Gibberellin stimulates synthesis of a protein kinase in dwarf pea epicotyls

Abstract

Application of gibberellin A 3 (GA 3, 1 μM) stimulated protein kinase activity two-fold in light-grown dwarf pea epicotyls. This response was almost completely blocked by the simultaneous application of abscisic acid (ABA, 10 μM). Likewise, this GA 3-mediated increase in protein kinase activity was strongly inhibited by the application of cycloheximide (CHI, 20 μg ml −1), indicating that de novo protein synthesis was necessary for the response. The enhancement of protein kinase activity was observed in crude and partially purified enzyme fractions prepared from untreated and GA 3-treated dwarf pea epicotyls. However, purified protein kinase (casein-Sepharose fraction) from untreated and GA 3-treated epicotyls showed no significant difference in their specific activities. These results clearly suggested that GA 3 stimulates protein kinase activity through de novo enzyme synthesis and not by enzyme activation. The M r of the untreated and GA 3-stimulated protein kinase was 70000. Analysis of the purified protein kinase on SDS-PAGE from untreated and GA 3-treated epicotyls showed a single protein-stained band with M r 67000, indicating that it is monomeric. Chemical characterization of the reaction product of the protein kinase showed 32P-label in phosphoserine and phosphothreonine residues of the substrate casein. Significantly, the tall pea epicotyls contained 2.9-fold higher activity of the M r 70000 protein kinase than did epicotyls from dwarfs. Thus, it seems likely that the protein kinases activity is stimulated by endogenous GA s. We have also observed a significant increase in the phosphorylation of endogenous proteins in GA 3-treated dwarf pea epicotyls over the untreated epicotyls.