Further analysis of necrotrophic mycoparasitism in the predator yeast Saccharomycopsis schoenii

• Applicant: Professor Dr. Jürgen Wendland
• Subject Area: Metabolism, Biochemistry and Genetics of Microorganisms ; General Genetics and Functional Genome Biology
• Term: since 2020
• Project identifier: Deutsche Forschungsgemeinschaft (DFG) - Project number 448656174

In the previous successful project period we characterized two predator yeast genes and their involvement in mycopathogenicity. Mutants in the Saccharomycopsis schoenii KIL1 (for KILler kinase 1) and STE12 genes are completely avirulent and cannot produce penetration pegs. This work has been published in PLOS Pathogens. In continuation of this work we propose the following research objectives: First, we will characterize two additional conserved MAP kinase effectors, TEC1 and FAR1, which are both upregulated under predation conditions. Functional analysis of these genes will clarify their roles in predation or penetration peg formation. For example, Far1 in S. cerevisiae promotes a cell-cycle block prior to mating and is required for mating projection (shmoo) formation. We observed a block in the S. schoenii cell cycle during predation, which could be dependent on Far1 in S. schoenii as well as forming the penetration peg in the direction of the prey. From the predator’s side the regulation of bud morphogenesis vs. peg formation and the switch from one to the other requires further study. Our previous studies have shown that an area close to the growing but tip can be converted to a penetration peg and that after the predation cycle bud growth is resumed at its previous position. Furthermore, electron microscopy images (Kreger-van Rij and Veenhuis, 1973) showed a ‘neck band’ at the base of penetration pegs. We will analyse the localization of the actin cytoskeleton in fixed cells and the localization of the GFP-tagged septin CDC3 throughout the cell cycle and during predation in vivo to clarify whether there is actin accumulation and a septin ring or other septin-containing structures at the penetration peg. The second objective centers around the characterization of a set of predation response genes that we termed CRE-genes (Cystein-Rich Effectors). In other plant pathogens cysteine-rich effectors have been identified with a role in virulence, whose functions, however, still remain elusive. CRE-genes are specific to the genus Saccharomycopsis, contain a signal peptide to promote secretion and contain eight highly conserved cysteine residues. Additionally, within the S. schoenii clade a longer CRE-gene has been found that harbors six further cysteine residues at conserved positions. Penetration of a prey via a penetration peg may alone not be sufficient to kill a prey cell. Therefore, the study of this gene set may elucidate how a necrotroph manages to kill prey cells. We will also look more closely into events within our model Saccharomyces cerevisiae prey cell. Our focus is on cell membrane biology of the prey cell to address the question whether (or for how long) the plasma membrane of the prey cells stays intact during predation. Using CRE-GFP we will elucidate if effector molecules need to enter the prey cell, and conversely, if, S. cerevisiae endocytosis mutants delay the predation process/time required for killing.

• DFG Programme: Research Grants