Limits...
A novel pathogenicity gene is required in the rice blast fungus to suppress the basal defenses of the host.

Chi MH, Park SY, Kim S, Lee YH - PLoS Pathog. (2009)

Bottom Line: For successful colonization and further reproduction in host plants, pathogens need to overcome the innate defenses of the plant.Targeted gene deletion of DES1 had no apparent effect on developmental morphogenesis, including vegetative growth, conidial germination, appressorium formation, and appressorium-mediated penetration.These results suggest that DES1 functions as a novel pathogenicity gene that regulates the activity of fungal proteins, compromising ROS-mediated plant defense.

View Article: PubMed Central - PubMed

Affiliation: Department of Agricultural Biotechnology, Center for Fungal Genetic Resources, and Center for Fungal Pathogenesis, Seoul National University, Seoul, Korea.

ABSTRACT
For successful colonization and further reproduction in host plants, pathogens need to overcome the innate defenses of the plant. We demonstrate that a novel pathogenicity gene, DES1, in Magnaporthe oryzae regulates counter-defenses against host basal resistance. The DES1 gene was identified by screening for pathogenicity-defective mutants in a T-DNA insertional mutant library. Bioinformatic analysis revealed that this gene encodes a serine-rich protein that has unknown biochemical properties, and its homologs are strictly conserved in filamentous Ascomycetes. Targeted gene deletion of DES1 had no apparent effect on developmental morphogenesis, including vegetative growth, conidial germination, appressorium formation, and appressorium-mediated penetration. Conidial size of the mutant became smaller than that of the wild type, but the mutant displayed no defects on cell wall integrity. The Deltades1 mutant was hypersensitive to exogenous oxidative stress and the activity and transcription level of extracellular enzymes including peroxidases and laccases were severely decreased in the mutant. In addition, ferrous ion leakage was observed in the Deltades1 mutant. In the interaction with a susceptible rice cultivar, rice cells inoculated with the Deltades1 mutant exhibited strong defense responses accompanied by brown granules in primary infected cells, the accumulation of reactive oxygen species (ROS), the generation of autofluorescent materials, and PR gene induction in neighboring tissues. The Deltades1 mutant displayed a significant reduction in infectious hyphal extension, which caused a decrease in pathogenicity. Notably, the suppression of ROS generation by treatment with diphenyleneiodonium (DPI), an inhibitor of NADPH oxidases, resulted in a significant reduction in the defense responses in plant tissues challenged with the Deltades1 mutant. Furthermore, the Deltades1 mutant recovered its normal infectious growth in DPI-treated plant tissues. These results suggest that DES1 functions as a novel pathogenicity gene that regulates the activity of fungal proteins, compromising ROS-mediated plant defense.

Show MeSH

Related in: MedlinePlus

DES1 is related to activity of extracellular peroxidase and laccase.(A) The discoloration of Congo Red was tested on medium containing 100 ppm of the dye at final concentration. Strains were inoculated on CM agar medium containing Congo Red. Discoloration was observed on day 4. Left: wild type, middle: DES1T-DNA, right: Δdes1. (B) Peroxidase activity measured by ABTS oxidizing test under H2O2 supplemented condition (see Materials and Methods for details). Black column: wild type, grey column: DES1T-DNA, white column: Δdes1. (C) Laccase activity measured by ABTS oxidizing test without H2O2. The strain scheme is same with panel B. Error bars represent standard deviation.
© Copyright Policy
Related In: Results  -  Collection


getmorefigures.php?uid=PMC2668191&req=5

ppat-1000401-g007: DES1 is related to activity of extracellular peroxidase and laccase.(A) The discoloration of Congo Red was tested on medium containing 100 ppm of the dye at final concentration. Strains were inoculated on CM agar medium containing Congo Red. Discoloration was observed on day 4. Left: wild type, middle: DES1T-DNA, right: Δdes1. (B) Peroxidase activity measured by ABTS oxidizing test under H2O2 supplemented condition (see Materials and Methods for details). Black column: wild type, grey column: DES1T-DNA, white column: Δdes1. (C) Laccase activity measured by ABTS oxidizing test without H2O2. The strain scheme is same with panel B. Error bars represent standard deviation.

Mentions: Since the conidial morphology of Δdes1and DES1T-DNA was altered, hypersensitivity to oxidative stress and reduced virulence may also be due to defects in cell wall composition, in spite of their insensitivities to osmotic stresses. To investigate this possibility, we added Nikkomycin Z, a chitin synthetase inhibitor, to germinating conidia. Treatment of Nikkomycin Z blocks conidial germination and induces protoplast-like swellings on cell wall-defective strains [48]. However, conidial germination of Δdes1and DES1T-DNA was not inhibited in high concentrations of Nikkomycin Z (100 µM) and swellings on germ tubes were not distinguishable from the wild type (Fig. S7A). We also tested the sensitivity of these strains to a cell wall-degrading enzyme. Enzyme-treated mycelia of Δdes1and DES1T-DNA released no more or less protoplasts than the wild type when observed over a time course (Fig. S7B). We also tested mycelial growth on Calcofluor white (CFW) and Congo Red (CR) amended media, which inhibit fungal cell wall assembly by binding chitin and β-1,4-glucans, respectively [49],[50]. The mycelial growth of Δdes1on CFW media (100 ppm) was little reduced (88% of the wild type) when compared with normal CM (96% of the wild type), and it was more severely reduced (70% of the wild type) on CR media (100 ppm). However, since degradation halo was observed around the wild-type colony and no degradation halo was observed around the Δdes1 colony (Fig. 7A), the growth defect on CR media was assumed to be due to the absence of CR-degrading activity rather than defects in cell wall composition. The DES1T-DNA colonies showed intermediated levels of CR discoloration (Fig. 7A).


