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Signaling governed by G proteins and cAMP is crucial for growth, secondary metabolism and sexual development in Fusarium fujikuroi.

Studt L, Humpf HU, Tudzynski B - PLoS ONE (2013)

Bottom Line: Here we studied the impact of the heterotrimeric G protein and the cAMP-mediated signaling network, including the regulatory subunits of the cAMP-dependent protein kinase (PKA), to study their effect on colony morphology, sexual development and regulation of bikaverins, fusarubins and GAs.In contrast, bikaverin biosynthesis is significantly reduced in ffg1 and ffg3 deletion mutants and positively regulated by FfAC and FfPKA1, while GA biosynthesis depends on the active FfAC and FfPKA2 in an FfG1- and FfG3-independent manner.In addition, we provide evidence that G Protein-mediated/cAMP signaling is important for growth in F. fujikuroi because deletion of ffg3, ffac and ffpka1 resulted in impaired growth on minimal and rich media.

View Article: PubMed Central - PubMed

Affiliation: Institut für Lebensmittelchemie, Westfälische Wilhelms-Universität, Münster, Germany.

ABSTRACT
The plant-pathogenic fungus Fusarium fujikuroi is a notorious rice pathogen causing hyper-elongation of infected plants due to the production of gibberellic acids (GAs). In addition to GAs, F. fujikuroi produces a wide range of other secondary metabolites, such as fusarins, fusaric acid or the red polyketides bikaverins and fusarubins. The recent availability of the fungal genome sequence for this species has revealed the potential of many more putative secondary metabolite gene clusters whose products remain to be identified. However, the complex regulation of secondary metabolism is far from being understood. Here we studied the impact of the heterotrimeric G protein and the cAMP-mediated signaling network, including the regulatory subunits of the cAMP-dependent protein kinase (PKA), to study their effect on colony morphology, sexual development and regulation of bikaverins, fusarubins and GAs. We demonstrated that fusarubin biosynthesis is negatively regulated by at least two Gα subunits, FfG1 and FfG3, which both function as stimulators of the adenylyl cyclase FfAC. Surprisingly, the primary downstream target of the adenylyl cyclase, the PKA, is not involved in the regulation of fusarubins, suggesting that additional, yet unidentified, cAMP-binding protein(s) exist. In contrast, bikaverin biosynthesis is significantly reduced in ffg1 and ffg3 deletion mutants and positively regulated by FfAC and FfPKA1, while GA biosynthesis depends on the active FfAC and FfPKA2 in an FfG1- and FfG3-independent manner. In addition, we provide evidence that G Protein-mediated/cAMP signaling is important for growth in F. fujikuroi because deletion of ffg3, ffac and ffpka1 resulted in impaired growth on minimal and rich media. Finally, sexual crosses of ffg1 mutants showed the importance of a functional FfG1 protein for development of perithecia in the mating strain that carries the MAT1-1 idiomorph.

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Analysis of growth behavior of the cAMP pathway mutants on different media compared to the WT.A) WT and cAMP pathway mutants (Δffg1, Δffg3, Δffac, Δffpka1, Δffpka2) were grown on solidified ICI medium with 6 mM sodium nitrate as sole nitrogen source. B) WT and cAMP pathway mutants on solidified complete medium (CM) and Czapek dox (CD, minimal medium). After 7 days radial growth rates of the respective mutants was determined. Experiments were done in triplicate. C) Radial growth of the respective mutants grown on CM or CD medium was measured [mm]. Mean values and standard deviations are shown in the diagram.
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pone-0058185-g006: Analysis of growth behavior of the cAMP pathway mutants on different media compared to the WT.A) WT and cAMP pathway mutants (Δffg1, Δffg3, Δffac, Δffpka1, Δffpka2) were grown on solidified ICI medium with 6 mM sodium nitrate as sole nitrogen source. B) WT and cAMP pathway mutants on solidified complete medium (CM) and Czapek dox (CD, minimal medium). After 7 days radial growth rates of the respective mutants was determined. Experiments were done in triplicate. C) Radial growth of the respective mutants grown on CM or CD medium was measured [mm]. Mean values and standard deviations are shown in the diagram.

Mentions: To study the involvement of the analyzed signaling components on fungal growth in F. fujikuroi, we grew the wild type and the different mutants for 7 days on both complete medium (CM) and minimal Czapek-Dox (CD) medium and determined the radial growth rates. When grown on CM, Δffg3, Δffac and Δffpka1 showed reduced growth rates, while no growth defects were observed for Δffg1 and Δffpka2 (fig 6A–B). The same tendency was observed when the indicated strains were grown on CD medium. Although Δffg1 grows in a wild type manner, Δffg3, Δffac and Δffpka1 mutants showed impaired growth. Surprisingly, while Δffpka2 showed normal growth on complete medium, radial growth was reduced about 25% on minimal medium (fig. 6A–B). When grown on synthetic ICI medium, Δffg1, Δffg3, and Δffac mutants secreted fusarubin into the medium, emphasizing our findings that the two Gα subunits and the adenylyl cyclase negatively affect fusarubin biosynthesis (fig. 6C). No significant differences were observed in liquid ICI medium regarding biomass accumulation of the single deletion mutants.


