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/ Substrate profiling of human vaccinia-related kinases identifies coilin, a Cajal body nuclear protein, as a phosphorylation target with neurological implications. Substrate profiling of human vaccinia-related kinases identifies coilin, a Cajal body nuclear protein, as a phosphorylation target with neurological implications. J Proteomics (1874-3919) 2011 imex curation Sanz-Garcia M., Vazquez-Cedeira M., Kellerman E., Renbaum P., Levy-Lahad E., Lazo P.A. 2011-09-27: P Lazo; Centro de Investigación del Cáncer, CSIC-Universidad de Salamanca, Spain pedro.lazo@usal.es,plazozbi@usal.es,pedro.lazo@csic.es 21-Dec-2011: Contacted by JYOTI. Only protein-protein interactions 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. pull down pull down anti tag western anti tag western blot anti tag western Sanz-Garcia M., Vazquez-Cedeira M., Kellerman E., Renbaum P., Levy-Lahad E., Lazo P.A. 2011-09-27: P Lazo; Centro de Investigación del Cáncer, CSIC-Universidad de Salamanca, Spain pedro.lazo@usal.es,plazozbi@usal.es,pedro.lazo@csic.es Accepted 2011-OCT-24 by MGT 21-Dec-2011: Contacted by JYOTI. J Proteomics (1874-3919) 2011 Only protein-protein interactions imex curation Substrate profiling of human vaccinia-related kinases identifies coilin, a Cajal body nuclear protein, as a phosphorylation target with neurological implications. Substrate profiling of human vaccinia-related kinases identifies coilin, a Cajal body nuclear protein, as a phosphorylation target with neurological implications. J Proteomics (1874-3919) 2011 imex curation Sanz-Garcia M., Vazquez-Cedeira M., Kellerman E., Renbaum P., Levy-Lahad E., Lazo P.A. 2011-09-27: P Lazo; Centro de Investigación del Cáncer, CSIC-Universidad de Salamanca, Spain pedro.lazo@usal.es,plazozbi@usal.es,pedro.lazo@csic.es 21-Dec-2011: Contacted by JYOTI. Only protein-protein interactions 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. confocal microscopy confocal microscopy immunostaining immunostaining Sanz-Garcia M., Vazquez-Cedeira M., Kellerman E., Renbaum P., Levy-Lahad E., Lazo P.A. 2011-09-27: P Lazo; Centro de Investigación del Cáncer, CSIC-Universidad de Salamanca, Spain pedro.lazo@usal.es,plazozbi@usal.es,pedro.lazo@csic.es Accepted 2011-OCT-24 by MGT 21-Dec-2011: Contacted by JYOTI. J Proteomics (1874-3919) 2011 Only protein-protein interactions imex curation Substrate profiling of human vaccinia-related kinases identifies coilin, a Cajal body nuclear protein, as a phosphorylation target with neurological implications. Substrate profiling of human vaccinia-related kinases identifies coilin, a Cajal body nuclear protein, as a phosphorylation target with neurological implications. J Proteomics (1874-3919) 2011 imex curation Sanz-Garcia M., Vazquez-Cedeira M., Kellerman E., Renbaum P., Levy-Lahad E., Lazo P.A. 2011-09-27: P Lazo; Centro de Investigación del Cáncer, CSIC-Universidad de Salamanca, Spain pedro.lazo@usal.es,plazozbi@usal.es,pedro.lazo@csic.es 21-Dec-2011: Contacted by JYOTI. Only protein-protein interactions 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 Sanz-Garcia M., Vazquez-Cedeira M., Kellerman E., Renbaum P., Levy-Lahad E., Lazo P.A. 2011-09-27: P Lazo; Centro de Investigación del Cáncer, CSIC-Universidad de Salamanca, Spain pedro.lazo@usal.es,plazozbi@usal.es,pedro.lazo@csic.es Accepted 2011-OCT-24 by MGT 21-Dec-2011: Contacted by JYOTI. J Proteomics (1874-3919) 2011 Only protein-protein interactions imex curation Substrate profiling of human vaccinia-related kinases identifies coilin, a Cajal body nuclear protein, as a phosphorylation target with neurological implications. Substrate profiling of human vaccinia-related kinases identifies coilin, a Cajal body nuclear protein, as a phosphorylation target with neurological implications. J Proteomics (1874-3919) 2011 imex curation Sanz-Garcia M., Vazquez-Cedeira M., Kellerman E., Renbaum P., Levy-Lahad E., Lazo P.A. 2011-09-27: P Lazo; Centro de Investigación del Cáncer, CSIC-Universidad de Salamanca, Spain pedro.lazo@usal.es,plazozbi@usal.es,pedro.lazo@csic.es 21-Dec-2011: Contacted by JYOTI. Only protein-protein interactions 