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/ Interaction of Cdc2 and Cdc18 with a fission yeast ORC2-like protein. Interaction of Cdc2 and Cdc18 with a fission yeast ORC2-like protein. Nature (0028-0836) 1996 imex curation Leatherwood J., Lopez-Girona A., Russell P. jyoti Only protein-protein interactions yeasx Saccharomyces cerevisiae (Baker's yeast) Baker's yeast 2 hybrid two hybrid 2H 2h Gal4 transcription regeneration two-hybrid yeast two hybrid 2-hybrid classical two hybrid classical two hybrid 2H two-hybrid 2-hybrid Gal4 transcription regeneration 2h yeast two hybrid Y2H Y-2H 2 hybrid predetermined predetermined participant predetermined Leatherwood J., Lopez-Girona A., Russell P. Accepted 2008-DEC-7 by JYOTI. Nature (0028-0836) 1996 jyoti Only protein-protein interactions imex curation Interaction of Cdc2 and Cdc18 with a fission yeast ORC2-like protein. Interaction of Cdc2 and Cdc18 with a fission yeast ORC2-like protein. Nature (0028-0836) 1996 imex curation Leatherwood J., Lopez-Girona A., Russell P. jyoti Only protein-protein interactions schpm Schizosaccharomyces pombe (fission yeast) Fission yeast pull down pull down western blot western blot Immuno blot Immuno blot Leatherwood J., Lopez-Girona A., Russell P. Accepted 2008-DEC-7 by JYOTI. Nature (0028-0836) 1996 jyoti Only protein-protein interactions imex curation cdc18_schpo Cell division control protein 18 cdc18 SPBC14C8.07c protein protein schpo Schizosaccharomyces pombe (strain ATCC 38366 / 972) Fission yeast MCETPIGCHTPRRCNRFIDSAALIDCTNKTNQREHSPSFSIEIPTTPSRKRTLASSHFQTPTKRIKYELGELQEEKTDLYPNFPAQLKENKKPKLPTTPQTPKTPKRTIQIVTPKSLNRTCNPVPFATRLLQSTPHRQLFPPTPSTPSTPSYNSTAKLSLRKSYRSAGVVGRENEKSIVESFFRQHLDANAGGALYVSGAPGTGKTVLLHNVLDHVVSDYPKVNVCYINCMTINEPKAIFEKIHSKIVKEEILENEDHHINFQCELESHFTQSANELYNPVIIVLDEMDHLIAREQQVLYTLFEWPSRPTSRLILVGIANALDMTDRFLPRLRTKHITPKLLSFTPYTAQEISTIIKARLKTAATTSEKNNPFTPIKSISEVSDDSINVVSQHADETPFIHPAAIELCARKVAASSGDLRKALDICRHAIELAEREWKAQHDNTLSSVDIPRASIAHVVRATSAMSQSASARLKNLGLQQKAILCTLVVCEKTSLSVADVFEKYSSLCLRDRLIYPLTSSEFCDVANSLETLAIIRLRTKQRNGKPQDRIISLLVPEMDVITAVGDIGTLKRFFDRR 74DBE9D5CF23504D 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 orc2_schpo Origin recognition complex subunit 2 orc2 orp2 SPBC685.09 protein protein schpo Schizosaccharomyces pombe (strain ATCC 38366 / 972) Fission yeast MLINEHVRERYHFLRVSLTRVAHLVFANSHESNDLKMVENVSSTPKKGVLEDPSTLTPEVVTPRTPGHRIIKAKGAYTKDRSAKRRRGRIEIERHLLGEFDDNVGSNSLLDVPLYSLEAEPLSPSVMLEDESMEGINQSPQGISVEKLGKEDNRSRSSTPASPSLESHEFSESREAGLSQSNGFEARSHGTGFDEYFDKFSQRKTSSNTLSQLPVLDNQVYLDTVKEICTETEQHTQTLVHFQSRNFHQWYFELVNNFNLLFYGFGSKEHFLSSFVEKKLPCFPIFVVKGYFPQLQLKNVLSSFLEFLEVTPAASVSDMLIQSLSIINSPSFSLGKIVFLVHNIDGESLIDERFQSALAAIASSKNVYFIASVDHVNFALLWDTSLESSFNFVMHDATTFARYYNETTYENSLGIGRANVSNKEKAIKHVLYSLPANSRGIFKFLLIHQLERMMDTQDFDARQGEKVGIEYRSFFQKCSAEFLCSNEPNFRSQLTEFFDHNIIESKRDVSNSEILWVPYPKNLLEILLEDMMEDV 7B04A316D94BAF88 cdc2-orc2 1 4 unspecified role unspecified role bait bait n-terminus required to bind necessary binding region required to bind deletion analysis deletion analysis certain certain sequence position certain certain certain sequence position certain 5 unspecified role unspecified role prey prey physical association physical association Figure 1. Potential targets of Cdc2 kinase in the fission yeast Schizosaccharomyces pombe were identified using two-hybrid screen in the budding yeast S. cerevisiae. A screen of 2 x 10-7 fission yeast cDNA clones yielded 112 positives representing seven genes. One, orp2+ (ORC2 related in pombe), encodes a protein which is 22% identical to the Orc2 subunit of S. cerevisiae ORC (Fig. 1). A Y2H also showed interaction between orc2 and cdc18 but the data was not shown in the paper. cdc2-orc2-1 2 4 unspecified role unspecified role prey prey schpm Schizosaccharomyces pombe (fission yeast) Fission yeast 5 unspecified role unspecified role bait bait region gst tag glutathione s tranferase tag glutathione S-tranferase tag glutathione S-tranferase tag gst tag anti tag western anti tag western blot anti tag western undetermined undetermined sequence position undetermined undetermined undetermined sequence position undetermined schpm Schizosaccharomyces pombe (fission yeast) Fission yeast physical association physical association Figure 2. To determine whether Cdc2 and Orp2 interact in S. pombe, functional GST-Orp2 fusion protein and a GST control were expressed in S. pombe and isolated by binding to GSH-Sepharose. Cdc2 was detected in association with the GST-Orp2 fusion but not with GST (Fig. 2a). An interaction between orc2 and cdc2 was detected. cdc18-orc2 2 3 unspecified role unspecified role prey prey region ha tag ha tag YPYDVPDYA epitope tag YPYDVPDYA epitope tag anti tag western anti tag western blot anti tag western undetermined undetermined sequence position undetermined undetermined undetermined sequence position undetermined schpm Schizosaccharomyces pombe (fission yeast) Fission yeast 5 unspecified role unspecified role bait bait region gst tag glutathione s tranferase tag glutathione S-tranferase tag glutathione S-tranferase tag gst tag anti tag western anti tag western blot anti tag western undetermined undetermined sequence position undetermined undetermined undetermined sequence position undetermined schpm Schizosaccharomyces pombe (fission yeast) Fission yeast physical association physical association Figure 4c. and was confirmed by finding that Cdc18 coprecipitated with GST-Orp2 in fission yeast lysates (Fig. 4c). An interaction between orc2 and cdc18 was detected.