Characterisation of an essential virulence factor in the maize pathogen Ustilago maydis

The biotrophic fungus Ustilago maydis causes tumours on all aerial parts of the crop plant maize. With the help of hundreds of small secreted proteins and other secreted molecules, so called effectors, this pathogen is able to subvert plant defence responses and to manipulate the host metabolism for its own needs. Despite their importance during infection, only very few fungal effectors have been functionally characterized. In a virulence screen for U. maydis effectors with strong impact on pathogenicity we could identify APATHOGENIC 1 (AP1), a small secreted fungal protein transcriptionally upregulated during the biotrophic stages of U. maydis. Mutant strains deleted in the ap1 gene grow normal during the saprophytic stages of the fungus but fail to establish biotrophy on the host plant. Initial microscopic analysis shows a failure at the early stages of penetration events and an inability to block plant defence responses. The complemented ap1 strain expressing an ectopically integrated genomic fragment of ap1 can induce tumour formation similar to wild type infections. The proposed careful functional characterisation of AP1 and the targeted host- sided processes might give important insights into the defence system of maize and the strategies of the pathogen. The fact that orthologs of ap1 can be found in all publically available smut genome sequences indicates that AP1 is a core effector with a likely important conserved function in this class of biotrophic pathogens.

The biotrophic pathogen Ustilago maydis, the causative agent of corn smut disease, infects one of the most important crops worldwide - Zea mays. U. maydis typically infects 1-5% of the plants within commercial maize fields, therefore lowering the yield and the quality of the crop plants. To successfully colonize its host, U. maydis secretes proteins, known as effectors which suppress plant defense responses and facilitate the establishment of biotrophy. The identification and understanding of targets on the pathogen side that are essential for virulence is an important building block in developing new sustainable strategies to inhibit plant diseases. During this study, we identified the U. maydis effector protein Cce1. Cce1 is essential for virulence and is upregulated during infection. Through microscopic analysis and in vitro assays, we show that Cce1 is secreted from hyphae during filamentous growth of the fungus. Strikingly, cce1 mutants are blocked at early stages of infection and induce callose deposition as a plant defense response. Cce1 is highly conserved among smut fungi and the Ustilago bromivora ortholog complemented the virulence defect of the SG200cce1 deletion strain. Surprisingly, Cce1 shows interactions with other core effectors essential for biotrophy establishment in Y2H assays but the biological significance of these interactions awaits future research.