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/ lito-2014-6 human-a_549 Homo sapiens Caucasian lung carcinoma cell line a_549 Human Caucasian lung carcinoma anti bait coip anti bait coimmunoprecipitation anti bait coip western blot western blot Immuno blot Immuno blot Lito P., Saborowski A., Yue J., Solomon M., Joseph E., Gadal S., Saborowski M., Kastenhuber E., Fellmann C., Ohara K., Morikami K., Miura T., Lukacs C., Ishii N., Lowe S., Rosen N. Cancer - Interactions investigated in the context of cancer MEK1-, KRas-, and NRas-encoding plasmids are stated to have been obtained from Biomyx; constructs containing these proteins have been entered as Human, as no equivalent mouse products seem to be available on the Biomyx website. rosenn@mskcc.org,lowes@mskcc.org Time of drug treatment Drug used Cancer Cell (1535-6108) 2014 Only protein-protein interactions imex curation lito-2014-1 Disruption of CRAF-Mediated MEK Activation Is Required for Effective MEK Inhibition in KRAS Mutant Tumors. 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 western blot western blot Immuno blot Immuno blot Lito P., Saborowski A., Yue J., Solomon M., Joseph E., Gadal S., Saborowski M., Kastenhuber E., Fellmann C., Ohara K., Morikami K., Miura T., Lukacs C., Ishii N., Lowe S., Rosen N. Cancer - Interactions investigated in the context of cancer MEK1-, KRas-, and NRas-encoding plasmids are stated to have been obtained from Biomyx; constructs containing these proteins have been entered as Human, as no equivalent mouse products seem to be available on the Biomyx website. rosenn@mskcc.org,lowes@mskcc.org Cancer Cell (1535-6108) 2014 Only protein-protein interactions imex curation lito-2014-4 Disruption of CRAF-Mediated MEK Activation Is Required for Effective MEK Inhibition in KRAS Mutant Tumors. in vitro In vitro spr surface plasmon resonance Optical biosensor BIAcore(r) BIAcore(r) Optical biosensor spr predetermined predetermined participant predetermined Lito P., Saborowski A., Yue J., Solomon M., Joseph E., Gadal S., Saborowski M., Kastenhuber E., Fellmann C., Ohara K., Morikami K., Miura T., Lukacs C., Ishii N., Lowe S., Rosen N. Cancer - Interactions investigated in the context of cancer MEK1-, KRas-, and NRas-encoding plasmids are stated to have been obtained from Biomyx; constructs containing these proteins have been entered as Human, as no equivalent mouse products seem to be available on the Biomyx website. rosenn@mskcc.org,lowes@mskcc.org Cancer Cell (1535-6108) 2014 Only protein-protein interactions imex curation lito-2014-2 Disruption of CRAF-Mediated MEK Activation Is Required for Effective MEK Inhibition in KRAS Mutant Tumors. human-a375 Human malignant melanoma a375 Human malignant melanoma ATCC catalog # CRL-1872 anti bait coip anti bait coimmunoprecipitation anti bait coip western blot western blot Immuno blot Immuno blot Lito P., Saborowski A., Yue J., Solomon M., Joseph E., Gadal S., Saborowski M., Kastenhuber E., Fellmann C., Ohara K., Morikami K., Miura T., Lukacs C., Ishii N., Lowe S., Rosen N. Cancer - Interactions investigated in the context of cancer MEK1-, KRas-, and NRas-encoding plasmids are stated to have been obtained from Biomyx; constructs containing these proteins have been entered as Human, as no equivalent mouse products seem to be available on the Biomyx website. rosenn@mskcc.org,lowes@mskcc.org