Characterization of factors involved in mating, morphogenesis and virulence in smut fungi

Ustilago hordei and Ustilago maydis represent a group of fungal pathogens that cause economically important smut diseases on cereals and grasses. To identify factors involved in pathogenesis, the mating-type locus (MAT) was characterized in U. hordei and a genetic suppression screen was utilized in U. maydis. In Ustilago hordei, mating and pathogenicity are controlled by the MAT locus, which contains two distinct gene complexes, a and b. In this study, the a and b regions were tagged with the recognition sequence for the restriction enzyme l-Scel and determined that the distance between the complexes is 500 kb in a MAT-1 strain and 430 kb in a MAT-2 strain. Characterization of the organization of the known genes within a and b provided evidence for non-homology and sequence inversion between MAT-1 and MAT-2. Antibiotic-resistance markers were also used to tag the a gene complex in MAT-1 strains (phleomycin) and the b gene complex in MAT-2 strains (hygromycin). Crosses were performed with these strains and progeny resistant to both antibiotics were recovered at a very low frequency suggesting that recombination is suppressed within the MAT region. Overall, the chromosome homologues carrying the MAT locus share features with primitive sex chromosomes, with the added twist that the MAT locus also controls pathogenicity. In many fungi, mating, pathogenicity and the morphological transition between budding and filamentous growth are regulated by conserved signaling mechanisms including the cAMP/protein kinase A (PKA) pathway and at least one MAP kinase pathway. In this study, suppressor mutants that restored budding growth to a constitutively filamentous U. maydis mutant with a defect in the gene encoding a catalytic subunit of PKA were identified. Complementation of one suppressor mutant unexpectedly identified the ras1 gene. Deletion of the ras1 gene in haploid cells altered cell morphology, eliminated pathogenicity on maize seedlings and revealed a role in the production of aerial hyphae during mating. An activated ras1 allele was also used to demonstrate that Ras1 promotes pseudohyphal growth via a MAPK cascade. These results reveal an additional level of cross-talk between the cAMP signaling pathway and a MAP kinase pathway influenced by Ras1.Science, Faculty ofMicrobiology and Immunology, Department ofGraduat