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/ http://www.ebi.ac.uk/ inada-2001-2 Keratin attenuates tumor necrosis factor-induced cytotoxicity through association with TRADD. 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 Inada H., Izawa I., Nishizawa M., Fujita E., Kiyono T., Takahashi T., Momoi T., Inagaki M. Apoptosis - Interactions involving proteins with a function related to apoptosis minagaki@aichi-cc.jp A human liver cDNA library was used. J. Cell Biol. (0021-9525) 2001 inada-2001-8 Keratin attenuates tumor necrosis factor-induced cytotoxicity through association with TRADD. human-mammary epithelium Human mammary epithelium mammary_epithelium Mammary epithelium coip coimmunoprecipitation co-immunoprecipitation CoIp immunoprecipitation Co-IP co-immunoprecipitation CoIp immunoprecipitation Co-IP coip western blot western blot Immuno blot Immuno blot Inada H., Izawa I., Nishizawa M., Fujita E., Kiyono T., Takahashi T., Momoi T., Inagaki M. Apoptosis - Interactions involving proteins with a function related to apoptosis minagaki@aichi-cc.jp A human liver cDNA library was used. J. Cell Biol. (0021-9525) 2001 inada-2001-3 Keratin attenuates tumor necrosis factor-induced cytotoxicity through association with TRADD. in vitro In vitro cosedimentation cosedimentation predetermined predetermined participant predetermined Inada H., Izawa I., Nishizawa M., Fujita E., Kiyono T., Takahashi T., Momoi T., Inagaki M. Apoptosis - Interactions involving proteins with a function related to apoptosis minagaki@aichi-cc.jp A human liver cDNA library was used. J. Cell Biol. (0021-9525) 2001 inada-2001-9 Keratin attenuates tumor necrosis factor-induced cytotoxicity through association with TRADD. human-sw13 human sw13 cells from bone sw13 SW1353 Species: human, Caucasian female 72 years old; Tissue: bone; Tumor: chondrosarcoma fluorescence imaging fluorescence microscopy fluorescence imaging immunostaining immunostaining Inada H., Izawa I., Nishizawa M., Fujita E., Kiyono T., Takahashi T., Momoi T., Inagaki M. Apoptosis - Interactions involving proteins with a function related to apoptosis minagaki@aichi-cc.jp A human liver cDNA library was used. J. Cell Biol. (0021-9525) 2001 inada-2001-1 Keratin attenuates tumor necrosis factor-induced cytotoxicity through association with TRADD. 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 Inada H., Izawa I., Nishizawa M., Fujita E., Kiyono T., Takahashi T., Momoi T., Inagaki M. Apoptosis - Interactions involving proteins with a function related to apoptosis minagaki@aichi-cc.jp A human liver cDNA library was used. J. Cell Biol. (0021-9525) 2001 inada-2001-4 Keratin attenuates tumor necrosis factor-induced cytotoxicity through association with TRADD. human-hela Homo sapiens epitheloid cervix carcinoma cells Human hela Human epitheloid cervix carcinoma HeLa cells derivation: cervix; epithelial; adenocarcinoma ATCC no:CCL-2 fluorescence imaging fluorescence microscopy fluorescence imaging immunostaining immunostaining Inada H., Izawa I., Nishizawa M., Fujita E., Kiyono T., Takahashi T., Momoi T., Inagaki M. Apoptosis - Interactions involving proteins with a function related to apoptosis minagaki@aichi-cc.jp A human liver cDNA library was used. J. Cell Biol. (0021-9525) 2001 inada-2001-5 Keratin attenuates tumor necrosis factor-induced cytotoxicity through association with TRADD. human-t24 Homo sapiens T24 bladder carcinoma cells t24 Human bladder carcinoma fluorescence imaging fluorescence microscopy fluorescence imaging immunostaining immunostaining Inada H., Izawa I., Nishizawa M., Fujita E., Kiyono T., Takahashi T., Momoi T., Inagaki M. Apoptosis - Interactions involving proteins with a function related to apoptosis minagaki@aichi-cc.jp