Regulation in tobacco callus of enzyme activities of the nicotine pathway

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Abstract
Nicotine synthesis was stimulated by reduction of the medium auxin concentration (induction medium) in callus tissue originating from Nicotiana tabacum cv. Samsun. The enzyme activities of the route ornithine to methylpyrroline, which are those of ornithine decarboxylase, putrescine methyltransferase and methylputrescine oxidase, were determined during callus growth in the induction medium and as a control under non-nicotine-stimulating conditions (growth medium). The enzymes were assayed by high-performance liquid chromatography. Whereas the activities of ornithine decarboxylase were very similar under nicotine-stimulating and non-stimulating conditions, those of putrescine methyltransferase and methyl-putrescine oxidase increased strongly in the induction medium. In addition, the pools of putrescine and methylputrescine were determined throughout the callus growth cycle. Both sets of data strongly confirm the supposition that putrescine methyl-transferase is the enzyme under stringent control for nicotine biosynthesis, whereas the subsequent methylputrescine oxidase is co-regulated, although less stringently.