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/ TGF-β-mediated Foxp3 gene expression is cooperatively regulated by Stat5, Creb, and AP-1 through CNS2. TGF-β-mediated Foxp3 gene expression is cooperatively regulated by Stat5, Creb, and AP-1 through CNS2. Journal of immunology (Baltimore, Md. : 1950) 2014 imex curation Ogawa C., Tone Y., Tsuda M., Peter C., Waldmann H., Tone M., Ogawa C., Tone Y., Tsuda M., Peter C., Waldmann H., Tone M., Ogawa C., Tone Y., Tsuda M., Peter C., Waldmann H., Tone M., Ogawa C., Tone Y., Tsuda M., Peter C., Waldmann H., Tone M. IBD - Inflammatory bowel disease email sent asking species for FALG-cFOS in figure 4C -Pull down -No Reply 14/9/15. as far as I can make out all the reporter expts are in EL4 cells, not (ordinary) T cells; so cannot add most possible causality statements EMSA Supershift experiment results have not been included due to lack of STAT5 antibody specificity. Masahide.Tone@csmc.edu 20-Dec-2016: Contacted by IntAct-Help. Only protein-protein interactions imex curation in vitro In vitro pull down pull down western blot western blot Immuno blot Immuno blot Ogawa C., Tone Y., Tsuda M., Peter C., Waldmann H., Tone M., Ogawa C., Tone Y., Tsuda M., Peter C., Waldmann H., Tone M., Ogawa C., Tone Y., Tsuda M., Peter C., Waldmann H., Tone M., Ogawa C., Tone Y., Tsuda M., Peter C., Waldmann H., Tone M. IBD - Inflammatory bowel disease email sent asking species for FALG-cFOS in figure 4C -Pull down -No Reply 14/9/15. as far as I can make out all the reporter expts are in EL4 cells, not (ordinary) T cells; so cannot add most possible causality statements EMSA Supershift experiment results have not been included due to lack of STAT5 antibody specificity. Masahide.Tone@csmc.edu Accepted 2016-NOV-01 AT 15:22 GMT AT 15:22 GMT by PPORRAS Journal of immunology (Baltimore, Md. : 1950) 2014 Only protein-protein interactions imex curation imex curation I have altered the Exp-mod slightly;however at Interaction level "Variable Expt. Conditions:1" is still visible - bug. Altered the H4-Ac antibody results to include "possible" and "observed ptm".; I dont know what you mean by "Variable condition for AcHA is missing". At Interaction level "Variable Expt. Conditions:1" is still visible - bug. Altered the H4-Ac antibody results to include "possible" and "observed ptm". EBI-11171087:- I have added a new causality statement "CHEBI:87103 decreases the expression of ENSMUSG00000048756 at protein level.". I have cloned interactions from fig 4(F) and 1A into a new Expt for Nat Treg cells, and altered THIS expt so it is for iTREg cells only. TGF-β-mediated Foxp3 gene expression is cooperatively regulated by Stat5, Creb, and AP-1 through CNS2. TGF-β-mediated Foxp3 gene expression is cooperatively regulated by Stat5, Creb, and AP-1 through CNS2. Journal of immunology (Baltimore, Md. : 1950) 2014 imex curation Ogawa C., Tone Y., Tsuda M., Peter C., Waldmann H., Tone M., Ogawa C., Tone Y., Tsuda M., Peter C., Waldmann H., Tone M., Ogawa C., Tone Y., Tsuda M., Peter C., Waldmann H., Tone M., Ogawa C., Tone Y., Tsuda M., Peter C., Waldmann H., Tone M. IBD - Inflammatory bowel disease email sent asking species for FALG-cFOS in figure 4C -Pull down -No Reply 14/9/15. as far as I can make out all the reporter expts are in EL4 cells, not (ordinary) T cells; so cannot add most possible