Search

Fungi in the basidiomycetous genus Amanita owe their high mammalian toxicity to the bicyclic octapeptide amatoxins such as α-amanitin. Amatoxins and the related phallotoxins (such as the heptapeptide phalloidin) are encoded by members of the "MSDIN" gene family and are synthesized on ribosomes as short (34- to 35-amino-acid) proproteins. Antiamanitin antibodies and confocal microscopy were used to determine the cellular and subcellular localizations of amanitin accumulation in basidiocarps (mushrooms) of the Eastern North American destroying angel (Amanita bisporigera). Consistent with previous studies, amanitin is present throughout the basidiocarp (stipe, pileus, lamellae, trama, and universal veil), but it is present in only a subset of cells within these tissues. Restriction of amanitin to certain cells is especially marked in the hymenium. Several lines of evidence implicate a specific prolyl oligopeptidase, A. bisporigera POPB (AbPOPB), in the initial processing of the amanitin and phallotoxin proproteins. The gene for AbPOPB is restricted taxonomically to the amatoxin-producing species of Amanita and is clustered in the genome with at least one expressed member of the MSDIN gene family. Immunologically, amanitin and AbPOPB show a high degree of colocalization, indicating that toxin biosynthesis and accumulation occur in the same cells and possibly in the same subcellular compartments.

Amatoxins, the lethal constituents of poisonous mushrooms in the genus Amanita, are bicyclic octapeptides. Two genes in A. bisporigera, AMA1 and PHA1, directly encode alpha-amanitin, an amatoxin, and the related bicyclic heptapeptide phallacidin, a phallotoxin, indicating that these compounds are synthesized on ribosomes and not by nonribosomal peptide synthetases. alpha-Amanitin and phallacidin are synthesized as proproteins of 35 and 34 amino acids, respectively, from which they are predicted to be cleaved by a prolyl oligopeptidase. AMA1 and PHA1 are present in other toxic species of Amanita section Phalloidae but are absent from nontoxic species in other sections. The genomes of A. bisporigera and A. phalloides contain multiple sequences related to AMA1 and PHA1. The predicted protein products of this family of genes are characterized by a hypervariable "toxin" region capable of encoding a wide variety of peptides of 7-10 amino acids flanked by conserved sequences. Our results suggest that these fungi have a broad capacity to synthesize cyclic peptides on ribosomes.