Cancer Cell (1535-6108) 2014 Only protein-protein interactions imex curation lito-2014-3 Disruption of CRAF-Mediated MEK Activation Is Required for Effective MEK Inhibition in KRAS Mutant Tumors. human-a_549 Homo sapiens Caucasian lung carcinoma cell line a_549 Human Caucasian lung carcinoma anti bait coip anti bait coimmunoprecipitation anti bait coip western blot western blot Immuno blot Immuno blot Lito P., Saborowski A., Yue J., Solomon M., Joseph E., Gadal S., Saborowski M., Kastenhuber E., Fellmann C., Ohara K., Morikami K., Miura T., Lukacs C., Ishii N., Lowe S., Rosen N. Cancer - Interactions investigated in the context of cancer MEK1-, KRas-, and NRas-encoding plasmids are stated to have been obtained from Biomyx; constructs containing these proteins have been entered as Human, as no equivalent mouse products seem to be available on the Biomyx website. rosenn@mskcc.org,lowes@mskcc.org Time of drug treatment Drug used Cancer Cell (1535-6108) 2014 Only protein-protein interactions imex curation lito-2014-5 Disruption of CRAF-Mediated MEK Activation Is Required for Effective MEK Inhibition in KRAS Mutant Tumors. in vitro In vitro x-ray diffraction x-ray crystallography X-ray X-ray x-ray diffraction predetermined predetermined participant predetermined Lito P., Saborowski A., Yue J., Solomon M., Joseph E., Gadal S., Saborowski M., Kastenhuber E., Fellmann C., Ohara K., Morikami K., Miura T., Lukacs C., Ishii N., Lowe S., Rosen N. Cancer - Interactions investigated in the context of cancer MEK1-, KRas-, and NRas-encoding plasmids are stated to have been obtained from Biomyx; constructs containing these proteins have been entered as Human, as no equivalent mouse products seem to be available on the Biomyx website. rosenn@mskcc.org,lowes@mskcc.org Cancer Cell (1535-6108) 2014 Only protein-protein interactions imex curation raf1_human RAF proto-oncogene serine/threonine-protein kinase RAF1 RAF Proto-oncogene c-RAF Raf-1 protein protein human Homo sapiens Human MEHIQGAWKTISNGFGFKDAVFDGSSCISPTIVQQFGYQRRASDDGKLTDPSKTSNTIRVFLPNKQRTVVNVRNGMSLHDCLMKALKVRGLQPECCAVFRLLHEHKGKKARLDWNTDAASLIGEELQVDFLDHVPLTTHNFARKTFLKLAFCDICQKFLLNGFRCQTCGYKFHEHCSTKVPTMCVDWSNIRQLLLFPNSTIGDSGVPALPSLTMRRMRESVSRMPVSSQHRYSTPHAFTFNTSSPSSEGSLSQRQRSTSTPNVHMVSTTLPVDSRMIEDAIRSHSESASPSALSSSPNNLSPTGWSQPKTPVPAQRERAPVSGTQEKNKIRPRGQRDSSYYWEIEASEVMLSTRIGSGSFGTVYKGKWHGDVAVKILKVVDPTPEQFQAFRNEVAVLRKTRHVNILLFMGYMTKDNLAIVTQWCEGSSLYKHLHVQETKFQMFQLIDIARQTAQGMDYLHAKNIIHRDMKSNNIFLHEGLTVKIGDFGLATVKSRWSGSQQVEQPTGSVLWMAPEVIRMQDNNPFSFQSDVYSYGIVLYELMTGELPYSHINNRDQIIFMVGRGYASPDLSKLYKNCPKAMKRLVADCVKKVKEERPLFPQILSSIELLQHSLPKINRSASEPSLHRAAHTEDINACTLTTSPRLPVF braf_human Serine/threonine-protein kinase B-raf BRAF BRAF1 RAFB1 v-Raf murine sarcoma viral oncogene homolog B1 p94 Proto-oncogene B-Raf protein protein human Homo sapiens Human MAALSGGGGGGAEPGQALFNGDMEPEAGAGAGAAASSAADPAIPEEVWNIKQMIKLTQEHIEALLDKFGGEHNPPSIYLEAYEEYTSKLDALQQREQQLLESLGNGTDFSVSSSASMDTVTSSSSSSLSVLPSSLSVFQNPTDVARSNPKSPQKPIVRVFLPNKQRTVVPARCGVTVRDSLKKALMMRGLIPECCAVYRIQDGEKKPIGWDTDISWLTGEELHVEVLENVPLTTHNFVRKTFFTLAFCDFCRKLLFQGFRCQTCGYKFHQRCSTEVPLMCVNYDQLDLLFVSKFFEHHPIPQEEASLAETALTSGSSPSAPASDSIGPQILTSPSPSKSIPIPQPFRPADEDHRNQFGQRDRSSSAPNVHINTIEPVNIDDLIRDQGFRGDGGSTTGLSATPPASLPGSLTNVKALQKSPGPQRERKSSSSSEDRNRMKTLGRRDSSDDWEIPDGQITVGQRIGSGSFGTVYKGKWHGDVAVKMLNVTAPTPQQLQAFKNEVGVLRKTRHVNILLFMGYSTKPQLAIVTQWCEGSSLYHHLHIIETKFEMIKLIDIARQTAQGMDYLHAKSIIHRDLKSNNIFLHEDLTVKIGDFGLATVKSRWSGSHQFEQLSGSILWMAPEVIRMQDKNPYSFQSDVYAFGIVLYELMTGQLPYSNINNRDQIIFMVGRGYLSPDLSKVRSNCPKAMKRLMAECLKKKRDERPLFPQILASIELLARSLPKIHRSASEPSLNRAGFQTEDFSLYACASPKTPIQAGGYGAFPVH