Effect of exogenous cytokinins,auxins and adenine on cytokinin <Emphasis Type="Italic">N</Emphasis>-glucosylation and cytokinin oxidase/dehydrogenase activity in de-rooted radish seedlings

The role of cytokinin N-glucosylation and degradation by cytokinin oxidase/dehydrogenase (CKX, EC 1.5.99.12) in response to application of exogenous auxins (2,4-dichlorophenoxyacetic acid 2,4-D] and -naphthaleneacetic acid NAA]) and cytokinins (N ⁶-benzyladenine BA] and trans-zeatin Z]) was investigated in de-rooted seedlings of Raphanus sativus L. cv. Rampouch. Both auxins applied for 24 h at 1 and 10 M concentration increased N-glucosylation of exogenously applied ³H]dihydrozeatin (DHZ) by up to 20%. The level of endogenous 7N-glucosides (of Z, isopentenyladenine iP] and DHZ) was increased by 2,4-D and NAA at 10 M concentration by 28 and 23%, respectively, the level of Z being decreased by 90 and 59%, respectively. 2,4-D and NAA suppressed CKX activity ca. by half. Exogenous cytokinins Z and BA applied at 1 and 10 M concentration stimulated 7N-glucosylation of ³H]DHZ (by up to 40%). BA both at 1 and 10 M, increased the level of endogenous Z by up to 35% and that of 7N-glucosides by up to 27%. BA application also strongly stimulated CKX activity (by up to 180%). Feeding with 1 and 10 M Z resulted in ca. 100-fold and 2000-fold increase of Z level, respectively. The main metabolite, Z7G, was increased ca. 6-fold and 60-fold, respectively. Levels of Z 9-glucoside (Z9G), trans-zeatin riboside (ZR) and Z O-glucoside (ZOG) were elevated to lesser extent. As compared to BA, Z had only negligible effect on CKX activity. Adenine (1–500 M) was preferentially 7N-glucosylated inhibiting competitively 7N-glucosylation of ³H]DHZ. At high concentrations (100–500 M) it increased endogenous levels of active cytokinins, especially of Z, however, it had no effect on CKX activity. Cytokinin N-glucosylation proved to be involved in down-regulation of active cytokinins in response to auxin and in the re-establishment of cytokinin homeostasis following application of exogenous cytokinins.