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/ http://www.ebi.ac.uk/ hao-2013-2 Gain of Interaction with IRS1 by p110a-Helical Domain Mutants Is Crucial for Their Oncogenic Functions. human-dld1 Human colorectal adenocarcinoma dld1 Human colon adenocarcinoma anti tag coip anti tag coimmunoprecipitation anti tag coip western blot western blot Immuno blot Immuno blot Hao Y., Wang C., Cao B., Hirsch B.M., Song J., Markowitz S.D., Ewing R.M., Sedwick D., Liu L., Zheng W., Wang Z. Cancer - Interactions investigated in the context of cancer All IRS1 primers were found to be of Human origin by blasting, except the His/GST IRS1 M-C2 sequence, whose species of origin could not be determined.(-these sequences were not murine or rattine in origin). Primers for the Myc-3YF construct (Y608F, Y628F, Y658F) which judging by the amino acid numbering given would have been assumed to have been Murine in origin, were in fact determined to be Human on blasting. The authors have not replied to any requests for further info. wzheng@ujs.edu.cn,zxw22@case.edu Cancer Cell (1535-6108) 2013 Only protein-protein interactions imex curation hao-2013-3 Gain of Interaction with IRS1 by p110a-Helical Domain Mutants Is Crucial for Their Oncogenic Functions. human-dld1 Human colorectal adenocarcinoma dld1 Human colon adenocarcinoma anti bait coip anti bait coimmunoprecipitation anti bait coip western blot western blot Immuno blot Immuno blot Hao Y., Wang C., Cao B., Hirsch B.M., Song J., Markowitz S.D., Ewing R.M., Sedwick D., Liu L., Zheng W., Wang Z. Cancer - Interactions investigated in the context of cancer All IRS1 primers were found to be of Human origin by blasting, except the His/GST IRS1 M-C2 sequence, whose species of origin could not be determined.(-these sequences were not murine or rattine in origin). Primers for the Myc-3YF construct (Y608F, Y628F, Y658F) which judging by the amino acid numbering given would have been assumed to have been Murine in origin, were in fact determined to be Human on blasting. The authors have not replied to any requests for further info. wzheng@ujs.edu.cn,zxw22@case.edu Cancer Cell (1535-6108) 2013 Only protein-protein interactions imex curation hao-2013-4 Gain of Interaction with IRS1 by p110a-Helical Domain Mutants Is Crucial for Their Oncogenic Functions. human-293 Homo sapiens transformed primary embryonal kidney cells Human 293 Transformed human primary embryonal kidney cells. The Cell type named 293 is synonymous with HEK-293. anti tag coip anti tag coimmunoprecipitation anti tag coip anti tag western anti tag western blot anti tag western Hao Y., Wang C., Cao B., Hirsch B.M., Song J., Markowitz S.D., Ewing R.M., Sedwick D., Liu L., Zheng W., Wang Z. Cancer - Interactions investigated in the context of cancer All IRS1 primers were found to be of Human origin by blasting, except the His/GST IRS1 M-C2 sequence, whose species of origin could not be determined.(-these sequences were not murine or rattine in origin). Primers for the Myc-3YF construct (Y608F, Y628F, Y658F) which judging by the amino acid numbering given would have been assumed to have been Murine in origin, were in fact determined to be Human on blasting. The authors have not replied to any requests for further info. wzheng@ujs.edu.cn,zxw22@case.edu Cancer Cell (1535-6108) 2013 Only protein-protein interactions imex curation hao-2013-6 Gain of Interaction with IRS1 by p110a-Helical Domain Mutants Is Crucial for Their Oncogenic Functions. in vitro In vitro pull down pull down western blot western blot Immuno blot Immuno blot Hao Y., Wang C., Cao B., Hirsch B.M., Song J., Markowitz S.D., Ewing R.M., Sedwick D., Liu L., Zheng W., Wang Z. Cancer - Interactions investigated in the context of cancer All IRS1 primers were found to be of Human origin by blasting, except the His/GST IRS1 M-C2 sequence, whose species of origin could not be determined.(-these sequences were not murine or rattine in origin). Primers for the Myc-3YF construct (Y608F, Y628F, Y658F) which judging by the amino acid numbering given would have been assumed to have been Murine in origin, were in fact determined to be Human on blasting. The authors have not replied to any requests for further info. wzheng@ujs.edu.cn,zxw22@case.edu Cancer Cell (1535-6108) 2013 Only protein-protein interactions imex curation hao-2013-7 Gain of Interaction with IRS1 by p110a-Helical Domain Mutants Is Crucial for Their Oncogenic Functions. human-vaco481 HUMAN VACO481- colon carcinoma with q546p mutation vaco481 VACO481 PMID: 10822375 useful anti bait coip anti bait coimmunoprecipitation anti bait coip western blot western blot Immuno blot Immuno blot Hao Y., Wang C., Cao B., Hirsch B.M., Song J., Markowitz S.D., Ewing R.M., Sedwick D., Liu L., Zheng W., Wang Z. Cancer - Interactions investigated in the context of cancer All IRS1 primers were found to be of Human origin by blasting, except the His/GST IRS1 M-C2 sequence, whose species of origin could not be determined.