Light-dependent functions of the Fusarium fujikuroi CryD DASH cryptochrome in development and secondary metabolism.
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ABSTRACT:
SUBMITTER: Castrillo M
PROVIDER: S-EPMC3623198 | biostudies-literature | 2013 Apr
REPOSITORIES: biostudies-literature
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Project description:The heterothallic ascomycete Fusarium fujikuroi is a notorious rice pathogen causing super-elongation of plants due to the production of terpene-derived gibberellic acids (GAs) that function as natural plant hormones. Additionally, F. fujikuroi is able to produce a variety of polyketide- and non-ribosomal peptide-derived metabolites such as bikaverins, fusarubins and fusarins as well as metabolites from yet unidentified biosynthetic pathways, e.g. moniliformin. The key enzymes needed for their production belong to the family of polyketide synthases (PKSs) and non-ribosomal peptide synthases (NRPSs) that are generally known to be post-translationally modified by a Sfp-type 4'phosphopantetheinyl transferase (PPTase). In this study we provide evidence that the F. fujikuroi Sfp-type PPTase FfPpt1 is essentially involved in lysine biosynthesis and production of bikaverins, fusarubins and fusarins, but not moniliformin as shown by analytical methods. Concomitantly, targeted Ffppt1 deletion mutants reveal an enhancement of terpene-derived metabolites like GAs and volatile substances such as ?-acorenol. Pathogenicity assays on rice roots using fluorescent labeled wild-type and Ffppt1 mutant strains indicate that lysine biosynthesis and iron acquisition but not PKS and NRPS metabolism is essential for establishment of primary infections of F. fujikuroi. Additionally, FfPpt1 is involved in conidiation and sexual mating recognition possibly by activating PKS- and/or NRPS-derived metabolites that could act as diffusible signals. Furthermore, the effect on iron acquisition of Ffppt1 mutants led us to identify a previously uncharacterized putative third reductive iron uptake system (FfFtr3/FfFet3) that is closely related to the FtrA/FetC system of A. fumigatus. Functional characterization provides evidence that both proteins are involved in iron acquisition and are liable to transcriptional repression of the homolog of the Aspergillus GATA-type transcription factor SreA under iron-replete conditions. Targeted deletion of the first Fusarium homolog of this GATA-type transcription factor-encoding gene, Ffsre1, strongly indicates its involvement in regulation of iron homeostasis and oxidative stress resistance.
| S-EPMC3360786 | biostudies-literature
Project description:The ascomycete fungus Fusarium fujikuroi (Gibberella fujikuroi MP-C) produces secondary metabolites of biotechnological interest, such as gibberellins, bikaverin, and carotenoids. Production of these metabolites is regulated by nitrogen availability and, in a specific manner, by other environmental signals, such as light in the case of the carotenoid pathway. A complex regulatory network controlling these processes is recently emerging from the alterations of metabolite production found through the mutation of different regulatory genes. Here we show the effect of the targeted mutation of the acyA gene of F. fujikuroi, coding for adenylyl cyclase. Mutants lacking the catalytic domain of the AcyA protein showed different phenotypic alterations, including reduced growth, enhanced production of unidentified red pigments, reduced production of gibberellins and partially derepressed carotenoid biosynthesis in the dark. The phenotype differs in some aspects from that of similar mutants of the close relatives F. proliferatum and F. verticillioides: contrary to what was observed in these species, ΔacyA mutants of F. fujikuroi showed enhanced sensitivity to oxidative stress (H(2)O(2)), but no change in heavy metal resistance or in the ability to colonize tomato tissue, indicating a high versatility in the regulatory roles played by cAMP in this fungal group.
