VdEGe1, a cell wall-degrading enzyme gene from Verticillium dahliae required for cotton pathogenesis

Verticillium wilt, caused by Verticillium dahliae, is a destructive disease threatening global cotton production, leading to severe yield and fiber quality losses. Cell wall-degrading enzymes (CWDEs) are known to be crucial for fungal pathogenicity, but the specific roles of individual endoglucanases V. dahliae virulence remain poorly understood. This study characterized an endoglucanase gene VdEGe1 to elucidate its function and mechanistic contribution to V. dahliae pathogenicity. Bioinformatic analyses were performed to identify endoglucanase genes in the V. dahliae genome, followed by phylogenetic clustering. Host-Induced Gene Silencing (HIGS) was employed to specifically inhibit VdEGe1 expression to evaluate the cotton resistance to Verticillium wilt. Knockout mutagenesis in V. dahliae was used to assess the roles of VdEGe1 in fungal growth, development, and pathogenicity. Secretory activity of VdEGe1 was validated using a yeast signal peptide trap system, and Agrobacterium-mediated transient expression in Nicotiana benthamiana were conducted to evaluate cell death induction. Secretome profiling was analyzed to decipher the pathogenic mechanisms associated with VdEGe1. VdEGe1 was classified into the glycoside hydrolase family 45 (GH45). HIGS of VdEGe1 significantly reduced Verticillium wilt symptoms, disease index, and fungal biomass in infected cotton. The ΔVdEGe1 knockout mutant exhibited impaired colony expansion, hyphal growth, sporulation, and spore germination, accompanied by severely attenuated virulence. VdEGe1 was confirmed as a secreted protein but did not trigger or suppress cell death in N. benthamiana. Secretome analysis identified 311 differentially expressed secretory proteins, with 156 being down-regulated. The VdEGe1 deletion mutant exhibited a significant down-regulation of mitochondrial-related proteins and several carbohydrate-active enzymes, accompanied by a decrease in intracellular ATP levels and a slower growth rate on media containing various carbon sources. VdEGe1 deletion leads to mitochondrial dysfunction and impaired carbon utilization, which induces energy starvation in V. dahliae, thereby compromising its growth, development and virulence. These findings provide novel molecular insights into V. dahliae-host interactions and lay a theoretical foundation for developing targeted control strategies against cotton Verticillium wilt.