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 Sanz-Garcia M., Vazquez-Cedeira M., Kellerman E., Renbaum P., Levy-Lahad E., Lazo P.A. 2011-09-27: P Lazo; Centro de Investigación del Cáncer, CSIC-Universidad de Salamanca, Spain pedro.lazo@usal.es,plazozbi@usal.es,pedro.lazo@csic.es Accepted 2011-OCT-24 by MGT 21-Dec-2011: Contacted by JYOTI. J Proteomics (1874-3919) 2011 Only protein-protein interactions imex curation Substrate profiling of human vaccinia-related kinases identifies coilin, a Cajal body nuclear protein, as a phosphorylation target with neurological implications. Substrate profiling of human vaccinia-related kinases identifies coilin, a Cajal body nuclear protein, as a phosphorylation target with neurological implications. J Proteomics (1874-3919) 2011 imex curation Sanz-Garcia M., Vazquez-Cedeira M., Kellerman E., Renbaum P., Levy-Lahad E., Lazo P.A. 2011-09-27: P Lazo; Centro de Investigación del Cáncer, CSIC-Universidad de Salamanca, Spain pedro.lazo@usal.es,plazozbi@usal.es,pedro.lazo@csic.es 21-Dec-2011: Contacted by JYOTI. Only protein-protein interactions in vitro In vitro peptide array peptide array autoradiography autoradiography Sanz-Garcia M., Vazquez-Cedeira M., Kellerman E., Renbaum P., Levy-Lahad E., Lazo P.A. The isoform VRK2B (Accession No =AJ512204, as given in Reference 14) does not correspond with 100% identity to any isoform in UniProt. 2011-09-27: P Lazo; Centro de Investigación del Cáncer, CSIC-Universidad de Salamanca, Spain Chip kinase assay was performed in vitro with different preparations of VRK1 (GST and His tagged) and VRK2A and VRK2B. The phosphorylated spots were detected with a BioRad FX Imager. pedro.lazo@usal.es,plazozbi@usal.es,pedro.lazo@csic.es Accepted 2011-OCT-24 by MGT 21-Dec-2011: Contacted by JYOTI. J Proteomics (1874-3919) 2011 Only protein-protein interactions imex curation Substrate profiling of human vaccinia-related kinases identifies coilin, a Cajal body nuclear protein, as a phosphorylation target with neurological implications. Substrate profiling of human vaccinia-related kinases identifies coilin, a Cajal body nuclear protein, as a phosphorylation target with neurological implications. J Proteomics (1874-3919) 2011 imex curation Sanz-Garcia M., Vazquez-Cedeira M., Kellerman E., Renbaum P., Levy-Lahad E., Lazo P.A. 2011-09-27: P Lazo; Centro de Investigación del Cáncer, CSIC-Universidad de Salamanca, Spain pedro.lazo@usal.es,plazozbi@usal.es,pedro.lazo@csic.es 21-Dec-2011: Contacted by JYOTI. Only protein-protein interactions in vitro In vitro protein kinase assay protein kinase assay weight autoradiogra molecular weight estimation by autoradiography weight autoradiogra Sanz-Garcia M., Vazquez-Cedeira M., Kellerman E., Renbaum P., Levy-Lahad E., Lazo P.A. 2011-09-27: P Lazo; Centro de Investigación del Cáncer, CSIC-Universidad de Salamanca, Spain Rejected 2011-OCT-13 by MGT. I have removed Fig 1 from this Expt as this has been entered elsewhere, and tags were not correct for Fig1. No GO terms entered. This is a Kinase assay, with autoradiography for detection and phosphorylation for Interaction Type . You have not entered an interaction showing the autophosphorylation of VRK1. (not sure if this is necessary, but I have had to do it before) These should be Neutral interactors, also for Biological Roles should use Enz/Enz Target. The S184A mutation for Coilin has not been entered.(Fig 2B). - now looking at Fig 6A - Coilin is not in any of the reactions- so the R358 mutation need not be added here. Fig 6 Interactions have not been entered - pull downs with Participant Detection by Wt autoradiography + VRK1 R358X mutant. pedro.lazo@usal.es,plazozbi@usal.es,pedro.lazo@csic.es Accepted 2011-OCT-24 by MGT J Proteomics (1874-3919) 2011 Only protein-protein interactions imex curation Substrate profiling of human vaccinia-related kinases identifies coilin, a Cajal body nuclear protein, as a phosphorylation target with neurological implications. Substrate profiling of human vaccinia-related kinases identifies coilin, a Cajal body nuclear protein, as a phosphorylation target with neurological implications. J Proteomics (1874-3919) 2011 