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/ p25rum1 promotes proteolysis of the mitotic B-cyclin p56cdc13 during G1 of the fission yeast cell cycle. p25rum1 promotes proteolysis of the mitotic B-cyclin p56cdc13 during G1 of the fission yeast cell cycle. EMBO J. (0261-4189) 1997 Correa-Bordes J., Gulli MP., Nurse P. nurse@icrf.icnet.uk 26-APR-2007: Contacted by JYOTI. orchard in vitro In vitro protein kinase assay protein kinase assay predetermined predetermined participant predetermined Correa-Bordes J., Gulli MP., Nurse P. nurse@icrf.icnet.uk Accepted 2007-APR-30 by ORCHARD. 26-APR-2007: Contacted by JYOTI. EMBO J. (0261-4189) 1997 orchard p25rum1 promotes proteolysis of the mitotic B-cyclin p56cdc13 during G1 of the fission yeast cell cycle. p25rum1 promotes proteolysis of the mitotic B-cyclin p56cdc13 during G1 of the fission yeast cell cycle. EMBO J. (0261-4189) 1997 Correa-Bordes J., Gulli MP., Nurse P. nurse@icrf.icnet.uk 26-APR-2007: Contacted by JYOTI. orchard in vitro In vitro anti bait coip anti bait coimmunoprecipitation anti bait coip predetermined predetermined participant predetermined Correa-Bordes J., Gulli MP., Nurse P. nurse@icrf.icnet.uk Accepted 2007-APR-30 by ORCHARD. 26-APR-2007: Contacted by JYOTI. EMBO J. (0261-4189) 1997 orchard cg23_schpo G2/mitotic-specific cyclin cdc13 cdc13 SPBC582.03 protein protein schpo Schizosaccharomyces pombe (strain ATCC 38366 / 972) Fission yeast MTTRRLTRQHLLANTLGNNDENHPSNHIARAKSSLHSSENSLVNGKKATVSSTNVPKKRHALDDVSNFHNKEGVPLASKNTNVRHTTASVSTRRALEEKSIIPATDDEPASKKRRQPSVFNSSVPSLPQHLSTKSHSVSTHGVDAFHKDQATIPKKLKKDVDERVVSKDIPKLHRDSVESPESQDWDDLDAEDWADPLMVSEYVVDIFEYLNELEIETMPSPTYMDRQKELAWKMRGILTDWLIEVHSRFRLLPETLFLAVNIIDRFLSLRVCSLNKLQLVGIAALFIASKYEEVMCPSVQNFVYMADGGYDEEEILQAERYILRVLEFNLAYPNPMNFLRRISKADFYDIQTRTVAKYLVEIGLLDHKLLPYPPSQQCAAAMYLAREMLGRGPWNRNLVHYSGYEEYQLISVVKKMINYLQKPVQHEAFFKKYASKKFMKASLFVRDWIKKNSIPLGDDADEDYTFHKQKRIQHDMKDEEW B2C267CF57F5FD36 B2C267CF57F5FD36 cdk1_schpo Cyclin-dependent kinase 1 cdc2 swo2 SPBC11B10.09 pi002 p34 protein kinase cdk1 Cell division control protein 2 Cell division protein kinase 1 protein protein schpo Schizosaccharomyces pombe (strain ATCC 38366 / 972) Fission yeast MENYQKVEKIGEGTYGVVYKARHKLSGRIVAMKKIRLEDESEGVPSTAIREISLLKEVNDENNRSNCVRLLDILHAESKLYLVFEFLDMDLKKYMDRISETGATSLDPRLVQKFTYQLVNGVNFCHSRRIIHRDLKPQNLLIDKEGNLKLADFGLARSFGVPLRNYTHEIVTLWYRAPEVLLGSRHYSTGVDIWSVGCIFAEMIRRSPLFPGDSEIDEIFKIFQVLGTPNEEVWPGVTLLQDYKSTFPRWKRMDLHKVVPNGEEDAIELLSAMLVYDPAHRISAKRALQQNYLRDFH D04EF14BA1492875 rum1_schpo Cyclin-dependent kinase inhibitor rum1 rum1 SPBC32F12.09 p25-rum1 protein protein schpo Schizosaccharomyces pombe (strain ATCC 38366 / 972) Fission yeast MEPSTPPMRGLCTPSTPESPGSFKGVIDASLEGNSSIMIDEIPESDLPAPQVSTFPPTPAKTPKKQLLPNLMLQDRSNSLERCMEEDREHNPFLSSSDNQLLSRKKRKPTPPPSDGLYYVFRGKRIKKSFRPGTDLSTFKPKLLFADSAPSSSSDNPTSSVDLNDYSQIGILPPNLNSIGNKMFSLKSRVPSSSSGSFVAPPPQMRLPAYSSPQKSRSNTKDENRHNLLR C5199FE345F7484A cdc13-rum1-3 2 5 unspecified