scholarly journals Pseudozyma saprotrophic yeasts have retained a large effector arsenal, including functional Pep1 orthologs

2018 ◽  
Author(s):  
Rahul Sharma ◽  
Bilal Ökmen ◽  
Gunther Doehlemann ◽  
Marco Thines
Keyword(s):  
Ustilago Maydis ◽  
Effector Proteins ◽  
Genetic Complementation ◽  
Smut Fungi ◽  
High Confidence ◽  
Positive Effects ◽  
Effector Gene ◽  
Sexual Recombination ◽  
Plant Surfaces ◽  
Plant Parasitic

SummaryThe basidiomycete smut fungi are predominantly plant parasitic, causing severe losses in some crops. Most species feature a saprotrophic haploid yeast stage, and several smut fungi are only known from this stage, with some isolated from habitats without suitable hosts, e.g. from Antarctica. Thus, these species are generally believed to be apathogenic, but recent findings that some of these might have a plant pathogenic sexual counterpart, casts doubts on the validity of this hypothesis. Here, four Pseudozyma genomes were re-annotated and compared to published smut pathogens and the well-characterised effector gene Pep1 from these species was checked for its ability to complement a Pep1 deletion strain of Ustilago maydis. It was found that 113 high-confidence putative effector proteins were conserved among smut and Pseudozyma genomes. Among these were several validated effector proteins, including Pep1. By genetic complementation we show that Pep1 homologs from the supposedly apathogenic yeasts restore virulence in Pep1-deficient mutants Ustilago maydis. Thus, it is concluded that Pseudozyma species have retained a suite of effectors. This hints at the possibility that Pseudozyma species have kept an unknown plant pathogenic stage for sexual recombination or that these effectors have positive effects when colonising plant surfaces.

scholarly journals A cell surface-exposed protein complex with an essential virulence function in Ustilago maydis

2021 ◽  
Author(s):  
Nicole Ludwig ◽  
Stefanie Reissmann ◽  
Kerstin Schipper ◽  
Carla Gonzalez ◽  
Daniela Assmann ◽  
...  
Keyword(s):  
Membrane Proteins ◽  
Protein Complex ◽  
Pathogenic Bacteria ◽  
Constitutive Expression ◽  
Pathogenic Fungi ◽  
Ustilago Maydis ◽  
Effector Proteins ◽  
Host Cells ◽  
Smut Fungi ◽  
Living Plant

AbstractPlant pathogenic fungi colonizing living plant tissue secrete a cocktail of effector proteins to suppress plant immunity and reprogramme host cells. Although many of these effectors function inside host cells, delivery systems used by pathogenic bacteria to translocate effectors into host cells have not been detected in fungi. Here, we show that five unrelated effectors and two membrane proteins from Ustilago maydis, a biotrophic fungus causing smut disease in corn, form a stable protein complex. All seven genes appear co-regulated and are only expressed during colonization. Single mutants arrest in the epidermal layer, fail to suppress host defence responses and fail to induce non-host resistance, two reactions that likely depend on translocated effectors. The complex is anchored in the fungal membrane, protrudes into host cells and likely contacts channel-forming plant plasma membrane proteins. Constitutive expression of all seven complex members resulted in a surface-exposed form in cultured U. maydis cells. As orthologues of the complex-forming proteins are conserved in smut fungi, the complex may become an interesting fungicide target.

scholarly journals Cross-species analysis between the maize smut fungi Ustilago maydis and Sporisorium reilianum highlights the role of transcriptional plasticity of effector orthologs for virulence and disease

2020 ◽  
Author(s):  
Weiliang Zuo ◽  
Deepak K Gupta ◽  
Jasper RL Depotter ◽  
Marco Thines ◽  
Gunther Doehlemann
Keyword(s):  
Ustilago Maydis ◽  
Effector Proteins ◽  
Differentially Expressed ◽  
Transcriptional Level ◽  
Smut Fungi ◽  
List Type ◽  
Sequencing Analysis ◽  
Distinct Disease ◽  
Sporisorium Reilianum ◽  
Transcriptional Plasticity

