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/ Cytoplasmic LEK1 is a regulator of microtubule function through its interaction with the LIS1 pathway. Cytoplasmic LEK1 is a regulator of microtubule function through its interaction with the LIS1 pathway. Proc. Natl. Acad. Sci. U.S.A. (0027-8424) 2005 imex curation Soukoulis V., Reddy S., Pooley RD., Feng Y., Walsh CA., Bader DM. david.bader@vanderbilt.edu 09-April-2009: Contacted by JYOTI. davet 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 nucleotide sequence nucleotide sequence identification sequence cloning sequence cloning nucleotide sequence Soukoulis V., Reddy S., Pooley RD., Feng Y., Walsh CA., Bader DM. david.bader@vanderbilt.edu Accepted 2009-MAY-1 by DAVET. 09-April-2009: Contacted by JYOTI. a whole mouse embryonic day 11 cDNA library was used. Proc. Natl. Acad. Sci. U.S.A. (0027-8424) 2005 davet Only protein-protein interactions imex curation Cytoplasmic LEK1 is a regulator of microtubule function through its interaction with the LIS1 pathway. Cytoplasmic LEK1 is a regulator of microtubule function through its interaction with the LIS1 pathway. Proc. Natl. Acad. Sci. U.S.A. (0027-8424) 2005 imex curation Soukoulis V., Reddy S., Pooley RD., Feng Y., Walsh CA., Bader DM. david.bader@vanderbilt.edu 09-April-2009: Contacted by JYOTI. davet 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 featur predetermined feature Soukoulis V., Reddy S., Pooley RD., Feng Y., Walsh CA., Bader DM. david.bader@vanderbilt.edu Accepted 2009-MAY-1 by DAVET. 09-April-2009: Contacted by JYOTI. Proc. Natl. Acad. Sci. U.S.A. (0027-8424) 2005 davet Only protein-protein interactions imex curation Cytoplasmic LEK1 is a regulator of microtubule function through its interaction with the LIS1 pathway. Cytoplasmic LEK1 is a regulator of microtubule function through its interaction with the LIS1 pathway. Proc. Natl. Acad. Sci. U.S.A. (0027-8424) 2005 imex curation Soukoulis V., Reddy S., Pooley RD., Feng Y., Walsh CA., Bader DM. david.bader@vanderbilt.edu 09-April-2009: Contacted by JYOTI. davet Only protein-protein interactions chlae-cos_7 Cercopithecus aethiops simian cells transformed with SV40 Grivet cos_7 cercopithecus aethiops, by transformation with SV40 anti tag coip anti tag coimmunoprecipitation anti tag coip anti tag western anti tag western blot anti tag western Soukoulis V., Reddy S., Pooley RD., Feng Y., Walsh CA., Bader DM. david.bader@vanderbilt.edu Accepted 2009-MAY-1 by DAVET. 09-April-2009: Contacted by JYOTI. Proc. Natl. Acad. Sci. U.S.A. (0027-8424) 2005 davet Only protein-protein interactions imex curation Cytoplasmic LEK1 is a regulator of microtubule function through its interaction with the LIS1 pathway. Cytoplasmic LEK1 is a regulator of microtubule function through its interaction with the LIS1 pathway. Proc. Natl. Acad. Sci. U.S.A. (0027-8424) 2005 imex curation Soukoulis V., Reddy S., Pooley RD., Feng Y., Walsh CA., Bader DM. david.bader@vanderbilt.edu 09-April-2009: Contacted by JYOTI. davet Only protein-protein interactions chlae-cos_7 Cercopithecus aethiops simian cells transformed with SV40 Grivet cos_7 cercopithecus aethiops, by transformation with SV40 fluorescence fluorescence technology fluorescence immunostaining immunostaining Soukoulis V., Reddy S., Pooley RD., Feng Y., Walsh CA., Bader DM. david.bader@vanderbilt.edu Accepted 2009-MAY-1 by DAVET. 