MINT MINT, Dpt of Biology, University of Rome Tor Vergata MINT http://mint.bio.uniroma2.it/mint url>http://mint.bio.uniroma2.it/mint/ https://mint.bio.uniroma2.it MINT-[0-9]+ https://mint.bio.uniroma2.it/index.php/results-interactions/?id=${ac} The Molecular INTeraction database (MINT) is a relational database designed to store interactions between biological molecules. http://mint.bio.uniroma2.it Goliath family E3 ligases regulate the recycling endosome pathway via VAMP3 ubiquitylation. Goliath family E3 ligases regulate the recycling endosome pathway via VAMP3 ubiquitylation. EMBO J. (0261-4189) 2013 imex curation Yamazaki Y., Schönherr C., Varshney G.K., Dogru M., Hallberg B., Palmer R.H. ruth.palmer@ucmp.umu.se 02-Apr-2014: Contacted by IntAct-Help. 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. fluorescence imaging fluorescence microscopy fluorescence imaging immunostaining immunostaining Yamazaki Y., Schönherr C., Varshney G.K., Dogru M., Hallberg B., Palmer R.H. ruth.palmer@ucmp.umu.se Accepted 2014-MAR-05 by MARTA EMBO J. (0261-4189) 2013 Only protein-protein interactions imex curation Goliath family E3 ligases regulate the recycling endosome pathway via VAMP3 ubiquitylation. Goliath family E3 ligases regulate the recycling endosome pathway via VAMP3 ubiquitylation. EMBO J. (0261-4189) 2013 imex curation Yamazaki Y., Schönherr C., Varshney G.K., Dogru M., Hallberg B., Palmer R.H. ruth.palmer@ucmp.umu.se 02-Apr-2014: Contacted by IntAct-Help. Only protein-protein interactions drome-wing disc Drosophila melanogaster wing disc wing_disc fluorescence imaging fluorescence microscopy fluorescence imaging immunostaining immunostaining Yamazaki Y., Schönherr C., Varshney G.K., Dogru M., Hallberg B., Palmer R.H. ruth.palmer@ucmp.umu.se Accepted 2014-MAR-05 by MARTA EMBO J. (0261-4189) 2013 Only protein-protein interactions imex curation q9vi20_drome gzl CG10277-RD Dmel\CG10277 godzilla godzilla[CG10277] CG10277 Dmel_CG10277 protein protein drome Drosophila melanogaster (Fruit fly) Fruit fly MSKRSCQILTLLGLCLVCHEATLVGGHVLVYRKATSQLIEEFNDLPAQFGPNLPSNGLKVYVVPARRPYYGCDSLDRPPHLKYPPSAKFVALVARGECVFERKIRVAQNASYSAVIVYNNEGDDLEQMSAENITGIRIPSVFVGHTTGKALATYFTTEVVLIINDELPFNINTQLILPFSILIGMCFIIMVIYMIYKCIREQRRLRRHRLPKSMLKKLPVLRYTKNNANNKYDTCVICLEDFIEDDKLRVLPCSHPYHTHCIDPWLTENRRVCPICKRKVFTKGEARASRSRQPSLDNVTDTDDDTTPLLQQQQSNGRQVGQVSSASSAGGAAGSSSSVAAAAVAGTTRHGTFRRGHAGRNPFEESQSSDDENALLASTVRPATSSGAHERINPFDRAPNLPAHLAEQLTESRRSVWSRINFASFFRRQPAVISVAAPPYLERVESGTSAMGLPVTGTIAVASPASNNILNPNLSGSFKDEDDMPPHRSIYEPIAISTPAADSATVDDSAFLQTPTQGGIGVAALPHSASDRQFLI 90E787655D4746A0 q9v3i2_drome Rab5 Drab5 AAF51265 BAA88244 DRab5 Dm Rab5 DmRab5 Dmel\CG3664 RAB5 RAB5a Rab Rab-5 dRab5 drab5 l(2)k08232 rab5 Dmel_CG3664 CG3664 protein protein drome Drosophila melanogaster (Fruit fly) Fruit fly MATTPRSGGASGTGTAQRPNGTSQNKSCQFKLVLLGESAVGKSSLVLRFVKGQFHEYQESTIGAAFLTQTICIEDTVVKFEIWDTAGQERYHSLAPMYYRGAQAAIVVYDIQNQDSFQRAKTWVKELHKQASPNIVIALAGNKADLSNIRVVEFDEAKQYAEENGLLFMETSAKTGMNVNDIFLAIAKKLPKNDGANNQGTSIRPTGTETNRPTNNCCK BFA3A28955B7F15D rn167_human E3 ubiquitin-protein ligase RNF167 RING-type E3 ubiquitin transferase