SummaryThe constitution and regulation of effector repertoires determines and shapes the outcome of the interaction with the host. Ustilago maydis and Sporisorium reilianum are two closely related smut fungi, which both infect maize, but cause distinct disease symptoms. Understanding how effector orthologs are regulated in these two pathogens can therefore provide insights to pathogen evolution and host adaption.We tracked the infection progress of U. maydis and S. reilianum in maize leaves, characterized two distinct infection stages for cross species RNA-sequencing analysis and identified 207 out of 335 one-to-one effector orthologs being differentially regulated during host colonization, while transcriptional plasticity of the effector orthologs correlated with the distinct disease development strategies.By using CRISPR-Cas9 mediated gene conversion, we identified two differentially expressed effector orthologs with conserved function between two pathogens. Thus, differential expression of functionally conserved genes contributes to species specific adaptation and symptom development. Interestingly, another differentially expressed orthogroup (UMAG_05318/sr1007) showed diverged protein function during speciation, providing a possible case for neofunctionalization.Together, we showed the diversification of effector genes in related pathogens can be caused both by plasticity on the transcriptional level, as well as through neofunctionalization of the encoded effector proteins.

scholarly journals A Novel Core Effector Vp1 Promotes Fungal Colonization and Virulence of Ustilago maydis

2021 ◽  
Vol 7 (8) ◽  
pp. 589
Author(s):  
Cuong V. Hoang ◽  
Chibbhi K. Bhaskar ◽  
Lay-Sun Ma
Keyword(s):  
Reverse Genetics ◽  
Signal Sequence ◽  
Protein Localization ◽  
Sequence Divergence ◽  
In Silico Analysis ◽  
Ustilago Maydis ◽  
Effector Proteins ◽  
Fungal Colonization ◽  
Smut Fungi ◽  
Fungal Virulence

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 cytoplasm, suggesting that the orthologs might have a distinct subcellular localization. Further complementation studies of the Vp1 orthologs in related smut fungi revealed that none of them could complement the virulence function of U. maydis Vp1, suggesting that UmVp1 could acquire a specialized function via sequence divergence.

scholarly journals Identification and Characterization of Two Transmembrane Proteins Required for Virulence of Ustilago maydis

2021 ◽  
Vol 12 ◽  
Author(s):  
Paul Weiland ◽  
Florian Altegoer
Keyword(s):  
Virulence Factors ◽  
Biochemical Analysis ◽  
Transmembrane Proteins ◽  
Ustilago Maydis ◽  
Effector Proteins ◽  
Smut Fungi ◽  
Initial Characterization ◽  
Associated Proteins ◽  
Identification And Characterization

Smut fungi comprise a large group of biotrophic phytopathogens infecting important crops such as wheat and corn. Through the secretion of effector proteins, the fungus actively suppresses plant immune reactions and modulates its host’s metabolism. Consequently, how soluble effector proteins contribute to virulence is already characterized in a range of phytopathogens. However, membrane-associated virulence factors have been much less studied to date. Here, we investigated six transmembrane (TM) proteins that show elevated gene expression during biotrophic development of the maize pathogen Ustilago maydis. We show that two of the six proteins, named Vmp1 and Vmp2 (virulence-associated membrane protein), are essential for the full virulence of U. maydis. The deletion of the corresponding genes leads to a substantial attenuation in the virulence of U. maydis. Furthermore, both are conserved in various related smuts and contain no domains of known function. Our biochemical analysis clearly shows that Vmp1 and Vmp2 are membrane-associated proteins, potentially localizing to the U. maydis plasma membrane. Mass photometry and light scattering suggest that Vmp1 mainly occurs as a monomer, while Vmp2 is dimeric. Notably, the large and partially unstructured C-terminal domain of Vmp2 is crucial for virulence while not contributing to dimerization. Taken together, we here provide an initial characterization of two membrane proteins as virulence factors of U. maydis.

scholarly journals Identification and characterization of two transmembrane proteins required for virulence of Ustilago maydis

2021 ◽  
Author(s):  
Paul Weiland ◽  
Florian Altegoer
Keyword(s):  
Virulence Factors ◽  
Biochemical Analysis ◽  
Transmembrane Proteins ◽  
Ustilago Maydis ◽  
Effector Proteins ◽  
Smut Fungi ◽  
Initial Characterization ◽  
Associated Proteins ◽  
Identification And Characterization