causality statements EMSA Supershift experiment results have not been included due to lack of STAT5 antibody specificity. Masahide.Tone@csmc.edu 20-Dec-2016: Contacted by IntAct-Help. Only protein-protein interactions imex curation mouse-t-cell Mouse T cell t-cell Peripheral blood T-lymphocytes ch-ip chromatin immunoprecipitation assay ch-ip q-pcr quantitative pcr q-pcr Ogawa C., Tone Y., Tsuda M., Peter C., Waldmann H., Tone M., Ogawa C., Tone Y., Tsuda M., Peter C., Waldmann H., Tone M., Ogawa C., Tone Y., Tsuda M., Peter C., Waldmann H., Tone M., Ogawa C., Tone Y., Tsuda M., Peter C., Waldmann H., Tone M. IBD - Inflammatory bowel disease Results from ChIps using STAT5 antibody have not been included due to lack of antibody specificity. email sent asking species for FALG-cFOS in figure 4C -Pull down -No Reply 14/9/15. as far as I can make out all the reporter expts are in EL4 cells, not (ordinary) T cells; so cannot add most possible causality statements iTreg (TGF-beta induced TReg cells) EMSA Supershift experiment results have not been included due to lack of STAT5 antibody specificity. Masahide.Tone@csmc.edu Accepted 2016-NOV-01 AT 15:22 GMT AT 15:22 GMT by PPORRAS Journal of immunology (Baltimore, Md. : 1950) 2014 Only protein-protein interactions imex curation imex curation I have altered the Exp-mod slightly;however at Interaction level "Variable Expt. Conditions:1" is still visible - bug. Altered the H4-Ac antibody results to include "possible" and "observed ptm".; I dont know what you mean by "Variable condition for AcHA is missing". At Interaction level "Variable Expt. Conditions:1" is still visible - bug. Altered the H4-Ac antibody results to include "possible" and "observed ptm". EBI-11171087:- I have added a new causality statement "CHEBI:87103 decreases the expression of ENSMUSG00000048756 at protein level.". I have cloned interactions from fig 4(F) and 1A into a new Expt for Nat Treg cells, and altered THIS expt so it is for iTREg cells only. TGF-β-mediated Foxp3 gene expression is cooperatively regulated by Stat5, Creb, and AP-1 through CNS2. TGF-β-mediated Foxp3 gene expression is cooperatively regulated by Stat5, Creb, and AP-1 through CNS2. Journal of immunology (Baltimore, Md. : 1950) 2014 imex curation Ogawa C., Tone Y., Tsuda M., Peter C., Waldmann H., Tone M., Ogawa C., Tone Y., Tsuda M., Peter C., Waldmann H., Tone M., Ogawa C., Tone Y., Tsuda M., Peter C., Waldmann H., Tone M., Ogawa C., Tone Y., Tsuda M., Peter C., Waldmann H., Tone M. IBD - Inflammatory bowel disease email sent asking species for FALG-cFOS in figure 4C -Pull down -No Reply 14/9/15. as far as I can make out all the reporter expts are in EL4 cells, not (ordinary) T cells; so cannot add most possible causality statements EMSA Supershift experiment results have not been included due to lack of STAT5 antibody specificity. Masahide.Tone@csmc.edu 20-Dec-2016: Contacted by IntAct-Help. Only protein-protein interactions imex curation mouse-t-cell Mouse T cell t-cell Peripheral blood T-lymphocytes ch-ip chromatin immunoprecipitation assay ch-ip q-pcr quantitative pcr q-pcr Ogawa C., Tone Y., Tsuda M., Peter C., Waldmann H., Tone M., Ogawa C., Tone Y., Tsuda M., Peter C., Waldmann H., Tone M., Ogawa C., Tone Y., Tsuda M., Peter C., Waldmann H., Tone M., Ogawa C., Tone Y., Tsuda M., Peter C., Waldmann