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/ Structural basis of inducible costimulator ligand costimulatory function: determination of the cell surface oligomeric state and functional mapping of the receptor binding site of the protein. Structural basis of inducible costimulator ligand costimulatory function: determination of the cell surface oligomeric state and functional mapping of the receptor binding site of the protein. Journal of immunology (Baltimore, Md. : 1950) 2006 imex curation Chattopadhyay K., Bhatia S., Fiser A., Almo SC., Nathenson SG. Fig4A + elsewhere- no aas are given for the ICOSL-ΔC-YFP construct - so have assumed it must full length minus aas 136-252 approx. nathenso@aecom.yu.edu,almo@aecom.yu.edu 27-Mar-2018: Contacted by IntAct-Help. Only protein-protein interactions imex curation in vitro In vitro comig non denat gel comigration in non denaturing gel electrophoresis comig non denat gel weight by comassie molecular weight estimation by coomasie staining weight by comassie Chattopadhyay K., Bhatia S., Fiser A., Almo SC., Nathenson SG. Fig4A + elsewhere- no aas are given for the ICOSL-ΔC-YFP construct - so have assumed it must full length minus aas 136-252 approx. nathenso@aecom.yu.edu,almo@aecom.yu.edu Accepted 2018-FEB-09 AT 08:45 GMT AT 08:45 GMT by PPORRAS Journal of immunology (Baltimore, Md. : 1950) 2006 Only protein-protein interactions imex curation imex curation Fig 2C:- I have altered Exp -prep to:- 1]sub-cellular prep, 2] over-expressed, and 3] altered nucl. transfection to a child term, that is more correct, as LipofectAMINE 2000 or Fugene6 were used for transfection. 4] Plasma membrane added as a GO term at Interaction level, as membranes were extracted after X-linking.; I have altered nucl. transfection to child term, as before, and added over-expressed to both participants.; altered; new interaction added - so one for each form.; Exp-Mod added. I have altered Exp-prep as before. Also added a Sufficient to bind in Fuzzy type, as they do not specify the exact sequence for this construct - if they used the ICOSL-V construct as before, presumably they would have stated this fact. (I had already added a Comment) Structural basis of inducible costimulator ligand costimulatory function: determination of the cell surface oligomeric state and functional mapping of the receptor binding site of the protein. Structural basis of inducible costimulator ligand costimulatory function: determination of the cell surface oligomeric state and functional mapping of the receptor binding site of the protein. Journal of immunology (Baltimore, Md. : 1950) 2006 imex curation Chattopadhyay K., Bhatia S., Fiser A., Almo SC., Nathenson SG. Fig4A + elsewhere- no aas are given for the ICOSL-ΔC-YFP construct - so have assumed it must full length minus aas 136-252 approx. nathenso@aecom.yu.edu,almo@aecom.yu.edu 27-Mar-2018: Contacted by IntAct-Help. Only protein-protein interactions imex curation in vitro In vitro far western blotting far western blotting Affinity blotting Affinity blotting western blot western blot Immuno blot Immuno blot Chattopadhyay K., Bhatia S., Fiser A., Almo SC., Nathenson SG. Fig4A + elsewhere- no aas are given for the ICOSL-ΔC-YFP construct - so have assumed it must full length minus aas 136-252 approx. native PAGE - non-denaturing. nathenso@aecom.yu.edu,almo@aecom.yu.edu Accepted 2018-FEB-09 AT 08:45 GMT AT 08:45 GMT by PPORRAS Journal of immunology (Baltimore, Md. : 1950) 2006 Only protein-protein interactions imex curation imex curation Fig 2C:- I have altered Exp -prep to:- 1]sub-cellular prep, 2] over-expressed, and 3] altered nucl. transfection to a child term, that is more correct, as LipofectAMINE 2000 or Fugene6 were used for transfection. 4] Plasma membrane added as a GO term at Interaction level, as membranes were extracted after X-linking.; I have altered nucl. transfection to child term, as before, and added over-expressed to both participants.; altered; new interaction added - so one for each form.; Exp-Mod added. I have altered Exp-prep as before. Also added a Sufficient to bind in Fuzzy type, as they do not specify the exact sequence for this construct - if they used the ICOSL-V construct as before, presumably they would have stated this fact. (I had already added a Comment) Structural basis of inducible costimulator ligand costimulatory function: determination of the cell surface oligomeric state and functional mapping of the receptor binding site of the protein. Structural basis of inducible costimulator ligand costimulatory function: determination of the cell surface oligomeric state and functional mapping of the receptor binding site of the protein. Journal of immunology (Baltimore, Md. : 1950) 2006 imex curation Chattopadhyay K., Bhatia S., Fiser A., Almo SC., Nathenson SG. Fig4A + elsewhere- no aas are given for the ICOSL-ΔC-YFP construct - so have assumed it must full length minus aas 136-252 approx. nathenso@aecom.yu.edu,almo@aecom.yu.edu 27-Mar-2018: Contacted by IntAct-Help. Only protein-protein interactions imex curation in vitro In vitro facs fluorescence-activated cell sorting FACS Flow cytometry facs antibody detection identification by antibody antibody detection Chattopadhyay K., Bhatia S., Fiser A., Almo SC., Nathenson SG. Fig4A + elsewhere- no aas are given for the ICOSL-ΔC-YFP construct - so have assumed it must full length minus aas 136-252 approx. native PAGE - non-denaturing. nathenso@aecom.yu.edu,almo@aecom.yu.edu Accepted 2018-FEB-09 AT 08:45 GMT AT 08:45 GMT by PPORRAS Journal of immunology (Baltimore, Md. : 1950) 2006 Only protein-protein interactions imex curation imex curation Fig 2C:- I have altered Exp -prep to:- 1]sub-cellular prep, 2] over-expressed, and 3] altered nucl. transfection to a child term, that is more correct, as LipofectAMINE 2000 or Fugene6 were used for transfection. 