A rust fungus effector directly binds plant pre‐mRNA splice site to reprogram alternative splicing and suppress host immunity

Alternative splicing (AS) is a crucial post‐transcriptional regulatory mechanism in plant resistance. However, whether and how plant pathogens target splicing in their host remains mostly unknown. For example, although infection by Puccinia striiformis f. sp. tritici (Pst), a pathogenic fungus that severely affects the yield of wheat worldwide, has been shown to significantly influence the levels of alternatively spliced transcripts in the host, the mechanisms that govern this process, and its functional consequence have not been examined. Here, we identified Pst_A23 as a new Pst arginine‐rich effector that localizes to host nuclear speckles, nuclear regions enriched in splicing factors. We demonstrated that transient expression of Pst_A23 suppresses plant basal defence dependent on the Pst_A23 nuclear speckle localization and that this protein plays an important role in virulence, stable silencing of which improves wheat stripe rust resistance. Remarkably, RNA‐Seq data revealed that AS patterns of 588 wheat genes are altered in Pst_A23‐overexpressing lines compared to control plants. To further examine the direct relationship between Pst_A23 and AS, we confirmed direct binding between two RNA motifs predicted from these altered splicing sites and Pst_A23 in vitro. The two RNA motifs we chose occur in the cis‐element of TaXa21‐H and TaWRKY53, and we validated that Pst_A23 overexpression results in decreased functional transcripts of TaXa21‐H and TaWRKY53 while silencing of TaXa21‐H and TaWRKY53 impairs wheat resistance to Pst. Overall, this represents formal evidence that plant pathogens produce ‘splicing’ effectors, which regulate host pre‐mRNA splicing by direct engagement of the splicing sites, thereby interfering with host immunity.