Ras proteins are monomeric G proteins that are ubiquitous in fungal cells and play important roles in fungal growth, virulence, and environmental responses. Botrytis cinerea is a phytopathogenic fungus that infects various crops. However, under specific environmental conditions, the overripe grapes infected by B. cinerea can be used to brew valuable noble rot wine. As a Ras protein, the role of Bcras2 in the environmental responses of B. cinerea is poorly understood. In this study, we deleted the Bcras2 gene using homologous recombination and examined its functions. Downstream genes regulated by Bcras2 were explored using RNA sequencing transcriptomics. It was found that ΔBcras2 deletion mutants showed significantly reduced growth rate, increased sclerotia production, decreased resistance to oxidative stress, and enhanced resistance to cell wall stress. Additionally, Bcras2 deletion promoted the expression of melanin-related genes in sclerotia and decreased the expression of melanin-related genes in conidia. The above results indicate that Bcras2 positively regulates growth, oxidative stress resistance, and conidial melanin-related genes expression, and negatively regulates sclerotia production, cell wall stress resistance and sclerotial melanin-related genes expression. These results revealed previously unknown functions of Bcras2 in environmental responses and melanin metabolism in B. cinerea.

The fungal pathogen Botrytis cinerea causes gray mold disease, with Bcras1 serving as a key virulence factor. However, the role of Bcras1 in the environmental responses and its downstream signaling pathways remains unclear. This study investigated the function of Bcras1 and its regulatory network using gene deletion and transcriptomic analysis. Results showed that Δ Bcras1 mutants exhibited severe defects in conidiation, sclerotia formation, acid production, and virulence, along with increased sensitivity to stress and enhanced melanin production. Transcriptome profiling revealed 2236 downregulated and 2149 upregulated genes in Δ Bcras1 , including decreased expression of conidiation-related genes ( Bcbck1 , Bos5 , and Bcnop53 ), sclerotia-related gene ( Bcatf1 ), and MAPK pathway-related genes ( Bcsak1 , Bcmkk1 , Bcbck1 , and Bcste50 ), as well as altered ROS-related genes and upregulated melanin biosynthesis genes ( Bcsmr1 and Bcscd1 ). Functional characterization of the downregulated MAPK gene Bcste50 showed that it similarly affects growth, development, and virulence. Our results demonstrate that Bcras1 regulates multiple pathways controlling fungal development, virulence, and stress adaptation, with Bcste50 acting as a potential downstream component in this regulatory network.