Systems Biology of the Secondary Metabolism in Filamentous Fungi

Doctoral thesis, 2018

Filamentous fungi constitute a rich reservoir of pharmaceutically relevant bioactive small molecules. These compounds, commonly referred to as secondary metabolites, are widely used as antibiotics for the treatment of microbial infections, but also as other pharmaceuticals such as immunosuppressors, cholesterol lowering agents and anticancer drugs. Although fungal derived antibiotics have been known for almost a century, genome sequencing has revealed that the biosynthetic potential of fungi is not fully exhausted. The Penicillium genus consists of around 350 accepted species, and many of these are well-known producers of pharmaceuticals and industrially exploited for this. The genus as a whole, however, is grossly understudied at the genomic level. To assess the potential for secondary metabolite biosynthesis in the Penicillium genus, we sequenced the genomes of ten species that produce diverse arrays of secondary metabolites in culture. One of the sequenced isolates was described as a new species, and we mapped secondary metabolites detected in culture to the corresponding biosynthetic gene clusters. The ten sequenced genomes were analyzed together with published Penicillium genomes, altogether 24, and we developed a pipeline to group biosynthetic gene clusters and map them to known pathways. We found a large untapped potential for biosynthesis of secondary metabolites, encoded in the genomes of these species, that potentially could fill the drug discovery pipeline. Based on our predictions, we experimentally identified a novel compound from the antifungal class of antibiotics called yanuthones. Since heterologous expression of secondary metabolite pathways has proved troublesome, the ten genome-sequenced Penicillium species were evaluated as cell factories in controlled bioreactor fermentations. Compared to an industrially relevant strain, the ten Penicillium species showed growth characteristics that encourage further exploration of their industrial potential. Transcriptome analysis of six of the species enabled the identification of a metabolic network that is responsible for precursor formation of secondary metabolites. This network provides important insight into the further industrial development of Penicillium cell factories, and could be used in designing metabolic engineering strategies for optimization of secondary metabolite production. Altogether this thesis provides novel insights into genetic and metabolic aspects of fungal secondary metabolism. Our findings propose that industrial production of secondary metabolites can be effectively established on the basis of native producers. Penicillium species constitute a rich source of drug leads, and possess promising physiological characteristics to be established as industrial production platforms.