Mitogenic activated protein kinase Kpp6 signaling in the phytopathogenic Fungus Ustilago maydis: identification of downstream elements (original) (raw)
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An unusual MAP kinase is required for efficient penetration of the plant surface by Ustilago maydis
The EMBO Journal, 2003
In Ustilago maydis, pathogenic development is controlled by a heterodimer of the two homeodomain proteins bW and bE. We have identi®ed by RNA ®ngerprinting a b-regulated gene, kpp6, which encodes an unusual MAP kinase. Kpp6 is similar to a number of other fungal MAP kinases involved in mating and pathogenicity, but contains an additional N-terminal domain unrelated to other proteins. Transcription of the kpp6 gene yields two transcripts differing in length, but encoding proteins of identical mass. One transcript is upregulated by the bW/bE heterodimer, while the other is induced after pheromone stimulation. kpp6 deletion mutants are attenuated in pathogenicity. kpp6 T355A,Y357F mutants carrying a non-activatable allele of kpp6 are more severely compromised in pathogenesis. These strains can still form appressoria, but are defective in the subsequent penetration of the plant cuticle. Kpp6 is expressed during all stages of the sexual life cycle except mature spores. We speculate that Kpp6 may respond to a plant signal and regulate the genes necessary for ef®cient penetration of plant tissue.
Molecular Microbiology, 2009
In the phytopathogenic fungus Ustilago maydis a conserved mitogen-activated-protein-kinase (MAPK) module regulates sexual and pathogenic development. Kpp2 is the central MAPK of this module and is required for transcriptional and morphological responses to pheromone. Upon perception of the pheromone signal Kpp2 is phosphorylated by the MAPK kinase Fuz7. Here we demonstrate that the MAPK Kpp6, which has a partially redundant function with Kpp2, is also phosphorylated by Fuz7. We show that Rok1, a putative dual specificity phosphatase for MAPK signalling, controls the phosphorylation of Kpp2 as well as of Kpp6. rok1 mutants display increased filamentation and are enhanced in virulence. The enhanced virulence is caused by more efficient appressorium formation as well as plant invasion. Overexpression of rok1 reduced conjugation hyphae formation and strongly attenuated pathogenicity. This places Rok1 in a negative feedback loop regulating Kpp2 and Kpp6 activity upon pheromone stimulation and plant colonization.
Fungal Genetics and Biology, 2002
Morphogenesis and pathogenesis are closely associated aspects of the life cycle of the fungal pathogen Ustilago maydis. In this fungus, the dimorphic switch from budding to filamentous growth coincides with the transition from non-pathogenic to pathogenic growth on maize. We have cloned and characterized the ukb1 gene that encodes a putative serine/threonine protein kinase with a role in budding and filamentous growth. Mutants defective in ukb1 were altered in bud site selection and produced lateral buds at a greater frequency than wild-type cells. Dikaryotic cells defective in ukb1 were capable of colonizing host tissue and growing with a filamentous morphology in planta. However, the mutants were incapable of inducing tumor formation and they failed to complete sexual development. In addition, the ukb1 gene influenced the ability of colonies to form aerial mycelia in response to environmental stimuli. Overall, the discovery of ukb1 reinforces the connection between morphogenesis and pathogenesis in U. maydis.
Fungal Genetics and Biology, 2008
The ability to respond to a changing environment separates successful organisms from their competitors. Thus, signal transduction is a crucial aspect of an organism's growth, development, differentiation, and reproduction. Nowhere is this more evident than in the co-evolution of obligate pathogens with their host organisms. The genome sequence of Ustilago maydis, the pathogen of maize, has provided a powerful tool in the assessment and characterization of signaling pathways for this organism. Inspection of the sequence reveals that while U. maydis has a streamlined gene content, it appears to contain a full repertoire of the standard signaling cascades present in other fungi. A full range of paralogues are present to provide redundancy of function on the one hand while, on the other, distinct strategies for survival. This review explores signaling based on the conserved mitogen-activated protein (MAP) kinase and cAMPdependent protein kinase A (PKA) pathways as well as ancillary functions, with emphasis on the unique aspects of the U. maydis approach to utilizing this architecture.
Plant Signaling & Behavior, 2013
Transcriptomic and biochemical analyses of the experimental pathosystem constituted by Ustilago maydis and Arabidopsis thaliana were performed. haploid or diploid strains of U. maydis inoculated in A. thaliana plantlets grew on the surface and within the plant tissues in the form of mycelium, inducing chlorosis, anthocyanin formation, malformations, necrosis and adventitious roots development, but not teliospores. Symptoms were more severe in plants inoculated with the haploid strain which grew more vigorously than the diploid strain. RNA extracted at different times post-infection was used for hybridization of one-channel microarrays that were analyzed focusing on the fungal genes involved in the general pathogenic process, biogenesis of the fungal cell wall and the secretome. In total, 3,537 and 3,299 genes were differentially expressed in the haploid and diploid strains, respectively. Differentially expressed genes were related to different functional categories and many of them showed a similar regulation occurring in U. maydis infecting maize. Our data suggest that the haploid strain behaves as a necrotrophic pathogen, whereas the diploid behaves as a biotrophic pathogen. The results obtained are evidence of the usefulness of the U. maydis-A. thaliana pathosystem for the analysis of the pathogenic mechanisms of U. maydis.
