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/ Regulators of PP2C phosphatase activity function as abscisic acid sensors. Regulators of PP2C phosphatase activity function as abscisic acid sensors. Science (0036-8075) 2009 imex curation Ma Y., Szostkiewicz I., Korte A., Moes D., Yang Y., Christmann A., Grill E. erwin.grill@wzw.tum.de 01-Sep-2009: Contacted by JYOTI. 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 Ma Y., Szostkiewicz I., Korte A., Moes D., Yang Y., Christmann A., Grill E. erwin.grill@wzw.tum.de Accepted 2009-JUL-9 by ORCHARD. 01-Sep-2009: Contacted by JYOTI. Science (0036-8075) 2009 orchard Only protein-protein interactions imex curation Regulators of PP2C phosphatase activity function as abscisic acid sensors. Regulators of PP2C phosphatase activity function as abscisic acid sensors. Science (0036-8075) 2009 imex curation Ma Y., Szostkiewicz I., Korte A., Moes D., Yang Y., Christmann A., Grill E. erwin.grill@wzw.tum.de 01-Sep-2009: Contacted by JYOTI. orchard Only protein-protein interactions arath Arabidopsis thaliana (Mouse-ear cress) Mouse-ear cress bifc bimolecular fluorescence complementation bifc tag fluorescence tag visualisation by fluorescence tag fluorescence Ma Y., Szostkiewicz I., Korte A., Moes D., Yang Y., Christmann A., Grill E. Expression constructs were introduced into protoplasts prepared from Arabidopsis thaliana plants. erwin.grill@wzw.tum.de Accepted 2009-JUL-9 by ORCHARD. 01-Sep-2009: Contacted by JYOTI. Science (0036-8075) 2009 orchard Only protein-protein interactions imex curation Regulators of PP2C phosphatase activity function as abscisic acid sensors. Regulators of PP2C phosphatase activity function as abscisic acid sensors. Science (0036-8075) 2009 imex curation Ma Y., Szostkiewicz I., Korte A., Moes D., Yang Y., Christmann A., Grill E. erwin.grill@wzw.tum.de 01-Sep-2009: Contacted by JYOTI. orchard Only protein-protein interactions in vitro In vitro itc isothermal titration calorimetry ITC ITC itc predetermined predetermined participant predetermined Ma Y., Szostkiewicz I., Korte A., Moes D., Yang Y., Christmann A., Grill E. erwin.grill@wzw.tum.de Accepted 2009-JUL-9 by ORCHARD. 01-Sep-2009: Contacted by JYOTI. Science (0036-8075) 2009 orchard Only protein-protein interactions imex curation pyl9_arath Abscisic acid receptor PYL9 At1g01360 PYL9 RCAR1 PYR1-like protein 9 ABI1-binding protein 4 Regulatory components of ABA receptor 1 F6F3.16 protein protein arath Arabidopsis thaliana (Mouse-ear cress) Mouse-ear cress MMDGVEGGTAMYGGLETVQYVRTHHQHLCRENQCTSALVKHIKAPLHLVWSLVRRFDQPQKYKPFVSRCTVIGDPEIGSLREVNVKSGLPATTSTERLELLDDEEHILGIKIIGGDHRLKNYSSILTVHPEIIEGRAGTMVIESFVVDVPQGNTKDETCYFVEALIRCNLKSLADVSERLASQDITQ DIP protein Q84MC7 original sequence version: 69 070D485311163568 p2c56_arath Protein phosphatase 2C 56 Protein ABSCISIC ACID-INSENSITIVE 1 ABI1 At4g26080 F20B18.190 Protein phosphatase 2C ABI1 protein protein arath Arabidopsis thaliana (Mouse-ear cress) Mouse-ear cress MEEVSPAIAGPFRPFSETQMDFTGIRLGKGYCNNQYSNQDSENGDLMVSLPETSSCSVSGSHGSESRKVLISRINSPNLNMKESAAADIVVVDISAGDEINGSDITSEKKMISRTESRSLFEFKSVPLYGFTSICGRRPEMEDAVSTIPRFLQSSSGSMLDGRFDPQSAAHFFGVYDGHGGSQVANYCRERMHLALAEEIAKEKPMLCDGDTWLEKWKKALFNSFLRVDSEIESVAPETVGSTSVVAVVFPSHIFVANCGDSRAVLCRGKTALPLSVDHKPDREDEAARIEAAGGKVIQWNGARVFGVLAMSRSIGDRYLKPSIIPDPEVTAVKRVKEDDCLILASDGVWDVMTDEEACEMARKRILLWHKKNAVAGDASLLADERRKEGKDPAAMSAAEYLSKLAIQRGSKDNISVVVVDLKPRRKLKSKPLN DIP protein P49597 original sequence version: 104 DIP protein P49597 original sequence version: 101 DIP protein P49597 original sequence version: 106 DIP protein P49597 original sequence version: 102 DIP protein P49597 original sequence version: 116 DIP protein P49597 original sequence version: 107 DIP protein P49597 original sequence