Induction of (+)-abscisic acid 8′ hydroxylase by (+)-abscisic acid in cultured maize cells

In vivo measurements of the enzymatic hydroxylation of the phytohormone (+)-abscisic acid to hormonally inactive (–)-phaseic acid in corn cell cultures revealed that (+)-abscisic acid 8′ hydroxylase activity was induced by (+)-abscisic acid treatment. This induction was blocked by the protein synthesis inhibitor cycloheximide and by the transcription inhibitor cordycepin. Following an induction treatment with abscisic acid, the amount of induced enzyme was measured by addition of both cycloheximide, to prevent further induction, and fresh abscisic acid as substrate for the induced enzyme. Phaseic acid production was related to the amount of enzyme induced. The experimental system was optimized for studying enzyme induction and the substrate specificity of the induced enzyme. The induced 8– hydroxylase was specific for (–)-abscisic acid, whereas the low basal activity also hydroxylated (–)-abscisic acid at the 7′ position to form 7′-hydroxy abscisic acid. After induction, the 8′ hydroxylase was rapidly degraded with a half-life of approximately 2 h. Phaseic and (′)-abscisic acid were weak inducers; however, the unstable oxidation product 8′-hydroxy abscisic acid exhibited 42% of the activity of (+)-abscisic acid. When corn cells were placed under water stress by increasing concentrations of mannitol in the culture medium, 8′ hydroxylase induction was suppressed. The experimental system described will be useful for further studies of the physiological and hormonal factors that modulate abscisic acid metabolism and for testing potential enzyme inhibitors and hormone analogues.