Utilising natural diversity of kinases to rationally engineer interactions with the angiosperm immune receptor ZAR1.

Abstract

The highly conserved angiosperm immune receptor HOPZ-ACTIVATED RESISTANCE1 (ZAR1) recognises the activity of diverse pathogen effector proteins by monitoring the ZED1-related kinase (ZRK) family. Understanding how ZAR1 achieves interaction specificity for ZRKs may allow for the expansion of the ZAR1-kinase recognition repertoire to achieve novel pathogen recognition outside of model species. We took advantage of the natural diversity of Arabidopsis thaliana kinases to probe the ZAR1-kinase interaction interface and found that A. thaliana ZAR1(AtZAR1) can interact with most ZRKs, except ZRK7. We found evidence of alternative splicing of ZRK7, resulting in a protein that can interact with AtZAR1. Despite high sequence conservation of ZAR1, interspecific ZAR1-ZRK pairings resulted in the autoactivation of cell death. We showed that ZAR1 interacts with a greater diversity of kinases than previously thought, while still possessing the capacity for specificity in kinase interactions. Finally, using AtZAR1-ZRK interaction data, we rationally increased ZRK10 interaction strength with AtZAR1, demonstrating the feasibility of the rational design of a ZAR1-interacting kinase. Overall, our findings advance our understanding of the rules governing ZAR1 interaction specificity, with promising future directions for expanding ZAR1 immunodiversity.