RNF167 RING finger protein 167 RING105 RNF167 LP2254 protein protein human Homo sapiens Human MHPAAFPLPVVVAAVLWGAAPTRGLIRATSDHNASMDFADLPALFGATLSQEGLQGFLVEAHPDNACSPIAPPPPAPVNGSVFIALLRRFDCNFDLKVLNAQKAGYGAAVVHNVNSNELLNMVWNSEEIQQQIWIPSVFIGERSSEYLRALFVYEKGARVLLVPDNTFPLGYYLIPFTGIVGLLVLAMGAVMIARCIQHRKRLQRNRLTKEQLKQIPTHDYQKGDQYDVCAICLDEYEDGDKLRVLPCAHAYHSRCVDPWLTQTRKTCPICKQPVHRGPGDEDQEEETQGQEEGDEGEPRDHPASERTPLLGSSPTLPTSFGSLAPAPLVFPGPSTDPPLSPPSSPVILV D87493CE24C2A418 goli_drome Protein goliath gol g1 GL Protein g1 CG2679 protein protein drome Drosophila melanogaster (Fruit fly) Fruit fly MYIRKTLLLACLVLLFGGLPPLTFPATTTTLVAAMSIANQDLERYFRPGNGTHSFSGMEDRIAVDVYNYAFLNWSYVEHGNMLCNMEFAQEQARYGEGKVLNVTGRLIHITATDNFSDDYACTPYIRGTLGAPIPDKGETWIALVRRGRCTFEEKVKHVYQQNAAGVIIYNDKQVMQLEKMQIKGKTRNIAAVITYQNIGQDLSLTLDKGYNVTISIIEGRRGVRTISSLNRTSVLFVSISFIVLMIISLVWLIFYYIQRFRYMQAKDQQSRNLCSVTKKAIMKIPTKTGKFSDEKDLDSDCCAICIEAYKPTDTIRILPCKHEFHKNCIDPWLIEHRTCPMCKLDVLKFYGYVFLGSEESILEYQPDPPQGLALVEARDESADLNRSRDFVVDFPRVFVLDSGCVVGAREMLFPCRIPERSQSSLSLRQARDWVSLMSNKLEEQQGLRSMRNDEMQQVIK 309F41D935BD78B7 eea1_human Early endosome antigen 1 EEA1 ZFYVE2 Endosome-associated protein p162 Zinc finger FYVE domain-containing protein 2 protein protein human Homo sapiens Human MLRRILQRTPGRVGSQGSDLDSSATPINTVDVNNESSSEGFICPQCMKSLGSADELFKHYEAVHDAGNDSGHGGESNLALKRDDVTLLRQEVQDLQASLKEEKWYSEELKKELEKYQGLQQQEAKPDGLVTDSSAELQSLEQQLEEAQTENFNIKQMKDLFEQKAAQLATEIADIKSKYDEERSLREAAEQKVTRLTEELNKEATVIQDLKTELLQRPGIEDVAVLKKELVQVQTLMDNMTLERERESEKLKDECKKLQSQYASSEATISQLRSELAKGPQEVAVYVQELQKLKSSVNELTQKNQTLTENLLKKEQDYTKLEEKHNEESVSKKNIQATLHQKDLDCQQLQSRLSASETSLHRIHVELSEKGEATQKLKEELSEVETKYQHLKAEFKQLQQQREEKEQHGLQLQSEINQLHSKLLETERQLGEAHGRLKEQRQLSSEKLMDKEQQVADLQLKLSRLEEQLKEKVTNSTELQHQLDKTKQQHQEQQALQQSTTAKLREAQNDLEQVLRQIGDKDQKIQNLEALLQKSKENISLLEKEREDLYAKIQAGEGETAVLNQLQEKNHTLQEQVTQLTEKLKNQSESHKQAQENLHDQVQEQKAHLRAAQDRVLSLETSVNELNSQLNESKEKVSQLDIQIKAKTELLLSAEAAKTAQRADLQNHLDTAQNALQDKQQELNKITTQLDQVTAKLQDKQEHCSQLESHLKEYKEKYLSLEQKTEELEGQIKKLEADSLEVKASKEQALQDLQQQRQLNTDLELRATELSKQLEMEKEIVSSTRLDLQKKSEALESIKQKLTKQEEEKKILKQDFETLSQETKIQHEELNNRIQTTVTELQKVKMEKEALMTELSTVKDKLSKVSDSLKNSKSEFEKENQKGKAAILDLEKTCKELKHQLQVQMENTLKEQKELKKSLEKEKEASHQLKLELNSMQEQLIQAQNTLKQNEKEEQQLQGNINELKQSSEQKKKQIEALQGELKIAVLQKTELENKLQQQLTQAAQELAAEKEKISVLQNNYEKSQETFKQLQSDFYGRESELLATRQDLKSVEEKLSLAQEDLISNRNQIGNQNKLIQELKTAKATLEQDSAKKEQQLQERCKALQDIQKEKSLKEKELVNEKSKLAEIEEIKCRQEKEITKLNEELKSHKLESIKEITNLKDAKQLLIQQKLELQGKADSLKAAVEQEKRNQQILKDQVKKEEEELKKEFIEKEAKLHSEIKEKEVGMKKHEENEAKLTMQITALNENLGTVKKEWQSSQRRVSELEKQTDDLRGEIAVLEATVQNNQDERRALLERCLKGEGEIEKLQTKVLELQRKLDNTTAAVQELGRENQSLQIKHTQALNRKWAEDNEVQNCMACGKGFSVTVRRHHCRQCGNIFCAECSAKNALTPSSKKPVRVCDACFNDLQG 51BA418F561E3411 vamp3_human Vesicle-associated membrane protein 3 Synaptobrevin-3 Cellubrevin VAMP3 SYB3 protein protein human Homo sapiens Human MSTGPTAATGSNRRLQQTQNQVDEVVDIMRVNVDKVLERDQKLSELDDRADALQAGASQFETSAAKLKRKYWWKNCKMWAIGITVLVIFIIIIIVWVVSS DIP protein Q15836 original sequence version: 141 347E9163157AFF42 cg10277_rd-eea1 1 7 immunostaining immunostaining immunostaining immunostaining unspecified role unspecified role neutral component neutral component physiological