Functional analysis of secondary metabolism gene clusters expressed in fungal appressoria

Species of the genus Colletotrichum cause devastating anthracnose or blight diseases on numerous crop plants worldwide. These pathogens use a hemibiotrophic infection strategy that involves the formation of specialized cell-types for initial penetration (appressoria), growth inside living cells (byotrophic hyphae) and tissue destruction (necrotrophic hyphae). The interaction between C. higginsianum and Arabidopsis provides an attractive model for studying the molecular basis of hemibiotrophy in which both partners can be geneticallymanipulated. The genome of C. higginsianum contains an unusually large number of genes encoding enzymes involved in secondary metabolism (SM) compared to other hemibiotrophs (e.g. Magnaporthe, Fusarium) and necrotrophs (e.g. Botrytis, Stagonospora). In particular,genes encoding terpene synthase, polyketide synthases and dimethyl-allyltransferases are all expanded in C. higginsianum. As in other fungi, the key enzyme genes are organized into clusters that also contain genes encoding accessory enzymes of the same biosynthetic pathway, efflux transporters and pathway-specific transcription factors. In C. higginsianum,47 such clusters were identified to date. A remarkable finding from RNA-Seq transcriptome profiling was that 17 SM clusters are specifically expressed only in planta by appressoria and biotrophic hyphae. Since each cluster potentially synthesizes one metabolite, this suggests appressoria and biotrophic hyphae deliver a cocktail of different metabolites to the first infected host cell. At this early stage host cells remain alive, raising the possibility that these molecules function similar to protein effectors in host manipulation, rather than as simple phytotoxins. Work is ongoing to decipher how the expression of these SM gene clusters is soprecisely regulated during the course of infection (e.g. transcription factors or chromatin remodelling), with a view to inducing the fungus to synthesize these compounds in large quantities in culture. The aim is to obtain sufficient material for identification and structural determination (mass spectrometry, NMR) and for functional assays to evaluate biological activities against plants, bacteria and fungi. O'Connell R et al., 2012. Nature Genetics 44, 1060