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/ An isoform of Arabidopsis myosin XI interacts with small GTPases in its C-terminal tail region. An isoform of Arabidopsis myosin XI interacts with small GTPases in its C-terminal tail region. J. Exp. Bot. (0022-0957) 2008 imex curation Hashimoto K., Igarashi H., Mano S., Takenaka C., Shiina T., Yamaguchi M., Demura T., Nishimura M., Shimmen T., Yokota E. yokota@sci.u-hyogo.ac.jp 14-Nov-2008: Contacted by JYOTI. Arabidopsis root cDNA library. orchard Only protein-protein interactions in vitro In vitro pull down pull down weight by comassie molecular weight estimation by coomasie staining weight by comassie Hashimoto K., Igarashi H., Mano S., Takenaka C., Shiina T., Yamaguchi M., Demura T., Nishimura M., Shimmen T., Yokota E. After staining with Coomassie brilliant blue (CBB), the intensity of each band of Rab proteins was measured using a densitometer. yokota@sci.u-hyogo.ac.jp Accepted 2008-NOV-15 by ORCHARD. 14-Nov-2008: Contacted by JYOTI. Arabidopsis root cDNA library. J. Exp. Bot. (0022-0957) 2008 orchard Only protein-protein interactions imex curation An isoform of Arabidopsis myosin XI interacts with small GTPases in its C-terminal tail region. An isoform of Arabidopsis myosin XI interacts with small GTPases in its C-terminal tail region. J. Exp. Bot. (0022-0957) 2008 imex curation Hashimoto K., Igarashi H., Mano S., Takenaka C., Shiina T., Yamaguchi M., Demura T., Nishimura M., Shimmen T., Yokota E. yokota@sci.u-hyogo.ac.jp 14-Nov-2008: Contacted by JYOTI. Arabidopsis root cDNA library. orchard 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 Hashimoto K., Igarashi H., Mano S., Takenaka C., Shiina T., Yamaguchi M., Demura T., Nishimura M., Shimmen T., Yokota E. yokota@sci.u-hyogo.ac.jp Accepted 2008-NOV-15 by ORCHARD. Arabidopsis root cDNA library. J. Exp. Bot. (0022-0957) 2008 orchard Only protein-protein interactions imex curation An isoform of Arabidopsis myosin XI interacts with small GTPases in its C-terminal tail region. An isoform of Arabidopsis myosin XI interacts with small GTPases in its C-terminal tail region. J. Exp. Bot. (0022-0957) 2008 imex curation Hashimoto K., Igarashi H., Mano S., Takenaka C., Shiina T., Yamaguchi M., Demura T., Nishimura M., Shimmen T., Yokota E. yokota@sci.u-hyogo.ac.jp 14-Nov-2008: Contacted by JYOTI. Arabidopsis root cDNA library. orchard 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 Hashimoto K., Igarashi H., Mano S., Takenaka C., Shiina T., Yamaguchi M., Demura T., Nishimura M., Shimmen T., Yokota E. yokota@sci.u-hyogo.ac.jp Accepted 2008-NOV-15 by ORCHARD. 14-Nov-2008: Contacted by JYOTI. Arabidopsis root cDNA library. J. Exp. Bot. (0022-0957) 2008 orchard Only protein-protein interactions imex curation myo6_arath Myosin-6 XI-2 AtMYA2 MYA2 F6B6.4 At5g43900 protein protein arath Arabidopsis thaliana (Mouse-ear cress) Mouse-ear cress