A human liver cDNA library was used. J. Cell Biol. (0021-9525) 2001 inada-2001-6 Keratin attenuates tumor necrosis factor-induced cytotoxicity through association with TRADD. canlf-mdck Canis familiaris (dog) kidney epithelial cells Canis lupus familiaris Dog Canis familiaris mdck Canine Cocker Spaniel kidney epithelial cell line fluorescence imaging fluorescence microscopy fluorescence imaging immunostaining immunostaining Inada H., Izawa I., Nishizawa M., Fujita E., Kiyono T., Takahashi T., Momoi T., Inagaki M. Apoptosis - Interactions involving proteins with a function related to apoptosis minagaki@aichi-cc.jp A human liver cDNA library was used. J. Cell Biol. (0021-9525) 2001 inada-2001-7 Keratin attenuates tumor necrosis factor-induced cytotoxicity through association with TRADD. human-mammary epithelium Human mammary epithelium mammary_epithelium Mammary epithelium fluorescence imaging fluorescence microscopy fluorescence imaging immunostaining immunostaining Inada H., Izawa I., Nishizawa M., Fujita E., Kiyono T., Takahashi T., Momoi T., Inagaki M. Apoptosis - Interactions involving proteins with a function related to apoptosis minagaki@aichi-cc.jp A human liver cDNA library was used. J. Cell Biol. (0021-9525) 2001 k1c18_human Keratin, type I cytoskeletal 18 KRT18 CYK18 Keratin-18 PIG46 Cytokeratin-18 Cell proliferation-inducing gene 46 protein protein protein human Homo sapiens Human MSFTTRSTFSTNYRSLGSVQAPSYGARPVSSAASVYAGAGGSGSRISVSRSTSFRGGMGSGGLATGIAGGLAGMGGIQNEKETMQSLNDRLASYLDRVRSLETENRRLESKIREHLEKKGPQVRDWSHYFKIIEDLRAQIFANTVDNARIVLQIDNARLAADDFRVKYETELAMRQSVENDIHGLRKVIDDTNITRLQLETEIEALKEELLFMKKNHEEEVKGLQAQIASSGLTVEVDAPKSQDLAKIMADIRAQYDELARKNREELDKYWSQQIEESTTVVTTQSAEVGAAETTLTELRRTVQSLEIDLDSMRNLKASLENSLREVEARYALQMEQLNGILLHLESELAQTRAEGQRQAQEYEALLNIKVKLEAEIATYRRLLEDGEDFNLGDALDSSNSMQTIQKTTTRRIVDGKVVSETNDTKVLRH tradd_human Tumor necrosis factor receptor type 1-associated DEATH domain protein TRADD TNFRSF1A-associated via death domain protein protein human Homo sapiens Human MAAGQNGHEEWVGSAYLFVESSLDKVVLSDAYAHPQQKVAVYRALQAALAESGGSPDVLQMLKIHRSDPQLIVQLRFCGRQPCGRFLRAYREGALRAALQRSLAAALAQHSVPLQLELRAGAERLDALLADEERCLSCILAQQPDRLRDEELAELEDALRNLKCGSGARGGDGEVASAPLQPPVPSLSEVKPPPPPPPAQTFLFQGQPVVNRPLSLKDQQTFARSVGLKWRKVGRSLQRGCRALRDPALDSLAYEYEREGLYEQAFQLLRRFVQAEGRRATLQRLVEALEENELTSLAEDLLGLTDPNGGLA k1c14_human Keratin, type I cytoskeletal 14 Cytokeratin-14 KRT14 Keratin-14 protein protein human Homo sapiens Human MTTCSRQFTSSSSMKGSCGIGGGIGGGSSRISSVLAGGSCRAPSTYGGGLSVSSSRFSSGGACGLGGGYGGGFSSSSSSFGSGFGGGYGGGLGAGLGGGFGGGFAGGDGLLVGSEKVTMQNLNDRLASYLDKVRALEEANADLEVKIRDWYQRQRPAEIKDYSPYFKTIEDLRNKILTATVDNANVLLQIDNARLAADDFRTKYETELNLRMSVEADINGLRRVLDELTLARADLEMQIESLKEELAYLKKNHEEEMNALRGQVGGDVNVEMDAAPGVDLSRILNEMRDQYEKMAEKNRKDAEEWFFTKTEELNREVATNSELVQSGKSEISELRRTMQNLEIELQSQLSMKASLENSLEETKGRYCMQLAQIQEMIGSVEEQLAQLRCEMEQQNQEYKILLDVKTRLEQEIATYRRLLEGEDAHLSSSQFSSGSQSSRDVTSSSRQIRTKVMDVHDGKVVSTHEQVLRTKN tnr1a_human Tumor necrosis factor receptor superfamily member 1A TNFRSF1A TNFAR TNFR1 p60 Tumor necrosis factor receptor 1 Tumor necrosis factor receptor type I p55 protein protein human Homo sapiens Human MGLSTVPDLLLPLVLLELLVGIYPSGVIGLVPHLGDREKRDSVCPQGKYIHPQNNSICCTKCHKGTYLYNDCPGPGQDTDCRECESGSFTASENHLRHCLSCSKCRKEMGQVEISSCTVDRDTVCGCRKNQYRHYWSENLFQCFNCSLCLNGTVHLSCQEKQNTVCTCHAGFFLRENECVSCSNCKKSLECTKLCLPQIENVKGTEDSGTTVLLPLVIFFGLCLLSLLFIGLMYRYQRWKSKLYSIVCGKSTPEKEGELEGTTTKPLAPNPSFSPTPGFTPTLGFSPVPSSTFTSSSTYTPGDCPNFAAPRREVAPPYQGADPILATALASDPIPNPLQKWEDSAHKPQSLDTDDPATLYAVVENVPPLRWKEFVRRLGLSDHEIDRLELQNGRCLREAQYSMLATWRRRTPRREATLELLGRVLRDMDLLGCLEDIEEALCGPAALPPAPSLLR k2c8_human Keratin, type II cytoskeletal 8 Keratin-8 KRT8 CYK8 Cytokeratin-8 Type-II keratin Kb8 protein protein human Homo sapiens Human