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 Pilt is a coiled-coil domain-containing protein that localizes at the trans-Golgi complex and regulates its structure. Pilt is a coiled-coil domain-containing protein that localizes at the trans-Golgi complex and regulates its structure. FEBS Lett. (0014-5793) 2012 Tamaki H., Sanda M., Katsumata O., Hara Y., Fukaya M., Sakagami H. yeast Saccharomyces cerevisiae 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 Tamaki H., Sanda M., Katsumata O., Hara Y., Fukaya M., Sakagami H. FEBS Lett. (0014-5793) 2012 Pilt is a coiled-coil domain-containing protein that localizes at the trans-Golgi complex and regulates its structure. Pilt is a coiled-coil domain-containing protein that localizes at the trans-Golgi complex and regulates its structure. FEBS Lett. (0014-5793) 2012 Tamaki H., Sanda M., Katsumata O., Hara Y., Fukaya M., Sakagami H. human-hela Homo sapiens epitheloid cervix carcinoma cells Human hela Human epitheloid cervix carcinoma HeLa cells derivation: cervix; epithelial; adenocarcinoma ATCC no:CCL-2 fluorescence imaging fluorescence microscopy fluorescence imaging predetermined predetermined participant predetermined Tamaki H., Sanda M., Katsumata O., Hara Y., Fukaya M., Sakagami H. FEBS Lett. (0014-5793) 2012 Pilt is a coiled-coil domain-containing protein that localizes at the trans-Golgi complex and regulates its structure. Pilt is a coiled-coil domain-containing protein that localizes at the trans-Golgi complex and regulates its structure. FEBS Lett. (0014-5793) 2012 Tamaki H., Sanda M., Katsumata O., Hara Y., Fukaya M., Sakagami H. in vitro In vitro pull down pull down predetermined predetermined participant predetermined Tamaki H., Sanda M., Katsumata O., Hara Y., Fukaya M., Sakagami H. FEBS Lett. (0014-5793) 2012 q9dcd5-2 Tight junction-associated protein 1 Tjap1 Pilt Protein incorporated later into tight junctions Tight junction protein 4 Tjp4 protein protein mouse Mus musculus Mouse MSSAAPAKKPYRKAPPEHRELRLEIPVSRLEQEESLTDAERMKLLQQENEELRKRLASATRRTEALERELEIGQDCLELELGQSREELDKFKDKFRRLQNSYTASQRTNQELEDKLHALIKKAEMDRKTLDWEIVELTNKLLDARNTINKLEELNERYRLDCNLAVQLLKCNKSHFRNHKLADLPCELQDMVRKHLRSGQEVASPSPSPSSSLSPGAVVPTSVIARVLEKPESLLLNSAQSGSAGRPLAEDVFVHVDMSGGDPASPPAPGSPNGECCSVSTAGGSPEEELPLPAFDKLSPYPTPSPPHPLYPGRKVIEFSEDKIRIPRNSPLPNCTYATRQAISLSLVEDGSERAHRSSVPSSPASAQGSPHHQPSPAPSALSAPASSASSEEDLLASWQRAFVDRTPPPAAVVQRTAFGRDSLPELQLHFSPGHSTAPPPSPHRERGLVLPAEPDSGFPQDEEEEMLNLPVSPEEERQSLLPDKEGTEEASGPSHVDGRAWPLPSPSRPQRSPKRMGVHHLHRKDSLTQAQEQGTVLS FD169E54AD29E4DA arf6_mouse ADP-ribosylation factor 6 Arf6 protein protein mouse Mus musculus Mouse MGKVLSKIFGNKEMRILMLGLDAAGKTTILYKLKLGQSVTTIPTVGFNVETVTYKNVKFNVWDVGGQDKIRPLWRHYYTGTQGLIFVVDCADRDRIDEARQELHRIINDREMRDAIILIFANKQDLPDAMKPHEIQEKLGLTRIRDRNWYVQPSCATSGDGLYEGLTWLTSNYKS 49E38E59AEA52B98 tjap1_mouse Tight junction-associated protein 1 Tight junction protein 4 Protein incorporated later into tight junctions Tjap1 Pilt Tjp4 protein protein mouse Mus musculus Mouse MSSAAPAKKPYRKAPPEHRELRLEIPVSRLEQEESLTDAERMKLLQQENEELRKRLASATRRTEALERELEIGQDCLELELGQSREELDKFKDKFRRLQNSYTASQRTNQELEDKLHALASLSHSWIFAIKKAEMDRKTLDWEIVELTNKLLDARNTINKLEELNERYRLDCNLAVQLLKCNKSHFRNHKLADLPCELQDMVRKHLRSGQEVASPSPSPSSSLSPGAVVPTSVIARVLEKPESLLLNSAQSGSAGRPLAEDVFVHVDMSGGDPASPPAPGSPNGECCSVSTAGGSPEEELPLPAFDKLSPYPTPSPPHPLYPGRKVIEFSEDKIRIPRNSPLPNCTYATRQAISLSLVEDGSERAHRSSVPSSPASAQGSPHHQPSPAPSALSAPASSASSEEDLLASWQRAFVDRTPPPAAVVQRTAFGRDSLPELQLHFSPGHSTAPPPSPHRERGLVLPAEPDSGFPQDEEEEMLNLPVSPEEERQSLLPDKEGTEEASGPSHVDGRAWPLPSPSRPQRSPKRMGVHHLHRKDSLTQAQEQGTVLS 267C34C0BC7EB85A goga2_human Golgin subfamily A member 2 GOLGA2 130 kDa cis-Golgi matrix protein GM130 autoantigen Golgin-95 protein protein human Homo sapiens