H., Tone M. IBD - Inflammatory bowel disease email sent asking species for FALG-cFOS in figure 4C -Pull down -No Reply 14/9/15. as far as I can make out all the reporter expts are in EL4 cells, not (ordinary) T cells; so cannot add most possible causality statements nTreg (natural TReg cells). EMSA Supershift experiment results have not been included due to lack of STAT5 antibody specificity. Masahide.Tone@csmc.edu Accepted 2016-NOV-01 AT 15:22 GMT AT 15:22 GMT by PPORRAS Journal of immunology (Baltimore, Md. : 1950) 2014 Only protein-protein interactions imex curation imex curation I have altered the Exp-mod slightly;however at Interaction level "Variable Expt. Conditions:1" is still visible - bug. Altered the H4-Ac antibody results to include "possible" and "observed ptm".; I dont know what you mean by "Variable condition for AcHA is missing". At Interaction level "Variable Expt. Conditions:1" is still visible - bug. Altered the H4-Ac antibody results to include "possible" and "observed ptm". EBI-11171087:- I have added a new causality statement "CHEBI:87103 decreases the expression of ENSMUSG00000048756 at protein level.". I have cloned interactions from fig 4(F) and 1A into a new Expt for Nat Treg cells, and altered THIS expt so it is for iTREg cells only. TGF-β-mediated Foxp3 gene expression is cooperatively regulated by Stat5, Creb, and AP-1 through CNS2. TGF-β-mediated Foxp3 gene expression is cooperatively regulated by Stat5, Creb, and AP-1 through CNS2. Journal of immunology (Baltimore, Md. : 1950) 2014 imex curation Ogawa C., Tone Y., Tsuda M., Peter C., Waldmann H., Tone M., Ogawa C., Tone Y., Tsuda M., Peter C., Waldmann H., Tone M., Ogawa C., Tone Y., Tsuda M., Peter C., Waldmann H., Tone M., Ogawa C., Tone Y., Tsuda M., Peter C., Waldmann H., Tone M. IBD - Inflammatory bowel disease email sent asking species for FALG-cFOS in figure 4C -Pull down -No Reply 14/9/15. as far as I can make out all the reporter expts are in EL4 cells, not (ordinary) T cells; so cannot add most possible causality statements EMSA Supershift experiment results have not been included due to lack of STAT5 antibody specificity. Masahide.Tone@csmc.edu 20-Dec-2016: Contacted by IntAct-Help. Only protein-protein interactions imex curation mouse-t-cell Mouse T cell t-cell Peripheral blood T-lymphocytes ch-ip chromatin immunoprecipitation assay ch-ip q-pcr quantitative pcr q-pcr Ogawa C., Tone Y., Tsuda M., Peter C., Waldmann H., Tone M., Ogawa C., Tone Y., Tsuda M., Peter C., Waldmann H., Tone M., Ogawa C., Tone Y., Tsuda M., Peter C., Waldmann H., Tone M., Ogawa C., Tone Y., Tsuda M., Peter C., Waldmann H., Tone M. IBD - Inflammatory bowel disease Results from ChIps using STAT5 antibody have not been included due to lack of antibody specificity. email sent asking species for FALG-cFOS in figure 4C -Pull down -No Reply 14/9/15. as far as I can make out all the reporter expts are in EL4 cells, not (ordinary) T cells; so cannot add most possible causality statements CD4+CD25− T cells were cultured with TGF-beta + anti-CD3 + anti-CD28 and IL-2 for up to 72 hours. EMSA Supershift experiment results have not been included due to lack of STAT5 antibody specificity. Masahide.Tone@csmc.edu Accepted 2016-NOV-01 AT 15:22 GMT AT 15:22 GMT by PPORRAS Journal of immunology (Baltimore, Md. : 1950) 2014 Only protein-protein