Functional Characterization of the Subtilase Gene Cg043 Downregulated by 4-Ethyl-1,2-dimethoxybenzene in the Growth and Pathogenicity of Colletotrichum gloeosporioides

Abstract

Colletotrichum gloeosporioides, the causative agent of anthracnose, poses a significant threat to agricultural production. Previous studies identified 4-ethyl-1,2-dimethoxybenzene as a potent antifungal compound that downregulates the expression of the subtilase gene Cg043, although the underlying molecular mechanism remains unclear. Here, we generated Cg043 knockout mutants (ΔCg043) and found that their sensitivity to 4-ethyl-1,2-dimethoxybenzene was significantly reduced, identifying Cg043 as a key molecular target. Phenotypic assays and transcriptomic analyses revealed that Cg043 downregulation inhibits hyphal growth, spore production, and germination while impairing cell wall and membrane integrity and reducing pathogenicity. Furthermore, functional verification of the signal peptide and subcellular localization analysis confirmed that Cg043 is a secreted protein specifically localized to the plant cell nucleus, suggesting its role in virulence. These findings elucidate a novel antifungal mechanism by which 4-ethyl-1,2-dimethoxybenzene suppresses the growth, development, and pathogenicity of C. gloeosporioides via Cg043 downregulation, highlighting a promising molecular target for sustainable anthracnose management.