Ozone-tolerant Bel B and ozone-sensitive Bel W3 tobacco cultivars were subjected to acute ozone fumigation (200 p.p.b. for 3 h) and the subcellular localization of H2O2 was then studied. H2O2 accumulated on the cell walls and plasma membrane of both cultivars but the accumulation pattern differed greatly. H2O2 production was high in both cultivars immediately after fumigation, but, in the tolerant Bel B cultivar, after 7 h was only detected in some spongy cells adjacent to epidermal cells. Instead, in the sensitive Bel W3 cultivar, accumulation was still abundant in the cell walls of palisade, spongy and epidermal cells at this time. Significant changes in apoplastic ascorbate pool were noted in both cultivars in the first hours after fumigation. As the reduced ascorbate content remained unchanged, the marked increase in total ascorbate must have originated from the striking increase in dehydroascorbate, particularly in the ozone-sensitive Bel W3. Exposure of plants to ozone resulted in a marked transient increase in both free and conjugated salicylic acid (SA) as well as an increase in the activity of benzoic acid 2-hydroxylase which catalyses SA biosynthesis. SA induction differed greatly in the two cultivars, in that: (1) SA accumulation was far greater in the ozone-sensitive Bel W3 cv. and (2) the maximum SA peak was delayed in Bel W3 and observed only 7 h after fumigation ended. These results suggest that a high SA content, as documented in the ozone-sensitive Bel W3 cultivar, could trigger the production of ROS with subsequent SA-mediated cell-death.

Alfalfa (Medicago sativa L.) N‐sufficient plants were fed 1·5 mM N in the form of NO3−, NH4+ or NO3− in conjunction with NH4+, or were N‐deprived for 2 weeks. The specific activity of phosphoenolpyruvate carboxylase (PEPC) from the non‐nodulated roots of N‐sufficient plants was increased in comparison with that of N‐deprived plants. The PEPC value was highest with NO3− nutrition, lowest with NH4+ and intermediate in plants that were fed mixed salts. The protein was more abundant in NO3−‐fed plants than in either NH4+‐ or N mixed‐fed plants. Nitrogen starvation decreased the level of PEPC mRNA, and nitrate was the N form that most stimulated PEPC gene expression. The malate content was significantly lower in NO3−‐deprived than in NO3−‐sufficient plants. Root malate accumulation was high in NO3−‐fed plants, but decreased significantly in plants that were fed with NH4+. The effect of malate on the desalted enzyme was also investigated. Root PEPC was not very sensitive to malate and PEPC activity was inhibited only by very high concentrations of malate. Asparagine and glutamine enhanced PEPC activity markedly in NO3−‐fed plants, but failed to affect plants that were either treated with other N types or N starved. Glutamate and citrate inhibited PEPC activity only at optimal pH. N‐nutrition also influenced root nitrate and ammonium accumulation. Nitrate accumulated in the roots of NO3−‐ and (NO3− + NH4+)‐fed plants, but was undetectable in those administered NH4+. Both the nitrate and the ammonium contents were significantly reduced in NO3−‐ and (NO3− + NH4+)‐starved plants. Root accumulation of free amino acids was strongly influenced by the type of N administered. It was highest in NH4+‐fed plants and the most abundant amides were asparagine and glutamine. It was concluded that root PEPC from alfalfa plants is N regulated and that nitrate exerts a strong influence on the PEPC enzyme by enhancing both PEPC gene expression and activity.

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