DIP Database of Interacting Proteins 611 Young Drive East; Los Angeles CA 90095; USA http://dip.doe-mbi.ucla.edu/ dip@mbi.ucla.edu 611 Young Drive East; Los Angeles CA 90095; USA http://dip.doe-mbi.ucla.edu/ shu-2007-4 Functional characterization of human PFTK1 as a cyclin-dependent kinase. human-293t Homo sapiens 293 cells transformed with SV40 large T antigen 293t 293 cells expressing SV40 large T antigen. The original designation of this cell line was 293tsA1609neo. It is not in Cabri,ATCC or HyperCLDB. anti tag coip anti tag coimmunoprecipitation anti tag coip western blot western blot Immuno blot Immuno blot Shu F., Lv S., Qin Y., Ma X., Wang X., Peng X., Luo Y., Xu BE., Sun X., Wu J. junwu@shanghaigenomics.com 3 human cDNA libraries - (heLa, human T/B lymphocyte and fetal brain. Proc. Natl. Acad. Sci. U.S.A. (0027-8424) 2007 IMEx Only protein-protein interactions imex curation shu-2007-1 Functional characterization of human PFTK1 as a cyclin-dependent kinase. 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 nucleotide sequence nucleotide sequence identification sequence cloning sequence cloning nucleotide sequence Shu F., Lv S., Qin Y., Ma X., Wang X., Peng X., Luo Y., Xu BE., Sun X., Wu J. junwu@shanghaigenomics.com 3 human cDNA libraries - (heLa, human T/B lymphocyte and fetal brain. Proc. Natl. Acad. Sci. U.S.A. (0027-8424) 2007 IMEx Only protein-protein interactions imex curation shu-2007-2 Functional characterization of human PFTK1 as a cyclin-dependent kinase. 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 Shu F., Lv S., Qin Y., Ma X., Wang X., Peng X., Luo Y., Xu BE., Sun X., Wu J. junwu@shanghaigenomics.com 3 human cDNA libraries - (heLa, human T/B lymphocyte and fetal brain. Proc. Natl. Acad. Sci. U.S.A. (0027-8424) 2007 IMEx Only protein-protein interactions imex curation shu-2007-5 Functional characterization of human PFTK1 as a cyclin-dependent kinase. human-sh_sy5y Homo sapiens SH-SY5Y cell line sh_sy5y Human neuroblastoma cells anti bait coip anti bait coimmunoprecipitation anti bait coip western blot western blot Immuno blot Immuno blot Shu F., Lv S., Qin Y., Ma X., Wang X., Peng X., Luo Y., Xu BE., Sun X., Wu J. junwu@shanghaigenomics.com 3 human cDNA libraries - (heLa, human T/B lymphocyte and fetal brain. Proc. Natl. Acad. Sci. U.S.A. (0027-8424) 2007 IMEx Only protein-protein interactions imex curation shu-2007-3 Functional characterization of human PFTK1 as a cyclin-dependent kinase. human-293t Homo sapiens 293 cells transformed with SV40 large T antigen 293t 293 cells expressing SV40 large T antigen. The original designation of this cell line was 293tsA1609neo. It is not in Cabri,ATCC or HyperCLDB. anti bait coip anti bait coimmunoprecipitation anti bait coip anti tag western anti tag western blot anti tag western Shu F., Lv S., Qin Y., Ma X., Wang X., Peng X., Luo Y., Xu BE., Sun X., Wu J. junwu@shanghaigenomics.com 3 human cDNA libraries - (heLa, human T/B lymphocyte and fetal brain. Proc. Natl. Acad. Sci. U.S.A. (0027-8424) 2007 IMEx Only protein-protein interactions imex curation shu-2007-6 Functional characterization of human PFTK1 as a cyclin-dependent kinase. in vitro In vitro protein kinase assay protein kinase assay predetermined predetermined participant predetermined Shu F., Lv S., Qin Y., Ma X., Wang X., Peng X., Luo Y., Xu BE., Sun X., Wu J. Additional data showing PFTK1 