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The transcription factor Ste12 mediates the regulatory role of the Tmk1 MAP kinase in mycoparasitism and vegetative hyphal fusion in the filamentous fungus Trichoderma atroviride.

Gruber S, Zeilinger S - PLoS ONE (2014)

Bottom Line: However, the transcription factors acting downstream of Tmk1 are hitherto unknown.Here we analyzed the functions of the T. atroviride Ste12 transcription factor whose orthologue in yeast is targeted by the Fus3 and Kss1 MAP kinases.Aerial hyphae formation and conidiation, in contrast, were found to be independent of Ste12.

View Article: PubMed Central - PubMed

Affiliation: Research Area Biotechnology and Microbiology, Institute of Chemical Engineering, Vienna University of Technology, Wien, Austria.

ABSTRACT
Mycoparasitic species of the fungal genus Trichoderma are potent antagonists able to combat plant pathogenic fungi by direct parasitism. An essential step in this mycoparasitic fungus-fungus interaction is the detection of the fungal host followed by activation of molecular weapons in the mycoparasite by host-derived signals. The Trichoderma atroviride MAP kinase Tmk1, a homolog of yeast Fus3/Kss1, plays an essential role in regulating the mycoparasitic host attack, aerial hyphae formation and conidiation. However, the transcription factors acting downstream of Tmk1 are hitherto unknown. Here we analyzed the functions of the T. atroviride Ste12 transcription factor whose orthologue in yeast is targeted by the Fus3 and Kss1 MAP kinases. Deletion of the ste12 gene in T. atroviride not only resulted in reduced mycoparasitic overgrowth and lysis of host fungi but also led to loss of hyphal avoidance in the colony periphery and a severe reduction in conidial anastomosis tube formation and vegetative hyphal fusion events. The transcription of several orthologues of Neurospora crassa hyphal fusion genes was reduced upon ste12 deletion; however, the Δste12 mutant showed enhanced expression of mycoparasitism-relevant chitinolytic and proteolytic enzymes and of the cell wall integrity MAP kinase Tmk2. Based on the comparative analyses of Δste12 and Δtmk1 mutants, an essential role of the Ste12 transcriptional regulator in mediating outcomes of the Tmk1 MAPK pathway such as regulation of the mycoparasitic activity, hyphal fusion and carbon source-dependent vegetative growth is suggested. Aerial hyphae formation and conidiation, in contrast, were found to be independent of Ste12.

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Comparative carbon source utilization profiles of the Δste12 and Δtmk1 mutants and the parental strain (WT).Strains were grown on 95 carbon sources (FF-plates) using the Biolog Phenotype Microarray system. Hierarchical clustering results are displayed as coloured mosaics attached to a dendrogram according to growth (OD750) after 48 hours of incubation. All data points are the average of three replicates. The threshold cut-off for all conditions was set at 0.225 which corresponded to growth of the parental strain on the water control. Red boxes indicate maximal growth (OD750 0.7–1.3), black boxes medium growth (OD750 0.55–0.7), and green boxes weak growth (OD750 0.2–0.55).
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pone-0111636-g003: Comparative carbon source utilization profiles of the Δste12 and Δtmk1 mutants and the parental strain (WT).Strains were grown on 95 carbon sources (FF-plates) using the Biolog Phenotype Microarray system. Hierarchical clustering results are displayed as coloured mosaics attached to a dendrogram according to growth (OD750) after 48 hours of incubation. All data points are the average of three replicates. The threshold cut-off for all conditions was set at 0.225 which corresponded to growth of the parental strain on the water control. Red boxes indicate maximal growth (OD750 0.7–1.3), black boxes medium growth (OD750 0.55–0.7), and green boxes weak growth (OD750 0.2–0.55).