imex curation Sanz-Garcia M., Vazquez-Cedeira M., Kellerman E., Renbaum P., Levy-Lahad E., Lazo P.A. 2011-09-27: P Lazo; Centro de Investigación del Cáncer, CSIC-Universidad de Salamanca, Spain pedro.lazo@usal.es,plazozbi@usal.es,pedro.lazo@csic.es 21-Dec-2011: Contacted by JYOTI. Only protein-protein interactions human-hela Homo sapiens epitheloid cervix carcinoma cells Human hela Human epitheloid cervix carcinoma HeLa cells derivation: cervix; epithelial; adenocarcinoma ATCC no:CCL-2 confocal microscopy confocal microscopy immunostaining immunostaining Sanz-Garcia M., Vazquez-Cedeira M., Kellerman E., Renbaum P., Levy-Lahad E., Lazo P.A. 2011-09-27: P Lazo; Centro de Investigación del Cáncer, CSIC-Universidad de Salamanca, Spain pedro.lazo@usal.es,plazozbi@usal.es,pedro.lazo@csic.es Accepted 2011-OCT-24 by MGT 21-Dec-2011: Contacted by JYOTI. J Proteomics (1874-3919) 2011 Only protein-protein interactions imex curation coil_human Coilin COIL CLN80 p80-coilin protein protein human Homo sapiens Human MAASETVRLRLQFDYPPPATPHCTAFWLLVDLNRCRVVTDLISLIRQRFGFSSGAFLGLYLEGGLLPPAESARLVRDNDCLRVKLEERGVAENSVVISNGDINLSLRKAKKRAFQLEEGEETEPDCKYSKKHWKSRENNNNNEKVLDLEPKAVTDQTVSKKNKRKNKATCGTVGDDNEEAKRKSPKKKEKCEYKKKAKNPKSPKVQAVKDWANQRCSSPKGSARNSLVKAKRKGSVSVCSKESPSSSSESESCDESISDGPSKVTLEARNSSEKLPTELSKEEPSTKNTTADKLAIKLGFSLTPSKGKTSGTTSSSSDSSAESDDQCLMSSSTPECAAGFLKTVGLFAGRGRPGPGLSSQTAGAAGWRRSGSNGGGQAPGASPSVSLPASLGRGWGREENLFSWKGAKGRGMRGRGRGRGHPVSCVVNRSTDNQRQQQLNDVVKNSSTIIQNPVETPKKDYSLLPLLAAAPQVGEKIAFKLLELTSSYSPDVSDYKEGRILSHNPETQQVDIEILSSLPALREPGKFDLVYHNENGAEVVEYAVTQESKITVFWKELIDPRLIIESPSNTSSTEPA DIP protein P38432 original sequence version: 96 879702B590F16849 vrk2_human Serine/threonine-protein kinase VRK2 VRK2 Vaccinia-related kinase 2 protein protein human Homo sapiens Human MPPKRNEKYKLPIPFPEGKVLDDMEGNQWVLGKKIGSGGFGLIYLAFPTNKPEKDARHVVKVEYQENGPLFSELKFYQRVAKKDCIKKWIERKQLDYLGIPLFYGSGLTEFKGRSYRFMVMERLGIDLQKISGQNGTFKKSTVLQLGIRMLDVLEYIHENEYVHGDIKAANLLLGYKNPDQVYLADYGLSYRYCPNGNHKQYQENPRKGHNGTIEFTSLDAHKGVALSRRSDVEILGYCMLRWLCGKLPWEQNLKDPVAVQTAKTNLLDELPQSVLKWAPSGSSCCEIAQFLVCAHSLAYDEKPNYQALKKILNPHGIPLGPLDFSTKGQSINVHTPNSQKVDSQKAATKQVNKAHNRLIEKKVHSERSAESCATWKVQKEEKLIGLMNNEAAQESTRRRQKYQESQEPLNEVNSFPQKISYTQFPNSFYEPHQDFTSPDIFKKSRSPSWYKYTSTVSTGITDLESSTGLWPTISQFTLSEETNADVYYYRIIIPVLLMLVFLALFFL 9F3F6FCC9280568F vrk1_human Serine/threonine-protein kinase VRK1 Vaccinia-related kinase 1 VRK1 protein protein human Homo sapiens Human MPRVKAAQAGRQSSAKRHLAEQFAVGEIITDMAKKEWKVGLPIGQGGFGCIYLADMNSSESVGSDAPCVVKVEPSDNGPLFTELKFYQRAAKPEQIQKWIRTRKLKYLGVPKYWGSGLHDKNGKSYRFMIMDRFGSDLQKIYEANAKRFSRKTVLQLSLRILDILEYIHEHEYVHGDIKASNLLLNYKNPDQVYLVDYGLAYRYCPEGVHKEYKEDPKRCHDGTIEFTSIDAHNGVAPSRRGDLEILGYCMIQWLTGHLPWEDNLKDPKYVRDSKIRYRENIASLMDKCFPEKNKPGEIAKYMETVKLLDYTEKPLYENLRDILLQGLKAIGSKDDGKLDLSVVENGGLKAKTITKKRKKEIEESKEPGVEDTEWSNTQTEEAIQTRSRTRKRVQK 5640C624BF059949 coil-vrk1-2 6 8 enzyme target enzyme target substrate substrate neutral component neutral component region myc tag myc tag EUKLISEED epitope tag EUKLISEED epitope tag undetermined undetermined sequence position undetermined undetermined undetermined sequence position undetermined 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. 10 enzyme enzyme neutral component neutral component region gst tag glutathione s tranferase tag glutathione S-tranferase tag glutathione S-tranferase tag gst tag undetermined undetermined sequence position undetermined undetermined undetermined sequence position undetermined region sufficient to bind sufficient binding region sufficient to bind certain certain sequence position certain certain certain sequence position certain 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 2, 6G. Phosphorylation of coilin and VRK1 observed vrk1-coil-1 1 8 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 10 unspecified role unspecified role bait bait region gst tag glutathione s tranferase tag glutathione S-tranferase tag glutathione S-tranferase tag gst tag undetermined undetermined sequence position undetermined undetermined undetermined sequence position undetermined physical association physical association 4A, 5B. vrk1-coil-2 3 8 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 10 