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/ Cell death in leukemia: passenger protein regulation by topoisomerase inhibitors. Cell death in leukemia: passenger protein regulation by topoisomerase inhibitors. Biochem. Biophys. Res. Commun. (0006-291X) 2007 imex curation Jahnke U., Higginbottom K., Newland AC., Cotter FE., Allen PD. Apoptosis - Interactions involving proteins with a function related to apoptosis Cancer - Interactions investigated in the context of cancer p.d.allen@qmul.ac.uk 12-Oct-2007: Contacted by JYOTI. Only protein-protein interactions human-k_562 Homo sapiens myeloid leukemia cells. k562 Myelogenous leukemia K562 cells fluorescence imaging fluorescence microscopy fluorescence imaging predetermined predetermined participant predetermined Jahnke U., Higginbottom K., Newland AC., Cotter FE., Allen PD. Apoptosis - Interactions involving proteins with a function related to apoptosis p.d.allen@qmul.ac.uk Accepted 2007-OCT-25 by JYOTI. Cell cycle Biochem. Biophys. Res. Commun. (0006-291X) 2007 Only protein-protein interactions imex curation Cell death in leukemia: passenger protein regulation by topoisomerase inhibitors. Cell death in leukemia: passenger protein regulation by topoisomerase inhibitors. Biochem. Biophys. Res. Commun. (0006-291X) 2007 imex curation Jahnke U., Higginbottom K., Newland AC., Cotter FE., Allen PD. Apoptosis - Interactions involving proteins with a function related to apoptosis Cancer - Interactions investigated in the context of cancer p.d.allen@qmul.ac.uk 12-Oct-2007: Contacted by JYOTI. Only protein-protein interactions human-k_562 Homo sapiens myeloid leukemia cells. k562 Myelogenous leukemia K562 cells anti bait coip anti bait coimmunoprecipitation anti bait coip western blot western blot Immuno blot Immuno blot Jahnke U., Higginbottom K., Newland AC., Cotter FE., Allen PD. Apoptosis - Interactions involving proteins with a function related to apoptosis Add GO term for mitotic catastrophe (requested) when available to interactions. K562 cells were untreated (control) and treated with 1 microg/ml etoposide and 20 mg/ml etoposide for 24 h and 48 h or only for 48 h. p.d.allen@qmul.ac.uk Accepted 2007-OCT-25 by JYOTI. 12-Oct-2007: Contacted by JYOTI. Biochem. Biophys. Res. Commun. (0006-291X) 2007 Only protein-protein interactions imex curation Cell death in leukemia: passenger protein regulation by topoisomerase inhibitors. Cell death in leukemia: passenger protein regulation by topoisomerase inhibitors. Biochem. Biophys. Res. Commun. (0006-291X) 2007 imex curation Jahnke U., Higginbottom K., Newland AC., Cotter FE., Allen PD. Apoptosis - Interactions involving proteins with a function related to apoptosis Cancer - Interactions investigated in the context of cancer p.d.allen@qmul.ac.uk 12-Oct-2007: Contacted by JYOTI. Only protein-protein interactions human-k_562 Homo sapiens myeloid leukemia cells. k562 Myelogenous leukemia K562 cells fluorescence imaging fluorescence microscopy fluorescence imaging predetermined predetermined participant predetermined Jahnke U., Higginbottom K., Newland AC., Cotter FE., Allen PD. Apoptosis - Interactions involving proteins with a function related to apoptosis K562 cells were treated with etoposide. p.d.allen@qmul.ac.uk Accepted 2007-OCT-25 by JYOTI. 