version: 124 DIP protein P49597 original sequence version: 161 4A4C54F04195F572 p2c16_arath Protein phosphatase 2C 16 HAB1 P2C-HA Protein phosphatase 2C HAB1 AtP2C-HA Protein HYPERSENSITIVE TO ABA 1 F28P22.4 At1g72770 protein protein arath Arabidopsis thaliana (Mouse-ear cress) Mouse-ear cress MEEMTPAVAMTLSLAANTMCESSPVEITQLKNVTDAADLLSDSENQSFCNGGTECTMEDVSELEEVGEQDLLKTLSDTRSGSSNVFDEDDVLSVVEDNSAVISEGLLVVDAGSELSLSNTAMEIDNGRVLATAIIVGESSIEQVPTAEVLIAGVNQDTNTSEVVIRLPDENSNHLVKGRSVYELDCIPLWGTVSIQGNRSEMEDAFAVSPHFLKLPIKMLMGDHEGMSPSLTHLTGHFFGVYDGHGGHKVADYCRDRLHFALAEEIERIKDELCKRNTGEGRQVQWDKVFTSCFLTVDGEIEGKIGRAVVGSSDKVLEAVASETVGSTAVVALVCSSHIVVSNCGDSRAVLFRGKEAMPLSVDHKPDREDEYARIENAGGKVIQWQGARVFGVLAMSRSIGDRYLKPYVIPEPEVTFMPRSREDECLILASDGLWDVMNNQEVCEIARRRILMWHKKNGAPPLAERGKGIDPACQAAADYLSMLALQKGSKDNISIIVIDLKAQRKFKTRT DIP protein Q9CAJ0 original sequence version: 64 DIP protein Q9CAJ0-1 original sequence version: 62 DIP protein Q9CAJ0-1 original sequence version: 77 DIP protein Q9CAJ0 original sequence version: 62 DIP protein Q9CAJ0 original sequence version: 65 DIP protein Q9CAJ0-1 original sequence version: 66 DIP protein Q9CAJ0 original sequence version: 81 DIP protein Q9CAJ0-1 original sequence version: 86 DIP protein Q9CAJ0-1 original sequence version: 108 DIP protein Q9CAJ0 original sequence version: 112 DIP protein Q9CAJ0-1 original sequence version: 81 B001BAF0DD8333E1 abscisic acid abscisic acid small molecule small molecule chemical synthesis Chemical synthesis (Chemical synthesis) abscisic acid (CHEBI:22152) is a apo carotenoid sesquiterpenoids (CHEBI:36758) , abscisic acid (CHEBI:22152) is a monocarboxylic acids (CHEBI:25384) IUPAC name (2Z,4E)-5-(1-hydroxy-2,6,6-trimethyl-4-oxocyclohex-2-en-1-yl)-3-methylpenta-2,4-dienoic acid abscisic acid (CHEBI:22152) is a apo carotenoid sesquiterpenoids (CHEBI:36758) , abscisic acid (CHEBI:22152) is a monocarboxylic acids (CHEBI:25384) IUPAC name (2Z,4E)-5-(1-hydroxy-2,6,6-trimethyl-4-oxocyclohex-2-en-1-yl)-3-methylpenta-2,4-dienoic acid p2c77_arath Protein phosphatase 2C 77 Protein ABSCISIC ACID-INSENSITIVE 2 ABI2 At5g57050 MHM17.19 Protein phosphatase 2C ABI2 PP2C77 protein protein arath Arabidopsis thaliana (Mouse-ear cress) Mouse-ear cress MDEVSPAVAVPFRPFTDPHAGLRGYCNGESRVTLPESSCSGDGAMKDSSFEINTRQDSLTSSSSAMAGVDISAGDEINGSDEFDPRSMNQSEKKVLSRTESRSLFEFKCVPLYGVTSICGRRPEMEDSVSTIPRFLQVSSSSLLDGRVTNGFNPHLSAHFFGVYDGHGGSQVANYCRERMHLALTEEIVKEKPEFCDGDTWQEKWKKALFNSFMRVDSEIETVAHAPETVGSTSVVAVVFPTHIFVANCGDSRAVLCRGKTPLALSVDHKPDRDDEAARIEAAGGKVIRWNGARVFGVLAMSRSIGDRYLKPSVIPDPEVTSVRRVKEDDCLILASDGLWDVMTNEEVCDLARKRILLWHKKNAMAGEALLPAEKRGEGKDPAAMSAAEYLSKMALQKGSKDNISVVVVDLKGIRKFKSKSLN 67CAAC76DA531A71 abi2-rcar1 1 4 unspecified role unspecified role prey prey 8 unspecified role unspecified role bait bait O04719:p.Gly168Asp mutation disrupting mutation disrupting interaction mutation disrupting certain certain sequence position certain certain certain sequence position certain G D Wed May 02 17:02:29 BST 2018 This feature has been corrected as a result of our quality control procedures. The original label was 'gly168asp' Tue Jun 30 17:43:57 BST 2020 This feature has been corrected as a result of our quality control procedures. The original label was 'p.Gly168Asp' physical association physical association Figure 2A We used the yeast two-hybrid system to screen for Arabidopsis proteins interacting with ABI2 (18). One of the interacting proteins (At1g01360.1) had no annotated function, and we named it regulatory component of ABA receptor 1 (RCAR1). abi1-rcar1 1 5 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 P49597:p.Gly180Asp mutation decreasing mutation decreasing interaction hotspot hotspot mutation decreasing certain certain sequence position certain certain