phosphorylated p53 could not be annotated as source of p53 not given in paper. junwu@shanghaigenomics.com 3 human cDNA libraries - (heLa, human T/B lymphocyte and fetal brain. Proc. Natl. Acad. Sci. U.S.A. (0027-8424) 2007 IMEx Only protein-protein interactions imex curation cdk14_human Cyclin-dependent kinase 14 CDK14 KIAA0834 PFTK1 Serine/threonine-protein kinase PFTAIRE-1 Cell division protein kinase 14 protein protein human Homo sapiens Human MCDLIEPQPAEKIGKMKKLRRTLSESFSRIALKKDDTTFDEICVTKMSTRNCQGMDSVIKPLDTIPEDKKVRVQRTQSTFDPFEKPANQVKRVHSENNACINFKTSSTGKESPKVRRHSSPSSPTSPKFGKADSYEKLEKLGEGSYATVYKGKSKVNGKLVALKVIRLQEEEGTPFTAIREASLLKGLKHANIVLLHDIIHTKETLTLVFEYVHTDLCQYMDKHPGGLHPDNVKLFLFQLLRGLSYIHQRYILHRDLKPQNLLISDTGELKLADFGLARAKSVPSHTYSNEVVTLWYRPPDVLLGSTEYSTCLDMWGVGCIFVEMIQGVAAFPGMKDIQDQLERIFLVLGTPNEDTWPGVHSLPHFKPERFTLYSSKNLRQAWNKLSYVNHAEDLASKLLQCSPKNRLSAQAALSHEYFSDLPPRLWELTDMSSIFTVPNVRLQPEAGESMRAFGKNNSYGKSLSNSKH cdn1a_human Cyclin-dependent kinase inhibitor 1 CDKN1A CAP20 CDKN1 CIP1 MDA6 PIC1 SDI1 WAF1 p21 CDK-interacting protein 1 Melanoma differentiation-associated protein 6 protein protein human Homo sapiens Human MSEPAGDVRQNPCGSKACRRLFGPVDSEQLSRDCDALMAGCIQEARERWNFDFVTETPLEGDFAWERVRGLGLPKLYLPTGPRRGRDELGGGRRPGTSPALLQGTAEEDHVDLSLSCTLVPRSGEQAEGSPGGPGDSQGRKRRQTSMTDFYHSKRRLIFSKRKP cdk4_human Cyclin-dependent kinase 4 CDK4 Cell division protein kinase 4 PSK-J3 protein protein human Homo sapiens Human MATSRYEPVAEIGVGAYGTVYKARDPHSGHFVALKSVRVPNGGGGGGGLPISTVREVALLRRLEAFEHPNVVRLMDVCATSRTDREIKVTLVFEHVDQDLRTYLDKAPPPGLPAETIKDLMRQFLRGLDFLHANCIVHRDLKPENILVTSGGTVKLADFGLARIYSYQMALTPVVVTLWYRAPEVLLQSTYATPVDMWSVGCIFAEMFRRKPLFCGNSEADQLGKIFDLIGLPPEDDWPRDVSLPRGAFPPRGPRPVQSVVPEMEESGAQLLLEMLTFNPHKRISAFRALQHSYLHKDEGNPE cdn2a_human Cyclin-dependent kinase inhibitor 2A CDKN2A CDKN2 Cyclin-dependent kinase 4 inhibitor A p16-INK4a Multiple tumor suppressor 1 MTS1 protein protein human Homo sapiens Human MEPAAGSSMEPSADWLATAAARGRVEEVRALLEAGALPNAPNSYGRRPIQVMMMGSARVAELLLLHGAEPNCADPATLTRPVHDAAREGFLDTLVVLHRAGARLDVRDAWGRLPVDLAEELGHRDVARYLRAAAGGTRGSNHARIDAAEGPSDIPD ccnd3_human G1/S-specific cyclin-D3 CCND3 protein protein human Homo sapiens Human MELLCCEGTRHAPRAGPDPRLLGDQRVLQSLLRLEERYVPRASYFQCVQREIKPHMRKMLAYWMLEVCEEQRCEEEVFPLAMNYLDRYLSCVPTRKAQLQLLGAVCMLLASKLRETTPLTIEKLCIYTDHAVSPRQLRDWEVLVLGKLKWDLAAVIAHDFLAFILHRLSLPRDRQALVKKHAQTFLALCATDYTFAMYPPSMIATGSIGAAVQGLGACSMSGDELTELLAGITGTEVDCLRACQEQIEAALRESLREASQTSSSPAPKAPRGSSSQGPSQTSTPTDVTAIHL ccnd1_human G1/S-specific cyclin-D1 CCND1 BCL1 PRAD1 PRAD1 oncogene BCL-1 oncogene B-cell lymphoma 1 protein protein protein human Homo sapiens Human MEHQLLCCEVETIRRAYPDANLLNDRVLRAMLKAEETCAPSVSYFKCVQKEVLPSMRKIVATWMLEVCEEQKCEEEVFPLAMNYLDRFLSLEPVKKSRLQLLGATCMFVASKMKETIPLTAEKLCIYTDNSIRPEELLQMELLLVNKLKWNLAAMTPHDFIEHFLSKMPEAEENKQIIRKHAQTFVALCATDVKFISNPPSMVAAGSVVAAVQGLNLRSPNNFLSYYRLTRFLSRVIKCDPDCLRACQEQIEALLESSLRQAQQNMDPKAAEEEEEEEEEVDLACTPTDVRDVDI cdk6_human Cyclin-dependent kinase 6 CDK6 Serine/threonine-protein kinase PLSTIRE Cell division protein kinase 6 CDKN6 protein protein human Homo sapiens Human MEKDGLCRADQQYECVAEIGEGAYGKVFKARDLKNGGRFVALKRVRVQTGEEGMPLSTIREVAVLRHLETFEHPNVVRLFDVCTVSRTDRETKLTLVFEHVDQDLTTYLDKVPEPGVPTETIKDMMFQLLRGLDFLHSHRVVHRDLKPQNILVTSSGQIKLADFGLARIYSFQMALTSVVVTLWYRAPEVLLQSSYATPVDLWSVGCIFAEMFRRKPLFRGSSDVDQLGKILDVIGLPGEEDWPRDVALPRQAFHSKSAQPIEKFVTDIDELGKDLLLKCLTFNPAKRISAYSALSHPYFQDLERCKENLDSHLPPSQNTSELNTA pftk1-cdkn1a-4 Interaction