AbstractSmut fungi comprise a large group of biotrophic phytopathogens infecting important crops such as wheat and corn. Through the secretion of effector proteins, the fungus actively suppresses plant immune reactions and modulates its host’s metabolism. Consequently, how soluble effector proteins contribute to virulence is already characterized in a range of phytopathogens. However, membrane-associated virulence factors have been much less studied to date. Here, we investigated six transmembrane (TM) proteins that show elevated gene expression during biotrophic development of the maize pathogen Ustilago maydis. We show that two of the six proteins, named Vmp1 and Vmp2 (virulence-associated membrane protein), are essential for the full virulence of U. maydis. The deletion of the corresponding genes lead to a substantial attenuation in the virulence of U. maydis. Furthermore, both are conserved in various related smuts and contain no domains of known function. Our biochemical analysis clearly shows that Vmp1 and Vmp2 are membrane-associated proteins, potentially localizing to the U. maydis plasma membrane. Mass photometry and light scattering suggest that Vmp1 mainly occurs as a monomer, while Vmp2 is dimeric. Notably, the large and partially unstructured C-terminal domain of Vmp2 is crucial for virulence while not contributing to dimerization. Taken together, we here provide an initial characterization of two membrane proteins as virulence factors of U. maydis.

Ultrastructure of meiosis and the spindle pole body cycle in freeze-substituted basidia of the smut fungi Ustilago maydis and Ustilago avenae

1992 ◽  
Vol 70 (3) ◽  
pp. 629-638 ◽  
Author(s):  
Kerry O'Donnell
Keyword(s):  
Electron Microscopy ◽  
Spindle Pole Body ◽  
Ustilago Maydis ◽  
Spindle Pole ◽  
Meiotic Spindle ◽  
Smut Fungi ◽  
Late Prophase ◽  
Prophase I ◽  
Pole Body ◽  
Middle Piece

Meiosis in the smut fungi Ustilago maydis and Ustilago avenae (Basidiomycota, Ustilaginales) was studied by electron microscopy of serial-sectioned freeze substituted basidia. At prophase I, a spindle pole body composed of two globular elements connected by a middle piece was attached to the extranuclear surface of each nucleus. Astral and spindle microtubules were initiated at each globular element at late prophase I to prometaphase I. During spindle initiation, the middle piece disappeared and interdigitating half-spindles entered the nucleoplasm, which was surrounded by discontinuous nuclear envelope together with perinuclear endoplasmic reticulum. Kinetochore pairs at metaphase I were analyzed to obtain a karyotype for each species. The meiotic spindle pole body replicational cycle is described. Key words: electron microscopy, freeze-substitution, meiosis, Ustilago, spindle pole body.

scholarly journals Where does the peanut smut pathogen, Thecaphora frezii, fit in the spectrum of smut diseases?

Plant Disease ◽  
2021 ◽  
Author(s):  
Silvina Arias ◽  
Verónica Sofía Mary ◽  
Pilar Andrea Velez ◽  
María Gisel Rodriguez ◽  
Santiago Nicolás Otaiza González ◽  
...  
Keyword(s):  
South America ◽  
Pest Control ◽  
Food Quality ◽  
Defense Mechanisms ◽  
Management Strategies ◽  
Ustilago Maydis ◽  
Immune Defense ◽  
Smut Fungi ◽  
Host Interactions ◽  
Commercial Species

Smut fungi, such as Ustilago maydis, have been studied extensively as a model for plant- pathogenic basidiomycetes. However, little attention has been paid to smut diseases of agronomic importance that are caused by species of the fungus Thecaphora spp., probably due to their more localized distribution. Peanut smut by T. frezii has been reported only in South America, with Argentina being the only country where this disease has been noted in commercial species. In this work, important advances in deciphering T. frezii specific biology/pathobiology in relation to the agronomically relevant potato (T. solani), wheat (U. tritici) and barley (U. nuda) smuts are presented. The state of knowledge of fungal effectors, functionally characterized to date in U. maydis and most recently in T. thlaspeos, as well as the potential to be present in other Thecaphora species involved in dicot-host interactions like T. frezii-peanut, is summarized. We also discuss the applicability and limitations of current available methods for the identification of smut fungi in different matrices, and the management strategies to reduce their impact on the agri-food quality. To conclude, we describe some of the challenges in elucidating T. frezii strategies which allow it to successfully infect the host, and tolerate or evade plant immune defense mechanisms, as well as analysis of other aspects related to pest control and their implications for human health.

scholarly journals Fungal Pathogen Emergence: Investigations with an Ustilago maydis × Sporisorium reilianum Hybrid