A novel pathogenicity gene is required in the rice blast fungus to suppress the basal defenses of the host.

Chi MH, Park SY, Kim S, Lee YH - PLoS Pathog. (2009)

DES1 is related to activity of extracellular peroxidase and laccase.(A) The discoloration of Congo Red was tested on medium containing 100 ppm of the dye at final concentration. Strains were inoculated on CM agar medium containing Congo Red. Discoloration was observed on day 4. Left: wild type, middle: DES1T-DNA, right: Δdes1. (B) Peroxidase activity measured by ABTS oxidizing test under H2O2 supplemented condition (see Materials and Methods for details). Black column: wild type, grey column: DES1T-DNA, white column: Δdes1. (C) Laccase activity measured by ABTS oxidizing test without H2O2. The strain scheme is same with panel B. Error bars represent standard deviation.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC2668191&req=5

ppat-1000401-g007: DES1 is related to activity of extracellular peroxidase and laccase.(A) The discoloration of Congo Red was tested on medium containing 100 ppm of the dye at final concentration. Strains were inoculated on CM agar medium containing Congo Red. Discoloration was observed on day 4. Left: wild type, middle: DES1T-DNA, right: Δdes1. (B) Peroxidase activity measured by ABTS oxidizing test under H2O2 supplemented condition (see Materials and Methods for details). Black column: wild type, grey column: DES1T-DNA, white column: Δdes1. (C) Laccase activity measured by ABTS oxidizing test without H2O2. The strain scheme is same with panel B. Error bars represent standard deviation.
Mentions: Since the conidial morphology of Δdes1and DES1T-DNA was altered, hypersensitivity to oxidative stress and reduced virulence may also be due to defects in cell wall composition, in spite of their insensitivities to osmotic stresses. To investigate this possibility, we added Nikkomycin Z, a chitin synthetase inhibitor, to germinating conidia. Treatment of Nikkomycin Z blocks conidial germination and induces protoplast-like swellings on cell wall-defective strains [48]. However, conidial germination of Δdes1and DES1T-DNA was not inhibited in high concentrations of Nikkomycin Z (100 µM) and swellings on germ tubes were not distinguishable from the wild type (Fig. S7A). We also tested the sensitivity of these strains to a cell wall-degrading enzyme. Enzyme-treated mycelia of Δdes1and DES1T-DNA released no more or less protoplasts than the wild type when observed over a time course (Fig. S7B). We also tested mycelial growth on Calcofluor white (CFW) and Congo Red (CR) amended media, which inhibit fungal cell wall assembly by binding chitin and β-1,4-glucans, respectively [49],[50]. The mycelial growth of Δdes1on CFW media (100 ppm) was little reduced (88% of the wild type) when compared with normal CM (96% of the wild type), and it was more severely reduced (70% of the wild type) on CR media (100 ppm). However, since degradation halo was observed around the wild-type colony and no degradation halo was observed around the Δdes1 colony (Fig. 7A), the growth defect on CR media was assumed to be due to the absence of CR-degrading activity rather than defects in cell wall composition. The DES1T-DNA colonies showed intermediated levels of CR discoloration (Fig. 7A).

Bottom Line: For successful colonization and further reproduction in host plants, pathogens need to overcome the innate defenses of the plant.Targeted gene deletion of DES1 had no apparent effect on developmental morphogenesis, including vegetative growth, conidial germination, appressorium formation, and appressorium-mediated penetration.These results suggest that DES1 functions as a novel pathogenicity gene that regulates the activity of fungal proteins, compromising ROS-mediated plant defense.

View Article: PubMed Central - PubMed

Affiliation: Department of Agricultural Biotechnology, Center for Fungal Genetic Resources, and Center for Fungal Pathogenesis, Seoul National University, Seoul, Korea.

ABSTRACT
For successful colonization and further reproduction in host plants, pathogens need to overcome the innate defenses of the plant. We demonstrate that a novel pathogenicity gene, DES1, in Magnaporthe oryzae regulates counter-defenses against host basal resistance. The DES1 gene was identified by screening for pathogenicity-defective mutants in a T-DNA insertional mutant library. Bioinformatic analysis revealed that this gene encodes a serine-rich protein that has unknown biochemical properties, and its homologs are strictly conserved in filamentous Ascomycetes. Targeted gene deletion of DES1 had no apparent effect on developmental morphogenesis, including vegetative growth, conidial germination, appressorium formation, and appressorium-mediated penetration. Conidial size of the mutant became smaller than that of the wild type, but the mutant displayed no defects on cell wall integrity. The Deltades1 mutant was hypersensitive to exogenous oxidative stress and the activity and transcription level of extracellular enzymes including peroxidases and laccases were severely decreased in the mutant. In addition, ferrous ion leakage was observed in the Deltades1 mutant. In the interaction with a susceptible rice cultivar, rice cells inoculated with the Deltades1 mutant exhibited strong defense responses accompanied by brown granules in primary infected cells, the accumulation of reactive oxygen species (ROS), the generation of autofluorescent materials, and PR gene induction in neighboring tissues. The Deltades1 mutant displayed a significant reduction in infectious hyphal extension, which caused a decrease in pathogenicity. Notably, the suppression of ROS generation by treatment with diphenyleneiodonium (DPI), an inhibitor of NADPH oxidases, resulted in a significant reduction in the defense responses in plant tissues challenged with the Deltades1 mutant. Furthermore, the Deltades1 mutant recovered its normal infectious growth in DPI-treated plant tissues. These results suggest that DES1 functions as a novel pathogenicity gene that regulates the activity of fungal proteins, compromising ROS-mediated plant defense.

Show MeSH
Related in: MedlinePlus