Construction of an Effector-Target Interaction Network for Identification of Immune-Related Effectors in Ralstonia pseudosolanacearum.

Bacterial wilt caused by Ralstonia pseudosolanacearum is a devastating soil-borne disease that results in significant losses of various crops worldwide. The type III secretion system (T3SS) secretes a suite of effectors into host cells, which is a major driver of disease development. These effectors do not act in isolation but form an interconnected network called the effectorome, functioning collectively through extensive crosstalk. Constructing large-scale effector-target interaction networks via high-throughput protein interaction assays is one of the primary approaches to studying effector function at the effectorome level. However, false positives inherent in high-throughput protein interaction methods hinder precise understanding of effector function. Here, we performed Gene Ontology (GO) enrichment analysis on a previously reported large-scale Ralstonia pseudosolanacearum effector-Arabidopsis thaliana target interaction network and found that effectors primarily interact with immune response-related proteins. Subsequently, we validated the interactions between immune-related target proteins and their corresponding effectors using one-to-one yeast two-hybrid assays, generating a validated immune-related effector-target subnetwork. Finally, analyses of reactive oxygen species (ROS) burst, callose deposition, and immune-related gene transcription revealed that the effectors in the subnetwork exhibit immune-suppressive functions, demonstrating the potential of the effector-target network in understanding the pathogenic mechanism at the effectorome level.