certain sequence position certain G D Wed May 02 14:42:01 BST 2018 This feature has been corrected as a result of our quality control procedures. The original label was 'gly180asp' Tue Jun 30 17:41:07 BST 2020 This feature has been corrected as a result of our quality control procedures. The original label was 'p.Gly180Asp' 4 unspecified role unspecified role prey prey physical association physical association Figure 2A The single point mutation present in abi1 also impaired RCAR1 binding, and the interaction was abrogated with a carboxyl terminally truncated ABI11-180. hab1-rcar1 1 4 unspecified role unspecified role prey prey 6 unspecified role unspecified role bait bait physical association physical association Supplemenatry Figure 1 There are more than 50 PP2Cs in Arabidopsis. Of these, HAB1, a PP2C involved in ABA signaling and structurally related to ABI1/2 (19), physically interacted with RCAR1 in the yeast assay. abi2-rcar1-1 2 4 unspecified role unspecified role neutral component neutral component region N-terminal region of YFP protein yfp tag yellow fluorescent protein tag YFP yellow fluorescent protein yellow fluorescent protein YFP yfp tag undetermined undetermined sequence position undetermined undetermined undetermined sequence position undetermined 8 unspecified role unspecified role neutral component neutral component region C-terminal region of YFP protein yfp tag yellow fluorescent protein tag YFP yellow fluorescent protein yellow fluorescent protein YFP yfp tag undetermined undetermined sequence position undetermined undetermined undetermined sequence position undetermined physical association physical association Figure 2B Supplementary Figure 2 Fluorescence-activated cell distribution analysis of RCAR1-YFPN� and ABI1-YFPC�transfected protoplasts provides evidence for the formation of functional YFP complexes abi1-rcar1-1 2 5 unspecified role unspecified role neutral component neutral component region C-terminal region of YFP protein yfp tag yellow fluorescent protein tag YFP yellow fluorescent protein yellow fluorescent protein YFP yfp tag undetermined undetermined sequence position undetermined undetermined undetermined sequence position undetermined 4 unspecified role unspecified role neutral component neutral component region N-terminal region of YFP protein yfp tag yellow fluorescent protein tag YFP yellow fluorescent protein yellow fluorescent protein YFP yfp tag undetermined undetermined sequence position undetermined undetermined undetermined sequence position undetermined physical association physical association Figure 2B abi2-rcar1-2 3 4 unspecified role unspecified role neutral component neutral component 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 drome-schneider_2 Drosophila melanogaster late embryo cells Fruit fly schneider_2 Established from the late embryo of Drosophila melanogaster Mixture of diploid and tetraploid. Originally had some XY cells now predominantly XX. 8 unspecified role unspecified role neutral component neutral component 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 Figure 3D Interaction analysis by isothermal titration calorimetry of RCAR1 + ABI2 aba-rcar1-1 3 7 unspecified role unspecified role prey prey 4 unspecified role unspecified role 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 drome-schneider_2 Drosophila melanogaster late embryo cells Fruit fly schneider_2 Established from the late embryo of Drosophila melanogaster Mixture of diploid and tetraploid. Originally had some XY cells now predominantly XX. direct interaction direct interaction Supp. Figure 4 The analysis for binding of (S)-ABA to RCAR1 yielded lower energy changes and a higher apparent Kd of ~0.66 � 0.08 �M ABA (fig. S4, A and B).