between PFTK1 and CDKN1A identified by coimmunoprecipitation 608993 N/A 608999 unspecified role unspecified role bait bait n-terminal flag tag flag tag FLAG-tagged FLAG DYKDDDDKV epitope tag DYKDDDDKV epitope tag FLAG FLAG-tagged 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 N/A 609000 unspecified role unspecified role prey prey region myc tag myc tag EUKLISEED epitope tag EUKLISEED epitope tag undetermined undetermined sequence position undetermined undetermined undetermined sequence position undetermined false physical association physical association false false false Fig. 2B pftk1-cdkn1a-5 Interaction between PFTK1 and CDKN1A identified by coimmunoprecipitation 608993 N/A 609000 unspecified role unspecified role bait bait n-terminal 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 608999 unspecified role unspecified role prey prey n-terminal flag tag flag tag FLAG-tagged FLAG DYKDDDDKV epitope tag DYKDDDDKV epitope tag FLAG FLAG-tagged 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 deletion analysis deletion analysis certain certain sequence position certain certain certain sequence position certain false physical association physical association false false false Fig. 2C pftk1-cdkn1a-6 Interaction between PFTK1 and CDKN1A identified by coimmunoprecipitation 608993 N/A 609000 unspecified role unspecified role prey prey region sufficient to bind sufficient binding region sufficient to bind deletion analysis deletion analysis certain certain sequence position certain certain certain sequence position certain false n-terminal myc tag myc tag EUKLISEED epitope tag EUKLISEED epitope tag deletion analysis deletion analysis n-term range n-terminal range n-term range n-term range n-terminal range n-term range false N/A 608999 unspecified role unspecified role bait bait n-terminal flag tag flag tag FLAG-tagged FLAG DYKDDDDKV epitope tag DYKDDDDKV epitope tag FLAG FLAG-tagged n-term range n-terminal range n-term range n-term range n-terminal range n-term range false physical association physical association false false false Fig. 2D cdk4-cdkn2a-2 Interaction between CDK4 and CDKN2A identified by coimmunoprecipitation 608993 N/A 609001 unspecified role unspecified role prey prey n-terminal ha tag ha tag YPYDVPDYA epitope tag YPYDVPDYA epitope tag n-term range n-terminal range n-term range n-term range n-terminal range n-term range false N/A 609002 unspecified role unspecified role bait bait n-terminal 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 physical association physical association false false false Fig. 2E pftk1-ccnd3-3 Interaction between PFTK1 and CCND3 identified by coimmunoprecipitation 608993 N/A 608999 unspecified role unspecified role bait bait n-terminal flag tag flag tag FLAG-tagged FLAG DYKDDDDKV epitope tag DYKDDDDKV epitope tag FLAG FLAG-tagged n-term range n-terminal range n-term range n-term range n-terminal range n-term range false N/A 609003 unspecified role unspecified role prey prey n-terminal ha tag ha tag YPYDVPDYA epitope tag YPYDVPDYA epitope tag n-term range n-terminal range n-term range n-term range n-terminal range n-term range false physical association physical association false false false Fig. 2F Weak interaction pftk1-ccnd3-4 Interaction between PFTK1, CCNA1 and CCND3 identified by coimmunoprecipitation 608993 N/A 608999 unspecified role unspecified role bait bait