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Summary: Recent studies have identified key genes that control the symbiotic interaction between Epichloë festucae and Lolium perenne. Here we report on the identification of specific E. festucae genes that control host infection. Deletion of setB, which encodes a homologue of the H3K36 histone methyltransferase Set2/KMT3, reduced histone H3K36 trimethylation and led to severe defects in colony growth and hyphal development. The E. festucae ΔclrD mutant, which lacks the gene encoding the homologue of the H3K9 methyltransferase KMT1, displays similar developmental defects. Both mutants are completely defective in their ability to infect L. perenne. Alleles that complement the culture and plant phenotypes of both mutants also complement the histone methylation defects. Co‐inoculation of either ΔsetB or ΔclrD with the wild‐type strain enables these mutants to colonize the host. However, successful colonization by the mutants resulted in death or stunting of the host plant. Transcriptome analysis at the early infection stage identified four fungal candidate genes, three of which encode small‐secreted proteins, that are differentially regulated in these mutants compared to wild type. Deletion of crbA, which encodes a putative carbohydrate binding protein, resulted in significantly reduced host infection rates by E. festucae. [ABSTRACT FROM AUTHOR]

Summary: Epichloë festucae is an endophytic fungus that forms a symbiotic association with Lolium perenne. Here we analysed how the metabolome of the ryegrass apoplast changed upon infection of this host with sexual and asexual isolates of E. festucae.A metabolite fingerprinting approach was used to analyse the metabolite composition of apoplastic wash fluid from uninfected and infected L. perenne. Metabolites enriched or depleted in one or both of these treatments were identified using a set of interactive tools. A genetic approach in combination with tandem MS was used to identify a novel product of a secondary metabolite gene cluster.Metabolites likely to be present in the apoplast were identified using MarVis in combination with the BioCyc and KEGG databases, and an in‐house Epichloë metabolite database. We were able to identify the known endophyte‐specific metabolites, peramine and epichloëcyclins, as well as a large number of unknown markers.To determine whether these methods can be applied to the identification of novel Epichloë‐derived metabolites, we deleted a gene encoding a NRPS (lgsA) that is highly expressed in planta. Comparative MS analysis of apoplastic wash fluid from wild‐type‐ vs mutant‐infected plants identified a novel Leu/Ile glycoside metabolite present in the former. [ABSTRACT FROM AUTHOR]

Structural features of genomes, including the three-dimensional arrangement of DNA in the nucleus, are increasingly seen as key contributors to the regulation of gene expression. However, studies on how genome structure and nuclear organisation influence transcription have so far been limited to a handful of model species. This narrow focus limits our ability to draw general conclusions about the ways in which three-dimensional structures are encoded, and to integrate information from three-dimensional data to address a broader gamut of biological questions. Here, we generate a complete and gapless genome sequence for the filamentous fungus, Epichloë festucae. We use Hi-C data to examine the three-dimensional organisation of the genome, and RNA-seq data to investigate how Epichloë genome structure contributes to the suite of transcriptional changes needed to maintain symbiotic relationships with the grass host. Our results reveal a genome in which very repeat-rich blocks of DNA with discrete boundaries are interspersed by gene-rich sequences that are almost repeat-free. In contrast to other species reported to date, the three-dimensional structure of the genome is anchored by these repeat blocks, which act to isolate transcription in neighbouring gene-rich regions. Genes that are differentially expressed in planta are enriched near the boundaries of these repeat-rich blocks, suggesting that their three-dimensional orientation partly encodes and regulates the symbiotic relationship formed by this organism. [ABSTRACT FROM AUTHOR]

Nodulisporic acids comprise a group of valuable indole diterpenes that exhibit potent insecticidal activities. We report the identification of a gene cluster in the genome of the filamentous fungus Hypoxylon pulicicidum (Nodulisporium sp.) that contains genes responsible for the biosynthesis of nodulisporic acids. Using Pénicillium paxilli as a heterologous host, and through pathway reconstitution experiments, we identified the function of four genes involved in the biosynthesis of the nodulisporic acid core compound, nodulisporic acid F (NAF). Two of these genes (nodM and nodW) are especially significant as they encode enzymes with previously unreported functionality: nodM encodes a 3-geranylgeranylindole epoxidase capable of catalyzing only a single epoxidation step to prime formation of the distinctive ring structure of nodulisporic acids, and nodW encodes the first reported gene product capable of introducing a carboxylic acid moiety to an indole diterpene core structure that acts as a reactive handle for further modification. Here, we present the enzymatic basis for the biosynthetic branch point that gives rise to nodulisporic acids. [ABSTRACT FROM AUTHOR]

Endophytes of the genusEpichloë(Clavicipitaceae, Ascomycota) frequently occur within cool-season grasses and form interactions with their hosts that range from mutualistic to antagonistic. ManyEpichloëspecies have arisen via interspecific hybridization, resulting in species with two or three subgenomes that retain all or nearly all of their original parental genomes, a process termed allopolyploidization. Here, we characterizeEpichloë hybrida, sp. nov., a mutualistic species that has increasingly become a model system for investigating allopolyploidy in fungi. TheEpichloëspecies so far identified as the closest known relatives of the two progenitors ofE. hybridaareE. festucaevar.loliiandE. typhina. We confirm that the nuclear genome ofE. hybridacontains two homeologs of most protein-coding genes fromE. festucaeandE. typhina, with genome-wide gene expression analysis indicating a slight bias in overall gene expression from theE. typhinasubgenome. Mitochondrial DNA is detectable only fromE. festucae, whereas ribosomal DNA is detectable only fromE. typhina. Inheriting ribosomal DNA from just one parent might be expected to preferentially favor interactions with ribosomal proteins from the same parent, but we find that ribosomal protein genes from both parental subgenomes are nearly all expressed equally inE. hybrida. Finally, we provide a comprehensive set of resources for this model system that are intended to facilitate further study of fungal hybridization by other researchers. [ABSTRACT FROM AUTHOR]