interactions imex curation imex curation I have altered the Exp-mod slightly;however at Interaction level "Variable Expt. Conditions:1" is still visible - bug. Altered the H4-Ac antibody results to include "possible" and "observed ptm".; I dont know what you mean by "Variable condition for AcHA is missing". At Interaction level "Variable Expt. Conditions:1" is still visible - bug. Altered the H4-Ac antibody results to include "possible" and "observed ptm". EBI-11171087:- I have added a new causality statement "CHEBI:87103 decreases the expression of ENSMUSG00000048756 at protein level.". I have cloned interactions from fig 4(F) and 1A into a new Expt for Nat Treg cells, and altered THIS expt so it is for iTREg cells only. creb1_mouse Cyclic AMP-responsive element-binding protein 1 Creb1 Creb-1 protein protein mouse Mus musculus Mouse MTMESGADNQQSGDAAVTEAENQQMTVQAQPQIATLAQVSMPAAHATSSAPTVTLVQLPNGQTVQVHGVIQAAQPSVIQSPQVQTVQISTIAESEDSQESVDSVTDSQKRREILSRRPSYRKILNDLSSDAPGVPRIEEEKSEEETSAPAITTVTVPTPIYQTSSGQYIAITQGGAIQLANNGTDGVQGLQTLTMTNAAATQPGTTILQYAQTTDGQQILVPSNQVVVQAASGDVQTYQIRTAPTSTIAPGVVMASSPALPTQPAEEAARKREVRLMKNREAARECRRKKKEYVKCLENRVAVLENQNKTLIEELKALKDLYCHKSD 00D1F26B00D48442 foxo3_mouse_gene Forkhead box protein O3 Foxo3 Fkhr2 Foxo3a gene gene mouse Mus musculus Mouse h4_mouse Histone H4 H4c11 H4c12 H4c14 H4c2 H4c3 H4c4 H4c6 H4c8 H4c9 H4f16 Hist1h4a Hist4h4 Hist1h4b Hist1h4c Hist1h4d Hist1h4f Hist1h4h Hist1h4i Hist1h4j Hist1h4k Hist2h4a Hist2h4 Hist1h4m H4-53 H4-12 H4c1 protein protein mouse Mus musculus Mouse MSGRGKGGKGLGKGGAKRHRKVLRDNIQGITKPAIRRLARRGGVKRISGLIYEETRGVLKVFLENVIRDAVTYTEHAKRKTVTAMDVVYALKRQGRTLYGFGG DIP protein P62806 original sequence version: 108 A9E5DFD3F8B97598 fos_mouse Proto-oncogene c-Fos Fos Cellular oncogene fos protein protein mouse Mus musculus Mouse MMFSGFNADYEASSSRCSSASPAGDSLSYYHSPADSFSSMGSPVNTQDFCADLSVSSANFIPTVTAISTSPDLQWLVQPTLVSSVAPSQTRAPHPYGLPTQSAGAYARAGMVKTVSGGRAQSIGRRGKVEQLSPEEEEKRRIRRERNKMAAAKCRNRRRELTDTLQAETDQLEDEKSALQTEIANLLKEKEKLEFILAAHRPACKIPDDLGFPEEMSVASLDLTGGLPEASTPESEEAFTLPLLNDPEPKPSLEPVKSISNVELKAEPFDDFLFPASSRPSGSETSRSVPDVDLSGSFYAADWEPLHSNSLGMGPMVTELEPLCTPVVTCTPGCTTYTSSFVFTYPEADSFPSCAAAHRKGSSSNEPSSDSLSSPTLLAL DIP protein P01101 original sequence version: 124 DIP protein P01101 original sequence version: 117 475966265952B624 wt_cns2_enhancer_probe ds dna double stranded deoxyribonucleic acid ds DNA ds dna mouse Mus musculus Mouse CCAGATGGACGTCACCTACCACATCCGCTAGC jun_mouse Transcription factor AP-1 Activator protein 1 Proto-oncogene c-Jun V-jun avian sarcoma virus 17 oncogene homolog AH119 Jun protein protein mouse Mus musculus Mouse MTAKMETTFYDDALNASFLQSESGAYGYSNPKILKQSMTLNLADPVGSLKPHLRAKNSDLLTSPDVGLLKLASPELERLIIQSSNGHITTTPTPTQFLCPKNVTDEQEGFAEGFVRALAELHSQNTLPSVTSAAQPVSGAGMVAPAVASVAGAGGGGGYSASLHSEPPVYANLSNFNPGALSSGGGAPSYGAAGLAFPSQPQQQQQPPQPPHHLPQQIPVQHPRLQALKEEPQTVPEMPGETPPLSPIDMESQERIKAERKRMRNRIAASKCRKRKLERIARLEEKVKTLKAQNSELASTANMLREQVAQLKQKVMNHVNSGCQLMLTQQLQTF DIP protein P05627 original sequence version: 134 2BE0E4025B8B1CA3 hist1h4h-foxo3-2 2 6 unspecified role unspecified role prey prey cns2_enhancer_region sufficient to bind sufficient binding region sufficient to bind undetermined undetermined sequence position undetermined undetermined undetermined sequence position undetermined 7 unspecified role unspecified role bait bait possible_acres ac1res