Establishment of the Inducible Tet-On System for the Activation of the Silent Trichosetin Gene Cluster in Fusarium fujikuroi

Toxins (Basel). 2017 Apr 5;9(4):126. doi: 10.3390/toxins9040126.


The PKS-NRPS-derived tetramic acid equisetin and its N-desmethyl derivative trichosetin exhibit remarkable biological activities against a variety of organisms, including plants and bacteria, e.g., Staphylococcus aureus. The equisetin biosynthetic gene cluster was first described in Fusarium heterosporum, a species distantly related to the notorious rice pathogen Fusarium fujikuroi. Here we present the activation and characterization of a homologous, but silent, gene cluster in F. fujikuroi. Bioinformatic analysis revealed that this cluster does not contain the equisetin N-methyltransferase gene eqxD and consequently, trichosetin was isolated as final product. The adaption of the inducible, tetracycline-dependent Tet-on promoter system from Aspergillus niger achieved a controlled overproduction of this toxic metabolite and a functional characterization of each cluster gene in F. fujikuroi. Overexpression of one of the two cluster-specific transcription factor (TF) genes, TF22, led to an activation of the three biosynthetic cluster genes, including the PKS-NRPS key gene. In contrast, overexpression of TF23, encoding a second Zn(II)₂Cys₆ TF, did not activate adjacent cluster genes. Instead, TF23 was induced by the final product trichosetin and was required for expression of the transporter-encoding gene MFS-T. TF23 and MFS-T likely act in consort and contribute to detoxification of trichosetin and therefore, self-protection of the producing fungus.

Keywords: Fungi; Fusarium fujikuroi; PKS-NRPS; biosynthesis; gene regulation; secondary metabolism.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Aspergillus / genetics
  • Cell Survival / drug effects
  • Fungal Proteins / genetics
  • Fusarium / genetics*
  • Fusarium / metabolism
  • Gene Expression Regulation, Fungal*
  • Hep G2 Cells
  • Humans
  • Multigene Family
  • Oryza / microbiology
  • Plant Diseases / microbiology
  • Promoter Regions, Genetic
  • Pyrrolidinones* / isolation & purification
  • Pyrrolidinones* / metabolism
  • Pyrrolidinones* / toxicity
  • Tetrahydronaphthalenes / toxicity
  • Transcription Factors / genetics


  • Fungal Proteins
  • Pyrrolidinones
  • Tetrahydronaphthalenes
  • Transcription Factors
  • trichosetin
  • equisetin