Die Bedeutung von Myo5 für das polare Wachstum, die pathogene Entwicklung und den Transport polarer Chitinsynthasen in Ustilago maydis

Table of Contents: Ustilago maydis is the causative agent of corn smut disease. A pre-requisite for the formation of the infectious dicaryotic filament is the fusion of two compatible sporidia. As a result of the pheromone stimulation by the partner cells form conjugation hyphae that grow towards each other and fuse at their tips. The ability to grow polar is crucial for both the mating process and the successful plant infection. Polar growth requires the delivery of growth components along the cytoskeleton. In this work it was shown, that the class V myosin Myo5 is of great importance for several stages during the sexual life cycle especially for the pheromone perception and the conjugation tube formation. In addition the growth of dicaryotic myo5ts hyphae was disturbed. Furthermore, myo5ts infection hyphae showed severe defects in polar growth during the early plant infection. Based on preliminary experiments chitin synthases were considered to be a potential cargo of Myo5. The genom of U. maydis codes for seven chitin synthases (Chs1-Chs7) and for one myosin chitin synthase (Mcs1). Chitin synthase deletion phenotypes were analysed and localized in vivo. It turned out that the deletion of chs5 and chs7 resulted in cell separation defects similar to the deletion of myo5. Dchs5- and Dchs7-cells were also impaired in the formation of conjugation tubes. In addition the deletion of chs7 led to impaired filament formation. The deletion of chs6 and mcs1 had no effect on filament formation but resulted in a complete loss of virulence. Similar to Myo5 Chs5, Chs6, Chs7 and Mcs1 localize to the growing bud tip. Inhibitor studies showed that their localization is actin dependent. In myo5ts cells Chs7 was mislocalized upon one hour temperature shift. Using the new established actin marker Fim1GFP a disruption of the actin cytoskeleton could be excluded. The phenotypic analyses as well as the localization and inhibitor studies indicate the participitation of Myo5 in Chs7 localization.