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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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Gibberellic acid (GA) content of cAMP pathway mutants compared to WT.The WT and cAMP pathway mutants (Δffg1, Δffg3, Δffac, Δffpka1, Δffpka2) were grown for 7 days in synthetic ICI medium under GA biosynthesis-favoring conditions (6 mM glutamine). GA3 and GA4/7 were extracted from 20 ml liquid culture (for details see material and methods). For comparison, GA content produced by the WT was set to 100%. Experiments were performed in triplicate.
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pone-0058185-g005: Gibberellic acid (GA) content of cAMP pathway mutants compared to WT.The WT and cAMP pathway mutants (Δffg1, Δffg3, Δffac, Δffpka1, Δffpka2) were grown for 7 days in synthetic ICI medium under GA biosynthesis-favoring conditions (6 mM glutamine). GA3 and GA4/7 were extracted from 20 ml liquid culture (for details see material and methods). For comparison, GA content produced by the WT was set to 100%. Experiments were performed in triplicate.

Mentions: GA biosynthesis is the ‘trade mark’ of F. fujikuroi, as only a few fungal species are able to produce these isoprenoid phytohormones. Therefore, we wanted to know if biosynthesis of GAs is also regulated via the G protein/cAMP pathway. We cultivated the wild type and mutant strains under GA biosynthesis-inducing conditions (6 mM glutamine) for 7 days. The quantity of produced GAs was analyzed by HPLC-UV. Interestingly, neither of the two Gα subunits, FfG1 and FfG3, was involved in regulation of GA biosynthesis. However, deletions of ffac and ffpka2 resulted in a significant reduction of GA biosynthesis of about 70% (fig. 5), while deletion of ffpka1 had no effect. These data indicate that FfAC can be activated by additional components other than the two Gα subunits, and that the two catalytic subunits of the PKA have distinct functions.


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)

Gibberellic acid (GA) content of cAMP pathway mutants compared to WT.The WT and cAMP pathway mutants (Δffg1, Δffg3, Δffac, Δffpka1, Δffpka2) were grown for 7 days in synthetic ICI medium under GA biosynthesis-favoring conditions (6 mM glutamine). GA3 and GA4/7 were extracted from 20 ml liquid culture (for details see material and methods). For comparison, GA content produced by the WT was set to 100%. Experiments were performed in triplicate.
© Copyright Policy
Related In: Results  -  Collection

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

pone-0058185-g005: Gibberellic acid (GA) content of cAMP pathway mutants compared to WT.The WT and cAMP pathway mutants (Δffg1, Δffg3, Δffac, Δffpka1, Δffpka2) were grown for 7 days in synthetic ICI medium under GA biosynthesis-favoring conditions (6 mM glutamine). GA3 and GA4/7 were extracted from 20 ml liquid culture (for details see material and methods). For comparison, GA content produced by the WT was set to 100%. Experiments were performed in triplicate.
Mentions: GA biosynthesis is the ‘trade mark’ of F. fujikuroi, as only a few fungal species are able to produce these isoprenoid phytohormones. Therefore, we wanted to know if biosynthesis of GAs is also regulated via the G protein/cAMP pathway. We cultivated the wild type and mutant strains under GA biosynthesis-inducing conditions (6 mM glutamine) for 7 days. The quantity of produced GAs was analyzed by HPLC-UV. Interestingly, neither of the two Gα subunits, FfG1 and FfG3, was involved in regulation of GA biosynthesis. However, deletions of ffac and ffpka2 resulted in a significant reduction of GA biosynthesis of about 70% (fig. 5), while deletion of ffpka1 had no effect. These data indicate that FfAC can be activated by additional components other than the two Gα subunits, and that the two catalytic subunits of the PKA have distinct functions.

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