09-April-2009: Contacted by JYOTI. Proc. Natl. Acad. Sci. U.S.A. (0027-8424) 2005 davet Only protein-protein interactions imex curation Cytoplasmic LEK1 is a regulator of microtubule function through its interaction with the LIS1 pathway. Cytoplasmic LEK1 is a regulator of microtubule function through its interaction with the LIS1 pathway. Proc. Natl. Acad. Sci. U.S.A. (0027-8424) 2005 imex curation Soukoulis V., Reddy S., Pooley RD., Feng Y., Walsh CA., Bader DM. david.bader@vanderbilt.edu 09-April-2009: Contacted by JYOTI. davet Only protein-protein interactions mouse-3t3 Mus musculus embryo cells 3t3 Mouse embryo fibroblast cells confocal microscopy confocal microscopy immunostaining immunostaining Soukoulis V., Reddy S., Pooley RD., Feng Y., Walsh CA., Bader DM. david.bader@vanderbilt.edu Accepted 2009-MAY-1 by DAVET. 09-April-2009: Contacted by JYOTI. Proc. Natl. Acad. Sci. U.S.A. (0027-8424) 2005 davet Only protein-protein interactions imex curation nde1_cerae_protein Nuclear distribution protein nudE homolog 1 protein protein chlae Chlorocebus aethiops Cercopithecus aethiops Grivet Green monkey An amino acid sequence which is not represented in UniProt similar to NDE1_HUMAN (AC: Q9NXR1). q155p7_mouse Cenpf Lek1 protein protein mouse Mus musculus Mouse MSWALEEWKEGLPSRALQKIQELEGQLEKLKKEKQQRQFQLDSLEAALQKQKQKVEDGKTEGADLKRENQRLMEICEHLEKSRQKLSHELQVKESQVNLQESQLSSCKKQIEKLEQELKRCKSEFERSQQVAQSADVSLNPCSTPQKLFATPLTPSSTYEDLKEKYNKEVEERKRLEEEVKALHAKKVSLPVSQATMNHRDIARHQASSSVFPWQQENTPSRLSSDALKTPLRRDGSAAHFLGEEVSPNKSSMKTGRGDCSSLPGEPHSAQLLHQAKAQNQDLKSKMTELELRLQGQEKEMRSQVNKCQDLQLQLEKTKVELIEKERILNKTRDEVVRSTAQYDQAAAKCTTLEQKLKTLTEELSCHRQNAESAKRSLEQRIKEKEKELQEELSRQHQSFQALDSEYTQMKTRLTQELQQVKHLHSTLQLELEKVTSVKQQLERNLEEIRLKLSRAEQALQASQVAENELRRSSEEMKKENSLIRSQSEQRTREVCHLEEELGKVKVSLSKSQNFAEEMKAKNTSQEIMLRDLQEKLNQQENSLTLEKLKLALADLERQRNCSQDLLKKREHHIDQLNNKLNKIEKEFETLLSALELKKKECEELKEEKNQISFWKIDSEKLINQIESEKEILLGKINHLETSLKTQQVSPDSNERIRTLEMERENFTVEIKNLQSMLDSKMVEIKTQKQAYLELQQKSESSDQKHQKEIENMCLKANKLTGQVESLECKLQLLSSEVVTKDQQYQDLRMEYETLRDLLKSRGSSLVTNEDNQRSSEDNQRSSEDNQRGSLAFEQQPAVSDSFANVMGRKGSINSERSDCSVDGGRSPEHIAILQNRVTSLESSLESQNQMNSDLQMRCEELLQIKGEVEENLSKAEQIHQNFVAETNQCISKLQEDAAVHQNIVAETLATLESKEKELQLLKEKLEAQQTEVQKLNKNNCLLEGTLKELQLLSDTLSSEKKEMNSIISLSKKNIEELTQANEALKEVNEALEQEKMNLLQKHEKITSCIAEQERSIAELSDQYKQERLQLLQRCEETEAVLEDLRGNYKTAQENNAKLECMLSECTALCENRKNELEQLKETFAKEQQEFLTKLAFAEEQNRKLMLELEIEQQTVRSEITNTNKHSMSATDGLRQECLTLNEEQNEQQNEVSNLTHENEQLMELTQTKHDSYLAVEPVENSVKATEDEIGKSSSQYQMDIDTKDISLDSYKAQLVHLEALVRILEVQLDQSEEENKKLHLELQTIREELETKSSQDPQSQARTGLKDCDTAEEKYVSMLQELSASQNENAHLQCSLQTAVNKLNELGKMCDVLRVEKLQLESELNDSRTECITATSQMTAEVEKLVSEMKMLNHESALSQNELMKDTSGGEFHDKANHSSVFLTPLDSSNFCEQMTLSSKEVRVHFAELQEKFSCLQSEHKILHDQHCEVSSKMSALRSYVDTLKAENSALSMSLRTLQGDLVKEGEPAAEGGHGLPLSFCGADSPSLTNFGETSFYKDVLEQTGDTCHLSLEGNASANSCDLDEEFSSSLEEETLTEKESPPAPGRTVEGLEVLCQVYLQSLKNLEEKTESQRIMKNKEIEELEQLLSSERKELSCLRKQYLSEKEQWQQKLTSVTLEMESKLAEEKQQTKTLSLELEVARLQLQELDLSSRSLLGTDLESVVRCQNDNYDIKESEVYISETTEKTPKQDTDQTCDKDIQQDLGLETSVTESETTRLTGEGCEEQPPKTNCEAPAEDKTQDCSECISELCSSSNVLVPMDVLEDQGSIQNLQLQKDTLNENLRLLPEVEDWDKKVESLLNEIMEADSKLSLQEVQLKMKIATCIQLEKIVKDLRKEKADLSEKLESLPCNQEVCLRVERSEEDLGFNLDMGANELLSKSTKDNATNTEDNYKEKFLDMERELTRIKSEKANIEHHILSVETNLEVVQAEKLCLERDTESKQKVIIDLKEELFTVISERNRLREELDNVSKESKALDQMSKKMKEKIEELESHQRESLRHIGAVESEVKDKADLIQTLSFNVGELTKDKAHLQEQLQNLQNDSQELSLAIGELEIQIGQLNKEKESLVKESQNFQIKLTESECEKQTISKALEVALKEKGEFAVQLSSAQEEVHQLRRGIEKLSVRIEADEKKHLSAVAKLKESQRESDSLKDTVETLERELERSEENQELAILDSENLKAEVETLKAQKDEMTKSLRIFELDLVTVRTERENLAKQLQEKQSRVSELDERCSSLRRLLEEKEQARVQMEEDSKSAMLMLQMQLKELREEVAALCNDQETLKAQEQSLDQPGEEVHHLKSSIRKLKVHIDADEKKHQNILEQLKESKHHADLLKDRVENLEQELILSEKNMIFQAEKSKAEIQTLKSEIQRMAQNLQDLQLELISTRSENENLIKELKKEQERVSDLETINPSIENLLKDKEQEKVQMKEEAKITVEMLQTQLKELNETVVSLCNDQEVSKTKEQNLGSQVQTLELEKAQLLQDLGEAKNKYIIFQSSVNALTQEVEAGKQKLEKGEKEIRTLKEQLKSQEQLVCKLAQVEGEQELWQKQKLELRNVTMALEQKVQVLQSENNTLQSTYEALQNSHKSLESELGLIKLEKVALVERVSTISGKEAELQRELRDMLQKTTQLSEDYNKEKNRLTEEVEVLREELQNTKAAHLKSVNQLEKELQRAQGKIKLMLKSCRQLEGEKEMLQKELSQLEAAQQQRAGSLVDSNVDEVMTENKALKETLEEKVKEADKYLDKYCSLLISHEELEKAKEILEIEVARLKSRQSRQDLQSSPLLNSSIPGPSPNTSVSEMKSASGQNKASGKRQRSSGVWEHGKRAAPSTAETFSKKSRKSDSKSTRPAEHEQETEFEPEGLPEVVKKGFADIPTGKTSPYILRRTTMATRTSPRFATQKLVGSSPSLGKENVVESSKPTAGGSRSQKVKVVQESSADSHTAFQELPAKSLTASNIPGRNSTESPREGLRAKRAYPASSPAAGPDPTNNENCRVQ 3CB74C4F24537155 nde1_mouse Nuclear distribution protein nudE homolog 1 Nde1 Nude protein protein mouse Mus musculus Mouse MEDSGKTFESEEEETNYWRDLAMTYKQRAENTQEELREFQEGSREYEAELEAQLQQIETRNRDLLSENNRLRMELESVKEKFEMQHSEGYRQISALEDDLAQTKAIKDQLQKYIRELEQANDDLERAKRATIMSLEDFEQRLNQAIERNAFLESELDEKENLLESVQRLKDEARDLRQELAVQQKQDKPRTPMPGSGQAKRTDMAVQATGSVPSTPVAHRGPSSGLNTPGMFRRGLDSSTSGTPLTPAARISALNIVGDLLRKVGALESKLASCRNFMYDQSPSRTSGPASGRGTKNRDGVDRRPGSTSVGDKGSGKRLEFGKPASEPASPALPSAQGVVKLLL C0A339C4F51EF8D9 gamma_tubulin_cerae gamma tubulin protein protein protein chlae Chlorocebus aethiops Cercopithecus aethiops Grivet Green monkey An amino acid sequence which is not represented in UniProt lis1_mouse Platelet-activating factor acetylhydrolase IB subunit beta PAF acetylhydrolase 45 kDa subunit PAF-AH alpha Lissencephaly-1 protein Pafah1b1 Lis1 Lis-1 Pafaha protein protein mouse Mus musculus Mouse MVLSQRQRDELNRAIADYLRSNGYEEAYSVFKKEAELDMNEELDKKYAGLLEKKWTSVIRLQKKVMELESKLNEAKEEFTSGGPLGQKRDPKEWIPRPPEKYALSGHRSPVTRVIFHPVFSVMVSASEDATIKVWDYETGDFERTLKGHTDSVQDISFDHSGKLLASCSADMTIKLWDFQGFECIRTMHGHDHNVSSVAIMPNGDHIVSASRDKTIKMWEVQTGYCVKTFTGHREWVRMVRPNQDGTLIASCSNDQTVRVWVVATKECKAELREHEHVVECISWAPESSYSSISEATGSETKKSGKPGPFLLSGSRDKTIKMWDVSTGMCLMTLVGHDNWVRGVLFHSGGKFILSCADDKTLRVWDYKNKRCMKTLNAHEHFVTSLDFHKTAPYVVTGSVDQTVKVWECR DIP protein P63005 original sequence version: 47 4DBF6A24A6B131CD cenpf-nde1 2 7 unspecified role unspecified role bait bait c-terminus required to bind necessary binding region required to bind deletion analysis deletion analysis certain certain sequence position certain certain certain sequence position certain 8 unspecified role unspecified role prey prey physical association physical association Fig 2A. Yeast