4] Plasma membrane added as a GO term at Interaction level, as membranes were extracted after X-linking.; I have altered nucl. transfection to child term, as before, and added over-expressed to both participants.; altered; new interaction added - so one for each form.; Exp-Mod added. I have altered Exp-prep as before. Also added a Sufficient to bind in Fuzzy type, as they do not specify the exact sequence for this construct - if they used the ICOSL-V construct as before, presumably they would have stated this fact. (I had already added a Comment) Structural basis of inducible costimulator ligand costimulatory function: determination of the cell surface oligomeric state and functional mapping of the receptor binding site of the protein. Structural basis of inducible costimulator ligand costimulatory function: determination of the cell surface oligomeric state and functional mapping of the receptor binding site of the protein. Journal of immunology (Baltimore, Md. : 1950) 2006 imex curation Chattopadhyay K., Bhatia S., Fiser A., Almo SC., Nathenson SG. Fig4A + elsewhere- no aas are given for the ICOSL-ΔC-YFP construct - so have assumed it must full length minus aas 136-252 approx. nathenso@aecom.yu.edu,almo@aecom.yu.edu 27-Mar-2018: Contacted by IntAct-Help. Only protein-protein interactions imex curation in vitro In vitro fret fluorescent resonance energy transfer FRET RET FRET analysis FRET RET FRET analysis fret tag fluorescence tag visualisation by fluorescence tag fluorescence Chattopadhyay K., Bhatia S., Fiser A., Almo SC., Nathenson SG. Fig4A + elsewhere- no aas are given for the ICOSL-ΔC-YFP construct - so have assumed it must full length minus aas 136-252 approx. Photobleaching-based FRET. nathenso@aecom.yu.edu,almo@aecom.yu.edu Accepted 2018-FEB-09 AT 08:45 GMT AT 08:45 GMT by PPORRAS Journal of immunology (Baltimore, Md. : 1950) 2006 Only protein-protein interactions imex curation imex curation Fig 2C:- I have altered Exp -prep to:- 1]sub-cellular prep, 2] over-expressed, and 3] altered nucl. transfection to a child term, that is more correct, as LipofectAMINE 2000 or Fugene6 were used for transfection. 4] Plasma membrane added as a GO term at Interaction level, as membranes were extracted after X-linking.; I have altered nucl. transfection to child term, as before, and added over-expressed to both participants.; altered; new interaction added - so one for each form.; Exp-Mod added. I have altered Exp-prep as before. Also added a Sufficient to bind in Fuzzy type, as they do not specify the exact sequence for this construct - if they used the ICOSL-V construct as before, presumably they would have stated this fact. (I had already added a Comment) Structural basis of inducible costimulator ligand costimulatory function: determination of the cell surface oligomeric state and functional mapping of the receptor binding site of the protein. Structural basis of inducible costimulator ligand costimulatory function: determination of the cell surface oligomeric state and functional mapping of the receptor binding site of the protein. Journal of immunology (Baltimore, Md. : 1950) 2006 imex curation Chattopadhyay K., Bhatia S., Fiser A., Almo SC., Nathenson SG. Fig4A + elsewhere- no aas are given for the ICOSL-ΔC-YFP construct - so have assumed it must full length minus aas 136-252 approx. nathenso@aecom.yu.edu,almo@aecom.yu.edu 27-Mar-2018: Contacted by IntAct-Help. Only protein-protein interactions imex curation in vitro In vitro crosslink cross-linking study crosslink weight by comassie molecular weight estimation by coomasie staining weight by comassie Chattopadhyay K., Bhatia S., Fiser A., Almo SC., Nathenson SG. Fig4A + elsewhere- no aas are given for the ICOSL-ΔC-YFP construct - so have assumed it must full length minus aas 136-252 approx. Crosslinker:BS3 (bis(sulfosuccinimidyl)suberate), in presence of 2-mercaptoethanol, thus reducing disulfide linkages. nathenso@aecom.yu.edu,almo@aecom.yu.edu Accepted 2018-FEB-09 AT 08:45 GMT AT 08:45 GMT by PPORRAS Journal of immunology (Baltimore, Md. : 1950) 2006 Only protein-protein interactions imex curation imex curation Fig 2C:- I have altered Exp -prep to:- 1]sub-cellular prep, 2] over-expressed, and 3] altered nucl. transfection to a child term, that is more correct, as LipofectAMINE 2000 or Fugene6 were used for transfection. 4] Plasma membrane added as a GO term at Interaction level, as membranes were extracted after X-linking.; I have altered nucl. transfection to child term, as before, and added over-expressed to both participants.; altered; new interaction added - so one for each form.; Exp-Mod added. I have altered Exp-prep as before. Also added a Sufficient to bind in Fuzzy type, as they do not specify the exact sequence for this construct - if they used the ICOSL-V construct as before, presumably they would have stated this fact. (I had already added a Comment) Structural basis of inducible costimulator ligand costimulatory function: determination of the cell surface oligomeric state and functional mapping of the receptor binding site of the protein. Structural basis of inducible costimulator ligand costimulatory function: determination of the cell surface oligomeric state and functional mapping of the receptor binding site of the protein. Journal of immunology (Baltimore, Md. : 1950) 2006 imex curation Chattopadhyay K., Bhatia S., Fiser A., Almo SC., Nathenson SG. Fig4A + elsewhere- no aas are given for the ICOSL-ΔC-YFP construct - so have assumed it must full length minus aas 136-252 approx. nathenso@aecom.yu.edu,almo@aecom.yu.edu 27-Mar-2018: Contacted by IntAct-Help. Only protein-protein interactions imex curation in vitro In vitro molecular sieving molecular sieving Gel Filtration Size Exclusion Chromatography Size Exclusion Chromatography Gel Filtration Sizing column predetermined predetermined participant predetermined Chattopadhyay K., Bhatia S., Fiser A., Almo SC., Nathenson SG. Fig4A + elsewhere- no aas are given for the ICOSL-ΔC-YFP construct - so have assumed it must full length minus aas 136-252 approx. nathenso@aecom.yu.edu,almo@aecom.yu.edu Accepted 2018-FEB-09 AT 08:45 GMT AT 08:45 GMT by PPORRAS Journal of immunology (Baltimore, Md. : 1950) 2006 Only protein-protein interactions imex curation imex curation Fig 2C:- I have altered Exp -prep to:- 1]sub-cellular prep, 2] over-expressed, and 3] altered nucl. transfection to a child term, that is more correct, as LipofectAMINE 2000 or Fugene6 were used for transfection. 