title: PYR/PYL/RCAR family members are major in-vivo ABI1 protein phosphatase 2C-interacting proteins in Arabidopsis.
abstract: Abscisic acid (ABA) mediates resistance to abiotic stress and controls developmental processes in plants. The group-A PP2Cs, of which ABI1 is the prototypical member, are protein phosphatases that play critical roles as negative regulators very early in ABA signal transduction. Because redundancy is thought to limit the genetic dissection of early ABA signalling, to identify redundant and early ABA signalling proteins, we pursued a proteomics approach. We generated YFP-tagged ABI1 Arabidopsis expression lines and identified in vivo ABI1-interacting proteins by mass-spectrometric analyses of ABI1 complexes. Known ABA signalling components were isolated including SnRK2 protein kinases. We confirm previous studies in yeast and now show that ABI1 interacts with the ABA-signalling kinases OST1, SnRK2.2 and SnRK2.3 in plants. Interestingly, the most robust in planta ABI1-interacting proteins in all LC-MS/MS experiments were nine of the 14 PYR/PYL/RCAR proteins, which were recently reported as ABA-binding signal transduction proteins, providing evidence for in vivo PYR/PYL/RCAR interactions with ABI1 in Arabidopsis. ABI1-PYR1 interaction was stimulated within 5 min of ABA treatment in Arabidopsis. Interestingly, in contrast, PYR1 and SnRK2.3 co-immunoprecipitated equally well in the presence and absence of ABA. To investigate the biological relevance of the PYR/PYLs, we analysed pyr1/pyl1/pyl2/pyl4 quadruple mutant plants and found strong insensitivities in ABA-induced stomatal closure and ABA-inhibition of stomatal opening. These findings demonstrate that ABI1 can interact with several PYR/PYL/RCAR family members in Arabidopsis, that PYR1-ABI1 interaction is rapidly stimulated by ABA in Arabidopsis and indicate new SnRK2 kinase-PYR/PYL/RCAR interactions in an emerging model for PYR/PYL/RCAR-mediated ABA signalling.
authors.affiliation: Division of Biological Sciences, Cell and Developmental Biology Section and Center for Molecular Genetics, University of California, San Diego, 9500 Gilman Drive, La Jolla, CA 92093-0116, USA.
authors.forename: Noriyuki
authors.initials: N
authors.lastname: Nishimura
authors.forename: Ali
authors.initials: A
authors.lastname: Sarkeshik
authors.forename: Kazumasa
authors.initials: K
authors.lastname: Nito
authors.forename: Sang-Youl
authors.initials: SY
authors.lastname: Park
authors.forename: Angela
authors.initials: A
authors.lastname: Wang
authors.forename: Paulo C
authors.initials: PC
authors.lastname: Carvalho
authors.forename: Stephen
authors.initials: S
authors.lastname: Lee
authors.forename: Daniel F
authors.initials: DF
authors.lastname: Caddell
authors.forename: Sean R
authors.initials: SR
authors.lastname: Cutler
authors.forename: Joanne
authors.initials: J
authors.lastname: Chory
authors.forename: John R
authors.initials: JR
authors.lastname: Yates
authors.forename: Julian I
authors.initials: JI
authors.lastname: Schroeder
mesh_terms: D000040:Abscisic Acid / Q000494:pharmacology; D017360:Arabidopsis / Q000187:drug effects / Q000235:genetics / Q000378:metabolism*; D029681:Arabidopsis Proteins / Q000235:genetics / Q000378:metabolism*; D015153:Blotting, Western; D002118:Calcium / Q000378:metabolism / Q000494:pharmacology; D008164:Luminescent Proteins / Q000235:genetics / Q000378:metabolism; D013058:Mass Spectrometry; D026901:Membrane Transport Proteins / Q000235:genetics / Q000378:metabolism*; D008856:Microscopy, Fluorescence; D009154:Mutation; D010749:Phosphoprotein Phosphatases / Q000235:genetics / Q000378:metabolism*; D010766:Phosphorylation; D019441:Plant Epidermis / Q000187:drug effects / Q000235:genetics / Q000378:metabolism; D010937:Plant Growth Regulators / Q000494:pharmacology; D054046:Plant Stomata / Q000187:drug effects / Q000235:genetics / Q000378:metabolism; D030821:Plants, Genetically Modified; D011485:Protein Binding; D011494:Protein Kinases / Q000235:genetics / Q000378:metabolism; D017346:Protein Serine-Threonine Kinases / Q000235:genetics / Q000378:metabolism; D040901:Proteomics
chemical_list: D029681:Arabidopsis Proteins; D008164:Luminescent Proteins; D026901:Membrane Transport Proteins; D010937:Plant Growth Regulators; C540976:AT4G17870 protein, Arabidopsis; C492978:SnRK2 protein, Arabidopsis; D000040:Abscisic Acid; D011494:Protein Kinases; C472139:OST1 protein, Arabidopsis; D017346:Protein Serine-Threonine Kinases; C087434:ABI1 protein, Arabidopsis; D010749:Phosphoprotein Phosphatases; D002118:Calcium