Peptides derived from Kex2-processed repeat proteins are widely distributed and highly diverse in the Fungi kingdom.

Background: Recently, a gene cluster responsible for biosynthesis of ustiloxin in Aspergillus flavus was identified as the first case of a ribosomally synthesized and post-translationally modified peptide (RiPP) synthetic pathway in Ascomycota. RiPPs are biosynthesized from precursor peptides, which are processed to produce the RiPP backbone (core peptides) for further modifications such as methylation and cyclization. Ustiloxin precursor peptide has two distinctive features: a signal peptide for translocation into the endoplasmic reticulum and highly repeated core sequences cleaved by Kex2 protease in the Golgi apparatus. On the basis of these characteristics, the ustiloxin-type RiPP precursor peptides or Kex2-processed repeat proteins (KEPs) in strains belonging to the Fungi kingdom were computationally surveyed, in order to investigate the distribution and putative functions of KEPs in fungal ecology.

Results: In total, 7878 KEPs were detected in 1345 of 1461 strains belonging to 8 phyla. The average number of KEPs per strain was 5.25 in Ascomycota and 5.30 in Basidiomycota, but only 1.35 in the class Saccharomycetes (Ascomycota) and 1.00 in the class Tremellomycetes (Basidiomycota). The KEPs were classified into 838 types and 2560 stand-alone ones, which had no homologs. Nearly 200 types were distributed in more than one genus, and 14 types in more than one phylum. These types included yeast α-mating factors and fungal pheromones. Genes for 22% KEPs were accompanied by genes for DUF3328-domain-containing proteins, which are indispensable for cyclization of the core peptides. DUF3328-domain-containing protein genes were located at an average distance of 3.09 genes from KEP genes. Genes for almost all (with three exceptions) KEPs annotated as yeast α-mating factors or fungal pheromones were not accompanied by DUF3328-domain-containing protein genes.

Conclusion: KEPs are widely distributed in the Fungi kingdom, but their repeated sequences are highly diverse. From these results and some examples, a hypothesis was raised that KEPs initially evolved as unmodified linear peptides (e.g., mating factors), and then those that adopted a modified cyclic form emerged (e.g., toxins) to utilize their strong bioactivity against predators and competitive microorganisms.