4] Plasma membrane added as a GO term at Interaction level, as membranes were extracted after X-linking.; I have altered nucl. transfection to child term, as before, and added over-expressed to both participants.; altered; new interaction added - so one for each form.; Exp-Mod added. I have altered Exp-prep as before. Also added a Sufficient to bind in Fuzzy type, as they do not specify the exact sequence for this construct - if they used the ICOSL-V construct as before, presumably they would have stated this fact. (I had already added a Comment) icos_human Inducible T-cell costimulator ICOS AILIM Activation-inducible lymphocyte immunomediatory molecule protein protein human Homo sapiens Human MKSGLWYFFLFCLRIKVLTGEINGSANYEMFIFHNGGVQILCKYPDIVQQFKMQLLKGGQILCDLTKTKGSGNTVSIKSLKFCHSQLSNNSVSFFLYNLDHSHANYYFCNLSIFDPPPFKVTLTGGYLHIYESQLCCQLKFWLPIGCAAFVVVCILGCILICWLTKKKYSSSVHDPNGEYMFMRAVNTAKKSRLTDVTL 214EC741C9BDC9FC icosl_human ICOS ligand ICOSLG B7 homolog 2 B7-like protein Gl50 B7-related protein 1 B7H2 B7RP1 ICOSL KIAA0653 protein protein human Homo sapiens Human MRLGSPGLLFLLFSSLRADTQEKEVRAMVGSDVELSCACPEGSRFDLNDVYVYWQTSESKTVVTYHIPQNSSLENVDSRYRNRALMSPAGMLRGDFSLRLFNVTPQDEQKFHCLVLSQSLGFQEVLSVEVTLHVAANFSVPVVSAPHSPSQDELTFTCTSINGYPRPNVYWINKTDNSLLDQALQNDTVFLNMRGLYDVVSVLRIARTPSVNIGCCIENVLLQQNLTVGSQTGNDIGERDKITENPVSTGEKNAATWSILAVLCLLVVVAVAIGWVCRDRCLQHSYAGAWAVSPETELTGHV 647934E21B55E34A icoslg-icoslg-1 6 8 unspecified role unspecified role neutral component neutral component purified purified signal+ecd sufficient to bind sufficient binding region sufficient to bind certain certain sequence position certain certain certain sequence position certain human-293t Homo sapiens 293 cells transformed with SV40 large T antigen 293t 293 cells expressing SV40 large T antigen. The original designation of this cell line was 293tsA1609neo. It is not in Cabri,ATCC or HyperCLDB. 8 unspecified role unspecified role neutral component neutral component purified purified signal+ecd sufficient to bind sufficient binding region sufficient to bind certain certain sequence position certain certain certain sequence position certain human-293t Homo sapiens 293 cells transformed with SV40 large T antigen 293t 293 cells expressing SV40 large T antigen. The original designation of this cell line was 293tsA1609neo. It is not in Cabri,ATCC or HyperCLDB. direct interaction direct interaction figure 1B icoslg-1 1 8 unspecified role unspecified role neutral component neutral component purified purified signal+ecd sufficient to bind sufficient binding region sufficient to bind certain certain sequence position certain certain certain sequence position certain human-293t Homo sapiens 293 cells transformed with SV40 large T antigen 293t 293 cells expressing SV40 large T antigen. The original designation of this cell line was 293tsA1609neo. It is not in Cabri,ATCC or HyperCLDB. 8 unspecified role unspecified role neutral component neutral component purified purified signal+ecd sufficient to bind sufficient binding region sufficient to bind certain certain sequence position certain certain certain sequence position certain human-293t Homo sapiens 293 cells transformed with SV40 large T antigen 293t 293 cells expressing SV40 large T antigen. The original designation of this cell line was 293tsA1609neo. It is not in Cabri,ATCC or HyperCLDB. direct interaction direct interaction figure 1B (inset) icoslg-4 2 7 unspecified role unspecified role bait bait conditioned medium conditioned medium ecd sufficient to bind sufficient binding region sufficient to bind certain certain sequence position certain certain certain sequence position certain c_terminus fc-igg tag fc-igg tag c-term range C-terminal range c-term range c-term range C-terminal range c-term range B2M_signal tag tag undetermined undetermined sequence position undetermined undetermined undetermined sequence position undetermined human-293t Homo sapiens 293 cells transformed with SV40 large T antigen 293t 293 cells expressing SV40 large T antigen. The original designation of this cell line was 293tsA1609neo. It is not in Cabri,ATCC or HyperCLDB. 8 unspecified role unspecified role prey prey purified purified signal+ecd sufficient to bind sufficient binding region sufficient to bind certain certain sequence position certain certain certain sequence position certain human-293t Homo sapiens 293 cells transformed with SV40 large T antigen 293t 293 cells expressing SV40 large T antigen. The original designation of this cell line was 293tsA1609neo. It is not in Cabri,ATCC or HyperCLDB. physical association physical association figure 1C monomeric ICOSL Both the oligomeric and monomeric forms of ICOSL-VC bound to soluble ICOS-Ig icoslg-5 5 8 unspecified role unspecified role neutral component neutral component purified purified signal+ecd sufficient to bind sufficient binding region sufficient to bind certain certain sequence position certain certain certain sequence position certain human-293t Homo sapiens 293 cells transformed with SV40 large T antigen 293t 293 cells expressing SV40 large T antigen. The original designation of this cell line was 293tsA1609neo. It is not in Cabri,ATCC or HyperCLDB. direct interaction direct interaction figure 1D icos-icoslg-2 3 7 unspecified role unspecified role neutral component neutral component over-expressed over expressed level over-expressed n transfection treat nucleic acid transfection by treatment n transfection treat conditioned medium conditioned medium ecd sufficient to bind sufficient binding region sufficient to bind certain certain sequence position certain certain certain sequence position certain c_terminus fc-igg tag fc-igg tag c-term range C-terminal range c-term range c-term range C-terminal range c-term range B2M_signal tag tag undetermined undetermined sequence position undetermined undetermined undetermined sequence position undetermined human-293t Homo sapiens 293 cells transformed with SV40 large T antigen 293t 293 cells expressing SV40 large T antigen. The original designation of this cell line was 293tsA1609neo. It is not in Cabri,ATCC or HyperCLDB. 