role unspecified role bait bait schpm Schizosaccharomyces pombe (fission yeast) Fission yeast 3 unspecified role unspecified role prey prey region 35s radiolabel 35s radiolabel 35S s35 radiolabelled S35 s35 radiolabelled 35S S35 undetermined undetermined sequence position undetermined undetermined undetermined sequence position undetermined schpm Schizosaccharomyces pombe (fission yeast) Fission yeast 4 unspecified role unspecified role ancillary ancillary region 35s radiolabel 35s radiolabel 35S s35 radiolabelled S35 s35 radiolabelled 35S S35 undetermined undetermined sequence position undetermined undetermined undetermined sequence position undetermined schpm Schizosaccharomyces pombe (fission yeast) Fission yeast association association Figure 4. We first tested whether p25rum1 could bind either p56cdc13 alone or p56cdc13 complexed with p34cdc2. Shows that rum1 can specifically associate with cdc13 alone or complexed with cdc2 in vitro. cdc2-rum1-1 1 5 enzyme target enzyme target substrate substrate unspecified role unspecified role schpm Schizosaccharomyces pombe (fission yeast) Fission yeast 4 enzyme enzyme unspecified role unspecified role schpm Schizosaccharomyces pombe (fission yeast) Fission yeast physical association physical association Figure 8. p25rum1 kinase activity was detected with both the anti-p34cdc2 and anti-cig1p antibodies, but not with the other two (Figure 8). This result indicates that p25rum1 can be phosphorylated in vitro by the p34cdc2 protein kinase activity associated with the cig1-encoded B-cyclin, but not with the other two B-cyclins. Rum1 kinase activity was detected with both anti-cdc2 and anti-cig1 antibodies. However, cig1 had been immunoprecipitated using cdc2, the authors state that the kinase activity it due to the formation a cig/cdc2 complex which phosphorylates rum1. Phosphorylation of Rum1 cdc13-rum1-4 2 5 unspecified role unspecified role bait bait schpm Schizosaccharomyces pombe (fission yeast) Fission yeast 3 unspecified role unspecified role prey prey region 35s radiolabel 35s radiolabel 35S s35 radiolabelled S35 s35 radiolabelled 35S S35 undetermined undetermined sequence position undetermined undetermined undetermined sequence position undetermined schpm Schizosaccharomyces pombe (fission yeast) Fission yeast physical association physical association Figure 4. It can be seen that p25rum1 could form specific complexes with p56cdc13 and with p34cdc2/p56cdc13 complexes but not with p34cdc2 alone. We next tested whether the destruction box which is required for B-cyclin proteolysis in mitosis was required for p25rum1 binding. The cdc13D90 mutant lacks this destruction box, but the mutant protein could still form an in vitro complex with p25rum1 (Figure 4). Shows that rum1 can specifically associate with cdc13 alone or complexed with cdc2 in vitro. Also deletion of the destruction box in the cdc13-delta-90 mutant did not inhibit its binding with rum1.