Adhesion of Ustilago maydis filaments at the onset of pathogenic development
Final Report Abstract
Our initial aim was to identify and to characterize proteins that mediating surface attachment in U. maydis, and to use the respective genes as reporters to identify regulatory components mediating the plant-surface specific expression. We have used candidate gene approaches based on (a) homology to proteins involved in adhesion, (b) protein structure and (c) microarray analysis, as well as a functional screen in S. cerevisiae for surface adhesion, expressing a full length U. maydis cDNA library in a nonadhering S. cerevisiae ∆FLO11 strain. Although the test systems used to monitor amd screen surface adhesion were proven to be functional, none of our attempts led to the identification of U. maydis adhesins. To ensure sufficient results, we designed a side project as an extension the surface attachment project, focusing now on secreted proteins with a potential function directly after penetration. We have identified two secreted effector proteins, Pst1 and Pst2, using a combination of transcriptome analysis during the early infection stages and functional genomics. Pst1 and Pst2 are both transferred from the fungus to plant cells and localize in plant nuclei. Based on this observation, we identified now Plp2 by means of a similar expression profile and the presence of secretion signals in combination with domains that suggest a nuclear localization. Plp2 was shown to be absolutely required for plant infection, ∆plp2 strains are stalled directly after plant penetration and elicit a strong plant defense reaction. The gene is expressed exclusively in plant cells. When plants are infected with a strain expressing a functional Pst2:GFP fusion protein, the GFP signal is localized in the plant nucleus. A HA-tagged Plp2 protein is detected specifically in purified plant nuclei, suggesting that the protein is transferred from the fungus into the plant cell and taken up into the nucleus. The presence of a nuclear localization signal and a potential transactivation domain, which are both required for protein function, suggest a function of Plp2 in the modulation of plant gene expression.