8 unspecified role unspecified role neutral component neutral component over-expressed over expressed level over-expressed n transfection treat nucleic acid transfection by treatment n transfection treat c_terminus yfp tag yellow fluorescent protein tag YFP yellow fluorescent protein yellow fluorescent protein YFP yfp tag c-term range C-terminal range c-term range c-term range C-terminal range c-term range O75144:p.Tyr53Ala mutation disrupting strength mutation disrupting interaction strength mutation disrupting strength certain certain sequence position certain certain certain sequence position certain Y A Wed May 02 15:11:41 BST 2018 This feature has been corrected as a result of our quality control procedures. The original label was 'tyr53ala' Tue Jun 30 20:01:23 BST 2020 This feature has been corrected as a result of our quality control procedures. The original label was 'p.Tyr53Ala' O75144:p.Val62Ala mutation decreasing strength mutation decreasing interaction strength mutation decreasing strength certain certain sequence position certain certain certain sequence position certain V A Wed May 02 19:56:36 BST 2018 This feature has been corrected as a result of our quality control procedures. The original label was 'val62ala' Tue Jun 30 16:41:43 BST 2020 This feature has been corrected as a result of our quality control procedures. The original label was 'p.Val62Ala' O75144:p.Tyr51Ala mutation disrupting strength mutation disrupting interaction strength mutation disrupting strength certain certain sequence position certain certain certain sequence position certain Y A Wed May 02 20:15:06 BST 2018 This feature has been corrected as a result of our quality control procedures. The original label was 'tyr51ala' Tue Jun 30 16:57:06 BST 2020 This feature has been corrected as a result of our quality control procedures. The original label was 'p.Tyr51Ala' crigr-cho chinese hamster ovary cell line cho chinese hamster ovary cell line Also called LR73 cells. physical association physical association figure 2A, 7A The presence of YFP- or CFP-tag at the C terminus of ICOSL did not affect its cell surface expression or binding to soluble ICOS-Ig icoslg-7 5 8 unspecified role unspecified role neutral component neutral component subcellular prep subcellular preparation subcellular prep over-expressed over expressed level over-expressed n transfection treat nucleic acid transfection by treatment n transfection treat c_terminus yfp tag yellow fluorescent protein tag YFP yellow fluorescent protein yellow fluorescent protein YFP yfp tag c-term range C-terminal range c-term range c-term range C-terminal range c-term range human-293t Homo sapiens 293 cells transformed with SV40 large T antigen 293t 293 cells expressing SV40 large T antigen. The original designation of this cell line was 293tsA1609neo. It is not in Cabri,ATCC or HyperCLDB. physical association physical association figure 2C Cross-linking shows presence of ICOSL oligomers of higher stoichiometry. Cross-linking shows presence of ICOSL oligomers of higher stoichiometry icoslg-6 4 8 unspecified role unspecified role neutral component neutral component over-expressed over expressed level over-expressed n transfection treat nucleic acid transfection by treatment n transfection treat deltaC_construct sufficient to bind sufficient binding region sufficient to bind range range range range c_terminus cfp tag cyan fluorescent protein tag CFP cyan fluorescent protein cyan fluorescent protein CFP cfp tag c-term range C-terminal range c-term range c-term range C-terminal range c-term range crigr-cho chinese hamster ovary cell line cho chinese hamster ovary cell line Also called LR73 cells. 8 unspecified role unspecified role neutral component neutral component over-expressed over expressed level over-expressed n transfection treat nucleic acid transfection by treatment n transfection treat deltaC_construct sufficient to bind sufficient binding region sufficient to bind range range range range c_terminus yfp tag yellow fluorescent protein tag YFP yellow fluorescent protein yellow fluorescent protein YFP yfp tag c-term range C-terminal range c-term range c-term range C-terminal range c-term range crigr-cho chinese hamster ovary cell line cho chinese hamster ovary cell line Also called LR73 cells. physical association physical association figure 2D-F,H.4 B/C The presence of ICOSL-YFP with a cysteine containing linker did not change the average FRET efficiency values of ICOSL. Fret efficiency was increased using an ICOSL construct missing the IgC domain, suggesting a higher degree of homo-association. icoslg-8 5 8 unspecified role unspecified role neutral component neutral component subcellular prep subcellular preparation subcellular prep over-expressed over expressed level over-expressed n transfection treat nucleic acid transfection by treatment n transfection treat c_terminus yfp tag yellow fluorescent protein tag YFP yellow fluorescent protein yellow fluorescent protein YFP yfp tag c-term range C-terminal range c-term range c-term range C-terminal range c-term range human-293t