg11His. BAIT gene NM_017763 rnf43-akap8l-1 2 5 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 4 unspecified role unspecified role bait bait region v5 tag v5 tag GKPIPNPLLGLDST epitope tag GKPIPNPLLGLDST epitope tag undetermined undetermined sequence position undetermined undetermined undetermined sequence position undetermined physical association physical association 6 B Westernblot analysis of the immunoprecipitate withanti- HA antibody revealed that RNF43 co-immunoprecipitated HAtagged HAP95 (HAP95-HA), rnf43-rps27a 2 4 unspecified role unspecified role bait bait region v5 tag v5 tag GKPIPNPLLGLDST epitope tag GKPIPNPLLGLDST epitope tag undetermined undetermined sequence position undetermined undetermined undetermined sequence position undetermined 6 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 physical association physical association 3 B we expressed RNF43-V5 along with HA-ubiquitin in 293 cells and the immunoprecipitates recovered with anti-V5 antibody were immunoblotted with anti-HA antibody. rnf43-rnf43 3 4 enzyme enzyme unspecified role unspecified role Q68DV7:p.[Cys290Ser;His292Ser] mutation decreasing mutation decreasing interaction hotspot hotspot mutation decreasing mutation analysis mutation analysis certain certain sequence position certain certain certain sequence position certain C S certain certain sequence position certain certain certain sequence position certain H S Wed May 02 20:08:37 BST 2018 This feature has been corrected as a result of our quality control procedures. The original label was 'cys290ser,his292ser' Tue Jun 30 18:54:50 BST 2020 This feature has been corrected as a result of our quality control procedures. The original label was 'p.[Cys290Ser;His292Ser]' n-terminal sufficient to bind sufficient binding region sufficient to bind deletion analysis deletion analysis 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) 9 unspecified role unspecified role ancillary ancillary 8 unspecified role unspecified role ancillary ancillary region his tag his tag Hexa-His-tag 6-His-tag Histidine-tag Hexa-His-tag Histidine-tag 6-His-tag undetermined undetermined sequence position undetermined undetermined undetermined sequence position undetermined region his tag his tag Hexa-His-tag 6-His-tag Histidine-tag Hexa-His-tag Histidine-tag 6-His-tag undetermined undetermined sequence position undetermined undetermined undetermined sequence position undetermined 4 enzyme target enzyme target substrate substrate unspecified role unspecified role Q68DV7:p.[Cys290Ser;His292Ser] mutation decreasing mutation decreasing interaction hotspot hotspot mutation decreasing mutation analysis mutation analysis certain certain sequence position certain certain certain sequence position certain C S certain certain sequence position certain certain certain sequence position certain H S Wed May 02 18:15:21 BST 2018 This feature has been corrected as a result of our quality control procedures. The original label was 'cys290ser,his292ser' Tue Jun 30 16:45:33 BST 2020 This feature has been corrected as a result of our quality control procedures. The original label was 'p.[Cys290Ser;His292Ser]' n-terminal sufficient to bind sufficient binding region sufficient to bind deletion analysis deletion analysis 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) ubiquitination ubiquitination reaction ubiquitination 5 Autoubiquitylation of RNF43 RNF43 is autoubiquitinated. TPEN - The presence of the metal chelating inhibitor TPEN resulted in the loss of polyubiquitination. The autoubiqutination of RNF43 was shown to be RING finger-dependent, as the mutations in this domain compromised autoubiqutination activity. rnf43-rnf43-1 3 4 enzyme enzyme unspecified role unspecified role Q68DV7:p.[Cys290Ser;His292Ser] mutation decreasing mutation decreasing interaction hotspot hotspot mutation decreasing mutation analysis mutation analysis certain certain sequence position certain certain certain sequence position certain C S certain certain sequence position certain certain certain sequence position certain H S Wed May 02 15:57:40 BST 2018 This feature has been corrected as a result of our quality control procedures. The original label was 'cys290ser,his292ser' Tue Jun 30 19:49:20 BST 2020 This feature has been corrected as a result of our quality control procedures. The original label was 'p.[Cys290Ser;His292Ser]' n-terminal sufficient to bind sufficient binding region sufficient to bind deletion analysis deletion analysis 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) 9 unspecified role unspecified role ancillary ancillary 7 unspecified role unspecified role ancillary ancillary region his tag his tag Hexa-His-tag 6-His-tag Histidine-tag Hexa-His-tag Histidine-tag 6-His-tag undetermined undetermined sequence position undetermined undetermined undetermined sequence position undetermined 4 enzyme target enzyme target substrate substrate unspecified role unspecified role Q68DV7:p.[Cys290Ser;His292Ser] mutation decreasing mutation decreasing interaction hotspot hotspot mutation decreasing mutation analysis mutation analysis certain certain sequence position certain certain certain sequence position certain C S certain certain sequence position certain certain certain sequence position certain H S Wed May 02 16:12:41 BST 2018 This feature has been corrected as a result of our quality control procedures. The original label was 'cys290ser,his292ser' Tue Jun 30 17:49:13 BST 2020 This feature has been corrected as a result of our quality control procedures. The original label was 'p.[Cys290Ser;His292Ser]' n-terminal sufficient to bind sufficient binding region sufficient to bind deletion analysis deletion analysis 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) ubiquitination ubiquitination reaction ubiquitination 5 Autoubiquitylation of RNF43 RNF43 is autoubiquitinated. TPEN - The presence of the metal chelating inhibitor TPEN resulted in the loss of polyubiquitination. The autoubiqutination of RNF43 was shown to be RING finger-dependent, as the mutations in this domain compromised autoubiqutination activity. rnf43-rps27a-1 2 6 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 4 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 Q68DV7:p.[Cys290Ser;His292Ser] mutation mutation mutation analysis mutation analysis certain certain sequence position certain certain certain sequence position certain C S certain certain sequence position certain certain certain sequence position certain H S Wed May 02 14:54:25 BST 2018 This feature has been corrected as a result of our quality control procedures. The original label was 'cys290ser,his292ser' Tue Jun 30 20:23:29 BST 2020 This feature has been corrected as a result of our quality control procedures. The original label was 'p.[Cys290Ser;His292Ser]' physical association physical association 3 C RNF43(M) was also ubiquitylated in vivo