No abstract

In Ustilago maydis, the causal agent of corn smut, the morphological transition from yeast to filamentous growth is inextricably linked to pathogenicity; budding haploid cells are saprobic and, upon mating of compatible strains, the fungus converts to dikaryotic filamentous growth and obligate parasitism. The filamentous dikaryon proliferates in the host plant, inducing tumor formation and undergoing additional morphological changes that eventually result in the production of melanized diploid teliospores. In an attempt to identify new trans-acting factors that regulate morphogenesis in U. maydis, we searched for the presence of common binding sequences in the promoter region of a set of 37 genes downregulated in the filamentous form. Putative cis-acting regulatory sequences fitting the consensus binding site for the Aspergillus nidulans transcription factor StuA were identified in 13 of these genes. StuA is a member of the APSES transcription factors which contain a highly conserved DNA-binding domain with a basic helix-loop-helix (bHLH)-like structure. This class of proteins comprises critical regulators of developmental processes in ascomycete fungi such as dimorphic growth, mating, and sporulation but has not been studied in any fungus of the phylum Basidiomycota. A search for StuA orthologs in the U. maydis genome identified a single closely related protein that we designated Ust1. Deletion of ust1 in budding haploid wild-type and solopathogenic strains led to filamentous growth and abolished mating, gall induction, and, consequently, in planta teliosporogenesis. Furthermore, cultures of ust1 null mutants produced abundant thick-walled, highly pigmented cells resembling teliospores which are normally produced only in planta. We showed that ssp1, a gene highly induced in teliospores produced in the host, is also abundantly expressed in cultures of ust1 null mutants containing these pigmented cells. Our results are consistent with a major role for ust1 in regulating dimorphism, virulence, and the sporulation program in U. maydis.

SUMMARYPlant pathogens of the genus Verticillium pose a threat to many important crops worldwide. They are soil-borne fungi which invade the plant systemically, causing wilt symptoms. We functionally characterized the APSES family transcription factor Vst1 in two Verticillium species, V. dahliae and V. nonalfalfae, which produce microsclerotia and melanized hyphae as resistant structures, respectively. We found that, in V. dahliae Dvst1 strains, microsclerotium biogenesis stalled after an initial swelling of hyphal cells and cultures were never pigmented. In V. nonalfalfae Dvst1, melanized hyphae were also absent. These results suggest that Vst1 controls melanin biosynthesis independent of its role in morphogenesis. The absence of vst1 also had a great impact on sporulation in both species, affecting the generation of the characteristic verticillate conidiophore structure and sporulation rates in liquid medium. In contrast with these key roles in development, Vst1 activity was dispensable for virulence. We performed a microarray analysis comparing global transcription patterns of wild-type and Dvst1 in V. dahliae. G-protein/cyclic adenosine monophosphate (G-protein/cAMP) signalling and mitogen-activated protein kinase (MAPK) cascades are known to regulate fungal morphogenesis and virulence. The microarray analysis revealed a negative interaction of Vst1 with G-protein/ cAMP signalling and a positive interaction with MAPK signalling. This analysis also identified Rho signalling as a potential regulator of morphogenesis in V. dahliae, positively interacting with Vst1. Furthermore, it exposed the association of secondary metabolism and development in this species, identifying Vst1 as a potential co-regulator of both processes. Characterization of the putative Vst1 targets identified in this study will aid in the dissection of specific aspects of development.