level physiological level endogenous level endogenous endogenous endogenous level fixed cell fixed cell 3 tag fluorescence tag visualisation by fluorescence tag fluorescence immunostaining immunostaining unspecified role unspecified role neutral component neutral component over-expressed over expressed level over-expressed fixed cell fixed cell tag gfp tag green fluorescent protein tag GFP green fluorescent protein GFP green fluorescent protein gfp tag undetermined undetermined sequence position undetermined undetermined undetermined sequence position undetermined colocalization colocalization f1d f2a A significant amount of Godzilla clearly colocalizes with endosomal markers, such as EEA1, upon overexpression (Figure 1D; Supplementary Figure S2). While less extensive than that observed with Godzilla, overlap with EEA1 is also observed with the related Goliath (Figures 1D and 2A). cg10277_rd-eea1-1 1 7 immunostaining immunostaining immunostaining immunostaining unspecified role unspecified role neutral component neutral component physiological level physiological level endogenous level endogenous endogenous endogenous level fixed cell fixed cell 6 tag fluorescence tag visualisation by fluorescence tag fluorescence immunostaining immunostaining unspecified role unspecified role neutral component neutral component over-expressed over expressed level over-expressed fixed cell fixed cell tag gfp tag green fluorescent protein tag GFP green fluorescent protein GFP green fluorescent protein gfp tag undetermined undetermined sequence position undetermined undetermined undetermined sequence position undetermined colocalization colocalization f1d f2a A significant amount of Godzilla clearly colocalizes with endosomal markers, such as EEA1, upon overexpression (Figure 1D; Supplementary Figure S2). While less extensive than that observed with Godzilla, overlap with EEA1 is also observed with the related Goliath (Figures 1D and 2A). cg10277_rd-rab5 2 4 tag fluorescence tag visualisation by fluorescence tag fluorescence immunostaining immunostaining unspecified role unspecified role neutral component neutral component over-expressed over expressed level over-expressed fixed cell fixed cell tag yfp tag yellow fluorescent protein tag YFP yellow fluorescent protein yellow fluorescent protein YFP yfp tag undetermined undetermined sequence position undetermined undetermined undetermined sequence position undetermined 3 immunostaining immunostaining immunostaining immunostaining unspecified role unspecified role neutral component neutral component physiological level physiological level endogenous level endogenous endogenous endogenous level fixed cell fixed cell colocalization colocalization f1e sf1e-g in agreement with these results, endogenous Godzilla protein in Drosophila tissues displays a vesicle localization overlapping with endosomes (Supplementary Figure S1E–G). To confirm the presence of endogenous Godzilla protein in the endosomal compartment, we expressed activated Rab5—Rab5Q88LYFP (Stenmark et al, 1994) to produce enlarged endosomes in a number of tissues. Expression