12-Oct-2007: Contacted by JYOTI. Biochem. Biophys. Res. Commun. (0006-291X) 2007 Only protein-protein interactions imex curation ince_human Inner centromere protein INCENP protein protein human Homo sapiens Human MGTTAPGPIHLLELCDQKLMEFLCNMDNKDLVWLEEIQEEAERMFTREFSKEPELMPKTPSQKNRRKKRRISYVQDENRDPIRRRLSRRKSRSSQLSSRRLRSKDSVEKLATVVGENGSVLRRVTRAAAAAAAATMALAAPSSPTPESPTMLTKKPEDNHTQCQLVPVVEIGISERQNAEQHVTQLMSTEPLPRTLSPTPASATAPTSQGIPTSDEESTPKKSKARILESITVSSLMATPQDPKGQGVGTGRSASKLRIAQVSPGPRDSPAFPDSPWRERVLAPILPDNFSTPTGSRTDSQSVRHSPIAPSSPSPQVLAQKYSLVAKQESVVRRASRRLAKKTAEEPAASGRIICHSYLERLLNVEVPQKVGSEQKEPPEEAEPVAAAEPEVPENNGNNSWPHNDTEIANSTPNPKPAASSPETPSAGQQEAKTDQADGPREPPQSARRKRSYKQAVSELDEEQHLEDEELQPPRSKTPSSPCPASKVVRPLRTFLHTVQRNQMLMTPTSAPRSVMKSFIKRNTPLRMDPKCSFVEKERQRLENLRRKEEAEQLRRQKVEEDKRRRLEEVKLKREERLRKVLQARERVEQMKEEKKKQIEQKFAQIDEKTEKAKEERLAEEKAKKKAAAKKMEEVEARRKQEEEARRLRWLQQEEEERRHQELLQKKKEEEQERLRKAAEAKRLAEQREQERREQERREQERREQERREQERREQERQLAEQERRREQERLQAERELQEREKALRLQKEQLQRELEEKKKKEEQQRLAERQLQEEQEKKAKEAAGASKALNVTVDVQSPACTSYQMTPQGHRAPPKINPDNYGMDLNSDDSTDDEAHPRKPIPTWARGTPLSQAIIHQYYHPPNLLELFGTILPLDLEDIFKKSKPRYHKRTSSAVWNSPPLQGARVPSSLAYSLKKH DIP protein Q9NQS7 original sequence version: 83 DIP protein Q9NQS7 original sequence version: 86 644D081DCC9035E8 aurkb_human Aurora kinase B AIRK2 Aurora- and IPL1-like midbody-associated protein 1 Aurora/IPL1-related kinase 2 STK-1 Serine/threonine-protein kinase 12 Serine/threonine-protein kinase 5 STK1 STK5 Aurora 1 AURKB AIK2 AIM1 ARK2 STK12 Serine/threonine-protein kinase aurora-B protein protein human Homo sapiens Human MAQKENSYPWPYGRQTAPSGLSTLPQRVLRKEPVTPSALVLMSRSNVQPTAAPGQKVMENSSGTPDILTRHFTIDDFEIGRPLGKGKFGNVYLAREKKSHFIVALKVLFKSQIEKEGVEHQLRREIEIQAHLHHPNILRLYNYFYDRRRIYLILEYAPRGELYKELQKSCTFDEQRTATIMEELADALMYCHGKKVIHRDIKPENLLLGLKGELKIADFGWSVHAPSLRRKTMCGTLDYLPPEMIEGRMHNEKVDLWCIGVLCYELLVGNPPFESASHNETYRRIVKVDLKFPASVPMGAQDLISKLLRHNPSERLPLAQVSAHPWVRANSRRVLPPSALQSVA DIP protein Q96GD4 original sequence version: 88 DIP protein Q96GD4 original sequence version: 113 DIP protein Q96GD4 original sequence version: 116 DIP protein Q96GD4 original sequence version: 122 DIP protein Q96GD4 original sequence version: 177 DIP protein Q96GD4-1 original sequence version: 177 A5ED13EF5A1FAFBF cdk1_human Cyclin-dependent kinase 1 p34 protein kinase Cell division protein kinase 1 CDK1 CDC2 Cell division control protein 2 homolog CDC28A CDKN1 P34CDC2 protein protein human Homo sapiens Human MEDYTKIEKIGEGTYGVVYKGRHKTTGQVVAMKKIRLESEEEGVPSTAIREISLLKELRHPNIVSLQDVLMQDSRLYLIFEFLSMDLKKYLDSIPPGQYMDSSLVKSYLYQILQGIVFCHSRRVLHRDLKPQNLLIDDKGTIKLADFGLARAFGIPIRVYTHEVVTLWYRSPEVLLGSARYSTPVDIWSIGTIFAELATKKPLFHGDSEIDQLFRIFRALGTPNNEVWPEVESLQDYKNTFPKWKPGSLASHVKNLDENGLDLLSKMLIYDPAKRISGKMALNHPYFNDLDNQIKKM DIP protein P06493 original sequence version: 185 DIP protein P06493 original sequence version: 162 DIP protein P06493 original sequence version: 184 DIP protein P06493 original sequence version: 97 DIP protein P06493 original sequence version: 192 942D79448EFE490A birc5_human Baculoviral IAP repeat-containing protein 5 Apoptosis inhibitor survivin Apoptosis inhibitor 4 BIRC5 API4 IAP4 protein protein human Homo sapiens Human MGAPTLPPAWQPFLKDHRISTFKNWPFLEGCACTPERMAEAGFIHCPTENEPDLAQCFFCFKELEGWEPDDDPIEEHKKHSSGCAFLSVKKQFEELTLGEFLKLDRERAKNKIAKETNNKKKEFEETAKKVRRAIEQLAAMD DIP protein O15392 original sequence version: 129 DIP protein O15392 original sequence version: 132 DIP