two-hybrid techniques were used to determine which region within the SRR of cytLEK1 interacted with NudE by dividing the SRR into the N domain (amino acids 1906�1964), the repeat (R) domain (amino acids 1964�2071), and the C domain (amino acids 2071�2149). Only those constructs containing the C domain, not necessarily the R domain itself, retained interaction with full-length NudE (Fig. 2A). cytLEK1 interacts with NDE1. The species of origin of cytLEK1 is assumed to be mouse since in the introduction the authors "Our laboratory has studied the structure, expression, and function of LEK1, a relatively large murine protein (>300 kDa) highly related to CMF1 (chicken) and CENP-F/mitosin (human) (16, 17)." cenpf-nde1-1 2 8 unspecified role unspecified role bait bait c-terminus required to bind necessary binding region required to bind deletion analysis deletion analysis certain certain sequence position certain certain certain sequence position certain 7 unspecified role unspecified role prey prey c-terminus sufficient to bind sufficient binding region sufficient to bind deletion analysis deletion analysis certain certain sequence position certain certain certain sequence position certain physical association physical association Fig 2A. Yeast two-hybrid techniques were used to determine which region within the SRR of cytLEK1 interacted with NudE by dividing the SRR into the N domain (amino acids 1906�1964), the repeat (R) domain (amino acids 1964�2071), and the C domain (amino acids 2071�2149). Only those constructs containing the C domain, not necessarily the R domain itself, retained interaction with full-length NudE (Fig. 2A). cytLEK1 interacts with NDE1. The species of origin of cytLEK1 is assumed to be mouse since in the introduction the authors "Our laboratory has studied the structure, expression, and function of LEK1, a relatively large murine protein (>300 kDa) highly related to CMF1 (chicken) and CENP-F/mitosin (human) (16, 17)." cenpf-nde1-2 3 8 unspecified role unspecified role prey prey region gfp tag green fluorescent protein tag GFP green fluorescent protein GFP green fluorescent protein gfp tag undetermined undetermined sequence position undetermined undetermined undetermined sequence position undetermined 7 unspecified role unspecified role bait bait c-terminus sufficient to bind sufficient binding region sufficient to bind deletion analysis deletion analysis certain certain sequence position certain certain certain sequence position certain region myc tag myc tag EUKLISEED epitope tag EUKLISEED epitope tag anti tag western anti tag western blot anti tag western undetermined undetermined sequence position undetermined undetermined undetermined sequence position undetermined c-terminus required to bind necessary binding region required to bind deletion analysis deletion analysis certain certain sequence position certain certain certain sequence position certain physical association physical association Fig 2B and 2C. GFP-mNudE coimmunoprecipitates with myc-SRR, confirming the two-hybrid data and demonstrating the interaction between cytLEK1 and NudE (Fig. 