monoacetylated residue Acetyl Acetylation (N terminus, N epsilon of Lysine, O of Serine) (Ac) Acetylation Ac1Res undetermined undetermined sequence position undetermined undetermined undetermined sequence position undetermined observed-ptm observed-ptm observed ptm observed-ptm association association 1A, 8 The 5' half of the CNS2 were highly acetylated in nTreg and TGF-beta activated iTReg cells, but not in CD3/28 activated cells. CHEBI:87662 -Stat5 inhibitor CHEBI:87103 JAK inhibitor CHEBI:87662 decreases the binding of P62806 to ENSMUSG00000048756 CHEBI:87103 inhibits the binding of P62806 to SMUSG00000048756 wt_cns2_enhancer_probe-jun-1 1 10 unspecified role unspecified role prey prey subcellular prep subcellular preparation subcellular prep mouse-t-cell Mouse T cell t-cell Peripheral blood T-lymphocytes 9 unspecified role unspecified role bait bait A9G,C10G,G11T,T12A,A14C,A21G,C22G,A23T,T24A mutation disrupting strength mutation disrupting interaction strength mutation disrupting strength certain certain sequence position certain certain certain sequence position certain ACAT GGTA certain certain sequence position certain certain certain sequence position certain ACGT GGTA certain certain sequence position certain certain certain sequence position certain A C region biotin tag biotin tag undetermined undetermined sequence position undetermined undetermined undetermined sequence position undetermined chemical synthesis Chemical synthesis (Chemical synthesis) physical association physical association 4C FLAG-c-Fos was also detected, but the origin of this construct is not stated. This interaction was seen in cells activated by anti-CD3/28, with and without TGF-beta, but not in non-activated cells. The Bait is stated to be a 102 bp DNA fragment containing three copies of the 32bp sequence given in Fig 4A. However the exact sequence is not stated, so the 32 bp sequence only has been entered here. jun-foxo3-1 2 6 unspecified role unspecified role prey prey cns2_enhancer_region sufficient to bind sufficient binding region sufficient to bind undetermined undetermined sequence position undetermined undetermined undetermined sequence position undetermined 10 unspecified role unspecified role bait bait association association 4 E Interaction was seen in nTReg and iTReg cells. hist1h4h-foxo3-3 4 6 unspecified role unspecified role prey prey cns2_enhancer_region sufficient to bind sufficient binding region sufficient to bind undetermined undetermined sequence position undetermined undetermined undetermined sequence position undetermined 7 unspecified role unspecified role bait bait possible_acres ac1res monoacetylated residue Acetyl Acetylation (N terminus, N epsilon of Lysine, O of Serine) (Ac) Acetylation Ac1Res undetermined undetermined sequence position undetermined undetermined undetermined sequence position undetermined observed-ptm observed-ptm observed ptm observed-ptm association association 7D Stat5 binding to this region was detected at 18hrs after stimulation and then gradually diminished;however binding of AcH4, c-Fos and Creb1 are at low levels at 18hrs, but increase by 48hrs, and are greatest at 72hrs. fos-foxo3-2 4 6 unspecified role unspecified role prey prey cns2_enhancer_region sufficient to bind sufficient binding region sufficient to bind undetermined undetermined sequence position