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/ ICOS ligation recruits the p50alpha PI3K regulatory subunit to the immunological synapse. ICOS ligation recruits the p50alpha PI3K regulatory subunit to the immunological synapse. Journal of immunology (Baltimore, Md. : 1950) 2008 imex curation Fos C., Salles A., Lang V., Carrette F., Audebert S., Pastor S., Ghiotto M., Olive D., Bismuth G., Nunès JA. Results from Figure 3 have not been entered as a chimeric mouse/human construct has been used and the results obtained are of dubious biological relevance. I have not entered interactions (Fig 1C) when there is any mention of L cells/L-B7.1, cells as it is unclear if these are Murine fibroblastic Ltk– cells (L cells) transfected with Human ICOS-L -see Ref 25, and ref 26 for the L-B7.1 cells where I cannot access full publication, so no idea of the origin of B7.1. Figure 1 states "resting T cells" under the LHS image - so are these Murine Fibroblastic cells, or Human T cells? Text states:- "As shown in Fig. 1C, (left panel), in resting T cells, p85a but not p50a is able to interact with CD28" - I only see any interaction for the L cells/L-B7.1 cells, not for the definitely human cells ( under NS). Mouse p85a Uniprot entry currently has NO isoforms, and given the ambiguity in cell lines used, and the fact I dont know if there would be enough evidence for Uniprot to add these isoforms anyway I have not requested an update. Interactions involving L cells have not been entered as the origin of B7.1 molecule in L-B7.1 cells cannot be traced and the species origin of cells used (L-cells) is unclear. Isoforms of p85a were mapped to Uniprot identifiers from reference 21. jacques.nunes@inserm.fr 09-May-2019: Contacted by IntAct-Help. Only protein-protein interactions imex curation human-t cells cd 4+ Homo sapiens t-cell_cd4+ t cell cd4+ anti bait coip anti bait coimmunoprecipitation anti bait coip western blot western blot Immuno blot Immuno blot Fos C., Salles A., Lang V., Carrette F., Audebert S., Pastor S., Ghiotto M., Olive D., Bismuth G., Nunès JA. Results from Figure 3 have not been entered as a chimeric mouse/human construct has been used and the results obtained are of dubious biological relevance. I have not entered interactions (Fig 1C) when there is any mention of L cells/L-B7.1, cells as it is unclear if these are Murine fibroblastic Ltk– cells (L cells) transfected with Human ICOS-L -see Ref 25, and ref 26 for the L-B7.1 cells where I cannot access full publication, so no idea of the origin of B7.1. Figure 1 states "resting T cells" under the LHS image - so are these Murine Fibroblastic cells, or Human T cells? Text states:- "As shown in Fig. 1C, (left panel), in resting T cells, p85a but not p50a is able to interact with CD28" - I only see any interaction for the L cells/L-B7.1 cells, not for the definitely human cells ( under NS). Mouse p85a Uniprot entry currently has NO isoforms, and given the ambiguity in cell lines used, and the fact I dont know if there would be enough evidence for Uniprot to add these isoforms anyway I have not requested an update. Interactions involving L cells have not been entered as the origin of B7.1 molecule in L-B7.1 cells cannot be traced and the species origin of cells used (L-cells) is unclear. Isoforms of p85a were mapped to Uniprot identifiers from reference 21. jacques.nunes@inserm.fr Accepted 2018-JUN-19 AT 12:52 BST AT 12:52 BST by PPORRAS Journal of immunology (Baltimore, Md. : 1950) 