Human MWPQPRLPPRPAMSEETRQSKLAAAKKKLREYQQRNSPGVPTGAKKKKKIKNGSNPETTTSGGCHSPEDTPKDNAATLQPSDDTVLPGGVPSPGASLTSMAASQNHDADNVPNLMDETKTFSSTESLRQLSQQLNGLVCESATCVNGEGPASSANLKDLESRYQQLAVALDSSYVTNKQLNITIEKLKQQNQEITDQLEEEKKECHQKQGALREQLQVHIQTIGILVSEKAELQTALAHTQHAARQKEGESEDLASRLQYSRRRVGELERALSAVSTQQKKADRYNKELTKERDALRLELYKNTQSNEDLKQEKSELEEKLRVLVTEKAGMQLNLEELQKKLEMTELLLQQFSSRCEAPDANQQLQQAMEERAQLEAHLGQVMESVRQLQMERDKYAENLKGESAMWRQRMQQMSEQVHTLREEKECSMSRVQELETSLAELRNQMAEPPPPEPPAGPSEVEQQLQAEAEHLRKELEGLAGQLQAQVQDNEGLSRLNREQEERLLELERAAELWGEQAEARRQILETMQNDRTTISRALSQNRELKEQLAELQSGFVKLTNENMEITSALQSEQHVKRELGKKLGELQEKLSELKETVELKSQEAQSLQQQRDQYLGHLQQYVAAYQQLTSEKEVLHNQLLLQTQLVDQLQQQEAQGKAVAEMARQELQETQERLEAATQQNQQLRAQLSLMAHPGEGDGLDREEEEDEEEEEEEAVAVPQPMPSIPEDLESREAMVAFFNSAVASAEEEQARLRGQLKEQRVRCRRLAHLLASAQKEPEAAAPAPGTGGDSVCGETHRALQGAMEKLQSRFMELMQEKADLKERVEELEHRCIQLSGETDTIGEYIALYQSQRAVLKERHREKEEYISRLAQDKEEMKVKLLELQELVLRLVGDRNEWHGRFLAAAQNPADEPTSGAPAPQELGAANQQGDLCEVSLAGSVEPAQGEAREGSPRDNPTAQQIMQLLREMQNPRERPGLGSNPCIPFFYRADENDEVKITVI BD067145C5092237 arf6-tjap1 1 Pilt 6 nucleotide sequence nucleotide sequence identification sequence cloning sequence cloning nucleotide sequence predetermined predetermined participant predetermined unspecified role unspecified role prey prey cdna library cdna library fusion protein fusion protein fusion protein undetermined undetermined sequence position undetermined undetermined undetermined sequence position undetermined Arf6(Q67L) 5 predetermined predetermined participant predetermined predetermined predetermined participant predetermined unspecified role unspecified role bait bait over-expressed over expressed level over-expressed living cell living cell fusion protein fusion protein fusion protein undetermined undetermined sequence position undetermined undetermined undetermined sequence position undetermined physical association physical association f1a mint Schematic representation of the clone 125 and Pilt mutants used to determine the region responsible for the interaction with Arf6. Note the presence of a 10-amino acid insertion in clone 125 compared with the amino acid sequence of TJAP1, a mouse orthologue of human Pilt. (B) Alignment of amino acid sequences between clone 125 and TJAP1 (accession number NM_028751). Nucleotide numbers on the left indicate the positions from the translation initiation sites. The nucleotide and amino acid sequences inserted into clone 125 are shown in the box. The sequence of clone 125 has been deposited in DDJ under the accession number AB712252. arf6-tjap1-1 1 Arf6(Q67L) 5 predetermined predetermined participant predetermined predetermined predetermined participant predetermined unspecified role unspecified role bait bait over-expressed over expressed level over-expressed living cell living cell fusion protein fusion protein fusion protein undetermined undetermined sequence position undetermined undetermined undetermined sequence position undetermined Pilt 4 predetermined predetermined participant predetermined predetermined predetermined participant predetermined unspecified role unspecified role prey prey over-expressed over expressed level over-expressed living cell living cell fusion protein fusion protein fusion protein undetermined undetermined sequence position undetermined undetermined undetermined sequence position undetermined binding site binding region binding-associated region binding region deletion analysis deletion analysis certain certain sequence position certain certain certain sequence position certain physical association physical association f1c f1d mint tjap1-arf6 3 Arf6(Q67L) 5 anti tag western anti tag western blot anti tag western predetermined predetermined participant predetermined