The secreted chorismate mutase Cmu1 effector
The secreted chorismate mutase Cmu1 of U. maydis is a translocated virulence promoting effector. Cmu1 in the cytosol of maize is proposed to lower chorismate levels in the chloroplast where it would serve as precursor for the biosynthesis of the plant defense hormone salicylic acid (SA). Lowering SA levels in U. maydis infected tissue which is beneficial for biotrophic growth. The crystal structure of Cmu1 was determined in collaboration with Prof. G. Bange (SYNMIKRO, Marburg).
By performing an immunoprecipitation of Cmu1-HA from infected leave tissues followed by mass spectrometry, the secreted maize protein ZmKwl1 could be identified as a specific interactor. ZmKwl1 is a member of the kiwellin family which has 13 members in maize. In vitro pull-down experiments confirmed the interaction between Cmu1 and Kwl1. Recombinant Kwl1 protein inhibited the chorismate mutase activity of Cmu1. The expression of Kwl1 was strongly induced upon infection by U. maydis, indicating that Kwl1 is likely a pathogenesis related protein. Hydrogen-Deuterium exchange mass spectrometry (HDX/MS) mapped the interaction interface between Cmu1 and Kwl1 to the loop region of Cmu1. Truncation of the loop in Cmu1 abolished the interaction with Kwl1. The respective mutant protein allowed only partial complementation of a cmu1 deletion mutant. This illustrates that the interaction between Cmu1 and Kwl1 is relevant for virulence of U. maydis. This was further corroborated by silencing Kwl1 in maize and demonstrating that silenced plants were more susceptible to U. maydis infection. Kwl1 specifically blocks the catalytic activity of Cmu1 whose activity is not subject to allosteric control by plant metabolites. The structure of the Cmu1-Kwl1 complex revealed an intimate interaction between the two proteins and showed that Kwl1 hinders substrate access to the active site of Cmu1. A phylogenetic analysis done in collaboration with Prof. S. Rensing (Philipps Universität Marburg) suggests that widely conserved kiwellin proteins in plants, which were previously recognized as important allergens for humans only, present a versatile scaffold specifically counteracting pathogen molecules.