title: Regulation of secondary metabolite production in the fungal tomato pathogen Cladosporium fulvum.
abstract: Cladosporium fulvum is a non-obligate biotrophic fungal tomato pathogen for which fifteen secondary metabolite (SM) gene clusters were previously identified in its genome. However, most of these SM biosynthetic pathways remain cryptic during growth in planta and in different in vitro conditions. The sole SM produced in vitro is the pigment cladofulvin. In this study, we attempted to activate cryptic pathways in order to identify new compounds produced by C. fulvum. For this purpose, we manipulated orthologues of the global regulators VeA, LaeA and HdaA known to regulate SM biosynthesis in other fungal species. In C. fulvum, deleting or over-expressing these regulators yielded no new detectable SMs. Yet, quantification of cladofulvin revealed that CfHdaA is an activator whilst CfVeA and CfLaeA seemed to act as repressors of cladofulvin production. In the wild type strain, cladofulvin biosynthesis was affected by the carbon source, with highest production under carbon limitation and traces only in presence of saccharose. Repression of cladofulvin production by saccharose was dependent on both CfVeA and CfLaeA. Deletion of CfVeA or CfLaeA caused production of sterile mycelia, whilst Δcfhdaa deletion mutants sporulated, suggesting that cladofulvin production is not linked to asexual reproduction. Profiling the transcription of these regulators showed that CfHdaA-mediated regulation of cladofulvin production is independent of both CfVeA and CfLaeA. Our data suggest CfLaeA directly affects cladofulvin production whilst the effect of CfVeA is indirect, suggesting a role for CfLaeA outside of the Velvet complex. In conclusion, our results showed that regulation of SM production in C. fulvum is different from other fungi and indicate that manipulation of global regulators is not a universal tool to discover new fungal natural products.
authors.affiliation: Laboratory of Phytopathology, Wageningen University, Droevendaalsesteeg 1, 6708PB Wageningen, The Netherlands.
authors.forename: Scott
authors.initials: S
authors.lastname: Griffiths
authors.affiliation: Laboratory of Phytopathology, Wageningen University, Droevendaalsesteeg 1, 6708PB Wageningen, The Netherlands.
authors.forename: Benedetta
authors.initials: B
authors.lastname: Saccomanno
authors.affiliation: Laboratory of Phytopathology, Wageningen University, Droevendaalsesteeg 1, 6708PB Wageningen, The Netherlands. Electronic address: pierre.dewit@wur.nl.
authors.forename: Pierre J G M
authors.initials: PJ
authors.lastname: de Wit
authors.affiliation: Laboratory of Phytopathology, Wageningen University, Droevendaalsesteeg 1, 6708PB Wageningen, The Netherlands; UMR 1345 IRHS-INRA, 42 rue Georges Morel, 49071 Beaucouzé Cedex, France. Electronic address: j.collemare@gmail.com.
authors.forename: Jérôme
authors.initials: J
authors.lastname: Collemare
mesh_terms: D016960:Agrobacterium tumefaciens / Q000235:genetics; D057465:Catabolite Repression; D002851:Chromatography, High Pressure Liquid; D002963:Cladosporium / Q000201:enzymology / Q000235:genetics / Q000378:metabolism*; D005656:Fungal Proteins / Q000235:genetics / Q000378:metabolism; D020869:Gene Expression Profiling; D005800:Genes, Fungal; D018551:Solanum lycopersicum / Q000382:microbiology*; D005810:Multigene Family; D010641:Phenotype; D060888:Real-Time Polymerase Chain Reaction; D064210:Secondary Metabolism; D017384:Sequence Deletion; D013395:Sucrose / Q000378:metabolism
chemical_list: D005656:Fungal Proteins; D013395:Sucrose
keywords: Cladofulvin; Cryptic biosynthetic pathways; Global regulator; HdaA; LaeA; VeA