mp2k1_human Dual specificity mitogen-activated protein kinase kinase 1 ERK activator kinase 1 MAPK/ERK kinase 1 MAP2K1 MEK1 PRKMK1 protein protein human Homo sapiens Human MPKKKPTPIQLNPAPDGSAVNGTSSAETNLEALQKKLEELELDEQQRKRLEAFLTQKQKVGELKDDDFEKISELGAGNGGVVFKVSHKPSGLVMARKLIHLEIKPAIRNQIIRELQVLHECNSPYIVGFYGAFYSDGEISICMEHMDGGSLDQVLKKAGRIPEQILGKVSIAVIKGLTYLREKHKIMHRDVKPSNILVNSRGEIKLCDFGVSGQLIDSMANSFVGTRSYMSPERLQGTHYSVQSDIWSMGLSLVEMAVGRYPIPPPDAKELELMFGCQVEGDAAETPPRPRTPGRPLSSYGMDSRPPMAIFELLDYIVNEPPPKLPSGVFSLEFQDFVNKCLIKNPAERADLKQLMVHAFIKRSDAEEVDFAGWLCSTIGLNQPSTPTHAAGV araf_human Serine/threonine-protein kinase A-Raf ARAF PKS PKS2 ARAF1 Proto-oncogene A-Raf-1 Proto-oncogene Pks Proto-oncogene A-Raf protein protein human Homo sapiens Human MEPPRGPPANGAEPSRAVGTVKVYLPNKQRTVVTVRDGMSVYDSLDKALKVRGLNQDCCVVYRLIKGRKTVTAWDTAIAPLDGEELIVEVLEDVPLTMHNFVRKTFFSLAFCDFCLKFLFHGFRCQTCGYKFHQHCSSKVPTVCVDMSTNRQQFYHSVQDLSGGSRQHEAPSNRPLNELLTPQGPSPRTQHCDPEHFPFPAPANAPLQRIRSTSTPNVHMVSTTAPMDSNLIQLTGQSFSTDAAGSRGGSDGTPRGSPSPASVSSGRKSPHSKSPAEQRERKSLADDKKKVKNLGYRDSGYYWEVPPSEVQLLKRIGTGSFGTVFRGRWHGDVAVKVLKVSQPTAEQAQAFKNEMQVLRKTRHVNILLFMGFMTRPGFAIITQWCEGSSLYHHLHVADTRFDMVQLIDVARQTAQGMDYLHAKNIIHRDLKSNNIFLHEGLTVKIGDFGLATVKTRWSGAQPLEQPSGSVLWMAAEVIRMQDPNPYSFQSDVYAYGVVLYELMTGSLPYSHIGCRDQIIFMVGRGYLSPDLSKISSNCPKAMRRLLSDCLKFQREERPLFPQILATIELLQRSLPKIERSASEPSLHRTQADELPACLLSAARLVP mp2k2_human Dual specificity mitogen-activated protein kinase kinase 2 MAP2K2 MEK2 MKK2 PRKMK2 ERK activator kinase 2 MAPK/ERK kinase 2 protein protein human Homo sapiens Human MLARRKPVLPALTINPTIAEGPSPTSEGASEANLVDLQKKLEELELDEQQKKRLEAFLTQKAKVGELKDDDFERISELGAGNGGVVTKVQHRPSGLIMARKLIHLEIKPAIRNQIIRELQVLHECNSPYIVGFYGAFYSDGEISICMEHMDGGSLDQVLKEAKRIPEEILGKVSIAVLRGLAYLREKHQIMHRDVKPSNILVNSRGEIKLCDFGVSGQLIDSMANSFVGTRSYMAPERLQGTHYSVQSDIWSMGLSLVELAVGRYPIPPPDAKELEAIFGRPVVDGEEGEPHSISPRPRPPGRPVSGHGMDSRPAMAIFELLDYIVNEPPPKLPNGVFTPDFQEFVNKCLIKNPAERADLKMLTNHTFIKRSEVEEVDFAGWLCKTLRLNQPGTPTRTAV p04049-1 RAF proto-oncogene serine/threonine-protein kinase 6C RAF1 RAF Proto-oncogene c-RAF Raf-1 protein protein human Homo sapiens Human MEHIQGAWKTISNGFGFKDAVFDGSSCISPTIVQQFGYQRRASDDGKLTDPSKTSNTIRVFLPNKQRTVVNVRNGMSLHDCLMKALKVRGLQPECCAVFRLLHEHKGKKARLDWNTDAASLIGEELQVDFLDHVPLTTHNFARKTFLKLAFCDICQKFLLNGFRCQTCGYKFHEHCSTKVPTMCVDWSNIRQLLLFPNSTIGDSGVPALPSLTMRRMRESVSRMPVSSQHRYSTPHAFTFNTSSPSSEGSLSQRQRSTSTPNVHMVSTTLPVDSRMIEDAIRSHSESASPSALSSSPNNLSPTGWSQPKTPVPAQRERAPVSGTQEKNKIRPRGQRDSSYYWEIEASEVMLSTRIGSGSFGTVYKGKWHGDVAVKILKVVDPTPEQFQAFRNEVAVLRKTRHVNILLFMGYMTKDNLAIVTQWCEGSSLYKHLHVQETKFQMFQLIDIARQTAQGMDYLHAKNIIHRDMKSNNIFLHEGLTVKIGDFGLATVKSRWSGSQQVEQPTGSVLWMAPEVIRMQDNNPFSFQSDVYSYGIVLYELMTGELPYSHINNRDQIIFMVGRGYASPDLSKLYKNCPKAMKRLVADCVKKVKEERPLFPQILSSIELLQHSLPKINRSASEPSLHRAAHTEDINACTLTTSPRLPVF ch5126766 CH5126766 small molecule small molecule chemical synthesis Chemical synthesis (Chemical synthesis) phosphoaminophosphonic acid-adenylate ester PHOSPHOAMINOPHOSPHONIC ACID-ADENYLATE ESTER small molecule small molecule chemical synthesis Chemical synthesis (Chemical synthesis) map2k1-araf-6 1546962 N/A 1546968 unspecified role unspecified role prey prey N/A 1546969 unspecified role unspecified role prey prey N/A 