Involvement of a Ca(2+)-dependent protein kinase component downstream to the gibberellin-binding phosphoprotein,...

Previously, we reported the identification of a gibberellin (GA)-binding protein in rice using ligand binding assay that was homologous to RuBisCO activase (Komatsu et al., FEBS Lett. 384, 167-171, 1996). Here, we provide an evidence for the involvement of protein kinases components downstream to the GA-binding phosphoprotein, RuBisCO activase in rice. Ca(2+)-dependent protein kinase activity was studied in subcellular fractions of leaf sheath from transgenic rice containing sense and antisense constructs of RuBisCO activase. In-gel kinase assay using histone III-S as a substrate showed constitutive induction of a 46- and 48-kDa Ca(2+)-dependent protein kinase activity in the sense transgenic plants. Kinase activities of these proteins were significantly reduced in the presence of uniconazole, a potent GA biosynthesis inhibitor, but one of them was strongly promoted by GA(3) treatment in transgenic plants carrying a smaller subunit of RuBisCO activase (OsrcaA1) compared to the larger subunit OsrcaA2. Also, in vitro phosphorylation studies using two-dimensional polyacrylamide gel showed changes in the degree of phosphorylation of several proteins in OsrcaA1- and OsrcaA2-sense transgenic rice. These studies suggest the presence of two independent cytosolic Ca(2+)-dependent protein kinase signaling components downstream to the GA-binding protein in rice suggesting their role in GA signaling.