(-these sequences were not murine or rattine in origin). Primers for the Myc-3YF construct (Y608F, Y628F, Y658F) which judging by the amino acid numbering given would have been assumed to have been Murine in origin, were in fact determined to be Human on blasting. The authors have not replied to any requests for further info. wzheng@ujs.edu.cn,zxw22@case.edu Cancer Cell (1535-6108) 2013 Only protein-protein interactions imex curation hao-2013-9 Gain of Interaction with IRS1 by p110a-Helical Domain Mutants Is Crucial for Their Oncogenic Functions. human Homo sapiens Human anti tag coip anti tag coimmunoprecipitation anti tag coip western blot western blot Immuno blot Immuno blot Hao Y., Wang C., Cao B., Hirsch B.M., Song J., Markowitz S.D., Ewing R.M., Sedwick D., Liu L., Zheng W., Wang Z. Cancer - Interactions investigated in the context of cancer The cell line used in these experiments is not known and the authors have not supplied the missing info when requested. All IRS1 primers were found to be of Human origin by blasting, except the His/GST IRS1 M-C2 sequence, whose species of origin could not be determined.(-these sequences were not murine or rattine in origin). Primers for the Myc-3YF construct (Y608F, Y628F, Y658F) which judging by the amino acid numbering given would have been assumed to have been Murine in origin, were in fact determined to be Human on blasting. The authors have not replied to any requests for further info. wzheng@ujs.edu.cn,zxw22@case.edu Cancer Cell (1535-6108) 2013 Only protein-protein interactions imex curation hao-2013-1 Gain of Interaction with IRS1 by p110a-Helical Domain Mutants Is Crucial for Their Oncogenic Functions. human-dld1 Human colorectal adenocarcinoma dld1 Human colon adenocarcinoma anti tag coip anti tag coimmunoprecipitation anti tag coip ms participant Identification by mass spectrometry ms participant Hao Y., Wang C., Cao B., Hirsch B.M., Song J., Markowitz S.D., Ewing R.M., Sedwick D., Liu L., Zheng W., Wang Z. Cancer - Interactions investigated in the context of cancer All IRS1 primers were found to be of Human origin by blasting, except the His/GST IRS1 M-C2 sequence, whose species of origin could not be determined.(-these sequences were not murine or rattine in origin). Primers for the Myc-3YF construct (Y608F, Y628F, Y658F) which judging by the amino acid numbering given would have been assumed to have been Murine in origin, were in fact determined to be Human on blasting. The authors have not replied to any requests for further info. wzheng@ujs.edu.cn,zxw22@case.edu Cancer Cell (1535-6108) 2013 Only protein-protein interactions imex curation hao-2013-5 Gain of Interaction with IRS1 by p110a-Helical Domain Mutants Is Crucial for Their Oncogenic Functions. in vitro In vitro anti tag coip anti tag coimmunoprecipitation anti tag coip western blot western blot Immuno blot Immuno blot Hao Y., Wang C., Cao B., Hirsch B.M., Song J., Markowitz S.D., Ewing R.M., Sedwick D., Liu L., Zheng W., Wang Z. Cancer - Interactions investigated in the context of cancer All IRS1 primers were found to be of Human origin by blasting, except the His/GST IRS1 M-C2 sequence, whose species of origin could not be determined.(-these sequences were not murine or rattine in origin). Primers for the Myc-3YF construct (Y608F, Y628F, Y658F) which judging by the amino acid numbering given would have been assumed to have been Murine in origin, were in fact determined to be Human on blasting. The authors have not replied to any requests for further info. wzheng@ujs.edu.cn,zxw22@case.edu Cancer Cell (1535-6108) 2013 Only protein-protein interactions imex curation hao-2013-8 Gain of Interaction with IRS1 by p110a-Helical Domain Mutants Is Crucial for Their Oncogenic Functions. in vitro In vitro phosphotransferase phosphotransferase assay phosphotransferase autoradiography autoradiography Hao Y., Wang C., Cao B., Hirsch B.M., Song J., Markowitz S.D., Ewing R.M., Sedwick D., Liu L., Zheng W., Wang Z. Cancer - Interactions investigated in the context of cancer All IRS1 primers were found to be of Human origin by blasting, except the His/GST IRS1 M-C2 sequence, whose species of origin could not be determined.