protein O15392-1 original sequence version: 141 DIP protein O15392-1 original sequence version: 143 9E7CADCDF2822286 aurkb-cdc2 1 5 unspecified role unspecified role neutral component neutral component 6 unspecified role unspecified role neutral component neutral component colocalization colocalization 3 A i In control K562 cells, Cdk1 was seen to co-localise with Aurora B (Fig. 3A) and survivin (Supplementary data) to the chromosomal centromeres during prophase (Fig. 3A i) and metaphase (Fig. 3A ii), to the midzone in anaphase (Fig. 3A iii) and to midbodies in telophase (Fig. 3A iv). Prophase aurkb-cdc2-1 1 5 unspecified role unspecified role neutral component neutral component 6 unspecified role unspecified role neutral component neutral component colocalization colocalization 3 A ii In control K562 cells, Cdk1 was seen to co-localise with Aurora B (Fig. 3A) and survivin (Supplementary data) to the chromosomal centromeres during prophase (Fig. 3A i) and metaphase (Fig. 3A ii), to the midzone in anaphase (Fig. 3A iii) and to midbodies in telophase (Fig. 3A iv). Metaphase aurkb-cdc2-2 1 6 unspecified role unspecified role neutral component neutral component 5 unspecified role unspecified role neutral component neutral component colocalization colocalization 3 A iii In control K562 cells, Cdk1 was seen to co-localise with Aurora B (Fig. 3A) and survivin (Supplementary data) to the chromosomal centromeres during prophase (Fig. 3A i) and metaphase (Fig. 3A ii), to the midzone in anaphase (Fig. 3A iii) and to midbodies in telophase (Fig. 3A iv). Anaphase birc5-cdc2-3 1 7 unspecified role unspecified role neutral component neutral component 6 unspecified role unspecified role neutral component neutral component colocalization colocalization S 3 In control K562 cells, Cdk1 was seen to co-localise with Aurora B (Fig. 3A) and survivin (Supplementary data) to the chromosomal centromeres during prophase (Fig. 3A i) and metaphase (Fig. 3A ii), to the midzone in anaphase (Fig. 3A iii) and to midbodies in telophase (Fig. 3A iv). Telophase birc5-cdc2 1 6 unspecified role unspecified role neutral component neutral component 7 unspecified role unspecified role neutral component neutral component colocalization colocalization S 3 In control K562 cells, Cdk1 was seen to co-localise with Aurora B (Fig. 3A) and survivin (Supplementary data) to the chromosomal centromeres during prophase (Fig. 3A i) and metaphase (Fig. 3A ii), to the midzone in anaphase (Fig. 3A iii) and to midbodies in telophase (Fig. 3A iv). Prophase birc5-cdc2-1 1 7 unspecified role unspecified role neutral component neutral component 6 unspecified role unspecified role neutral component neutral component colocalization colocalization S 3 In control K562 cells, Cdk1 was seen to co-localise with Aurora B (Fig. 3A) and survivin (Supplementary data) to the chromosomal centromeres during prophase (Fig. 3A i) and metaphase (Fig. 3A ii), to the midzone in anaphase (Fig. 3A iii) and to midbodies in telophase (Fig. 3A iv). Metaphase birc5-cdc2-2 1 7 unspecified role unspecified role neutral component neutral component 6 unspecified role unspecified role neutral component neutral component colocalization colocalization S 3 In control K562 cells, Cdk1 was seen to co-localise with Aurora B (Fig. 3A) and survivin (Supplementary data) to the chromosomal