A Ralstonia solanacearum Effector Targets Splicing Factor SR34a to Reprogram Alternative Splicing and Regulate Plant Immunity

Alternative splicing is a critical post-transcriptional regulatory mechanism in eukaryotes. While infection with <i>Ralstonia solanacearum</i> GMI1000 significantly alters plant alternative splicing patterns, the underlying molecular mechanisms remain unclear. Herein, the effect of the GMI1000 Type III secretion system effectors on alternative splicing in the tomato cultivar Heinz 1706 was investigated. The RNA-seq analysis confirmed genome-wide alternative splicing changes induced by the Type III secretion system in tomato, including 1386 differential alternatively spliced events across 1023 genes, many of which are associated with plant defense. Seven nucleus-localized Type III effectors were transiently expressed in an RLPK splicing reporter system transgenic tobacco, identifying RipP2 as an effector that modulates alternative splicing levels. Sequence analysis, protein-protein interaction assays, and AlphaFold2 structural predictions revealed that RipP2 interacted with the tomato splicing factor <i>SR34a</i>. Furthermore, RipP2 acetylated a conserved lysine at position 132 within the SWQDLKD motif of <i>SR34a</i>, regulating its splicing pattern in defense-related genes and modulating plant immunity. This study elucidates how the "RipP2-<i>SR34a</i> module" influences plant immune responses by regulating the alternative splicing of immune-related genes, providing new insights into pathogen-plant interactions and splicing regulation.