(-these sequences were not murine or rattine in origin). Primers for the Myc-3YF construct (Y608F, Y628F, Y658F) which judging by the amino acid numbering given would have been assumed to have been Murine in origin, were in fact determined to be Human on blasting. The authors have not replied to any requests for further info. wzheng@ujs.edu.cn,zxw22@case.edu Cancer Cell (1535-6108) 2013 Only protein-protein interactions imex curation 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 pk3ca_human Phosphatidylinositol 4,5-bisphosphate 3-kinase catalytic subunit alpha isoform Phosphoinositide-3-kinase catalytic alpha polypeptide Serine/threonine protein kinase PIK3CA PIK3CA Phosphatidylinositol 4,5-bisphosphate 3-kinase 110 kDa catalytic subunit alpha Phosphoinositide 3-kinase alpha protein protein human Homo sapiens Human MPPRPSSGELWGIHLMPPRILVECLLPNGMIVTLECLREATLITIKHELFKEARKYPLHQLLQDESSYIFVSVTQEAEREEFFDETRRLCDLRLFQPFLKVIEPVGNREEKILNREIGFAIGMPVCEFDMVKDPEVQDFRRNILNVCKEAVDLRDLNSPHSRAMYVYPPNVESSPELPKHIYNKLDKGQIIVVIWVIVSPNNDKQKYTLKINHDCVPEQVIAEAIRKKTRSMLLSSEQLKLCVLEYQGKYILKVCGCDEYFLEKYPLSQYKYIRSCIMLGRMPNLMLMAKESLYSQLPMDCFTMPSYSRRISTATPYMNGETSTKSLWVINSALRIKILCATYVNVNIRDIDKIYVRTGIYHGGEPLCDNVNTQRVPCSNPRWNEWLNYDIYIPDLPRAARLCLSICSVKGRKGAKEEHCPLAWGNINLFDYTDTLVSGKMALNLWPVPHGLEDLLNPIGVTGSNPNKETPCLELEFDWFSSVVKFPDMSVIEEHANWSVSREAGFSYSHAGLSNRLARDNELRENDKEQLKAISTRDPLSEITEQEKDFLWSHRHYCVTIPEILPKLLLSVKWNSRDEVAQMYCLVKDWPPIKPEQAMELLDCNYPDPMVRGFAVRCLEKYLTDDKLSQYLIQLVQVLKYEQYLDNLLVRFLLKKALTNQRIGHFFFWHLKSEMHNKTVSQRFGLLLESYCRACGMYLKHLNRQVEAMEKLINLTDILKQEKKDETQKVQMKFLVEQMRRPDFMDALQGFLSPLNPAHQLGNLRLEECRIMSSAKRPLWLNWENPDIMSELLFQNNEIIFKNGDDLRQDMLTLQIIRIMENIWQNQGLDLRMLPYGCLSIGDCVGLIEVVRNSHTIMQIQCKGGLKGALQFNSHTLHQWLKDKNKGEIYDAAIDLFTRSCAGYCVATFILGIGDRHNSNIMVKDDGQLFHIDFGHFLDHKKKKFGYKRERVPFVLTQDFLIVISKGAQECTKTREFERFQEMCYKAYLAIRQHANLFINLFSMMLGSGMPELQSFDDIAYIRKTLALDKTEQEALEYFMKQMNDAHHGGWTTKMDWIFHTIKQHALN irs1_human Insulin receptor substrate 1 IRS1 protein protein human Homo sapiens Human MASPPESDGFSDVRKVGYLRKPKSMHKRFFVLRAASEAGGPARLEYYENEKKWRHKSSAPKRSIPLESCFNINKRADSKNKHLVALYTRDEHFAIAADSEAEQDSWYQALLQLHNRAKGHHDGAAALGAGGGGGSCSGSSGLGEAGEDLSYGDVPPGPAFKEVWQVILKPKGLGQTKNLIGIYRLCLTSKTISFVKLNSEAAAVVLQLMNIRRCGHSENFFFIEVGRSAVTGPGEFWMQVDDSVVAQNMHETILEAMRAMSDEFRPRSKSQSSSNCSNPISVPLRRHHLNNPPPSQVGLTRRSRTESITATSPASMVGGKPGSFRVRASSDGEGTMSRPASVDGSPVSPSTNRTHAHRHRGSARLHPPLNHSRSIPMPASRCSPSATSPVSLSSSSTSGHGSTSDCLFPRRSSASVSGSPSDGGFISSDEYGSSPCDFRSSFRSVTPDSLGHTPPARGEEELSNYICMGGKGPSTLTAPNGHYILSRGGNGHRCTPGTGLGTSPALAGDEAASAADLDNRFRKRTHSAGTSPTITHQKTPSQSSVASIEEYTEMMPAYPPGGGSGGRLPGHRHSAFVPTRSYPEEGLEMHPLERRGGHHRPDSSTLHTDDGYMPMSPGVAPVPSGRKGSGDYMPMSPKSVSAPQQIINPIRRHPQRVDPNGYMMMSPSGGCSPDIGGGPSSSSSSSNAVPSGTSYGKLWTNGVGGHHSHVLPHPKPPVESSGGKLLPCTGDYMNMSPVGDSNTSSPSDCYYGPEDPQHKPVLSYYSLPRSFKHTQRPGEPEEGARHQHLRLSTSSGRLLYAATADDSSSSTSSDSLGGGYCGARLEPSLPHPHHQVLQPHLPRKVDTAAQTNSRLARPTRLSLGDPKASTLPRAREQQQQQQPLLHPPEPKSPGEYVNIEFGSDQSGYLSGPVAFHSSPSVRCPSQLQPAPREEETGTEEYMKMDLGPGRRAAWQESTGVEMGRLGPAPPGAASICRPTRAVPSSRGDYMTMQMSCPRQSYVDTSPAAPVSYADMRTGIAAEEVSLPRATMAAASSSSAASASPTGPQGAAELAAHSSLLGGPQGPGGMSAFTRVNLSPNRNQSAKVIRADPQGCRRRHSSETFSSTPSATRVGNTVPFGAGAAVGGGGGSSSSSEDVKRHSSASFENVWLRPGELGGAPKEPAKLCGAAGGLENGLNYIDLDLVKDFKQCPQECTPEPQPPPPPPPHQPLGSGESSSTRRSSEDLSAYASISFQKQPEDRQ pi Phosphatidylinositol phosphatidylinositols PI PtdIns small molecule small molecule chemical synthesis Chemical synthesis (Chemical synthesis) pik3ca-irs1-2 1390006 n/a 1390015 unspecified role unspecified role prey prey n/a 1390016 unspecified role unspecified role bait bait P42336:p.Glu545Lys mutation mutation certain certain sequence position certain certain certain sequence position certain false c_terminus flag tag flag tag FLAG-tagged FLAG DYKDDDDKV epitope tag DYKDDDDKV epitope tag FLAG FLAG-tagged c-term range C-terminal range c-term range c-term range C-terminal range c-term range false n/a 1390017 unspecified role unspecified role prey prey association association false false false 1Ci, supp 1C, 6Bi A protein of ~170 kDa was detected using Flag tagged E545K knock-in (KI) cells, but not using Wild Type cells. pik3ca-irs1-3 1390006 n/a 1390016 unspecified role unspecified role bait bait c_terminus flag tag flag tag FLAG-tagged FLAG