| S-EPMC3266886 | biostudies-literature
Project description:In Fusarium fujikuroi, the biosynthesis of gibberellins (GAs) and bikaverin is under control of AreA-mediated nitrogen metabolite repression. Thus far, the signaling components acting upstream of AreA and regulating its nuclear translocation are unknown. In Saccharomyces cerevisiae, the target of rapamycin (TOR) proteins, Tor1p and Tor2p, are key players of nutrient-mediated signal transduction to control cell growth. In filamentous fungi, probably only one TOR kinase-encoding gene exists. However, nothing is known about its function. Therefore, we investigated the role of TOR in the GA-producing fungus F. fujikuroi in order to determine whether TOR plays a role in nitrogen regulation, especially in the regulation of GA and bikaverin biosynthesis. We cloned and characterized the F. fujikuroi tor gene. However, we were not able to create knockout mutants, suggesting that TOR is essential for viability. Inhibition of TOR by rapamycin affected the expression of AreA-controlled secondary metabolite genes for GA and bikaverin biosynthesis, as well as genes involved in transcriptional and translational regulation, ribosome biogenesis, and autophagy. Deletion of fpr1 encoding the FKBP12-homologue confirmed that the effects of rapamycin are due to the specific inhibition of TOR. Interestingly, the expression of most of the TOR target genes has been previously shown to be also affected in the glutamine synthetase mutant, although in the opposite way. We demonstrate here for the first time in a filamentous fungus that the TOR kinase is involved in nitrogen regulation of secondary metabolism and that rapamycin affects also the expression of genes involved in translation control, ribosome biogenesis, carbon metabolism, and autophagy.
| S-EPMC1595341 | biostudies-literature
Project description: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.
| S-EPMC3585259 | biostudies-literature
Project description:Histone modifications are crucial for the regulation of secondary metabolism in various filamentous fungi. Here we studied the involvement of histone deacetylases (HDACs) in secondary metabolism in the phytopathogenic fungus Fusarium fujikuroi, a known producer of several secondary metabolites, including phytohormones, pigments, and mycotoxins. Deletion of three Zn(2+)-dependent HDAC-encoding genes, ffhda1, ffhda2, and ffhda4, indicated that FfHda1 and FfHda2 regulate secondary metabolism, whereas FfHda4 is involved in developmental processes but is dispensable for secondary-metabolite production in F. fujikuroi. Single deletions of ffhda1 and ffhda2 resulted not only in an increase or decrease but also in derepression of metabolite biosynthesis under normally repressing conditions. Moreover, double deletion of both the ffhda1 and ffhda2 genes showed additive but also distinct phenotypes with regard to secondary-metabolite biosynthesis, and both genes are required for gibberellic acid (GA)-induced bakanae disease on the preferred host plant rice, as ?ffhda1 ?ffhda2 mutants resemble the uninfected control plant. Microarray analysis with a ?ffhda1 mutant that has lost the major HDAC revealed differential expression of secondary-metabolite gene clusters, which was subsequently verified by a combination of chemical and biological approaches. These results indicate that HDACs are involved not only in gene silencing but also in the activation of some genes. Chromatin immunoprecipitation with the ?ffhda1 mutant revealed significant alterations in the acetylation state of secondary-metabolite gene clusters compared to the wild type, thereby providing insights into the regulatory mechanism at the chromatin level. Altogether, manipulation of HDAC-encoding genes constitutes a powerful tool to control secondary metabolism in filamentous fungi.
| S-EPMC3837819 | biostudies-literature
Project description:GATA-type transcription factors (TFs) such as the nitrogen regulators AreA and AreB, or the light-responsive TFs WC-1 and WC-2, play global roles in fungal growth and development. The conserved GATA TF NsdD is known as an activator of sexual development and key repressor of conidiation in Aspergillus nidulans, and as light-regulated repressor of macroconidia formation in Botrytis cinerea. In the present study, we functionally characterized the NsdD ortholog in Fusarium fujikuroi, named Csm1. Deletion of this gene resulted in elevated microconidia formation in the wild-type (WT) and restoration of conidiation in the non-sporulating velvet mutant Δvel1 demonstrating that Csm1 also plays a role as repressor of conidiation in F. fujikuroi. Furthermore, biosynthesis of the PKS-derived red pigments, bikaverin and fusarubins, is de-regulated under otherwise repressing conditions. Cross-species complementation of the Δcsm1 mutant with the B. cinerea ortholog LTF1 led to full restoration of WT-like growth, conidiation and pigment formation. In contrast, the F. fujikuroi CSM1 rescued only the defects in growth, the tolerance to H2O2 and virulence, but did not restore the light-dependent differentiation when expressed in the B. cinerea Δltf1 mutant. Microarray analysis comparing the expression profiles of the F. fujikuroi WT and the Δcsm1 mutant under different nitrogen conditions revealed a strong impact of this GATA TF on 19 of the 47 gene clusters in the genome of F. fujikuroi. One of the up-regulated silent gene clusters is the one containing the sesquiterpene cyclase-encoding key gene STC1. Heterologous expression of STC1 in Escherichia coli enabled us to identify the product as the volatile bioactive compound (-)-germacrene D.
