α‐costic acid, the main sesquiterpenoid isolated from Dittrichia viscosa (L) Greuter, induces oxidative stress and autophagy in tomato

Dittrichia viscosa (L.) Greuter, a perennial plant in the Asteraceae, has strong allelopathic activity due to the high content of various secondary metabolites. The bicyclic sesquiterpenoid alpha-costic acid is the most abundant secondary metabolite of D. viscosa. Its remarkable insecticidal, antiparasitic, and phytotoxic activities point to its potential use as a natural herbicide, but information on its mode of action is lacking. To shed light on the mechanism of action of alpha-costic acid in plant cells, we investigated the phytotoxicity of alpha-costic acid in tomato plants (Solanum lycopersicum L.) through in vivo assays, the underlying cellular effects using biochemical assays, and the effect on subcellular organelles using confocal microscopy on tomato protoplasts incubated with organelle-specific fluorescent probes. In vivo tests showed that alpha-costic acid inhibited the growth of tomato seedlings and induced chlorosis and spot lesions in leaves. Biochemical assays demonstrated that alpha-costic acid caused ion leakage, chlorophyll depletion, H2O2 overproduction, callose deposition, and membrane lipid peroxidation. Confocal microscopy demonstrated that alpha-costic acid determined ROS overproduction and network disruption in mitochondria, singlet oxygen overproduction in chloroplasts, vacuole disintegration, and autophagosome formation. Overall, our data are consistent with a model according to which alpha-costic acid phytotoxicity is related to oxidative stress in mitochondria and chloroplasts that induces extensive membrane damage, ultimately resulting in cell death associated with autophagy