Signaling governed by G proteins and cAMP is crucial for growth, secondary metabolism and sexual development in Fusarium fujikuroi.

Studt L, Humpf HU, Tudzynski B - PLoS ONE (2013)

Analysis of growth behavior of the cAMP pathway mutants on different media compared to the WT.A) WT and cAMP pathway mutants (Δffg1, Δffg3, Δffac, Δffpka1, Δffpka2) were grown on solidified ICI medium with 6 mM sodium nitrate as sole nitrogen source. B) WT and cAMP pathway mutants on solidified complete medium (CM) and Czapek dox (CD, minimal medium). After 7 days radial growth rates of the respective mutants was determined. Experiments were done in triplicate. C) Radial growth of the respective mutants grown on CM or CD medium was measured [mm]. Mean values and standard deviations are shown in the diagram.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC3585259&req=5

pone-0058185-g006: Analysis of growth behavior of the cAMP pathway mutants on different media compared to the WT.A) WT and cAMP pathway mutants (Δffg1, Δffg3, Δffac, Δffpka1, Δffpka2) were grown on solidified ICI medium with 6 mM sodium nitrate as sole nitrogen source. B) WT and cAMP pathway mutants on solidified complete medium (CM) and Czapek dox (CD, minimal medium). After 7 days radial growth rates of the respective mutants was determined. Experiments were done in triplicate. C) Radial growth of the respective mutants grown on CM or CD medium was measured [mm]. Mean values and standard deviations are shown in the diagram.
Mentions: To study the involvement of the analyzed signaling components on fungal growth in F. fujikuroi, we grew the wild type and the different mutants for 7 days on both complete medium (CM) and minimal Czapek-Dox (CD) medium and determined the radial growth rates. When grown on CM, Δffg3, Δffac and Δffpka1 showed reduced growth rates, while no growth defects were observed for Δffg1 and Δffpka2 (fig 6A–B). The same tendency was observed when the indicated strains were grown on CD medium. Although Δffg1 grows in a wild type manner, Δffg3, Δffac and Δffpka1 mutants showed impaired growth. Surprisingly, while Δffpka2 showed normal growth on complete medium, radial growth was reduced about 25% on minimal medium (fig. 6A–B). When grown on synthetic ICI medium, Δffg1, Δffg3, and Δffac mutants secreted fusarubin into the medium, emphasizing our findings that the two Gα subunits and the adenylyl cyclase negatively affect fusarubin biosynthesis (fig. 6C). No significant differences were observed in liquid ICI medium regarding biomass accumulation of the single deletion mutants.

Bottom Line: Here we studied the impact of the heterotrimeric G protein and the cAMP-mediated signaling network, including the regulatory subunits of the cAMP-dependent protein kinase (PKA), to study their effect on colony morphology, sexual development and regulation of bikaverins, fusarubins and GAs.In contrast, bikaverin biosynthesis is significantly reduced in ffg1 and ffg3 deletion mutants and positively regulated by FfAC and FfPKA1, while GA biosynthesis depends on the active FfAC and FfPKA2 in an FfG1- and FfG3-independent manner.In addition, we provide evidence that G Protein-mediated/cAMP signaling is important for growth in F. fujikuroi because deletion of ffg3, ffac and ffpka1 resulted in impaired growth on minimal and rich media.

View Article: PubMed Central - PubMed

Affiliation: Institut für Lebensmittelchemie, Westfälische Wilhelms-Universität, Münster, Germany.

ABSTRACT
The plant-pathogenic fungus Fusarium fujikuroi is a notorious rice pathogen causing hyper-elongation of infected plants due to the production of gibberellic acids (GAs). In addition to GAs, F. fujikuroi produces a wide range of other secondary metabolites, such as fusarins, fusaric acid or the red polyketides bikaverins and fusarubins. The recent availability of the fungal genome sequence for this species has revealed the potential of many more putative secondary metabolite gene clusters whose products remain to be identified. However, the complex regulation of secondary metabolism is far from being understood. Here we studied the impact of the heterotrimeric G protein and the cAMP-mediated signaling network, including the regulatory subunits of the cAMP-dependent protein kinase (PKA), to study their effect on colony morphology, sexual development and regulation of bikaverins, fusarubins and GAs. We demonstrated that fusarubin biosynthesis is negatively regulated by at least two Gα subunits, FfG1 and FfG3, which both function as stimulators of the adenylyl cyclase FfAC. Surprisingly, the primary downstream target of the adenylyl cyclase, the PKA, is not involved in the regulation of fusarubins, suggesting that additional, yet unidentified, cAMP-binding protein(s) exist. In contrast, bikaverin biosynthesis is significantly reduced in ffg1 and ffg3 deletion mutants and positively regulated by FfAC and FfPKA1, while GA biosynthesis depends on the active FfAC and FfPKA2 in an FfG1- and FfG3-independent manner. In addition, we provide evidence that G Protein-mediated/cAMP signaling is important for growth in F. fujikuroi because deletion of ffg3, ffac and ffpka1 resulted in impaired growth on minimal and rich media. Finally, sexual crosses of ffg1 mutants showed the importance of a functional FfG1 protein for development of perithecia in the mating strain that carries the MAT1-1 idiomorph.

Show MeSH
Related in: MedlinePlus