of activated Rab5–Rab5Q88LYFP in the wing disc leads to the accumulation of endogenous Godzilla protein on the resulting enlarged endosomes (Figure 1E). cg10277_rd-eea1-2 1 3 tag fluorescence tag visualisation by fluorescence tag fluorescence immunostaining immunostaining unspecified role unspecified role neutral component neutral component over-expressed over expressed level over-expressed fixed cell fixed cell tag ha tag ha tag YPYDVPDYA epitope tag YPYDVPDYA epitope tag undetermined undetermined sequence position undetermined undetermined undetermined sequence position undetermined 7 immunostaining immunostaining immunostaining immunostaining unspecified role unspecified role neutral component neutral component physiological level physiological level endogenous level endogenous endogenous endogenous level fixed cell fixed cell colocalization colocalization sf3 A significant amount of Godzilla clearly colocalizes with endosomal markers, such as EEA1, upon overexpression (Figure 1D; Supplementary Figure S2). While less extensive than that observed with Godzilla, overlap with EEA1 is also observed with the related Goliath (Figures 1D and 2A). eea1-gol 1 6 tag fluorescence tag visualisation by fluorescence tag fluorescence immunostaining immunostaining unspecified role unspecified role neutral component neutral component over-expressed over expressed level over-expressed fixed cell fixed cell tag ha tag ha tag YPYDVPDYA epitope tag YPYDVPDYA epitope tag undetermined undetermined sequence position undetermined undetermined undetermined sequence position undetermined 7 immunostaining immunostaining immunostaining immunostaining unspecified role unspecified role neutral component neutral component physiological level physiological level endogenous level endogenous endogenous endogenous level fixed cell fixed cell colocalization colocalization sf3 A significant amount of Godzilla clearly colocalizes with endosomal markers, such as EEA1, upon overexpression (Figure 1D; Supplementary Figure S2). While less extensive than that observed with Godzilla, overlap with EEA1 is also observed with the related Goliath (Figures 1D and 2A). cg10277_rd-eea1-3 1 3 tag fluorescence tag visualisation by fluorescence tag fluorescence immunostaining immunostaining unspecified role unspecified role neutral component neutral component over-expressed over expressed level over-expressed fixed cell fixed cell tag gfp tag green fluorescent protein tag GFP green fluorescent protein GFP green fluorescent protein gfp tag undetermined undetermined sequence position undetermined undetermined undetermined sequence position undetermined 8 tag fluorescence tag visualisation by fluorescence tag fluorescence immunostaining immunostaining unspecified role unspecified role neutral component neutral component over-expressed over expressed level over-expressed fixed cell fixed cell tag gfp tag green fluorescent protein tag GFP green fluorescent protein GFP green fluorescent protein gfp tag undetermined undetermined sequence position undetermined undetermined undetermined sequence position undetermined