2B). Importantly, GFP-mNudE is unable to be coimmunoprecipitated with myc-N+R, which lacks the C domain (Fig. 2C). The C domain of cytLEK1 is required for interaction with NDE1. The species of origin of cytLEK1 is assumed to be mouse since in the introduction the authors "Our laboratory has studied the structure, expression, and function of LEK1, a relatively large murine protein (>300 kDa) highly related to CMF1 (chicken) and CENP-F/mitosin (human) (16, 17)." cenpf-nde1-3 5 8 unspecified role unspecified role neutral component neutral component 7 unspecified role unspecified role neutral component neutral component physical association physical association Fig 3C and 3D. CytLEK1 and NudE exhibited a similar distribution in our confocal studies (Fig. 3C). Deconvolution analysis of confocal images (Fig. 3D) further defined the highest amount of colocalization to be near the nucleus, as described above for the two other LIS1 pathway members. cytLEK1 colocalizes with NDE1 at the perinuclear region of the cytoplasm. cenpf-lis1 5 10 unspecified role unspecified role neutral component neutral component 7 unspecified role unspecified role neutral component neutral component physical association physical association Fig 3B. Deconvolution analysis similarly demonstrated a high degree of colocalization between cytLEK1 and LIS1, especially in perinuclear regions. (Fig. 3B). The cytLEK1 interacts with LIS1. The species of origin of cytLEK1 is assumed to be mouse since in the introduction the authors "Our laboratory has studied the structure, expression, and function of LEK1, a relatively large murine protein (>300 kDa) highly related to CMF1 (chicken) and CENP-F/mitosin (human) (16, 17)." cenpf-nde1-4 4 8 unspecified role unspecified role neutral component neutral component region gfp tag green fluorescent protein tag GFP green fluorescent protein GFP green fluorescent protein gfp tag undetermined undetermined sequence position undetermined undetermined undetermined sequence position undetermined 7 unspecified role unspecified role neutral component neutral component c-terminus required to bind necessary binding region required to bind undetermined undetermined sequence position undetermined undetermined undetermined sequence position undetermined 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 Fig 4A. Importantly, expression of the C domain alone directs this colocalization at the centrosome in COS-7 and 3T3 fibroblasts (Fig. 4A), The C-domain colocalizes with NudE in COS-7 cells. Expression of the C domain alone directs this colocalization at the centrosome in COS-7 cells. The species of origin of cytLEK1 is assumed to be mouse since in the introduction the authors "Our laboratory has studied the structure, expression, and function of LEK1, a relatively large murine protein (>300 kDa) highly related to CMF1 (chicken) and CENP-F/mitosin (human) (16, 17)." cenpf-nde1-5 4 7 unspecified role unspecified role neutral component neutral component c-terminus required to bind necessary binding region required to bind c-term range C-terminal range c-term range c-term range C-terminal