2008 Only protein-protein interactions imex curation imex curation ICOS ligation recruits the p50alpha PI3K regulatory subunit to the immunological synapse. ICOS ligation recruits the p50alpha PI3K regulatory subunit to the immunological synapse. Journal of immunology (Baltimore, Md. : 1950) 2008 imex curation Fos C., Salles A., Lang V., Carrette F., Audebert S., Pastor S., Ghiotto M., Olive D., Bismuth G., Nunès JA. Results from Figure 3 have not been entered as a chimeric mouse/human construct has been used and the results obtained are of dubious biological relevance. I have not entered interactions (Fig 1C) when there is any mention of L cells/L-B7.1, cells as it is unclear if these are Murine fibroblastic Ltk– cells (L cells) transfected with Human ICOS-L -see Ref 25, and ref 26 for the L-B7.1 cells where I cannot access full publication, so no idea of the origin of B7.1. Figure 1 states "resting T cells" under the LHS image - so are these Murine Fibroblastic cells, or Human T cells? Text states:- "As shown in Fig. 1C, (left panel), in resting T cells, p85a but not p50a is able to interact with CD28" - I only see any interaction for the L cells/L-B7.1 cells, not for the definitely human cells ( under NS). Mouse p85a Uniprot entry currently has NO isoforms, and given the ambiguity in cell lines used, and the fact I dont know if there would be enough evidence for Uniprot to add these isoforms anyway I have not requested an update. Interactions involving L cells have not been entered as the origin of B7.1 molecule in L-B7.1 cells cannot be traced and the species origin of cells used (L-cells) is unclear. Isoforms of p85a were mapped to Uniprot identifiers from reference 21. jacques.nunes@inserm.fr 09-May-2019: Contacted by IntAct-Help. Only protein-protein interactions imex curation human-t cells cd 4+ Homo sapiens t-cell_cd4+ t cell cd4+ confocal microscopy confocal microscopy immunocytochemistry immunocytochemistry Fos C., Salles A., Lang V., Carrette F., Audebert S., Pastor S., Ghiotto M., Olive D., Bismuth G., Nunès JA. Results from Figure 3 have not been entered as a chimeric mouse/human construct has been used and the results obtained are of dubious biological relevance. I have not entered interactions (Fig 1C) when there is any mention of L cells/L-B7.1, cells as it is unclear if these are Murine fibroblastic Ltk– cells (L cells) transfected with Human ICOS-L -see Ref 25, and ref 26 for the L-B7.1 cells where I cannot access full publication, so no idea of the origin of B7.1. Figure 1 states "resting T cells" under the LHS image - so are these Murine Fibroblastic cells, or Human T cells? Text states:- "As shown in Fig. 1C, (left panel), in resting T cells, p85a but not p50a is able to interact with CD28" - I only see any interaction for the L cells/L-B7.1 cells, not for the definitely human cells ( under NS). Mouse p85a Uniprot entry currently has NO isoforms, and given the ambiguity in cell lines used, and the fact I dont know if there would be enough evidence for Uniprot to add these isoforms anyway I have not requested an update. Activated T cells were transfected with the GFP-p50a construct and mixed with mature Dendritic Cells. Interactions involving L cells have not been entered as the origin of B7.1 molecule in L-B7.1 cells cannot be traced and the species origin of cells used (L-cells) is unclear. Isoforms of p85a were mapped to Uniprot identifiers from reference 21. jacques.nunes@inserm.fr Accepted 2018-JUN-19 