MVANFNPSVGSFVWVEDPDEAWIDGEVVQVNGDEIKVLCTSGKHVVTKISNAYPKDVEAPASGVDDMTRLAYLHEPGVLQNLHSRYDINEIYTYTGSILIAVNPFRRLPHLYSSHMMAQYKGASLGELSPHPFAVADAAYRQMINDGVSQSILVSGESGAGKTESTKLLMRYLAYMGGRAAAEGRSVEQKVLESNPVLEAFGNAKTVRNNNSSRFGKFVEIQFDEKGRISGAAIRTYLLERSRVCQVSDPERNYHCFYMLCAAPQEDVKKFKLEEPKKYHYLNQSKCLELDSINDAEEYHATRRAMDVVGISTEEQDAIFSVVAAILHIGNIEFAKGEEIDSSIPKDDKSLFHLKTAAELLSCDEKALEDSLCKRIMVTRDETITKTLDPEAATLSRDALAKVMYSRLFDWLVDKINSSIGQDHDSKYLIGVLDIYGFESFKTNSFEQFCINLTNEKLQQHFNQHVFKMEQEEYKKEEINWSYIEFVDNQDILDLIEKKPGGIIALLDEACMFPRSTHETFAQKLYQTFKTHKRFTKPKLARSDFTICHYAGDVTYQTELFLDKNKDYVIAEHQALLNSSSCSFVASLFPPMSDDSKQSKFSSIGTRFKQQLVSLLEILNTTEPHYIRCIKPNNLLKPGIFENENILQQLRCGGVMEAIRISCAGYPTRKHFDEFLARFGILAPEVLVKNSDDPAACKKLLDKVGLEGYQIGKTKVFLRAGQMADLDTRRTEVLGRSASIIQRKVRSYLAKKSFIVLRNSAKQIQSVCRGYLARSVYEGMRREAAALKIQRDLRRFLARKAYTELYSAAVSVQAGMRGMVARKELCFRRQTKAAIIIQTWCRGYLARLHYRKLKKAAITTQCAWRSKVARGELRKLKMAARETGALQAAKNKLEKQVEELTWRLQLEKRIRTDLEEAKKQESAKAQSSLEELQLKCKETEALLIKEREAAKKIAETAPIIKEIPVVDQELMDKITNENEKLKSMVSSLEMKIGETEKKLQETTKISQDRLNQALEAESKLVKLKTAMQRLEEKILDMEAEKKIMHQQTISTPVRTNLGHPPTAPVKNLENGHQTNLEKEFNEAEFTTPVDGKAGKSAAERQIMNVDALIDCVKDNIGFSNGKPVAAFTIYKCLLHWKCFESEKTNVFDRLIQMIGSAIENEDDNSHLAYWLTSTSALLFLLQKSLKTNGSGATQSKKPPASTSLFGRMAMSFRSSPASGNLAAAAEAAALAVVRPVEAKYPALLFKQQLAAYVEKMFGMVRDNLKRELSTLLSLCIQAPRSSKGGMLRSGRSFGKDSPAVHWQSIIDGLNSLLVTLKENHVPLVLIQKIYSQTFSYINVQLFNSLLLRKECCTFSNGEFVKSGLAELELWCCQAKEYSGPSWEELKHIRQAVGFLVIHQKYRISYDEIANDLCPVLSVQQLYRICTLYWDDSYNTRSVSQEVISSMRTLMTEESNDADSDSFLLDDDSSIPFSIDDISSSMEEKDFVGIKPAEELLENPAFVFLH B344E4668DEC6A8D rabd1_arath Ras-related protein RABD1 RABD1 ATFP8 Ras-related protein ATFP8 F26K24.2 At3g11730 protein protein arath Arabidopsis thaliana (Mouse-ear cress) Mouse-ear cress MSNEYDYLFKLLLIGDSSVGKSCLLLRFADDAYIDSYISTIGVDFKIRTIEQDGKTIKLQIWDTAGQERFRTITSSYYRGAHGIIIVYDCTEMESFNNVKQWLSEIDRYANESVCKLLIGNKNDMVESKVVSTETGRALADELGIPFLETSAKDSINVEQAFLTIAGEIKKKMGSQTNANKTSGPGTVQMKGQPIQQNNGGCCGQ 8842C2B074B71AAD rac2a_arath Ras-related protein RABC2a RABC2A RAB18B At5g03530 Ras-related protein Rab18B F12E4.310 protein protein arath Arabidopsis thaliana (Mouse-ear cress) Mouse-ear cress MGSSSGQSGYDLSFKILLIGDSGVGKSSLLVSFISSSVEDLAPTIGVDFKIKQLTVGGKRLKLTIWDTAGQERFRTLTSSYYRGAQGIILVYDVTRRETFTNLVDVWGKEIELYSTNQECVRMLVGNKVDRESERGVSREEGIALAKELNCMFLECSARTRQNVEQCFEELALKIMEVPSLLEEGSSAVKRNILKQKPEHQTNTQSGCCS 551337D936809B5B mya2-rabd1 2 4 unspecified role unspecified role bait bait c-terminal sufficient to bind sufficient binding region sufficient to bind certain certain sequence position certain certain certain sequence position certain 5 unspecified role unspecified role prey prey physical association physical association An Arabidopsis root cell cDNA library with the tail region of MYA2, aa1049-aa1505 (f1), lacking the coiled-0coil domain, as shown schematically in Fig. 1A was screened. After screening 1.2x106 clones, DNA fragments encoding AtRabD1 and AtRabC2a, members of the small GTPase Rab family, were identified. mya2-rabc2a 2 4 unspecified role unspecified role bait bait c-terminal sufficient to bind sufficient binding region sufficient to bind certain certain sequence position certain certain certain sequence position certain 6 unspecified role unspecified role prey prey physical association physical association An Arabidopsis root cell cDNA library with the tail region of MYA2, aa1049-aa1505 (f1), lacking the coiled-0coil domain, as shown schematically in Fig. 1A was screened. After screening 1.2x106 clones, DNA fragments encoding AtRabD1 and AtRabC2a, members of the small GTPase Rab family, were identified. mya2-rabd1-1 3 4 unspecified role unspecified role bait bait region sufficient to bind sufficient binding region sufficient to bind certain certain sequence position certain certain certain sequence position certain 