Inositol Polyphosphate Binding Specificity of the Jasmonate Receptor Complex.

Recent findings that receptor complexes for auxin and jasmonate bind inositol polyphosphates stimulated the idea that plant hormone perception is regulated by inositol-derived molecules (Tan et al., 2007; Sheard et al., 2010). Inositol polyphosphates regulate critical cellular functions in eukaryotic cells (Munnik and Nielsen, 2011; Munnik and Vermeer, 2010; Gillaspy, 2013; Tsui and York, 2010; Kuo et al., 2014; Lee et al., 2015), and the discovery that thesemolecules bind to plant hormone receptors provides an interesting case model to study plant hormone perception. For instance, the ASK1-TIR1 component of the auxin receptor complex was copurified and cocrystallized with insect cellderived inositol hexakisphosphate (InsP6; Tan et al., 2007). TIR1 mutants defective in InsP6 binding failed to interact with the IAA7 transcriptional repressor in the presence of auxin in yeast two-hybrid assays and in pull-down experiments using tagged-recombinant Aux/IAA protein (Calderón Villalobos et al., 2012), suggesting that InsP6 binding might be important for auxin receptor function. Interestingly, the ASK1-COI1 component of the jasmonate receptor complex also copurified with inositol polyphosphate (Sheard et al., 2010). Here, NMR analyses revealed that insect cellpurified, nondialyzed protein contained either Dand/or L-myo-inositol-1,2,4,5,6-pentakisphosphate (Sheard et al., 2010), also referred to as Ins(1,2,4,5,6)P5 or short InsP5 [3-OH] and Ins(2,3,4,5,6)P5 or short InsP5 [1-OH], respectively. Unfortunately, NMR cannot discriminate between enantiomers; therefore, the structure of the insect-purified InsP5 isomer remains unresolved. Dialyzed ASK1-COI1 protein depleted of inositol polyphosphate failed to reconstitute the jasmonate receptor complex in vitro,while addition of InsP5 [3-OH] robustly stimulated complex formation (Sheard et al., 2010). Interestingly, Ins(1,4,5,6)P4 and InsP6 also stimulated complex formation, although InsP6 stimulated with lower efficiency (Sheard et al., 2010). Other InsP5 isomers (including the possible alternative InsP5 [1-OH] enantiomer) were not tested in this study. In plants, three InsP5 species with distinct chromatographic mobilities have been identified (StevensonPaulik et al., 2005; Hanke et al., 2012; Laha et al., 2015; Brearley and Hanke, 1996). Among them, only the isomeric nature of the symmetrical molecule InsP5 [2-OH] was determined,while the identity of the other two InsP5 isomers remains unknown (Stevenson-Paulik et al., 2005; Brearley and Hanke, 1996). Independent work in amoeba and in a pancreatoma cell line showed that inositol polyphosphates can be further phosphorylated at an existing phosphate position to give rise to inositol pyrophosphates, molecules such as InsP7 and InsP8 that contain energy-rich diphosphate bonds and have important cellular functions in amoeba, animal, and yeast cells (Menniti et al., 1993; Stephens et al., 1993; Shears et al., 2012; Mulugu et al., 2007; Wilson et al., 2013; Thota and Bhandari, 2015). Inositol pyrophosphates have also been detected in different plant species (Desai et al., 2014; Lemtiri-Chlieh et al., 2000; Brearley and Hanke, 1996; Laha et al., 2015), and recent work suggests an important function of these molecules in regulating jasmonate-dependent responses (Laha et al., 2015). Jasmonate perception is regulated by COI1, the F-box component of an SCF ubiquitin E3 ligase complex. COI1 recruits JasmonateZIM-domain (JAZ) transcriptional 1 This work was supported by Emmy Noether Grant SCHA 1274/2-1 and SFB 1101/TP A05 from the Deutsche Forschungsgemeinschaft to G.S. Efforts of D.L. and P.J. were supported by the Deutscher Akademischer Austauschdienst and the Landesgraduiertenförderung Baden-Württemberg, respectively. N.Z. is a Howard Hughes Medical Institute Investigator and is supported by National Institutes of Health Grant R01CA107134 and the National Science Foundation. 2 These authors contributed equally to the article. * Address correspondence to [email protected]. G.S. and D.L. designed the research; D.L. andN.P. performedmost of the experiments; M.D. performed the molecular docking experiments; P.J. performed the immunoblot analyses and the Botrytis experiments; H.M. purified COI1-ASK1 from insect cells; S.T.B. helped with the analyses of the IC50 experiments; G.S., D.L., and N.Z. analyzed and interpreted the data; G.S. prepared the structural figures and wrote the article. [OPEN] Articles can be viewed without a subscription. www.plantphysiol.org/cgi/doi/10.1104/pp.16.00694