A metabolomics-guided approach to discover Fusarium graminearum metabolites after removal of a repressive histone modification.

Many secondary metabolites are produced by biosynthetic gene clusters (BGCs) that are repressed during standard growth conditions, which complicates the discovery of novel bioactive compounds. In the genus Fusarium, many BGCs reside in chromatin enriched for trimethylated histone 3 lysine 27 (H3K27me3), a modification correlated with transcriptional gene silencing. Here we report on our progress in assigning metabolites to genes by using a strain lacking the H3K27 methyltransferase, Kmt6.

Destabilization of chromosome structure by histone H3 lysine 27 methylation.

Chromosome and genome stability are important for normal cell function as instability often correlates with disease and dysfunction of DNA repair mechanisms. Many organisms maintain supernumerary or accessory chromosomes that deviate from standard chromosomes. The pathogenic fungus Zymoseptoria tritici has as many as eight accessory chromosomes, which are highly unstable during meiosis and mitosis, transcriptionally repressed, show enrichment of repetitive elements, and enrichment with heterochromatic histone methylation marks, e.

Application of the Cre/lox System to Construct Auxotrophic Markers for Quantitative Genetic Analyses in Fusarium graminearum.

The bacteriophage P1 Cre/lox system has been utilized in diverse fungi for marker recycling and exchange, generation of targeted chromosome translocations, and targeted deletion of interstitial chromosome segments. Here we show the application of this tool in the wheat and maize pathogen, Fusarium graminearum. We explored three different ways to introduce Cre into strains with floxed genes, namely transformation with an episomal or integrative plasmid (pLC28), fusion of protoplasts of strains carrying floxed genes with strains expressing Cre by forcing heterokaryons, and crosses between strains with floxed genes and strains expressing Cre to isolate progeny in which the target genes had been deleted during the cross.

Structural reorganization of the fungal endoplasmic reticulum upon induction of mycotoxin biosynthesis.

Compartmentalization of metabolic pathways to particular organelles is a hallmark of eukaryotic cells. Knowledge of the development of organelles and attendant pathways under different metabolic states has been advanced by live cell imaging and organelle specific analysis. Nevertheless, relatively few studies have addressed the cellular localization of pathways for synthesis of fungal secondary metabolites, despite their importance as bioactive compounds with significance to medicine and agriculture.

Histone modifications rather than the novel regional centromeres of Zymoseptoria tritici distinguish core and accessory chromosomes.

Background: Supernumerary chromosomes have been found in many organisms. In fungi, these "accessory" or "dispensable" chromosomes are present at different frequencies in populations and are usually characterized by higher repetitive DNA content and lower gene density when compared to the core chromosomes. In the reference strain of the wheat pathogen, Zymoseptoria tritici, eight discrete accessory chromosomes have been found.

Chromatin analyses of Zymoseptoria tritici: Methods for chromatin immunoprecipitation followed by high-throughput sequencing (ChIP-seq).

The presence or absence of specific transcription factors, chromatin remodeling machineries, chromatin modification enzymes, post-translational histone modifications and histone variants all play crucial roles in the regulation of pathogenicity genes. Chromatin immunoprecipitation (ChIP) followed by high-throughput sequencing (ChIP-seq) provides an important tool to study genome-wide protein-DNA interactions to help understand gene regulation in the context of native chromatin. ChIP-seq is a convenient in vivo technique to identify, map and characterize occupancy of specific DNA fragments with proteins against which specific antibodies exist or which can be epitope-tagged in vivo.

Epigenetic control of effector gene expression in the plant pathogenic fungus Leptosphaeria maculans.

Plant pathogens secrete an arsenal of small secreted proteins (SSPs) acting as effectors that modulate host immunity to facilitate infection. SSP-encoding genes are often located in particular genomic environments and show waves of concerted expression at diverse stages of plant infection. To date, little is known about the regulation of their expression.

The Fusarium graminearum histone H3 K27 methyltransferase KMT6 regulates development and expression of secondary metabolite gene clusters.

The cereal pathogen Fusarium graminearum produces secondary metabolites toxic to humans and animals, yet coordinated transcriptional regulation of gene clusters remains largely a mystery. By chromatin immunoprecipitation and high-throughput DNA sequencing (ChIP-seq) we found that regions with secondary metabolite clusters are enriched for trimethylated histone H3 lysine 27 (H3K27me3), a histone modification associated with gene silencing. H3K27me3 was found predominantly in regions that lack synteny with other Fusarium species, generally subtelomeric regions.

Genetic manipulation of the Fusarium fujikuroi fusarin gene cluster yields insight into the complex regulation and fusarin biosynthetic pathway.

In this work, the biosynthesis and regulation of the polyketide synthase/nonribosomal peptide synthetase (PKS/NRPS)-derived mutagenic mycotoxin fusarin C was studied in the fungus Fusarium fujikuroi. The fusarin gene cluster consists of nine genes (fus1-fus9) that are coexpressed under high-nitrogen and acidic pH conditions. Chromatin immunoprecipitation revealed a correlation between high expression and enrichment of activating H3K9-acetylation marks under inducing conditions.

Deciphering the cryptic genome: genome-wide analyses of the rice pathogen Fusarium fujikuroi reveal complex regulation of secondary metabolism and novel metabolites.

The fungus Fusarium fujikuroi causes "bakanae" disease of rice due to its ability to produce gibberellins (GAs), but it is also known for producing harmful mycotoxins. However, the genetic capacity for the whole arsenal of natural compounds and their role in the fungus' interaction with rice remained unknown. Here, we present a high-quality genome sequence of F.

Library preparation and data analysis packages for rapid genome sequencing.

High-throughput sequencing (HTS) has quickly become a valuable tool for comparative genetics and genomics and is now regularly carried out in laboratories that are not connected to large sequencing centers. Here we describe an updated version of our protocol for constructing single- and paired-end Illumina sequencing libraries, beginning with purified genomic DNA. The present protocol can also be used for "multiplexing," i.

Centromeres of filamentous fungi.

How centromeres are assembled and maintained remains one of the fundamental questions in cell biology. Over the past 20 years, the idea of centromeres as precise genetic loci has been replaced by the realization that it is predominantly the protein complement that defines centromere localization and function. Thus, placement and maintenance of centromeres are excellent examples of epigenetic phenomena in the strict sense.