Mitochondrial complex II as a self-resistance probe uncovers antifungal 2-pyridones from Penicillium oxalicum

Abstract

Mitochondrial complex II (SDH), a key enzyme in the tricarboxylic acid cycle and oxidative phosphorylation, represents a validated target for agricultural fungicide development. Capitalizing on filamentous fungi as valuable sources of bioactive metabolites, we employed the subunit B of SDH as a probe in the self-resistance gene-directed strategy to rapidly discover antifungal natural products. The apn biosynthetic gene cluster (BGC), harboring the SDHB homolog gene apnS, was identified in Penicillium oxalicum. Overexpression of the pathway-specific transcription factor gene apnH successfully activated the silent apn BGC in a marine-derived P. oxalicum strain, yielding four new 2-pyridones (1-3 and 9) and five known analogues (4-8). Antifungal profiling revealed broad-spectrum activity of 2-pyridones against phytopathogenic fungi. Importantly, compound 6 showed strong in vivo activity against B. cinerea-induced gray mold in tomatoes, with inhibition levels similar to those of carbendazim. Concurrently, gene knockout-complementation and heterologous expression confirmed apnS as a self-resistance gene. Leveraging this mechanism, we established a selectable system for fungal genetic transformation by combining apnS expression with 2-pyridone exposure. This work enables efficient discovery of antifungal 2-pyridones by targeting energy metabolism pathways, providing valuable lead compounds for agrochemical and food preservative development.