We show that above a certain threshold concentration, ozone leads to leaf injury in tomato (Lycopersicon esculentum). Ozone-induced leaf damage was preceded by a rapid increase in 1-aminocyclopropane-1-carboxylic acid (ACC) synthase activity, ACC content, and ethylene emission. Changes in mRNA levels of specific ACC synthase, ACC oxidase, and ethylene receptor genes occurred within 1 to 5 h. Expression of the genes encoding components of ethylene biosynthesis and perception, and biochemistry of ethylene synthesis suggested that ozone-induced ethylene synthesis in tomato is under biphasic control. In transgenic plants containing anLE-ACO1 promoter-β-glucuronidase fusion construct, β-glucuronidase activity increased rapidly at the beginning of the O3 exposure and had a spatial distribution resembling the pattern of extracellular H2O2 production at 7 h, which coincided with the cell death pattern after 24 h. Ethylene synthesis and perception were required for active H2O2 production and cell death resulting in visible tissue damage. The results demonstrate a selective ozone response of ethylene biosynthetic genes and suggest a role for ethylene, in combination with the burst of H2O2production, in regulating the spread of cell death.

SummaryStress ethylene emission is positively correlated with ozone sensitivity in various plant species, indicating that ethylene may be involved in the control of ozone damage. This study shows that ozone exposure of tomato plants for 5 h at 85 nl 1-1 and above leads to leaf injury within 24 h. 1-aminocyclopropane-l-carboxylic acid (ACC) content and ACC synthase activity were accordingly elevated within 1-2 h. Pre-treatment of leaves with inhibitors of ACC synthase and ACC oxidase significantly inhibited the evolution of ethylene and reduced ozone-induced visible damage. Transcript levels for only one out of three SadenosyI-L-methionine (SAM) synthetase genes (SAM3), and one out of four ACC synthase genes (LE-ACS2) were induced by ozone (maximum at 2 h). Treatment with protein kinase (K-252a) and phosphatase inhibitors (calyculin A) revealed that ACC synthase activity was additionally regulated by protein phosphorylation/dephosphorylation. Transcripts of ACC oxidase (pTOM13 cDNA probe) displayed the fastest response of the parameters tested (maximum at 30 min), suggesting a regulatory role for ACC oxidase in ethylene formation of ozone-exposed plants. The results demonstrate a highly selective ozone response by ethylene biosynthetic genes which resembles that of plant-pathogen interactions.

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