Mentions: In order to get additional insights into the phenotypic consequences resulting from ste12 deletion and to learn more on a putative involvement of Ste12 in the Tmk1 MAPK pathway, we performed comparative nutrient profiling of Δste12 and Δtmk1 mutants. The carbon-source utilization profile of T. atroviride on 95 different carbon sources has been characterized previously resulting in four clusters [33]. Clusters I (comprising mainly monosaccharides and polyols, but also γ-amino-butyric acid and N-acetyl-D-glucosamine), II (several monosaccharides, some oligosaccharides and arylglucosides), and III (mainly disaccharides and oligosaccharides, arylglucosides and L-amino acids) contain carbon sources allowing fast, moderate and slow growth, respectively, while carbon sources only allowing very poor or no growth at all are contained in cluster IV (containing several L-amino acids, peptides, amines, TCA-intermediates, aliphatic organic acids). Analysis of growth of the Δste12 and Δtmk1 mutants on the 95 carbon sources revealed similar carbon utilization profiles for both mutants which, however, significantly differed from the parental strain (Fig. 3). Amongst the cluster I-III carbon sources, D-mannose, D-ribose, dextrin, salicin, amygdalin, L-arabinose, succinic acid, and β-methyl-D-glucoside only allowed reduced growth of both mutants compared to the parental strain. On the other hand, both, Δste12 and Δtmk1, exhibited enhanced growth on 2-keto-D-gluconic acid, glycerol, maltotriose, quinic acid and the amino acids and amino acid derivatives L-glutamic acid, L-asparagine, L-aspartic acid, L-threonine, L-pyroglutamic acid, and L-alanyl-glycine. In addition to this congruent behavior, also differences in the nutritional profiles between Δste12 and Δtmk1 mutants were evident. While on i-erythritol, the Δtmk1 mutant showed similar growth than the parental strain, the Δste12 mutant showed significantly reduced growth. Vice versa, deletion of tmk1, but not ste12, resulted in reduced growth on γ-amino-butyric acid which, together with N-acetyl-D-glucosamine, was the best carbon source for the T. atroviride parental strain. On the other hand, the Δtmk1 mutant exhibited better growth compared to the ste12 deletion mutant and the parental strain on the monosaccharides D-trehalose, D-xylose, and D-fructose, the disaccharide cellobiose and the tetrasaccharide stachyose, while Δste12 grew better on the polyols D-mannitol and D-arabitol.


The transcription factor Ste12 mediates the regulatory role of the Tmk1 MAP kinase in mycoparasitism and vegetative hyphal fusion in the filamentous fungus Trichoderma atroviride.

Gruber S, Zeilinger S - PLoS ONE (2014)

Comparative carbon source utilization profiles of the Δste12 and Δtmk1 mutants and the parental strain (WT).Strains were grown on 95 carbon sources (FF-plates) using the Biolog Phenotype Microarray system. Hierarchical clustering results are displayed as coloured mosaics attached to a dendrogram according to growth (OD750) after 48 hours of incubation. All data points are the average of three replicates. The threshold cut-off for all conditions was set at 0.225 which corresponded to growth of the parental strain on the water control. Red boxes indicate maximal growth (OD750 0.7–1.3), black boxes medium growth (OD750 0.55–0.7), and green boxes weak growth (OD750 0.2–0.55).
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4214791&req=5