1546970 unspecified role unspecified role bait bait association association false false false 6D The MEK inhibitor PD0325901 Trametinib - a newer MEK inhibitor map2k1-braf-26 1546962 N/A 1546969 unspecified role unspecified role prey prey N/A 1546970 unspecified role unspecified role bait bait physical association physical association false false false 6D The MEK inhibitor PD0325901 Trametinib - a newer MEK inhibitor weak interaction 0 hours PD0325901 map2k1-araf-7 1546962 N/A 1546969 unspecified role unspecified role prey prey N/A 1546970 unspecified role unspecified role bait bait N/A 1546968 unspecified role unspecified role prey prey association association false false false 6D The MEK inhibitor PD0325901 Trametinib - a newer MEK inhibitor weak interaction Time of Trametinib treatment (hours)-3 map2k1-braf-27 1546962 N/A 1546970 unspecified role unspecified role bait bait N/A 1546968 unspecified role unspecified role prey prey physical association physical association false false false 6D The MEK inhibitor PD0325901 Trametinib - a newer MEK inhibitor weak interaction 48 hours Trametinib map2k1-araf-4 1546963 n/a 1546971 unspecified role unspecified role prey prey n/a 1546968 unspecified role unspecified role prey prey n/a 1546969 unspecified role unspecified role prey prey n/a 1546970 unspecified role unspecified role bait bait region flag tag flag tag FLAG-tagged FLAG DYKDDDDKV epitope tag DYKDDDDKV epitope tag FLAG FLAG-tagged undetermined undetermined sequence position undetermined undetermined undetermined sequence position undetermined false Q02750:p.[Ser218Glu;Ser222Glu] mutation decreasing mutation decreasing interaction hotspot hotspot mutation decreasing certain certain sequence position certain certain certain sequence position certain false certain certain sequence position certain certain certain sequence position certain false Q02750:p.[Met308Ala;Ile310Ala] mutation disrupting mutation disrupting interaction mutation disrupting 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 4 A/D, 6A, 7G, Supp figure 4 A MEK inhibitors: Selumetinib and PD0325901 CH5126766 - a newer MEK inhibitor. Trametinib -a newer MEK inhibitor, disrupts the interaction of MEK1 with ARAF and CRAF. In cells co-transfected with K-RAS G12V this interaction was completely disrupted, but was only partially reduced in the presence of CH5126766 or PD0325901. Co-transfection of BRAF V600E reduced this interaction,and the presence of CH5126766 or PD0325901 further reduced the interaction , but in the presence of CH5126766 the interaction was increased. map2k2-araf-4 1546963 n/a 1546971 unspecified role unspecified role prey prey n/a 1546972 unspecified role unspecified role bait bait region flag tag flag tag FLAG-tagged FLAG DYKDDDDKV epitope tag DYKDDDDKV epitope tag FLAG FLAG-tagged undetermined undetermined sequence position undetermined undetermined undetermined sequence position undetermined false n/a 1546969 unspecified role unspecified role prey prey n/a 1546968 unspecified role unspecified role prey prey association association false false false CH5126766 - a newer MEK inhibitor. (PD0325901) K-RAS G12V transfection:- this interaction was completely disrupted, but was only partially reduced in the presence of CH5126766 or PD0325901. BRAF V600E transfection: MEK2 interacted with ARAF, BRAF and CRAF, but no untransfected