Novel Cyclic Peptides from Lethal Amanita Mushrooms through a Genome-Guided Approach.

Shengwen Zhou,Pengcheng Yuan,Zhu L Yang,Xincan Li,Hong Luo,Zuo H Chen,Xuan Li,Yunjiao Lüli,Guohong Li

doi:10.3390/jof7030204

Abstract

Most species in the genus Amanita are ectomycorrhizal fungi comprising both edible and poisonous mushrooms. Some species produce potent cyclic peptide toxins, such as α-amanitin, which places them among the deadliest organisms known to mankind. These toxins and related cyclic peptides are encoded by genes of the “MSDIN” family (named after the first five amino acid residues of the precursor peptides), and it is largely unknown to what extent these genes are expressed in the basidiocarps. In the present study, Amanita rimosa and Amanita exitialis were sequenced through the PacBio and Illumina techniques. Together with our two previously sequenced genomes, Amanita subjunquillea and Amanita pallidorosea, in total, 46 previously unknown MSDIN genes were discovered. The expression of over 80% of the MSDIN genes was demonstrated in A. subjunquillea. Through a combination of genomics and mass spectrometry, 12 MSDIN genes were shown to produce novel cyclic peptides. To further confirm the results, three of the cyclic peptides were chemically synthesized. The tandem mass spectrometry (MS/MS) spectra of the natural and the synthetic peptides shared a majority of the fragment ions, demonstrating an identical structure between each peptide pair. Collectively, the results suggested that the genome-guided approach is reliable for identifying novel cyclic peptides in Amanita species and that there is a large peptide reservoir in these mushrooms.

Highlights

The genus Amanita (Persoon 1797) is double-faced: on one hand, it harbors some of the best-known gourmet mushrooms such as Amanita caesarea, once favored by emperors of Rome [1]; on the other hand, it causes over 90% of deadly mushroom poisonings worldwide because most have little knowledge to distinguish the edible from the deadly [2,3,4]
It has been shown that amatoxins and phallotoxins in Amanita are synthesized on ribosomes, which represents the first ribosomal cyclic peptide discovered in the Fungi kingdom [15]
Two deadly Amanita species (A. rimosa and A. subjunquillea (As). exitialis) were sequenced, and, in total, 46 novel MSDIN genes discovered in four draft genomes

Summary

Introduction

The genus Amanita (Persoon 1797) is double-faced: on one hand, it harbors some of the best-known gourmet mushrooms such as Amanita caesarea, once favored by emperors of Rome [1]; on the other hand, it causes over 90% of deadly mushroom poisonings worldwide because most have little knowledge to distinguish the edible from the deadly [2,3,4]. It has been shown that amatoxins and phallotoxins in Amanita are synthesized on ribosomes, which represents the first ribosomal cyclic peptide discovered in the Fungi kingdom [15] They are encoded by the “MSDIN” gene family as precursor peptides of 34–37 amino acids. Newly sequenced genomes of lethal Amanita species show that the MSDIN genes by far outnumber the count of known cyclic peptides, indicating there could be a much larger potential for novel cyclic peptides in these mushrooms. It is mostly unknown whether these MSDINs are translated into cyclic peptides

Methods

Results

Discussion

Conclusion