Integrated proteomics and metabolomics analysis reveals the antifungal mechanism of the C-coordinated O-carboxymethyl chitosan Cu(II) complex

Abstract

With wide application in agriculture, copper fungicides have undergone three stages of development: inorganic copper, synthetic organic copper, and natural organic copper. Using chitin/chitosan (CS) as a substrate, the natural organic copper fungicide C-coordinated O-carboxymethyl chitosan Cu(II) complex (O-CSLn-Cu) was developed in the laboratory. Taking Phytophthora capsici Leonian as an example, we explored the antifungal mechanism of O-CSLn-Cu by combining tandem mass tag (TMT)-based proteomics with non-targeted liquid chromatography-mass spectrometry (LC-MS)-based metabolomics. A total of 1172 differentially expressed proteins were identified by proteomics analysis. According to the metabolomics analysis, 93 differentially metabolites were identified. Acetyl-CoA-related and membrane localized proteins showed significant differences in the proteomics analysis. Most of the differential expressed metabolites were distributed in the cytoplasm, followed by mitochondria. The integrated analysis revealed that O-CSLn-Cu could induce the “Warburg effect”, with increased glycolysis in the cytoplasm and decreased metabolism in the mitochondria. Therefore, P. capsici Leonian had to compensate for ATP loss in the TCA cycle by increasing the glycolysis rate. However, this metabolic shift could not prevent the death of P. capsici Leonian. To verify this hypothesis, a series of biological experiments, such as scanning electron microscopy (SEM), transmission electron microscopy (TEM), and enzyme activity measurements were carried out. The results suggest that O-CSLn-Cu causes mitochondrial injury, which consequently leads to excessive ROS levels and insufficient ATP levels, thereby killing P. capsici Leonian.