Project description:In fungi, sexual compatibility is controlled by mating type loci that prevent self-fertilization. In the plant pathogenic fungus Ustilago maydis, the b mating type locus encodes a pair of unrelated homeodomain proteins, termed bE and bW. After fusion of two compatible, haploid cells, the bE and bW proteins form a heterodimeric complex, but only if they are derived from different, compatible alleles. The active bE/bW complex is required and sufficient to initiate pathogenic development of U. maydis, which is prerequisite for sexual reproduction of the fungus. However, the role of the b heterodimer during later stages of pathogenic development was unclear. To analyze b function during in planta development, we generated a temperature-sensitive bE allele (bEts) encoding a protein with a single amino acid alteration at the border of the homeodomain. This mutation leads to a stop in pathogenic development at the restrictive temperature in planta, while bEts strains show normal development at permissive temperature. At restrictive temperature, hyphae develop enlarged, bulbous cells at their tips that contain multiple nuclei, indicating a severe defect in cell division. DNA array analysis of bEts mutant strains in planta revealed a b-dependent regulation of genes coding for secreted proteins that were shown to influence fungal virulence. Our data demonstrate that in U. maydis the b heterodimer is not only essential to establish the heterodikaryon after mating of two compatible sporidia and to initiate fungal pathogenicity, but also to sustain in planta proliferation and ensure sexual reproduction. Maize plants were infected with a mixture of either FB1 and FB2 (wild-type), or RAb1ts and RAb2ts (temperature-sensitive b heterodimer) and kept at 22°C (permissive conditions). Control samples of FB1/FB2 and RAb1ts and RAb2ts were taken 5 days post inoculation at 22 °C (1 replicate each), to see if strains expressing the bts heterodimer show wildtype-like biotrophic development. To measure the impact of the b heterodimer on pathogenic development, infected plants were shifted for 9 hours to resrictive conditions (31°C) 111 hours post infection. After the temperature shift FB1/FB2 (3 replicates) infections developed normal, whereas RAb1ts/RAb2ts (3 replicates) infections were not able to further proliferate in planta because of a none-functional b heterodimer.
Project description:In fungi, sexual compatibility is controlled by mating type loci that prevent self-fertilization. In the plant pathogenic fungus Ustilago maydis, the b mating type locus encodes a pair of unrelated homeodomain proteins, termed bE and bW. After fusion of two compatible, haploid cells, the bE and bW proteins form a heterodimeric complex, but only if they are derived from different, compatible alleles. The active bE/bW complex is required and sufficient to initiate pathogenic development of U. maydis, which is prerequisite for sexual reproduction of the fungus. However, the role of the b heterodimer during later stages of pathogenic development was unclear. To analyze b function during in planta development, we generated a temperature-sensitive bE allele (bEts) encoding a protein with a single amino acid alteration at the border of the homeodomain. This mutation leads to a stop in pathogenic development at the restrictive temperature in planta, while bEts strains show normal development at permissive temperature. At restrictive temperature, hyphae develop enlarged, bulbous cells at their tips that contain multiple nuclei, indicating a severe defect in cell division. DNA array analysis of bEts mutant strains in planta revealed a b-dependent regulation of genes coding for secreted proteins that were shown to influence fungal virulence. Our data demonstrate that in U. maydis the b heterodimer is not only essential to establish the heterodikaryon after mating of two compatible sporidia and to initiate fungal pathogenicity, but also to sustain in planta proliferation and ensure sexual reproduction.
Project description:Ustilago maydis is a plant-pathogenic fungus that establishes a biotrophic relationship with its host Zea mays. The biotrophic interaction is initiated upon host penetration, and involves expansion of the host plasma membrane around hyphae, which is thought to facilitate the exchange of nutrients and virulence factors. Transcriptional regulators involved in the establishment of an infectious dikaryon and penetration into the host have been identified, however, regulators involved in the post-penetration stages remained to be elucidated. In the study we report the identification of an Ustilago maydis forkhead transcription factor, Fox1, which is exclusively expressed during biotrophic development. Deletion of fox1 results in reduced virulence and impaired tumour development in planta. Microarray analyses of Δfox1-infected plant tissue identified Fox1 as a transcriptional activator, involved in the expression of secreted effectors required for virulence.
Project description:Ustilago maydis is a plant-pathogenic fungus that establishes a biotrophic relationship with its host Zea mays. The biotrophic interaction is initiated upon host penetration, and involves expansion of the host plasma membrane around hyphae, which is thought to facilitate the exchange of nutrients and virulence factors. Transcriptional regulators involved in the establishment of an infectious dikaryon and penetration into the host have been identified, however, regulators involved in the post-penetration stages remained to be elucidated. In the study we report the identification of an Ustilago maydis forkhead transcription factor, Fox1, which is exclusively expressed during biotrophic development. Deletion of fox1 results in reduced virulence and impaired tumour development in planta. Δfox1 hyphae induce plant defences including the overproduction and accumulation of H2O2 in and around infected cells. This oxidative burst acts as an intercellular signal, which elicits a specific host defence response phenotypically represented by the encasement of proliferating hyphae in extensions of the plant cell wall. Maize microarrays experiments were performed to identify genes involved in the observed plant defence responses on leaf tissue infected with U. maydis strain SG200∆fox1 4 dpi.