Homo sapiens 293 cells transformed with SV40 large T antigen 293t 293 cells expressing SV40 large T antigen. The original designation of this cell line was 293tsA1609neo. It is not in Cabri,ATCC or HyperCLDB. physical association physical association figure 2Ci Cross-linking shows presence of ICOSL oligomers of higher stoichiometry. Cross-linking shows presence of ICOSL oligomers of higher stoichiometry icoslg-9 5 8 unspecified role unspecified role neutral component neutral component subcellular prep subcellular preparation subcellular prep over-expressed over expressed level over-expressed n transfection treat nucleic acid transfection by treatment n transfection treat c_terminus yfp tag yellow fluorescent protein tag YFP yellow fluorescent protein yellow fluorescent protein YFP yfp tag c-term range C-terminal range c-term range c-term range C-terminal range c-term range human-293t Homo sapiens 293 cells transformed with SV40 large T antigen 293t 293 cells expressing SV40 large T antigen. The original designation of this cell line was 293tsA1609neo. It is not in Cabri,ATCC or HyperCLDB. physical association physical association figure 2Cii Cross-linking shows presence of ICOSL oligomers of higher stoichiometry. Cross-linking shows presence of ICOSL oligomers of higher stoichiometry icos-icoslg-1 3 7 unspecified role unspecified role neutral component neutral component over-expressed over expressed level over-expressed n transfection treat nucleic acid transfection by treatment n transfection treat c_terminus yfp tag yellow fluorescent protein tag YFP yellow fluorescent protein yellow fluorescent protein YFP yfp tag c-term range C-terminal range c-term range c-term range C-terminal range c-term range human-293t Homo sapiens 293 cells transformed with SV40 large T antigen 293t 293 cells expressing SV40 large T antigen. The original designation of this cell line was 293tsA1609neo. It is not in Cabri,ATCC or HyperCLDB. 8 unspecified role unspecified role neutral component neutral component over-expressed over expressed level over-expressed n transfection treat nucleic acid transfection by treatment n transfection treat conditioned medium conditioned medium c_terminus fc-igg tag fc-igg tag c-term range C-terminal range c-term range c-term range C-terminal range c-term range IgV_domain sufficient to bind sufficient binding region sufficient to bind certain certain sequence position certain certain certain sequence position certain human-293t Homo sapiens 293 cells transformed with SV40 large T antigen 293t 293 cells expressing SV40 large T antigen. The original designation of this cell line was 293tsA1609neo. It is not in Cabri,ATCC or HyperCLDB. physical association physical association figure 3 B/C, 4A Both the recombinant proteins, ICOSL-VC-Ig and ICOSL-V-Ig, exhibited similar equilibrium binding to ICOS, indicating that the IgV domain of ICOSL alone could bind to ICOS. The IgV domain of ICOSL alone could bind to ICOS, however ICOSL-V-Ig was found to bind at least 8- to 10-fold less well to ICOS compared with ICOSL-VC-Ig. icos-icoslg-3 3 7 unspecified role unspecified role neutral component neutral component over-expressed over expressed level over-expressed n transfection treat nucleic acid transfection by treatment n transfection treat conditioned medium conditioned medium ecd sufficient to bind sufficient binding region sufficient to bind certain certain sequence position certain certain certain sequence position certain c_terminus fc-igg tag fc-igg tag c-term range C-terminal range c-term range c-term range C-terminal range c-term range B2M_signal tag tag undetermined undetermined sequence position undetermined undetermined undetermined sequence position undetermined human-293t Homo sapiens 293 cells transformed with SV40 large T antigen 293t 293 cells expressing SV40 large T antigen. The original designation of this cell line was 293tsA1609neo. It is not in Cabri,ATCC or HyperCLDB. 8 unspecified role unspecified role neutral component neutral component over-expressed over expressed level over-expressed n transfection treat nucleic acid transfection by treatment n transfection treat c_terminus yfp tag yellow fluorescent protein tag YFP yellow fluorescent protein yellow fluorescent protein YFP yfp tag c-term range C-terminal range c-term range c-term range C-terminal range c-term range O75144:p.Tyr53Ala mutation disrupting strength mutation disrupting interaction strength mutation disrupting strength certain certain sequence position certain certain certain sequence position certain Y A Wed May 02 18:17:19 BST 2018 This feature has been corrected as a result of our quality control procedures. The original label was 'tyr53ala' Tue Jun 30 17:42:36 BST 2020 This feature has been corrected as a result of our quality control procedures. The original label was 'p.Tyr53Ala' O75144:p.Gln55Ala mutation decreasing strength mutation decreasing interaction strength mutation decreasing strength certain certain sequence position certain certain certain sequence position certain Q A Wed May 02 20:06:21 BST 2018 This feature has been corrected as a result of our quality control procedures. The original label was 'gln55ala' Tue Jun 30 17:59:28 BST 2020 This feature has been corrected as a result of our quality control procedures. The original label was 'p.Gln55Ala' O75144:p.Ser59Ala mutation with no effect mutation with no effect mutation with no effect mutation not affecting interaction certain certain sequence position certain certain certain sequence position certain S A Wed May 02 20:02:41 BST 2018 This feature has been corrected as a result of our quality control procedures. The original label was 'ser59ala' Tue Jun 30 19:59:41 BST 2020 This feature has been corrected as a result of our quality control procedures. The original label was 'p.Ser59Ala' O75144:p.Thr61Ala mutation with no effect mutation with