Identification and engineering of the cytochalasin gene cluster from Aspergillus clavatus NRRL 1

Cytochalasins are a group of fungal secondary metabolites with diverse structures and bioactivities, including cytochalasin E produced by Aspergillus clavatus, which is a potent anti-angiogenic agent. Here, we report the identification and characterization of the cytochalasin gene cluster from A. clavatus NRRL 1. As a producer of cytochalasin E and K, the genome of A. clavatus was analyzed and the ∼30 kb ccs gene cluster was identified based on the presence of a polyketide synthase–nonribosomal peptide synthetases (PKS–NRPS) and a putative Baeyer–Villiger monooxygenase (BVMO). Deletion of the central PKS–NRPS gene, ccsA, abolished the production of cytochalasin E and K, confirming the association between the natural products and the gene cluster. Based on bioinformatic analysis, a putative biosynthetic pathway is proposed. Furthermore, overexpression of the pathway specific regulator ccsR elevated the titer of cytochalasin E from 25 mg/L to 175 mg/L. Our results not only shed light on the biosynthesis of cytochalasins, but also provided genetic tools for increasing and engineering the production. ► We identified the gene cluster for cytochalasin E/K in Aspergillus clavatus genome. ► Involvement of a PKS–NRPS gene ( ccsA) was confirmed by targeted gene disruption. ► A biosynthetic route for cytochalasin E/K was proposed based on gene cluster analyses. ► Overexpression of the ccsR regulator gene improved cytochalasin production. ► Genome mining revealed ccs-like gene clusters among other sequenced fungal genomes.