pone-0111636-g003: Comparative carbon source utilization profiles of the Δste12 and Δtmk1 mutants and the parental strain (WT).Strains were grown on 95 carbon sources (FF-plates) using the Biolog Phenotype Microarray system. Hierarchical clustering results are displayed as coloured mosaics attached to a dendrogram according to growth (OD750) after 48 hours of incubation. All data points are the average of three replicates. The threshold cut-off for all conditions was set at 0.225 which corresponded to growth of the parental strain on the water control. Red boxes indicate maximal growth (OD750 0.7–1.3), black boxes medium growth (OD750 0.55–0.7), and green boxes weak growth (OD750 0.2–0.55).
Mentions: In order to get additional insights into the phenotypic consequences resulting from ste12 deletion and to learn more on a putative involvement of Ste12 in the Tmk1 MAPK pathway, we performed comparative nutrient profiling of Δste12 and Δtmk1 mutants. The carbon-source utilization profile of T. atroviride on 95 different carbon sources has been characterized previously resulting in four clusters [33]. Clusters I (comprising mainly monosaccharides and polyols, but also γ-amino-butyric acid and N-acetyl-D-glucosamine), II (several monosaccharides, some oligosaccharides and arylglucosides), and III (mainly disaccharides and oligosaccharides, arylglucosides and L-amino acids) contain carbon sources allowing fast, moderate and slow growth, respectively, while carbon sources only allowing very poor or no growth at all are contained in cluster IV (containing several L-amino acids, peptides, amines, TCA-intermediates, aliphatic organic acids). Analysis of growth of the Δste12 and Δtmk1 mutants on the 95 carbon sources revealed similar carbon utilization profiles for both mutants which, however, significantly differed from the parental strain (Fig. 3). Amongst the cluster I-III carbon sources, D-mannose, D-ribose, dextrin, salicin, amygdalin, L-arabinose, succinic acid, and β-methyl-D-glucoside only allowed reduced growth of both mutants compared to the parental strain. On the other hand, both, Δste12 and Δtmk1, exhibited enhanced growth on 2-keto-D-gluconic acid, glycerol, maltotriose, quinic acid and the amino acids and amino acid derivatives L-glutamic acid, L-asparagine, L-aspartic acid, L-threonine, L-pyroglutamic acid, and L-alanyl-glycine. In addition to this congruent behavior, also differences in the nutritional profiles between Δste12 and Δtmk1 mutants were evident. While on i-erythritol, the Δtmk1 mutant showed similar growth than the parental strain, the Δste12 mutant showed significantly reduced growth. Vice versa, deletion of tmk1, but not ste12, resulted in reduced growth on γ-amino-butyric acid which, together with N-acetyl-D-glucosamine, was the best carbon source for the T. atroviride parental strain. On the other hand, the Δtmk1 mutant exhibited better growth compared to the ste12 deletion mutant and the parental strain on the monosaccharides D-trehalose, D-xylose, and D-fructose, the disaccharide cellobiose and the tetrasaccharide stachyose, while Δste12 grew better on the polyols D-mannitol and D-arabitol.

Bottom Line: However, the transcription factors acting downstream of Tmk1 are hitherto unknown.Here we analyzed the functions of the T. atroviride Ste12 transcription factor whose orthologue in yeast is targeted by the Fus3 and Kss1 MAP kinases.Aerial hyphae formation and conidiation, in contrast, were found to be independent of Ste12.

View Article: PubMed Central - PubMed

Affiliation: Research Area Biotechnology and Microbiology, Institute of Chemical Engineering, Vienna University of Technology, Wien, Austria.

ABSTRACT
Mycoparasitic species of the fungal genus Trichoderma are potent antagonists able to combat plant pathogenic fungi by direct parasitism. An essential step in this mycoparasitic fungus-fungus interaction is the detection of the fungal host followed by activation of molecular weapons in the mycoparasite by host-derived signals. The Trichoderma atroviride MAP kinase Tmk1, a homolog of yeast Fus3/Kss1, plays an essential role in regulating the mycoparasitic host attack, aerial hyphae formation and conidiation. However, the transcription factors acting downstream of Tmk1 are hitherto unknown. Here we analyzed the functions of the T. atroviride Ste12 transcription factor whose orthologue in yeast is targeted by the Fus3 and Kss1 MAP kinases. Deletion of the ste12 gene in T. atroviride not only resulted in reduced mycoparasitic overgrowth and lysis of host fungi but also led to loss of hyphal avoidance in the colony periphery and a severe reduction in conidial anastomosis tube formation and vegetative hyphal fusion events. The transcription of several orthologues of Neurospora crassa hyphal fusion genes was reduced upon ste12 deletion; however, the Δste12 mutant showed enhanced expression of mycoparasitism-relevant chitinolytic and proteolytic enzymes and of the cell wall integrity MAP kinase Tmk2. Based on the comparative analyses of Δste12 and Δtmk1 mutants, an essential role of the Ste12 transcriptional regulator in mediating outcomes of the Tmk1 MAPK pathway such as regulation of the mycoparasitic activity, hyphal fusion and carbon source-dependent vegetative growth is suggested. Aerial hyphae formation and conidiation, in contrast, were found to be independent of Ste12.

Show MeSH
Related in: MedlinePlus