Polyamine metabolism and ethylene biosynthesis in normal and habituated sugar beet callus

Abstract

The optimal assay conditions and the trend with time in culture (28 days) of arginine decarboxylase (ADE; EC 4.1.1.19), omithine decarboxylase (ODC; EC 4.1.1.17) and diamine oxidase (DAO; EC 1.4.3.6) activities in habituated (H) and normal (N) auxin‐ and cytokinin‐requiring sugar beet callus were compared. Although the response to variations in buffer pH and EDTA and pyridoxal phosphate (PLP) concentrations varied for ADC and ODC activities between the two callus types, pH 8.3, 50 μM PLP and 5 mM EDTA were generally optimal or near‐optimal for both H and N callus. In most cases the addition of ornithine or arginine in the ADC and ODC assays, respectively, given to block the interconversion between the two substrates, resulted in lower 14CO2 recovery. DAO activity was very differently affected in H and N callus by the presence of polyvinylpyrrolidone in the extration buffer. However, in both cases, this activity increased with time in culure. ADC activity was always predominant in both cell lines and always higher in N callus. In the latter, ADC activity rose sharply between days 14 and 21 and then leveled off while in H callus it incresed steadily from day 14 onwards. ODC activity was also higher in N callus and peaked sharply on day 21 while in H callus it was not detectable in the second half of the culture period. In both cell lines this activity was low or nil on day 28. 3,4‐[14C]‐methionine incorporation into ethylene and polyamines was also compared in N and H callus. In the latter, ethylene synthesis was lower and [14C]‐spermidine formation higher than in N callus. This is in accord with the significantly higher spermidine titres found in H callus.