DYKDDDDKV epitope tag DYKDDDDKV epitope tag FLAG FLAG-tagged c-term range C-terminal range c-term range c-term range C-terminal range c-term range false n/a 1390015 unspecified role unspecified role prey prey physical association physical association false false false 1Cii, supp 1C. A protein of ~170 kDa was detected using Flag tagged E545K knock-in (KI) cells, but not using Wild Type cells. pik3ca-irs1-7 1390006 n/a 1390015 unspecified role unspecified role prey prey n/a 1390016 unspecified role unspecified role bait bait P42336:p.Glu545Lys mutation increasing mutation increasing interaction mutation increasing certain certain sequence position certain certain certain sequence position certain false c_terminus flag tag flag tag FLAG-tagged FLAG DYKDDDDKV epitope tag DYKDDDDKV epitope tag FLAG FLAG-tagged c-term range C-terminal range c-term range c-term range C-terminal range c-term range false abd_domain required to bind necessary binding region required to bind range range range range false physical association physical association false false false 2B, supp 2B.i The p110α E545K-IRS1 interaction also did not require p85 because a truncated p110α E545K lacking the ABD domain required for p85 binding still associated with IRS1. pik3ca-irs1-16 1390006 n/a 1390016 unspecified role unspecified role bait bait region sufficient to bind sufficient binding region sufficient to bind range range range range false range range range range false c_terminus flag tag flag tag FLAG-tagged FLAG DYKDDDDKV epitope tag DYKDDDDKV epitope tag FLAG FLAG-tagged c-term range C-terminal range c-term range c-term range C-terminal range c-term range false P42336:p.Glu545Lys mutation increasing mutation increasing interaction mutation increasing certain certain sequence position certain certain certain sequence position certain false n/a 1390017 unspecified role unspecified role prey prey n_terminus myc tag myc tag EUKLISEED epitope tag EUKLISEED epitope tag n-term range n-terminal range n-term range n-term range n-terminal range n-term range false association association false false false 2B, supp 2B.ii, The p110α E545K-IRS1 interaction also did not require p85 because a truncated p110α E545K lacking the ABD domain required for p85 binding still associated with IRS1. pik3ca-irs1-9 1390006 n/a 1390017 unspecified role unspecified role prey prey n_terminus myc tag myc tag EUKLISEED epitope tag EUKLISEED epitope tag n-term range n-terminal range n-term range n-term range n-terminal range n-term range false n/a 1390015 unspecified role unspecified role prey prey n/a 1390016 unspecified role unspecified role bait bait c_terminus flag tag flag tag FLAG-tagged FLAG DYKDDDDKV epitope tag DYKDDDDKV epitope tag FLAG FLAG-tagged c-term range C-terminal range c-term range c-term range C-terminal range c-term range false abd_domain required to bind necessary binding region required to bind range range range range false association association false false false 2B, supp 2B. iv The p110α E545K-IRS1 interaction also did not require p85 because a truncated p110α E545K lacking the ABD domain required for p85 binding still associated with IRS1. pik3ca-irs1-13 1390006 n/a 1390017 unspecified role unspecified role prey prey n/a 1390016 unspecified role unspecified role bait bait P42336:p.Glu545Lys mutation mutation certain certain sequence position certain certain certain sequence position certain false c_terminus flag tag flag tag FLAG-tagged FLAG DYKDDDDKV epitope tag DYKDDDDKV epitope tag FLAG FLAG-tagged c-term range C-terminal range c-term range c-term range C-terminal range c-term range false n/a 1390016 inhibitor inhibitor unspecified role unspecified role P42336:p.Glu545Lys mutation mutation certain certain sequence position certain certain certain sequence position certain false region sufficient to bind sufficient binding region sufficient to bind certain certain sequence position certain certain certain sequence position certain false asp549,ser553 cross linker cross linker crosslinker crosslinker certain certain sequence position certain certain certain sequence position certain false certain certain sequence position certain certain certain sequence position certain false MOD:01067 -linker attached to peptide in Fmoc peptide synthesis association association false false false 6Bii The stapled mutant peptide effectively disrupted IRS1-p110α E545K interaction.The stapled mutant peptide did not interfere with cellular