undetermined undetermined undetermined sequence position undetermined 8 unspecified role unspecified role bait bait association association 7D Stat5 binding to this region was detected at 18hrs after stimulation and then gradually diminished;however binding of AcH4, c-Fos and Creb1 are at low levels at 18hrs, but increase by 48hrs, and are greatest at 72hrs. creb1-foxo3-1 4 6 unspecified role unspecified role prey prey cns2_enhancer_region sufficient to bind sufficient binding region sufficient to bind undetermined undetermined sequence position undetermined undetermined undetermined sequence position undetermined 5 unspecified role unspecified role bait bait association association 7Di Stat5 binding to this region was detected at 18hrs after stimulation and then gradually diminished;however binding of AcH4, c-Fos and Creb1 are at low levels at 18hrs, but increase by 48hrs, and are greatest at 72hrs. fos-foxo3-1 2 6 unspecified role unspecified role prey prey cns2_enhancer_region sufficient to bind sufficient binding region sufficient to bind undetermined undetermined sequence position undetermined undetermined undetermined sequence position undetermined 8 unspecified role unspecified role bait bait association association 8 CHEBI:87103 JAK inhibitor CHEBI:87662 STAT5 inhibitor CHEBI:87662 decreases the binding of P01101 to ENSMUSG00000048756 CHEBI:87103 inhibits binding of P01101 to ENSMUSG00000048756 creb1-foxo3-2 2 6 unspecified role unspecified role prey prey cns2_enhancer_region sufficient to bind sufficient binding region sufficient to bind undetermined undetermined sequence position undetermined undetermined undetermined sequence position undetermined 5 unspecified role unspecified role bait bait association association 8 CHEBI:87103 - JAK inhibitor CHEBI:87662 STAT5 inhibitor CHEBI:87103 inhibits binding of Q01147 to ENSMUSG00000048756 CHEBI:87662 decreases the binding of Q01147 to ENSMUSG00000048756 CHEBI:87103 decreases the expression of ENSMUSG00000048756 at protein level. hist1h4h-foxo3-4 3 7 unspecified role unspecified role bait bait possible_acres ac1res monoacetylated residue Acetyl Acetylation (N terminus, N epsilon of Lysine, O of Serine) (Ac) Acetylation Ac1Res undetermined undetermined sequence position undetermined undetermined undetermined sequence position undetermined observed-ptm observed-ptm observed ptm observed-ptm 6 unspecified role unspecified role prey prey cns2_enhancer_region sufficient to bind sufficient binding region sufficient to bind undetermined undetermined sequence position undetermined undetermined undetermined sequence position undetermined association association 1A The 5' half of the CNS2 were highly acetylated in nTreg and TGF-beta activated iTReg cells, but not in CD3/28 activated cells. CHEBI:87662 -Stat5 inhibitor CHEBI:87103 JAK inhibitor CHEBI:87662 decreases the binding of P62806 to ENSMUSG00000048756 CHEBI:87103 inhibits the binding of P62806 to SMUSG00000048756 jun-foxo3-2 3 6 unspecified role unspecified role prey prey cns2_enhancer_region sufficient to bind sufficient binding region sufficient to bind undetermined undetermined sequence position undetermined undetermined undetermined sequence position undetermined 10 unspecified role unspecified role bait bait association association 4 F Interaction was seen in nTReg and iTReg cells.