RasGEFs Play Essential Roles in the Biology of Sclerotinia sclerotiorum and Botrytis cinerea and Can Be Used as HIGS Targets for Disease Management

Sclerotinia sclerotiorum is a notorious soilborne fungal pathogen that causes serious yield losses in many economically important crops such as canola, beans, lettuce, and sunflower. The diseases it causes, commonly known as Sclerotinia stem rot or white mold, are difficult to control due to the pathogen's wide host range and the persistence of the sclerotia. Host-induced gene silencing (HIGS) has recently emerged as a promising strategy for managing fungal diseases, including those caused by S. sclerotiorum. However, effective HIGS targets are limited due to the incomplete knowledge about the pathogen's survival and virulence strategies. In this study, through combined forward and reverse genetic analyses, we identified and characterized four Ras guanine nucleotide exchange factors (RasGEFs) in S. sclerotiorum. These proteins were found to be essential for sclerotia development, compound appressoria functionality, and virulence to varying degrees. In parallel, four corresponding RasGEF homologs were characterized in Botrytis cinerea, a necrotrophic pathogen from the Sclerotiniaceae family, in which they contribute to fungal growth, conidiation, and virulence. Among them, RasGEFa plays a predominant role, as its deletion in both fungi resulted in the most severe developmental and virulence defects, and it appears to function upstream of the SMK1 MAPK pathway. Furthermore, plant hosts expressing double-stranded RNA targeting SsRasGEFa and SsRasGEFc showed significantly reduced disease symptoms, demonstrating the potential of HIGS as a strategy for controlling these pathogens. Taken together, our findings revealed that these RasGEFs play conserved and crucial roles in fungal development and virulence and represent promising molecular targets for durable HIGS-based crop protection.

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