A chicory (Cichorium intybus L. var. Witloof) line, previously selected in vitro for tolerance to the sulfonylurea herbicide chlorsulfuron and regenerated into resistant fertile plants showing a 1500 to 2000 higher resistance than wild-type, was characterized in order to understand the biochemical basis of the resistance. Transcription, specific activity level and properties of acetolactate synthase (ALS, EC 184.108.40.206), the first enzyme in branched-chain amino acid synthesis and the target of sulfonylurea and imidazolinone herbicides, were evaluated in plant tissues and compared with those of a sensitive control. Whilst Northern blot analysis detected similar levels of ALS-mRNA, a significant difference was found in ALS specific activity. However, its higher lever in sensitive plants, could not account for the selected tolerance. When ALS activity from wild-type and resistant plants was measured in the presence of increasing chrorsulfuron concentrations, a striking variation in enzyme sensitivity was found. In the two cases 50%-inhibition was achieved at: 21 nmol/L and 12 mu mol/L (R = 570), respectively, suggesting that the tolerant line could owe its resistance to a mutational event resulting in a decreased affinity of the target enzyme. Mutant ALS displayed cross-tolerance to the imidazolinone herbicide imazamethabenz, a 125-fold higher level of which was required to cause 50%-inhibition of enzyme activity. Herbicide tolerance was accompanied also by a reduced sensitivity to the regulative feedback inhibition by leucine and valine, with concentrations causing 50%-inhibition about: 10 and 3-fold higher, respectively, than those found for the wild-type enzyme.
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