A global atlas of fungal biosynthetic gene clusters reveals the diversification of diketopiperazine biosynthesis

Background Fungi represent one of the largest and most promising reservoirs of structurally diverse natural products. However, the global biosynthetic potential of fungi significantly expanded but remains underexplored. Results Here, we presented the most comprehensive fungal biosynthetic gene cluster (BGC) atlas that comprised 303,983 BGCs predicted from 13,125 fungal genomes, revealing many less-explored taxa encoding large biosynthetic diversity. The fungal BGCs were organized into 43,984 gene cluster families (GCFs), with 99.6% remaining uncharacterized and 91.7% being genus-specific. Gene-centric analysis has revealed the presence of 359 cyclodipeptide synthases of three distinct subcategories and 9,482 nonribosomal peptide synthetases (NRPSs) responsible for diketopiperazine biosynthesis in the fungal BGC atlas. Interestingly, 304 type one CDPSs with high homology to bacterial CDPSs were discovered in fungi for the first time, exclusively found in Fusarium . A mass spectrometry-guided approach resulted in the isolation of eighteen indole diketopiperazine alkaloids, including three novel ones, from an Aspergillus strain. Bioinformatics analysis confirmed that these compounds are synthesized by an NRPS protein and several post-modification enzymes. Conclusions The study presents the most comprehensive fungal BGC atlas and highlights the diversification of diketopiperazine biosynthesis in fungi, laying a crucial foundation for the exploration of specific types of natural products from fungi.