| S-EPMC5483468 | biostudies-literature
Project description:In Fusarium fujikuroi, the production of gibberellins and bikaverin is repressed by nitrogen sources such as glutamine or ammonium. Sensing and uptake of ammonium by specific permeases play key roles in nitrogen metabolism. Here, we describe the cloning of three ammonium permease genes, mepA, mepB, and mepC, and their participation in ammonium uptake and signal transduction in F. fujikuroi. The expression of all three genes is strictly regulated by the nitrogen regulator AreA. Severe growth defects of DeltamepB mutants on low-ammonium medium and methylamine uptake studies suggest that MepB functions as the main ammonium permease in F. fujikuroi. In DeltamepB mutants, nitrogen-regulated genes such as the gibberellin and bikaverin biosynthetic genes are derepressed in spite of high extracellular ammonium concentrations. mepA mepB and mepC mepB double mutants show a similar phenotype as DeltamepB mutants. All three F. fujikuroi mep genes fully complemented the Saccharomyces cerevisiae mep1 mep2 mep3 triple mutant to restore growth on low-ammonium medium, whereas only MepA and MepC restored pseudohyphal growth in the mep2/mep2 mutant. Overexpression of mepC in the DeltamepB mutants partially suppressed the growth defect but did not prevent derepression of AreA-regulated genes. These studies provide evidence that MepB functions as a regulatory element in a nitrogen sensing system in F. fujikuroi yet does not provide the sensor activity of Mep2 in yeast, indicating differences in the mechanisms by which nitrogen is sensed in S. cerevisiae and F. fujikuroi.
| S-EPMC2238153 | biostudies-literature
Project description:The fungus Fusarium fujikuroi (Gibberella fujikuroi MP-C) produces metabolites of biotechnological interest, such as gibberellins, bikaverins, and carotenoids. Gibberellin and bikaverin productions are induced upon nitrogen exhaustion, while carotenoid accumulation is stimulated by light. We evaluated the effect of nitrogen availability on carotenogenesis in comparison with bikaverin and gibberellin production in the wild type and in carotenoid-overproducing mutants (carS). Nitrogen starvation increased carotenoid accumulation in all strains tested. In carS strains, gibberellin and bikaverin biosynthesis patterns differed from those of the wild type and paralleled the expression of key genes for both pathways, coding for geranylgeranyl pyrophosphate (GGPP) and kaurene synthases for the former and a polyketide synthase for the latter. These results suggest regulatory connections between carotenoid biosynthesis and nitrogen-controlled biosynthetic pathways in this fungus. Expression of gene ggs1, which encodes a second GGPP synthase, was also derepressed in the carS mutants, suggesting the participation of Ggs1 in carotenoid biosynthesis. The carS mutations did not affect genes for earlier steps of the terpenoid pathway, such as fppS or hmgR. Light induced carotenoid biosynthesis in the wild type and carRA and carB levels in the wild-type and carS strains irrespective of nitrogen availability.