unspecified role unspecified role bait bait physical association physical association 4C coil-vrk1-1 7 8 unspecified role unspecified role neutral component neutral component 10 unspecified role unspecified role neutral component neutral component colocalization colocalization 3 vrk1-coil-4 5 10 enzyme enzyme bait bait region gst tag glutathione s tranferase tag glutathione S-tranferase tag glutathione S-tranferase tag gst tag undetermined undetermined sequence position undetermined undetermined undetermined sequence position undetermined 8 enzyme target enzyme target substrate substrate prey prey region binding region binding-associated region binding region certain certain sequence position certain certain certain sequence position certain chemical synthesis Chemical synthesis (Chemical synthesis) direct interaction direct interaction 1 vrk1-coil-5 5 10 enzyme enzyme bait bait 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 8 enzyme target enzyme target substrate substrate prey prey region binding region binding-associated region binding region certain certain sequence position certain certain certain sequence position certain chemical synthesis Chemical synthesis (Chemical synthesis) direct interaction direct interaction 1 vrk2-coil 5 9 enzyme enzyme bait bait region gst tag glutathione s tranferase tag glutathione S-tranferase tag glutathione S-tranferase tag gst tag undetermined undetermined sequence position undetermined undetermined undetermined sequence position undetermined 8 enzyme target enzyme target substrate substrate prey prey region binding region binding-associated region binding region certain certain sequence position certain certain certain sequence position certain chemical synthesis Chemical synthesis (Chemical synthesis) direct interaction direct interaction 1 coil-vrk1 4 10 unspecified role unspecified role prey prey 8 unspecified role unspecified role bait bait region myc tag myc tag EUKLISEED epitope tag EUKLISEED epitope tag undetermined undetermined sequence position undetermined undetermined undetermined sequence position undetermined physical association physical association 4B vrk1-1 6 10 putative self putative self putative self putative self region gst tag glutathione s tranferase tag glutathione S-tranferase tag glutathione S-tranferase tag gst tag undetermined undetermined sequence position undetermined undetermined undetermined sequence position undetermined n terminal sufficient to bind sufficient binding region sufficient to bind certain certain sequence position certain certain certain sequence position certain 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 true 2, 6G. Phosphorylation of GST- VRK1 observed According to OMIM the mutation Arg 358 results in a truncated protein. coil-vrk1-3 2 8 unspecified role unspecified role neutral component neutral component 10 unspecified role unspecified role neutral component neutral component region ha tag ha tag YPYDVPDYA epitope tag YPYDVPDYA epitope tag undetermined undetermined sequence position undetermined undetermined undetermined sequence position undetermined region sufficient to bind sufficient binding region sufficient to bind certain certain sequence position certain certain certain sequence position certain colocalization colocalization 5A The mutation R358X results in the distribution of VRK1 in the cytoplasm due to loss of NLS. Even under these conditions interaction between VRK1 and Coilin can be observed coil-vrk1-4 7 10 unspecified role unspecified role neutral component neutral component region sufficient to bind sufficient binding region sufficient to bind certain certain sequence position certain certain certain sequence position certain region ha tag ha tag YPYDVPDYA epitope tag YPYDVPDYA epitope tag undetermined undetermined sequence position undetermined undetermined undetermined sequence position undetermined 8 unspecified role unspecified role neutral component neutral component colocalization colocalization 5A The mutation R358X results in the distribution of VRK1 in the cytoplasm due to loss of NLS. Even under these conditions interaction between VRK1 and Coilin can be observed.