Arabidopsis inositol polyphosphate kinase AtIpk2β is phosphorylated by CPK4 and positively modulates ABA signaling

Arabidopsis inositol polyphosphate kinase 213 (AtIpk2 beta) has multiple functions in plant development and in responding to abiotic stress. Although some related clues suggested a potential role of AtIpk2 beta in ABA signaling, the defined evidence was still lack. Here we discovered that a key ABA signaling component calcium-dependent protein kinase 4 (CPK4) can interact with AtIpk2 beta under ABA treated conditions through affinity purification and mass spectrometry detection. The interaction between CPK4 and AtIpk2 beta were further confirmed by yeast two hybrid and bimolecular fluorescence complementation assays. Expression of AtIpk2 beta also can be rapidly induced by ABA. In addition, we found that CPK4 can phosphorylate AtIpk2 beta in vitro and identified five novel phosphorylation sites of AtIpk2 beta by CPK4 kinase, including Tyr46, Ser48, Ser51, Thr128, Ser147. Overexpression of AtIpk2 beta in Arabidopsis was more sensitive to ABA in seed germination, primary root inhibition, ABA-responsive gene expression than wild type plants, whereas knockout mutant atipk2 beta exhibited no significant difference. The AtIpk2 beta variants containing Tyr46, Thr128, Ser147 mutated to Ala cannot complement the yeast mutant ipk2 growth in high temperature, suggesting that those three amino acid residues are critical for AtIpk2 beta. These findings provide insight into the modulation of ABA signaling by AtIpk2 beta. (C) 2017 Elsevier Inc. All rights reserved.