Project description:Histone modifications have been shown to be crucial for secondary metabolism in various filamentous fungi. Here we studied the influence of histone acetylation on secondary metabolite production in the phytopathogenic fungus Fusarium fujikuroi, a known producer of several secondary metabolites including pigments and mycotoxins. Deletion of the classical HDACs FfHdF1, FfHdF2 and FfHdF3 indicated that FfHdF1 and FfHdF2 are major regulators of secondary metabolism, whereas FfHdF3 is involved in developmental processes but dispensable for secondary metabolite production in F. fujikuroi. Microarray analysis with the major HDAC FfHdF2 revealed differential regulation of several secondary metabolite gene clusters, subsequently verified by a combination of chemical and biological approaches. These results indicate that HDACs are responsible for gene silencing but also gene activation. Chromatin immunoprecipitation assays with M-NM-^TffhdF2 revealed significant alterations regarding the acetylation state in the landscape of secondary metabolite gene clusters thereby providing insights into the regulatory mechanism. In addition, the class I HDAC FfHdF1 also has major impact on secondary metabolism in F. fujikuroi. Furthermore, deletion of both ffhdF1 and ffhdF2 resulted in de-repression of secondary metabolites under normally repressing conditions. Thus, manipulation of HDAC encoding genes might provide a powerful tool for the activation of cryptic secondary metabolites. Investigation of whole genome gene expression of the Fusarium fujikuroi wild type IMI58289, M-NM-^TffhdF2 mutant under nitrogen starvation and nitrogen sufficient conditions. In this study we hybridized in total 12 microarrays using total RNA recovered from a wild-type culture of F. fujikuroi IMI58289 and M-NM-^TffhdF2 mutant culture. All cultures were grown on a 6 mM Gln (10%) and a 60 mM Gln medium (100%). For each combination of culture and medium a biological replicate was created. Each chip measures the expression level of 14,397 genes from F. fujikuroi IMI58289 with eight 60-mer probes.
Project description:Histone modifications have been shown to be crucial for secondary metabolism in various filamentous fungi. Here we studied the influence of histone acetylation on secondary metabolite production in the phytopathogenic fungus Fusarium fujikuroi, a known producer of several secondary metabolites including pigments and mycotoxins. Deletion of the classical HDACs FfHdF1, FfHdF2 and FfHdF3 indicated that FfHdF1 and FfHdF2 are major regulators of secondary metabolism, whereas FfHdF3 is involved in developmental processes but dispensable for secondary metabolite production in F. fujikuroi. Microarray analysis with the major HDAC FfHdF2 revealed differential regulation of several secondary metabolite gene clusters, subsequently verified by a combination of chemical and biological approaches. These results indicate that HDACs are responsible for gene silencing but also gene activation. Chromatin immunoprecipitation assays with ΔffhdF2 revealed significant alterations regarding the acetylation state in the landscape of secondary metabolite gene clusters thereby providing insights into the regulatory mechanism. In addition, the class I HDAC FfHdF1 also has major impact on secondary metabolism in F. fujikuroi. Furthermore, deletion of both ffhdF1 and ffhdF2 resulted in de-repression of secondary metabolites under normally repressing conditions. Thus, manipulation of HDAC encoding genes might provide a powerful tool for the activation of cryptic secondary metabolites. Investigation of whole genome gene expression of the Fusarium fujikuroi wild type IMI58289, ΔffhdF2 mutant under nitrogen starvation and nitrogen sufficient conditions.
Project description:Investigation of whole genome gene expression level differences of Fusarium fujikuroi between wild-type and a Ffvel1 (velvet) deletion mutant in liquid medium with minimal nitrogen between 24 hr, 72 hr and 120 hr of growth using an array based on a F. verticillioides gene call set. Fusarium fujikuroi produces a number of secondary metabolites including gibberellins, bikaverin, fumonisin and fusarin C that are influenced by nitrogen availability and the velvet global regulatory complex. A twelve chip study using total RNA recovered from six cultures of wild-type Fusarium fujikuroi and six cultures of Ffvel1 F. fujikuroi deletion mutant. Each chip measures the expression level of over 13,000 putative genes with twelve 60-mer probes per sequence.
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.
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)
Project description:We performed genome-wide transcriptome analyses of the Fusarium fujikuroi wild type compared to the ∆nsd1 mutant Nsd1 is a GATA type transcription factor which has a major impact on growth, conidiation and secondary metabolism in F. fujikuroi
Project description:Investigation of whole genome gene expression level differences of Fusarium fujikuroi between wild-type and a Ffvel1 (velvet) deletion mutant in liquid medium with minimal nitrogen between 24 hr, 72 hr and 120 hr of growth using an array based on a F. verticillioides gene call set. Fusarium fujikuroi produces a number of secondary metabolites including gibberellins, bikaverin, fumonisin and fusarin C that are influenced by nitrogen availability and the velvet global regulatory complex.
Project description:Post-translational modification of histones is a crucial mode of transcriptional regulation in eukaryotes. A well-described acetylation modifier of certain lysine residues is the Spt-Ada-Gcn5 acetyltransferase (SAGA) complex assembled around the histone acetyltransferase Gcn5 in Saccharomyces cerevisiae. We identified and characterized the SAGA complex in the rice pathogen Fusarium fujikuroi, well-known for producing a large variety of secondary metabolites (SMs). By using a co-immunoprecipitation approach, almost all of the S. cerevisiae SAGA complex components have been identified, except for the ubiquitinating DUBm module and the chromodomain containing Chd1. Deletion of GCN5 led to impaired growth, loss of conidiation and alteration of SM biosynthesis. Furthermore, we show that in F. fujikuroi Gcn5 is essential for the acetylation of several histone 3 lysines, i.e. H3K4, H3K9, H3K18 and H3K27. A genome-wide microarray analysis revealed differential expression of about 30% of the genome with an enrichment of genes involved in primary and secondary metabolism, transport and histone modification. HPLC-based analysis of known SMs revealed significant alterations in the Δgcn5 mutant. While most SM genes were activated by Gcn5 activity, the biosynthesis of the pigment bikaverin was strongly increased upon GCN5 deletion underlining the diverse roles of the SAGA complex in F. fujikuroi. Investigation of whole genome gene expression of the Fusarium fujikuroi wild type IMI58289 and the Δgcn5 mutant under nitrogen starvation and nitrogen sufficient conditions.
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.