p110α-p85 interaction. irs1-pik3ca-10 1390006 n/a 1390015 unspecified role unspecified role prey prey region ha tag ha tag YPYDVPDYA epitope tag YPYDVPDYA epitope tag undetermined undetermined sequence position undetermined undetermined undetermined sequence position undetermined false n/a 1390017 unspecified role unspecified role bait bait purified purified n_terminus myc tag myc tag EUKLISEED epitope tag EUKLISEED epitope tag n-term range n-terminal range n-term range n-term range n-terminal range n-term range false region sufficient to bind sufficient binding region sufficient to bind certain certain sequence position certain certain certain sequence position certain false certain certain sequence position certain certain certain sequence position certain false n/a 1390016 unspecified role unspecified role prey prey purified purified P42336:p.Glu545Lys mutation increasing mutation increasing interaction mutation increasing certain certain sequence position certain certain certain sequence position certain false c_terminus flag tag flag tag FLAG-tagged FLAG DYKDDDDKV epitope tag DYKDDDDKV epitope tag FLAG FLAG-tagged c-term range C-terminal range c-term range c-term range C-terminal range c-term range false association association false false false Supp 3H A deletion construct of IRS1 (IRS1Δ) devoid of both AA 607–623 and AA 936–962 regions greatly reduced its interaction with p110α E545K. The IRS1 deletion construct did not affect the IRS1-p85 interaction. irs1-pik3ca-11 1390006 n/a 1390017 unspecified role unspecified role bait bait purified purified region required to bind necessary binding region required to bind certain certain sequence position certain certain certain sequence position certain false certain certain sequence position certain certain certain sequence position certain false n_terminus myc tag myc tag EUKLISEED epitope tag EUKLISEED epitope tag n-term range n-terminal range n-term range n-term range n-terminal range n-term range false n/a 1390016 unspecified role unspecified role prey prey purified purified P42336:p.Glu545Lys mutation mutation certain certain sequence position certain certain certain sequence position certain false association association false false false Supp 5D A deletion construct of IRS1 (IRS1Δ) devoid of both AA 607–623 and AA 936–962 regions greatly reduced its interaction with p110α. irs1-pik3ca 1390007 n/a 1390016 unspecified role unspecified role prey prey P42336:p.Glu545Lys mutation increasing mutation increasing interaction mutation increasing certain certain sequence position certain certain certain sequence position certain false n/a 1390017 unspecified role unspecified role bait bait physical association physical association false false false 1Di, supp1D. irs1-pik3ca-1 1390007 n/a 1390017 unspecified role unspecified role bait bait n/a 1390016 unspecified role unspecified role prey prey P42336:p.Glu545Lys mutation increasing mutation increasing interaction mutation increasing certain certain sequence position certain certain certain sequence position certain false c_terminus flag tag flag tag FLAG-tagged FLAG DYKDDDDKV epitope tag DYKDDDDKV epitope tag FLAG FLAG-tagged c-term range C-terminal range c-term range c-term range C-terminal range c-term range false physical association physical association false false false 1Dii, supp 1D irs1-pik3ca-5 1390007 n/a 1390015 unspecified role unspecified role prey prey n/a 1390016 unspecified role unspecified role prey prey P42336:p.Glu545Lys mutation increasing mutation increasing interaction mutation increasing certain certain sequence position certain certain certain sequence position certain false n/a 1390017 unspecified role unspecified role bait bait association association false false false 2A, supp 2A Insulin and pervanadate p110α E545K, but not the WT p110α, robustly associated with IRS1 under serum-starvation conditions in which tyrosine phosphorylation of IRS1 was undetectable. pik3ca-irs1-10 1390007 n/a 1390015 unspecified role unspecified role prey prey n/a 1390017 unspecified role unspecified role prey prey n/a 1390016 unspecified role unspecified role bait bait P42336:p.Glu545Lys mutation mutation certain certain sequence position certain certain certain sequence position certain false association association false false false 2E, supp 2D Ablation of p85α and p85β enhanced rather than inhibited the interaction between p110α E545K and IRS1. irs1-pik3ca-3 1390008 n/a 1390017 