no effect mutation with no effect mutation not affecting interaction certain certain sequence position certain certain certain sequence position certain T A Wed May 02 16:58:40 BST 2018 This feature has been corrected as a result of our quality control procedures. The original label was 'thr61ala' Tue Jun 30 17:37:58 BST 2020 This feature has been corrected as a result of our quality control procedures. The original label was 'p.Thr61Ala' O75144:p.Val62Ala mutation decreasing strength mutation decreasing interaction strength mutation decreasing strength certain certain sequence position certain certain certain sequence position certain V A Wed May 02 15:12:59 BST 2018 This feature has been corrected as a result of our quality control procedures. The original label was 'val62ala' Tue Jun 30 17:45:42 BST 2020 This feature has been corrected as a result of our quality control procedures. The original label was 'p.Val62Ala' O75144:p.[Gln55Ala;Val62Ala] mutation disrupting strength mutation disrupting interaction strength mutation disrupting strength certain certain sequence position certain certain certain sequence position certain Q A certain certain sequence position certain certain certain sequence position certain V A Wed May 02 16:32:33 BST 2018 This feature has been corrected as a result of our quality control procedures. The original label was 'gln55ala,val62ala' Tue Jun 30 15:41:15 BST 2020 This feature has been corrected as a result of our quality control procedures. The original label was 'p.[Gln55Ala;Val62Ala]' O75144:p.Thr64Ala mutation with no effect mutation with no effect mutation with no effect mutation not affecting interaction certain certain sequence position certain certain certain sequence position certain T A Wed May 02 19:13:21 BST 2018 This feature has been corrected as a result of our quality control procedures. The original label was 'thr64ala' Tue Jun 30 16:37:30 BST 2020 This feature has been corrected as a result of our quality control procedures. The original label was 'p.Thr64Ala' O75144:p.His66Ala mutation decreasing strength mutation decreasing interaction strength mutation decreasing strength certain certain sequence position certain certain certain sequence position certain H A Wed May 02 15:39:28 BST 2018 This feature has been corrected as a result of our quality control procedures. The original label was 'his66ala' Tue Jun 30 18:48:42 BST 2020 This feature has been corrected as a result of our quality control procedures. The original label was 'p.His66Ala' O75144:p.Val76Ala mutation decreasing strength mutation decreasing interaction strength mutation decreasing strength certain certain sequence position certain certain certain sequence position certain V A Wed May 02 17:01:32 BST 2018 This feature has been corrected as a result of our quality control procedures. The original label was 'val76ala' Tue Jun 30 16:27:03 BST 2020 This feature has been corrected as a result of our quality control procedures. The original label was 'p.Val76Ala' O75144:p.Asp77Ala mutation with no effect mutation with no effect mutation with no effect mutation not affecting interaction certain certain sequence position certain certain certain sequence position certain D A Wed May 02 17:47:28 BST 2018 This feature has been corrected as a result of our quality control procedures. The original label was 'asp77ala' Tue Jun 30 19:51:57 BST 2020 This feature has been corrected as a result of our quality control procedures. The original label was 'p.Asp77Ala' O75144:p.Tyr80Ala mutation disrupting strength mutation disrupting interaction strength mutation disrupting strength certain certain sequence position certain certain certain sequence position certain Y A Wed May 02 15:12:33 BST 2018 This feature has been corrected as a result of our quality control procedures. The original label was 'tyr80ala' Tue Jun 30 20:06:37 BST 2020 This feature has been corrected as a result of our quality control procedures. The original label was 'p.Tyr80Ala' O75144:p.Ser87Ala mutation with no effect mutation with no effect mutation with no effect mutation not affecting interaction certain certain sequence position certain certain certain sequence position certain S A Wed May 02 16:37:34 BST 2018 This feature has been corrected as a result of our quality control procedures. The original label was 'ser87ala' Tue Jun 30 19:29:01 BST 2020 This feature has been corrected as a result of our quality control procedures. The original label was 'p.Ser87Ala' O75144:p.Leu47Ala mutation with no effect mutation with no effect mutation with no effect mutation not affecting interaction certain certain sequence position certain certain certain sequence position certain L A Wed May 02 15:07:22 BST 2018 This feature has been corrected as a result of our quality control procedures. The original label was 'leu47ala' Tue Jun 30 19:04:06 BST 2020 This feature has been corrected as a result of our quality control procedures. The original label was 'p.Leu47Ala' O75144:p.Tyr51Ala mutation disrupting strength mutation disrupting interaction strength mutation disrupting strength certain certain sequence position certain certain certain sequence position certain Y A Wed May 02 20:16:03 BST 2018 This feature has been corrected as a result of our quality control procedures. The original label was 'tyr51ala' Tue Jun 30 17:48:41 BST 2020 This feature has been corrected as a result of our quality control procedures. The original label was 'p.Tyr51Ala' O75144:p.Tyr53Phe mutation disrupting strength mutation disrupting interaction strength mutation disrupting strength certain certain sequence position certain certain certain sequence position certain Y F Wed May 02 15:52:27 BST 2018 This feature has been corrected as a result of our quality control procedures. The original label was 'tyr53phe' Tue Jun 30 16:21:46 BST 2020 This feature has been