Ustilago maydis fox1 mediated gene expression during pathogenic development

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. Maize plants were infected with a mixture of either FB1 and FB2 (wild-type), or FB1∆fox1 and FB2∆fox1 crossings, to measure the impact of fox1 on pathogenic development. Early Golden Bantam maize plants were grown in a phytochamber in a 15h/9h light-dark cycle; light period started/ended with 1h ramping of light intensity. Maize plants were kept at 28°C (light) and 20° (dark). Plantlets were individually sown in pots with potting soil (Fruhstorfer Pikiererde) and infected 7 days after sowing, 1 h before end of the light period. Infected leaf tumor material from at least 10 plants was collected 5 days post infection, 1 h before the end of the light period and directly frozen in liquid nitrogen for RNA-extraction. RNA samples were extracted from infected leaf tissue 5 days after infection.