unspecified role unspecified role bait bait n_terminus myc tag myc tag EUKLISEED epitope tag EUKLISEED epitope tag n-term range n-terminal range n-term range n-term range n-terminal range n-term range false n/a 1390016 unspecified role unspecified role prey prey P42336:p.Glu545Lys mutation increasing mutation increasing interaction mutation increasing certain certain sequence position certain certain certain sequence position certain false c_terminus flag tag flag tag FLAG-tagged FLAG DYKDDDDKV epitope tag DYKDDDDKV epitope tag FLAG FLAG-tagged c-term range C-terminal range c-term range c-term range C-terminal range c-term range false P42336:p.Glu542Lys mutation increasing mutation increasing interaction mutation increasing certain certain sequence position certain certain certain sequence position certain false P42336:p.Asn345Lys mutation increasing mutation increasing interaction mutation increasing certain certain sequence position certain certain certain sequence position certain false P42336:p.Met1043Ile mutation increasing mutation increasing interaction mutation increasing certain certain sequence position certain certain certain sequence position certain false P42336:p.Gln546Lys mutation increasing mutation increasing interaction mutation increasing certain certain sequence position certain certain certain sequence position certain false P42336:p.Glu545Gly mutation increasing mutation increasing interaction mutation increasing certain certain sequence position certain certain certain sequence position certain false P42336:p.Glu545Ala mutation increasing mutation increasing interaction mutation increasing certain certain sequence position certain certain certain sequence position certain false P42336:p.Lys111Asn mutation increasing mutation increasing interaction mutation increasing certain certain sequence position certain certain certain sequence position certain false physical association physical association false false false 1E, Supp1E. The presence of the following p110 mutations did not result in a statistically significant interaction with IRS1 :- H1047R, H1047L, G1049R, R88Q, C420R, P539R. The authors state that p110 molecules containing these mutations have "failed to interact with IRS1". irs1-pik3ca-9 1390008 n/a 1390015 unspecified role unspecified role prey prey region ha tag ha tag YPYDVPDYA epitope tag YPYDVPDYA epitope tag undetermined undetermined sequence position undetermined undetermined undetermined sequence position undetermined false n/a 1390016 unspecified role unspecified role prey prey purified purified P42336:p.Glu545Lys mutation increasing mutation increasing interaction mutation increasing certain certain sequence position certain certain certain sequence position certain false c_terminus flag tag flag tag FLAG-tagged FLAG DYKDDDDKV epitope tag DYKDDDDKV epitope tag FLAG FLAG-tagged c-term range C-terminal range c-term range c-term range C-terminal range c-term range false n/a 1390017 unspecified role unspecified role bait bait purified purified region sufficient to bind sufficient binding region sufficient to bind certain certain sequence position certain certain certain sequence position certain false certain certain sequence position certain certain certain sequence position certain false region myc tag myc tag EUKLISEED epitope tag EUKLISEED epitope tag undetermined undetermined sequence position undetermined undetermined undetermined sequence position undetermined false association association false false false 3G/H, Supp 3D–S3G A deletion construct of IRS1 (IRS1Δ) devoid of both AA 607–623 and AA 936–962 regions greatly reduced its interaction with p110α E545K. Single region deletions could not interrupt the IRS1 mutant p110α interaction. The IRS1 deletion construct did not affect the IRS1-p85 interaction. irs1-pik3ca-6 1390009 n/a 1390016 unspecified role unspecified role prey prey purified purified P42336:p.Glu545Lys mutation mutation certain certain sequence position certain certain certain sequence position certain false human-dld1 Human colorectal adenocarcinoma dld1 Human colon adenocarcinoma n/a 1390017 unspecified role unspecified role bait bait purified purified region sufficient to bind sufficient binding region sufficient to bind certain certain sequence position certain certain certain sequence position certain false certain certain sequence position certain certain certain sequence position certain false region his tag