Soluble phospholipase A 2 activity is induced before oxylipin accumulation in tobacco mosaic virus‐infected tobacco leaves and is contributed by patatin‐like enzymes

Summary Recent evidence suggests that oxidized lipid‐derived molecules play significant roles in inducible plant defence responses against microbial pathogens, either by directly deterring parasite multiplication, or as signals involved in the induction of sets of defence genes. The synthesis of these oxylipins was hypothesized to be initiated by the phospholipase A 2 ‐mediated release of unsaturated fatty acids from membrane lipids. Here, we demonstrate that, in tobacco leaves reacting hypersensitively to tobacco mosaic virus, a strong increase in soluble phospholipase A 2 (PLA 2 ) activity occurs at the onset of necrotic lesion appearance. This rapid PLA 2 activation occurred before the accumulation of 12‐oxophytodienoic and jasmonic acids, two fatty acid‐derived defence signals. Three PLA 2 isoforms were separated and the most active enzyme was partially purified, its N‐terminal sequence displaying similarity with patatin, the major storage protein in potato tubers. Three related tobacco patatin‐like cDNAs, called NtPat1 , NtPat2 and NtPat3 , were cloned, with NtPat2 encoding the PLA 2 isolated from infected leaves. RT–PCR experiments showed a rapid transcriptional activation of the three NtPat genes in virus‐infected leaves, preceding the increase in PLA 2 activity. Recombinant NtPat1 and NtPat3 enzymes were active in an assay using labelled bacterial membranes, and also displayed high bona fide PLA 2 activity on phosphatidylcholine substrate. These results point to a possible new role of patatin‐like phospholipases in inducible plant defence responses. The induction kinetics together with the enzymatic activity data indicate that the NtPat proteins may provide precursors for oxylipin synthesis during the hypersensitive response to pathogens.