colocalization colocalization f5c In order to confirm whether VAMP3 is a potential target of Godzilla, we investigated VAMP3 localization in Godzilla‐transfected HEK293 cells. VAMP3 is known to localize on both early and recycling endosomes and is involved in endocytic recycling (McMahon et al, 1993; Galli et al, 1994). In both control and ligase‐dead Godzilla‐transfected cells, VAMP3 displays a similar intracellular distribution, partially overlapping with ligase‐dead Godzilla; suggesting an early and recycling endosome distribution (Figure 5C). In contrast, in cells expressing wild‐type Godzilla VAMP3 is found on enlarged endosomes, where it colocalizes with Godzilla (Figure 5C). cg10277_rd-vamp3 1 8 immunostaining immunostaining immunostaining immunostaining unspecified role unspecified role neutral component neutral component physiological level physiological level endogenous level endogenous endogenous endogenous level fixed cell fixed cell 3 tag fluorescence tag visualisation by fluorescence tag fluorescence immunostaining immunostaining unspecified role unspecified role neutral component neutral component over-expressed over expressed level over-expressed fixed cell fixed cell tag gfp tag green fluorescent protein tag GFP green fluorescent protein GFP green fluorescent protein gfp tag undetermined undetermined sequence position undetermined undetermined undetermined sequence position undetermined colocalization colocalization f5c In order to confirm whether VAMP3 is a potential target of Godzilla, we investigated VAMP3 localization in Godzilla‐transfected HEK293 cells. VAMP3 is known to localize on both early and recycling endosomes and is involved in endocytic recycling (McMahon et al, 1993; Galli et al, 1994). In both control and ligase‐dead Godzilla‐transfected cells, VAMP3 displays a similar intracellular distribution, partially overlapping with ligase‐dead Godzilla; suggesting an early and recycling endosome distribution (Figure 5C). In contrast, in cells expressing wild‐type Godzilla VAMP3 is found on enlarged endosomes, where it colocalizes with Godzilla (Figure 5C). eea1-gol-1 1 7 immunostaining immunostaining immunostaining immunostaining unspecified role unspecified role neutral component neutral component physiological level physiological level endogenous level endogenous endogenous endogenous level fixed cell fixed cell 5 tag fluorescence tag visualisation by fluorescence tag fluorescence immunostaining immunostaining unspecified role unspecified role neutral component neutral component over-expressed over expressed level over-expressed fixed cell fixed cell tag gfp tag green fluorescent protein tag GFP green fluorescent protein GFP green fluorescent protein gfp tag undetermined undetermined sequence position undetermined undetermined undetermined sequence position undetermined colocalization colocalization f6a Consistent with this we found that expression of human RNF167 also led to the accumulation of giant endosomes that were positive for EEA1, FK2 and VAMP3, with a concomitant loss of Rab11‐positive recycling endosomes (Figure 6A and B).