C60 Fullerols Enhance Copper Toxicity and Alter the Leaf Metabolite and Protein Profile in Cucumber

Abiotic and biotic stress induce the production of reactive oxygen species (ROS), which limit crop production. Little is known about ROS reduction through the application of exogenous scavengers. In this study, C60 fullerol, a free radical scavenger, was foliar applied to three-week-old cucumber plants (1 or 2 mg/plant) before exposure to copper ions (5 mg/plant). Results showed that C60 fullerols augmented Cu toxicity by increasing the influx of Cu ions into cells (170% and 511%, respectively, for 1 and 2 mg of C60 fullerols/plant). We further use metabolomics and proteomics to investigate the mechanism of plant response to C60 fullerols. Metabolomics revealed that C60 fullerols up-regulated antioxidant metabolites including 3-hydroxyflavone, 1,2,4-benzenetriol, and methyl trans-cinnamate, among others, while it down-regulated cell membrane metabolites (linolenic and palmitoleic acid). Proteomics analysis revealed that C60 fullerols up-regulated chloroplast proteins involved in water photolysis (PSII protein), light-harvesting (CAB), ATP production (ATP synthase), pigment fixation (Mg-PPIX), and electron transport ( Cyt b6f). Chlorophyll fluorescence measurement showed that C60 fullerols significantly accelerated the electron transport rate in leaves (13.3% and 9.4%, respectively, for 1 and 2 mg C60 fullerols/plant). The global view of the metabolic pathway network suggests that C60 fullerols accelerated electron transport rate, which induced ROS overproduction in chloroplast thylakoids. Plant activated antioxidant and defense pathways to protect the cell from ROS damaging. The revealed benefit (enhance electron transport) and risk (alter membrane composition) suggest a cautious use of C60 fullerols for agricultural application.