Tagging pathogenicity genes inUstilago maydis by restriction enzyme-mediated integration (REMI)

Bölker, Michael; Böhnert, Heidi U.; Braun, Karl Heinz; Görl, Johannes; Kahmann, Regine

doi:10.1007/bf02423450

Cited by 188 publications

(155 citation statements)

References 25 publications

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“…Infections with both combinations caused normal disease symptoms (Table 1). To exclude the possibility that our results reflect a cell fusion defect of the Dpmt4 strains, we generated a deletion of pmt4 in the solopathogenic SG200 strain (a1 mfa2 bE1bW2), which is able to infect plants without mating as it possesses both a and b loci active (Bö lker et al, 1995). Deletion of pmt4 in SG200 also resulted in a total loss of infectivity (Table 1).…”

Section: Pmt4 Is Required For Virulence On Maizementioning

confidence: 99%

TheO-Mannosyltransferase PMT4 Is Essential for Normal Appressorium Formation and Penetration inUstilago maydis

2009

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In Saccharomyces cerevisiae, the PMT, KRE2/MNT1, and MNN1 mannosyltransferase protein families catalyze the steps of the O-mannosylation pathway, sequentially adding mannoses to target proteins. We have identified members of all three families and analyzed their roles in pathogenesis of the maize smut fungus Ustilago maydis. Furthermore, we have shown that PMT4, one of the three PMT family members in U. maydis, is essential for tumor formation in Zea mays. Significantly, PMT4 seems to be required only for pathogenesis and is dispensable for other aspects of the U. maydis life cycle. We subsequently show that the deletion of pmt4 results in a strong reduction in the frequency of appressorium formation, with the few appressoria that do form lacking the capacity to penetrate the plant cuticle. Our findings suggest that the O-mannosylation pathway plays a key role in the posttranslational modification of proteins involved in the pathogenic development of U. maydis. The fact that PMT homologs are not found in plants may open new avenues for the development of fungal control strategies. Moreover, the discovery of a highly specific requirement for a single O-mannosyltransferase should aid in the identification of the proteins directly involved in fungal plant penetration, thus leading to a better understanding of plant-fungi interactions.

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Section: Pmt4 Is Required For Virulence On Maizementioning

confidence: 99%

TheO-Mannosyltransferase PMT4 Is Essential for Normal Appressorium Formation and Penetration inUstilago maydis

2009

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“…To address this question we exchanged the native wee1 promoter with the scp promoter, a weak U. maydis constitutive promoter that produces low levels of expression (Bölker et al, 1995a). The abundance of the mRNA produced by this promoter did not vary as a function of the nutritional conditions, and was similar to the wee1 levels produced in wild-type cells growing in rich medium (YPD, Fig.…”

Section: Wee1 Controls the Cdk1-clb2 Complexmentioning

confidence: 99%

Inhibitory phosphorylation of a mitotic cyclin-dependent kinase regulates the morphogenesis, cell size and virulence of the smut fungusUstilago maydis

Sgarlata

Pérez-Martı́n

2005

Journal of Cell Science

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The regulation of cyclin-dependent kinase (CDK) activity through inhibitory phosphorylation seems to play an important role in the eukaryotic cell cycle. We have investigated the influence that inhibitory phosphorylation of the catalytic subunit of mitotic CDK has on cell growth and pathogenicity of the corn smut fungus Ustilago maydis. This model pathogen is worthy of attention since it is well suited to analyze the relationships between the cell cycle, morphogenesis and pathogenicity. We set out to study these relationships by producing a cdk1 mutant allele that was refractory to inhibitory phosphorylation. The expression of this mutant in U. maydis cells dramatically altered their morphology. Since this kind of mutation makes the CDK catalytic subunit resistant to regulation by Wee1-related kinases in other organisms, we characterized the orthologous Wee1 kinase from U. maydis. We found that Wee1 is essential in U. maydis. Overexpression of wee1 produces cell cycle arrest in G2, the target of Wee1 apparently being the Cdk1/Clb2 complex, which is required specifically for the onset of mitosis. Given the connection between the cell cycle control and pathogenesis in U. maydis, we also analyzed whether cells with impaired inhibitory phosphorylation of Cdk1 were able to infect plants. We found that inhibitory phosphorylation was required for mating, a prerequisite to initiate pathogenic development. By examining plant-specific expression of the constitutively unphosphorylated cdk1 AF allele, we also found that appropriate levels of inhibitory phosphorylation were required at stages of infection subsequent to penetration by the fungus. These data reinforces the connections between cell cycle, morphogenesis and virulence in this smut fungus.

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“…By increasing the target sites for recombination, REMI usually improves the overall transformation rate and frequency of single copy integration events [49,55,[59][60][61][62][63][64] although that is not always the case [50,65]. Conditions of transformation contribute significantly to the variable success rate of REMI.…”

Section: Gene Tagging By Direct Dna Transfermentioning

confidence: 99%

Approaches to functional genomics in filamentous fungi

et al. 2006

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The study of gene function in filamentous fungi is a field of research that has made great advances in very recent years. A number of transformation and gene manipulation strategies have been developed and applied to a diverse and rapidly expanding list of economically important filamentous fungi and oomycetes. With the significant number of fungal genomes now sequenced or being sequenced, functional genomics promises to uncover a great deal of new information in coming years. This review discusses recent advances that have been made in examining gene function in filamentous fungi and describes the advantages and limitations of the different approaches.

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Tagging pathogenicity genes inUstilago maydis by restriction enzyme-mediated integration (REMI)

Cited by 188 publications

References 25 publications

TheO-Mannosyltransferase PMT4 Is Essential for Normal Appressorium Formation and Penetration inUstilago maydis

TheO-Mannosyltransferase PMT4 Is Essential for Normal Appressorium Formation and Penetration inUstilago maydis

Inhibitory phosphorylation of a mitotic cyclin-dependent kinase regulates the morphogenesis, cell size and virulence of the smut fungusUstilago maydis

Approaches to functional genomics in filamentous fungi

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