n-terminal flag tag flag tag FLAG-tagged FLAG DYKDDDDKV epitope tag DYKDDDDKV epitope tag FLAG FLAG-tagged n-term range n-terminal range n-term range n-term range n-terminal range n-term range false N/A 609000 unspecified role unspecified role prey prey n-terminal 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 609003 unspecified role unspecified role prey prey n-terminal ha tag ha tag YPYDVPDYA epitope tag YPYDVPDYA 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 Fig. 2F pftk1-cdkn1a-1 Interaction between PFTK1 and CDKN1A identified by Y2H 608994 N/A 608999 unspecified role unspecified role bait bait N/A 609000 unspecified role unspecified role prey prey physical association physical association false false false Table 1 Protein fragments only encoded but no detail given as to length pftk1-ccnd3-1 Interaction between PFTK1 and CCND3 identified by Y2H 608994 N/A 609003 unspecified role unspecified role prey prey N/A 608999 unspecified role unspecified role bait bait physical association physical association false false false Table 1 Protein fragments only encoded but no detail given as to length pftk1-cdkn1a-2 Interaction between PFTK1 and CDKN1A identified by Y2H 608995 N/A 608999 unspecified role unspecified role bait bait N/A 609000 unspecified role unspecified role prey prey physical association physical association false false false Fig. 1 pftk1-ccnd3-2 Interaction between PFTK1 and CCND3 identified by Y2H 608995 N/A 609003 unspecified role unspecified role prey prey N/A 608999 unspecified role unspecified role bait bait physical association physical association false false false Fig. 1 pftk1-ccnd3-5 Interaction between PFTK1 and CCND3 identified by coimmunoprecipitation 608996 N/A 609003 unspecified role unspecified role prey prey N/A 608999 unspecified role unspecified role bait bait physical association physical association false false false Fig. 3C Results confirmed by immunodepletion cdk6-ccdn1-1 CCND1 and CCND3 interact with CDK6 608996 N/A 609003 unspecified role unspecified role prey prey N/A 609004 unspecified role unspecified role prey prey N/A 609005 unspecified role unspecified role bait bait association association false false false Fig 3C pftk1-cdkn1a-3 Interaction between PFTK1 and CDKN1A identified by coimmunoprecipitation 608997 N/A 609000 unspecified role unspecified role bait bait N/A 608999 unspecified role unspecified role prey prey n-terminal flag tag flag tag FLAG-tagged FLAG DYKDDDDKV epitope tag DYKDDDDKV epitope tag FLAG FLAG-tagged n-term range n-terminal range n-term range n-term range n-terminal range n-term range false physical association physical association false false false Fig. 2A pftk1-1 Autophosphorylation of PFTK1 608998 N/A 608999 putative self putative self putative self putative self O94921:p.Lys164Ala mutation mutation mutation analysis mutation analysis certain certain sequence position certain certain certain sequence position certain false human-293t Homo sapiens 293 cells transformed with SV40 large T antigen 293t 293 cells expressing SV40 large T antigen. The original designation of this cell line was 293tsA1609neo. It is not in Cabri,ATCC or HyperCLDB. phosphorylation phosphorylation reaction phosphorylation false true false Fig. 4a Activity enhanced when cells transfected with CCND3 and inhibted when transfected with p21Cip1 prior to immunoprecipitation of kinase