centromeres during prophase (Fig. 3A i) and metaphase (Fig. 3A ii), to the midzone in anaphase (Fig. 3A iii) and to midbodies in telophase (Fig. 3A iv). Anaphase aurkb-cdc2-3 3 5 unspecified role unspecified role neutral component neutral component 6 unspecified role unspecified role neutral component neutral component colocalization colocalization 3 B i and ii Aurora B (Fig. 3B i and ii) and survivin (Fig. 3B iii) were again localised to the centromeres on chromosomes. Cdk1 co-localised with Aurora B and survivin at centromeres. birc5-cdc2-4 3 7 unspecified role unspecified role neutral component neutral component 6 unspecified role unspecified role neutral component neutral component colocalization colocalization 3 B iii Aurora B (Fig. 3B i and ii) and survivin (Fig. 3B iii) were again localised to the centromeres on chromosomes. Cdk1 co-localised with Aurora B and survivin at centromeres. cdc2-aurkb 2 6 unspecified role unspecified role bait bait 5 unspecified role unspecified role prey prey physical association physical association 4 A c Immunoprecipitating Cdk1 and staining for Aurora B and survivin revealed a similar pattern to whole cell lysates. The amount of Aurora B (Fig. 4A c) and survivin (Fig. 4A d) binding to Cdk1 increased in cells arrested at the G2 checkpoint both at 24 (lane 2) and 48 h (lane 5) compared to controls (lanes 1 and 4). Etoposide - Treatment with etoposide increased the amount of Aurora kinase B coimmunoprecipitated with Cdk1. The level of Aurora kinase B coimmunoprecipitated with Cdk1 was increased in cells arrested at G2 and cell that undergo mitotic catastrophe. cdc2-birc5 2 6 unspecified role unspecified role bait bait 7 unspecified role unspecified role prey prey physical association physical association 4 A d Immunoprecipitating Cdk1 and staining for Aurora B and survivin revealed a similar pattern to whole cell lysates. The amount of Aurora B (Fig. 4A c) and survivin (Fig. 4A d) binding to Cdk1 increased in cells arrested at the G2 checkpoint both at 24 (lane 2) and 48 h (lane 5) compared to controls (lanes 1 and 4). Etoposide - Treatment with etoposide increased the amount of Survivin coimmunoprecipitated with Cdk1. The level of Survivin coimmunoprecipitated with Cdk1 was increased in cells arrested at G2 and cell that undergo mitotic catastrophe. cdc2-incenp 2 6 unspecified role unspecified role bait bait 4 unspecified role unspecified role prey prey physical association physical association Etoposide - the level of INCENP bound to Cdk1 increased with etoposide treatment. 4 A e The level of INCENP coimmunoprecipitated with Cdk1 was increased in cells arrested at G2 and cell that undergo mitotic catastrophe. aurkb-birc5 2 7 unspecified role unspecified role prey prey 5 unspecified role unspecified role bait bait physical association physical association 4 A f When Aurora B was immunoprecipitated it was demonstrated that survivin bound to Aurora B in control cells (Fig. 4A f lane 4). Survivin binding to Aurora B was increased in G2 cell cycle arrested cells (lane 5) and cells induced to die post genotoxic insult (lane 6). Etoposide - Treatment with etoposide increases the amount of Survivin coimmunoprecipitated with Aurora kinase B. Survivin binding to Aurora kinase B was increased in G2 cell cycle arrested cells and cells induced to die by DNA damage.