Indole-diterpene gene cluster from Aspergillus flavus.

Aflatrem is a potent tremorgenic mycotoxin produced by the soil fungus Aspergillus flavus and is a member of a large structurally diverse group of secondary metabolites known as indole-diterpenes. By using degenerate primers for conserved domains of fungal geranylgeranyl diphosphate synthases, we cloned two genes, atmG and ggsA (an apparent pseudogene), from A. flavus. Adjacent to atmG are two other genes, atmC and atmM. These three genes have 64 to 70% amino acid sequence similarity and conserved synteny with a cluster of orthologous genes, paxG, paxC, and paxM, from Penicillium paxilli which are required for indole-diterpene biosynthesis. atmG, atmC, and atmM are coordinately expressed, with transcript levels dramatically increasing at the onset of aflatrem biosynthesis. A genomic copy of atmM can complement a paxM deletion mutant of P. paxilli, demonstrating that atmM is a functional homolog of paxM. Thus, atmG, atmC, and atmM are necessary, but not sufficient, for aflatrem biosynthesis by A. flavus. This provides the first genetic evidence for the biosynthetic pathway of aflatrem in A. flavus.