Genome analysis and elucidation of the biosynthetic pathway for the cRAS inhibitor rasfonin in Cephalotrichum gorgonifer

Abstract

Abstract Background Fungi are important sources for bioactive compounds that find their applications in many important sectors like in the pharma-, food- or agricultural industries. In an environmental monitoring project for fungi involved in soil nitrogen cycling we also isolated Cephalotrichum gorgonifer (strain NG_p51). In the course of strain characterization work we found that this strain is able to naturally produce high amounts of rasfonin, a polyketide inducing autophagy, apoptosis, necroptosis in human cell lines and shows anti-tumor activity in RAS-dependent cancer cells. Results In order to elucidate the biosynthetic pathway of rasfonin, the strain was genome sequenced, annotated, submitted to transcriptome analysis and genetic transformation was established. Biosynthetic gene cluster (BGC) prediction revealed the existence of 22 BGCs of which the majority was not expressed under our experimental conditions. In silico prediction revealed two BGCs with a suite of enzymes possibly involved in rasfonin biosynthesis. Experimental verification by gene-knock out of the key enzyme genes showed that one of the predicted BGCs is indeed responsible for rasfonin biosynthesis. Conclusions The results of this study lay the ground for molecular biology focused research in Cephalotrichum gorgonifer. Furthermore, strain engineering and heterologous expression of the rasfonin BGC is now possible which allow both the construction of rasfonin high producing strains and biosynthesis of rasfonin derivates for diverse applications.