AT 12:52 BST AT 12:52 BST by PPORRAS Journal of immunology (Baltimore, Md. : 1950) 2008 Only protein-protein interactions imex curation imex curation ICOS ligation recruits the p50alpha PI3K regulatory subunit to the immunological synapse. ICOS ligation recruits the p50alpha PI3K regulatory subunit to the immunological synapse. Journal of immunology (Baltimore, Md. : 1950) 2008 imex curation Fos C., Salles A., Lang V., Carrette F., Audebert S., Pastor S., Ghiotto M., Olive D., Bismuth G., Nunès JA. Results from Figure 3 have not been entered as a chimeric mouse/human construct has been used and the results obtained are of dubious biological relevance. I have not entered interactions (Fig 1C) when there is any mention of L cells/L-B7.1, cells as it is unclear if these are Murine fibroblastic Ltk– cells (L cells) transfected with Human ICOS-L -see Ref 25, and ref 26 for the L-B7.1 cells where I cannot access full publication, so no idea of the origin of B7.1. Figure 1 states "resting T cells" under the LHS image - so are these Murine Fibroblastic cells, or Human T cells? Text states:- "As shown in Fig. 1C, (left panel), in resting T cells, p85a but not p50a is able to interact with CD28" - I only see any interaction for the L cells/L-B7.1 cells, not for the definitely human cells ( under NS). Mouse p85a Uniprot entry currently has NO isoforms, and given the ambiguity in cell lines used, and the fact I dont know if there would be enough evidence for Uniprot to add these isoforms anyway I have not requested an update. Interactions involving L cells have not been entered as the origin of B7.1 molecule in L-B7.1 cells cannot be traced and the species origin of cells used (L-cells) is unclear. Isoforms of p85a were mapped to Uniprot identifiers from reference 21. jacques.nunes@inserm.fr 09-May-2019: Contacted by IntAct-Help. Only protein-protein interactions imex curation human-jurkat Homo sapiens jurkat Human T cell leukemia anti tag coip anti tag coimmunoprecipitation anti tag coip anti tag western anti tag western blot anti tag western Fos C., Salles A., Lang V., Carrette F., Audebert S., Pastor S., Ghiotto M., Olive D., Bismuth G., Nunès JA. Results from Figure 3 have not been entered as a chimeric mouse/human construct has been used and the results obtained are of dubious biological relevance. I have not entered interactions (Fig 1C) when there is any mention of L cells/L-B7.1, cells as it is unclear if these are Murine fibroblastic Ltk– cells (L cells) transfected with Human ICOS-L -see Ref 25, and ref 26 for the L-B7.1 cells where I cannot access full publication, so no idea of the origin of B7.1. Figure 1 states "resting T cells" under the LHS image - so are these Murine Fibroblastic cells, or Human T cells? Text states:- "As shown in Fig. 1C, (left panel), in resting T cells, p85a but not p50a is able to interact with CD28" - I only see any interaction for the L cells/L-B7.1 cells, not for the definitely human cells ( under NS). Mouse p85a Uniprot entry currently has NO isoforms, and given the ambiguity in cell lines used, and the fact I dont know if there would be enough evidence for Uniprot to add these isoforms anyway I have not requested an update. Interactions involving L cells have not been entered as the origin of B7.1 molecule in L-B7.1 cells cannot be traced and the species origin of cells used (L-cells) is unclear. Isoforms of p85a were mapped to Uniprot identifiers from reference 21. jacques.nunes@inserm.fr