corrected as a result of our quality control procedures. The original label was 'p.Tyr53Phe' O75144:p.Tyr51Phe mutation decreasing strength mutation decreasing interaction strength mutation decreasing strength certain certain sequence position certain certain certain sequence position certain Y F Wed May 02 16:54:37 BST 2018 This feature has been corrected as a result of our quality control procedures. The original label was 'tyr51phe' Tue Jun 30 18:18:37 BST 2020 This feature has been corrected as a result of our quality control procedures. The original label was 'p.Tyr51Phe' O75144:p.Leu98Ala mutation with no effect mutation with no effect mutation with no effect mutation not affecting interaction certain certain sequence position certain certain certain sequence position certain L A Wed May 02 15:31:31 BST 2018 This feature has been corrected as a result of our quality control procedures. The original label was 'leu98ala' Tue Jun 30 17:26:49 BST 2020 This feature has been corrected as a result of our quality control procedures. The original label was 'p.Leu98Ala' O75144:p.Leu114Ala mutation decreasing strength mutation decreasing interaction strength mutation decreasing strength certain certain sequence position certain certain certain sequence position certain L A Wed May 02 19:53:11 BST 2018 This feature has been corrected as a result of our quality control procedures. The original label was 'leu114ala' Tue Jun 30 17:59:33 BST 2020 This feature has been corrected as a result of our quality control procedures. The original label was 'p.Leu114Ala' O75144:p.Leu116Ala mutation disrupting rate mutation disrupting interaction rate mutation disrupting rate certain certain sequence position certain certain certain sequence position certain L A Wed May 02 19:07:47 BST 2018 This feature has been corrected as a result of our quality control procedures. The original label was 'leu116ala' Tue Jun 30 17:02:03 BST 2020 This feature has been corrected as a result of our quality control procedures. The original label was 'p.Leu116Ala' O75144:p.Leu120Ala mutation decreasing strength mutation decreasing interaction strength mutation decreasing strength certain certain sequence position certain certain certain sequence position certain L A Wed May 02 17:36:53 BST 2018 This feature has been corrected as a result of our quality control procedures. The original label was 'leu120ala' Tue Jun 30 17:35:57 BST 2020 This feature has been corrected as a result of our quality control procedures. The original label was 'p.Leu120Ala' O75144:p.Gly121Ala mutation with no effect mutation with no effect mutation with no effect mutation not affecting interaction certain certain sequence position certain certain certain sequence position certain G A Wed May 02 20:10:49 BST 2018 This feature has been corrected as a result of our quality control procedures. The original label was 'gly121ala' Tue Jun 30 18:24:46 BST 2020 This feature has been corrected as a result of our quality control procedures. The original label was 'p.Gly121Ala' O75144:p.Phe122Ala mutation disrupting strength mutation disrupting interaction strength mutation disrupting strength certain certain sequence position certain certain certain sequence position certain F A Wed May 02 15:58:19 BST 2018 This feature has been corrected as a result of our quality control procedures. The original label was 'phe122ala' Tue Jun 30 17:33:03 BST 2020 This feature has been corrected as a result of our quality control procedures. The original label was 'p.Phe122Ala' O75144:p.Gln123Ala mutation with no effect mutation with no effect mutation with no effect mutation not affecting interaction certain certain sequence position certain certain certain sequence position certain Q A Wed May 02 20:26:12 BST 2018 This feature has been corrected as a result of our quality control procedures. The original label was 'gln123ala' Tue Jun 30 17:29:13 BST 2020 This feature has been corrected as a result of our quality control procedures. The original label was 'p.Gln123Ala' O75144:p.Glu124Ala mutation decreasing strength mutation decreasing interaction strength mutation decreasing strength certain certain sequence position certain certain certain sequence position certain E A Wed May 02 17:12:11 BST 2018 This feature has been corrected as a result of our quality control procedures. The original label was 'glu124ala' Tue Jun 30 17:07:22 BST 2020 This feature has been corrected as a result of our quality control procedures. The original label was 'p.Glu124Ala' O75144:p.Phe138Ala mutation with no effect mutation with no effect mutation with no effect mutation not affecting interaction certain certain sequence position certain certain certain sequence position certain F A Wed May 02 18:34:56 BST 2018 This feature has been corrected as a result of our quality control procedures. The original label was 'phe138ala' Tue Jun 30 17:24:06 BST 2020 This feature has been corrected as a result of our quality control procedures. The original label was 'p.Phe138Ala' O75144:p.Pro141Ala mutation with no effect mutation with no effect mutation with no effect mutation not affecting interaction certain certain sequence position certain certain certain sequence position certain P A Wed May 02 16:26:28 BST 2018 This feature has been corrected as a result of our quality control procedures. The original label was 'pro141ala' Tue Jun 30 18:04:50 BST 2020 This feature has been corrected as a result of our quality control procedures. The original label was 'p.Pro141Ala' O75144:p.Val143Ala mutation with no effect mutation with no effect mutation with no effect mutation not affecting interaction certain certain sequence position certain certain certain sequence position certain V A Wed May 02 16:27:56 BST 2018 This feature has been corrected as a result of our quality control procedures. The original label was 'val143ala' Tue