Regulation and Functional Analysis of Bioprotective Metabolite Genes from the Grass Symbiont Epichloe festucae

Epichloe endophytes (Neotyphodium/Epichloe species) are biotrophic fungi that systemically colonise the intercellular spaces of leaves of grasses to form mutualistic symbiotic associations. The production of secondary metabolites by these fungi confers bioprotective benefits to the grass. However, in pastoral ecosystems some of these metabolites are toxic to grazing mammals. We have cloned and functionally analysed genes for the synthesis of three classes of the bioprotective molecules, peramine, lolitrem B and ergovaline. A single gene, perA, encoding a non-ribosomal peptide synthetase is required for peramine biosynthesis. Complex gene clusters with 10 and 11 genes, respectively, are required for lolitrem B (ltm genes) and ergovaline (eas genes) biosynthesis. The biochemical function of these genes is being elucidated by a systematic deletion analysis combined with chemical analysis of intermediates that accumulate in planta. These experiments allow us to propose biosynthetic schemes for the synthesis of the metabolites. Symbiota of these ‘knock-out’ mutants are being used to examine biological function of the metabolites. Spatial and temporal patterns of gene expression are being examined in planta using promoter fusions with the GUS reporter gene. An overview of recent progress to date combined with some original data are presented.