Eukaryotic Cell, 2003
In the phytopathogenic fungus Ustilago maydis, pheromone-mediated cell fusion is a prerequisite for the generation of the infectious dikaryon. The pheromone signal elevates transcription of the pheromone genes and elicits formation of conjugation hyphae. Cyclic AMP and mitogen-activated protein kinase (MAPK) signaling are involved in this process. The MAPK cascade is presumed to be composed of Ubc4 (MAPK kinase kinase), Fuz7 (MAPK kinase), and Ubc3/Kpp2 (MAPK). We isolated the kpp4 gene and found it to be allelic to ubc4. Epistasis analyses with constitutively active alleles of kpp4 and fuz7 substantiate that Kpp4, Fuz7, and Kpp2/Ubc3 are components of the same module. Moreover, we demonstrate that Fuz7 activates Kpp2 and shows interactions in vitro. Signaling via this cascade regulates expression of pheromone-responsive genes, presumably through acting on the transcription factor Prf1. Interestingly, the same cascade is needed for conjugation tube formation, and this process does not involve Prf1. In addition, fuz7 as well as kpp4 deletion strains are nonpathogenic, while kpp2 deletion mutants are only attenuated in pathogenesis. Here we show that strains expressing the unphosphorylatable allele kpp2 T182A/Y184F are severely affected in tumor induction and display defects in early infection-related differentiation.
Applied and environmental microbiology, 1995
A mutant of Ustilago maydis (VR43) with single-gene resistance to the dicarboximide fungicide vinclozolin was previously isolated and characterized. A genomic library was constructed, and an 8.7-kb resistance-conferring fragment was isolated by sib selection. Sequencing this fragment, we identified an 1,218-bp open reading frame, which, if disrupted by deletion, no longer confers resistance. Analyses of the data in GenBank demonstrated a high degree of homology between the product of the 1,218-bp open reading frame, referred to as the adr-1 gene, and Ser (Thr) protein kinases.
Molecular Plant-Microbe Interactions, 2000
In the corn smut fungus Ustilago maydis, mating of two haploid sporidia is a prerequisite for subsequent colonization of the host. Cyclic AMP (cAMP) and pheromone signals have been implicated in this developmental program. The cAMP pathway is also needed for subsequent fungal development in planta, as null mutants in any component of the pathway fail to form tumors. Here we show that moderate activation of the pathway conferred either by mutation in the Gα subunit or by mutation in the regulatory subunit of the protein kinase A influences tumor morphology. In the resulting tumors, the amount of fungal material is drastically reduced and fungal development is arrested at the stage of sporogenic hyphae. We conclude that tight regulation of the cAMP pathway is crucial for fungal development within the plant but does not interfere with the tumor induction process.
A Novel Core Effector Vp1 Promotes Fungal Colonization and Virulence of Ustilago maydis
2021
The biotrophic fungus Ustilago maydis secretes a plethora of uncharacterized effector proteins and causes smut disease in maize. Among the effector genes that are up-regulated during the biotrophic growth in maize, we identified vp1 (virulence promoting 1), which has an expression that was up-regulated and maintained at a high level throughout the life cycle of the fungus. We characterized Vp1 by applying in silico analysis, reverse genetics, phenotypic assessment, microscopy, and protein localization and provided a fundamental understanding of the Vp1 protein in U. maydis. The reduction in fungal virulence and colonization in the vp1 mutant suggests the virulence-promoting function of Vp1. The deletion studies on the NLS (nuclear localization signal) sequence and the protein localization study revealed that the C-terminus of Vp1 is processed after secretion in plant apoplast and could localize to the plant nucleus. The Ustilago hordei ortholog UhVp1 lacks NLS localized in the plant...
Ustilago maydis: dissecting the molecular interface between pathogen and plant
PLoS pathogens, 2012
Fungal diseases of plants represent one of the most eminent threats to agriculture. Given the food needs of a growing world population and that more and more crops are devoted to fuel production, the necessity to develop crops with better resistance to disease is increasing. To accomplish this, the mechanisms that plant pathogenic fungi use to colonize plants need to be elucidated. As of now, there are only few examples/models in which this can be done on a functional, genome-wide level, taking into account both the pathogen and its host plant [1]. The fungus Ustilago maydis (U. maydis) is one of these examples. It is a member of the smut fungi: a large group of parasites infecting mostly grasses, including several important crop plants such as maize ( , wheat, barley, and sugar cane. Smut fungi are biotrophs, i.e., parasites that need the living host plant to complete their sexual life cycle [2,3]. They do not establish prominent feeding structures like the related, haustoria-forming rust fungi. During penetration, the host plasma membrane invaginates and completely encases the intracellular hyphae ( , establishing an extended interaction zone [4] mediating the exchange of molecules between fungus and host. In contrast to most smut fungi that cause a systemic infection, remaining symptomless until the plant flowers, U. maydis can infect all above-ground parts of the maize plant but fails to spread systemically. U. maydis induces local tumors in which spores develop ( ) -a unique feature that allows detection of symptoms in corn seedlings less than a week after syringe infection with high levels of inoculum. This, together with the toolbox developed for reverse genetics, cell biology, and functional studies, has contributed to its status as a model for biotrophic basidiomycete fungi . Here the current level of our understanding of the elaborate molecular crosstalk between U. maydis and its host plant will be discussed.