his tag Hexa-His-tag 6-His-tag Histidine-tag Hexa-His-tag Histidine-tag 6-His-tag undetermined undetermined sequence position undetermined undetermined undetermined sequence position undetermined false ecoli Escherichia coli (strain K12) physical association physical association false false false 3B,3D No binding was observed using the WT cells. irs1-pik3ca-7 1390009 n/a 1390017 unspecified role unspecified role bait bait purified purified region 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 false region sufficient to bind sufficient binding region sufficient to bind certain certain sequence position certain certain certain sequence position certain false certain certain sequence position certain certain certain sequence position certain false ecoli Escherichia coli (strain K12) n/a 1390016 unspecified role unspecified role prey prey purified purified P42336:p.Glu545Lys mutation mutation certain certain sequence position certain certain certain sequence position certain false human-dld1 Human colorectal adenocarcinoma dld1 Human colon adenocarcinoma physical association physical association false false false 3F pik3ca-irs1-14 1390010 n/a 1390017 unspecified role unspecified role prey prey n/a 1390016 unspecified role unspecified role bait bait P42336:p.Gln546Pro mutation mutation certain certain sequence position certain certain certain sequence position certain false n/a 1390015 unspecified role unspecified role prey prey association association false false false 6C pik3ca-irs1-15 1390010 n/a 1390016 inhibitor inhibitor unspecified role unspecified role P42336:p.Glu545Lys mutation mutation certain certain sequence position certain certain certain sequence position certain false asp549,ser553 cross linker cross linker crosslinker crosslinker certain certain sequence position certain certain certain sequence position certain false certain certain sequence position certain certain certain sequence position certain false MOD:01067 -linker attached to peptide in Fmoc peptide synthesis region sufficient to bind sufficient binding region sufficient to bind certain certain sequence position certain certain certain sequence position certain false n/a 1390016 unspecified role unspecified role bait bait c_terminus flag tag flag tag FLAG-tagged FLAG DYKDDDDKV epitope tag DYKDDDDKV epitope tag FLAG FLAG-tagged c-term range C-terminal range c-term range c-term range C-terminal range c-term range false P42336:p.Gln546Pro mutation mutation certain certain sequence position certain certain certain sequence position certain false n/a 1390017 unspecified role unspecified role prey prey association association false false false 6Ci The p110α E545K stapled peptide also disrupted protein interaction between p110α Q546P and IRS1 in Vaco481 CRC cells. pik3ca-irs1-6 1390011 n/a 1390015 unspecified role unspecified role prey prey n/a 1390017 unspecified role unspecified role prey prey n_terminus myc tag myc tag EUKLISEED epitope tag EUKLISEED epitope tag n-term range n-terminal range n-term range n-term range n-terminal range n-term range false n/a 1390016 unspecified role unspecified role bait bait P42336:p.Glu545Lys mutation increasing mutation increasing interaction mutation increasing certain certain sequence position certain certain certain sequence position certain false c_terminus flag tag flag tag FLAG-tagged FLAG DYKDDDDKV epitope tag DYKDDDDKV epitope tag FLAG FLAG-tagged c-term range C-terminal range c-term range c-term range C-terminal range c-term range false association association false false false 2B, supp 2B. The p110α E545K-IRS1 interaction also did not require p85 because a truncated p110α E545K lacking the ABD domain required for p85 binding still associated with IRS1. pik3ca-irs1-17 1390011 n/a 1390017 unspecified role unspecified role prey prey n_terminus myc tag myc tag EUKLISEED epitope tag EUKLISEED epitope tag n-term range n-terminal range n-term range n-term range n-terminal range n-term range false n/a 1390016 unspecified role unspecified role bait bait c_terminus flag tag flag tag FLAG-tagged FLAG DYKDDDDKV epitope tag DYKDDDDKV epitope tag FLAG FLAG-tagged c-term range C-terminal range c-term range c-term range C-terminal range c-term range false P42336:p.Glu545Lys mutation increasing mutation increasing interaction mutation increasing certain certain sequence position certain certain certain sequence position certain false region sufficient to bind sufficient