Jun 30 19:32:49 BST 2020 This feature has been corrected as a result of our quality control procedures. The original label was 'p.Val143Ala' O75144:p.Gly195Ala mutation with no effect mutation with no effect mutation with no effect mutation not affecting interaction certain certain sequence position certain certain certain sequence position certain G A Wed May 02 19:15:32 BST 2018 This feature has been corrected as a result of our quality control procedures. The original label was 'gly195ala' Tue Jun 30 16:22:40 BST 2020 This feature has been corrected as a result of our quality control procedures. The original label was 'p.Gly195Ala' O75144:p.Leu196Ala mutation with no effect mutation with no effect mutation with no effect mutation not affecting interaction certain certain sequence position certain certain certain sequence position certain L A Wed May 02 14:31:03 BST 2018 This feature has been corrected as a result of our quality control procedures. The original label was 'leu196ala' Tue Jun 30 19:09:33 BST 2020 This feature has been corrected as a result of our quality control procedures. The original label was 'p.Leu196Ala' O75144:p.Asn70Gln mutation with no effect mutation with no effect mutation with no effect mutation not affecting interaction certain certain sequence position certain certain certain sequence position certain N Q Wed May 02 15:33:18 BST 2018 This feature has been corrected as a result of our quality control procedures. The original label was 'asn70gln' Tue Jun 30 18:40:53 BST 2020 This feature has been corrected as a result of our quality control procedures. The original label was 'p.Asn70Gln' O75144:p.Asn102Gln mutation decreasing strength mutation decreasing interaction strength mutation decreasing strength certain certain sequence position certain certain certain sequence position certain N Q Wed May 02 16:37:42 BST 2018 This feature has been corrected as a result of our quality control procedures. The original label was 'asn102gln' Tue Jun 30 17:28:35 BST 2020 This feature has been corrected as a result of our quality control procedures. The original label was 'p.Asn102Gln' O75144:p.Asn137Gln mutation with no effect mutation with no effect mutation with no effect mutation not affecting interaction certain certain sequence position certain certain certain sequence position certain N Q Wed May 02 16:08:03 BST 2018 This feature has been corrected as a result of our quality control procedures. The original label was 'asn137gln' Tue Jun 30 17:55:55 BST 2020 This feature has been corrected as a result of our quality control procedures. The original label was 'p.Asn137Gln' O75144:p.Asn186Gln mutation with no effect mutation with no effect mutation with no effect mutation not affecting interaction certain certain sequence position certain certain certain sequence position certain N Q Wed May 02 15:21:05 BST 2018 This feature has been corrected as a result of our quality control procedures. The original label was 'asn186gln' Tue Jun 30 15:42:30 BST 2020 This feature has been corrected as a result of our quality control procedures. The original label was 'p.Asn186Gln' O75144:p.Asn225Gln mutation with no effect mutation with no effect mutation with no effect mutation not affecting interaction certain certain sequence position certain certain certain sequence position certain N Q Wed May 02 15:37:59 BST 2018 This feature has been corrected as a result of our quality control procedures. The original label was 'asn225gln' Tue Jun 30 19:43:14 BST 2020 This feature has been corrected as a result of our quality control procedures. The original label was 'p.Asn225Gln' human-293t Homo sapiens 293 cells transformed with SV40 large T antigen 293t 293 cells expressing SV40 large T antigen. The original designation of this cell line was 293tsA1609neo. It is not in Cabri,ATCC or HyperCLDB. physical association physical association figure 6A, table 1 Disrupting mutations show equal or less than 5% binding. I think the nos of mutations are correct - no expression for N173Q mutant. icos-icoslg-4 2 8 unspecified role unspecified role prey prey purified purified signal+ecd sufficient to bind sufficient binding region sufficient to bind certain certain sequence position certain certain certain sequence position certain human-293t Homo sapiens 293 cells transformed with SV40 large T antigen 293t 293 cells expressing SV40 large T antigen. The original designation of this cell line was 293tsA1609neo. It is not in Cabri,ATCC or HyperCLDB. 7 unspecified role unspecified role bait bait conditioned medium conditioned medium ecd sufficient to bind sufficient binding region sufficient to bind certain certain sequence position certain certain certain sequence position certain c_terminus fc-igg tag fc-igg tag c-term range C-terminal range c-term range c-term range C-terminal range c-term range B2M_signal tag tag undetermined undetermined sequence position undetermined undetermined undetermined sequence position undetermined human-293t Homo sapiens 293 cells transformed with SV40 large T antigen 293t 293 cells expressing SV40 large T antigen. The original designation of this cell line was 293tsA1609neo. It is not in Cabri,ATCC or HyperCLDB. 8 unspecified role unspecified role prey prey purified purified signal+ecd sufficient to bind sufficient binding region sufficient to bind certain certain sequence position certain certain certain sequence position certain human-293t Homo sapiens 293 cells transformed with SV40 large T antigen 293t 293 cells expressing SV40 large T antigen. The original designation of this cell line was 293tsA1609neo. It is not in Cabri,ATCC or HyperCLDB. 104 110 physical association physical association figure 1Ci oligomeric ICOSL Both the oligomeric and monomeric forms of ICOSL-VC bound to soluble ICOS-Ig. Both the oligomeric and monomeric forms of ICOSL-VC bound to soluble ICOS-Ig