5 unspecified role unspecified role prey prey physical association physical association To determine the region in the MYA2 tail responsible for the interaction with each AtRab, their interaction with a series of deletion forms of the MYA2 tail, aa1049�aa1450 (f2 in Fig. 1A), aa900�aa1221 (f3 in Fig. 1A), in which the coiled-coil domain was included, and aa1195�aa1505 (f4 in Fig. 1A) was examined. The association of each AtRab to each truncation of the MYA2 tail is summarized in Fig. 1B. AtRabD1 interacted with a truncation of MYA2 tail, aa1049�aa1450 (f2) lacking 55 amino acids in the C-terminus Figure 1 mya2-rabc2a-1 3 6 unspecified role unspecified role prey prey 4 unspecified role unspecified role bait bait c-terminal required to bind necessary binding region required to bind certain certain sequence position certain certain certain sequence position certain physical association physical association By contrast, AtRabC2a interacted only with a full-length MYA2 tail (f1), but not with its truncated forms, where 55 amino acids from the C-terminus or 146 amino acids from N-terminus were deleted. Therefore, both tail termini appear to be necessary for the interaction with AtRabC2a. The authors note that in contrast to RabD1, "RabC2a interacted only with a full-length MYA2 tail (f1), but not with its truncated forms, where 55 amino acids from the C-terminus or 146 amino acids from N-terminus were deleted. Therefore, both tail termini appear to be necessary for the interaction with AtRabC2a". Figure 1 mya2-rabd1-2 1 4 unspecified role unspecified role bait bait region sufficient to bind sufficient binding region sufficient to bind certain certain sequence position certain certain certain sequence position certain 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 ecoli Escherichia coli (strain K12) 5 unspecified role unspecified role prey prey 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 ecoli Escherichia coli (strain K12) direct interaction direct interaction These results indicate that AtRabD1 and AtRabC2a protein tagged with His6 interacted weakly with the MYA2 tails fused with GST in a GTP dependent manner. Figure 4 Binding of RabD1 (Q9ZRE2) protein bound to the MYA protein region in the presence of GTP 2.6 fold higher than when GTP was not present. Guanosine-5'-triphosphate (GTP) mya2-rabc2a-2 1 4 unspecified role unspecified role bait bait c-terminal sufficient to bind sufficient binding region sufficient to bind certain certain sequence position certain certain certain sequence position certain 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 6 unspecified role unspecified role prey prey 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 physical association physical association These results indicate that AtRabD1 and AtRabC2a protein tagged with His6 interacted weakly with the MYA2 tails fused with GST in a GTP dependent manner. The authors note that RabC2a only displayed GTP dependent interaction with the c-terminus of the MYA2 protein (1049 to 1505 amino acids) and not the region 1405-1450 amino acids. They state that the binding of the AtRabC2a protein bound to the MYA2 tail (1049-1505 amino acids) coated beads in the presence of GTP 1.7-fold higher than that in the absence of GTP. Figure 2 Figure 3 Figure 4 Guanosine-5'-triphosphate (GTP)