MSIRVTQKSYKVSTSGPRAFSSRSYTSGPGSRISSSSFSRVGSSNFRGGLGGGYGGASGMGGITAVTVNQSLLSPLVLEVDPNIQAVRTQEKEQIKTLNNKFASFIDKVRFLEQQNKMLETKWSLLQQQKTARSNMDNMFESYINNLRRQLETLGQEKLKLEAELGNMQGLVEDFKNKYEDEINKRTEMENEFVLIKKDVDEAYMNKVELESRLEGLTDEINFLRQLYEEEIRELQSQISDTSVVLSMDNSRSLDMDSIIAEVKAQYEDIANRSRAEAESMYQIKYEELQSLAGKHGDDLRRTKTEISEMNRNISRLQAEIEGLKGQRASLEAAIADAEQRGELAIKDANAKLSELEAALQRAKQDMARQLREYQELMNVKLALDIEIATYRKLLEGEESRLESGMQNMSIHTKTTSGYAGGLSSAYGGLTSPGLSYSLGSSFGSGAGSSSFSRTSSSRAVVVKKIETRDGKLVSESSDVLPK tradd-krt18-1 Y2H interaction between tradd and krt18 655238 N/A 655247 unspecified role unspecified role prey prey N/A 655248 unspecified role unspecified role bait bait c-terminal sufficient to bind sufficient binding region sufficient to bind predetermined featur predetermined feature predetermined featur certain certain sequence position certain certain certain sequence position certain false physical association physical association false false false Both the c-terminal and full-length Tradd were able to interact with Krt18 but there was strongest interaction with the Tradd c-terminal fragment. Figure 1B tradd-krt14 Y2H interaction between tradd and krt14 655238 N/A 655249 unspecified role unspecified role prey prey N/A 655248 unspecified role unspecified role bait bait c-terminal sufficient to bind sufficient binding region sufficient to bind predetermined featur predetermined feature predetermined featur certain certain sequence position certain certain certain sequence position certain false physical association physical association false false false Both the c-terminal and full-length Tradd were able to interact with Krt14 but there was strongest interaction with the Tradd c-terminal fragment. Figure 1B tradd-krt18-2 Y2H interaction between tradd and krt18 655238 N/A 655247 unspecified role unspecified role bait bait n-terminal required to bind necessary binding region required to bind deletion analysis deletion analysis certain certain sequence position certain certain certain sequence position certain false N/A 655248 unspecified role unspecified role prey prey c-terminal sufficient to bind sufficient binding region sufficient to bind predetermined featur predetermined feature predetermined featur certain certain sequence position certain certain certain sequence position certain false physical association physical association false false false The coil Ia region of the rod domain of Krt18 specifically interacted with both full-length TRADD (TRADD-F) and the c-terminal region of TRADD (TRADD-C). Figure 1C tradd-krt18-8 Coimmunoprecipitation showing interaction between human tradd and krt18 655239 N/A 655248 unspecified role unspecified role bait bait human-mammary epithelium Human mammary epithelium mammary_epithelium Mammary epithelium N/A 655247 unspecified role unspecified role prey prey human-mammary epithelium Human mammary epithelium mammary_epithelium Mammary epithelium physical association physical association false false false K18 was coimmunoprecipitated with TRADD, using the anti-TRADD antibody. This coprecipitation was barely visible in the 100 ng/ml TNF-treated HMEC samples suggesting that TRADD is specifically dissociated from K18 by TNF. Figure 4B, 5B, 6B tradd-tradd-3 Coimmunoprecipitation showing interaction between human tradd and itself 655239 N/A 655248 unspecified role unspecified role prey prey human-mammary epithelium Human mammary epithelium mammary_epithelium Mammary epithelium N/A 655248 unspecified role unspecified role bait bait human-mammary epithelium Human mammary epithelium mammary_epithelium Mammary epithelium physical association physical association false false false TRADD was coimmunoprecipitated with TRADD, using the anti-TRADD antibody. Figure 4B tradd-tnfr1 Coimmunoprecipitation showing interaction