Ozone has dramatic effects on the regulation of the prechorismate pathway in tobacco (Nicotiana tabacum L.cv. Bel W3)

The accumulation of aromatic secondary metabolites is a well-known element of the plant response to ozone. Most of these metabolites are synthesized via the three aromatic amino acids phenylalanine, tyrosine and tryptophan. Before branching, the biosynthetic pathway to the three amino acids shares seven enzymatic steps, called the prechorismate pathway, catalysed by 3-deoxy-D: -arabino-heptulosonate-7-phosphate (DAHP) synthase [EC 2.5.1.54], 3-dehydroquinate synthase [EC 4.2.3.4], 3-dehydroquinate dehydratase [EC 4.2.1.10]-shikimate 5-dehydrogenase [EC 1.1.1.25], shikimate kinase [EC 2.7.1.71], 5-enolpyruvylshikimate 3-phosphate synthase [EC 2.5.1.19] and chorismate synthase [EC 4.2.3.5]). We have studied the transcript level of these enzymes and the aromatic metabolite profile in the ozone sensitive tobacco cultivar BelW3 (Nicotiana tabacum L. cv Bel W3), when exposed to an acute ozone pulse (160 nl l(-1), 5 h). Specific cDNA-fragments of the corresponding six genes were isolated from tobacco Bel W3 and used as probes for determining the expression of the prechorismate pathway genes. The fully expanded leaves of ozone treated plants, which developed symptoms like necrotic leaf spots and accumulation of aromatic metabolites, showed a clear induction of the shikimate pathway genes; indicating, that this induction is linked to the development of the symptoms. Distinct kinetics and magnitudes were observed in tobacco leaves for the ozone dependent enhanced mRNA accumulation of the aforementioned genes in BelW3. The strongest and earliest induction due to ozone treatment could be observed for DAHP synthase. An isoform-specific analysis of the transcripts showed a strong induction on transcript level only for one of three isoforms, which was followed by the induction of the DAHP synthase also on protein level. The different induction kinetics of the prechorismate pathway genes indicate that their regulation in response to ozone might be regulated by different signals, for example, ethylene, reactive oxygen species or salicylic acid, which also occur with different kinetics and thus may play different roles in the plant response to ozone