| S-EPMC2620702 | biostudies-literature
Project description:Investigation of whole genome gene expression of the Fusarium fujikuroi wild type IMI58289 under gibberellin-inducing and -repressing conditions. Fusarium fujikuroi is a biotechnologically important fungus due to its almost unique ability to produce gibberellic acids (GAs), a family of phytohormones. The fungus was already described about 100 years ago as the causative agent of Bakanae (foolish seedling) disease of rice. Beside GAs, the fungus is known to produce some pigments and mycotoxins, but for only eight products the biosynthetic genes are known. Here we present a high-quality genome sequence of the first member of the Gibberella fujikuroi species complex (GFC), that allowed de novo genome assembly with 12 scaffolds corresponding to the 12 chromosomes. In this work, we focused on identification of all potential secondary metabolism-related gene clusters and their regulation in response to nitrogen availability by transcriptome, proteome, HPLC-FLPC and ChIP-seq analyses. We show that most of the cluster genes are regulated in a nitrogen-dependent manner, and that expression profiles fit to proteome and ChIP-seq data for some but not all clusters. Comparison with genomes of all available Fusarium species, including the recently sequenced F. mangiferae and F. circinatum, showed only a small number of common gene clusters and provides new insights into the divergence of secondary metabolism in the genus Fusarium. Phylogenetic analyses suggest that some gene clusters were acquired by horizontal gene transfer, while others were present in ancient Fusarim species and have evolved differently by gene duplications and losses. One PKS and one NRPS gene cluster are unique for F. fujikuroi. Their products were identified by combining overexpression of cluster genes with HPLC-FLPC -based product analyses. In planta, expression studies suggest a specific role of the PKS19 product in rice infection. Our results indicate that comparative genomics together with the used genome-wide experimental approaches is a powerful tool to uncover new secondary metabolites and to understand their regulation on the transcript, protein and epigenetic levels. In this study, we hybridized in total 15 microarrays using total RNA recovered from wild-type cultures of F. fujikuroi IMI58289. Two cultures were grown on a 6 mM Gln medium. Additionally, two technical replicates were created. Four cultures were grown on a 60 mM Gln medium. Again, two technical replicates were created. On a 6 mM NO3 medium, three cultures were grown, and two cultures on a 120 mM NO3 medium, with no technical replicates. Each chip measures the expression level of 14,397 genes from F. fujikuroi IMI58289 with eight 60-mer probes.
2013-07-01 | E-GEOD-43745 | biostudies-arrayexpress
Project description:Fusarium fujikuroi is a biotechnologically important fungus due to its almost unique ability to produce gibberellic acids (GAs), a family of phytohormones. The fungus was described about 100 years ago as the causative agent of Bakanae (M-bM-^@M-^\foolish seedlingM-bM-^@M-^]) disease of rice. Apart from GAs, the fungus is known to produce pigments and mycotoxins, but the biosynthetic genes are known for only eight products. Here we present a high-quality genome sequence of the first member of the Gibberella fujikuroi species complex (GFC) that allowed de novo genome assembly with 12 scaffolds corresponding to the 12 chromosomes. In this work we focused on identification of all potential secondary metabolism-related gene clusters and their regulation in response to nitrogen availability by transcriptome, proteome, HPLC-FTMS and ChIP-seq analyses. We show that most of the cluster genes are regulated in a nitrogen-dependent manner, and that expression profiles fit to proteome and ChIP-seq data for some but not all clusters. Comparison with genomes of all available Fusarium species, including the recently sequenced F. mangiferae and F. circinatum, showed only a small number of common gene clusters and provides new insights into the divergence of secondary metabolism in the genus Fusarium. Phylogenetic analyses suggest that some gene clusters were acquired by horizontal gene transfer, while others were present in ancient Fusarim species and have evolved differently by gene duplications and losses. One polyketide synthase (PKS) and one non-ribosomal peptide synthetase (NRPS) gene cluster are unique for F. fujikuroi. Their products were identified by combining overexpression of cluster genes with HPLC-FTMS-based analyses. In planta expression studies suggest a specific role of the PKS19 product in rice infection. Our results indicate that comparative genomics together with the used genome-wide experimental approaches is a powerful tool to uncover new secondary metabolites and to understand their regulation at the transcriptional, translational and epigenetic levels. Examination of 3 different histone modifications, with 2 growth conditions for one of the modifications (Total of 4 samples)
2013-10-16 | E-GEOD-46033 | biostudies-arrayexpress