Phylogenetic Investigation of Isocyanide Synthases in the Fungal Kingdom, with a Focus on Lichenized Fungi

Fungi are prolific producers of secondary metabolites with promising pharmaceutical and industrial applications. The most common classes of secondary metabolites produced by fungi include polyketide synthases, non-ribosomal peptide synthases, terpenes, and other peptides. Recently, genome mining and bioinformatic pipelines for gene characterization, comparison, and clustering have become increasingly popular in identifying genes that encode useful novel natural products and predicting their bioactivity. These approaches have unraveled another class of biosynthetic gene clusters (BGCs), isocyanide synthases (ICSs)—involved in metal homeostasis, microbial competition, and pathogenesis—which are common in saprophytic and pathogenic fungi. However, their presence and diversity in symbiotic, lichen-forming fungi and the evolutionary relationships to those present in non-lichenized fungi have remained unexplored. To explore the presence and diversity of ICSs in lichen-forming fungi (LFF), we extracted and aligned ICS sequences from 232 genomes (121 from LFFs and 111 from non-LFFs). We also included representative ICSs from non-lichenized fungi, constructing a maximum-likelihood tree with 1000 bootstrap replicates. Our analysis revealed that: 1) ICSs are unevenly distributed across the fungal kingdom and are absent in Basidiomycota and in early-diverging lineages of Pezizomycotina (Pezizomycetes, Orbiliomycetes); 2) about 45% of lichens have at least one copy of an ICS gene; 3) ICSs form four major clades in LFFs, of which three (LFF-ICS1, LFF-ICS2, and LFF-ICS3) are putatively novel and exclusive to lichens; and 4) lichens possess putative copper-responsive metabolite (crm) gene clusters, which are known to be induced under copper-starvation conditions.