Accepted 2018-JUN-19 AT 12:52 BST AT 12:52 BST by PPORRAS Journal of immunology (Baltimore, Md. : 1950) 2008 Only protein-protein interactions imex curation imex curation p27986-3 Phosphatidylinositol 3-kinase regulatory subunit alpha p46 PIK3R1 GRB1 Phosphatidylinositol 3-kinase 85 kDa regulatory subunit alpha protein protein human Homo sapiens Human MHNLQTLPPKPPKPTTVANNGMNNNMSLQDAEWYWGDISREEVNEKLRDTADGTFLVRDASTKMHGDYTLTLRKGGNNKLIKIFHRDGKYGFSDPLTFSSVVELINHYRNESLAQYNPKLDVKLLYPVSKYQQDQVVKEDNIEAVGKKLHEYNTQFQEKSREYDRLYEEYTRTSQEIQMKRTAIEAFNETIKIFEEQCQTQERYSKEYIEKFKREGNEKEIQRIMHNYDKLKSRISEIIDSRRRLEEDLKKQAAEYREIDKRMNSIKPDLIQLRKTRDQYLMWLTQKGVRQKKLNEWLGNENTEDQYSLVEDDEDLPHHDEKTWNVGSSNRNKAENLLRGKRDGTFLVRESSKQGCYACSVVVDGEVKHCVINKTATGYGFAEPYNLYSSLKELVLHYQHTSLVQHNDSLNVTLAYPVYAQQRR EF2D6D4D668C62D8 p27986-2 Phosphatidylinositol 3-kinase regulatory subunit alpha PIK3R1 GRB1 Phosphatidylinositol 3-kinase 85 kDa regulatory subunit alpha AS53 protein protein human Homo sapiens Human MYNTVWNMEDLDLEYAKTDINCGTDLMFYIEMDPPALPPKPPKPTTVANNGMNNNMSLQDAEWYWGDISREEVNEKLRDTADGTFLVRDASTKMHGDYTLTLRKGGNNKLIKIFHRDGKYGFSDPLTFSSVVELINHYRNESLAQYNPKLDVKLLYPVSKYQQDQVVKEDNIEAVGKKLHEYNTQFQEKSREYDRLYEEYTRTSQEIQMKRTAIEAFNETIKIFEEQCQTQERYSKEYIEKFKREGNEKEIQRIMHNYDKLKSRISEIIDSRRRLEEDLKKQAAEYREIDKRMNSIKPDLIQLRKTRDQYLMWLTQKGVRQKKLNEWLGNENTEDQYSLVEDDEDLPHHDEKTWNVGSSNRNKAENLLRGKRDGTFLVRESSKQGCYACSVVVDGEVKHCVINKTATGYGFAEPYNLYSSLKELVLHYQHTSLVQHNDSLNVTLAYPVYAQQRR 33BB0F88AA47CB3F pk3cd_human Phosphatidylinositol 4,5-bisphosphate 3-kinase catalytic subunit delta isoform PIK3CD Phosphatidylinositol 4,5-bisphosphate 3-kinase 110 kDa catalytic subunit delta protein protein human Homo sapiens Human MPPGVDCPMEFWTKEENQSVVVDFLLPTGVYLNFPVSRNANLSTIKQLLWHRAQYEPLFHMLSGPEAYVFTCINQTAEQQELEDEQRRLCDVQPFLPVLRLVAREGDRVKKLINSQISLLIGKGLHEFDSLCDPEVNDFRAKMCQFCEEAAARRQQLGWEAWLQYSFPLQLEPSAQTWGPGTLRLPNRALLVNVKFEGSEESFTFQVSTKDVPLALMACALRKKATVFRQPLVEQPEDYTLQVNGRHEYLYGSYPLCQFQYICSCLHSGLTPHLTMVHSSSILAMRDEQSNPAPQVQKPRAKPPPIPAKKPSSVSLWSLEQPFRIELIQGSKVNADERMKLVVQAGLFHGNEMLCKTVSSSEVSVCSEPVWKQRLEFDINICDLPRMARLCFALYAVIEKAKKARSTKKKSKKADCPIAWANLMLFDYKDQLKTGERCLYMWPSVPDEKGELLNPTGTVRSNPNTDSAAALLICLPEVAPHPVYYPALEKILELGRHSECVHVTEEEQLQLREILERRGSGELYEHEKDLVWKLRHEVQEHFPEALARLLLVTKWNKHEDVAQMLYLLCSWPELPVLSALELLDFSFPDCHVGSFAIKSLRKLTDDELFQYLLQLVQVLKYESYLDCELTKFLLDRALANRKIGHFLFWHLRSEMHVPSVALRFGLILEAYCRGSTHHMKVLMKQGEALSKLKALNDFVKLSSQKTPKPQTKELMHLCMRQEAYLEALSHLQSPLDPSTLLAEVCVEQCTFMDSKMKPLWIMYSNEEAGSGGSVGIIFKNGDDLRQDMLTLQMIQLMDVLWKQEGLDLRMTPYGCLPTGDRTGLIEVVLRSDTIANIQLNKSNMAATAAFNKDALLNWLKSKNPGEALDRAIEEFTLSCAGYCVATYVLGIGDRHSDNIMIRESGQLFHIDFGHFLGNFKTKFGINRERVPFILTYDFVHVIQQGKTNNSEKFERFRGYCERAYTILRRHGLLFLHLFALMRAAGLPELSCSKDIQYLKDSLALGKTEEEALKHFRVKFNEALRESWKTKVNWLAHNVSKDNRQ A38B5D1A1081A3D0 A38B5D1A1081A3D0 p27986-1 Phosphatidylinositol 3-kinase regulatory subunit alpha Phosphatidylinositol 3-kinase 85 kDa regulatory subunit alpha PIK3R1 GRB1 protein protein human Homo sapiens Human MSAEGYQYRALYDYKKEREEDIDLHLGDILTVNKGSLVALGFSDGQEARPEEIGWLNGYNETTGERGDFPGTYVEYIGRKKISPPTPKPRPPRPLPVAPGSSKTEADVEQQALTLPDLAEQFAPPDIAPPLLIKLVEAIEKKGLECSTLYRTQSSSNLAELRQLLDCDTPSVDLEMIDVHVLADAFKRYLLDLPNPVIPAAVYSEMISLAPEVQSSEEYIQLLKKLIRSPSIPHQYWLTLQYLLKHFFKLSQTSSKNLLNARVLSEIFSPMLFRFSAASSDNTENLIKVIEILISTEWNERQPAPALPPKPPKPTTVANNGMNNNMSLQDAEWYWGDISREEVNEKLRDTADGTFLVRDASTKMHGDYTLTLRKGGNNKLIKIFHRDGKYGFSDPLTFSSVVELINHYRNESLAQYNPKLDVKLLYPVSKYQQDQVVKEDNIEAVGKKLHEYNTQFQEKSREYDRLYEEYTRTSQEIQMKRTAIEAFNETIKIFEEQCQTQERYSKEYIEKFKREGNEKEIQRIMHNYDKLKSRISEIIDSRRRLEEDLKKQAAEYREIDKRMNSIKPDLIQLRKTRDQYLMWLTQKGVRQKKLNEWLGNENTEDQYSLVEDDEDLPHHDEKTWNVGSSNRNKAENLLRGKRDGTFLVRESSKQGCYACSVVVDGEVKHCVINKTATGYGFAEPYNLYSSLKELVLHYQHTSLVQHNDSLNVTLAYPVYAQQRR