0000000000729596
AUTHOR
Alexander Yemelin
Histidine kinases mediate differentiation, stress response, and pathogenicity in Magnaporthe oryzae.
The aim of this study is a functional characterization of 10 putative histidine kinases (HIKs)-encoding genes in the phytopathogenic fungus Magnaporthe oryzae. Two HIKs were found to be required for pathogenicity in the fungus. It was found that the mutant strains ΔMohik5 and ΔMohik8 show abnormal conidial morphology and furthermore ΔMohik5 is unable to form appressoria. Both HIKs MoHik5p and MoHik8p appear to be essential for pathogenicity since the mutants fail to infect rice plants. MoSln1p and MoHik1p were previously reported to be components of the HOG pathway in M. oryzae. The ΔMosln1 mutant is more susceptible to salt stress compared to ΔMohik1, whereas ΔMohik1 appears to be stronger…
Rapid adaptation of signaling networks in the fungal pathogen Magnaporthe oryzae
Abstract Background One fundamental question in biology is how the evolution of eukaryotic signaling networks has taken place. “Loss of function” (lof) mutants from components of the high osmolarity glycerol (HOG) signaling pathway in the filamentous fungus Magnaporthe oryzae are viable, but impaired in osmoregulation. Results After long-term cultivation upon high osmolarity, stable individuals with reestablished osmoregulation capacity arise independently from each of the mutants with inactivated HOG pathway. This phenomenon is extremely reproducible and occurs only in osmosensitive mutants related to the HOG pathway – not in other osmosensitive Magnaporthe mutants. The major compatible so…
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Additional file 7: Figure S6. qPCR results of selected genes. qRT-PCR analysis of putative genes in MoWT, the “lof” mutants ΔMohog1 and ΔMohog1(adapted). The M. oryzae cultures were grown for 96 h in CM at 26 °C and 100 rpm. Each of the cultures was separated into two samples, one mixed with 0.5 M KCl and one untreated control further grown in CM at 26 °C and 100 rpm). Samples were taken after 25 min. The RNA was isolated from the mycelium samples and the results of transcript abundance given relative to quantification in the MoWT untreated control. Three biological replicates were used of each.
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Additional file 3: Figure S2. Investigation of the â adaptation-frequencyâ in Magnaporthe oryzae mutants with inactivated components of the HOG signaling cascade.
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Additional file 4: Figure S3. Mycelium dry weight of the Magnaporthe oryzae wildtype strain, mutants with inactivated components of the HOG signaling cascade and the “adapted” strains after growth in liquid culture upon sorbitol-stress. The fungal colonies were grown in 250 ml complete medium inclusive 1,5 M sorbitol for 6 d at 26 °C and 120 rpm. Error bars represent the standard deviation of three biological replicates of each strain.
Fungicide resistance towards fludioxonil conferred by overexpression of the phosphatase gene Mo PTP 2 in Magnaporthe oryzae
The fungicide fludioxonil causes hyperactivation of the Hog1p MAPK within the high-osmolarity glycerol signaling pathway essential for osmoregulation in pathogenic fungi. The molecular regulation of MoHog1p phosphorylation is not completely understood in pathogenic fungi. Thus, we identified and characterized the putative MoHog1p-interacting phosphatase gene MoPTP2 in the filamentous rice pathogen Magnaporthe oryzae. We found overexpression of MoPTP2 conferred fludioxonil resistance in M. oryzae, whereas the 'loss of function' mutant ΔMoptp2 was more susceptible toward the fungicide. Additionally, quantitative phosphoproteome profiling of MoHog1p phosphorylation revealed lower phosphorylati…
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Additional file 6: Figure S5. VENN diagram of putative structural variations in promotor [A] and in coding sequences (CDS) [B] within the genome of ΔMohog1, ΔMohog1(adapted) and ΔMopbs2(adapted). Numbers in the intersection regions represent overlapping SNPs among the strains. Numbers in parentheses show the corresponding relative percentage of genes harbouring the SNPs.
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Additional file 5: Figure S4. Pathogenicity assay of the MoWT, the lof mutants and the â adaptedâ strains. The plant infection assays were carried out as described in experimental procedures. The error bars represent the standard deviation of three experiments with three replicates each.
The exceptionality of stress response in Magnaporthe oryzae: a set of “salt stress-induced” genes unique to the rice blast fungus
The ability of pathogens to signal perception and adaptation to environmental changes is an important prerequisite for successful colonization of the host organism. Filamentous phytopathogenic fungi, for example, have to cope with rapid changes in the environment during invasive growth in planta. Consequently, they have evolved a range of specific factors contributing to environmental adaptation facilitating host invasion. In addition to conserved pathways, including genes participating in stress response, unique/individual genes within the pathogens might represent determinants of pathogenicity. Therefore, identification of unique genes could provide a set of excellent candidates for novel…
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Additional file 2: Figure S1. Schematic presentation and verification of the MoWT, the lof-mutants and the adapted strains by southern hybridization within the Magnaporthe oryzae genome. Southern blot analysis of gene deletion/disruption mutants in M. oryzae with gene specific probes. Genomic DNA of M. oryzae strain 70â 15 and the mutants was isolated and restricted with restriction enzymes. The probes which we used for hybridization with the genomic DNA of the wildtype strain and the corresponding mutant strains were always identical.
High osmolarity glycerol (HOG) signalling in Magnaporthe oryzae: Identification of MoYPD1 and its role in osmoregulation, fungicide action, and pathogenicity
AbstractThis study comprises a first functional analysis of an YPD1-homologue in filamentous phytopathogenic fungi and its role in the HOG signalling pathway. We generated a gene deletion mutant of the gene MoYPD1 in Magnaporthe oryzae and characterized the resulting mutant strain. We have shown that MoYpd1p is a component of the phosphorelay system acting in the HOG pathway due to its Y2H protein interaction with the HKs MoHik1p and MoSln1p as well as with the response regulator MoSsk1p. Fungicidal activity of fludioxonil was reported to be based on the inhibition of MoHik1p resulting in hyperactivation of the HOG signalling pathway and lethality. Western analysis proved that both, osmotic…
Evidence of a New MoYpd1p Phosphotransferase Isoform in the Multistep Phosphorelay System of Magnaporthe oryzae
Different external stimuli are perceived by multiple sensor histidine kinases and transmitted by phosphorylation via the phosphotransfer protein Ypd1p in the multistep phosphorelay system of the high osmolarity glycerol signaling pathway of filamentous fungi. How the signal propagation takes place is still not known in detail since multiple sensor histidine kinase genes in most filamentous fungi are coded in the genome, whereas only one gene for Ypd1p exists. That raises the hypothesis that various Ypd1p isoforms are produced from a single gene sequence, perhaps by alternative splicing, facilitating a higher variability in signal transduction. We found that the mRNA of MoYPD1 in the rice bl…
Identification of factors involved in dimorphism and pathogenicity of Zymoseptoria tritici
A forward genetics approach was applied in order to investigate the molecular basis of morphological transition in the wheat pathogenic fungus Zymoseptoria tritici. Z. tritici is a dimorphic plant pathogen displaying environmentally regulated morphogenetic transition between yeast-like and hyphal growth. Considering the infection mode of Z. tritici, the switching to hyphal growth is essential for pathogenicity allowing the fungus the host invasion through natural openings like stomata. We exploited a previously developed Agrobacterium tumefaciens-mediated transformation (ATMT) to generate a mutant library by insertional mutagenesis including more than 10,000 random mutants. To identify gene…
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Additional file 1: SVs summary.