binding region sufficient to bind range range range range false range range range range false n/a 1390016 inhibitor inhibitor unspecified role unspecified role asp549,ser553 cross linker cross linker crosslinker crosslinker certain certain sequence position certain certain certain sequence position certain false certain certain sequence position certain certain certain sequence position certain false association association false false false supp 6D. the stapled mutant peptide also inhibited the interaction between IRS1 and ΔABD p110α E545K. pik3ca-irs1 1390012 n/a 1390017 unspecified role unspecified role prey prey n/a 1390016 unspecified role unspecified role bait bait c_terminus flag tag flag tag FLAG-tagged FLAG DYKDDDDKV epitope tag DYKDDDDKV epitope tag FLAG FLAG-tagged c-term range C-terminal range c-term range c-term range C-terminal range c-term range false P42336:p.Glu545Lys mutation mutation certain certain sequence position certain certain certain sequence position certain false physical association physical association false false false 1B A protein of ~170 kDa was detected using Flag tagged E545K knock-in (KI) cells, but not using Wild Type cells. pik3ca-irs1-11 1390013 n/a 1390016 unspecified role unspecified role bait bait purified purified region his tag his tag Hexa-His-tag 6-His-tag Histidine-tag Hexa-His-tag Histidine-tag 6-His-tag undetermined undetermined sequence position undetermined undetermined undetermined sequence position undetermined false glu545lys required to bind necessary binding region required to bind certain certain sequence position certain certain certain sequence position certain false region myc tag myc tag EUKLISEED epitope tag EUKLISEED epitope tag undetermined undetermined sequence position undetermined undetermined undetermined sequence position undetermined false spofr-sf_9 Spodoptera frugiperda insect cells Fall armyworm sf_9 insect cells Derived from pupal ovarian tissue of the fall armyworm Cloned from the parent cell line, IPLB-SF-21-AE (SF-21) derived from immature ovaries of fall armyworm Spodoptera frugiperda pupae cells are susceptible to Baculovirus infection n/a 1390017 unspecified role unspecified role prey prey purified purified spofr-sf_9 Spodoptera frugiperda insect cells Fall armyworm sf_9 insect cells Derived from pupal ovarian tissue of the fall armyworm Cloned from the parent cell line, IPLB-SF-21-AE (SF-21) derived from immature ovaries of fall armyworm Spodoptera frugiperda pupae cells are susceptible to Baculovirus infection direct interaction direct interaction false false false 2G, supp 2F recombinant IRS1 bound directly in vitro with p110α E545K but not WT p110α. pi-pik3ca-1 1390014 n/a 1390015 ancillary ancillary unspecified role unspecified role ni_sh2_domains sufficient to bind sufficient binding region sufficient to bind certain certain sequence position certain certain certain sequence position certain false spofr-sf_9 Spodoptera frugiperda insect cells Fall armyworm sf_9 insect cells Derived from pupal ovarian tissue of the fall armyworm Cloned from the parent cell line, IPLB-SF-21-AE (SF-21) derived from immature ovaries of fall armyworm Spodoptera frugiperda pupae cells are susceptible to Baculovirus infection n/a 1390016 enzyme enzyme unspecified role unspecified role purified purified glu545lys required to bind necessary binding region required to bind certain certain sequence position certain certain certain sequence position certain false n_terminus his tag his tag Hexa-His-tag 6-His-tag Histidine-tag Hexa-His-tag Histidine-tag 6-His-tag n-term range n-terminal range n-term range n-term range n-terminal range n-term range false spofr-sf_9 Spodoptera frugiperda insect cells Fall armyworm sf_9 insect cells Derived from pupal ovarian tissue of the fall armyworm Cloned from the parent cell line, IPLB-SF-21-AE (SF-21) derived from immature ovaries of fall armyworm Spodoptera frugiperda pupae cells are susceptible to Baculovirus infection n/a 1390018 enzyme target enzyme target substrate substrate unspecified role unspecified role chemical synthesis Chemical synthesis (Chemical synthesis) phosphorylation phosphorylation reaction phosphorylation false false false Supp 6 J/K The stapled mutant peptide has no effect on the kinase activity of recombinant p110α, either alone or in the presence of IRS1, in vitro. Full-length IRS1 proteins also had no effect on the lipid kinase activity of p110α E545K, in vitro.