B9DAD8416C33140F icos_human Inducible T-cell costimulator ICOS AILIM Activation-inducible lymphocyte immunomediatory molecule protein protein human Homo sapiens Human MKSGLWYFFLFCLRIKVLTGEINGSANYEMFIFHNGGVQILCKYPDIVQQFKMQLLKGGQILCDLTKTKGSGNTVSIKSLKFCHSQLSNNSVSFFLYNLDHSHANYYFCNLSIFDPPPFKVTLTGGYLHIYESQLCCQLKFWLPIGCAAFVVVCILGCILICWLTKKKYSSSVHDPNGEYMFMRAVNTAKKSRLTDVTL 214EC741C9BDC9FC p85a_human Phosphatidylinositol 3-kinase regulatory subunit alpha PIK3R1 GRB1 Phosphatidylinositol 3-kinase 85 kDa regulatory subunit alpha protein protein human Homo sapiens Human MSAEGYQYRALYDYKKEREEDIDLHLGDILTVNKGSLVALGFSDGQEARPEEIGWLNGYNETTGERGDFPGTYVEYIGRKKISPPTPKPRPPRPLPVAPGSSKTEADVEQQALTLPDLAEQFAPPDIAPPLLIKLVEAIEKKGLECSTLYRTQSSSNLAELRQLLDCDTPSVDLEMIDVHVLADAFKRYLLDLPNPVIPAAVYSEMISLAPEVQSSEEYIQLLKKLIRSPSIPHQYWLTLQYLLKHFFKLSQTSSKNLLNARVLSEIFSPMLFRFSAASSDNTENLIKVIEILISTEWNERQPAPALPPKPPKPTTVANNGMNNNMSLQDAEWYWGDISREEVNEKLRDTADGTFLVRDASTKMHGDYTLTLRKGGNNKLIKIFHRDGKYGFSDPLTFSSVVELINHYRNESLAQYNPKLDVKLLYPVSKYQQDQVVKEDNIEAVGKKLHEYNTQFQEKSREYDRLYEEYTRTSQEIQMKRTAIEAFNETIKIFEEQCQTQERYSKEYIEKFKREGNEKEIQRIMHNYDKLKSRISEIIDSRRRLEEDLKKQAAEYREIDKRMNSIKPDLIQLRKTRDQYLMWLTQKGVRQKKLNEWLGNENTEDQYSLVEDDEDLPHHDEKTWNVGSSNRNKAENLLRGKRDGTFLVRESSKQGCYACSVVVDGEVKHCVINKTATGYGFAEPYNLYSSLKELVLHYQHTSLVQHNDSLNVTLAYPVYAQQRR DIP protein P27986 original sequence version: 181 DIP protein P27986 original sequence version: 184 DIP protein P27986-1 original sequence version: 135 DIP protein P27986 original sequence version: 135 DIP protein P27986-1 original sequence version: 145 DIP protein P27986-1 original sequence version: 147 DIP protein P27986 original sequence version: 191 B9DAD8416C33140F cd28_human T-cell-specific surface glycoprotein CD28 CD28 TP44 protein protein human Homo sapiens Human MLRLLLALNLFPSIQVTGNKILVKQSPMLVAYDNAVNLSCKYSYNLFSREFRASLHKGLDSAVEVCVVYGNYSQQLQVYSKTGFNCDGKLGNESVTFYLQNLYVNQTDIYFCKIEVMYPPPYLDNEKSNGTIIHVKGKHLCPSPLFPGPSKPFWVLVVVGGVLACYSLLVTVAFIIFWVRSKRSRLLHSDYMNMTPRRPGPTRKHYQPYAPPRDFAAYRS 1D9B6552A5878D0F icos-pik3r1-5 1 5 unspecified role unspecified role prey prey physiological level physiological level endogenous level endogenous endogenous endogenous level 9 unspecified role unspecified role prey prey physiological level physiological level endogenous level endogenous endogenous endogenous level 8 unspecified role unspecified role bait bait physiological level physiological level endogenous level endogenous endogenous endogenous level association association Figure 1C Activated T cells (stimulated by anti-CD3 and CD28 to induce ICOS expression) and then either re-stimulated or not re-stimulated using the same antibodies. icos-pik3r1-6 1 4 unspecified role unspecified role prey prey physiological level physiological level endogenous level endogenous endogenous endogenous level 5 unspecified role unspecified role prey prey physiological level physiological level endogenous level endogenous endogenous endogenous level 9 unspecified role unspecified role prey prey physiological level physiological level endogenous level endogenous endogenous endogenous level 8 unspecified role unspecified role bait bait physiological level physiological level endogenous level endogenous endogenous endogenous level association association Figure 1Ci Activated T cells (stimulated by anti-CD3 and CD28 to induce ICOS