between human tradd and tnfr1 655239 N/A 655248 unspecified role unspecified role bait bait human-mammary epithelium Human mammary epithelium mammary_epithelium Mammary epithelium N/A 655250 unspecified role unspecified role prey prey physical association physical association false false false The coprecipitation of K18 with TRADD was dissociated by TNF suggesting that TRADD binds with TNFR1. Figure 5B krt8-krt18-3 Cosedimentation assay showing interaction between human krt18 and krt8 655240 N/A 655247 unspecified role unspecified role neutral component neutral component in vitro In vitro N/A 655251 unspecified role unspecified role neutral component neutral component in vitro In vitro physical association physical association false false false Krt18 assembled with K8 independantly of GST or GST-Tradd. Figure 2A, 2B tradd-krt18-3 Cosedimentation assay showing interaction between human tradd and krt18/krt8 filaments 655240 N/A 655248 unspecified role unspecified role neutral component neutral component 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 false ecoli Escherichia coli (strain K12) N/A 655247 unspecified role unspecified role neutral component neutral component in vitro In vitro N/A 655251 unspecified role unspecified role neutral component neutral component in vitro In vitro physical association physical association false false false Because Tradd interacted with K18, but not with K8, in the two-hybrid analysis, the Tradd cosedimention with the K8/18 polymer via direct binding to K18. In addition, Tradd did not sediment with vimentin. Figure 2A, 2B tradd-krt18-9 Colocalization showing interaction between human tradd and krt18 655241 N/A 655247 unspecified role unspecified role 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 false human-sw13 human sw13 cells from bone sw13 SW1353 Species: human, Caucasian female 72 years old; Tissue: bone; Tumor: chondrosarcoma N/A 655248 unspecified role unspecified role neutral component neutral component human-sw13 human sw13 cells from bone sw13 SW1353 Species: human, Caucasian female 72 years old; Tissue: bone; Tumor: chondrosarcoma N/A 655251 unspecified role unspecified role 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 false human-sw13 human sw13 cells from bone sw13 SW1353 Species: human, Caucasian female 72 years old; Tissue: bone; Tumor: chondrosarcoma colocalization colocalization false false false In K8/ 18-transfected SW13 cells, TRADD was recruited to overexpressed K8/18 filaments Figure 7C tradd-krt18-10 Colocalization showing interaction between human tradd and krt18 655241 N/A 655248 unspecified role unspecified role prey prey human-sw13 human sw13 cells from bone sw13 SW1353 Species: human, Caucasian female 72 years old; Tissue: bone; Tumor: chondrosarcoma N/A 655247 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 false n-terminus sufficient to bind sufficient binding region sufficient to bind certain certain sequence position certain certain certain sequence position certain false human-sw13 human sw13 cells from bone sw13 SW1353 Species: human, Caucasian female 72 years old; Tissue: bone; Tumor: chondrosarcoma colocalization colocalization false false false In cells overexpressing the NH2 terminus of K18, TRADD markedly colocalized with the overexpressed NH2 terminus Figure 7C tradd-krt18 Y2H interaction between tradd and krt18 655242 N/A 655248 unspecified role unspecified role prey prey c-terminal sufficient to bind sufficient binding region sufficient to bind full dna sequence full identification by DNA sequencing full dna sequence certain certain sequence position certain certain certain sequence position certain false N/A 655247 unspecified role unspecified role bait bait physical