ABSTRACT
The melleolides are structurally unique and bioactive natural products of the basidiomycete genus Armillaria. Here, we report on cytotoxic effects of melleolides from Armillaria mellea towards non-transformed human primary monocytes and human cancer cell lines, respectively. In contrast to staurosporine or pretubulysin that are less cytotoxic for monocytes, the cytotoxic potency of the active melleolides in primary monocytes is comparable to that in cancer cells. The onset of the cytotoxic effects of melleolides was rapid (within <1 h), as compared to the apoptosis inducer staurosporine, the protein biosynthesis inhibitor cycloheximide, and the DNA transcription inhibitor actinomycin D (>5 h, each). Side-by-side comparison with the detergent triton X-100 and staurosporine in microscopic and flow cytometric analysis studies as well as analysis of the viability of mitochondria exclude cell lysis and apoptosis as relevant or primary mechanisms. Our results rather point to necrotic features of cell death mediated by an as yet elusive but rapid mechanism.
Subject(s)
Apoptosis/drug effects , Monocytes/drug effects , Cell Survival/drug effects , Cells, Cultured , HeLa Cells , Humans , K562 Cells , Microscopy , Monocytes/cytology , Monocytes/metabolism , Oligopeptides/chemistry , Oligopeptides/isolation & purification , Oligopeptides/toxicity , Staurosporine/chemistry , Staurosporine/isolation & purification , Staurosporine/toxicity , Structure-Activity RelationshipABSTRACT
The fungal genus Armillaria is unique in that it is the only natural source of melleolide antibiotics, i.e., protoilludene alcohols esterified with orsellinic acid or its derivatives. This class of natural products is known to exert antimicrobial and cytotoxic effects. Here, we present a refined relationship between the structure and the antimicrobial activity of the melleolides. Using both agar diffusion and broth dilution assays, we identified the Δ(2,4)-double bond of the protoilludene moiety as a key structural feature for antifungal activity against Aspergillus nidulans, Aspergillus flavus, and Penicillium notatum. These findings contrast former reports on cytotoxic activities and may indicate a different mode of action towards susceptible fungi. We also report the isolation and structure elucidation of five melleolides (6'-dechloroarnamial, 6'-chloromelleolide F, 10-hydroxy-5'-methoxy-6'-chloroarmillane, and 13-deoxyarmellides A and B), along with the finding that treatment with an antifungal melleolide impacts transcription of A. nidulans natural product genes.
Subject(s)
Anti-Bacterial Agents/isolation & purification , Anti-Bacterial Agents/pharmacology , Anti-Infective Agents/isolation & purification , Anti-Infective Agents/pharmacology , Antifungal Agents/isolation & purification , Antifungal Agents/pharmacology , Armillaria/chemistry , Sesquiterpenes/isolation & purification , Sesquiterpenes/pharmacology , Anti-Bacterial Agents/chemistry , Anti-Infective Agents/chemistry , Antifungal Agents/chemistry , Aspergillus/drug effects , Fungi/drug effects , Molecular Structure , Nuclear Magnetic Resonance, Biomolecular , Resorcinols , Sesquiterpenes/chemistry , Structure-Activity RelationshipABSTRACT
Little is known about polyketide biosynthesis in mushrooms (basidiomycota). In this study, we investigated the iterative type I polyketide synthase (PKS) ArmB of the tree pathogen Armillaria mellea, a producer of cytotoxic melleolides (i.e., polyketides esterified with various sesquiterpene alcohols). Heterologously produced ArmB showed orsellinic acid (OA) synthase activity in vitro. Further, we demonstrate cross-coupling activity of ArmB, which forms OA esters with various alcohols. Using a tricyclic Armillaria sesquiterpene alcohol, we reconstituted the biosynthesis of melledonol. Intermolecular transesterification reactions may represent a general mechanism of fungal PKSs to create structural diversity of small molecules. Phylogenetic network construction of thioesterase domains of both basidiomycetes and ascomycetes suggests that the fungal nonreducing PKS family has likely evolved from an ancient OA synthase and has gained versatility by adopting Claisen-like cyclase or transferase activity.
Subject(s)
Anti-Bacterial Agents/biosynthesis , Polyketide Synthases/metabolism , Sesquiterpenes/metabolism , Anti-Bacterial Agents/chemistry , Armillaria/enzymology , Armillaria/metabolism , Esterification , Evolution, Molecular , Polyketide Synthases/classification , Polyketide Synthases/genetics , Resorcinols/chemistry , Resorcinols/metabolism , Sesquiterpenes/chemistryABSTRACT
Melleolide sesquiterpene aryl esters are secondary products of the mushroom genus Armillaria. We compared the cytotoxicity of eleven melleolides--five thereof are new natural products--against four human cancer cell lines. Armillaridin, 4-O-methylarmillaridin, and dehydroarmillylorsellinate were most active, at IC(50) = 3.0, 4.1 and 5.0 µM, respectively, against Jurkat T cells for the former two compounds, and K-562 cells for the latter. Dehydroarmillylorsellinate did not inhibit respiration and RNA-synthesis of K-562 cells at 5 µM. However, replication of DNA dropped to 35% after 120 min at this concentration, and translational activity also decreased.
Subject(s)
Anti-Bacterial Agents/pharmacology , Salicylates/pharmacology , Sesquiterpenes/pharmacology , Anti-Bacterial Agents/chemistry , Basidiomycota/chemistry , Cell Line, Tumor , DNA/drug effects , Humans , In Vitro Techniques , Magnetic Resonance Spectroscopy , Molecular Structure , Salicylates/chemistry , Sesquiterpenes/chemistryABSTRACT
A remarkable feature of filamentous fungi is their ability to produce small yet structurally complex and often bioactive natural products. In this mini-review, we cover advances in the research on fungal secondary metabolites, particularly mycotoxins, and focus on biosynthetic aspects as well as on the complex regulatory mechanisms which control the expression of biosynthetic genes. We also highlight the increasing impact of genomics and transcriptomics, which help explore the realm of secondary metabolism of fungi.