expression) and then either re-stimulated using the anti-CD3 and anti-ICOS antibodies. icos-pik3r1-7 1 5 unspecified role unspecified role prey prey physiological level physiological level endogenous level endogenous endogenous endogenous level 9 unspecified role unspecified role prey prey physiological level physiological level endogenous level endogenous endogenous endogenous level 10 unspecified role unspecified role bait bait physiological level physiological level endogenous level endogenous endogenous endogenous level association association Figure 1Ciii Activated T cells (stimulated by anti-CD3 and CD28 to induce ICOS expression) and then either re-stimulated or not re-stimulated using the same antibodies. icos-pik3r1-8 1 4 unspecified role unspecified role prey prey physiological level physiological level endogenous level endogenous endogenous endogenous level 5 unspecified role unspecified role prey prey physiological level physiological level endogenous level endogenous endogenous endogenous level 9 unspecified role unspecified role prey prey physiological level physiological level endogenous level endogenous endogenous endogenous level 10 unspecified role unspecified role bait bait physiological level physiological level endogenous level endogenous endogenous endogenous level association association Figure 1Civ Activated T cells (stimulated by anti-CD3 and CD28 to induce ICOS expression) and then either re-stimulated using the anti-CD3 and anti-ICOS antibodies. 2423rawtetg-1 3 6 unspecified role unspecified role prey prey over-expressed over expressed level over-expressed region myc tag myc tag EUKLISEED epitope tag EUKLISEED epitope tag undetermined undetermined sequence position undetermined undetermined undetermined sequence position undetermined 7 unspecified role unspecified role bait bait over-expressed over expressed level over-expressed n_terminus gfp tag green fluorescent protein tag GFP green fluorescent protein GFP green fluorescent protein gfp tag n-term range n-terminal range n-term range n-term range n-terminal range n-term range physical association physical association figure 2B pik3r1-pik3cd-5 3 6 unspecified role unspecified role prey prey over-expressed over expressed level over-expressed region myc tag myc tag EUKLISEED epitope tag EUKLISEED epitope tag undetermined undetermined sequence position undetermined undetermined undetermined sequence position undetermined 4 unspecified role unspecified role bait bait over-expressed over expressed level over-expressed n_terminus gfp tag green fluorescent protein tag GFP green fluorescent protein GFP green fluorescent protein gfp tag n-term range n-terminal range n-term range n-term range n-terminal range n-term range physical association physical association figure 2Bi icos-pik3r1-9 2 8 unspecified role unspecified role neutral component neutral component physiological level physiological level endogenous level endogenous endogenous endogenous level 4 tag fluorescence tag visualisation by fluorescence tag fluorescence immunocytochemistry immunocytochemistry unspecified role unspecified role neutral component neutral component over-expressed over expressed level over-expressed n_terminus gfp tag green fluorescent protein tag GFP green fluorescent protein GFP green fluorescent protein gfp tag n-term range n-terminal range n-term range n-term range n-terminal range n-term range colocalization colocalization figure 4 ICOS and p50a colocalized at the contact zone.