control result is shown. The presence of PD0325901 did not appear to affect this interaction. In the presence of Trametinib the interaction of MEK2 and BRAF still took place at a reduced level. It is not clear if any of these interactions were with endogenous wild-type BRAF, or the V600E mutant, or both. Supp fig 4B Trametinib -a newer MEK inhibitor, disrupts the interaction of MEK2 with ARAF and CRAF. braf-map2k1-1 1546964 n/a 1546970 unspecified role unspecified role unspecified role unspecified role 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 n/a 1546969 unspecified role unspecified role unspecified role unspecified role n_terminus gst tag glutathione s tranferase tag glutathione S-tranferase tag glutathione S-tranferase tag gst tag n-term range n-terminal range n-term range n-term range n-terminal range n-term range false kinase_domain sufficient to bind sufficient binding region sufficient to bind certain certain sequence position certain certain certain sequence position certain false direct interaction direct interaction false false false 6 E/F Trametinib is a MEK inhibitor that reduces the affinity of MEK and RAF. raf1-map2k1-2 1546964 n/a 1546970 unspecified role unspecified role prey prey 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 n/a 1546973 unspecified role unspecified role bait bait n_terminus gst tag glutathione s tranferase tag glutathione S-tranferase tag glutathione S-tranferase tag gst tag n-term range n-terminal range n-term range n-term range n-terminal range n-term range false kinase_domain sufficient to bind sufficient binding region sufficient to bind certain certain sequence position certain certain certain sequence position certain false P04049-1:p.Tyr340_Tyr341delinsAspAsp mutation mutation certain certain sequence position certain certain certain sequence position certain false direct interaction direct interaction false false false 6 E/F Trametinib is a MEK inhibitor that reduces the affinity of MEK and RAF. map2k1-araf-1 1546965 n/a 1546970 unspecified role unspecified role bait bait n/a 1546969 unspecified role unspecified role prey prey n/a 1546971 unspecified role unspecified role prey prey n/a 1546968 unspecified role unspecified role prey prey association association false false false 4 B/C, 6B The MEK inhibitor PD0325901 reduced the level of MEK1 in complex with BRAF and ARAF. map2k1-araf-2 1546966 n/a 1546969 unspecified role unspecified role prey prey n/a 1546970 unspecified role unspecified role bait bait n/a 1546968 unspecified role unspecified role prey prey n/a 1546971 unspecified role unspecified role prey prey association association false false false 4 C, 6C The MEK inhibitor PD0325901 Trametinib - a newer MEK inhibitor ch5126766-map2k1-1 1546967 n/a 1546974 allosteric effector allosteric effector neutral component neutral component chemical synthesis Chemical synthesis (Chemical synthesis) n/a 1546975 unspecified role unspecified role neutral component neutral component chemical synthesis Chemical synthesis (Chemical synthesis) n/a 1546970 unspecified role unspecified role neutral component neutral component 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 regions 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) direct interaction direct interaction false false false Table S1 2.7 A R-factor: working - 21.7%, Free R-factor: 25.0%,