2021 ◽  
Vol 7 (8) ◽  
pp. 672
Author(s):  
Emilee R. M. Storfie ◽  
Barry J. Saville
Keyword(s):  
Zea Mays ◽  
Fungal Pathogen ◽  
Fungal Pathogens ◽  
Crop Production ◽  
Virulence Genes ◽  
Ustilago Maydis ◽  
Smut Fungi ◽  
Transcript Levels ◽  
Sporisorium Reilianum ◽  
Molecular Aspects

The emergence of new fungal pathogens threatens sustainable crop production worldwide. One mechanism by which new pathogens may arise is hybridization. To investigate hybridization, the related smut fungi, Ustilago maydis and Sporisorium reilianum, were selected because they both infect Zea mays, can hybridize, and tools are available for their analysis. The hybrid dikaryons of these fungi grew as filaments on plates but their colonization and virulence in Z. mays were reduced compared to the parental dikaryons. The anthocyanin induction caused by the hybrid dikaryon infections was distinct, suggesting its interaction with the host was different from that of the parental dikaryons. Selected virulence genes previously characterized in U. maydis and their predicted S. reilianum orthologs had altered transcript levels during hybrid infection of Z. mays. The downregulated U. maydis effectors, tin2, pit2, and cce1, and transcription factors, rbf1, hdp2, and nlt1, were constitutively expressed in the hybrid. Little impact was observed with increased effector expression; however, increased expression of rbf1 and hdp2, which regulate early pathogenic development by U. maydis, increased the hybrid’s capacity to induce symptoms including the rare induction of small leaf tumors. These results establish a base for investigating molecular aspects of smut fungal hybrid pathogen emergence.

Detection of DsRNA Viruses in Isolates of Australian Smut Fungi and Their Serological Relationship to Viruses Found in Ustilago maydis From the U.S.A.

1991 ◽  
Vol 39 (1) ◽  
pp. 59 ◽  
Author(s):  
MG Francki ◽  
GC Kirby
Keyword(s):  
Gel Electrophoresis ◽  
Ustilago Maydis ◽  
Smut Fungi ◽  
Agarose Gel ◽  
Agarose Gel Electrophoresis ◽  
Serological Relationship ◽  
Killer Activity ◽  
Virus Like Particle ◽  
Low Salt ◽  
Dsrna Viruses

DsRNA was detected in two killer strains of smut fungi collected in Australia. This was determined by digestion of dsRNA preparations with RNase in the presence of high and low salt concentrations and detection of the nucleic acid using agarose gel electrophoresis. Isometric particles approximately 40 nm in diameter were detected using electron microscopy in the same isolates. These particles were shown to be serologically related but not identical to the P1 Ustilago maydis. virus-like particle (VLP) as determined by immunodiffusion using an antiserum to the P1 VLP. This shows that VLPs similar to those in North America also occur in Australia. Neither isometric particles nor dsRNA were detected in three other killer strains of smut fungi or in 11 sensitive strains. This indicates that there is a lack of correlation between killer activity and the presence of a dsRNA VLP in some Australian isolates of smut fungi.

scholarly journals Mating Type Loci of Sporisorium reilianum: Novel Pattern with Three a and Multiple b Specificities

2005 ◽  
Vol 4 (8) ◽  
pp. 1317-1327 ◽  
Author(s):  
Jan Schirawski ◽  
Bernadette Heinze ◽  
Martin Wagenknecht ◽  
Regine Kahmann
Keyword(s):  
Mating Type ◽  
Molecular Basis ◽  
Plant Invasion ◽  
Ustilago Maydis ◽  
Smut Fungi ◽  
Symptom Development ◽  
Receptor System ◽  
Mating Partner ◽  
Sporisorium Reilianum ◽  
High Degree

ABSTRACT Sporisorium reilianum and Ustilago maydis are two closely related smut fungi, which both infect maize but differ fundamentally in their mode of plant invasion and site of symptom development. As a prelude to studying the molecular basis of these differences, we have characterized the mating type loci of S. reilianum. S. reilianum has two unlinked mating type loci, a and b. Genes in both loci and adjacent regions show a high degree of synteny to the corresponding genes of U. maydis. The b locus occurs in at least five alleles and encodes two subunits of a heterodimeric homeodomain transcription factor, while the a locus encodes a pheromone/receptor system. However, in contrast to that of U. maydis, the a locus of S. reilianum exists in three alleles containing two active pheromone genes each. The alleles of the a locus appear to have arisen through recent recombination events within the locus itself. This has created a situation where each pheromone is specific for recognition by only one mating partner.

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