association physical association false false false Both Krt8 and Krt18 were used as bait on the human liver cDNA library, but only Krt18 picked up the Tradd c-terminal fragment. Figure 1A. tradd-krt18-4 Colocalization showing interaction between human tradd and krt18 655243 N/A 655247 unspecified role unspecified role prey prey human-hela Homo sapiens epitheloid cervix carcinoma cells Human hela Human epitheloid cervix carcinoma HeLa cells derivation: cervix; epithelial; adenocarcinoma ATCC no:CCL-2 N/A 655248 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 false human-hela Homo sapiens epitheloid cervix carcinoma cells Human hela Human epitheloid cervix carcinoma HeLa cells derivation: cervix; epithelial; adenocarcinoma ATCC no:CCL-2 colocalization colocalization false false false HeLa cells were fixed and doubly stained with anti-myc polyclonal antibody and anti-K18 monoclonal antibody. Overexpressed myc�TRADD exhibited a filamentous pattern, and these myc�TRADD filaments markedly colocalized with K8/18 filaments. Figure 3A tradd-krt18-5 Colocalization showing interaction between human tradd and krt18 655244 N/A 655247 unspecified role unspecified role prey prey human-t24 Homo sapiens T24 bladder carcinoma cells t24 Human bladder carcinoma N/A 655248 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 false human-t24 Homo sapiens T24 bladder carcinoma cells t24 Human bladder carcinoma colocalization colocalization false false false T24 cells were fixed and doubly stained with anti-myc polyclonal antibody and anti-K18 monoclonal antibody. Overexpressed myc�TRADD exhibited a filamentous pattern, and these myc�TRADD filaments markedly colocalized with K8/18 filaments. Figure 3A tradd-krt18-6 Colocalization showing interaction between human tradd and krt18 655245 N/A 655248 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 false canlf-mdck Canis familiaris (dog) kidney epithelial cells Canis lupus familiaris Dog Canis familiaris mdck Canine Cocker Spaniel kidney epithelial cell line N/A 655247 unspecified role unspecified role prey prey canlf-mdck Canis familiaris (dog) kidney epithelial cells Canis lupus familiaris Dog Canis familiaris mdck Canine Cocker Spaniel kidney epithelial cell line colocalization colocalization false false false MDCK cells were fixed and doubly stained with anti-myc polyclonal antibody and anti-K18 monoclonal antibody. Overexpressed myc�TRADD exhibited a filamentous pattern, and these myc�TRADD filaments markedly colocalized with K8/18 filaments. Figure 3A tradd-krt18-7 Colocalization showing interaction between human tradd and krt18 655246 N/A 655247 unspecified role unspecified role prey prey human-mammary epithelium Human mammary epithelium mammary_epithelium Mammary epithelium N/A 655248 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 false human-mammary epithelium Human mammary epithelium mammary_epithelium Mammary epithelium colocalization colocalization false false false In immortalized human mammary epithelial cells (HMEC) myc�TRADD filaments markedly colocalized with K8/18 filaments. Figure 3A, 4C, 5A, 6A To further confirm the colocalization of TRADD with K18, HMEC cells were transfected with a K8 deletion mutant to disrupt K8/18 filaments. Cells expressing the myc-tagged K8 deletion mutant (aa 1-239 of K8) showed disorganization of K8/18 filaments and the disappearance of the filamentous distribution of TRADD. These results confirm the colocalization of TRADD with K8/18 filaments, and suggest that K8/18 filament organization affects the subcellular distribution of TRADD. On treatment with TNF, TRADD dissociated from K8/18 filaments, and dispersed diffusely in the cytoplasm. Other stimuli did not alter subcellular distribution of TRADD.