range c-term range region myc tag myc tag EUKLISEED epitope tag EUKLISEED epitope tag undetermined undetermined sequence position undetermined undetermined undetermined sequence position undetermined 6 unspecified role unspecified role neutral component neutral component physical association physical association Fig 4C. Additionally, myc-C colocalizes with endogenous NudE protein present at the centrosome and perinuclear region (Fig. 4C), The C domain also colocalizes with endogenous NudE. The species of origin of cytLEK1 is assumed to be mouse since in the introduction the authors "Our laboratory has studied the structure, expression, and function of LEK1, a relatively large murine protein (>300 kDa) highly related to CMF1 (chicken) and CENP-F/mitosin (human) (16, 17)." cenpf-gammatubulin 4 7 unspecified role unspecified role neutral component neutral component c-terminus required to bind necessary binding region required to bind c-term range C-terminal range c-term range c-term range C-terminal range c-term range region myc tag myc tag EUKLISEED epitope tag EUKLISEED epitope tag undetermined undetermined sequence position undetermined undetermined undetermined sequence position undetermined 9 unspecified role unspecified role neutral component neutral component physical association physical association Fig 4B. Importantly, expression of the C domain alone directs this colocalization at the centrosome in COS-7 and 3T3 fibroblasts (Fig. 4A), as confirmed by endogenous gamma-tubulin staining (Fig. 4B). The C domain also colocalizes with endogenous gamma-tubulin. The species of origin of cytLEK1 is assumed to be mouse since in the introduction the authors "Our laboratory has studied the structure, expression, and function of LEK1, a relatively large murine protein (>300 kDa) highly related to CMF1 (chicken) and CENP-F/mitosin (human) (16, 17)." cenpf-nde1-7 1 8 unspecified role unspecified role prey prey 7 unspecified role unspecified role bait bait c-terminus cytoplasmic LEK1 (cytLEK1) sufficient to bind sufficient binding region sufficient to bind certain certain sequence position certain certain certain sequence position certain c-terminus required to bind necessary binding region required to bind deletion analysis deletion analysis certain certain sequence position certain certain certain sequence position certain physical association physical association Fig 2A. Yeast two-hybrid techniques were used to determine which region within the SRR of cytLEK1 interacted with NudE by dividing the SRR into the N domain (amino acids 1906�1964), the repeat (R) domain (amino acids 1964�2071), and the C domain (amino acids 2071�2149). Only those constructs containing the C domain, not necessarily the R domain itself, retained interaction with full-length NudE (Fig. 2A). cytLEK1 interacts with NDE1. The species of origin of cytLEK1 is assumed to be mouse since in the introduction the authors "Our laboratory has studied the structure, expression, and function of LEK1, a relatively large murine protein (>300 kDa) highly related to CMF1 (chicken) and CENP-F/mitosin (human) (16, 17)."