unspecified role unspecified role prey prey over-expressed over expressed level over-expressed cell lysate cell lysate P62331:p.Thr44Asn mutation disrupting interaction mutation disrupting mutation disrupting interaction mutation disrupting mutation analysis mutation analysis certain certain sequence position certain certain certain sequence position certain T N Tue Jul 12 10:13:46 BST 2016 This feature has been corrected as a result of our quality control procedures. The original label was 'mutation disrupting' Wed May 02 18:02:09 BST 2018 This feature has been corrected as a result of our quality control procedures. The original label was 'thr44asn' Tue Jun 30 16:21:09 BST 2020 This feature has been corrected as a result of our quality control procedures. The original label was 'p.Thr44Asn' ha tag ha tag ha tag YPYDVPDYA epitope tag YPYDVPDYA epitope tag undetermined undetermined sequence position undetermined undetermined undetermined sequence position undetermined Pilt 6 predetermined predetermined participant predetermined predetermined predetermined participant predetermined unspecified role unspecified role bait bait purified purified gst tag 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 physical association physical association f1e mint An in vitro GST pull down assay demonstrated that GST-Pilt(1-549) efficiently pulled down Arf6(Q67L)-HA and wild-type Arf6-HA incubated with GTPγS, but not Arf6(T44N)-HA, a GDP-locked mutant (Fig. 1E). golga2-tjap1 2 GM130 7 immunostaining immunostaining predetermined predetermined participant predetermined unspecified role unspecified role neutral component neutral component physiological level physiological level endogenous level endogenous endogenous endogenous level fixed cell fixed cell Pilt 6 anti tag immunost anti tag immunostaining anti tag immunost predetermined predetermined participant predetermined unspecified role unspecified role neutral component neutral component over-expressed over expressed level over-expressed fixed cell fixed cell myc tag myc tag myc tag EUKLISEED epitope tag EUKLISEED epitope tag undetermined undetermined sequence position undetermined undetermined undetermined sequence position undetermined binding site binding region binding-associated region binding region certain certain sequence position certain certain certain sequence position certain colocalization colocalization mint f4 sf2 To determine the region of Pilt responsible for the Golgi targeting, the localizations of 7 various Pilt mutants transfected in COS-7 cells were examined by immunostaining with antibodies against c-Myc and GM130 (Fig. 4). When c-Myc-Pilt(1-549) was expressed at a low level, it was closely associated with GM130 as described for endogenous Pilt (Fig. 4B). However, when expressed at moderate to high levels, it exhibited predominant cytoplasmic distribution that masked underlying structures, suggesting the Golgi localization of Pilt in a saturable manner. When expressed at a weak level, c-Myc-Pilt(176-549) was efficiently targeted to the Golgi, while c-Myc-Pilt(1-175) failed to be targeted to the Golgi and was distributed as aggregates unrelated to GM130 (Fig. 4B). Further deletion revealed that c-Myc-Pilt(176-439), (176-439), (311-549) and (311-439) exhibited perinuclear Golgi patterns (Fig. 4B). Therefore, the C-terminal two-thirds, particularly a central region corresponding to amino acids 301-439, are suggested to be important for the Golgi localization. The predominant localization of Pilt at the Golgi complex was also true for other cell types including HeLa cells, intestinal absorptive cells, epididymal epithelial cells and adrenal chromaffin cells (Supplementary Fig. 2)